Journal of Translational Medicine

Mesothelin-based CAR-T cells exhibit potent antitumor activity against ovarian cancer

Abstract Background Ovarian cancer (OC) is characterized by its rapid growth and spread which, accompanied by a low 5-year survival rate, necessitates the development of improved treatments. In ovarian cancer, the selective overexpression of Mucin-16 (MUC16, CA125) in tumor cells highlights its potential as a promising target for developing anti-tumor therapies. However, the potential effectiveness of CAR-T cell therapy that targets MUC16 in ovarian cancer cells is unknown. Methods The expression of MUC16 in viable OC cells was detected using immunofluorescence and flow cytometry techniques. A MSLN-CAR construct, comprising the MUC16-binding polypeptide region of mesothelin (MSLN), a CD8 hinge spacer and transmembrane domain, 4-1BB, and CD3ζ endo-domains; was synthesized and introduced into T cells using lentiviral particles. The cytotoxicity of the resultant CAR-T cells was evaluated in vitro using luciferase assays. Cytokine release by CAR-T cells was measured using enzyme-linked immunosorbent assays. The anti-tumor efficacy of the CAR-T cells was subsequently assessed in mice through both systemic and local administration protocols. Results MSLN-CAR T cells exhibited potent cytotoxicity towards OVCAR3 cells and their stem-like cells that express high levels of MUC16. Also, MSLN-CAR T cells were inefficient at killing SKOV3 cells that express low levels of MUC16, but were potently cytotoxic to such cells overexpressing MUC16. Moreover, MSLN-CAR T cells delivered via tail vein or peritoneal injection could shrink OVCAR3 xenograft tumors in vivo, with sustained remission observed following peritoneal delivery of MSLN-CAR T cells. Conclusions Collectively, these results suggested that MSLN-CAR T cells could potently eliminate MUC16- positive ovarian cancer tumor cells both in vitro and in vivo, thereby providing a promising therapeutic intervention for MUC16-positive patients.

RPL22L1 fosters malignant features of cervical cancer via the modulation of DUSP6-ERK axis

Cervical cancer remains one of the leading causes of cancer-related deaths among women globally, and there is still a need to research molecular targets that can be used for prognosis assessment and personalized molecular therapies. Here, we investigate the role of potential molecular target ribosomal L22-like 1 (RPL22L1) on cervical cancer, identify its potential mechanisms, and explore its related applications in prognosis and molecular therapies. Multiple cervical cancer cohorts online, tissue microarrays and clinical tissue specimens were analyzed for the association between RPL22L1 expression and patient outcomes. Functional and molecular biology studies of cell and mice models were used to clarify the effects and potential mechanisms of RPL22L1 on cervical cancer. RPL22L1 is highly expressed in both cervical adenocarcinoma and squamous cell carcinoma, and its expression is significantly associated with histology grade, clinical stage, recurrence, vascular space involvement, tumor sizes and poor prognosis. In vitro and in vivo experiment revealed that RPL22L1 overexpression significantly promoted cervical cancer cell proliferation, migration, invasion, tumorigenicity and Sorafenib resistance, which were attenuated by RPL22L1 knockdown. Mechanistically, RPL22L1 competitively binds to ERK phosphatase DUSP6, leading to excessive activation of ERK. The combined application of ERK inhibitors can effectively inhibit RPL22L1 overexpressing cervical cancer cells both in vivo and in vitro. RPL22L1 promotes malignant biological behavior of cervical cancer cells by competitively binding with DUSP6, thereby activating the ERK pathway. The combined use of Sorafenib and an ERK inhibitor is a potentially effective molecular targeted therapy for RPL22L1-high cervical cancer.

Eight-lncRNA signature of cervical cancer were identified by integrating DNA methylation, copy number variation and transcriptome data

AbstractBackgroundCopy number variation (CNV) suggests genetic changes in malignant tumors. Abnormal expressions of long non-coding RNAs (lncRNAs) resulted from genomic and epigenetic abnormalities play a driving role in tumorigenesis of cervical cancer. However, the role of lncRNAs-related CNV in cervical cancer remained largely unclear.MethodsThe data of messenger RNAs (mRNAs), DNA methylation, and DNA copy number were collected from 292 cervical cancer specimens. The prognosis-related subtypes of cervical cancer were determined by multi-omics integration analysis, and protein-coding genes (PCGs) and lncRNAs with subtype-specific expressions were identified. The CNV pattern of the subtype-specific lncRNAs was analyzed to identify the subtype-specific lncRNAs. A prognostic risk model based on lncRNAs was established by least absolute shrinkage and selection operator (LASSO).ResultsMulti-omics integration analysis identified three molecular subtypes incorporating 617 differentially expressed lncRNAs and 1395 differentially expressed PCGs. The 617 lncRNAs were found to intersect with disease-related lncRNAs. Functional enrichment showed that 617 lncRNAs were mainly involved in tumor metabolism, immunity and other pathways, such as p53 and cAMP signaling pathways, which are closely related to the development of cervical cancer. Finally, according to CNV pattern consistent with differential expression analysis, we established a lncRNAs-based signature consisted of 8 lncRNAs, namely, RUSC1-AS1, LINC01990, LINC01411, LINC02099, H19, LINC00452, ADPGK-AS1, C1QTNF1-AS1. The interaction of the 8 lncRNAs showed a significantly poor prognosis of cervical cancer patients, which has also been verified in an independent dataset.ConclusionOur study expanded the network of CNVs and improved the understanding on the regulatory network of lncRNAs in cervical cancer, providing novel biomarkers for the prognosis management of cervical cancer patients.

Dietary and circulating omega-6 fatty acids and their impact on cardiovascular disease, cancer risk, and mortality: a global meta-analysis of 150 cohorts and meta-regression

Targeting ubiquitin-specific protease 14 reduces metastatic potential and metabolic activity in cervical cancer via direct modulation of monocarboxylate transporter-4

Cervical cancer (CC) remains a major health burden in low- and middle-income countries, where HPV vaccination coverage is suboptimal. Ubiquitin-specific peptidase 14 (USP14), a proteasome-associated deubiquitinase, has emerged as a potential driver of tumorigenesis, but its role in CC and impact on tumor metabolism remain poorly defined. In-silico analysis of RNA and proteomics data from publicly available datasets was performed to assess the expression and prognostic value of USP14. Expression was validated in CC cell lines (HeLa, SiHa, CaSki) in comparison to non-transformed MCF10A cells. Functional studies involved USP14 knockdown (siRNA) or inhibition (IU1), followed by various cell viability, metastasis and invasion assays. Comparative proteomics was employed to identify USP14 interacting partners, with a particular focus on monocarboxylate transporter 4 (MCT4). USP14-MCT4 interactions were confirmed by co-immunoprecipitation, ubiquitination assays, and molecular dynamics simulations. Immunohistochemistry was performed on retrospective cervical squamous cell carcinoma tissues to correlate USP14 and MCT4 expression. High USP14 expression correlated with poor overall survival in CC patients and was elevated in CC cell lines. Genetic or pharmacological inhibition of USP14 significantly reduced CC cell proliferation, metastasis, and invasion. Proteomic profiling identified MCT4, a key lactate transporter in tumor metabolism, as a novel USP14 interactor. USP14 knockdown decreased MCT4 protein levels and stability, suggesting deubiquitination-dependent regulation. Immunoprecipitation confirmed direct binding between USP14 and MCT4, and molecular dynamics simulations revealed distinct binding interfaces. In patient tissues, USP14 and MCT4 expression were positively correlated. This study uncovers MCT4 as a novel substrate of USP14, linking USP14 activity to metabolic reprogramming in CC. The USP14-MCT4 axis represents a previously unrecognized oncogenic pathway with therapeutic potential. Targeting USP14 could simultaneously impair tumor growth and disrupt lactate metabolism, offering a promising strategy for CC treatment.

Radiology-based artificial intelligence for predicting targeted therapy response in pan-cancer: a comprehensive review

Rac1 in gastric cancer: a molecular driver of invasion, EMT, and therapeutic resistance

Abstract Gastric cancer (GC) ranks as the fifth most common cancer worldwide and is the third main cause of cancer-related mortality, posing a substantial burden to global public health. Research suggests that targeted therapy and immunotherapy may become more effective treatment options for advanced, unresectable, or metastatic gastric cancer. Ras-related C3 botulinum toxin substrate 1 (Rac1), a small GTP-binding protein within the Rac subfamily of the Rho GTPase family, is a critical molecule that promotes cancer cell invasion and metastasis by regulating signal transmission and promoting cell polarity. It has emerged as a key driver of tumor development and metastasis in several malignancies, including breast, lung, prostate, ovarian, gastric, and pancreatic cancers. This review summarizes the structure, regulatory dynamics, and signaling mechanisms of Rac1 in gastric cancer growth, epithelial-to-mesenchymal transition (EMT), and metastasis, as well as the roles of factors such as hypoxia, oxidative stress, and H. pylori infection. Additionally, it highlights small-molecule inhibitors targeting Rac1, miRNAs capable of suppressing Rac1, and ongoing research on Rac1-related immunotherapy. The potential of Rac1 as a therapeutic biomarker in gastric cancer and the remaining challenges in this area are also discussed. This review advances the understanding of Rac1’s role in gastric cancer, provides a theoretical foundation for further studies, and supports the development of precision medicine for this disease.

Unlocking the mitochondrial functional code: unraveling the pathogenesis of ovarian cancer and innovative targets to inhibit malignant behavior

This article focuses on ovarian cancer (OC), the most lethal gynecological malignancy, whose risk is influenced by multiple factors, including genetic background, age, reproductive history, parity, obesity status, and smoking habits. Mitochondria, as the core organelles in eukaryotic cells, play a pivotal role in the initiation and progression of OC. In recent years, growing evidence has revealed a close relationship between mitochondrial dysfunction and the accelerated proliferation, enhanced invasiveness, metastasis ability, and therapy resistance of OC. This paper provides an in-depth analysis of the complex interactions between mitochondrial dysfunction and the malignant biological characteristics of OC, as well as the underlying regulatory mechanisms of mitochondrial function. Furthermore, it elaborates on how genetic variations, regulatory factors, and the tumor microenvironment (TME) influence mitochondrial function and drive the malignant progression of OC. Additionally, this paper comprehensively summarizes therapeutic strategies targeting mitochondrial dysfunction in OC, aiming to provide novel insights and strategies for clinical research and treatment.

Role of KLF5 in enhancing ovarian cancer stemness and PARPi resistance: mechanisms and therapeutic targeting

Abstract Background Ovarian cancer (OC) often presents at advanced stages with poor prognosis. Although poly(ADP-ribose) polymerase inhibitors (PARPi) offer clinical benefits, resistance remains a major challenge. This study investigates the role of KLF5 in regulating OC cell stemness and contributing to PARPi resistance. Methods Gene expression analysis was conducted on OC cell lines and their PARPi-resistant counterparts. qRT-PCR and Western blotting assessed the expression levels of stemness markers and KLF5. IHC evaluated KLF5 expression in ovarian cancer tissue samples. Sphere formation and ALDH activity assays were used to evaluate stemness. Chromatin immunoprecipitation (ChIP) investigated KLF5’s binding to the Vimentin promoter. The effects of the KLF5 inhibitor ML264 were tested in vitro using cell viability and apoptosis assays, and in vivo using a xenograft mouse model to evaluate tumor growth and response to PARPi treatment. Results PARPi-resistant OC cells showed elevated stemness, indicated by increased SOX2, KLF4, Nanog, and OCT4 expression. KLF5 was significantly upregulated in these cells and linked to poor clinical outcomes. PARPi-resistant cells formed larger and more numerous spheres and had higher ALDH activity. KLF5 bound to the Vimentin promoter, upregulating its expression. Inhibition of KLF5 with ML264 reduced stemness features, decreased Vimentin expression, and resensitized resistant cells to PARPi. In vivo, ML264-treated mice with PARPi-resistant tumors exhibited reduced tumor growth and increased sensitivity to PARPi. Conclusion KLF5 enhances stemness and contributes to PARPi resistance in ovarian cancer through Vimentin regulation. Targeting KLF5 offers a promising therapeutic strategy to overcome resistance and improve patient outcomes.

Multi-dimensional characterization of cellular states reveals clinically relevant immunological subtypes and therapeutic vulnerabilities in ovarian cancer

Diverse cell types and cellular states in the tumor microenvironment (TME) are drivers of biological and therapeutic heterogeneity in ovarian cancer (OV). Characterization of the diverse malignant and immunology cellular states that make up the TME and their associations with clinical outcomes are critical for cancer therapy. However, we are still lack of knowledge about the cellular states and their clinical relevance in OV. We manually collected the comprehensive transcriptomes of OV samples and characterized the cellular states and ecotypes based on a machine-learning framework. The robustness of the cellular states was validated in independent cohorts and single-cell transcriptomes. The functions and regulators of cellular states were investigated. Meanwhile, we thoroughly examined the associations between cellular states and various clinical factors, including clinical prognosis and drug responses. We depicted and characterized an immunophenotypic landscape of 3,099 OV samples and 80,044 cells based on a machine learning framework. We identified and validated 32 distinct transcriptionally defined cellular states from 12 cell types and three cellular communities or ecotypes, extending the current immunological subtypes in OV. Functional enrichment and upstream transcriptional regulator analyses revealed cancer hallmark-related pathways and potential immunological biomarkers. We further investigated the spatial patterns of identified cellular states by integrating the spatially resolved transcriptomes. Moreover, prognostic landscape and drug sensitivity analysis exhibited clinically relevant immunological subtypes and therapeutic vulnerabilities. Our comprehensive analysis of TME helps leveraging various immunological subtypes to highlight new directions and targets for the treatment of cancer.

The G-quadruplex ligand CX-5461: an innovative candidate for disease treatment

Development of model for identifying homologous recombination deficiency (HRD) status of ovarian cancer with deep learning on whole slide images

Homologous recombination deficiency (HRD) refers to the dysfunction of homologous recombination repair (HRR) at the cellular level. The assessment of HRD status has the important significance for the formulation of treatment plans, efficacy evaluation, and prognosis prediction of patients with ovarian cancer. This study aimed to construct a deep learning-based classifier for identifying tumor regions from whole slide images (WSIs) and stratify the HRD status of patients with ovarian cancer (OC). The deep learning models were trained on 205 H&E-stained sections which contained 205 ovarian cancer patients, 64 were found to have HRD status while 141 had homologous recombination proficiency (HRP) status from two institutions Memorial Sloan Kettering Cancer Center (MSKCC) and Zhongda Hospital, Southeast University. The framework includes tumor regions identification by UNet + + and subtypes of ovarian cancer classifier construction. Referring to the EasyEnsemble, we classified the HRP patients into three distributed subsets. These three subsets of HRP patients were combined with the HRD patients to establish three new training groups for subsequent model construction. The three models were integrated into a single model named Ensemble Model. The UNet + + algorithm segmented tumor regions with 81.8% accuracy, 85.9% recall, 83.8% dice score and 68.3% IoU. The AUC of the Ensemble Model was 0.769 (Precision = 0.800, Recall = 0.727, F1-score = 0.762) in the study. The most discriminative features between HRD and HRP comprised S_mean_dln_obtuse_ratio, S_mean_dln_acute_ratio and mean_Graph_T-S_Betweenness_normed. The models we constructed enables accurate discrimination between tumor and non-tumor tissues in ovarian cancer as well as the prediction of HRD status for patients with ovarian cancer.

Association of pre- and post-diagnosis dietary total antioxidant capacity (TAC) and composite dietary antioxidant index (CDAI) with overall survival in patients with ovarian cancer: a prospective cohort study

Abstract Background The evidence on the relationship of dietary antioxidant nutrients with the survival of ovarian cancer (OC) remains scarce. Objective This study aimed to investigate these associations in a prospective cohort of Chinese patients with OC. Methods In this prospective cohort study, patients with epithelial OC completed a food frequency questionnaire at diagnosis and 12 months post-diagnosis, and were followed from 2015 to 2023. Dietary total antioxidant capacity (TAC) and composite dietary antioxidant index (CDAI) were calculated based on specific antioxidant nutrients. We examined the associations of pre-diagnosis, post-diagnosis, and changes from pre-diagnosis to post-diagnosis in TAC, CDAI, and representative antioxidant nutrients with overall survival (OS) among patients with OC. Multivariable Cox proportional-hazards models were applied to calculate the hazard ratios (HR) and 95% confidence intervals (CI). Dose–response relationships were evaluated by restricted cubic splines. Results Among the total 560 patients with OC, there were 211 (37.68%) deaths during a median follow-up of 44.40 (interquartile range: 26.97–61.37) months. High pre-diagnosis TAC (HR = 0.58; 95% CI 0.38–0.8) and vitamin C intake (HRT3 vs. T1 = 0.36; 95% CI 0.21–0.61), and post-diagnosis TAC (HR = 0.57; 95% CI 0.37–0.8), CDAI (HR = 0.57; 95% CI 0.33–0.9), and β-carotene intake (HRT3 vs. T1 = 0.55; 95% CI 0.32–0.97) were significantly associated with improved OS. Compared to patients with constantly low pre- and post-diagnosis TAC and CDAI, those with consistently higher TAC (HRMedium-Medium vs. Low-Low = 0.53; 95% CI 0.29–0.97; HRHigh-High vs. Low-Low = 0.40; 95% CI 0.16–0.94) and CDAI (HRHigh-High vs. Low-Low = 0.33; 95% CI 0.12–0.88) experienced better OS. Conclusion High pre- and post-diagnosis TAC, and post-diagnosis CDAI were associated with improved OC survival, suggesting that consistent high-intake of antioxidant-rich food may be beneficial for the prognosis of OC.

UBE2J1 is identified as a novel plasma cell-related gene involved in the prognosis of high-grade serous ovarian cancer

Immune cells within tumor tissues play important roles in remodeling the tumor microenvironment, thus affecting tumor progression and the therapeutic response. The current study was designed to identify key markers of plasma cells and explore their role in high-grade serous ovarian cancer (HGSOC). We utilized single-cell sequencing data from the Gene Expression Omnibus (GEO) database to identify key immune cell types within HGSOC tissues and to extract related markers via the Seurat package. The effects of immune cell markers on prognosis were analyzed via univariate Cox regression, least absolute shrinkage and selection operator (LASSO) and gene set variation analysis (GSVA) of bulk sequencing data from The Cancer Genome Atlas (TCGA)-HGSOC cohort. Finally, the effects of key markers on HGSOC cells were evaluated via Cell Counting Kit-8 (CCK-8), Transwell, colony formation, wound healing, immunofluorescence and in vivo tumor growth assays. At the single-cell level, we detected a significant increase in the proportion of plasma cells in HGSOC samples compared to that in normal ovarian samples. Within HGSOC tissues, these plasma cells were found to interact with CD8 + T cells, fibroblasts and endothelial cells. In addition, patients in the high-plasma cell-related score group had better survival rates and higher epithelial‒mesenchymal transition (EMT), apoptosis and immune scores. Moreover, univariate Cox and LASSO regression analyses revealed that ubiquitin-conjugating enzyme E2 J1 (UBE2J1) is a prognostic marker in HGSOC. Further functional studies revealed that overexpression of UBE2J1 promoted cell proliferation, invasion, migration and colony formation, whereas UBE2J1 knockdown attenuated the abovementioned cellular behaviors. Additionally, UBE2J1 overexpression promoted EMT, as evidenced by alterations in the protein expression levels of N-cadherin, snail family transcriptional repressor 2 (Slug), Twist family BHLH transcription factor 1 (Twist 1) and E-cadherin. Moreover, we found that UBE2J1 silencing was able to inhibit the tumor growth in vivo. Overall, this study elucidated the role of plasma cells and revealed UBE2J1 as a novel oncogene in HGSOC, uncovering new mechanisms related to HGSOC tumorigenesis and promising therapeutic targets for HGSOC patients.

Two-sample Mendelian Randomization to evaluate the causal relationship between inflammatory arthritis and female-specific cancers

Abstract Background There is evidence that inflammatory arthritis in the form of ankylosing spondylitis (AS), psoriatic arthritis (PsA), and rheumatoid arthritis are both positively and negatively associated with certain female-specific cancers. However, the study results are very heterogeneous. Methods Based on up to 375,814 European women, we performed an iterative two-sample Mendelian randomization to assess causal effects of the occurrence of the inflammatory arthritis on the risk of female-specific cancer in form of breast, endometrial, and ovarian cancer sites as well as their subtypes. Evidence was strengthened by using similar exposures for plausibility or by replication with a subsequent meta-analysis. P-values were Bonferroni adjusted. Results Genetic liability to AS was associated with ovarian cancer (OR = 1.03; 95% CI: [1.01; 1.04]; $$\:{P}_{adj}$$=0.029) and liability to PsA with breast cancer (OR = 1.02; CI: [1.01; 1.04]; $$\:{P}_{adj}$$=0.002). Subgroup analyses revealed that the high-grade serous ovarian cancer (OR = 1.04; CI: [1.02; 1.06]; $$\:{P}_{adj}$$=0.015) and the ER- breast cancer (OR = 1.04; CI: [1.01; 1.07]; $$\:{P}_{adj}$$=0.118) appeared to drive the observed associations, respectively. No further associations were found between the remaining inflammatory arthritis phenotypes and female-specific cancers. Conclusions This study suggests that AS is a risk factor for ovarian cancer, while PsA is linked to an increased breast cancer risk. These results are important for physicians caring women with inflammatory arthritis to advise their patients on cancer screening and preventive measures.

The application of HER2 and CD47 CAR-macrophage in ovarian cancer

Abstract Background The chimeric antigen receptor (CAR)-T therapy has a limited therapeutic effect on solid tumors owing to the limited CAR-T cell infiltration into solid tumors and the inactivation of CAR-T cells by the immunosuppressive tumor microenvironment. Macrophage is an important component of the innate and adaptive immunity, and its unique phagocytic function has been explored to construct CAR macrophages (CAR-Ms) against solid tumors. This study aimed to investigate the therapeutic application of CAR-Ms in ovarian cancer. Methods In this study, we constructed novel CAR structures, which consisted of humanized anti-HER2 or CD47 scFv, CD8 hinge region and transmembrane domains, as well as the 4-1BB and CD3ζ intracellular domains. We examined the phagocytosis of HER2 CAR-M and CD47 CAR-M on ovarian cancer cells and the promotion of adaptive immunity. Two syngeneic tumor models were used to estimate the in vivo antitumor activity of HER2 CAR-M and CD47 CAR-M. Results We constructed CAR-Ms targeting HER2 and CD47 and verified their phagocytic ability to ovarian cancer cells in vivo and in vitro. The constructed CAR-Ms showed antigen-specific phagocytosis of ovarian cancer cells in vitro and could activate CD8+ cytotoxic T lymphocyte (CTL) to secrete various anti-tumor factors. For the in vivo model, mice with human-like immune systems were used. We found that CAR-Ms enhanced CD8+ T cell activation, affected tumor-associated macrophage (TAM) phenotype, and led to tumor regression. Conclusions We demonstrated the inhibition effect of our constructed novel HER2 CAR-M and CD47 CAR-M on target antigen-positive ovarian cancer in vitro and in vivo, and preliminarily verified that this inhibitory effect is due to phagocytosis, promotion of adaptive immunity and effect on tumor microenvironment.

Unraveling the interplay of CD8 + T cells and microRNA signaling in cancer: implications for immune dysfunction and therapeutic approaches

Repurposing anti-mesothelin CAR-NK immunotherapy against colorectal cancer

KLF14 directly downregulates the expression of GPX4 to exert antitumor effects by promoting ferroptosis in cervical cancer

Cervical cancer is the fourth leading cause of cancer-related death among women worldwide, and effective therapeutic strategies for its treatment are limited. Recent studies have indicated that ferroptosis, a form of regulated cell death, is a promising therapeutic strategy. KLF14 has been shown to regulate both cell proliferation and apoptosis in cervical cancer. However, its role in modulating lipid peroxidation and ferroptosis remains largely unexplored and enigmatic. SiHa and HeLa cells were transduced with lentiviral vectors to overexpress KLF14. Protein levels were analyzed via western blotting and immunohistochemistry (IHC). LDH assays, calcein-AM/propidium iodide (PI) staining, and generation of cell growth curves using a real-time cell analysis (RTCA) system were used to detect cell damage and proliferation. Cellular ROS, lipid ROS, transmission electron microscopy (TEM), and Fe KLF14 is abnormally expressed in various tumors and downregulated in cervical cancer. Overexpression of KLF14 induced ferroptosis and inhibited cell proliferation in vitro as well as xenograft tumorigenicity in vivo. Mechanistic studies revealed that KLF14 binds to the promoter of GPX4, suppressing its transcriptional activity and thereby decreasing its expression, which contributes to the induction of ferroptosis. Truncation and point mutation analyses of the GPX4 promoter revealed multiple binding sites for KLF14 within the - 1000 bp to + 35 bp region, which are responsible for its inhibitory effect on GPX4 transcription. Additionally, deletion of the zinc finger motif in KLF14 abolished its inhibitory effect on GPX4 promoter activity and cell proliferation. Our data revealed a previously unidentified function of KLF14 in promoting ferroptosis, which results in the suppression of cell proliferation. Mechanistically, we revealed a novel regulatory mechanism by which KLF14 targets GPX4. These findings suggest a novel strategy to induce ferroptosis through the targeting of KLF14 in human cervical cancer cells.

Association between pre-diagnosis and post-diagnosis Alternate Mediterranean Diet and ovarian cancer survival: evidence from a prospective cohort study

There is currently a lack of comprehensive evidence regarding the correlation between Alternate Mediterranean Diet (AMED) and the survival of patients with ovarian cancer (OC). This prospective cohort study first assessed the association of AMED, not only pre-diagnosis and post-diagnosis but also the change from pre-diagnosis to post-diagnosis with OC survival. A total of 560 OC patients were included in the study, and their dietary intake was assessed using a reliable 111-item food frequency questionnaire. The overall survival (OS) of the patients was monitored through active follow-up and review of medical records until February 16th, 2023. Cox proportional hazard regression models were utilized to compute the hazard ratios (HRs) and their corresponding 95% confidence intervals (CIs). Out of the total 560 patients with OC, 211 (37.68%) succumbed during a median follow-up period of 44.40 months (interquartile range: 26.97-61.37). Comparative analysis indicated a significant association between the highest tertiles of pre-diagnosis (HR = 0.59; 95% CI 0.38-0.90; P High pre-diagnosis and post-diagnosis AMED was associated with an improved OS in patients with OC, suggesting that maintaining a consistently high intake of AMED could potentially benefit the prognosis of OC.

Effect of the mRNA decapping enzyme scavenger (DCPS) inhibitor RG3039 on glioblastoma

Enhanced anti-tumor efficacy with multi-transgene armed mesenchymal stem cells for treating peritoneal carcinomatosis

Abstract Background Mesenchymal stem cells (MSCs) have garnered significant interest for their tumor-tropic property, making them potential therapeutic delivery vehicles for cancer treatment. We have previously shown the significant anti-tumour activity in mice preclinical models and companion animals with naturally occurring cancers using non-virally engineered MSCs with a therapeutic transgene encoding cytosine deaminase and uracil phosphoribosyl transferase (CDUPRT) and green fluorescent protein (GFP). Clinical studies have shown improved response rate with combinatorial treatment of 5-fluorouracil and Interferon-beta (IFNb) in peritoneal carcinomatosis (PC). However, high systemic toxicities have limited the clinical use of such a regime. Methods In this study, we evaluated the feasibility of intraperitoneal administration of non-virally engineered MSCs to co-deliver CDUPRT/5-Flucytosine prodrug system and IFNb to potentially enhance the cGAS-STING signalling axis. Here, MSCs were engineered to express CDUPRT or CDUPRT-IFNb. Expression of CDUPRT and IFNb was confirmed by flow cytometry and ELISA, respectively. The anti-cancer efficacy of the engineered MSCs was evaluated in both in vitro and in vivo model. ES2, HT-29 and Colo-205 were cocultured with engineered MSCs at various ratio. The cell viability with or without 5-flucytosine was measured with MTS assay. To further compare the anti-cancer efficacy of the engineered MSCs, peritoneal carcinomatosis mouse model was established by intraperitoneal injection of luciferase expressing ES2 stable cells. The tumour burden was measured through bioluminescence tracking. Results Firstly, there was no changes in phenotypes of MSCs despite high expression of the transgene encoding CDUPRT and IFNb (CDUPRT-IFNb). Transwell migration assays and in-vivo tracking suggested the co-expression of multiple transgenes did not impact migratory capability of the MSCs. The superiority of CDUPRT-IFNb over CDUPRT expressing MSCs was demonstrated in ES2, HT-29 and Colo-205 in-vitro. Similar observations were observed in an intraperitoneal ES2 ovarian cancer xenograft model. The growth of tumor mass was inhibited by ~ 90% and 46% in the mice treated with MSCs expressing CDUPRT-IFNb or CDUPRT, respectively. Conclusions Taken together, these results established the effectiveness of MSCs co-expressing CDUPRT and IFNb in controlling and targeting PC growth. This study lay the foundation for the development of clinical trial using multigene-armed MSCs for PC.

Transcript and protein signatures derived from shared molecular interactions across cancers are associated with mortality

Abstract Background Characterization of shared cancer mechanisms have been proposed to improve therapy strategies and prognosis. Here, we aimed to identify shared cell–cell interactions (CCIs) within the tumor microenvironment across multiple solid cancers and assess their association with cancer mortality. Methods CCIs of each cancer were identified by NicheNet analysis of single-cell RNA sequencing data from breast, colon, liver, lung, and ovarian cancers. These CCIs were used to construct a shared multi-cellular tumor model (shared-MCTM) representing common CCIs across cancers. A gene signature was identified from the shared-MCTM and tested on the mRNA and protein level in two large independent cohorts: The Cancer Genome Atlas (TCGA, 9185 tumor samples and 727 controls across 22 cancers) and UK biobank (UKBB, 10,384 cancer patients and 5063 controls with proteomics data across 17 cancers). Cox proportional hazards models were used to evaluate the association of the signature with 10-year all-cause mortality, including sex-specific analysis. Results A shared-MCTM was derived from five individual cancers. A shared gene signature was extracted from this shared-MCTM and the most prominent regulatory cell type, matrix cancer-associated fibroblast (mCAF). The signature exhibited significant expression changes in multiple cancers compared to controls at both mRNA and protein levels in two independent cohorts. Importantly, it was significantly associated with mortality in cancer patients in both cohorts. The highest hazard ratios were observed for brain cancer in TCGA (HR [95%CI] = 6.90[4.64–10.25]) and ovarian cancer in UKBB (5.53[2.08–8.80]). Sex-specific analysis revealed distinct risks, with a higher mortality risk associated with the protein signature score in males (2.41[1.97–2.96]) compared to females (1.84[1.44–2.37]). Conclusion We identified a gene signature from a comprehensive shared-MCTM representing common CCIs across different cancers and revealed the regulatory role of mCAF in the tumor microenvironment. The pathogenic relevance of the gene signature was supported by differential expression and association with mortality on both mRNA and protein levels in two independent cohorts.

Cancer-associated fibroblast-secreted FGF7 as an ovarian cancer progression promoter

Abstract Background Ovarian cancer (OC) is distinguished by its aggressive nature and the limited efficacy of current treatment strategies. Recent studies have emphasized the significant role of cancer-associated fibroblasts (CAFs) in OC development and progression. Methods Employing sophisticated machine learning techniques on bulk transcriptomic datasets, we identified fibroblast growth factor 7 (FGF7), derived from CAFs, as a potential oncogenic factor. We investigated the relationship between FGF7 expression and various clinical parameters. A series of in vitro experiments were undertaken to evaluate the effect of CAFs-derived FGF7 on OC cell activities, such as proliferation, migration, and invasion. Single-cell transcriptomic analysis was also conducted to elucidate the interaction between FGF7 and its receptor. Detailed mechanistic investigations sought to clarify the pathways through which FGF7 fosters OC progression. Results Our findings indicate that higher FGF7 levels correlate with advanced tumor stages, increased vascular invasion, and poorer prognosis. CAFs-derived FGF7 significantly enhanced OC cell proliferation, migration, and invasion. Single-cell analysis and in vitro studies revealed that CAFs-derived FGF7 inhibits the ubiquitination and degradation of hypoxia-inducible factor 1 alpha (HIF-1α) via FGFR2 interaction. Activation of the FGF7/HIF-1α pathway resulted in the upregulation of mesenchymal markers and downregulation of epithelial markers. Importantly, in vivo treatment with neutralizing antibodies targeting CAFs-derived FGF7 substantially reduced tumor growth. Conclusion Neutralizing FGF7 in the medium or inhibiting HIF-1α signaling reversed the effects of FGF7-mediated EMT, emphasizing the dependence of FGF7-mediated EMT on HIF-1α activation. These findings suggest that targeting the FGF7/HIF-1α/EMT axis may offer new therapeutic opportunities to intervene in OC progression.

ZSWIM4 inhibition improves chemosensitivity in epithelial ovarian cancer cells by suppressing intracellular glycine biosynthesis

Abstract Background Zinc finger SWIM-type containing 4 (ZSWIM4) induces drug resistance in breast cancer cells. However, its role in epithelial ovarian cancer (EOC) remains unknown. In this study, we aimed to investigate the clinical significance of ZSWIM4 expression in EOC and develop new clinical therapeutic strategies for EOC. Methods ZSWIM4 expression in control and EOC tumor tissues was examined using immunohistochemistry. Lentiviral transduction, Cell Counting Kit-8 assay, tumorsphere formation assay, flow cytometry, western blotting, and animal xenograft model were used to assess the role of ZSWIM4 in chemotherapy. Cleavage Under Targets and Tagmentation (CUT&Tag) assays, chromatin immunoprecipitation assays, and luciferase reporter assays were used to confirm FOXK1-mediated upregulation of ZSWIM4 expression. The mechanism by which ZSWIM4 inhibition improves chemosensitivity was evaluated using RNA-sequencing. A ZSWIM4-targeting inhibitor was explored by virtual screening and surface plasmon resonance analysis. Patient-derived organoid (PDO) models were constructed from EOC tumor tissues with ZSWIM4 expression. Results ZSWIM4 was overexpressed in EOC tumor tissues and impaired patient prognoses. Its expression correlated positively with EOC recurrence. ZSWIM4 expression was upregulated following carboplatin treatment, which, in turn, contributed to chemoresistance. Silencing ZSWIM4 expression sensitized EOC cells to carboplatin treatment in vitro and in vivo. FOXK1 could bind to the GTAAACA sequence of the ZSWIM4 promoter region to upregulate ZSWIM4 transcriptional activity and FOXK1 expression increased following carboplatin treatment, leading to an increase in ZSWIM4 expression. Mechanistically, ZSWIM4 knockdown downregulated the expression of several rate-limiting enzymes involved in glycine synthesis, causing a decrease in intracellular glycine levels, thus enhancing intracellular reactive oxygen species production induced by carboplatin treatment. Compound IPN60090 directly bound to ZSWIM4 protein and exerted a significant chemosensitizing effect in both EOC cells and PDO models. Conclusions ZSWIM4 inhibition enhanced EOC cell chemosensitivity by ameliorating intracellular glycine metabolism reprogramming, thus providing a new potential therapeutic strategy for EOC.

m6A methylation-mediated regulation of LncRNA MEG3 suppresses ovarian cancer progression through miR-885-5p and the VASH1 pathway

Abstract Background Ovarian cancer poses a serious threat to women's health. Due to the difficulty of early detection, most patients are diagnosed with advanced-stage disease or peritoneal metastasis. We found that LncRNA MEG3 is a novel tumor suppressor, but its role in tumor occurrence and development is still unclear. Methods We investigated the expression level of MEG3 in pan-cancer through bioinformatics analysis, especially in gynecological tumors. Function assays were used to detect the effect of MEG3 on the malignant phenotype of ovarian cancer. RIP, RNA pull-down, MeRIP-qPCR, actinomycin D test were carried out to explore the m6A methylation-mediated regulation on MEG3. Luciferase reporter gene assay, PCR and Western blot were implemented to reveal the potential mechanism of MEG3. We further confirmed the influence of MEG3 on tumor growth in vivo by orthotopic xenograft models and IHC assay. Results In this study, we discovered that MEG3 was downregulated in various cancers, with the most apparent downregulation in ovarian cancer. MEG3 inhibited the proliferation, migration, and invasion of ovarian cancer cells. Overexpression of MEG3 suppressed the degradation of VASH1 by negatively regulating miR-885-5p, inhibiting the ovarian cancer malignant phenotype. Furthermore, we demonstrated that MEG3 was regulated at the posttranscriptional level. YTHDF2 facilitated MEG3 decay by recognizing METTL3‑mediated m6A modification. Compared with those injected with vector control cells, mice injected with MEG3 knockdown cells showed larger tumor volumes and faster growth rates. Conclusion We demonstrated that MEG3 is influenced by METTL3/YTHDF2 methylation and restrains ovarian cancer proliferation and metastasis by binding miR-885-5p to increase VASH1 expression. MEG3 is expected to become a therapeutic target for ovarian cancer.

Circulating small extracellular vesicles microRNAs plus CA-125 for treatment stratification in advanced ovarian cancer

Abstract Background No residual disease (R0 resection) after debulking surgery is the most critical independent prognostic factor for advanced ovarian cancer (AOC). There is an unmet clinical need for selecting primary or interval debulking surgery in AOC patients using existing prediction models. Methods RNA sequencing of circulating small extracellular vesicles (sEVs) was used to discover the differential expression microRNAs (DEMs) profile between any residual disease (R0, n = 17) and no residual disease (non-R0, n = 20) in AOC patients. We further analyzed plasma samples of AOC patients collected before surgery or neoadjuvant chemotherapy via TaqMan qRT-PCR. The combined risk model of residual disease was developed by logistic regression analysis based on the discovery-validation sets. Results Using a comprehensive plasma small extracellular vesicles (sEVs) microRNAs (miRNAs) profile in AOC, we identified and optimized a risk prediction model consisting of plasma sEVs-derived 4-miRNA and CA-125 with better performance in predicting R0 resection. Based on 360 clinical human samples, this model was constructed using least absolute shrinkage and selection operator (LASSO) and logistic regression analysis, and it has favorable calibration and discrimination ability (AUC:0.903; sensitivity:0.897; specificity:0.910; PPV:0.926; NPV:0.871). The quantitative evaluation of Net Reclassification Improvement (NRI) and Integrated Discrimination Improvement (IDI) suggested that the additional predictive power of the combined model was significantly improved contrasted with CA-125 or 4-miRNA alone (NRI = 0.471, IDI = 0.538, p < 0.001; NRI = 0.122, IDI = 0.185, p < 0.01). Conclusion Overall, we established a reliable, non-invasive, and objective detection method composed of circulating tumor-derived sEVs 4-miRNA plus CA-125 to preoperatively anticipate the high-risk AOC patients of residual disease to optimize clinical therapy.

RUNX1 knockdown induced apoptosis and impaired EMT in high-grade serous ovarian cancer cells

AbstractOvarian cancer is the leading cause of death from gynecologic illnesses worldwide. High-grade serous ovarian cancer (HGSOC) is a gynecological tumor that accounts for roughly 70% of ovarian cancer deaths in women. Runt-related transcription factor 1(RUNX1) proteins were identified with overexpression in the HGSOC. However, the roles of RUNX1 in the development of HGSOC are poorly understood. In this study, combined with whole-transcriptome analysis and multiple research methods, RUNX1 was identified as vital in developing HGSOC. RUNX1 knockdown inhibits the physiological function of ovarian cancer cells and regulates apoptosis through the FOXO1-Bcl2 axis. Down-regulated RUNX1 impairs EMT function through the EGFR-AKT-STAT3 axis signaling. In addition, RUNX1 knockdown can significantly increase the sensitivity to clinical drug therapy for ovarian cancer. It is strongly suggested that RUNX1 work as a potential diagnostic and therapeutic target for HGSOC patients with better prognoses and treatment options. It is possible to generate novel potential targeted therapy strategies and translational applications for serous ovarian carcinoma patients with better clinical outcomes.

Intratumoral Collinsella aerofaciens exhibits antitumor activity in endometrial carcinoma through activation of the p53 signaling pathway

The intratumoral microbiota plays dual roles in cancer progression and suppression, but its composition and functional mechanisms in endometrial carcinoma (EC) remain incompletely defined. This study aimed to characterize the EC intratumoral microbiome, elucidate microbial spatial localization, and identify bacteria with tumor-suppressive properties. Tumor and adjacent normal tissues from patients with EC were analyzed using 5R 16S rRNA sequencing to profile microbial communities, with fluorescence in situ hybridization (FISH) validating bacterial localization. Spatial transcriptomics (ST), single-cell RNA sequencing (scRNA-seq), and FISH were integrated to map microbiota-niche cell interactions. RNA sequencing was performed on EC cells treated with bacterial supernatant. Fecal microbiota transplantation (FMT) from EC patients to mice was used to assess gut-tumor microbial crosstalk. Collinsella aerofaciens (C. aerofaciens), Haloamaerobium gallinarum, and Massilia oculi were enriched in adjacent normal tissues, while Bacteroides vulgatus (B. vulgatus) and Delfia tsuruhatensis dominated tumor tissues. Tumors exhibited reduced microbial richness versus normal tissues. C. aerofaciens localized predominantly to smooth muscle cells and modulated the tumor microenvironment, as revealed by FISH and ST-scRNA-seq integration. RNA sequencing suggested that C. aerofaciens suppressed EC progression by activating the p53 signaling pathway. FMT experiments demonstrated gut microbiota-driven remodeling of the tumor microbiome. This study identifies C. aerofaciens as a novel tumor-suppressive bacterium in EC, with mechanistic evidence linking its activity to p53 pathway activation. Gut microbiota modulates intratumoral microbial composition, suggesting potential dual-target therapeutic strategies for EC.

C/D box small nucleolar RNA SNORD104 promotes endometrial cancer by regulating the 2ʹ-O-methylation of PARP1

Abstract Background Small nucleolar RNAs (snoRNAs) are dysregulated in many cancers, although their exact role in tumor genesis and progression remains unclear. Methods The expression profiles of snoRNAs in endometrial cancer (EC) tissues were analyzed using data from The Cancer Genome Atlas, and SNORD104 was identified as an upregulated snoRNA in EC. The tumorigenic role of SNORD104 in EC was established in CCK8, colony formation, EdU, apoptosis, Transwell, and in vivo xenograft experiments. The molecular mechanisms of SNORD104 were analyzed by RNA immunoprecipitation (RIP), Nm-seq, RTL-P assay, RNA stability assay, qRT-PCR, and western blotting. Results Antisense oligonucleotide (ASO)-mediated knockdown of SNORD104 in Ishikawa cells significantly inhibited their proliferation, colony formation ability, migration, and invasion in vitro and increased apoptosis. On the other hand, overexpression of SNORD104 promoted EC growth in vivo and in vitro. RIP assay showed that SNORD104 binds to the 2ʹ-O-methyltransferase fibrillarin (FBL), and according to the results of Nm-seq and RTL-P assay, SNORD104 upregulated PARP1 (encoding poly (ADP-ribose) polymerase 1) 2ʹ-O-methylation. The binding of FBL to PARP1 mRNA was also verified by RIP assay. Furthermore, SNORD104 expression was positively correlated with PARP1 expression in EC tissues. In the presence of actinomycin D, SNORD104 increased the stability of PARP1 mRNA and promoted its nuclear localization. Finally, silencing FBL or PARP1 in the HEC1B cells overexpressing SNORD104 inhibited their proliferative and clonal capacities and increased apoptosis rates. Conclusions SNORD104 enhances PARP1 mRNA stability and translation in the EC cells by upregulating 2ʹ-O-methylation and promotes tumor growth.

Identification and validation of the important role of KIF11 in the development and progression of endometrial cancer

Human kinesin family member 11 (KIF11) plays a vital role in regulating the cell cycle and is implicated in the tumorigenesis and progression of various cancers, but its role in endometrial cancer (EC) is still unclear. Our current research explored the prognostic value, biological function and targeting strategy of KIF11 in EC through approaches including bioinformatics, machine learning and experimental studies. The GSE17025 dataset from the GEO database was analyzed via the limma package to identify differentially expressed genes (DEGs) in EC. Functional enrichment analysis of the DEGs was conducted using Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analyses. DEGs were further screened for hub genes through protein-protein interaction (PPI) network analysis and machine learning. The role of the hub gene KIF11 in EC was analyzed using clinical data from the TCGA database. The expression of KIF11 in EC was subsequently validated in clinical samples. In vitro experiments were utilized to evaluate the effects of KIF11 on biological functions such as proliferation, migration, apoptosis, and the cell cycle in endometrial cancer cells. A total of 877 DEGs, which are widely involved in important biological processes such as cell division, tubulin binding, and the cell cycle, were identified. Through PPI network analysis and machine learning, KIF11 was selected as the hub gene for subsequent analysis and experimental validation. An analysis of TCGA data revealed that KIF11 is highly expressed in EC and is associated with tumor grade, stage, and a low survival rate. The overexpression of KIF11 in tumor tissues was further confirmed in EC patient samples. KIF11 knockdown had inhibitory effects on cell proliferation, migration and invasion. Flow cytometry analysis revealed that KIF11 knockdown induced G2/M phase arrest and promoted apoptosis in EC cells. Our study demonstrated that KIF11 was upregulated in EC and was strongly associated with a poor prognosis. Notably, we found that reduced KIF11 expression inhibited EC cell proliferation, migration and invasion. KIF11 knockdown caused more EC cells to arrest in the G2/M phase and undergo apoptosis. The findings of our study emphasized that KIF11 may be a promising prognostic biomarker and therapeutic target for EC patients.

The OncoLifeS data-biobank for oncology: a comprehensive repository of clinical data, biological samples, and the patient's perspective.

Understanding cancer heterogeneity, its temporal evolution over time, and the outcomes of guided treatment depend on accurate data collection in a context of routine clinical care. We have developed a hospital-based data-biobank for oncology, entitled OncoLifeS (Oncological Life Study: Living well as a cancer survivor), that links routine clinical data with preserved biological specimens and quality of life assessments. The aim of this study is to describe the organization and development of a data-biobank for cancer research. We have enrolled 3704 patients aged ≥ 18 years diagnosed with cancer, of which 45 with hereditary breast-ovarian cancer (70% participation rate) as of October 24th, 2019. The average age is 63.6 ± 14.2 years and 1892 (51.1%) are female. The following data are collected: clinical and treatment details, comorbidities, lifestyle, radiological and pathological findings, and long-term outcomes. We also collect and store various biomaterials of patients as well as information from quality of life assessments. Embedding a data-biobank in clinical care can ensure the collection of high-quality data. Moreover, the inclusion of longitudinal quality of life data allows us to incorporate patients' perspectives and inclusion of imaging data provides an opportunity for analyzing raw imaging data using artificial intelligence (AI) methods, thus adding new dimensions to the collected data.

TEFM facilitates uterine corpus endometrial carcinoma progression by activating ROS-NFκB pathway

Mitochondrial transcription elongation factor (TEFM) is a recently discovered factor involved in mitochondrial DNA replication and transcription. Previous studies have reported that abnormal TEFM expression can disrupt the assembly of mitochondrial respiratory chain and thus mitochondrial function. However, the role of TEFM on Uterine corpus endometrial carcinoma (UCEC) progression remains unclear. The present study aims to investigate the expression of TEFM in tumor tissue of UCEC and the effect of abnormal TEFM expression on malignant phenotype of UCEC cells. The expressions of TEFM were measured in tumor tissues and cell lines of UCEC by immunohistochemistry, Western blotting, and real-time quantitative PCR assays. Besides, the effects of TEFM knockdown or overexpression on UCEC cell growth, metastasis, apoptosis, and autophagy were also determined using EdU, colony formation, flow cytometry, TUNEL, and transmission electron microscopy assays. Xenograft model was used to confirm the role of TEFM on proliferative potential of UECE cells in vivo. Our bioinformatics analysis of CPTAC data showed that TEFM is abnormally overexpressed in UCEC and its upregulation was significantly associated with poor survival of patients with UCEC. We found that TEFM upregulation significantly promoted the growth and metastasis of UCEC cells. Mechanically, TEFM upregulation impaired the function of mitochondria, decreased their membrane potential and activated the AKT-NFκB pathway by promoting reactive oxygen species (ROS) production, leading to enhanced intracellular autophagy and thus UCEC growth and metastasis. This study demonstrates that TEFM positively regulates autophagy to promote the growth and metastasis of UCEC cells, which provides a potential prognostic biomarker and therapeutic target for the treatment of UCEC.

An integrated approach of network pharmacology, molecular docking, and experimental verification uncovers kaempferol as the effective modulator of HSD17B1 for treatment of endometrial cancer

Abstract Background Endometrial cancer (EC) is one of the most common gynecological malignancies globally, and the development of innovative, effective drugs against EC remains a key issue. Phytoestrogen kaempferol exhibits anti-cancer effects, but the action mechanisms are still unclear. Method MTT assays, colony-forming assays, flow cytometry, scratch healing, and transwell assays were used to evaluate the proliferation, apoptosis, cell cycle, migration, and invasion of both ER-subtype EC cells. Xenograft experiments were used to assess the effects of kaempferol inhibition on tumor growth. Next-generation RNA sequencing was used to compare the gene expression levels in vehicle-treated versus kaempferol-treated Ishikawa and HEC-1-A cells. A network pharmacology and molecular docking technique were applied to identify the anti-cancer mechanism of kaempferol, including the building of target-pathway network. GO analysis and KEGG pathway enrichment analysis were used to identify cancer-related targets. Finally, the study validated the mRNA and protein expression using real-time quantitative PCR, western blotting, and immunohistochemical analysis. Results Kaempferol was found to suppress the proliferation, promote apoptosis, and limit the tumor-forming, scratch healing, invasion, and migration capacities of EC cells. Kaempferol inhibited tumor growth and promotes apoptosis in a human endometrial cancer xenograft mouse model. No significant toxicity of kaempferol was found in human monocytes and normal cell lines at non-cytotoxic concentrations. No adverse effects or significant changes in body weight or organ coefficients were observed in 3–7 weeks’ kaempferol-treated animals. The RNA sequencing, network pharmacology, and molecular docking approaches identified the overall survival-related differentially expressed gene HSD17B1. Interestingly, kaempferol upregulated HSD17B1 expression and sensitivity in ER-negative EC cells. Kaempferol differentially regulated PPARG expression in EC cells of different ER subtypes, independent of its effect on ESR1. HSD17B1 and HSD17B1-associated genes, such as ESR1, ESRRA, PPARG, AKT1, and AKR1C1\2\3, were involved in several estrogen metabolism pathways, such as steroid binding, 17-beta-hydroxysteroid dehydrogenase (NADP+) activity, steroid hormone biosynthesis, and regulation of hormone levels. The molecular basis of the effects of kaempferol treatment was evaluated. Conclusions Kaempferol is a novel therapeutic candidate for EC via HSD17B1-related estrogen metabolism pathways. These results provide new insights into the efficiency of the medical translation of phytoestrogens.

Circulating tumor DNA as a prognostic marker in high-risk endometrial cancer

Abstract Background Currently, there is no reliable blood-based marker to track tumor recurrence in endometrial cancer (EC) patients. Liquid biopsies, specifically, circulating tumor DNA (ctDNA) analysis emerged as a way to monitor tumor metastasis. The objective of this study was to examine the feasibility of ctDNA in recurrence surveillance and prognostic evaluation of high-risk EC. Methods Tumor tissues from nine high-risk EC patients were collected during primary surgery and tumor DNA was subjected to next generation sequencing to obtain the initial mutation spectrum using a 78 cancer-associated gene panel. Baseline and serial post-operative plasma samples were collected and droplet digital PCR (ddPCR) assays for patient-specific mutations were developed to track the mutations in the ctDNA in serial plasma samples. Log-rank test was used to assess the association between detection of ctDNA before or after surgery and disease-free survival. Results Somatic mutations were identified in all of the cases. The most frequent mutated genes were PTEN , FAT4 , ARID1A , TP53 , ZFHX3 , ATM , and FBXW7 . For each patient, personalized ddPCR assays were designed for one-to-three high-frequent mutations. DdPCR analysis and tumor panel sequencing had a high level of agreement in the assessment of the mutant allele fractions in baseline tumor tissue DNA. CtDNA was detected in 67% (6 of 9) of baseline plasma samples, which was not predictive of disease-free survival (DFS). CtDNA was detected in serial post-operative plasma samples (ctDNA tracking) of 44% (4 of 9) of the patients, which predicted tumor relapse. The DFS was a median of 9 months (ctDNA detected) versus median DFS undefined (ctDNA not detected), with a hazard ratio of 17.43 (95% CI, 1.616–188.3). The sensitivity of post-operative ctDNA detection in estimating tumor relapse was 100% and specificity was 83.3%, which was superior to CA125 or HE4. Conclusions Personalized ctDNA detection was effective and stable for high-risk EC. CtDNA tracking in post-operative plasma is valuable for predicting tumor recurrence.

Glucose metabolic reprogramming and its therapeutic potential in obesity-associated endometrial cancer

AbstractEndometrial cancer (EC) is a common gynecological cancer that endangers women health. Although substantial progresses of EC management have been achieved in recent years, the incidence of EC still remains high. Obesity has been a common phenomenon worldwide that increases the risk of EC. However, the mechanism associating obesity and EC has not been fully understood. Metabolic reprogramming as a remarkable characteristic of EC is currently emerging. As the primary factor of metabolic syndrome, obesity promotes insulin resistance, hyperinsulinemia and hyperglycaemia. This metabolic disorder remodels systemic status, which increases EC risk and is related with poor prognosis. Glucose metabolism in EC cells is complex and mediated by glycolysis and mitochondria to ensure energy requirement. Factors that affect glucose metabolism may have an impact on EC initiation and progression. In this study, we review the glucose metabolic reprogramming of EC not only systemic metabolism but also inherent tumor cell metabolism. In particular, the role of glucose metabolic regulation in malignant properties of EC will be focused. Understanding of metabolic profile and glucose metabolism-associated regulation mechanism in EC may provide novel perspective for treatment.

Integrated multi-omics analyses and functional validation reveal TTK as a novel EMT activator for endometrial cancer

Abstract Background Cancer-testis antigens (CTAs) are often expressed in tumor and testicular tissues but not in other normal tissues. To date, there has been no comprehensive study of the expression and clinical significance of CTA genes associated with endometrial cancer (EC) development. Additionally, the clinical relevance, biological role, and molecular mechanisms of the CTA gene TTK protein kinase (TTK) in EC are yet to be fully understood. Methods Using bioinformatics methods, we comprehensively investigated the genomic, transcriptomic, and epigenetic changes associated with aberrant TTK overexpression in EC samples from the TCGA database. We further investigated the mechanisms of the lower survival associated with TTK dysregulation using single-cell data of EC samples from the GEO database. Cell functional assays were used to confirm the biological roles of TTK in EC cells. Results We identified 80 CTA genes that were more abundant in EC than in normal tissues, and high expression of TTK was significantly linked with lower survival in EC patients. Furthermore, ROC analysis revealed that TTK could accurately distinguish stage I EC tissues from benign endometrial samples, suggesting that TTK has the potential to be a biomarker for early EC detection. We found TTK overexpression was more prevalent in EC patients with high-grade, advanced tumors, serous carcinoma, and TP53 alterations. Furthermore, in EC tissue, TTK expression showed a strong positive correlation with EMT-related genes. With single-cell transcriptome data, we identified a proliferative cell subpopulation with high expression of TTK and known epithelial–mesenchymal transition (EMT)-related genes and transcription factors. When proliferative cells were grouped according to TTK expression levels, the overexpressed genes in the TTKhigh group were shown to be functionally involved in the control of chemoresistance. Utilizing shRNA to repress TTK expression in EC cells resulted in substantial decreases in cell proliferation, invasion, EMT, and chemoresistance. Further research identified microRNA-21 (miR-21) as a key downstream regulator of TTK-induced EMT and chemoresistance. Finally, the TTK inhibitor AZ3146 was effective in reducing EC cell growth and invasion and enhancing the apoptosis of EC cells generated by paclitaxel. Conclusion Our findings establish the clinical significance of TTK as a new biomarker for EC and an as-yet-unknown carcinogenic function. This present study proposes that the therapeutic targeting of TTK might provide a viable approach for the treatment of EC.

The BRD4 inhibitor JQ1 suppresses tumor growth by reducing c-Myc expression in endometrial cancer

Abstract Background Endometrial cancer (EC) is the most common gynecological malignancy in developed countries. Efficacy of the bromodomain 4 (BRD4) inhibitor JQ1 has been reported for the treatment of various human cancers, but its potential impact on EC remains unclear. We therefore aimed to elucidate the function of BRD4 and the effects of JQ1 in EC in vivo and in vitro. Methods The mRNA expression of BRD4 was evaluated using datasets from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO). BRD4 protein expression in EC tissues was measured using immunohistochemistry (IHC) assays. The effects of JQ1 on EC were determined by using MTT and colony formation assays, flow cytometry and xenograft mouse models. The underlying mechanism was also examined by western blot and small interfering RNA (siRNA) transfection. Results BRD4 was overexpressed in EC tissues, and the level of BRD4 expression was strongly related to poor prognosis. The BRD4-specific inhibitor JQ1 suppressed cell proliferation and colony formation and triggered cell apoptosis, cell cycle arrest, and changes in the expression of proteins in related signaling pathways. Moreover, JQ1 decreased the protein expression of BRD4 and c-Myc, and knockdown of BRD4 or c-Myc reduced the viability of EC cells. Intraperitoneal administration of JQ1 (50 mg/kg) significantly suppressed the tumorigenicity of EC cells in a xenograft mouse model. Conclusion Our results demonstrate that BRD4 is a potential marker of EC and that the BRD4 inhibitor JQ1 is a promising chemotherapeutic agent for the treatment of EC.

CircESRP1 enhances metastasis and epithelial–mesenchymal transition in endometrial cancer via the miR-874-3p/CPEB4 axis

AbstractBackgroundMetastasis is critical for endometrial cancer (EC) progression and prognosis. Accumulating evidence suggests that circular RNAs (circRNAs) can operate as independent functional entities. However, the functional regulatory mechanisms of circRNAs in EC remain unclear.MethodsThe levels of circESRP1, miR-874-3p, and CPEB4 mRNA in EC tissues and cells were determined by qRT-PCR. Sanger sequencing, PCR with divergent primers, an actinomycin D assay, and RNase R treatment were applied to verify the circular properties. Fluorescence in situ hybridization (FISH) and nuclear-cytoplasmic fractionation were used to determine the localization of circESRP1. CCK-8, EdU incorporation, colony formation, Transwell, and wound healing assays were applied to assess the effects of circESRP1 on cell proliferation, migration, and invasion. The mutual regulatory mechanism of ceRNAs was investigated using dual-luciferase reporter, RNA pulldown, RNA immunoprecipitation (RIP), and Western blot assays. The biological effects were further validated in vivo in nude mouse xenograft models.ResultscircESRP1 was highly expressed in EC tissues and cells and was mainly localized in the cytoplasm. Silencing circESRP1 inhibited the proliferation, migration, and invasion of EC cells in vitro and in vivo; however, overexpression of circESRP1 had the opposite effects. Mechanistically, circESRP1 sponged miR-874-3p to upregulate CPEB4 expression and ultimately contribute to EC cell proliferation and metastasis. Furthermore, circESRP1 regulated tumour growth in xenograft models.ConclusionsCircESRP1 can interact with miR-874-3p to regulate EMT in endometrial cancer via the miR-874-3p/CPEB4 axis. CircESRP1 may serve as a promising therapeutic target for endometrial cancer.

Feasibility and utility of MRI and dynamic 18F-FDG-PET in an orthotopic organoid-based patient-derived mouse model of endometrial cancer

Abstract Background Pelvic magnetic resonance imaging (MRI) and whole-body positron emission tomography-computed tomography (PET-CT) play an important role at primary diagnostic work-up and in detecting recurrent disease in endometrial cancer (EC) patients, however the preclinical use of these imaging methods is currently limited. We demonstrate the feasibility and utility of MRI and dynamic 18F-fluorodeoxyglucose (FDG)-PET imaging for monitoring tumor progression and assessing chemotherapy response in an orthotopic organoid-based patient-derived xenograft (O-PDX) mouse model of EC. Methods 18 O-PDX mice (grade 3 endometrioid EC, stage IIIC1), selectively underwent weekly T2-weighted MRI (total scans = 32), diffusion-weighted MRI (DWI) (total scans = 9) and dynamic 18F-FDG-PET (total scans = 26) during tumor progression. MRI tumor volumes (vMRI), tumor apparent diffusion coefficient values (ADCmean) and metabolic tumor parameters from 18F-FDG-PET including maximum and mean standard uptake values (SUVmax/SUVmean), metabolic tumor volume (MTV), total lesion glycolysis (TLG) and metabolic rate of 18F-FDG (MRFDG) were calculated. Further, nine mice were included in a chemotherapy treatment study (treatment; n = 5, controls; n = 4) and tumor ADCmean-values were compared to changes in vMRI and cellular density from histology at endpoint. A Mann–Whitney test was used to evaluate differences between groups. Results Tumors with large tumor volumes (vMRI) had higher metabolic activity (MTV and TLG) in a clear linear relationship (r2 = 0.92 and 0.89, respectively). Non-invasive calculation of MRFDG from dynamic 18F-FDG-PET (mean MRFDG = 0.39 μmol/min) was feasible using an image-derived input function. Treated mice had higher tumor ADCmean (p = 0.03), lower vMRI (p = 0.03) and tumor cellular density (p = 0.02) than non-treated mice, all indicating treatment response. Conclusion Preclinical imaging mirroring clinical imaging methods in EC is highly feasible for monitoring tumor progression and treatment response in the present orthotopic organoid mouse model.

Novel triple-target panels utilizing methylation-sensitive restriction enzyme-based quantitative PCR for detecting advanced cervical precancers and cancers among high-risk HPV-positive women

Optimal triage option for high-risk HPV-positive (hrHPV) women remains uncertain. We aimed to utilize methylation-sensitive restriction enzyme-based quantitative PCR (MSRE-qPCR) technique and develop triple-target human gene methylation panels to improve detection of advanced cervical precancers and cancers (CIN3 +) among hrHPV women. Eighteen candidate genes were detected by MSRE-qPCR in cervical samples from hrHPV women. All possible triple-target panels from these genes were generated by logistic regression models with repeated ten-fold cross-validation on a training set of 1223 women (180 CIN3 +; 1043 < CIN3). Panels with the top two AUCs for CIN3 + on a validation set of 937 women (69 CIN3 +; 868 < CIN3) were ultimately selected for qPCR reconstruction, retesting, and retraining. Triage performance, screening efficiency and risk stratification of the selected panels were then compared with traditional triage strategies (cytology [ASCUS as the threshold, atypical squamous cells of undetermined significance], HPV16/18 genotyping, HPV16/18 genotyping combined with cytology [ASCUS]) within the validation set. Two panels were finally identified and validated for CIN3 + detection. Panel 1 includes JAM3, PCDHGB7, and SORCS1; while Panel 2 consists of PAX1, ZNF671, and ASCL1. Compared to traditional triage strategies, both panels demonstrated superior AUCs (Panel 1: 0.799; Panel 2: 0.790; Cytology: 0.532; HPV16/18 genotyping: 0.589; HPV16/18 + Cytology: 0.515; all P Two triple-target human gene methylation panels were successfully developed, each integrated into a single MSRE-qPCR system for one-tube detection. Both panels outperformed current triage strategies, indicating their potential as alternatives, though external validation among diverse settings is needed before clinical application.

Associations among HPV persistence, the vaginal microbiome, and cervical cancer recurrence

In the US, up to 60% of cervical cancer (CxCa) survivors will have persistent HPV infection, the causative agent of CxCa, and up to 35% will develop recurrent local CxCa within 4 years after chemo-radiation therapy. Preliminary studies suggest healthy vaginal microbiome (VM) could affect the acquisition, persistence, and clearance of HPV. Through longitudinal studies, we investigate associations between the dynamics of VM, HPV persistence, cancer recurrence (CR), and outcomes in gynecologic cancer survivors who completed cancer treatments. We enrolled 49 patients with Stage IB-IIIC CxCa who completed radiation therapy (RT) alone or concurrent chemoradiation (chemoRT). VM sequencing and HPV typing were performed on samples obtained at baseline (T0, pre-treatment), T1, T2, and T3 (3, 6, and 12 months after the completion of cancer treatment). Patients were evaluated for CR up to 4 years following the end of treatment. Among all patients, 33% (16/49) had CxCa recurrence within 2-3 years post-therapy. The vaginal microbiome exhibited high diversity, Prevotella-dominant communities; only 20% were dominated by lactobacilli at any time. Post-treatment hrHPV was detected in 17 out of 41 women (41.5%) with follow-up samples. We identified key taxa, such as Prevotella species, which were highly associated with CxCa recurrence and post-treatment detection of hrHPV. Our findings link Prevotella-dominant, high diversity vaginal microbiome communities with post-therapy hrHPV persistence and cervical cancer recurrence in gynecologic cancer survivors. Such findings warrant further research into the role of the microbiome in modulating cervical cancer progression and response to therapy, suggesting modulation of the microbiome with probiotics or other methods could be considered a novel approach to improve cervical cancer treatment outcomes.

Endometrial cancer: mapping the global landscape of research

Abstract Background From a global viewpoint, endometrial cancer belongs to the most common female cancers. Despite the heavy burden of diseases and numerous unanswered questions, no detailed pictures of the global structure of endometrial cancer research are available so far. Therefore, this malignancy was reviewed using the New Quality and Quantity Indices in Science (NewQIS) protocol. Methods Using NewQIS, we identified endometrial carcinoma related research published in the Web of Science from 1900–2015 (P1) and from 2016–2020 (P2). Item analysis was performed with regard to research activity. Also, semi-qualitative aspects and socio-economic benchmarks were visualized using density equalizing mapping. Results In total, 9,141 from 1900–2015 and 4,593 from 2016–2020 endometrial cancer related studies were identified with the USA having the largest numbers of publications, citations, institutions, as well as the highest country-specific h-Index concerning endometrial cancer research in both periods. In contrast to other fields of cancer research, the two East Asian countries Japan and China followed concerning total research activities until 2015. From 2016 until 2020, China was found in short distance to the USA and was ranked second. In the socio-economic analysis, European countries were in prominent positions. Greece published 579.83 endometrial carcinoma-related articles per billion US-$ GDP, Finland (527.29), Sweden (494.65), Israel (493.75), and Norway (367.85) followed in the ranking. Density equalizing mapping visualized that large parts of Africa, Asia and South America with a high burden of disease played almost no visible role in the endometrial cancer research activities. Conclusions Endometrial cancer research activity is continuously increasing from a global viewpoint. However, the majority of original articles is published by authors based in high-income countries. Together with the finding that the research field of public health does only play a minimal role, our study points to the necessity that global health aspects should be introduced to endometrial cancer research.

Integrated single-cell, spatial, and bulk transcriptomics reveal a chromatin regulator-TME prognostic framework guiding precision therapy in cervical cancer

Chromatin regulators (CRs) play a critical role in tumorigenesis, drug response, and prognosis, with dysregulation of chromatin regulator genes (CRGs) potentially disrupting the tumor immune microenvironment (TME) and influencing immune responses in cervical cancer. However, the prognostic and therapeutic implications of integrating CRGs and TME parameters in cervical cancer remain poorly understood. This study used prognostic CRGs and TME cell signatures identified through Cox regression and Kaplan-Meier survival analyses to construct the CRG and TME score. The CRG score for each cell was calculated by integrating single-cell and spatial transcriptome, which were subsequently integrated to develop a CRG-TME classifier for prognostic prediction. The prognosis, somatic mutations, immune characteristics, therapeutic benefits, and drug sensitivity were then analyzed across CRG-TME defined subgroups. Both CRG score and TME score demonstrated prognostic value, and the high CRG score group showed distinct alterations in TME signaling dynamics and increased intensity of intercellular communication. Patients in CRG This study develops a novel CRG-TME classifier to predict prognosis and therapy response in cervical cancer and unveils the inhibitory effect and underlying mechanism of TSPYL2 on cervical cancer progression.

A novel bispecific affitoxin simultaneously targeting E7 of HPV16/18 types: superior anti-tumor activity and EMT reversal in HPV-driven cervical cancer therapy

Sustained infection with high-risk HPV of the 16 and 18 types is accounted for nearly 75% of cervical cancer (CC), but now there is an absence of agents aimed at eradicating HPV infections. Notwithstanding, affibody-based affitoxins may represent a breakthrough in tumor-targeted therapy. Our previous work has constructed a bispecific affibody simultaneously targeting the early oncogenic proteins E7 of HPV16 and 18 types (named as Z Three forms of the affitoxin constructs (GrB-Z Two bispecific affitoxins of Z This work has developed a bispecific affitoxin that simultaneously targets HPV16- and HPV18-type CC cells, with a significant dual-functional advantage, combining the targeted inhibitory of the affibody with the cytotoxicity of the toxin molecule. Our research offers a novel design and approach for targeted therapy in HPV-driven cervical cancer.

Spatial transcriptomics reveals unique metabolic profile and key oncogenic regulators of cervical squamous cell carcinoma

As a prevalent and deadly malignant tumor, the treatment outcomes for late-stage patients with cervical squamous cell carcinoma (CSCC) are often suboptimal. Previous studies have shown that tumor progression is closely related with tumor metabolism and microenvironment reshaping, with disruptions in energy metabolism playing a critical role in this process. To delve deeper into the understanding of CSCC development, our research focused on analyzing the tumor microenvironment and metabolic characteristics across different regions of tumor tissue. Utilizing spatial transcriptomics (ST) sequencing technology, we conducted a study on FFPE (formalin-fixed paraffin-embedded) tumor samples from CSCC patients. Coupled with single-cell RNA sequencing (scRNA-seq) data after deconvolution, we described spatial distribution maps of tumor leading edge and core regions in detail. Tumor tissues were classified into hypermetabolic and hypometabolic regions to analyze the metabolism profiles and tumor differentiation degree across different spatial areas. We also employed The Cancer Genome Atlas (TCGA) database to examine the analysis results of ST data. Our findings indicated a more complex tumor microenvironment in hypermetabolic regions. Cell-cell communication analysis showed that various cells in tumor microenvironment were influenced by the signalling molecule APP released by cancer cells and higher expression of APP was observed in hypermetabolic regions. Furthermore, our results revealed the correlation between APP and the transcription factor TRPS1. Both APP and TRPS1 demonstrated significant effects on cancer cell proliferation, migration, and invasion, potentially contributing to tumor progression. Utilizing ST, scRNA-seq, and TCGA database, we examined the spatial metabolic profiles of CSCC tissues, including metabolism distribution, metabolic variations, and the relationship between metabolism and tumor differentiation degree. Additionally, potential cancer-promoting factors were proposed, offering a valuable foundation for the development of more effective treatment strategies for CSCC.

Application of PARP inhibitors combined with immune checkpoint inhibitors in ovarian cancer

The advent of polyadenosine diphosphate ribose polymerase inhibitors (PARPi) has brought about significant changes in the field of ovarian cancer treatment. However, in 2022, Rucaparib, Olaparib, and Niraparib, had their marketing approval revoked for third-line and subsequent therapies due to an increased potential for adverse events. Consequently, the exploration of new treatment modalities remains imperative. Recently, the integration of PARPi with immune checkpoint inhibitors (ICIs) has emerged as a potential remedy option within the context of ovarian cancer. This article offers a comprehensive examination of the mechanisms and applications of PARPi and ICIs in the treatment of ovarian cancer. It synthesizes the existing evidence supporting their combined use and discusses key considerations that merit attention in ongoing development efforts.

SORBS3-β suppresses lymph node metastasis in cervical cancer by promoting the ubiquitination of β-catenin

Cervical cancer (CC) is a prevalent gynecological malignancy, with lymph node metastasis (LNM) serving as a critical factor influencing patient prognosis. SORBS3, an adaptor protein with two known isoforms (α and β), has been implicated in tumor suppression, but the specific roles of its isoforms in CC metastasis remains unexplored. This study aimed to identify the functional isoform of SORBS3 driving LNM suppression and elucidate its mechanisms. Proteomic analysis of clinical CC tissues and metastatic lymph nodes revealed progressive downregulation of SORBS3. The mRNA and protein levels of SORBS3-α and SORBS3-β were subsequently examined in normal cervical epithelial and CC cell lines. Functional studies, including siRNA-mediated knockdown of SORBS3-α, lentiviral-mediated overexpression and knockdown of SORBS3-β, Transwell migration, lymphangiogenesis assays, and in vivo footpad xenograft models, were conducted to evaluate the role of SORBS3 isoforms in LNM. SORBS3 DNA methylation mechanisms were analyzed by MSP and Targeted Bisulfite sequencing. Mechanistic insights were derived from Co-IP, ubiquitination assays, RNA-seq, and LC-MS/MS. Knockdown of SORBS3-α had no effect on CC cell migration, invasion, or lymphangiogenesis. In contrast, SORBS3-β overexpression markedly suppressed CC cell invasion, lymphangiogenesis, and adhesion to lymphatic endothelial cells, whereas its knockdown significantly promoted these phenotypes. Promoter hypermethylation driven by DNMT-1 inhibited SORBS3 expression in CC. SORBS3- β directly binds to β-catenin and recruits UBA1 to enhance its ubiquitination and degradation, thereby inhibiting Wnt/β-catenin signaling. This inhibition reduced accumulation of β-catenin and downregulated the pro-lymphangiogenic gene VEGFC, ultimately suppressing lymphangiogenesis and LNM. In vivo, SORBS3-β overexpression attenuated lymphatic metastasis in nude mice, whereas its knockdown promoted metastasis. SORBS3-β is the major isoform of SORBS3 that inhibits lymphatic metastasis of cervical cancer by degrading β-catenin through UBA1-mediated ubiquitination, blocking Wnt/β-catenin signaling and downstream lymphangiogenesis pathways, thereby inhibiting lymphatic metastasis. Our findings elucidate key molecular mechanisms underlying cervical cancer lymph node metastasis, offering potential therapeutic targets.metastasis.

A gene-based predictive model for lymph node metastasis in cervical cancer: superior performance over imaging techniques

Abstract Objective Lymph node metastasis (LNM) critically impacts the prognosis and treatment decisions of cervical cancer patients. The accuracy and sensitivity of current imaging techniques, such as CT and MRI, are limited in assessing lymph node status. This study aims to develop a more accurate and efficient method for predicting LNM. Methods Three independent cohorts were merged and divided into training and internal validation groups, with our cohort and those from other centers serving as external validation. A predictive model for LNM in cervical cancer was established using the LASSO regression and multivariate logistic regression. The diagnostic performance of the predictive model was compared with that of CT/MRI in terms of accuracy, sensitivity, specificity, and AUC. Results Using RNA-seq data, four independent predictive genes (MAPT, EPB41L1, ACSL5, and PRPF4B) were identified through LASSO regression and multivariate logistic regression, and a predictive model was constructed to calculate the LNM risk score. Compared with CT/MRI, the model demonstrated higher diagnostic efficiency, with an accuracy of 0.840 and sensitivity of 0.804, compared to CT/MRI’s accuracy of 0.713 and sensitivity of 0.587. The predictive model corrected 81% of misdiagnoses by CT/MRI, demonstrating significant improvements in accuracy and sensitivity. Conclusion The predictive model developed in this study, based on gene expression data, significantly improves the preoperative assessment accuracy of LNM in cervical cancer. Compared to traditional imaging techniques, this model shows superior sensitivity and accuracy. This study provides a robust foundation for developing precise diagnostic tools, paving the way for future clinical applications in individualized treatment planning.

Ligand-based adoptive T cell targeting CA125 in ovarian cancer

Abstract Background Ovarian cancer (OC) is a highly aggressive gynecological malignancy prevalent worldwide. Most OC cases are typically diagnosed at advanced stages, which has led to a 5-year overall survival rate of less than 35% following conventional treatment. Furthermore, immune checkpoint inhibitor therapy has shown limited efficacy in the treatment of patients with OC, and CAR-T therapy has also demonstrated modest results owing to inadequate T cell infiltration. Therefore, novel strategies must be developed to enhance T cell persistence and trafficking within the OC tumor microenvironment. Methods In this study, we developed a novel adoptive T-cell therapy for ovarian cancer based on a chimeric antigen receptor structure. We used a ligand-receptor binding motif to enhance the therapeutic effect of targeting CA125. Since mesothelin can naturally bind to CA125 with high affinity, we concatenated the core-binding fragment of mesothelin with the 4-1BB and CD3ζ signal fragments to assemble a novel CA125-targeting chimeric receptor (CR). The CAR structure targeting CA125 derived from the 4H11 antibody was also constructed. CR- and CAR-encoding RNA were electroporated into T cells to evaluate their antitumor activity both in vitro and in vivo. Results While CR-T or CAR-T cells exhibited moderate activity against two ovarian cancer cell lines, T cells co-expressing CR and CAR exhibited a superior killing effect compared to T cells expressing either CR or CAR alone. Furthermore, upon interaction with ovarian tumors, the ability of CR and CAR T cells to release activation markers and functional cytokines increased significantly. Similarly, CR and CAR co-expressing T cells persistently controlled the growth of transplanted ovarian cancer tumors in NSG mice and significantly prolonged the overall survival of tumor-challenged mice. Transcriptome sequencing revealed that the survival and cytotoxicity of T cells co-expressing CR and CAR were significantly altered compared with those of T cells expressing either CR or CAR. Conclusion Our findings demonstrate that CA125 targeting CR and CAR can synergistically kill ovarian cancer cells, indicating that CA125 targeting by the two binding motifs simultaneously in tumors may improve the therapeutic outcomes of ovarian cancer treatment.

PD-L1+ CD49f+ CD133+ Circulating tumor cells predict outcome of patients with vulvar or cervical cancer after radio- and chemoradiotherapy

Abstract Background Monitoring individual therapy responses of patients with cancer represents a major clinical challenge providing the basis to early identify metastases and cancer relapse. We previously demonstrated that radio- or chemoradiotherapy affects the systemic cellular milieu of patients with vulvar or cervical cancer and creates individual post-therapeutic environments associated with cancer relapse. Circulating tumor cells (CTCs) in the systemic milieu are related to metastases and relapse; however, their quantitative and phenotypic characteristics during therapy of patients with vulvar and cervical cancer are still unknown. Methods In this prospective, longitudinal study, we verified the presence of CTCs via immunofluorescence and systemically characterized CTCs by flow cytometry from the blood of 40 patients with vulvar and 115 patients with cervical cancer receiving surgery, adjuvant radiotherapy (aRT), chemoradiotherapy (aCRT) or primary chemoradiotherapy (pCRT) and linked the presence of different CTC subpopulations with individual outcome of disease. Results Pre-therapeutic cytokeratin+ CD45− CTC numbers significantly correlated with tumor FIGO stages, lymph node metastases and relapse. While surgery only did not significantly alter CTC occurrence, aRT and aCRT as well as pCRT differentially decreased or increased CTCs in patients with both tumor entities compared to baseline levels. Therapy-mediated increased CTC numbers were directly linked with subsequent cancer recurrence on follow-up. Phenotypic characterization of CTCs revealed enhanced expression of the stem cell marker CD133 as well as the integrin α6 (CD49f) after aRT, aCRT and pCRT. Furthermore, the aRT, aCRT and pCRT cohorts exhibited increased proportions of Programmed Cell Death Protein Ligand (PD-L1) expressing cells among post-therapeutic CTCs. Notably, post-therapeutic PD-L1+ CD49f+ CD133+ numbers ≥ 5/ml in patients with vulvar cancer and ≥ 2/ml in patients with cervical cancer were associated with reduced recurrence-free survival on follow-up. Conclusion Our study unravels individual therapy-induced changes in CTC phenotypic characteristics and occurrence in the patients’ blood and their association with cancer relapse. Our results may help to explain differences in the individual courses of disease of patients with vulvar and cervical cancer and suggest PD-L1, CD49f and CD133 as targets for immunotherapy in vulvar and cervical cancer.

Pathological and molecular insights into intravenous leiomyomatosis: an integrative multi-omics study

Intravenous leiomyomatosis (IVL) is a histologically well differentiated smooth muscle tumor with aggressive behavior, capable of extending throughout the venous system. Understanding how IVL occurs and develops is really important for diagnosing and treating it. Unfortunately, because IVL is quite rare, there aren't many comprehensive studies available. In our research, we carried out an extensive multi-omics study, gathering tissue samples from IVL cases, uterine fibroid, and normal myometrium. The single-cell RNA sequencing analysis revealed a notable difference in cell composition between IVL and uterine fibroid. Additionally, H&E staining demonstrated more frequent hydropic changes and hyalinization in IVL tissues, along with a reduced vascular density compared to both normal myometrium and uterine fibroid. In our proteomics analysis of eight paired samples of IVL and normal myometrium fresh frozen tissue, we identified proteins that were differentially expressed, mainly related to focal adhesions and regulation of the actin cytoskeleton. The most frequently involved chromosomes included deletions in 10q22.2, 10q24.32, 13q14, and 13q21-31. Correlation analyses highlighted chromosome 10q as the most frequent cytoband, with corresponding proteins involved in regulating focal adhesions and the cytoskeleton. Integrated analysis between pathological and clinical characteristics indicated that chromosome 10q deletion and vascular morphology in IVL could serve as important markers predicting aggressive behavior. Our study sheds light on the pathological and molecular changes linked to IVL, which could pave the way for new treatment approaches.

Predictive models for personalized precision medical intervention in spontaneous regression stages of cervical precancerous lesions

Abstract Background During the prolonged period from Human Papillomavirus (HPV) infection to cervical cancer development, Low-Grade Squamous Intraepithelial Lesion (LSIL) stage provides a critical opportunity for cervical cancer prevention, giving the high potential for reversal in this stage. However, there is few research and a lack of clear guidelines on appropriate intervention strategies at this stage, underscoring the need for real-time prognostic predictions and personalized treatments to promote lesion reversal. Methods We have established a prospective cohort. Since 2018, we have been collecting clinical data and pathological images of HPV-infected patients, followed by tracking the progression of their cervical lesions. In constructing our predictive models, we applied logistic regression and six machine learning models, evaluating each model’s predictive performance using metrics such as the Area Under the Curve (AUC). We also employed the SHAP method for interpretative analysis of the prediction results. Additionally, the model identifies key factors influencing the progression of the lesions. Results Model comparisons highlighted the superior performance of Random Forests (RF) and Support Vector Machines (SVM), both in clinical parameter and pathological image-based predictions. Notably, the RF model, which integrates pathological images and clinical multi-parameters, achieved the highest AUC of 0.866. Another significant finding was the substantial impact of sleep quality on the spontaneous clearance of HPV and regression of LSIL. Conclusions In contrast to current cervical cancer prediction models, our model’s prognostic capabilities extend to the spontaneous regression stage of cervical cancer. This model aids clinicians in real-time monitoring of lesions and in developing personalized treatment or follow-up plans by assessing individual risk factors, thus fostering lesion spontaneous reversal and aiding in cervical cancer prevention and reduction. Graphical Abstract

Insights into spheroid formation: interaction of ovarian cancer cells with macrophage populations in the tumor microenvironment

Treating advanced ovarian cancer (OC) is challenging due to the immunosuppressive tumor microenvironment. This study investigates tumor-immune cell interactions using organotypic spheroid models that simulate the in vivo microenvironment. A dual-model spheroid system was established combining serous adenocarcinoma SKOV-3 cells with monocytes, pro-inflammatory (MΦ1) or anti-inflammatory (MΦ2) macrophages, or their derived exosomes (EXOs). In Model A, immune cells or EXOs were co-seeded with tumor cells to replicate early heterotypic aggregation. In Model B, immune cells or EXOs were introduced 24 h post-spheroid formation to simulate immune infiltration into established spheroids. Spheroid morphology was quantified by diameter and circularity, while the distribution of immune cells and EXOs was assessed via fluorescence intensity profiling in 2D and 3D. epithelial-to-mesenchymal transition (EMT) marker expression was analyzed to assess tumor cell phenotypic changes. Spheroids formed with SKOV-3 cells and ThP-1 monocytes developed a dense monocyte-enriched outer layer. Macrophage subtypes differentially influenced spheroid morphology: MΦ2 macrophages promoted the formation of multiple, loosely organized spheroids and increased N-cadherin expression, indicative of enhanced EMT. Similarly, MΦ-EXOs modulated EMT marker expression, underscoring the contribution of both direct cell interactions and paracrine signaling in regulating spheroid dynamics. Macrophages and their exosomes play a critical role in modulating the architecture and functional behavior of spheroids, reflecting two key aspects of OC progression: the formation of immune cell-enriched spheroids in ascitic fluid and tumor-immune interactions at peritoneal metastatic sites. This model provides a clinically relevant platform for preclinical testing of therapeutic strategies targeting peritoneal dissemination in OC.

T cell-engaging CD276xCD3 bispecific antibody for treatment of endometrial cancer

Abstract Background Endometrial cancer ranks among the most prevalent gynecological malignancies, with a notable increase in incidence, especially among women under 40. Although most patients are diagnosed at an early stage and have an excellent prognosis, the outcome for metastatic and recurrent cases remains poor. Current treatment for advanced-stage disease includes chemotherapy, hormonal therapy and checkpoint inhibitors. The clinical response rate to immunotherapy varies depending on the molecular subtype of endometrial carcinoma. Novel immunotherapeutic strategies are needed to improve patient survival, particularly across molecular subtypes. CD276 (cluster of differentiation 276, B7-H3) is emerging as a promising immunotherapy target due to its expression across multiple tumor types. Therapeutic targeting of CD276 may enhance immune cell infiltration into the tumor site by affecting its expression on tumor cells and tumor vasculature, which addresses a critical challenge for the successful treatment of solid tumors. Methods We developed a novel, IgG-based CD276xCD3 bispecific antibody termed CC-3, which has demonstrated pronounced preclinical efficacy in stimulating T cell antitumor responses and is presently undergoing evaluation in a Phase I clinical trial (NCT05999396). In this study, CC-3-induced T cell activation and proliferation was analyzed using flow cytometry. We also used a LegendPlex assay to measure the secretion of cytokines and effector molecules induced by CC-3. Finally, these processes culminated in target cell lysis which was analyzed using a flow cytometry-based cytotoxicity assay. Results CD276 is abundantly expressed in endometrial cancer. Treatment with CC-3 activated T cells, stimulated degranulation, and induced the secretion of cytokines and effector molecules, demonstrating CC-3-mediated T cell reactivity against endometrial cancer cells. Furthermore, CC-3 promoted robust T cell proliferation and memory T cell subset formation, culminating in potent target cell lysis. Conclusion Overall, our findings highlight the potential of CC-3 for clinical evaluation as a therapeutic option for patients with endometrial cancer.

Glutamine-driven metabolic reprogramming promotes CAR-T cell function through mTOR-SREBP2 mediated HMGCS1 upregulation in ovarian cancer

Chimeric antigen receptor T (CAR-T) cell therapy holds promise for cancer treatment, but its efficacy is often hindered by metabolic constraints in the tumor microenvironment. This study investigates the role of glutamine in enhancing CAR-T cell function against ovarian cancer. Metabolomic profiling of blood samples from ovarian cancer patients treated with MSLN-CAR-T cells was conducted to identify metabolic changes. In vitro, glutamine pretreatment was applied to CAR-T cells, and their proliferation, CAR expression, tumor lysis, and cytokine production (TNF-α, IFN-γ) were assessed. Mechanistic studies focused on the mTOR-SREBP2 pathway and its effect on HMGCS1 expression, membrane stability and immune synapse formation. In vivo, the antitumor effects and memory phenotype of glutamine-pretreated CAR-T cells were evaluated. Elevated glutamine levels were observed in the blood of ovarian cancer patients who responded to MSLN-CAR-T cell treatment. Glutamine pretreatment enhanced CAR-T cell proliferation, CAR expression, tumor lysis, and cytokine production. Mechanistically, glutamine activated the mTOR-SREBP2 pathway, upregulating HMGCS1 and promoting membrane stability and immune synapse formation. In vivo, glutamine-pretreated CAR-T cells exhibited superior tumor infiltration, sustained antitumor activity, and preserved memory subsets. Our findings highlight glutamine-driven metabolic rewiring via the mTOR-SREBP2-HMGCS1 axis as a strategy to augment CAR-T cell efficacy in ovarian cancer. NCT05372692.

Integrated multi-omic analysis of low-grade ovarian serous carcinoma collected from short and long-term survivors

Abstract Background Low-grade serous ovarian cancer (LGSOC) is a rare disease that occurs more frequently in younger women than those with high-grade disease. The current treatment is suboptimal and a better understanding of the molecular pathogenesis of this disease is required. In this study, we compared the proteogenomic analyses of LGSOCs from short- and long-term survivors (defined as &lt; 40 and &gt; 60 months, respectively). Our goal was to identify novel mutations, proteins, and mRNA transcripts that are dysregulated in LGSOC, particularly in short-term survivors. Methods Initially, targeted sequencing of 409 cancer-related genes was performed on 22 LGSOC and 6 serous borderline ovarian tumor samples. Subsequently, whole-genome sequencing analysis was performed on 14 LGSOC samples (7 long-term survivors and 7 short-term survivors) with matched normal tissue samples. RNA sequencing (RNA-seq), quantitative proteomics, and phosphoproteomic analyses were also performed. Results We identified single-nucleotide variants (SNVs) (range: 5688–14,833 per sample), insertion and deletion variants (indels) (range: 880–1065), and regions with copy number variants (CNVs) (range: 62–335) among the 14 LGSOC samples. Among all SNVs and indels, 2637 mutation sites were found in the exonic regions. The allele frequencies of the detected variants were low (median12%). The identified recurrent nonsynonymous missense mutations included KRAS, NRAS, EIF1AX, UBR5, and DNM3 mutations. Mutations in DNM3 and UBR5 have not previously been reported in LGSOC. For the two samples, somatic DNM3 nonsynonymous missense mutations in the exonic region were validated using Sanger sequencing. The third sample contained two missense mutations in the intronic region of DNM3, leading to a frameshift mutation detected in RNA transcripts in the RNA-seq data. Among the 14 LGSOC samples, 7754 proteins and 9733 phosphosites were detected by global proteomic analysis. Some of these proteins and signaling pathways, such as BST1, TBXAS1, MPEG1, HBA1, and phosphorylated ASAP1, are potential therapeutic targets. Conclusions This is the first study to use whole-genome sequencing to detect somatic mutations in LGSOCs with matched normal tissues. We detected and validated novel mutations in DNM3, which were present in 3 of the 14 samples analyzed. Additionally, we identified novel indels, regions with CNVs, dysregulated mRNA, dysregulated proteins, and phosphosites that are more prevalent in short-term survivors. This integrated proteogenomic analysis can guide research into the pathogenesis and treatment of LGSOC.

Genome-wide DNA methylation in relation to ARID1A deficiency in ovarian clear cell carcinoma

Abstract Background The poor chemo-response and high DNA methylation of ovarian clear cell carcinoma (OCCC) have attracted extensive attentions. Recently, we revealed the mutational landscape of the human kinome and additional cancer-related genes and found deleterious mutations in ARID1A , a component of the SWI/SNF chromatin-remodeling complex, in 46% of OCCC patients. The present study aims to comprehensively investigate whether ARID1A loss and genome-wide DNA methylation are co-regulated in OCCC and identify putative therapeutic targets epigenetically regulated by ARID1A. Methods DNA methylation of ARID1A mt/ko and ARID1A wt OCCC tumors and cell lines were analyzed by Infinium MethylationEPIC BeadChip. The clustering of OCCC tumors in relation to clinical and mutational status of tumors were analyzed by hierarchical clustering analysis of genome-wide methylation. GEO expression profiles were used to identify differentially methylated (DM) genes and their expression level in ARID1A mt/ko vs ARID1A wt OCCCs. Combining three pre-ranked GSEAs, pathways and leading-edge genes epigenetically regulated by ARID1A were revealed. The leading-edge genes that passed the in-silico validation and showed consistent ARID1A -related methylation change in tumors and cell lines were regarded as candidate genes and finally verified by bisulfite sequencing and RT-qPCR. Results Hierarchical clustering analysis of genome-wide methylation showed two clusters of OCCC tumors. Tumor stage, ARID1A/PIK3CA mutations and TP53 mutations were significantly different between the two clusters. ARID1A mutations in OCCC did not cause global DNA methylation changes but were related to DM promoter or gene-body CpG islands of 2004 genes. Three pre-ranked GSEAs collectively revealed the significant enrichment of EZH2- and H3K27me3-related gene-sets by the ARID1A -related DM genes. 13 Leading-edge DM genes extracted from the enriched gene-sets passed the expression-based in-silico validation and showed consistent ARID1A -related methylation change in tumors and cell lines. Bisulfite sequencing and RT-qPCR analysis showed promoter hypermethylation and lower expression of IRX1 , TMEM101 and TRIP6 in ARID1A mt compared to ARID1A wt OCCC cells, which was reversed by 5-aza-2′-deoxycytidine treatment. Conclusions Our study shows that ARID1A loss is related to the differential methylation of a number of genes in OCCC. ARID1A -dependent DM genes have been identified as key genes of many cancer-related pathways that may provide new candidates for OCCC targeted treatment.

TMEM176B inhibits ovarian cancer progression by regulating EMT via the Wnt/β-catenin signaling pathway

Abstract Background Ovarian cancer (OC) is recognized as one of the deadliest forms of gynecological cancer, approximately two-thirds of patients have already developed metastasis when they are diagnosed. The function of transmembrane protein 176B (TMEM176B) in the progression of OC remains elusive. This study aimed to investigate the role and molecular mechanism of TMEM176B on OC proliferation and metastasis. Method Expression of TMEM176B in OC and normal tissues were determined from the TCGA, GTEx, and CPTAC databases, and verified by patient-derived tissue samples. We analysed the prognostic relevance of TMEM176B in OC via Kaplan‒Meier (K‒M) survival curves and receiver operating characteristic (ROC) curves. Subsequent in vitro assays, including the CCK8 assay, colony formation assay, wound healing assay, and transwell assay, were performed to detect the influence of TMEM176B on cell proliferation and metastasis. Furthermore, a tumorigenesis study in nude mice was conducted to confirm the suppressive impact of TMEM176B on OC. RNA sequencing (RNA-seq) was utilized to uncover the mechanisms of TMEM176B on OC progression. Spearman correlation analysis was used to calculate the correlations between TMEM176B and cell adhesion, DNA replication, and the Wnt/β-catenin pathway. Finally, the role of TMEM176B in regulating the epithelial-mesenchymal transition (EMT) depending on the Wnt/β-catenin pathway was evaluated using LiCl agonist. Result The mRNA expression of TMEM176B was significantly downregulated in OC tissues, with lower TMEM176B correlating with a worse prognosis. Moreover, higher tumor stage and tumor grade were associated with a lower TMEM176B protein level. Consistent with these findings, OC tissues exhibited significantly reduced of TMEM176B compared to normal ovarian tissue from patients. In vitro studies indicated that TMEM176B knockdown increased both the proliferation, metastasis and EMT levels of OC cells, while TMEM176B overexpression had the opposite effects. In vivo investigations reinforced that TMEM176B significantly inhibited the progression of OC. RNA-seq analysis demonstrated that TMEM176B enhanced cell adhesion, diminished DNA replication, and suppressed EMT through the regulation of the Wnt/β-catenin signaling pathway, effectively obstructing the proliferation and metastasis of OC cells and impeding the disease’s progression. Conclusions TMEM176B inhibited EMT in OC cells by controlling the activation of the Wnt/β-catenin pathway. This mechanism underscored the diagnostic and prognostic potential of TMEM176B for OC and highlights its tumor-suppressive properties as a promising therapeutic candidate.

Machine learning-derived diagnostic model of epithelial ovarian cancer based on gut microbiome signatures

Prior studies have elucidated that alterations in gut microbiota are associated with a spectrum of tumors and metabolic disorders. However, the diagnostic value of gut microbiota in epithelial ovarian cancer remains insufficiently investigated. A total of 34 patients with a diagnosis of epithelial ovarian cancer (EOC), 15 patients with benign ovarian tumors (TB), and 30 healthy volunteers (NOR) were enrolled in this study. Fecal samples were collected, followed by sequencing of the V3-V4 region of the 16S rRNA gene. The clinical data and pathological characteristics were comprehensively recorded for further analysis, PICRUSt2 was utilized to conduct an analysis of microbial functional predictions, WGCNA networks were constructed by integrating microbiome and clinical data. LEfSe analysis was employed to identify microbial diagnostic markers, LASSO and SVM analyses were used to screen microbial diagnostic markers in conjunction with the Cally index, to establish a Microbial-Cally diagnostic model. Bootstrap resampling was utilized for the internal validation of the model, whereas the Hosmer-Lemeshow test and decision curve analysis (DCA) were employed to evaluate the diagnostic performance of the model. Plasma samples were subjected to untargeted metabolomics profiling, followed by differential analysis to identify key metabolites that are significantly altered in epithelial ovarian cancer. At the same time, Spearman correlation analysis was used to study the association between key microbiota and differential metabolites. The supernatants from Escherichia coli and Bifidobacterium cultures were co-cultured with SKOV3 cells. Cell proliferation, migration, and invasion were evaluated using Cell Counting Kit-8 (CCK-8) assay, Transwell migration and invasion assays. Apoptosis was assessed by flow cytometry analysis of fluorescence signals from Annexin V and propidium iodide (PI) staining. Compared to Nor and TB populations, individuals diagnosed with EOC demonstrated a significantly diminished gut microbiota diversity when contrasted with both normal controls and those presenting benign conditions. Specifically, the relative abundance of Bilophila, Bifidobacterium, and other probiotics was significantly reduced in patients diagnosed with epithelial ovarian cancer (EOC), while Escherichia and Shigella demonstrated a marked enrichment within this cohort. Differential microorganisms were identified through the application of machine learning techniques to delineate the characteristic microbial profiles associated with the EOC patients. A significant correlation was identified between the Cally index and microorganisms. In conclusion, we utilized microbial biomarkers alongside the Cally to establish a diagnostic model for epithelial ovarian cancer, receiver operating characteristic (ROC) curve Area Under Curve (AUC) of 0.976 (95%CI 0.943-1.00), The AUC obtained from the Bootstrap internal validation was 0.974. The Hosmer-Lemeshow test revealed a robust concordance between the observed probabilities and the predicted probabilities generated by the model. The decision curve analysis revealed that the model provided a significant net clinical benefit. A total of 233 differential metabolites were identified between the EOC group and the NT (NOR and TB) groups. Among these, eight specific metabolites (HMDB0243492, C09265, HMDB0242046, HMDB0240606, C04171, HMDB0060557, HMDB0252797, and C21412) were exclusively derived from the microbiome. Notably, metabolite HMDB0240606 exhibited a significant positive correlation with Escherichia coli and Shigella, while it showed a significant negative correlation with Ruminococcus. In vitro studies demonstrated that Bifidobacterium possessed anti-tumor activity, whereas Escherichia coli exhibited pro-tumor activity. This study provides the inaugural comprehensive analysis of gut microbiota composition and its differential profiles among patients with epithelial ovarian cancer, those with benign ovarian tumors, and healthy controls in Hunan province, China.

Cold atmospheric plasma drives USP49/HDAC3 axis mediated ferroptosis as a novel therapeutic strategy in endometrial cancer via reinforcing lactylation dependent p53 expression

Endometrial cancer ranks among the most common gynecological cancers, with increasing rates of incidence and death. Cold atmospheric plasma (CAP) has become a promising novel therapeutic approach for cancer treatment. Nevertheless, the specific impact of CAP on endometrial cancer remains inadequately characterized. This study aimed to investigate the effect of CAP on the progression of endometrial cancer and reveal its specific regulatory mechanisms. Colony formation, EdU, wound-healing, and transwell assay were used to detect the effect of CAP on endometrial cancer progression. Proteomics is employed to identify potential targets and signaling pathways through which CAP impacts endometrial cancer cells. MDA, lipid ROS, and JC-1 MMP assays were used to detect ferroptosis. Immunoprecipitation-mass spectrometry, co-immunoprecipitation, immunofluorescence co-localization, and molecular docking were used to analyze USP49 and HDAC3 interactions. The tumor xenografts model determined that CAP inhibits endometrial cancer growth in vivo. This study observed a significant inhibitory effect of CAP on the proliferation and migration of endometrial cancer cells and reported for the first time that CAP induces ferroptosis in endometrial cancer cells. Mechanistically, CAP activated the transcription of p53 by modulating HDAC3 mediated the histone H3K18 lactylation, resulting in upregulation of p53 driving cell ferroptosis. The interaction between USP49 and HDAC3 was validated through mass spectrometry and co-immunoprecipitation experiments. The regulation of HDAC3 by CAP is contingent upon USP49, wherein the down-regulation of USP49 augments the ubiquitination of HDAC3, consequently diminishing its protein stability. Furthermore, animal models with transplanted tumors corroborated the inhibitory impact of CAP on endometrial cancer in vivo. Our findings illustrate the suppressive effect of CAP treatment on endometrial cancer and uncover a novel regulatory mechanism mediated by CAP. Specifically, CAP modulates the ferroptosis pathway through the HDAC3/H3K18la/p53 axis, presenting a novel therapeutic approach for endometrial cancer treatment.

A Pronectin™ AXL-targeted first-in-class bispecific T cell engager (pAXLxCD3ε) for ovarian cancer

AbstractBackgroundPronectins™ are a new class of fibronectin-3-domain 14th-derived (14Fn3) antibody mimics that can be engineered as bispecific T cell engager (BTCE) to redirect immune effector cells against cancer. We describe here the in vitro and in vivo activity of a Pronectin™ AXL-targeted first-in-class bispecific T cell engager (pAXLxCD3ε) against Epithelial Ovarian Cancer (EOC).MethodspAXLxCD3ε T-cell mediated cytotoxicity was evaluated by flow cytometry and bioluminescence. pAXLxCD3ε mediated T-cell infiltration, activation and proliferation were assessed by immunofluorescence microscopy and by flow cytometry. Activity of pAXLxCD3ε was also investigated in combination with poly-ADP ribose polymerase inhibitors (PARPi). In vivo antitumor activity of pAXLxCD3ε was evaluated in immunocompromised (NSG) mice bearing intraperitoneal or subcutaneous EOC xenografts and immunologically reconstituted with human peripheral blood mononuclear cells (PBMC).ResultspAXLxCD3ε induced dose-dependent cytotoxicity by activation of T lymphocytes against EOC cells, regardless of their histologic origin. The addition of PARPi to cell cultures enhanced pAXLxCD3ε cytotoxicity. Importantly, in vivo, pAXLxCD3ε was highly effective against EOC xenografts in two different NSG mouse models, by inhibiting the growth of tumor cells in ascites and subcutaneous xenografts. This effect translated into a significantly prolonged survival of treated animals.ConclusionpAXLxCD3ε is an active therapeutics against EOC cells providing a rational for its development as a novel agent in this still incurable disease. The preclinical validation of a first-in-class agent opens the way to the development of a new 14Fn3-based scaffold platform for the generation of innovative immune therapeutics against cancer.

Single-cell transcriptomics in ovarian cancer identify a metastasis-associated cell cluster overexpressed RAB13

Abstract Background Metastasis, the leading cause of cancer-related death in patients diagnosed with ovarian cancer (OC), is a complex process that involves multiple biological effects. With the continuous development of sequencing technology, single-cell sequence has emerged as a promising strategy to understand the pathogenesis of ovarian cancer. Methods Through integrating 10 × single-cell data from 12 samples, we developed a single-cell map of primary and metastatic OC. By copy-number variations analysis, pseudotime analysis, enrichment analysis, and cell–cell communication analysis, we explored the heterogeneity among OC cells. We performed differential expression analysis and high dimensional weighted gene co-expression network analysis to identify the hub genes of C4. The effects of RAB13 on OC cell lines were validated in vitro. Results We discovered a cell subcluster, referred to as C4, that is closely associated with metastasis and poor prognosis in OC. This subcluster correlated with an epithelial–mesenchymal transition (EMT) and angiogenesis signature and RAB13 was identified as the key marker of it. Downregulation of RAB13 resulted in a reduction of OC cells migration and invasion. Additionally, we predicted several potential drugs that might inhibit RAB13. Conclusions Our study has identified a cell subcluster that is closely linked to metastasis in OC, and we have also identified RAB13 as its hub gene that has great potential to become a new therapeutic target for OC.

Function and mechanism of GBP1 in the development and progression of cervical cancer

AbstractGuanylate binding protein 1 (GBP1) is the most concerned member of the GBP family, which has a series of effects such as anti-infection and anti-angiogenesis. Its role in malignant tumors including cervical cancer is still controversial. We aim to explore the effects of GBP1 on cervical cancer through bioinformatics and related experiments. In this study, we first found that GBP1 was generally expressed in cervical cancer in various online databases and was closely related to immune invasion. Secondly, we used multicolor immunofluorescence technology to verify the expression of GBP1 in cervical cancer tissues and its relationship with immune invasion, and explored its relationship with the prognosis of patients with cervical cancer. Knockdown and overexpression assays of GBP1 in vitro were used to prove GBP1 as a potential oncogene of cervical cancer, and its carcinogenicity was verified by in vivo experiment. In order to explore the potential mechanism of GBP1 in promoting cancer, RNA-seq was performed on GBP1 overexpression and knockdown expression cell lines, and GBP1 knockdown and overexpression were found to be associated with many RNA alternative splicing events, suggesting that GBP1 maybe a RNA binding protein (RBP) which affect the biological characteristics of cervical cancer cells through the alternative splicing pathway. However, the later RNA binding protein immunoprecipitation (RIP) assay proved that GBP1 was not a direct alternative splicing factor, while the co-immunoprecipitation (CoIP)-mass spectroscopy (MS) assay combined with protein protein interaction (PPI) analysis proved that 8 alternative splicing factors including Heterogeneous Nuclear Ribonucleoprotein K (HNRNPK) were interacting proteins of GBP1. Combined with the existing reports and the results of RNA-seq alternative splicing analysis, it is speculated that GBP1 may regulate the alternative splicing of CD44 protein by binding to interacting protein-HNRNPK, and thus play a role in promoting cancer in cervical cancer.

Characteristics of vaginal microbiota in various cervical intraepithelial neoplasia: a cross-sectional study

Abstract Background Precancerous lesions of cervical cancer exhibit characteristics indicative of natural progression. To prevent overtreatment of patients whose cervical intraepithelial neoplasia (CIN) in regression and to predict the onset of invasive cervical cancer at an early stage, we've identified the vaginal microbiome as a potential key factor, which is associated with both HPV infection and the various cervical intraepithelial neoplasia. This study aims to investigate the microbiome characteristics of patients with various cervical intraepithelial neoplasia. Methods Utilizing high-throughput 16S ribosomal RNA (16S rRNA) sequencing technology, a description of the characteristics and community composition of Vaginal Microbiota (VMB) was conducted among 692 Chinese women infected with the High-risk Human Papillomavirus (HR-HPV). Results As the grade of the lesions increased, the proportions of Lactobacillus and Pseudomonas demonstrated a significant declining trend, while the proportions of Gardnerella , Dialister , and Prevotella significantly increased. The diversity of the VMB was more significant in high-grade CIN. Furthermore, KEGG pathway enrichment analysis indicates that high-grade cervical intraepithelial neoplasia can inhibit various pathways, including those of phosphotransferase system, transcription factors, Fructose and mannose metabolism, amino sugar and nucleotide sugar metabolism, and galactose metabolism, which may contribute to the development of early cervical cancer symptoms. Conclusion Patients with CIN exhibit a distinct vaginal microbial profile characterized by a decrease in Lactobacillus and Pseudomonas , and an increase in Gardnerella , Prevotella , and Dialister . The proliferation and diminution of these two types of microbial communities are interrelated, suggesting a mutual restraint and balance among them. Disruption of this regulatory balance could potentially lead to the onset of cervical lesions and carcinogenesis. Retrospectively registered: This study was approved by the Ethics Committee of the Beijing Chaoyang Hospital affiliated with the Capital Medical University (NO.2023-S-415).

Characterization of tumor microenvironment and tumor immunology based on the double-stranded RNA-binding protein related genes in cervical cancer

Abstract Background Cervical cancer is one of the most common gynecological cancers threatening women’s health worldwide. Double-stranded RNA-binding proteins (dsRBPs) regulate innate immunity and are therefore believed to be involved in virus-related malignancies, however, their role in cervical cancer is not well known. Methods We performed RNA-seq of tumor samples from cervical cancer patients in local cohort and also assessed the RNA-seq and clinical data derived from public datasets. By using single sample Gene Set Enrichment Analysis (ssGSEA) and univariate Cox analysis, patients were stratified into distinct dsRBP clusters. Stepwise Cox and CoxBoost were performed to construct a risk model based on optimal dsRBPs clusters-related differentially expressed genes (DEGs), and GSE44001 and CGCI-HTMCP-CC were employed as two external validation cohorts. Single cell RNA sequencing data from GSE168652 and Scissor algorithm were applied to evaluated the signature-related cell population. Results The expression of dsRBP features was found to be associated with HPV infection and carcinogenesis in CESC. However, only Adenosine deaminases acting on RNA (ADAR) and Dicer, Drosha, and Argonautes (DDR) exhibited significant correlations with the overall survival (OS) of CESC patients. Based on these findings, CESC patients were divided into three dsRBP clusters. Cluster 3 showed superior OS but lower levels of ADAR and DDR. Additionally, Cluster 3 demonstrated enhanced innate immunity, with significantly higher activity in cancer immunity cycles, immune scores, and levels of tumor-infiltrating immune cells, particularly CD8+ T cells. Furthermore, a risk model based on nine dsRBP cluster-related DEGs was established. The accuracy of survival prediction for 1 to 5 years was consistently above 0.78, and this model’s robust predictive capacity was confirmed by two external validation sets. The low-risk group exhibited significantly higher levels of immune checkpoints, such as PDCD1 and CTLA4, as well as a higher abundance of CD8+ T cells. Analysis of single-cell sequencing data revealed a significant association between the dsRBP signature and glycolysis. Importantly, low-risk patients showed improved OS and a higher response rate to immunotherapy, along with enduring clinical benefits from concurrent chemoradiotherapy. Conclusions dsRBP played a crucial role in the regulation of prognosis and tumor immunology in cervical cancer, and its prognostic signature provides a strategy for risk stratification and immunotherapy evaluation.

Establishment of a high-fidelity patient-derived xenograft model for cervical cancer enables the evaluation of patient’s response to conventional and novel therapies

Abstract Background Recurrent or metastatic cervical cancer (r/m CC) often has poor prognosis owing to its limited treatment options. The development of novel therapeutic strategies has been hindered by the lack of preclinical models that accurately reflect the biological and genomic heterogeneity of cervical cancer (CC). Herein, we aimed to establish a large patient-derived xenograft (PDX) biobank for CC, evaluate the consistency of the biologic indicators between PDX and primary tumor tissues of patients, and explore its utility for assessing patient’s response to conventional and novel therapies. Methods Sixty-nine fresh CC tumor tissues were implanted directly into immunodeficient mice to establish PDX models. The concordance of the PDX models with their corresponding primary tumors (PTs) was compared based on the clinical pathological features, protein biomarker levels, and genomic features through hematoxylin &amp; eosin staining, immunohistochemistry, and whole exome sequencing, respectively. Moreover, the clinical information of CC patients, RNA transcriptome and immune phenotyping of primary tumors were integrated to identify the potential parameters that could affect the success of xenograft engraftment. Subsequently, PDX model was evaluated for its capacity to mirror patient’s response to chemotherapy. Finally, PDX model and PDX-derived organoid (PDXO) were utilized to evaluate the therapeutic efficacy of neratinib and adoptive cell therapy (ACT) combination strategy for CC patients with human epidermal growth factor receptor 2 (HER2) mutation. Results We established a PDX biobank for CC with a success rate of 63.8% (44/69). The primary features of established PDX tumors, including clinicopathological features, the expression levels of protein biomarkers including Ki67, α-smooth muscle actin, and p16, and genomics, were highly consistent with their PTs. Furthermore, xenograft engraftment was likely influenced by the primary tumor size, the presence of follicular helper T cells and the expression of cell adhesion-related genes in primary tumor tissue. The CC derived PDX models were capable of recapitulating the patient’s response to chemotherapy. In a PDX model, a novel therapeutic strategy, the combination of ACT and neratinib, was shown to effectively inhibit the growth of PDX tumors derived from CC patients with HER2-mutation. Conclusions We established by far the largest PDX biobank with a high engraftment rate for CC that preserves the histopathological and genetic characteristics of patient’s biopsy samples, recapitulates patient’s response to conventional therapy, and is capable of evaluating the efficacy of novel therapeutic modalities for CC.

The promoting effect and mechanism of Nrf2 on cell metastasis in cervical cancer

Abstract Background Cervical cancer (CC) has poor prognosis and high mortality rate for its metastasis during the disease progression. Epithelial-mesenchymal transition (EMT) and anoikis are initial and pivotal steps during the metastatic process. Although higher levels of Nrf2 are associated with aggressive tumor behaviors in cervical cancer, the detailed mechanism of Nrf2 in cervical cancer metastasis, especially EMT and anoikis, remains unclear. Methods Immunohistochemistry (IHC) was used to examine Nrf2 expression in CC. Wound healing assay and transwell analysis were used to evaluate the migration ability of CC cells. Western blot, qTR-PCR and immunofluorescent staining were used to verify the expression level of Nrf2, the EMT associated markers and anoikis associated proteins. Flow cytometry assays and cell counting were used to detect the apoptosis of cervical cancer cells. The lung and lymph node metastatic mouse model were established for studies in vivo. The interaction between Nrf2 and Snail1 was confirmed by rescue-of-function assay. Results When compared with cervical cancer patients without lymph node metastasis, Nrf2 was highly expressed in patients with lymph node metastasis. And Nrf2 was proved to enhance the migration ability of HeLa and SiHa cells. In addition, Nrf2 was positively correlated with EMT processes and negatively associated with anoikis in cervical cancer. In vivo, a xenograft assay also showed that Nrf2 facilitated both pulmonary and lymphatic distant metastasis of cervical cancer. Rescue-of-function assay further revealed the mechanism that Nrf2 impacted the metastasis of CC through Snail1. Conclusion Our fundings established Nrf2 plays a crucial role in the metastasis of cervical cancer by enhancing EMT and resistance to anoikis by promoting the expression of Snail1, with potential value as a therapeutic candidate.

The epigenetic factor CHD4 contributes to metastasis by regulating the EZH2/β-catenin axis and acts as a therapeutic target in ovarian cancer

Abstract Background The overall survival rate of patients with advanced ovarian cancer (OC) has remained static for several decades. Advanced ovarian cancer is known for its poor prognosis due to extensive metastasis. Epigenetic alterations contribute to tumour progression and therefore are of interest for potential therapeutic strategies. Methods Following our previous study, we identified that CHD4, a chromatin remodelling factor, plays a strong role in ovarian cancer cell metastasis. We investigated the clinical significance of CHD4 through TCGA and GEO database analyses and explored the effect of CHD4 expression modulation and romidepsin treatment on the biological behaviour of ovarian cancer through CCK-8 and transwell assays. Bioluminescence imaging of tumours in xenografted mice was applied to determine the therapeutic effect of romidepsin. GSEA and western blotting were used to screen the regulatory mechanism of CHD4. Results In ovarian cancer patient specimens, high CHD4 expression was associated with a poor prognosis. Loss of function of CHD4 in ovarian cancer cells induced suppression of migration and invasion. Mechanistically, CHD4 knockdown suppressed the expression of EZH2 and the nuclear accumulation of β-catenin. CHD4 also suppressed the metastasis of ovarian cancer cells and prevented disease progression in a mouse model. To inhibit the functions of CHD4 that are mediated by histone deacetylase, we evaluated the effect of the HDAC1/2 selective inhibitor romidepsin. Our findings indicated that treatment with romidepsin suppressed the progression of metastases in vitro and in vivo. Conclusions Collectively, our results uncovered an oncogenic function of CHD4 in ovarian cancer and provide a rationale for clinical trials of romidepsin in ovarian cancer patients.

Development of a 3D functional assay and identification of biomarkers, predictive for response of high-grade serous ovarian cancer (HGSOC) patients to poly-ADP ribose polymerase inhibitors (PARPis): targeted therapy

Abstract Background Poly(ADP-ribose) polymerase inhibitors (PARPis) specifically target homologous recombination deficiency (HRD) cells and display good therapeutic effect in women with advanced-stage BRCA1/2-mutated breast and epithelial ovarian cancer (EOC). However, about 50% of high grade serous ovarian cancers (HGSOC) present with HRD due to epigenetic BRCA1 inactivation, as well as genetic/epigenetic inactivation(s) of other HR genes, a feature known as “BRCAness”. Therefore, there is a potential for extending the use of PARPis to these patients if HR status can be identified. Methods We have developed a 3D (spheroid) functional assay to assess the sensitivity of two PARPis (niraparib and olaparib) in ascites-derived primary cell cultures (AsPCs) from HGSOC patients. A method for AsPCs preparation was established based on a matrix (agarose), allowing for easy isolation and successive propagation of monolayer and 3D AsPCs. Based on this method, we performed cytotoxicity assays on 42 AsPCs grown both as monolayers and spheroids. Results The response to PARPis treatment in monolayer AsPCs, was significantly higher, compared to 3D AsPCs, as 88% and 52% of the monolayer AsPCs displayed sensitivity to niraparib and olaparib respectively, while 66% of the 3D AsPCs were sensitive to niraparib and 38% to olaparib, the latter being more consistent with previous estimates of HRD (40%–60%) in EOC. Moreover, niraparib displayed a significantly stronger cytotoxic effect in both in 3D and monolayer AsPCs, which was confirmed by consecutive analyses of the HR pathway activity (γH2AX foci formation) in PARPis-sensitive and resistant AsPCs. Global gene expression comparison of 6 PARPi-resistant and 6 PARPi-sensitive 3D AsPCs was indicative for the predominant downregulation of numerous genes and networks with previously demonstrated roles in EOC chemoresistance, suggesting that the PARPis-sensitive AsPCs could display enhanced sensitivity to other chemotherapeutic drugs, commonly applied in cancer management. Microarray data validation identified 24 potential gene biomarkers associated with PARPis sensitivity. The differential expression of 7 selected biomarkers was consecutively confirmed by immunohistochemistry in matched EOC tumor samples. Conclusion The application of this assay and the potential biomarkers with possible predictive significance to PARPis therapy of EOC patients now need testing in the setting of a clinical trial.

A comprehensive custom panel evaluation for routine hereditary cancer testing: improving the yield of germline mutation detection

Abstract Background In the context of our Regional Program of Hereditary Cancer, individuals fulfilling the criteria are tested for germline mutations to subsequently establish the clinical management. Our standard diagnostic approach focuses on sequencing a few classic high-risk genes, a method that frequently renders uninformative genetic results. This study aims to examine the improved yield offered by an On-Demand panel. Methods We designed an On-Demand panel for the analysis of 35-genes associated with inherited cancer susceptibility in a total of 128 cases of Hereditary Breast and Ovarian Cancer (HBOC) and Hereditary Nonpolyposis Colorectal Cancer (HNPCC). Results Eighteen deleterious mutations were detected, in both routinely (BRCA2, MLH1, MSH2, PMS2) and non-routinely (ATM, BLM, BRIP1, CHEK2, MUTYH) tested genes. The screening extended to 35 genes rendered by patients carrying several- up to 6-Variants of Unknown Significance (VUS). Moreover, we confirmed the splicing disruption at RNA level for a not previously reported BRIP1 splicing mutation. Using an On-Demand panel, we identified 18 pathogenic mutation carriers, seven of which would have gone unnoticed with traditional analysis. Conclusions Our results reinforce the utility of NGS gene panels in the diagnostic routine to increase the performance of genetic testing, especially in individuals from families with overlapping cancer phenotypes.

Accumulation of blood-circulating PD-L1-expressing M-MDSCs and monocytes/macrophages in pretreatment ovarian cancer patients is associated with soluble PD-L1

AbstractBackgroundPrevious studies have shown clinical relevance of programmed death-ligand 1 (PD-L1) and soluble PD-L1 (sPD-L1) in human cancers. However, still contradictory results exist. Our aim was evaluation of PD-L1-expressing monocytic myeloid-derived suppressor cells (M-MDSCs), monocytes/macrophages (MO/MA), tumour cells (TC) and immune/inflammatory cells (IC) as well as investigation of the sPD-L1 in ovarian cancer (OC) patients.MethodsThe group of 74 pretreatment women were enrollment to the study. The expression of PD-L1 on M-MDSCS and MO/MA was assessed by flow cytometry. The profile of sPD-L1 was examined with ELISA. The expression of PD-L1 in mononuclear cells (MCs) was analyzed using real time PCR. PD-L1 immunohistochemical analysis was prepared on TC and IC. An in silico validation of prognostic significance of PD-L1 mRNA expression was performed based microarray datasets.ResultsOC patients had significantly higher frequency of MO/MA versus M-MDSC in the blood, ascites and tumour (each p &lt; 0.0001). In contrast, PD-L1 expression was higher on M-MDSCs versus MO/MA in the blood and ascites (each p &lt; 0.0001), but not in the tumour (p &gt; 0.05). Significantly higher accumulation of blood-circulating M-MDSC, MO/MA, PD-L1+M-MDSC, PD-L1+MO/MA and sPD-L1 was observed in patients versus control (p &lt; 0.001, p &lt; 0.05, p &lt; 0.001, p &lt; 0.001 and p &lt; 0.0001, respectively). Accumulation of these factors was clinicopathologic-independent (p &gt; 0.05). The expression of PD-L1 was significantly higher on IC versus TC (p &lt; 0.0001) and was clinicopathologic-independent (p &gt; 0.05) except higher level of PD-L1+TC in the endometrioid versus mucinous tumours. Interestingly, blood-circulating sPD-L1 positively correlated with PD-L1+M-MDSCs (p = 0.03) and PD-L1+MO/MA (p = 0.02) in the blood but not with these cells in the ascites and tumours nor with PD-L1+TC/IC (each p &gt; 0.05). PD-L1 and sPD-L1 were not predictors of overall survival (OS; each p &gt; 0.05). Further validation revealed no association between PD-L1 mRNA expression and OS in large independent OC patient cohort (n = 655, p &gt; 0.05).ConclusionsAlthough PD-L1 may not be a prognostic factor for OC, our study demonstrated impaired immunity manifested by up-regulation of PD-L1/sPD-L1. Furthermore, there was a positive association between PD-L1+myeloid cells and sPD-L1 in the blood, suggesting that sPD-L1 may be a noninvasive surrogate marker for PD-L1+myeloid cells immunomonitoring in OC. Overall, these data should be under consideration during future clinical studies/trials.

Establishment of a novel glycolysis-related prognostic gene signature for ovarian cancer and its relationships with immune infiltration of the tumor microenvironment

Abstract Background Glycolysis affects tumor growth, invasion, chemotherapy resistance, and the tumor microenvironment. In this study, we aimed to construct a glycolysis-related prognostic model for ovarian cancer and analyze its relationship with the tumor microenvironment’s immune cell infiltration. Methods We obtained six glycolysis-related gene sets for gene set enrichment analysis (GSEA). Ovarian cancer data from The Cancer Genome Atlas (TCGA) database and two Gene Expression Omnibus (GEO) datasets were divided into two groups after removing batch effects. We compared the tumor environments' immune components in high-risk and low-risk groups and analyzed the correlation between glycolysis- and immune-related genes. Then, we generated and validated a predictive model for the prognosis of ovarian cancer using the glycolysis-related genes. Results Overall, 27/329 glycolytic genes were associated with survival in ovarian cancer, 8 of which showed predictive value. The tumor cell components in the tumor microenvironment did not differ between the high-risk and low-risk groups; however, the immune score differed significantly between groups. In total, 13/24 immune cell types differed between groups, including 10 T cell types and three other immune cell types. Eight glycolysis-related prognostic genes were related to the expression of multiple immune-related genes at varying degrees, suggesting a relationship between glycolysis and immune response. Conclusions We identified eight glycolysis-related prognostic genes that effectively predicted survival in ovarian cancer. To a certain extent, the newly identified gene signature was related to the tumor microenvironment, especially immune cell infiltration and immune-related gene expression. These findings provide potential biomarkers and therapeutic targets for ovarian cancer.

Dynamic changes of angiopoietins and endothelial nitric oxide supply during fluid resuscitation for major gyn-oncological surgery: a prospective observation

Abstract Background Despite goal-directed hemodynamic therapy, vascular function may deteriorate during surgery for advanced abdominal tumor masses. Fluid administration has been shown to be associated with distinct changes in serum levels of functional proteins. We sought to determine how serum total protein and angiopoietin (ANG) levels change during major abdominal tumor surgery. In addition, ex vivo endothelial nitric oxide synthase (eNOS) activation as well as NO bioavailability in vivo were assessed. Methods 30 patients scheduled for laparotomy for late-stage ovarian or uterine cancer were prospectively included. Advanced hemodynamic monitoring as well as protocol-driven goal-directed fluid optimization were performed. Total serum protein, ANG-1, -2, and soluble TIE2 were determined pre-, intra-, and postoperatively. Phosphorylation of eNOS was assessed in microvascular endothelial cells after incubation with patient serum, and microvascular reactivity was determined in vivo by near-infrared spectroscopy and arterial vascular occlusion. Results Cardiac output as well as preload gradually decreased during surgery and were associated with a median total fluid intake of 12.8 (9.7–15.4) mL/kg*h and a postoperative fluid balance of 6710 (4113–9271) mL. Total serum protein decreased significantly from baseline (66.5 (56.4–73.3) mg/mL) by almost half intraoperatively (42.7 (36.8–51.5) mg/mL, p &lt; 0.0001) and remained at low level. While ANG-1 showed no significant dilutional change (baseline: 12.7 (11.9–13.9) ng/mL, postop.: 11.6 (10.8 –13.5) ng/mL, p = 0.06), serum levels of ANG-2 were even increased postoperatively (baseline: 2.2 (1.6–2.6) ng/mL vs. postop.: 3.4 (2.3–3.8) ng/mL, p &lt; 0.0001), resulting in a significant shift in ANG-2 to ANG-1 ratio. Ex vivo phosphorylation of eNOS was decreased depending on increased ANG-2 levels and ANG-2/1 ratio (Spearman r = − 0.37, p = 0.007). In vivo, increased ANG-2 levels were associated with impaired capillary recruitment and NO bioavailability (Spearman r = − 0.83, p = 0.01). Conclusions Fluid resuscitation-associated changes in serum vascular mediator profile during abdominal tumor surgery were accompanied by impaired eNOS activity ex vivo as well as reduced NO bioavailability in vivo. Our results may explain disturbed microvascular function in major surgery despite goal-directed hemodynamic optimization.

Applying low coverage whole genome sequencing to detect malignant ovarian mass

AbstractTo evaluate whether low coverage whole genome sequencing is suitable for the detection of malignant pelvic mass and compare its diagnostic value with traditional tumor markers. We enrolled 63 patients with a pelvic mass suspicious for ovarian malignancy. Each patient underwent low coverage whole genome sequencing (LCWGS) and traditional tumor markers test. The pelvic masses were finally confirmed via pathological examination. The copy number variants (CNVs) of whole genome were detected and the Stouffers Z-scores for each CNV was extracted. The risk of malignancy (RM) of each suspicious sample was calculated based on the CNV counts and Z-scores, which was subsequently compared with ovarian cancer markers CA125 and HE4, and the risk of ovarian malignancy algorithm (ROMA). Receiver Operating Characteristic Curve (ROC) were used to access the diagnostic value of variables. As confirmed by pathological diagnosis, 44 (70%) patients with malignancy and 19 patients with benign mass were identified. Our results showed that CA125 and HE4, the CNV, the mean of Z-scores (Zmean), the max of Z-scores (Zmax), the RM and the ROMA were significantly different between patients with malignant and benign masses. The area under curve (AUC) of CA125, HE4, CNV, Zmax, and Zmean was 0.775, 0.866, 0.786, 0.685 and 0.725 respectively. ROMA and RM showed similar AUC (0.876 and 0.837), but differed in sensitivity and specificity. In the validation cohort, the AUC of RM was higher than traditional serum markers. In conclusion, we develop a LCWGS based method for the identification of pelvic mass of suspicious ovarian cancer. LCWGS shows accurate result and could be complementary with the existing diagnostic methods.

miR-18a-5p derived from mesenchymal stem cells-extracellular vesicles inhibits ovarian cancer cell proliferation, migration, invasion, and chemotherapy resistance

Abstract Objective Ovarian cancer (OC) is a major threat to women’s health. Mesenchymal stem cells (MSCs) are key regulators in cellular communication by secreting extracellular vesicles (EVs) that are involved in OC. This study probed into the mechanism of human MSCs derived-EVs (hMSC-EVs) in regulating OC cell growth and chemotherapy resistance. Methods hMSCs and EVs were isolated and identified. After adding EVs, the uptake of EVs by OC CAOV3/ES2 cells (for in vitro studies), and cell proliferation, migration, and invasion were detected. Downregulated miRNAs in hMSC-EVs were screened and miR-18a-5p expression in OC patients was detected. The prognosis of OC patients was analyzed. Binding sites of miR-18a-5p and NACC1 were predicted and validated. NACC1 expression in OC tissues was measured by RT-qPCR, and its correlation with miR-18a-5p was analyzed by Pearson method. AKT/mTOR pathway activation was assessed by WB. The cisplatin sensitivity of EVs-treated CAOV3 cells was evaluated via MTT assay and tested by tumor formation assay in nude mice. Results hMSC-EVs suppressed OC cell proliferation, migration, and invasion. miR-18a-5p was downregulated in OC and miR-18a-5p low expression was associated with a poor prognosis. EV-encapsulated miR-18a-5p targeted NACC1. NACC1 was upregulated in OC tissues. miR-18a-5p knockdown and NACC1 overexpression both annulled the inhibition of hMSC-EVs on OC cell growth. AKT and mTOR were elevated in OC and NACC1 activated the AKT/mTOR pathway in OC cells. hMSC-EVs promoted cisplatin sensitivity of OC cells by carrying miR-18a-5p. Conclusion hMSC-EVs-derived miR-18a-5p inhibits OC cell proliferation, migration, invasion, and chemotherapy resistance.

FOXA1 can be modulated by HDAC3 in the progression of epithelial ovarian carcinoma

Abstract FOXA1 is associated with malignant tumors, but the function of FOXA1 in EOC is unclear. HDAC3 can influence the proliferation, migration and invasion ability of EOC. In this study, we wanted to explore the function of FOXA1 in ovarian cancer and the relationship between HDAC3 and FOXA1.The expression of HDAC3 and FOXA1 was detected by immunohistochemical staining of primary lesions from 127 epithelial ovarian carcinoma patients. A proliferation assay, a Transwell assay, an apoptosis assay and animal experiments were used to assess the proliferation, invasion and apoptosis abilities of ovarian cancer cells before and after transfection with FOXA1. The relevance of the in vitro findings was confirmed in xenografts. The H-scores for FOXA1 and HDAC3 staining in FIGO stage III-IV were noticeably higher and predicted adverse clinical outcomes in patients with ovarian cancer. The expression level of HDAC3 was significantly correlated with the expression level of FOXA1. Invasion, proliferation and apoptosis capacity and tumor formation were decreased in the FOXA1-knockdown cells. Experiments in xenografts confirmed that HDAC3 mediated tumor formation. In conclusion, FOXA1 can be modulated by HDAC3 through the Wnt/β-catenin signaling pathway, and FOXA1 plays essential roles in the proliferation, apoptosis and invasion of EOC cell lines and xenograft experiments.

Exosomal lncRNA UCA1 modulates cervical cancer stem cell self-renewal and differentiation through microRNA-122-5p/SOX2 axis

Abstract Background There is growing evidence discussing the role of long non-coding RNAs (lncRNAs) in cervical cancer (CC). We performed this study to explore the impact of exosomal lncRNA urothelial cancer-associated 1 (UCA1) in CC stem cells by sponging microRNA-122-5p (miR-122-5p) and regulating SOX2 expression. Methods CC stem cells (CD133+CaSki) and exosomes were extracted and identified. The synthesized UCA1- and miR-122-5p-related sequences were transfected into CaSki cells, CaSki cells-derived exosomes were extracted and then co-cultured with CD133+CaSki cells. The functional roles of UCA1 and miR-122-5p in self-renewal and differentiation ability of CC stem cells were determined using ectopic expression, knockdown/depletion and reporter assay experiments. An in vivo experiment was performed to verify the in vitro results. Results Up-regulated UCA1 and SOX2 and down-regulated miR-122-5p were found in CaSki-Exo. Exosomes promoted invasion, migration, proliferation and restrained apoptosis of CD133+CaSki cells. Silencing UCA1 or up-regulating miR-122-5p degraded SOX2 expression, and reduced invasion, migration and proliferation of CD133+CaSki cells while advanced apoptosis and suppressed the tumor volume and weight in nude mice. Conclusion Our study provides evidence that CaSki-Exo can promote the self-renewal and differentiation ability of CC stem cells while silencing UCA1 or up-regulating miR-122-5p restrains self-renewal and differentiation of CC stem cells.

LINC01089 in cancer: multifunctional roles and therapeutic implications

AbstractLINC01089 is a prime example of a long non-coding RNA that plays a pivotal role in the progression of human cancers. The gene encoding this lncRNA is located on 12q24.31. LINC01089 has been demonstrated to exert tumor-suppressive effects in various cancers, including colorectal cancer, gastric cancer, lung cancer, ovarian cancer, cervical cancer, papillary thyroid carcinoma, breast cancer, and osteosarcoma. However, its role in hepatocellular carcinoma shows significant discrepancies across different studies. In this review, we systematically explore the functions of LINC01089 in human cancers through bioinformatics analysis, clinical studies, animal models, and fundamental experimental research. Furthermore, we delve into the biological mechanisms and functions of LINC01089, and discuss its potential as a future biomarker and therapeutic target in detail.

Functional genetic variants of GEN1 predict overall survival of Chinese epithelial ovarian cancer patients

Abstract Background Inherited variations in DNA double-strand break (DSB) repair pathway are known to influence ovarian cancer occurrence, progression and treatment response. Despite its significance, survival-associated genetic variants within the DSB pathway remain underexplored. Methods In the present study, we performed a two-phase analysis of 19,290 single-nucleotide polymorphisms (SNPs) in 199 genes in the DSB repair pathway from a genome-wide association study (GWAS) dataset and explored their associations with overall survival (OS) in 1039 Han Chinese epithelial ovarian carcinoma (EOC) patients. After utilizing multivariate Cox regression analysis with bayesian false-discovery probability for multiple test correction, significant genetic variations were identified and subsequently underwent functional prediction and validation. Results We discovered a significant association between poor overall survival and the functional variant GEN1 rs56070363 C &gt; T (CT + TT vs. TT, adjusted hazard ratio (HR) = 2.50, P &lt; 0.001). And the impact of GEN1 rs56070363 C &gt; T on survival was attributed to its reduced binding affinity to hsa-miR-1287-5p and the resultant upregulation of GEN1 mRNA expression. Overexpression of GEN1 aggregated EOC cell proliferation, invasion and migration presumably by influencing the expression of immune inhibitory factors, thereby elevating the proportion of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs) and then constructing an immunosuppressive tumor microenvironment. Conclusions In conclusion, GEN1 rs56070363 variant could serve as a potential predictive biomarker and chemotherapeutic target for improving the survival of EOC patients.

Targeting interleukin-6 as a treatment approach for peritoneal carcinomatosis

AbstractPeritoneal carcinomatosis (PC) is a complex manifestation of abdominal cancers, with a poor prognosis and limited treatment options. Recent work identifying high concentrations of the cytokine interleukin-6 (IL-6) and its soluble receptor (sIL-6-Rα) in the peritoneal cavity of patients with PC has highlighted this pathway as an emerging potential therapeutic target. This review article provides a comprehensive overview of the current understanding of the potential role of IL-6 in the development and progression of PC. We discuss mechansims by which the IL-6 pathway may contribute to peritoneal tumor dissemination, mesothelial adhesion and invasion, stromal invasion and proliferation, and immune response modulation. Finally, we review the prospects for targeting the IL-6 pathway in the treatment of PC, focusing on common sites of origin, including ovarian, gastric, pancreatic, colorectal and appendiceal cancer, and mesothelioma.

CD4+CCR8+ Tregs in ovarian cancer: a potential effector Tregs for immune regulation

Abstract Background Tregs are key drivers of immunosuppression in solid tumors. As an important chemokine receptor on Tregs, the regulatory effect of CCR8 on tumor immunity has received more and more attention. However, the current research on CCR8 in the immune microenvironment of ovarian cancer has not been clear. Methods Bioinformatics analysis was used to compare the transcriptome differences between CD4+ T cells in the peripheral circulation and infiltrated in ovarian tumor tissues. RT-PCR was used to detect the expression levels of chemokine receptor-related differential genes on CD4+ T cells in peripheral blood and ovarian tumor tissues. Multiparameter flow cytometry was used to detect the proportion and phenotypic characteristics of CD4+CCR8+ Tregs and CD4+CCR8− Tregs in different sample types. The expression level of CCR8 ligands was detected at multiple levels. To explore the important role of CCR8-CCL1 and CCR8-CCL18 axis in the migration and invasion of CD4+CCR8+ Tregs into ovarian tumor tissues by establishing a chemotaxis system in vitro. Results In this study, significantly different gene expression profiles were found between peripheral circulating CD4+ T cells and infiltrating CD4+ T cells in ovarian tumor tissues, in which chemokine-chemokine receptor signaling pathway was significantly enriched in all three groups of differential genes. The expression level of CCR8 in infiltrating CD4+ T cells of ovarian cancer tissue was significantly higher than that in peripheral blood of healthy controls and ovarian cancer patients, and high expression of CCR8 was significantly correlated with advanced tumor stage and poor differentiation. CD4+CCR8+ Tregs are the main type of infiltrating CD4+ Tregs in ovarian tumor tissues, which have stronger immunosuppressive phenotypes, secrete more inhibitory cytokines and have stronger proliferation ability. The ligands CCL1 and CCL18 corresponding to CCR8 were significantly overexpressed in ovarian tumor tissues, and the CCR8-CCL1 and CCR8-CCL18 axis played a key role in the migration and infiltration of CD4+CCR8+ Tregs into ovarian tumor tissues. Conclusions The results of this study may help to understand the phenotypic characteristics and recruitment process of Tregs in the tumor, and provide new ideas for improving the immunosuppressive status of the ovarian cancer microenvironment.

Kynurenine in IDO1high cancer cell-derived extracellular vesicles promotes angiogenesis by inducing endothelial mitophagy in ovarian cancer

Abstract Background Mitophagy, a prominent cellular homeostasis process, has been implicated in modulating endothelial cell function. Emerging evidence suggests that extracellular vesicles (EVs) participate in intercellular communication, which could modulate tumor angiogenesis, a hallmark of ovarian cancer (OC) progression. However, the underlying mechanisms through how EVs regulate endothelial mitophagy associated with tumor angiogenesis during OC development remain obscure. Methods The effect of cancer cell-derived EVs on endothelial mitophagy and its correlation with tumor angiogenesis and OC development were explored by in vitro and in vivo experiments. Multi-omics integration analysis was employed to identify potential regulatory mechanisms of cancer cell-derived EVs on endothelial mitophagy, which is involved in tumor angiogenesis associated with OC development. These insights were then further corroborated through additional experiments. An orthotopic OC mouse model was constructed to assess the antiangiogenic and therapeutic potential of the Indoleamine 2,3 dioxygenase-1 (IDO1) inhibitor. Results Cancer cell-derived EVs promoted tumor angiogenesis via the activation of endothelial mitophagy, contributing to the growth and metastasis of OC. The aberrantly high expression of IDO1 mediated abnormal tryptophan metabolism in cancer cells and promoted the secretion of l-kynurenine (L-kyn)-enriched EVs, with associated high levels of L-kyn in EVs isolated from both the tumor tissues and patient plasma in OC. EVs derived from IDO1high ovarian cancer cells elevated nicotinamide adenine dinucleotide (NAD +) levels in endothelial cells via delivering L-kyn. Besides, IDO1high ovarian cancer cell-derived EVs upregulated sirt3 expression in endothelial cells by increasing acetylation modification. These findings are crucial for promoting endothelial mitophagy correlated with tumor angiogenesis. Notably, both endothelial mitophagy and tumor angiogenesis could be suppressed by the IDO1 inhibitor in the orthotopic OC mouse model. Conclusions Together, our findings unveil a mechanism of mitophagy in OC angiogenesis and indicate the clinical relevance of EV enriched L-kyn as a potential biomarker for tumorigenesis and progression. Additionally, IDO1 inhibitors might become an alternative option for OC adjuvant therapy. Graphical Abstract

ANGPTL4 accelerates ovarian serous cystadenocarcinoma carcinogenesis and angiogenesis in the tumor microenvironment by activating the JAK2/STAT3 pathway and interacting with ESM1

Abstract Background Ovarian cancer (OC) is a malignant neoplasm that displays increased vascularization. Angiopoietin-like 4 (ANGPTL4) is a secreted glycoprotein that functions as a regulator of cell metabolism and angiogenesis and plays a critical role in tumorigenesis. However, the precise role of ANGPTL4 in the OC microenvironment, particularly its involvement in angiogenesis, has not been fully elucidated. Methods The expression of ANGPTL4 was confirmed by bioinformatics and IHC in OC. The potential molecular mechanism of ANGPTL4 was measured by RNA-sequence. We used a series of molecular biological experiments to measure the ANGPTL4-JAK2-STAT3 and ANGPTL4-ESM1 axis in OC progression, including MTT, EdU, wound healing, transwell, xenograft model, oil red O staining, chick chorioallantoic membrane assay and zebrafish model. Moreover, the molecular mechanisms were confirmed by Western blot, Co-IP and molecular docking. Results Our study demonstrates a significant upregulation of ANGPTL4 in OC specimens and its strong association with unfavorable prognosis. RNA-seq analysis affirms that ANGPTL4 facilitates OC development by driving JAK2-STAT3 signaling pathway activation. The interaction between ANGPTL4 and ESM1 promotes ANGPTL4 binding to lipoprotein lipase (LPL), thereby resulting in reprogrammed lipid metabolism and the promotion of OC cell proliferation, migration, and invasion. In the OC microenvironment, ESM1 may interfere with the binding of ANGPTL4 to integrin and vascular-endothelial cadherin (VE-Cad), which leads to stabilization of vascular integrity and ultimately promotes angiogenesis. Conclusion Our findings underscore that ANGPTL4 promotes OC development via JAK signaling and induces angiogenesis in the tumor microenvironment through its interaction with ESM1.

Prediction of complete regression in fertility-sparing patients with endometrial cancer and apical hyperplasia: the GLOBAL model in a large Chinese cohort

Abstract Background Fertility preservation treatment is increasingly essential for patients with apical endometrial hyperplasia (AEH) and early endometrial cancer (EEC) worldwide. Complete regression (CR) is the main endpoint of this treatment. Accurately predicting CR and implementing appropriate interventions during treatment are crucial for these patients. Methods We conducted a retrospective study involving 193 patients diagnosed with atypical AEH or EEC, enrolled from January 2012 to March 2022 at our center. We evaluated 24 clinical parameters as candidate predictors and employed LASSO regression to develop a prediction model for CR. Subsequently, a nomogram was constructed to predict CR after the treatment. We evaluated the performance of the nomogram using receiver operator characteristic (ROC) curve and decision curve analysis (DCA) to assess its predictive accuracy. Additionally, we employed cumulative curves to determine the CR rate among patients. Results Out of the 193 patients, 173 achieved CR after undergoing fertility preservation treatment. We categorized features with similar properties and provided a list of formulas based on their coefficients. The final model, named GLOBAL (including basic information, characteristics, blood pressure, glucose metabolism, lipid metabolism, immunohistochemistry, histological type, and medication), comprised eight variables identified using LASSO regression. A nomogram incorporating these eight risk factors was developed to predict CR. The GLOBAL model exhibited an AUC of 0.907 (95% CI 0.828–0.969). Calibration plots demonstrated a favorable agreement between the predicted probability by the GLOBAL model and actual observations in the cohort. The cumulative curve analysis revealed varying cumulative CR rates among patients in the eight subgroups. Categorized analysis demonstrated significant diversity in the effects of the GLOBAL model on CR among patients with different total points (p &lt; 0.05). Conclusion We have developed and validated a model that significantly enhances the predictive accuracy of CR in AEH and EEC patients seeking fertility preservation treatment.

Deep learning model for classifying endometrial lesions

Abstract Background Hysteroscopy is a commonly used technique for diagnosing endometrial lesions. It is essential to develop an objective model to aid clinicians in lesion diagnosis, as each type of lesion has a distinct treatment, and judgments of hysteroscopists are relatively subjective. This study constructs a convolutional neural network model that can automatically classify endometrial lesions using hysteroscopic images as input. Methods All histopathologically confirmed endometrial lesion images were obtained from the Shengjing Hospital of China Medical University, including endometrial hyperplasia without atypia, atypical hyperplasia, endometrial cancer, endometrial polyps, and submucous myomas. The study included 1851 images from 454 patients. After the images were preprocessed (histogram equalization, addition of noise, rotations, and flips), a training set of 6478 images was input into a tuned VGGNet-16 model; 250 images were used as the test set to evaluate the model’s performance. Thereafter, we compared the model’s results with the diagnosis of gynecologists. Results The overall accuracy of the VGGNet-16 model in classifying endometrial lesions is 80.8%. Its sensitivity to endometrial hyperplasia without atypia, atypical hyperplasia, endometrial cancer, endometrial polyp, and submucous myoma is 84.0%, 68.0%, 78.0%, 94.0%, and 80.0%, respectively; for these diagnoses, the model’s specificity is 92.5%, 95.5%, 96.5%, 95.0%, and 96.5%, respectively. When classifying lesions as benign or as premalignant/malignant, the VGGNet-16 model’s accuracy, sensitivity, and specificity are 90.8%, 83.0%, and 96.0%, respectively. The diagnostic performance of the VGGNet-16 model is slightly better than that of the three gynecologists in both classification tasks. With the aid of the model, the overall accuracy of the diagnosis of endometrial lesions by gynecologists can be improved. Conclusions The VGGNet-16 model performs well in classifying endometrial lesions from hysteroscopic images and can provide objective diagnostic evidence for hysteroscopists.

The intra-tumoral microbiome as a potential biomarker of response to external beam radiation therapy in cervical cancer

We aimed to determine the potential predictive value of the intra-tumoral microbiome as a marker of the response to external beam radiation therapy (EBRT) in cervical cancer (CC). A prospective longitudinal trial of 36 CC patients receiving pelvic radiotherapy was designed to investigate microbial characteristic signatures and diversity (alpha and beta) of multiple sites (tumor, vaginal, gut, urethral, and oral) in the superior response (SR) and inferior response (IR) groups of CC patients by 16S rRNA sequencing. Utilized the least absolute shrinkage and selection operator (LASSO) logistic regression method to analyze clinicopathological factors that potentially influenced the efficacy of EBRT. LEfSe analysis highlighted the microbiome features that best distinguished the categorized patient samples. Selected parameters were validated with Spearman correlation analysis, receiver operating characteristic (ROC) area under the curve (AUC) analysis and Kaplan-Meier survival analysis. Firstly, in our cohort, LASSO logistic regression analysis revealed no association between clinicopathological factors and EBRT efficacy. Subsequently, we employed 16S rRNA sequencing to compare microbiome differences across multiple sites and their correlations with major clinicopathological factors. We discovered that the intra-tumoral microbiome was independent of clinicopathologic features and represented the most direct and reliable reflection of the microbial differences between the SR and IR groups. We found lower alpha diversity in the tumor microbiome of SR group and identified the most relevant microbiome taxa (Bifidobacteriaceae, Beijerinckiaceae, and Orbaceae) associated with the efficacy of the response to EBRT in CC patients. We then conducted ROC analysis, finding that specific microbial taxa had an AUC of 0.831 (95% CI, 0.667-0.995), indicating the potential of these taxa as biomarkers for predicting EBRT efficacy. Kaplan-Meier survival analysis showed a better prognosis for patients with lower alpha diversity and higher relative abundance of Bifidobacteriaceae. Our data suggested that intra-tumoral specific microbiome taxa and lower alpha diversity may play an important role in the CC patient sensitivity to EBRT and offer novel potential biomarkers for predicting the response to EBRT efficacy.

Single-cell RNA sequencing highlights the unique tumor microenvironment of small cell neuroendocrine cervical carcinoma

Small cell neuroendocrine cervical carcinoma is a highly aggressive tumor characterized by early metastasis, a high recurrence rate, and poor prognosis. This study represents the first instance of single-cell sequencing conducted on small cell neuroendocrine carcinoma of the cervix worldwide. Analysis of gene expression regulatory networks revealed that the transcription factor TFF3 drived up-regulation of ELF3. Furthermore, our findings indicated that the neuroendocrine marker genes and gene regulatory networks associated with small cell neuroendocrine cervical carcinoma differed from those observed in lung, small intestine, and liver neuroendocrine carcinoma within the GEO database, suggesting tissue-specific origins for these malignancies. Overall, this study addresses a significant research in understanding small cell neuroendocrine cervical carcinoma in vivo and provides valuable insights for guiding radiotherapy, chemotherapy, and targeted therapy.

Targeting immune microenvironment in cervical cancer: current research and advances

The immune microenvironment plays a critical role in pathogenesis and treatment response of cervical cancer. This review comprehensively examines the cellular and molecular components of the tumor immune microenvironment (TIME) in cervical cancer, encompassing patterns of immune cell infiltration (T cells, B cells, NK cells, DCs, TAMs), immune checkpoint molecules, and cytokine/chemokine networks. We emphasize recent advances in understanding TIME heterogeneity, enabled by high-resolution spatial mapping and single-cell sequencing technologies, focusing specifically on differences across disease stages and treatment approaches. The review systematically evaluates immunotherapeutic strategies, such as immune checkpoint inhibitors, adoptive cell therapies, and therapeutic vaccines, discussing their mechanisms of action, clinical efficacy, and challenges. By synthesizing insights from both preclinical and clinical studies, our aim is to offer a translational perspective on targeting the TIME to enhance outcomes for cervical cancer patients.

The biomarker potential of circPOLD1 and its binding protein YBX1 in cervical carcinogenesis

Cervical cancer progresses through distinct precancerous stages, making early screening and intervention crucial for prevention. However, conventional screening modalities, such as cytology and HPV testing, face challenges related to sensitivity, specificity, and resource dependency. Circular RNAs (circRNAs), owing to their high stability and tissue-specific expression, have emerged as promising biomarkers, though their role in cervical carcinogenesis remains underexplored. In particular, the clinical utility of circRNAs for optimizing cervical cancer screening and early diagnosis has yet to be established. This study aimed to investigate the dynamic expression profiles of circRNAs across various stages of cervical cancer progression and identify potential biomarkers to enhance early detection. CircRNA sequencing was performed on cervical tissues spanning normal cervical epithelium (NCE), high-grade squamous intraepithelial lesions (HSIL), and cervical squamous cell carcinoma (CSCC). Functional assays, including cell viability, colony formation, and apoptosis, were performed to assess the oncogenic potential of circPOLD1 and its interaction with YBX1 in cervical cancer cells. BaseScope and immunohistochemistry (IHC) were applied to tissue microarrays for clincial validation and ROC curve analysis evaluated the diagnostic performance of circPOLD1 in serum as a liquid biopsy marker. CircRNA profiling revealed a progressive increase in circPOLD1 expression from NCE to HSIL and CSCC. Mechanistically, circPOLD1 functioned as an oncogene by binding to and phosphorylating YBX1, activating the AKT/mTOR/HIF-1α pathway to enhance glycolysis-driven tumorigenesis. BaseScope and IHC confirmed the stage-specific elevation of circPOLD1 and YBX1 in cervical lesions. The circPOLD1/YBX1 multi-marker panel demonstrated superior diagnostic performance, achieving an AUC of 0.951 for LSIL+ and 0.817 for HSIL+ detection. Furthermore, serum circPOLD1 levels exhibited a progressive increase across disease stages, underscoring its potential as a non-invasive biomarker. circPOLD1 and YBX1 synergistically drive cervical carcinogenesis and exhibit stage-specific expression patterns. Their combined detection significantly enhanced the accuracy for cervical cancer screening and dynamic monitoring. The successful application of BaseScope and IHC highlights the immediate translational potential of these biomarkers, paving the way for refined risk stratification, improved therapeutic targeting, and reduced cervical cancer burden through early intervention.

Apoptotic cell-derived extracellular vesicles-MTA1 confer radioresistance in cervical cancer by inducing cellular dormancy

Radioresistance presents a major challenge in the treatment of cervical cancer (CC). Apoptotic tumor cells can create an "onco-regenerative niche," contributing to radioresistance. However, the intercellular signaling mechanisms mediating the transfer of radioresistance from apoptotic to surviving cancer cells remain unclear. The role of apoptotic tumor cell-derived extracellular vesicles (apoEVs) in mediating radioresistance was investigated through integrated bioinformatics and experimental approaches. The GSE236738 dataset was analyzed to identify potential regulators, with subsequent validation of apoEV-MTA1 function using in vitro and in vivo models. Mechanistic studies focused on caspase-3 activation, p-STAT1 signaling pathway, and dormancy-associated protein networks. Furthermore, therapeutic strategies targeting MTA1 and its downstream signaling were evaluated for radiosensitization potential. MTA1 was identified as a critical factor enriched in and transferred by apoEVs from apoptotic tumor cells to neighboring CC cells. Caspase-3 activation facilitated the nuclear export and encapsulation of MTA1 in apoEVs. Transferred MTA1 retained transcriptional activity, activated the p-STAT1 signaling pathway, and induced cellular dormancy via NR2F1, a key dormancy regulator, resulting in increased radioresistance. Knockdown of MTA1 in apoEVs or inhibition of p-STAT1 in recipient cells enhanced radiosensitivity. Furthermore, apoEV-MTA1 promoted tumor radioresistance and reduced survival rates in irradiated cervical cancer mouse model. This study demonstrates that apoEV-MTA1 confers radioresistance in CC by promoting cellular dormancy via the p-STAT1/NR2F1 signaling axis. Targeting this pathway could improve radiosensitivity and provide a promising therapeutic strategy for CC patients.

Molecule interacting with CasL-2 enhances tumor progression and alters radiosensitivity in cervical cancer

Cervical cancer is a common malignancy among women, and radiotherapy remains a primary treatment modality across all disease stages. However, resistance to radiotherapy frequently results in treatment failure, highlighting the need to identify novel therapeutic targets to improve clinical outcomes. The expression of molecule interacting with CasL-2 (MICAL2) was confirmed in cervical cancer tissues and cell lines through western blotting (WB) and immunohistochemistry (IHC). Siha and Hela cells were used to examine the regulatory and biological functions of MICAL2 via knockdown and overexpression experiments. Assays including MTT, colony formation, wound healing, transwell migration, and sphere formation were employed, along with WB analysis. DNA damage in irradiated cells with MICAL2 knockdown or overexpression was evaluated using the comet assay, while γ-H2AX and Rad51 protein levels were detected by WB. In vivo experiments validated the tumorigenic and radioresistance functions of MICAL2. Additionally, the relationship between MICAL2 expression and radiotherapy response was analyzed in 62 patients with cervical cancer by assessing tumor regression and MICAL2 levels six months post-treatment. MICAL2 expression was significantly elevated in cervical cancer tissues and cells. Functional analyses demonstrated that MICAL2 promotes cell proliferation, migration, and invasion by activating the MAPK and PI3K/AKT pathways, as confirmed through both in vitro and in vivo experiments. Silencing MICAL2 increased DNA damage, impeded DNA repair, and enhanced radiosensitivity. Among the 62 patients with cervical cancer, elevated MICAL2 expression was associated with a lower complete response rate to radiotherapy (25.6% vs. 60.9% in those with low expression), reduced progression-free survival, and advanced cancer stage (*p < 0.05). MICAL2 plays a critical role in tumor progression and radiotherapy resistance in cervical cancer. These findings provide a foundation for developing targeted therapies to improve treatment outcomes in this population.

Suppressive stroma-immune prognostic signature impedes immunotherapy in ovarian cancer and can be reversed by PDGFRB inhibitors

Abstract Background As the most lethal gynecologic cancer, ovarian cancer (OV) holds the potential of being immunotherapy-responsive. However, only modest therapeutic effects have been achieved by immunotherapies such as immune checkpoint blockade. This study aims to propose a generalized stroma-immune prognostic signature (SIPS) to identify OV patients who may benefit from immunotherapy. Methods The 2097 OV patients included in the study were significant with high-grade serous ovarian cancer in the III/IV stage. The 470 immune-related signatures were collected and analyzed by the Cox regression and Lasso algorithm to generalize a credible SIPS. Correlations between the SIPS signature and tumor microenvironment were further analyzed. The critical immunosuppressive role of stroma indicated by the SIPS was further validated by targeting the major suppressive stroma component (CAFs, Cancer-associated fibroblasts) in vitro and in vivo. With four machine-learning methods predicting tumor immune subtypes, the stroma-immune signature was upgraded to a 23-gene signature. Results The SIPS effectively discriminated the high-risk individuals in the training and validating cohorts, where the high SIPS succeeded in predicting worse survival in several immunotherapy cohorts. The SIPS signature was positively correlated with stroma components, especially CAFs and immunosuppressive cells in the tumor microenvironment, indicating the critical suppressive stroma-immune network. The combination of CAFs’ marker PDGFRB inhibitors and frontline PARP inhibitors substantially inhibited tumor growth and promoted the survival of OV-bearing mice. The stroma-immune signature was upgraded to a 23-gene signature to improve clinical utility. Several drug types that suppress stroma-immune signatures, such as EGFR inhibitors, could be candidates for potential immunotherapeutic combinations in ovarian cancer. Conclusions The stroma-immune signature could efficiently predict the immunotherapeutic sensitivity of OV patients. Immunotherapy and auxiliary drugs targeting stroma could enhance immunotherapeutic efficacy in ovarian cancer.

Development of a prognostic immune cell-based model for ovarian cancer using multiplex immunofluorescence

Ovarian cancer is the most lethal gynecological malignancy, often diagnosed at advanced stages with poor prognosis. The tumor microenvironment (TME) plays a critical role in disease progression and treatment response. This study aimed to construct a prognostic model for ovarian cancer patients by evaluating the tumor immune landscape using multiplex immunofluorescence (mIF) staining, which focused on the spatial distribution and interactions of immune cells within the TME. Formalin-fixed paraffin-embedded (FFPE) tissues from 129 ovarian cancer patients were analyzed using mIF to assess the expression of PD-L1(Programmed death-ligand 1, PD-L1), CD8(Cluster of Differentiation 8, CD8), TOX (Thymocyte Selection-Associated HMG Box, TOX), CD68(Cluster of Differentiation 68, CD68), and CK (Cytokeratin, CK). The Vectra Polaris quantitative pathology imaging system and Inform software were employed for image and spatial analysis. The LASSO Cox regression model was used for feature selection, and Kaplan-Meier survival analysis was performed to evaluate the prognostic significance of immune cell markers. A nomogram was developed to predict overall survival (OS) based on clinical parameters and the Immune Cell Related Prognostic Index (ICRPI). High percentages of CD8 + T cells, CD68 + macrophages, and CD68 + PD-L1 + macrophages were significantly associated with poor OS. Moreover, a high percentage of CD8 + T cells, CD68 + macrophages was significantly associated with poor disease-free survival (DFS). Spatial analysis revealed that a higher average count of CD68 + PD-L1 + macrophages within 30 μm of CD8 + T cells correlated with worse prognosis. The ICRPI model, incorporating CD68+, CD68 + PD-L1+, and spatial variables, effectively stratified patients into high- and low-risk groups, with high-risk patients showing significantly poorer OS. This study highlights the prognostic value of immune cell spatial distribution in ovarian cancer. The ICRPI model, integrating immune cell markers and spatial analysis, provides a novel framework for predicting patient outcomes. Further validation in prospective studies is warranted to confirm the clinical utility of this model.

Protein tyrosine phosphatase receptor type kappa (PTPRK) revisited: evolving insights into structure, function, and pathology

Vaginal microbial profile of cervical cancer patients receiving chemoradiotherapy: the potential involvement of Lactobacillus iners in recurrence

AbstractThe vaginal microbiome is an immune defense against reproductive diseases and can serve as an important biomarker for cervical cancer. However, the intrinsic relationship between the recurrence and the vaginal microbiome in patients with cervical cancer before and after concurrent chemoradiotherapy is poorly understood. Here, we analyzed 125 vaginal microbial profiles from a patient cohort of stage IB–IVB cervical cancer using 16S metagenomic sequencing and deciphered the microbial composition and functional characteristics of the recurrent and non-recurrent both before and after chemoradiotherapy. We demonstrated that the abundance of beneficial bacteria and stability of the microbial community in the vagina decreased in the recurrence group, implying the unique characteristics of the vaginal microbiome for recurrent cervical cancer. Moreover, using machine learning, we identified Lactobacillus iners as the most important biomarker, combined with age and other biomarkers (such as Ndongobacter massiliensis, Corynebacterium pyruviciproducens ATCC BAA-1742, and Prevotella buccalis), and could predict cancer recurrence phenotype before chemoradiotherapy. This study prospectively employed rigorous bioinformatics analysis and highlights the critical role of vaginal microbiota in post-treatment cervical cancer recurrence, identifying promising biomarkers with prognostic significance in the context of concurrent chemoradiotherapy for cervical cancer. The role of L. iners in determining chemoradiation resistance in cervical cancer warrants further detailed investigation. Our results expand our understanding of cervical cancer recurrence and help develop better strategies for prognosis prediction and personalized therapy.

CEBPG suppresses ferroptosis through transcriptional control of SLC7A11 in ovarian cancer

Abstract Background Ovarian cancer (OC) has high mortality and poor prognosis for lacking of specific biomarkers and typical clinical symptoms in the early stage. CEBPG is an important regulator in tumor development, yet it is unclear exactly how it contributes to the progression of OC. Methods TCGA and tissue microarrays with immunohistochemical staining (IHC) were used to examine CEBPG expression in OC. A variety of in vitro assays were conducted, including colony formation, proliferation, migration, and invasion. The orthotopic OC mouse model was established for in vivo studies. Ferroptosis was detected by observing mitochondrial changes with electron microscopy, detecting ROS expression, and detecting cell sensitivity to drugs by CCK8 assay. The interaction between CEBPG and SLC7A11 was confirmed by CUT&amp;Tag and dual luciferase reporter assays. Results A significantly higher expression level of CEBPG in OC when compared with benign tissues of ovary, and that high CEBPG expression level was also tightly associated with poor prognosis of patients diagnosed with OC, as determined by analysis of datasets and patient samples. Conversely, knockdown of CEBPG inhibited OC progression using experiments of OC cell lines and in vivo orthotopic OC-bearing mouse model. Importantly, CEBPG was identified as a new participator mediating ferroptosis evasion in OC cells using RNA-sequencing, which could contribute to OC progression. The CUT&amp;Tag and dua luciferase reporter assays further revealed the inner mechanism that CEBPG regulated OC cell ferroptosis through transcriptional control of SLC7A11. Conclusions Our findings established CEBPG as a novel transcriptional regulator of OC ferroptosis, with potential value in predicting clinical outcomes and as a therapeutic candidate.

SPOP promotes cervical cancer progression by inducing the movement of PD-1 away from PD-L1 in spatial localization

Abstract Background Metastasis is a major obstacle in the treatment of cervical cancer (CC), and SPOP-mediated regulatory effects are involved in metastasis. However, the mechanisms have not been fully elucidated. Methods Proteomic sequencing and SPOP immunohistochemistry (IHC) were performed for the pelvic lymph node (pLN)-positive and non-pLN groups of CC patients. The corresponding patients were stratified by SPOP expression level for overall survival (OS) and relapse-free survival (RFS) analysis. In vitro and in vivo tests were conducted to verify the causal relationship between SPOP expression and CC metastasis. Multiplex immunofluorescence (m-IF) and the HALO system were used to analyse the mechanism, which was further verified by in vitro experiments. Results SPOP is upregulated in CC with pLN metastasis and negatively associated with patient outcome. In vitro and in vivo, SPOP promotes CC proliferation and metastasis. According to m-IF and HALO analysis, SPOP may promote CC metastasis by promoting the separation of PD-1 from PD-L1. Finally, it was further verified that SPOP can achieve immune tolerance by promoting the movement of PD-1 away from PD-L1 in spatial location and function. Conclusion This study shows that SPOP can inhibit the immune microenvironment by promoting the movement of PD-1 away from PD-L1, thereby promoting pLN metastasis of CC and resulting in worse OS and RFS.

USP39 interacts with SIRT7 to promote cervical squamous cell carcinoma by modulating autophagy and oxidative stress via FOXM1

Abstract Background Sirtuin 7 (SIRT7) is an oncogene that promotes tumor progression in various malignancies, however, its role and regulatory mechanism in cervical squamous cell carcinoma (CSCC) is unknown. Herein, we attempted to investigate the functional role and molecular mechanism of SIRT7 underlying CSCC progression. Methods SIRT7 expression was evaluated in CSCC cells using various assays. We then used a series of function gain-and-loss experiments to determine the role of SIRT7 in CSCC progression. Furthermore, mechanism experiments were conducted to assess the interaction between SIRT7/USP39/FOXM1 in CSCC cells. Additionally, rescue assays were conducted to explore the regulatory function of USP39/FOXM1 in CSCC cellular processes. Results SIRT7 was highly expressed in CSCC patient tissues and cell lines. SIRT7 deficiency showed significant repression on the proliferation, and autophagy of CSCC cells in vitro and tumorigenesis in vivo. Similarly, apoptosis and ROS production in CSCC cells were accelerated after the SIRT7 knockdown. Moreover, SIRT7 and USP39 were found colocalized in the cell nucleus. Interestingly, SIRT7 was revealed to deacetylate USP39 to promote its protein stability in CSCC cells. USP39 protein was also verified to be upregulated in CSCC tissues and cells. USP39 silencing showed suppressive effects on CSCC cell growth. Mechanistically, USP39 was revealed to upregulate SIRT7 by promoting the transcriptional activity of FOXM1. Rescue assays also indicated that SIRT7 promoted autophagy and inhibited ROS production in CSCC cells by regulating USP39/FOXM1. Conclusion The SIRT7/USP39/FOXM1 positive feedback network regulates autophagy and oxidative stress in CSCC, thus providing a new direction for CSCC-targeted therapy. Graphical Abstract

Cytosine–phosphate–guanine oligodeoxynucleotides alleviate radiation-induced kidney injury in cervical cancer by inhibiting DNA damage and oxidative stress through blockade of PARP1/XRCC1 axis

Abstract Background Radiotherapy can cause kidney injury in patients with cervical cancer. This study aims to investigate the possible molecular mechanisms by which CpG-ODNs (Cytosine phosphate guanine-oligodeoxynucleotides) regulate the PARP1 (poly (ADP-ribose) polymerase 1)/XRCC1 (X-ray repair cross-complementing 1) signaling axis and its impact on radiation kidney injury (RKI) in cervical cancer radiotherapy. Methods The GSE90627 dataset related to cervical cancer RKI was obtained from the Gene Expression Omnibus (GEO) database. Bioinformatics databases and R software packages were used to analyze the target genes regulated by CpG-ODNs. A mouse model of RKI was established by subjecting C57BL/6JNifdc mice to X-ray irradiation. Serum blood urea nitrogen (BUN) and creatinine levels were measured using an automated biochemical analyzer. Renal tissue morphology was observed through HE staining, while TUNEL staining was performed to detect apoptosis in renal tubular cells. ELISA was conducted to measure levels of oxidative stress-related factors in mouse serum and cell supernatant. An in vitro cell model of RKI was established using X-ray irradiation on HK-2 cells for mechanism validation. RT-qPCR was performed to determine the relative expression of PARP1 mRNA. Cell proliferation activity was assessed using the CCK-8 assay, and Caspase 3 activity was measured in HK-2 cells. Immunofluorescence was used to determine γH2AX expression. Results Bioinformatics analysis revealed that the downstream targets regulated by CpG-ODNs in cervical cancer RKI were primarily PARP1 and XRCC1. CpG-ODNs may alleviate RKI by inhibiting DNA damage and oxidative stress levels. This resulted in significantly decreased levels of BUN and creatinine in RKI mice, as well as reduced renal tubular and glomerular damage, lower apoptosis rate, decreased DNA damage index (8-OHdG), and increased levels of antioxidant factors associated with oxidative stress (SOD, CAT, GSH, GPx). Among the CpG-ODNs, CpG-ODN2006 had a more pronounced effect. CpG-ODNs mediated the inhibition of PARP1, thereby suppressing DNA damage and oxidative stress response in vitro in HK-2 cells. Additionally, PARP1 promoted the formation of the PARP1 and XRCC1 complex by recruiting XRCC1, which in turn facilitated DNA damage and oxidative stress response in renal tubular cells. Overexpression of either PARP1 or XRCC1 reversed the inhibitory effects of CpG-ODN2006 on DNA damage and oxidative stress in the HK-2 cell model and RKI mouse model. Conclusion CpG-ODNs may mitigate cervical cancer RKI by blocking the activation of the PARP1/XRCC1 signaling axis, inhibiting DNA damage and oxidative stress response in renal tubule epithelial cells.

Circular RNA circ0001955 promotes cervical cancer tumorigenesis and metastasis via the miR-188-3p/NCAPG2 axis

Abstract Background Circular RNAs (circRNAs) are known to play a crucial role in a variety of malignancies. However, the precise role of circRNAs in cervical squamous cell carcinoma (CSCC) remains largely unknown. Methods The expression of circ0001955 was determined by real-time quantitative PCR and fluorescence in situ hybridization. To examine the effects of circ0001955 on CSCC metastasis and growth, functional experiments were conducted in vitro and in vivo. Mechanistically, nucleocytoplasmic separation, dual luciferase reporter assay, RNA antisense purification experiments, and rescue experiments were performed to confirm the interaction between circ0001955, miR-188-3p, and NCAPG2 in CSCC. Results Here, we demonstrated that a circRNA derived from the CSNK1G1 gene (circ0001955) is significantly upregulated in CSCC. The overexpression of circ0001955 promotes tumor proliferation and metastasis, whereas the knockdown of circ0001955 exerts the opposite effects. Mechanistically, circ0001955 competitively binds miR-188-3p and prevents miR-188-3p from reducing the levels of NCAPG2, activating the AKT/mTOR signaling pathway to induce epithelial mesenchymal transformation. Notably, the application of an inhibitor of mTOR significantly antagonized circ0001955-mediated CSCC tumorigenesis. Conclusion circ0001955 promotes CSCC tumorigenesis and metastasis via the miR-188-3p/NCAPG2 axis which would provide an opportunity to search new therapeutic targets for CSCC. Graphical Abstract

Niraparib exhibits a synergistic anti-tumor effect with PD-L1 blockade by inducing an immune response in ovarian cancer

Abstract Background Immune checkpoint blockades (ICBs) therapy showed limited efficacy in ovarian cancer management. Increasing evidence indicated that conventional and targeted therapies could affect tumor-associated immune responses and increase the effectiveness of immunotherapy. However, the effects of Niraparib, one of the poly (ADP) ribose polymerase (PARP) inhibitors, on the immune response remains unclear. Delineating the crosstalk between cytotoxic anticancer agents and cancer-associated immunity may lead to more efficient combinatorial strategies. Methods Programmed death ligand 1 (PD-L1) expression in human ovarian cancer cells after PARP inhibitors treatment was examined by western blotting (WB) and flow cytometry. The expression of poly ADP-ribose polymerase (PARP1), PD-L1, and CD8 in human ovarian cancer tissues was detected by immunohistochemistry(IHC). The effect of Niraparib and PD-L1 blockade in ovarian cancer progression was investigated in vivo. The changes of immune cells and cytokines in vitro and in vivo were detected by flow cytometry and enzyme-linked immunosorbent assay (ELISA). Changes of cGAS/STING signal pathway after Niraparib treatment were determined by WB, ELISA. Results Niraparib upregulated membrane PD-L1 and total PD-L1 expression in ovarian cancer cells and had a synergistic effect with PD-L1 blockade in vivo. In clinical patient samples, Niraparib augmented cytotoxic CD8+T cell proportion and function. In vivo and vitro, Niraparib can also increase the proportion of T cells and combined with PD-L1 blockade could further enhance the effect. Besides, Niraparib activated the cGAS-STING pathway, increasing the levels of cytokines such as CCL5 and CXCL10, which played a vital role in augmenting the infiltration and activation of cytotoxic T cells. Conclusions Niraparib could modulate the immune response via the activation of the cGAS/STING pathway, and combination with PD-L1 blockade could further enhance the effect. These results provide a sound theoretical basis for clinical treatment.

Down-regulation of PADI2 prevents proliferation and epithelial-mesenchymal transition in ovarian cancer through inhibiting JAK2/STAT3 pathway in vitro and in vivo, alone or in combination with Olaparib

Abstract Background Epithelial ovarian cancer (EOC) is the most lethal disease among female genital malignant tumors. Peptidylarginine deiminase type II(PADI II) has been shown to enhance a variety of cancers carcinogenesis, including ovarian cancer. The purpose of this study was to investigate the biological role of PADI2 in ovarian cancer (OC) and the relative mechanism. Methods Gene Expression Profiling Interactive Analysis (GEPIA) (https://gepia.pku.cn/) and ONCOMINE (https://www.oncomine.org/) were used to analyze PADI2 Gene Expression data. The survival curve for the PADI2 gene was generated by using the online Kaplan–Meier mapping site (https://www.kmplot.com/). We conducted MTT assay, cloning formation assay and EdU cell proliferation assay to detect the cell activity of PADI2 knockdown A2780 and SKOV3 ovarian cancer cells treated with Olaparib. Cell migration and invasion were observed by would healing and transwell assay. The pathway changes after the treatment of PADI2 were detected by transcriptome sequencing and western blot. The role of PADI2 combined with Olaparib treatment in vivo was studied in nude mouse model bearing ovarian cancer tumor. Results We investigated the role of PADI2 on EOC in vitro and in vivo. PADI2 was upregulated in ovarian cancer samples and high PADI2 expression was correlated with poor outcome. Downregulating PADI2 suppressed colony formation, proliferation, migration and invasion of A2780 and SKOV3 cells. Furthermore, downregulating PADI2 and Olaparib combination treatment attenuated the viability, migration and invasion of A2780 and SKOV3 cells. We identified differentially expressed genes in A2780-shPADI2 and SKOV3-shPADI2 cell by transcriptome sequencing analysis and verified that downregulating PADI2 and Olaparib combination treatment suppresses EMT and JAK2/STAT3 signaling pathway in A2780 and SKOV3 cells in vitro and in vivo. Conclusions Downregulation of PADI2 and Olaparib combination treatment attenuated the proliferation, migration and invasion of A2780 and SKOV3 cells by inhibiting the EMT through JAK2/STAT3 signaling pathway.

Patients with stage IV epithelial ovarian cancer: understanding the determinants of survival

Abstract Background The most appropriate management for patients with stage IV ovarian cancer remains unclear. Our objective was to understand the main determinants associated with survival and to discuss best surgical management. Methods Data of 1038 patients with confirmed ovarian cancer treated between 1996 and 2016 were extracted from maintained databases of 7 French referral gynecologic oncology institutions. Patients with stage IV diseases were selected for further analysis. The Kaplan–Meier method was used to estimate the survival distribution. A Cox proportional hazards model including all the parameters statistically significant in univariable analysis, was used to account for the influence of multiple variables. Results Two hundred and eight patients met our inclusion criteria: 65 (31.3%) never underwent debulking surgery, 52 (25%) underwent primary debulking surgery (PDS) and 91 (43.8%) neoadjuvant chemotherapy and interval debulking surgery (NACT-IDS). Patients not operated had a significantly worse overall survival than patients that underwent PDS or NACT–IDS (p &lt; 0.001). In multivariable analysis, three factors were independent predictors of survival: upfront surgery (HR 0.32 95% CI 0.14–0.71, p = 0.005), postoperative residual disease = 0 (HR 0.37 95% CI 0.18–0.75, p = 0.006) and association of Carboplatin and Paclitaxel regimen (HR 0.45 95% CI 0.25–0.80, p = 0.007). Conclusions Presence of distant metastases should not refrain surgeons from performing radical procedures, whenever the patient is able to tolerate. Maximal surgical efforts should be done to minimize residual disease as it is the main determinant of survival.

FCGBP promotes ovarian cancer progression via activation of IL-6/JAK-STAT signaling pathway

Ovarian cancer is among the deadliest gynecological malignancies, primarily due to late-stage diagnosis and poor prognosis. Novel biomarkers and therapeutic targets are urgently needed to enhance early detection and treatment efficacy. Fc fragment of IgG-binding protein (FCGBP), a mucin-like glycoprotein, has been associated with various cancers, but its specific role in ovarian cancer progression has not been well-defined. This study aimed to investigate the clinical relevance, functional role, and underlying mechanisms of FCGBP in ovarian cancer progression. Gene expression profiles from multiple public datasets were analyzed to identify differentially expressed genes. Weighted gene co-expression network analysis was performed to correlate FCGBP expression with clinical traits. Single-cell RNA sequencing and pseudotime trajectory analyses were used to examine FCGBP expression dynamics. FCGBP expression was validated in ovarian cancer tissues using quantitative PCR, western blotting, and immunohistochemistry. Functional assays, including proliferation, migration, invasion, and colony formation, were conducted in SKOV3 and ES-2 ovarian cancer cell lines with FCGBP knockdown. The molecular mechanism was explored using dual-luciferase reporter assays and co-immunoprecipitation. Enzyme-linked immunosorbent assays and western blotting assessed cytokine levels and pathway activation. An in vivo xenograft mouse model was used to evaluate tumorigenic effects. FCGBP expression was significantly elevated in ovarian cancer tissues and correlated with advanced tumor stage and poor prognosis. Single-cell analysis showed FCGBP expression peaked in terminally differentiated epithelial cancer cells. Silencing FCGBP significantly reduced proliferation, migration, invasion, and colony formation in vitro, and suppressed tumor growth and improved survival in vivo. Mechanistically, FCGBP enhanced interleukin-6 expression by interacting with NF-kappaB subunit p65, leading to activation of the JAK-STAT signaling pathway. Rescue experiments confirmed that exogenous interleukin-6 could restore the tumor-promoting effects lost upon FCGBP knockdown. Our findings establish FCGBP as a crucial oncogenic regulator in ovarian cancer, acting through the IL-6-mediated activation of the JAK-STAT signaling pathway. FCGBP holds promise as a novel diagnostic biomarker and therapeutic target, potentially improving early diagnosis, prognosis, and management of ovarian cancer.

Single-cell analysis revealed that IL4I1 promoted ovarian cancer progression

Abstract Background Ovarian cancer was one of the leading causes of female deaths. Patients with OC were essentially incurable and portends a poor prognosis, presumably because of profound genetic heterogeneity limiting reproducible prognostic classifications. Methods We comprehensively analyzed an ovarian cancer single-cell RNA sequencing dataset, GSE118828, and identified nine major cell types. Relationship between the clusters was explored with CellPhoneDB. A malignant epithelial cluster was confirmed using pseudotime analysis, CNV and GSVA. Furthermore, we constructed the prediction model (i.e., RiskScore) consisted of 10 prognosis-specific genes from 2397 malignant epithelial genes using the LASSO Cox regression algorithm based on public datasets. Then, the prognostic value of Riskscore was assessed with Kaplan–Meier survival analysis and time-dependent ROC curves. At last, a series of in-vitro assays were conducted to explore the roles of IL4I1, an important gene in Riskscore, in OC progression. Results We found that macrophages possessed the most interaction pairs with other clusters, and M2-like TAMs were the dominant type of macrophages. C0 was identified as the malignant epithelial cluster. Patients with a lower RiskScore had a greater OS (log-rank P &lt; 0.01). In training set, the AUC of RiskScore was 0.666, 0.743 and 0.809 in 1-year, 3-year and 5-year survival, respectively. This was also validated in another two cohorts. Moreover, downregulation of IL4I1 inhibited OC cells proliferation, migration and invasion. Conclusions Our work provide novel insights into our understanding of the heterogeneity among OCs, and would help elucidate the biology of OC and provide clinical guidance in prognosis for OC patients.

Extracellular vesicle-encapsulated microRNA-424 exerts inhibitory function in ovarian cancer by targeting MYB

AbstractBackgroundRecent studies have suggested a crucial role of mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) in ovarian cancer treatment. We, therefore, set out to explore the mechanism through which MSC-derived EVs delivered microRNA-424 (miR-424) to influence the development of ovarian cancer.MethodsBioinformatics analyses were first performed to screen ovarian cancer-related differentially expressed genes and to predict regulatory miRNAs. Then, dual-luciferase reporter gene assay was carried out to verify the relationship between miR-424 and MYB. Subsequently, the characterized MSCs and isolated EVs were co-cultured with ovarian cancer cells, followed by determination of the expression patterns of miR-424, MYB, vascular endothelial growth factor (VEGF), and VEGF receptor (VEGFR), respectively. In addition, the effects of EVs-delivered miR-424 on the proliferation, migration, invasion and tube formation of ovarian cancer cells were assessed using gain- and loss-of-function approaches. Lastly, tumor xenograft was induced in nude mice to illustrate the influence of EVs-loaded miR-424 on ovarian cancer in vivo.ResultsOur data exhibited that MYB was highly-expressed and miR-424 was poorly-expressed in ovarian cancer. More importantly, MYB was identified as a target gene of miR-424. Additionally, the transfer of miR-424 by MSC-derived EVs was found to repress the proliferation, migration, and invasion of ovarian cancer cells, with a reduction in the expressions of VEGF and VEGFR. Furthermore, MSC-derived EVs over-expressing miR-424 could inhibit the proliferation, migration, and tube formation of human umbilical vein endothelial cells, and also suppressed tumorigenesis and angiogenesis of ovarian tumors in vivo.ConclusionCollectively, our findings indicate that MSC-derived EVs transfer miR-424 to down-regulate MYB, which ultimately led to the inhibition of the tumorigenesis and angiogenesis of ovarian cancer. Hence, this study offers a potential prognostic marker and a therapeutic target for ovarian cancer.

Platinum-resistance in epithelial ovarian cancer: an interplay of epithelial–mesenchymal transition interlinked with reprogrammed metabolism

Abstract Background Epithelial ovarian cancer is the most lethal gynaecological cancer worldwide. Chemotherapy resistance represents a significant clinical challenge and is the main reason for poor ovarian cancer prognosis. We identified novel expression of markers related to epithelial mesenchymal transitions (EMT) in a carboplatin resistant ovarian cancer cell line by proteomics. This was validated in the platinum resistant versus sensitive parental cell lines, as well as platinum resistant versus sensitive human ovarian cancer patient samples. The prognostic significance of the different proteomics-identified marker proteins in prognosis prediction on survival as well as their correlative association and influence on immune cell infiltration was determined by public domain data bases. Methods We explored the proteomic differences between carboplatin-sensitive OVCAR5 cells (parental) and their carboplatin-resistant counterpart, OVCAR5 CBPR cells. qPCR and western blots were performed to validate differentially expressed proteins at the mRNA and protein levels, respectively. Association of the identified proteins with epithelial–mesenchymal transition (EMT) prompted the investigation of cell motility. Cellular bioenergetics and proliferation were studied to delineate any biological adaptations that facilitate cancer progression. Expression of differentially expressed proteins was assessed in ovarian tumors obtained from platinum-sensitive (n = 15) versus platinum-resistant patients (n = 10), as well as matching tumors from patients at initial diagnosis and following relapse (n = 4). Kaplan–Meier plotter and Tumor Immune Estimation Resource (TIMER) databases were used to determine the prognostic significance and influence of the different proteomics-identified proteins on immune cell infiltration in the tumor microenvironment (TME). Results Our proteomics study identified 2422 proteins in both cell lines. Of these, 18 proteins were upregulated and 14 were downregulated by ≥ twofold (p &lt; 0.05) in OVCAR5 CBPR cells. Gene ontology enrichment analysis amongst upregulated proteins revealed an overrepresentation of biological processes consistent with EMT in the resistant cell line. Enhanced mRNA and/or protein expression of the identified EMT modulators including ITGA2, TGFBI, AKR1B1, ITGAV, ITGA1, GFPT2, FLNA and G6PD were confirmed in OVCAR5 CBPR cells compared to parental OVCAR5 cell line. Consistent with the altered EMT profile, the OVCAR5 CBPR cells demonstrated enhanced migration and reduced proliferation, glycolysis, and oxidative phosphorylation. The upregulation of G6PD, AKR1B1, ITGAV, and TGFβ1 in OVCAR5 CBPR cells was also identified in the tumors of platinum-resistant compared to platinum-sensitive high grade serous ovarian cancer (HGSOC) patients. Matching tumors of relapsed versus newly diagnosed HGSOC patients also showed enhanced expression of AKR1B1, ITGAV, TGFβ1 and G6PD protein in relapsed tumors. Among the identified proteins, significant enhanced expression of GFPT2, FLNA, TGFBI (CDGG1), ITGA2 predicted unfavorable prognosis in ovarian cancer patients. Further analysis suggested that the expression of TGFBI to correlate positively with the expression of identified and validated proteins such as GFPT2, FLNA, G6PD, ITGAV, ITGA1 and ITGA2; and with the infiltration of CD8+ T cells, macrophages, neutrophils, and dendritic cells in the TME. Conclusions Our research demonstrates proteomic-based discovery of novel EMT-related markers with an altered metabolic profile in platinum-resistant versus sensitive ovarian cancer cell lines. The study also confirms the expression of selected identified markers in the tumors of platinum-resistant versus sensitive, and in matching relapsed versus newly diagnosed HGSOC patients. The study provides insights into the metabolic adaptation of EMT-induced carboplatin resistant cells that confers on them reduced proliferation to provide effective migratory advantage; and the role of some of these identified proteins in ovarian cancer prognosis. These observations warrant further investigation of these novel target proteins in platinum-resistant patients.

Hsa_circ_0000520 overexpression increases CDK2 expression via miR-1296 to facilitate cervical cancer cell proliferation

AbstractBackgroundCircular RNA (circRNA) has been demonstrated to participate in cervical cancer development. In this study, we analyzed the role of hsa_circ_0000520 in cervical cancer.MethodsFifty-two pairs of cervical cancer and adjacent normal tissue samples were collected, and five human cervical cancer cell lines were obtained followed by the detection of hsa_circ_0000520 expression. Nuclear-cytoplasmic isolation and fluorescence in situ hybridization were performed to analyze the subcellular localization of hsa_circ_0000520 while linear RNA was digested by RNase R. Gain- or loss-of function experiments on hsa_circ_0000520 were performed, followed by detection of cell proliferation and cell cycle by EdU, Cell Counting Kit-8, colony formation assay, and flow cytometry respectively.ResultsHsa_circ_0000520 and cyclin-dependent kinase 2 (CDK2) were highly expressed in cervical cancer tissues. Binding sites between microRNA-1296 (miR-1296) and hsa_circ_0000520 or CDK2 were verified. Antibody to Argonaute 2 (Ago2) could precipitate hsa_circ_0000520, indicating that hsa_circ_0000520 could competitively bind to miR-1296 via Ago2. Silencing hsa_circ_0000520 inhibited cervical cancer cell proliferation and promoted the inhibitory effects of miR-1296 on CDK2, thereby blocking cell cycle progression and promoting apoptosis.ConclusionThese results support the premise that targeting hsa_circ_0000520 can be a potential approach to combat cervical cancer.

Blocking TIGIT/CD155 signalling reverses CD8+ T cell exhaustion and enhances the antitumor activity in cervical cancer

AbstractObjectiveTIGIT/CD155 has attracted widespread attention as a new immune checkpoint and a potential target for cancer immunotherapy. In our study, we evaluated the role of TIGIT/CD155 checkpoints in the progression of cervical cancer.MethodsThe expression of CD155 and TIGIT in cervical cancer tissues was detected using flow cytometry, immunohistochemistry (IHC) and gene expression profiling. In vivo and in vitro experiments have proven that blocking TIGIT/CD155 restores the ability of CD8+T cells to produce cytokines. Changes in the NF-κB and ERK pathways were detected using western blotting (WB) after blocking TIGIT/CD155 signalling.ResultsTIGIT expression was elevated in patients with cervical cancer. High TIGIT expression in CD8+T lymphocytes from patients with cervical cancer promotes the exhaustion of CD8+T lymphocytes. In addition, CD155 is expressed at high levels in cervical cancer tissues and is negatively correlated with the level of infiltrating CD8+T cells. We found that TIGIT, upon binding to CD155 and being phosphorylated, inhibited NF-κB and ERK activation by recruiting SHIP-1, resulting in the downregulation of cytokine production. Blocking TIGIT in activated CD8+T cells attenuates the inhibitory effect of SHIP-1 on CD8+T cells and enhances the activation of NF-κB and ERK. In vivo and in vitro experiments have proven that blocking TIGIT/CD155 restores the ability of CD8+T cells to produce cytokines. Injecting the blocking antibody TIGIT in vivo inhibits tumour growth and enhances CD8+T lymphocyte function. Treatment with a combination of TIGIT and PD-1 inhibitors further increases the efficacy of the TIGIT blocking antibody.ConclusionsOur research shows that TIGIT/CD155 is a potential therapeutic target for cervical cancer.

Clinical validation of full HR-HPV genotyping HPV Selfy assay according to the international guidelines for HPV test requirements for cervical cancer screening on clinician-collected and self-collected samples

Abstract Background According to international guidelines, Human Papillomavirus (HPV) DNA tests represent a valid alternative to Pap Test for primary cervical cancer screening, provided that they guarantee balanced clinical sensitivity and specificity for cervical intraepithelial neoplasia grade 2 or more (CIN2+) lesions. The study aimed to assess whether HPV Selfy (Ulisse BioMed – Trieste, Italy), a full-genotyping HPV DNA test that detects and differentiates 14 high-risk HPV (HR-HPV) types, meets the criteria for primary cervical cancer screening described in the international guidelines, on clinician-collected as well as on self-collected samples. Methods For each participant woman, consecutively referring to Azienda Sanitaria Universitaria Giuliano Isontina (Trieste, Italy) and CRO—National Cancer Institute (Aviano, Italy) for the cervical cancer screening program, the following samples were tested: (a) a clinician-collected cervical specimen, analyzed with the reference test (Hybrid Capture®2 test, HC2) and HPV Selfy; and (b) a self-collected vaginal sample, analyzed with HPV Selfy. Enrolled women were also asked to fulfill a questionnaire about self-sampling acceptability. As required by guidelines, a non-inferiority test was conducted to compare the clinical performance of the test under evaluation with its reference test. Results HPV Selfy clinical sensitivity and specificity resulted non-inferior to those of HC2. By analysis of a total of 889 cervical liquid-based cytology samples from a screening population, of which 98 were from women with CIN2+, HPV Selfy showed relative sensitivity and specificity for CIN2+ of 0.98 and 1.00 respectively (non-inferiority score test: P  = 0.01747 and P  = 0.00414, respectively); the test reached adequate intra- and inter-laboratory reproducibility. Moreover, we demonstrated that the performance of HPV Selfy on self-collected vaginal samples was non-inferior to the performance obtained on clinician-collected cervical specimen (0.92 relative sensitivity and 0.97 relative specificity). Finally, through HPV Selfy genotyping, we were able to describe HPV types prevalence in the study population. Conclusions HPV Selfy fulfills all the requirements of the international Meijer’s guidelines and has been clinically validated for primary cervical cancer screening purposes. Moreover, HPV Selfy has also been validated for self-sampling according to VALHUDES guidelines. Therefore, at date, HPV Selfy is the only full-genotyping test validated both for screening purposes and for self-sampling. Trial registration ASUGI Trieste n. 16008/2018; CRO Aviano n.17149/2018

The genomic landscape of invasive stratified mucin-producing carcinoma of the uterine cervix: the first description based on whole-exome sequencing

Unraveling the role of M2 TAMs in ovarian cancer dynamics: a systematic review

Tumor-associated macrophages (TAMs) represent the dominant immune cell subset within the ovarian tumor microenvironment, which exhibits remarkable plasticity. They can readily undergo polarization toward an immunosuppressive M2-like phenotype in response to various factors secreted by tumor cells, playing a pivotal role in ovarian cancer advancement and the development of resistance to chemotherapy. M2 TAMs promote the invasiveness of ovarian cancer cells and their resistance to therapeutic agents through the secretion of numerous tumor-promoting factors, including cytokines, chemokines, enzymes, and exosomes. Furthermore, M2 TAMs significantly contribute to the peritoneal metastasis of cancer cells by aiding in the formation of spheroids and facilitating adhesion at metastatic sites. Concurrently, TAMs can suppress immune responses by interacting with lymphocytes, natural killer cells, and dendritic cells, thereby fostering an immunosuppressive milieu. Extensive research has indicated that M2-like TAMs exert detrimental effects in ovarian tumors, with elevated levels correlating positively with diminished overall survival rates. This review seeks to summarize the existing knowledge regarding the mechanisms through which TAMs affect both the progression of ovarian cancer and the development of chemotherapy resistance, underscoring the necessity for innovative therapeutic approaches aimed at targeting these cells within the tumor microenvironment.

Patient-derived organoids predict responses to chemotherapy and PARP inhibitors in advanced ovarian cancer

Transmembrane protein TMEM98 as a multifunctional regulator in cancer: from signaling pathways to translational implications

Role of tumor microenvironment in ovarian cancer metastasis and clinical advancements

Abstract Ovarian cancer (OC) is the most lethal gynecological malignancy worldwide, characterized by heterogeneity at the molecular, cellular and anatomical levels. Most patients are diagnosed at an advanced stage, characterized by widespread peritoneal metastasis. Despite optimal cytoreductive surgery and platinum-based chemotherapy, peritoneal spread and recurrence of OC are common, resulting in poor prognoses. The overall survival of patients with OC has not substantially improved over the past few decades, highlighting the urgent necessity of new treatment options. Unlike the classical lymphatic and hematogenous metastasis observed in other malignancies, OC primarily metastasizes through widespread peritoneal seeding. Tumor cells (the “seeds”) exhibit specific affinities for certain organ microenvironments (the “soil”), and metastatic foci can only form when there is compatibility between the “seeds” and “soil.” Recent studies have highlighted the tumor microenvironment (TME) as a critical factor influencing the interactions between the “seeds” and “soil,” with ascites and the local peritoneal microenvironment playing pivotal roles in the initiation and progression of OC. Prior to metastasis, the interplay among tumor cells, immunosuppressive cells, and stromal cells leads to the formation of an immunosuppressive pre-metastatic niche in specific sites. This includes characteristic alterations in tumor cells, recruitment and functional anomalies of immune cells, and dysregulation of stromal cell distribution and function. TME-mediated crosstalk between cancer and stromal cells drives tumor progression, therapy resistance, and metastasis. In this review, we summarize the current knowledge on the onset and metastatic progression of OC. We provide a comprehensive discussion of the characteristics and functions of TME related to OC metastasis, as well as its association with peritoneal spread. We also outline ongoing relevant clinical trials, aiming to offer new insights for identifying potential effective biomarkers and therapeutic targets in future clinical practice.

LINC02888 promotes HGSOC progression and immune evasion via PPIB-mediated stabilization of LAPTM5 mRNA and inhibition of RIG-I-like receptor signaling

High-grade serous ovarian cancer (HGSOC) remains highly lethal due to late diagnosis and chemoresistance. Long noncoding RNAs (lncRNAs) have emerged as critical regulators of tumor progression. This study investigates the role of the cytoplasmic lncRNA LINC02888 in HGSOC. We identified overexpression of LINC02888 in HGSOC tissues and cell lines through whole-transcriptome sequencing, qPCR, in situ hybridization, and analyses of TCGA/GTEx datasets. Functional assays including CCK-8, EdU, colony formation, wound healing, Transwell migration/invasion, and flow cytometry were performed after modulating LINC02888 expression. RNA pull-down coupled with mass spectrometry identified the RNA-binding protein PPIB as an interactor of LINC02888. RNA-seq following both LINC02888 and PPIB knockdown, along with Western blot, qPCR analyses and functional assays, revealed that LAPTM5 is a downstream effector. Furthermore, RIP-qPCR, deletion mapping, and ActD assays confirmed that LINC02888 binds PPIB, which stabilizes LAPTM5 mRNA via its 3'UTR. The effect on the RIG-I-like receptor pathway was evaluated by measuring the expression levels of RIG-I, IRF7, and ISG15. Finally, in vivo, the critical role of the LINC02888/PPIB/ LAPTM5 axis in ovarian tumor progression were explored by axillary subcutaneous injection of specifically transfected OVCAR3 cells into nude mice. LINC02888 was significantly upregulated in HGSOC tissues and correlated with advanced disease and poor prognosis. Silencing LINC02888 inhibited proliferation, migration, and invasion, while inducing apoptosis; conversely, overexpression promoted these oncogenic behaviors. Mechanistically, LINC02888 interacts with PPIB to stabilize LAPTM5 mRNA via its 3'UTR, resulting in elevated LAPTM5 protein levels and suppression of RIG-I-like receptor signaling. Rescue experiments confirmed the critical role of the LINC02888-PPIB-LAPTM5 axis in HGSOC progression in vivo and vitro. Our findings reveal that LINC02888 drives HGSOC progression through a mechanism involving PPIB-mediated stabilization of LAPTM5 mRNA, which suppresses RIG-I-like receptor signaling. It also appropriately suggests that inhibiting this innate immune pathway may contribute to an immunosuppressive tumor microenvironment, making the LINC02888-PPIB-LAPTM5 axis a promising therapeutic target for this aggressive malignancy.

Spatial transcriptome reveals histology-correlated immune signature learnt by deep learning attention mechanism on H&amp;E-stained images for ovarian cancer prognosis

Abstract Background The ability to predict the prognosis of patients with ovarian cancer can greatly improve disease management. However, the knowledge on the mechanism of the prediction is limited. We sought to deconvolute the attention feature learnt by a deep learning convolutional neural networks trained with whole-slide images (WSIs) of hematoxylin-and-eosin (H&amp;E)–stained tumor samples using spatial transcriptomic data. Methods In this study, 773 WSIs of H&amp;E-stained tumor sections from 335 patients with treatment naïve high-grade serous ovarian cancer who were included in The Cancer Genome Atlas (TCGA) Pan-Cancer study were used to train, and validate, and to test a ResNet101 CNN model modified with attention mechanism. WSIs from patients in an independent cohort were used to further evaluate the model. Results The prognostic value of the predicted H&amp;E-based survival scores from the trained model on patient survival was evaluated. The attention signals learnt by the model were then examined their correlation with immune signatures using spatial transcriptome. After validating the model with the testing datasets, pathway enrichment analysis showed that the H&amp;E—based survival score significantly correlated with certain immune signatures and this was validated spatially using spatial transcriptome data generated from ovarian cancer FFPE samples by correlating the selected signature and attention signal. Conclusions In conclusion, attention mechanism might be useful to identify regions for their specific immune activities. This could guide future pathological study for the useful immunological features that are important in modulating the prognosis of ovarian cancer patients.

Colonization by Porphyromonas gingivalis in cervical squamous cell carcinomas promotes metastasis through FimA/CD151/ITGB1 signaling

Cervicovaginal microbiota dysbiosis has been implicated in the progression of cervical cancer. Recent studies have reported an increased abundance of Porphyromonas species in the vaginal microbiota of cervical cancer patients. Porphyromonas gingivalis (P. gingivalis), a key periodontal pathogen, has been associated with adverse pregnancy outcomes and bacterial vaginosis; however, its potential role in the progression of cervical squamous cell carcinoma (CSCC) remains largely unexplored. We employed immunohistochemistry (IHC), 16S rRNA fluorescence in situ hybridization (FISH), and immunofluorescence to detect P. gingivalis in CSCC tissues. The association between P. gingivalis colonization and CSCC patient survival outcomes was assessed. Transwell and wound healing assays were used to assess the migration and invasion abilities of CSCC cells. In vivo xenograft mouse models were established to evaluate the impact of P. gingivalis on tumor growth and metastasis. Pull-down assays were employed to investigate interactions between the P. gingivalis fimbrial protein FimA and host cell membrane proteins. RNA sequencing and Western blotting were utilized to identify signaling pathways activated by P. gingivalis in host cells. Porphyromonas gingivalis was detected immunohistochemically in 63% of CSCC tumor tissues, with significantly higher colonization in tumors compared to adjacent nontumor tissues (P < 0.0001). The presence of P. gingivalis was significantly associated with advanced tumor stage (P < 0.01), distant metastasis (P < 0.05), lymph node metastasis (P < 0.01), and poor survival outcomes (P = 0.0257, HR = 3.167) in CSCC patients. P. gingivalis preferentially adhered to CSCC cells and promoted cell migration and invasion. In this study, animal models revealed that P. gingivalis promoted lung and lymph node metastasis in CSCC without affecting tumor growth. Pull-down assays revealed that FimA interacts with CD151 and integrin β1 (ITGB1), which are highly expressed in CSCC cells. Knockdown of CD151 and ITGB1 significantly reduced P. gingivalis adhesion to CSCC cells (P < 0.01) and suppressed its effects on cell migration and invasion (P< 0.05). P. gingivalis treatment activated the JNK/paxillin pathway and triggered actin cytoskeleton reorganization. This study identifies P. gingivalis as a tumor-associated bacterium that promotes CSCC metastasis through direct interaction between its fimbrial adhesin FimA and the host CD151/ITGB1 complex. This interaction activates JNK/paxillin signaling, induces cytoskeletal reorganization, and enhances the metastatic capacity of CSCC cells. Targeting this microbial-host interaction may provide a novel therapeutic intervention for P. gingivalis-driven CSCC metastasis.

Evaluation of pathological response to neoadjuvant chemotherapy in locally advanced cervical cancer

AbstractNeoadjuvant chemotherapy (NACT) is a viable therapeutic option for women diagnosed locally advanced cervical cancer (LACC). However, the factors influencing pathological response are still controversial. We collected pair specimens of 185 LACC patients before and after receiving NACT and conducted histological evaluation. 8 fresh tissues pre-treatment were selected from the entire cohort to conducted immune gene expression profiling. A novel pathological grading system was established by comprehensively assessing the percentages of viable tumor, inflammatory stroma, fibrotic stroma, and necrosis in the tumor bed. Then, 185 patients were categorized into either the good pathological response (GPR) group or the poor pathological response (PPR) group post-NACT, with 134 patients (72.4%, 134/185) achieving GPR. Increasing tumor-infiltrating lymphocytes (TILs) and tumor-infiltrating lymphocytes volume (TILV) pre-treatment were correlated with GPR, with TILV emerging as an independent predictive factor for GPR. Additionally, CIBERSORT analysis revealed noteworthy differences in the expression of immune makers between cPR and non-cPR group. Furthermore, a significantly heightened density of CD8 + T cells and a reduced density of FOXP3 + T cells were observed in GPR than PPR. Importantly, patients exhibiting GPR or inflammatory type demonstrated improved overall survival and disease-free survival. Notably, stromal type was an independent prognostic factor in multivariate analysis. Our study indicates the elevated TILV in pre-treatment specimens may predict a favorable response to NACT, while identifying stromal type in post-treatment specimens as an independent prognostic factor. Moreover, we proposed this pathological grading system in NACT patients, which may offer a more comprehensive understanding of treatment response and prognosis.

Co-evolution of vaginal microbiome and cervical cancer

Abstract Background Exploration of adaptive evolutionary changes at the genetic level in vaginal microbial communities during different stages of cervical cancer remains limited. This study aimed to elucidate the mutational profile of the vaginal microbiota throughout the progression of cervical disease and subsequently establish diagnostic models. Methods This study utilized a metagenomic dataset consisting of 151 subjects classified into four categories: invasive cervical cancer (CC) (n = 42), cervical intraepithelial neoplasia (CIN) (n = 43), HPV-infected (HPVi) patients without cervical lesions (n = 34), and healthy controls (n = 32). The analysis focused on changes in microbiome abundance and extracted information on genetic variation. Consequently, comprehensive multimodal microbial signatures associated with CC, encompassing taxonomic alterations, mutation signatures, and enriched metabolic functional pathways, were identified. Diagnostic models for predicting CC were established considering gene characteristics based on single nucleotide variants (SNVs). Results In this study, we screened and analyzed the abundances of 18 key microbial strains during CC progression. Additionally, 71,6358 non-redundant mutations were identified, predominantly consisting of SNVs that were further annotated into 25,773 genes. Altered abundances of SNVs and mutation types were observed across the four groups. Specifically, there were 9847 SNVs in the HPV-infected group and 14,892 in the CC group. Furthermore, two distinct mutation signatures corresponding to the benign and malignant groups were identified. The enriched metabolic pathways showed limited similarity with only two overlapping pathways among the four groups. HPVi patients exhibited active nucleotide biosynthesis, whereas patients with CC demonstrated a significantly higher abundance of signaling and cellular-associated protein families. In contrast, healthy controls showed a distinct enrichment in sugar metabolism. Moreover, biomarkers based on microbial SNV abundance displayed stronger diagnostic capability (cc.AUC = 0.87) than the species-level biomarkers (cc.AUC = 0.78). Ultimately, the integration of multimodal biomarkers demonstrated optimal performance for accurately identifying different cervical statuses (cc.AUC = 0.86), with an acceptable performance (AUC = 0.79) in the external testing set. Conclusions The vaginal microbiome exhibits specific SNV evolution in conjunction with the progression of CC, and serves as a specific biomarker for distinguishing between different statuses of cervical disease.

Relationship between vaginal and oral microbiome in patients of human papillomavirus (HPV) infection and cervical cancer

Abstract Background The aim of this study was to assess the microbial variations and biomarkers in the vaginal and oral environments of patients with human papillomavirus (HPV) and cervical cancer (CC) and to develop novel prediction models. Materials and methods This study included 164 samples collected from both the vaginal tract and oral subgingival plaque of 82 women. The participants were divided into four distinct groups based on their vaginal and oral samples: the control group (Z/KZ, n = 22), abortion group (AB/KAB, n = 17), HPV-infected group (HP/KHP, n = 21), and cervical cancer group (CC/KCC, n = 22). Microbiota analysis was conducted using full-length 16S rDNA gene sequencing with the PacBio platform. Results The vaginal bacterial community in the Z and AB groups exhibited a relatively simple structure predominantly dominated by Lactobacillus. However, CC group shows high abundances of anaerobic bacteria and alpha diversity. Biomarkers such as Bacteroides, Mycoplasma, Bacillus, Dialister , Porphyromonas , Anaerococcus , and Prevotella were identified as indicators of CC. Correlations were established between elevated blood C-reactive protein (CRP) levels and local/systemic inflammation, pregnancy, childbirth, and abortion, which contribute to unevenness in the vaginal microenvironment. The altered microbial diversity in the CC group was confirmed by amino acid metabolism. Oral microbial diversity exhibited an inverse pattern to that of the vaginal microbiome, indicating a unique relationship. The microbial diversity of the KCC group was significantly lower than that of the KZ group, indicating a link between oral health and cancer development. Several microbes, including Fusobacterium , Campylobacter , Capnocytophaga , Veillonella , Streptococcus , Lachnoanaerobaculum , Propionibacterium , Prevotella , Lactobacillus , and Neisseria, were identified as CC biomarkers. Moreover, periodontal pathogens were associated with blood CRP levels and oral hygiene conditions. Elevated oral microbial amino acid metabolism in the CC group was closely linked to the presence of pathogens. Positive correlations indicated a synergistic relationship between vaginal and oral bacteria. Conclusion HPV infection and CC impact both the vaginal and oral microenvironments, affecting systemic metabolism and the synergy between bacteria. This suggests that the use of oral flora markers is a potential screening tool for the diagnosis of CC.

Weighted gene coexpression network analysis and machine learning reveal oncogenome associated microbiome plays an important role in tumor immunity and prognosis in pan-cancer

Abstract Background For many years, the role of the microbiome in tumor progression, particularly the tumor microbiome, was largely overlooked. The connection between the tumor microbiome and the tumor genome still requires further investigation. Methods The TCGA microbiome and genome data were obtained from Haziza et al.’s article and UCSC Xena database, respectively. Separate WGCNA networks were constructed for the tumor microbiome and genomic data after filtering the datasets. Correlation analysis between the microbial and mRNA modules was conducted to identify oncogenome associated microbiome module (OAM) modules, with three microbial modules selected for each tumor type. Reactome analysis was used to enrich biological processes. Machine learning techniques were implemented to explore the tumor type-specific enrichment and prognostic value of OAM, as well as the ability of the tumor microbiome to differentiate TP53 mutations. Results We constructed a total of 182 tumor microbiome and 570 mRNA WGCNA modules. Our results show that there is a correlation between tumor microbiome and tumor genome. Gene enrichment analysis results suggest that the genes in the mRNA module with the highest correlation with the tumor microbiome group are mainly enriched in infection, transcriptional regulation by TP53 and antigen presentation. The correlation analysis of OAM with CD8+ T cells or TAM1 cells suggests the existence of many microbiota that may be involved in tumor immune suppression or promotion, such as Williamsia in breast cancer, Biostraticola in stomach cancer, Megasphaera in cervical cancer and Lottiidibacillus in ovarian cancer. In addition, the results show that the microbiome-genome prognostic model has good predictive value for short-term prognosis. The analysis of tumor TP53 mutations shows that tumor microbiota has a certain ability to distinguish TP53 mutations, with an AUROC value of 0.755. The tumor microbiota with high importance scores are Corallococcus, Bacillus and Saezia. Finally, we identified a potential anti-cancer microbiota, Tissierella, which has been shown to be associated with improved prognosis in tumors including breast cancer, lung adenocarcinoma and gastric cancer. Conclusion There is an association between the tumor microbiome and the tumor genome, and the existence of this association is not accidental and could change the landscape of tumor research.

A traditional gynecological medicine inhibits ovarian cancer progression and eliminates cancer stem cells via the LRPPRC–OXPHOS axis

Abstract Background Ovarian cancer (OC) is the most lethal malignant gynecological tumor type for which limited therapeutic targets and drugs are available. Enhanced mitochondrial oxidative phosphorylation (OXPHOS), which enables cell growth, migration, and cancer stem cell maintenance, is a critical driver of disease progression and a potential intervention target of OC. However, the current OXPHOS intervention strategy mainly suppresses the activity of the electron transport chain directly and cannot effectively distinguish normal tissues from cancer tissues, resulting in serious side effects and limited efficacy. Methods We screened natural product libraries to investigate potential anti-OC drugs that target OXPHOS. Additionally, LC-MS, qRT-PCR, western-blot, clonogenic assay, Immunohistochemistry, wound scratch assay, and xenograft model was applied to evaluate the anti-tumor mechanism of small molecules obtained by screening in OC. Results Gossypol acetic acid (GAA), a widely used gynecological medicine, was screened out from the drug library with the function of suppressing OXPHOS and OC progression by targeting the leucine-rich pentatricopeptide repeat containing (LRPPRC) protein. Mechanically, LRPPRC promotes the synthesis of OXPHOS subunits by binding to RNAs encoded by mitochondrial DNA. GAA binds to LRPPRC directly and induces LRPPRC rapid degradation in a ubiquitin-independent manner. LRPPRC was overexpressed in OC, which is highly correlated with the poor outcomes of OC and could promote the malignant phenotype of OC cells in vitro and in vivo. GAA management inhibits cell growth, clonal formation, and cancer stem cell maintenance in vitro, and suppresses subcutaneous graft tumor growth in vivo. Conclusions Our study identified a therapeutic target and provided a corresponding inhibitor for OXPHOS-based OC therapy. GAA inhibits OC progression by suppressing OXPHOS complex synthesis via targeting LRPPRC protein, supporting its potential utility as a natural therapeutic agent for ovarian cancer.

A patient stratification signature mirrors the immunogenic potential of high grade serous ovarian cancers

Abstract Background While high-grade serous ovarian cancer (HGSC) has proven largely resistant to immunotherapy, sporadic incidents of partial and complete response have been observed in clinical trials and case reports. These observations suggest that a molecular basis for effective immunity may exist within a subpopulation of HGSC. Herein, we developed an algorithm, CONSTRU (Computing Prognostic Marker Dependencies by Successive Testing of Gene-Stratified Subgroups), to facilitate the discovery and characterization of molecular backgrounds of HGSC that confer resistance or susceptibility to protective anti-tumor immunity. Methods We used CONSTRU to identify genes from tumor expression profiles that influence the prognostic power of an established immune cytolytic activity signature (CYTscore). From the identified genes, we developed a stratification signature (STRATsig) that partitioned patient populations into tertiles that varied markedly by CYTscore prognostic power. The tertile groups were then analyzed for distinguishing biological, clinical and immunological properties using integrative bioinformatics approaches. Results Patient survival and molecular measures of immune suppression, evasion and dysfunction varied significantly across STRATsig tertiles in validation cohorts. Tumors comprising STRATsig tertile 1 (S-T1) showed no immune-survival benefit and displayed a hyper-immune suppressed state marked by activation of TGF-β, Wnt/β-catenin and adenosine-mediated immunosuppressive pathways, with concurrent T cell dysfunction, reduced potential for antigen presentation, and enrichment of cancer-associated fibroblasts. By contrast, S-T3 tumors exhibited diminished immunosuppressive signaling, heightened antigen presentation machinery, lowered T cell dysfunction, and a significant CYTscore-survival benefit that correlated with mutational burden in a manner consistent with anti-tumor immunoediting. These tumors also showed elevated activity of DNA damage/repair, cell cycle/proliferation and oxidative phosphorylation, and displayed greater proportions of Th1 CD4 + T cells. In these patients, but not those of S-T1 or S-T2, validated predictors of immunotherapy response were prognostic of longer patient survival. Further analyses showed that STRATsig tertile properties were not explained by known HGSC molecular or clinical subtypes or singular immune mechanisms. Conclusions STRATsig is a composite of parallel immunoregulatory pathways that mirrors tumor immunogenic potential. Approximately one-third of HGSC cases classify as S-T3 and display a hypo-immunosuppressed and antigenic molecular composition that favors immunologic tumor control. These patients may show heightened responsiveness to current immunotherapies.

snRNA-seq of human ovaries reveals heat shock proteins are associated with obesity related cancer risk

Obesity significantly impacts female reproductive health and increases the risk of gynecological tumors. However, the specific transcriptional changes that occur in the ovarian microenvironment during obesity-induced stress and the relationship between obesity and ovarian cancer remain unclear. Our study investigated the single-cell landscape of the ovarian cortex in individuals with varying BMI levels by snRNA-seq, revealing weight-stage related cellular composition deviations and expression profile irregularities. Using single-cell high-dimensional Weighted Gene Co-expression Network Analysis (hdWGCNA), we identified distinct obesity-related gene modules within various subpopulations of stroma cells and blood vascular endothelial cells. Notably, we observed a negative correlation between BMI and heat shock protein (HSP) family genes. Specifically, we found that HSPD1 might function as a potential regulator of ovarian carcinogenesis and progression under conditions of obesity, as supported by our co-analysis with data from three bulk RNA-seq ovarian cancer databases. Our findings suggested that lower expression of HSPD1 indicated a poorer prognosis for ovarian cancer. Our study identified a cluster of genes in ovarian cells that are suppressed by obesity, including those belonging to HSP family genes. These findings provide valuable insights for investigating the link between obesity and ovarian diseases.

Stromal cartilage oligomeric matrix protein as a tumorigenic driver in ovarian cancer via Notch3 signaling and epithelial-to-mesenchymal transition

Abstract Background Cartilage oligomeric matrix protein (COMP), an extracellular matrix glycoprotein, is vital in preserving cartilage integrity. Further, its overexpression is associated with the aggressiveness of several types of solid cancers. This study investigated COMP’s role in ovarian cancer, exploring clinicopathological links and mechanistic insights. Methods To study the association of COMP expression in cancer cells and stroma with clinicopathological features of ovarian tumor patients, we analyzed an epithelial ovarian tumor cohort by immunohistochemical analysis. Subsequently, to study the functional mechanisms played by COMP, an in vivo xenograft mouse model and several molecular biology techniques such as transwell migration and invasion assay, tumorsphere formation assay, proximity ligation assay, and RT-qPCR array were performed. Results Based on immunohistochemical analysis of epithelial ovarian tumor tissues, COMP expression in the stroma, but not in cancer cells, was linked to worse overall survival (OS) of ovarian cancer patients. A xenograft mouse model showed that carcinoma-associated fibroblasts (CAFs) expressing COMP stimulate the growth and metastasis of ovarian tumors through the secretion of COMP. The expression of COMP was upregulated in CAFs stimulated with TGF-β. Functionally, secreted COMP by CAFs enhanced the migratory capacity of ovarian cancer cells. Mechanistically, COMP activated the Notch3 receptor by enhancing the Notch3-Jagged1 interaction. The dependency of the COMP effect on Notch was confirmed when the migration and tumorsphere formation of COMP-treated ovarian cancer cells were inhibited upon incubation with Notch inhibitors. Moreover, COMP treatment induced epithelial-to-mesenchymal transition and upregulation of active β-catenin in ovarian cancer cells. Conclusion This study suggests that COMP secretion by CAFs drives ovarian cancer progression through the induction of the Notch pathway and epithelial-to-mesenchymal transition.

Targeting mitochondria: a novel approach for treating platinum-resistant ovarian cancer

Ovarian cancer is a prevalent gynecologic malignancy with the second-highest mortality rate among gynecologic malignancies. Platinum-based chemotherapy is the first-line treatment for ovarian cancer; however, a majority of patients with ovarian cancer experience relapse and develop platinum resistance following initial treatment. Despite extensive research on the mechanisms of platinum resistance at the nuclear level, the issue of platinum resistance in ovarian cancer remains largely unresolved. It is noteworthy that mitochondrial DNA (mtDNA) exhibits higher affinity for platinum compared to nuclear DNA (nDNA). Mutations in mtDNA can modulate tumor chemosensitivity through various mechanisms, including DNA damage responses, shifts in energy metabolism, maintenance of Reactive Oxygen Species (ROS) homeostasis, and alterations in mitochondrial dynamics. Concurrently, retrograde signals produced by mtDNA mutations and their subsequent cascades establish communication with the nucleus, leading to the reorganization of the nuclear transcriptome and governing the transcription of genes and signaling pathways associated with chemoresistance. Furthermore, mitochondrial translocation among cells emerges as a crucial factor influencing the effectiveness of chemotherapy in ovarian cancer. This review aims to explore the role and mechanism of mitochondria in platinum resistance, with a specific focus on mtDNA mutations and the resulting metabolic reprogramming, ROS regulation, changes in mitochondrial dynamics, mitochondria-nucleus communication, and mitochondrial transfer.

Cytotoxic lymphocytes-related gene ITK from a systematic CRISPR screen could predict prognosis of ovarian cancer patients with distant metastasis

Abstract Background Ovarian cancer, a highly metastatic malignancy, has benefited tremendously from advances in modern human genomics. However, the genomic variations related to the metastasis remains unclear. Methods We filtered various significant genes (n = 6722) associated with metastasis within a large-scale functional genomic CRISPR/Cas9 knock-out library including 122,756 single guide RNAs, and identified ITK (IL2 Inducible T Cell Kinase) as a potential cancer suppressor gene for ovarian cancer metastasis. Downstream bioinformatic analysis was performed for ITK using public databases. Results We found that patients in low-ITK group had poor prognosis and more distant metastasis than those in high-ITK group in TCGA and GEO databases. We also demonstrated that ITK combined with the clinical factors could accurately predict prognosis through multiple Cox regression analysis and ROC analysis. Moreover, alterations correlated with distant metastasis emereged with significantly increased expression in SAMRCD1 in low-ITK group, but CD244 and SOCS1 in high-ITK group. Integrated analysis revealed dysregulated molecular processes including predominantly oncogenic signaling pathways in low-ITK group but immune related pathways in high-ITK group, which suggested ITK might inhibit distant metastasis in ovarian cancer. Furtherly, deconvolution of the cellular composition of all samples validated the close correlation between ITK and immune related function especially for cytotoxic lymphocytes. Conclusions Together, these data provide insights into the potential role of ITK, with implications for the future development of tansformative ovarian cancer therapeutics.

TMEM119 facilitates ovarian cancer cell proliferation, invasion, and migration via the PDGFRB/PI3K/AKT signaling pathway

Abstract Background Ovarian cancer (OV) is the deadliest gynecological cancer. Transmembrane protein 119 (TMEM119) has been reported as oncogene in several human cancers. However, the function of TMEM119 in OV is still poorly known. Methods Western blot and qRT-PCR were used to analyze TMEM119 levels. Transwell assays, wound healing assays, CCK-8 assays and EdU cell proliferation assays were designed to explore the function and potential mechanism of TMEM119 in malignant biological behaviors in OV. Results TMEM119 was observed to be overexpressed in OV tissues and associated with poor survival in OV patients. Knockdown and overexpression experiments demonstrated that TMEM119 promoted proliferation, invasion, and migration in OV cells in vitro. TMEM119 mRNA expression was related to the pathways of focal adhesion according to Gene Set Enrichment Analyses and was correlated with the mRNA expression level of platelet-derived growth factor receptor beta (PDGFRB). TMEM119 exerted oncogenic effects partially by regulating the expression of PDGFRB and by activating the PI3K/AKT signaling pathway. Conclusions Collectively, our findings highlight the potential role of TMEM119 in the malignant biological behavior of OV, which may serve as a potential biomarker and a therapeutic candidate for OV.

Novel and recurrent BRCA1/BRCA2 germline mutations in patients with breast/ovarian cancer: a series from the south of Tunisia

Abstract Background The incidence of breast cancer (BC) and/or ovarian cancer (OC) is increasing in Tunisia especially in young women and mostly those with family history. However, the spectrum of BRCA mutations remains little explored in Tunisian patients in particular in the southern region. Methods We sequenced the entire coding regions of BRCA1 and BRCA2 genes using next generation sequencing (NGS) in 134 selected patients with BC and/or OC. Results Among the 134 patients, 19 (14.17%) carried pathogenic mutations (10 are BRCA1 mutation carriers and 9 are BRCA2 mutation carriers) that are mainly frameshift index (76.9%). Interestingly, 5 out of the 13 variants (38.46%) were found at least twice in unrelated patients, as the c.1310-1313 delAAGA in BRCA2 and the c.5030_5033 delCTAA that has been identified in 4/98 BC patients and in 3/15 OC patients from unrelated families with strong history of cancer. Besides recurrent mutations, 6 variant (4 in BRCA1 and 2 in BRCA2 ) were not reported previously. Furthermore, 3 unrelated patients carried the VUS c.9976A &gt; T, (K3326*) in BRCA2 exon 27. BRCA carriers correlated significantly with tumor site (p = 0.029) and TNBC cases (p = 0.008). In the groups of patients aged between 31 and 40, and 41–50 years, BRCA1 mutations occurred more frequently in patients with OC than those with BC, and conversely BRCA2 carriers are mostly affected with BC (p = 0.001, and p = 0.044 respectively). Conclusions The overall frequency of the BRCA germline mutations was 14.17% in patients with high risk of breast/ovarian cancer. We identified recurrent mutations as the c.1310_1313 delAAGA in BRCA2 gene and the c.5030_5033 delCTAA in BRCA1 gene that were found in 4% and 20% of familial BC and OC respectively. Our data will contribute in the implementation of genetic counseling and testing for families with high-risk of BC and/or OC.

FOXO1 mitigates the SMAD3/FOXL2C134W transcriptomic effect in a model of human adult granulosa cell tumor

Abstract Background Adult granulosa cell tumor (aGCT) is a rare type of stromal cell malignant cancer of the ovary characterized by elevated estrogen levels. aGCTs ubiquitously harbor a somatic mutation in FOXL2 gene, Cys134Trp (c.402C &lt; G); however, the general molecular effect of this mutation and its putative pathogenic role in aGCT tumorigenesis is not completely understood. We previously studied the role of FOXL2 C134W , its partner SMAD3 and its antagonist FOXO1 in cellular models of aGCT. Methods In this work, seeking more comprehensive profiling of FOXL2 C134W transcriptomic effects, we performed an RNA-seq analysis comparing the effect of FOXL2 WT /SMAD3 and FOXL2 C134W /SMAD3 overexpression in an established human GC line (HGrC1), which is not luteinized, and bears normal alleles of FOXL2 . Results Our data shows that FOXL2 C134W /SMAD3 overexpression alters the expression of 717 genes. These genes include known and novel FOXL2 targets ( TGFB2 , SMARCA4 , HSPG2 , MKI67 , NFKBIA ) and are enriched for neoplastic pathways (Proteoglycans in Cancer, Chromatin remodeling, Apoptosis, Tissue Morphogenesis, Tyrosine Kinase Receptors). We additionally expressed the FOXL2 antagonistic Forkhead protein, FOXO1. Surprisingly, overexpression of FOXO1 mitigated 40% of the altered genome-wide effects specifically related to FOXL2 C134W , suggesting it can be a new target for aGCT treatment. Conclusions Our transcriptomic data provide novel insights into potential genes (FOXO1 regulated) that could be used as biomarkers of efficacy in aGCT patients.

A combination of molecular and clinical parameters provides a new strategy for high-grade serous ovarian cancer patient management

Abstract Background High-grade serous carcinoma (HGSC) is the most common and deadly subtype of ovarian cancer. Although most patients will initially respond to first-line treatment with a combination of surgery and platinum-based chemotherapy, up to a quarter will be resistant to treatment. We aimed to identify a new strategy to improve HGSC patient management at the time of cancer diagnosis (HGSC-1LTR). Methods A total of 109 ready-available formalin-fixed paraffin-embedded HGSC tissues obtained at the time of HGSC diagnosis were selected for proteomic analysis. Clinical data, treatment approach and outcomes were collected for all patients. An initial discovery cohort (n = 21) were divided into chemoresistant and chemosensitive groups and evaluated using discovery mass-spectrometry (MS)-based proteomics. Proteins showing differential abundance between groups were verified in a verification cohort (n = 88) using targeted MS-based proteomics. A logistic regression model was used to select those proteins able to correctly classify patients into chemoresistant and chemosensitive. The classification performance of the protein and clinical data combinations were assessed through the generation of receiver operating characteristic (ROC) curves. Results Using the HGSC-1LTR strategy we have identified a molecular signature (TKT, LAMC1 and FUCO) that combined with ready available clinical data (patients’ age, menopausal status, serum CA125 levels, and treatment approach) is able to predict patient response to first-line treatment with an AUC: 0.82 (95% CI 0.72–0.92). Conclusions We have established a new strategy that combines molecular and clinical parameters to predict the response to first-line treatment in HGSC patients (HGSC-1LTR). This strategy can allow the identification of chemoresistance at the time of diagnosis providing the optimization of therapeutic decision making and the evaluation of alternative treatment strategies. Thus, advancing towards the improvement of patient outcome and the individualization of HGSC patients’ care.

Whole-cell tumor vaccines desialylated to uncover tumor antigenic Gal/GalNAc epitopes elicit anti-tumor immunity

Abstract Background Aberrant sialoglycans on the surface of tumor cells shield potential tumor antigen epitopes, escape recognition, and suppress activation of immunocytes. α2,3/α2,6Gal- and α2,6GalNAc (Gal/GalNAc)-linked sialic acid residues of sialoglycans could affect macrophage galactose-type lectins (MGL) mediated-antigen uptake and presentation and promote sialic acid-binding immunoglobulin-like lectins (Siglecs) mediated-immunosuppression. Desialylating sialoglycans on tumor cells could present tumor antigens with Gal/GalNAc residues and overcome glyco-immune checkpoints. Thus, we explored whether vaccination with desialylated whole-cell tumor vaccines (DWCTVs) triggers anti-tumor immunity in ovarian cancer (OC). Methods Sialic acid (Sia) and Gal/GalNAc residues on OC A2780, OVCAR3, and ID8 cells treated with α2-3 neuraminidase (α2-3NA) and α2-6NA, and Sigec-9 or Siglec-E and MGL on DCs pulsed with desialylated OC cells were identified using flow cytometry (FCM); RT-qPCR determined IFNG expression of T cells, TRBV was sequenced using Sanger sequencing and cytotoxicity of αβ T cells was measured with LDH assay; Anti-tumor immunity in vivo was validated via vaccination with desialylated whole-cell ID8 vaccine (ID8 DWCTVs). Results Gal/GalNAc but not Sia residues were significantly increased in the desialylated OC cells. α2-3NA-modified DWCTV increased MGL but decreased Siglec-9 or Siglec E expression on DCs. MGLbright/Siglec-9dim DCs significantly up-regulated IFNG expression and CD4/CD8 ratio of T cells and diversified the TCR repertoire of αβ T-cells that showed enhanced cytotoxic activity. Vaccination with α2-3NA-modified ID8 DWCTVs increased MGLbright/Siglec-Edim DCs in draining lymph nodes, limited tumor growth, and extended survival in tumor-challenged mice. Conclusion Desialylated tumor cell vaccine could promote anti-tumor immunity and provide a strategy for OC immunotherapy in a clinical setting.

Associations between dietary fiber intake and mortality from all causes, cardiovascular disease and cancer: a prospective study

Abstract Objective Several studies suggest that dietary fiber intake may reduce mortality risk, but this might depend on the fiber types and the evidence regarding the role of soluble fiber or insoluble fiber on death risk remain limited and inconsistent. Therefore, this study aimed to comprehensively evaluate multiple types of dietary fiber intake on mortality from all causes, cardiovascular disease and cancer in the large-scale Prostate, Lung, Colorectal, and Ovarian Cancer (PLCO) Screening Trial. Methods A multivariate Cox proportional hazards model was used to estimate hazard ratios (HRs) and 95% confidence intervals (CIs). Results This study finally included 86,642 participants with 17,536 all-cause deaths, 4842 cardiovascular deaths and 5760 cancer deaths identified after a total of 1,444,068 follow-up years. After adjusting for potential confounders, dietary total fiber intake was statistically significantly inversely associated with all-cause death (Q5 vs Q1: HR 0.71, 95% CI 0.66–0.75; P for trend &lt; 0.001), cardiovascular death (Q5 vs Q1: HR 0.73, 95% CI 0.65–0.83; P for trend &lt; 0.001) and cancer mortality (Q5 vs Q1: HR 0.77, 95% CI 0.69–0.86; P for trend &lt; 0.001). Similar results were observed for both insoluble and soluble fiber intake. Restricted cubic spline model analysis suggested that there was a nonlinear association of dietary fiber intake with mortality risk (all P for nonlinearity &lt; 0.05). Conclusions In this large nationally representative sample of US adult population, intakes of total fiber, soluble fiber, and insoluble fiber were associated with lower risks of all-cause, cardiovascular and cancer mortality.

Establishment of a piglet model for peritoneal metastasis of ovarian cancer

Abstract Background A piglet model for peritoneal metastasis (PM) of ovarian cancer was developed. It will contribute to establishing innovative chemotherapeutical and surgical strategies without any limitation on rodent models. Methods A total of 12 four- to five-week-old piglets of 7 to 8 kg were used. Two phases of ovarian cancer cell injections were performed with laparoscopic surgery. In phase I trial, 5.0 × 106 SK-OV-3 cells in 0.1 ml suspension were inoculated into the omentum, peritoneum, and uterine horns of two piglets twice with a one-week interval. In the phase II trial, 5.0 × 106 SNU-008 cells in 0.1 ml suspension were injected only into uterine horns within the same time frame because tumor implantation after inoculation of SK-OV-3 cells was not observed at the omentum or peritoneum in the phase I trial. Modified peritoneal cancer index (PCI) score was used to monitor tumorigenesis up to 4 weeks after inoculation. Tumor tissues disseminated in the peritoneum 4 weeks after injection were used for histological examination with hematoxylin and eosin (H&amp;E) and paired-box gene 8 (PAX-8) staining. Results In the phase I trial, two piglets showed PM with modified PCI scores of 5 and 4 at 3 weeks after the first inoculation, which increased to 14 and 15 after 4 weeks, respectively. In the phase II trial, PM was detected in eight of ten piglets, which showed modified PCI scores of 6 to 12 at 4 weeks after the first inoculation. The overall incidence of PM from the total of 12 piglets after inoculation was 75%. Immunohistochemical H&amp;E and PAX-8 staining confirmed metastatic tumors. Conclusions This study provides strong evidence that piglets can be employed as a model for PM by inoculating ovarian cancer cell lines from humans. Using two cell lines, the PM rate is 75%.

Platinum-induced mitochondrial OXPHOS contributes to cancer stem cell enrichment in ovarian cancer

Abstract Background Platinum based agents—cisplatin and carboplatin in combination with taxanes are used for the treatment of ovarian cancer (OC) patients. However, the majority of OC patients develop recurrent, platinum resistant disease that is uniformly fatal. Platinum treatment enriches for chemoresistant aldehyde dehydrogenase (ALDH) + ovarian cancer stem cells (OCSCs), which contribute to tumor recurrence and disease relapse. Acquired platinum resistance also includes metabolic reprograming and switching to oxidative phosphorylation (OXPHOS). Chemosensitive cells rely on glycolysis while chemoresistant cells have the ability to switch between glycolysis and OXPHOS, depending on which pathway drives a selective advantage for growth and chemoresistance. High expression of genes involved in OXPHOS and high production of mitochondrial ROS are characteristics of OCSCs, suggesting that OCSCs favor OXPHOS over glycolysis. Based on connections between OCSCs, chemoresistance and OXPHOS, we hypothesize that platinum treatment induces changes in metabolism that contribute to platinum-induced enrichment of OCSCs. Methods The effect of cisplatin on mitochondrial activity was assessed by JC1 staining and expression of OXPHOS genes by RT-qPCR. Cisplatin-induced changes in Sirtuin 1 (SIRT1) levels and activity were assessed by western blot. Small molecule inhibitors of mitochondrial complex I and SIRT1 were used to determine if their enzymatic activity contributes to the platinum-induced enrichment of OCSCs. The percentage of ALDH + OCSCs in OC cells and tumor tissue from xenograft models across different treatment conditions was analyzed using ALDEFLUOR assay and flow cytometry. Results We demonstrate that platinum treatment increases mitochondrial activity. Combined treatment of platinum agents and OXPHOS inhibitors blocks the platinum-induced enrichment of ALDH + OCSCs in vitro and in vivo. Furthermore, platinum treatment increases SIRT1 levels and subsequent deacetylase activity, which likely contributes to the increase in platinum-induced mitochondrial activity. Conclusions These findings on metabolic pathways altered by platinum-based chemotherapy have uncovered key targets that can be exploited therapeutically to block the platinum-induced enrichment of OCSCs, ultimately improving the survival of OC patients.

Hsa_circ_0000497 and hsa_circ_0000918 contributed to peritoneal metastasis of ovarian cancer via ascites

Abstract Purpose As a common complication of epithelial ovarian cancer (EOC), malignant ascites contributes to the peritoneal metastasis of EOC. CircRNAs play essential roles in tumor metastasis. However, no circRNAs have been reported to be involved in EOC peritoneal metastasis via ascites. Methods Total of 22 samples from 9 EOC patients containing primary lesions (T), tumor cells from ascites (ASC), and metastatic lesions (M) were included for RNA sequencing to identify differentially expressed circRNAs and mRNAs among different tumors. Bioinformatic analyses, including single-sample Gene Set Enrichment Analysis and soft cluster analysis, were performed to find circRNAs potentially correlated with ascitic metastasis. Wound healing and transwell analysis were performed to evaluate tumor cells metastasis in vitro. Quantitative real-time PCR and western-blot were used for gene expression evaluation. Results According to transcriptomic analysis, ASC showed mesenchymal phenotype while T and M showed epithelial phenotype. 10 circRNAs were differentially expressed among ASC, T, and M. Among them, hsa_circ_0000497 and hsa_circ_0000918 were significantly up-regulated in ASC. Functional analysis showed that both hsa_circ_0000497 and hsa_circ_0000918 promoted metastasis of EOC via epithelial-mesenchymal transition (EMT) in vitro. The regulatory network construction identified 8 miRNAs and 19 mRNAs, and 7 miRNAs and 17 mRNAs as potential downstream target genes of hsa_circ_0000497 and hsa_circ_0000918, respectively, which may play pivotal roles in EOC ascitic metastasis. Conclusions circRNAs (hsa_circ_0000497 and hsa_circ_0000918) contribute to metastasis of EOC via ascites by regulating EMT. These circRNAs may serve as novel potential therapeutic targets or prognostic biomarkers for EOC peritoneal metastasis.

In vivo detection of dysregulated choline metabolism in paclitaxel-resistant ovarian cancers with proton magnetic resonance spectroscopy

Abstract Background Chemoresistance gradually develops during treatment of epithelial ovarian cancer (EOC). Metabolic alterations, especially in vivo easily detectable metabolites in paclitaxel (PTX)-resistant EOC remain unclear. Methods Xenograft models of the PTX-sensitive and PTX-resistant EOCs were built. Using a combination of in vivo proton-magnetic resonance spectroscopy (1H-MRS), metabolomics and proteomics, we investigated the in vivo metabolites and dysregulated metabolic pathways in the PTX-resistant EOC. Furthermore, we analyzed the RNA expression to validate the key enzymes in the dysregulated metabolic pathway. Results On in vivo 1H-MRS, the ratio of (glycerophosphocholine + phosphocholine) to (creatine + phosphocreatine) ((GPC + PC) to (Cr + PCr))(i.e. Cho/Cr) in the PTX-resistant tumors (1.64 [0.69, 4.18]) was significantly higher than that in the PTX-sensitive tumors (0.33 [0.10, 1.13]) (P = 0.04). Forty-five ex vivo metabolites were identified to be significantly different between the PTX-sensitive and PTX-resistant tumors, with the majority involved of lipids and lipid-like molecules. Spearman’s correlation coefficient analysis indicated in vivo and ex vivo metabolic characteristics were highly consistent, exhibiting the highest positive correlation between in vivo GPC + PC and ex vivo GPC (r = 0.885, P &lt; 0.001). These metabolic data suggested that abnormal choline concentrations were the results from the dysregulated glycerophospholipid metabolism, especially choline metabolism. The proteomics data indicated that the expressions of key enzymes glycerophosphocholine phosphodiesterase 1 (GPCPD1) and glycerophosphodiester phosphodiesterase 1 (GDE1) were significantly lower in the PTX-resistant tumors compared to the PTX-sensitive tumors (both P &lt; 0.01). Decreased expressions of GPCPD1 and GDE1 in choline metabolism led to an increased GPC levels in the PTX-resistant EOCs, which was observed as an elevated total choline (tCho) on in vivo 1H-MRS. Conclusions These findings suggested that dysregulated choline metabolism was associated with PTX-resistance in EOCs and the elevated tCho on in vivo 1H-MRS could be as an indicator for the PTX-resistance in EOCs.

RNA-binding proteins in ovarian cancer: a novel avenue of their roles in diagnosis and treatment

AbstractOvarian cancer (OC), an important cause of cancer-related death in women worldwide, is one of the most malignant cancers and is characterized by a poor prognosis. RNA-binding proteins (RBPs), a class of endogenous proteins that can bind to mRNAs and modify (or even determine) the amount of protein they can generate, have attracted great attention in the context of various diseases, especially cancers. Compelling studies have suggested that RBPs are aberrantly expressed in different cancer tissues and cell types, including OC tissues and cells. More specifically, RBPs can regulate proliferation, apoptosis, invasion, metastasis, tumorigenesis and chemosensitivity and serve as potential therapeutic targets in OC. Herein, we summarize what is currently known about the biogenesis, molecular functions and potential roles of human RBPs in OC and their prospects for application in the clinical treatment of OC.

Connecting METTL3 and intratumoural CD33+ MDSCs in predicting clinical outcome in cervical cancer

Abstract Background Methyltransferase-like 3 (METTL3) is a member of the m 6 A methyltransferase family and acts as an oncogene in cancers. Recent studies suggest that host innate immunity is regulated by the enzymes controlling m 6 A epitranscriptomic changes. Here, we aim to explore the associations between the levels of METTL3 and CD33 + myeloid-derived suppressor cells (MDSCs) in tumour tissues and the survival of patients with cervical cancer (CC). Methods Specimens of paraffin embedded tumour from 197 CC patients were collected. The expression levels of METTL3 and CD33 were measured by immunohistochemical (IHC) staining. The clinical associations of the IHC variants were analysed by Pearson’s or Spearman’s chi-square tests. Overall survival (OS) and disease-free survival (DFS) were estimated by the Kaplan–Meier method and log-rank test. Hazard ratios (HRs) and independent significance were obtained via Cox proportional hazards models for multivariate analyses. METTL3 in CD33 + cells or CC-derived cells was knocked down by METTL3-specific siRNA, and MDSC induction in vitro was performed in a co-culture system in the presence of METTL3-siRNA and METTL3-knockdown-CC-derived cells compared with that of the corresponding controls. Results We found that tumour tissues displayed increased levels of METTL3 and CD33 + MDSCs compared with tumour-adjacent tissues from the same CC patients. Importantly, METTL3 expression was positively related to the density of CD33 + cells in tumour tissues ( P  = 0.011). We further found that the direct CD33 + CD11b + HLA-DR − MDSC induction and tumour-derived MDSC induction in vitro were decreased in the absence of METTL3. The level of METTL3 in tumour microenvironments was significantly related to advanced tumour stage. The levels of METTL3 and CD33 + MDSCs in tumour tissues were notably associated with reduced DFS or OS. Cox model analysis revealed that the level of METTL3 in tumour cells was an independent factor for patient survival, specifically for DFS (HR = 3.157, P  = 0.022) and OS (HR = 3.271, P  = 0.012), while the CD33 + MDSC number was an independent predictor for DFS (HR: 3.958, P  = 0.031). Interestingly, in patients with advanced-disease stages (II–IV), METTL3 in tumour cells was an independent factor for DFS (HR = 6.725, P  = 0.010) and OS (HR = 5.140, P  = 0.021), while CD33 + MDSC density was an independent factor for OS (HR = 8.802, P  = 0.037). Conclusion Our findings suggest that CD33 + MDSC expansion is linked to high levels of METTL3 and that METTL3 and CD33 + MDSCs are independent prognostic factors in CC.

Prevalence of antibodies against a cyclic peptide mimicking the FG loop of the human papillomavirus type 16 capsid among Tunisian women

Abstract Background In the past decade, cervical cancer has gone from being the second to the fourth most common cancer in women worldwide, but remains the second most common in developing countries. This cancer is most commonly caused by high-risk types of human papillomavirus (HPV), mainly type 16 (HPV16), which are sexually transmitted. This study aimed to investigate the usefulness of a cyclic synthetic peptide designed from the major L1 capsid protein of HPV16 for detecting anti-HPV16 antibodies. Methods We designed and synthetized a peptide that corresponds to the full sequence of the surface-exposed FG loop. We tested the antigenicity of the linear and the cyclic peptides against HPV16 L1 monoclonal antibodies. We used ELISA to detect anti-peptide antibodies in sera and cervical secretions of 179 Tunisian women, and we applied polymerase chain reaction and direct sequencing methods to detect and genotype HPV DNA. Results Both the linear and the cyclic peptides were recognized by the same neutralizing monoclonal antibodies, but the cyclic peptide was more reactive with human sera. The prevalence of the anti-peptide antibodies in sera was higher in women with low-grade squamous intraepithelial lesions (LGSIL) than in women with high-grade squamous intraepithelial lesions (HGSIL) (44% and 15%, respectively). This contrasts with HPV16 DNA prevalence. Compared to women from the general population, systemic IgG prevalence was significantly higher among sex workers (25%; P  = 0.002) and women with LGSIL (44%; P  = 0.001). In addition, systemic IgA and cervical IgG prevalence was higher among sex workers only ( P  = 0.002 and P  = 0.001, respectively). We did not observe anti-peptide IgG antibodies in women with a current HPV16 infection. Conclusion Anti-peptide IgG in sera or in cervical secretions could be markers of an effective natural immunization against HPV16. This may open novel perspectives for monitoring vaccinated women and for the design of synthetic peptide-based vaccines.

Low molecular weight serum cell-free DNA concentration is associated with clinicopathologic indices of poor prognosis in women with uterine cancer

Abstract Background Serum cell-free DNA (cfDNA) holds promise as a non-invasive cancer biomarker. The objective of this study was to evaluate the association of cfDNA concentration with clinicopathologic variables of poor prognosis and overall survival among women with uterine cancer compared to benign cancer-free controls. Methods cfDNA was extracted from the serum of 91 women with multiple uterine cancer histologies and 22 post-menopausal controls without cancer. Low molecular weight (LMW) cfDNA was separated from contaminating genomic high molecular weight cfDNA using paramagnetic bead purification and its concentration was measured using fluorometric quantification. Clinicopathologic data was abstracted from the electronic medical record. The association between serum cfDNA concentration, clinicopathologic variables, and overall survival was assessed using linear regression modelling, Cox proportional hazards modelling, and the Kaplan–Meier method. Results Median total serum cfDNA concentration for the cohort was 69.2 ng/mL (IQR 37.4, 132.3) and median LMW cfDNA concentration was 23.8 ng/mL (IQR 14.9, 44.4). There were no significant differences in total serum cfDNA concentration with any clinicopathologic variables. However, LMW cfDNA concentration was significantly higher in serum of women with cancer (25.8 ng/mL IQR 16.0, 49.6) compared to benign controls (15.5 ng/mL IQR 9.3, 25.8 ng/mL) (p &lt; 0.01). It is also significantly higher among women with early stage cancer than benign controls (p &lt; 0.01). There were also significant associations between LMW cfDNA concentration and stage of cancer (p = 0.01) and histology (p = 0.02). Patients with leiomyosarcoma and carcinosarcoma had higher cfDNA concentrations than those with endometrioid cancer. Over a median follow-up of 51.9 months, 75th percentile for overall survival for women with cancer was 24.0 months. Higher LMW cfDNA concentrations is associated with lower survival among women with cancer (p &lt; 0.01). Conclusions Serum LMW cfDNA concentration is associated with overall survival in women with uterine cancer, and it is higher among women with uterine cancer compared to those of controls.

Identification of an immune classification for cervical cancer and integrative analysis of multiomics data

Abstract Background To understand the molecular mechanisms of the antitumour response, we analysed the immune landscape of cervical cancer to identify novel immune molecular classes. Methods We established a stable immune molecular classification using a nonnegative matrix factorization algorithm and validated the correlation in two validation sets of 249 samples. Results Approximately 78% of cervical cancers (CCs) (228/293) were identified to show significant enrichment in immune cells (e.g., CD8 T cells and macrophages), a type I IFN response, enhanced cytolytic activity and high PDCD1, and these CCs were referred to as the “immune class”. We further identified two subtypes of the immune class: active immune and exhausted subtypes. Although the active immune subtype was characterized by enrichment of IFN signatures and better survival, the exhausted subtype expressed activated stroma, a wound healing signature, enhanced M2 macrophages and absence of CD8 T cells and the TGF-β response signature. Integrative analysis of multiomics data identified EGFR, JUN, MYC, FN1 and SERPINE1 as key modulators of the tumour immune microenvironment and potential targets for combination therapies which was validated in two validation sets. Conclusions Our study introduces a novel immune classification that might predict ideal candidates to receive immunotherapy or specific combination therapies.

The intratumour microbiota and neutrophilic inflammation in squamous cell vulvar carcinoma microenvironment

Abstract Background A causal link between microbiota composition (dysbiosis) and oncogenesis has been demonstrated for several types of cancer. Neutrophils play a role in both immune protection against bacterial threats and carcinogenesis. This study aimed to characterise intratumoral bacteria in vulvar squamous cell carcinoma (VSCC) and their putative effect on neutrophil recruitment and cancer progression. Methods Clinical material was obtained from 89 patients with VSCC. Next-generation sequencing (NGS) of 16S rRNA and quantitative polymerase chain reaction (qPCR) were used to detect bacterial species in VSCC. To verify neutrophil activation, CD66b expression in tumour specimens was analysed by immunohistochemistry (IHC). Subsequently, IHC was applied to detect the main neutrophil serine proteases (NSPs), cathepsin G (CTSG), neutrophil elastase (ELANE), and proteinase 3 (PRTN3) in VSCC. Results Fusobacterium nucleatum and Pseudomonas aeruginosa were identified as tumour-promoting bacteria, and their presence was found to be associated with a shorter time to progression in VSCC patients. Furthermore, high abundance of CD66b, the neutrophil activation marker, in VSCC samples, was found to relate to poor survival of patients with VSCC. The selected NSPs were shown to be expressed in vulvar tumours, also within microabscess. The increased numbers of microabscesess were correlated with poor survival in VSCC patients. Conclusions Our results show that neutrophilic inflammation seem to be permissive for tumour-promoting bacteria growth in VSCC. The findings provide new therapeutic opportunities, such as based on shifting the balance of neutrophil populations to those with antitumorigenic activity and on targeting NSPs produced by activated neutrophils at the inflammation sites.

A phase 2 study of combined chemo-immunotherapy with cisplatin-pembrolizumab and radiation for unresectable vulvar squamous cell carcinoma

Abstract Background Unresectable or metastatic vulvar cancer has relatively poor outcomes despite chemotherapy-sensitized radiation therapy and combination cytotoxic therapy. Despite the virus-associated and immunogenic nature of this disease, novel immunotherapy options that exploit this advantage are currently lacking. Platinum agents such as cisplatin have been shown to prime dendritic cells for T-cell costimulation, promote downregulation of inhibitory checkpoint molecules, and sensitize tumor cells to cytotoxic T-cell killing. Radiation therapy has also been shown to promote immunogenetic cell death as monotherapy and in combination with cisplatin. In combination with pembrolizumab, cisplatin-sensitized radiation is hypothesized to increase overall response rates and recurrence-free survival in patients with vulvar cancer, via induction of an anti-tumor inflammatory response. Methods We propose a single-arm phase II clinical trial of pembrolizumab combined with cisplatin-sensitized radiation therapy for women with unresectable, locally advanced, or metastatic vulvar cancer. The first three patients with locally advanced or unresectable disease will receive cycle 1 of pembrolizumab followed by a break and resumption of pembrolizumab at cycle 4 and as part of a safety cohort. All other patients, including the fourth patient with locally advanced/unresectable disease, will receive weekly cisplatin and pembrolizumab every 3 weeks, concurrently with daily radiation therapy. Following the completion of Cis-RT, patients will continue pembrolizumab maintenance for a total of 12 cycles. Archived tissue will be used for HPV status, MSI status, PD-L1, and TIL stratification post hoc. Imaging will be performed at baseline and every 3 cycles (21-day cycles) as per standard-of-care. Laboratory analysis will occur on the first day of each cycle. Discussion The combination of cisplatin-sensitized radiation and immune checkpoint blockade has not been evaluated in the upfront setting for vulvar cancer. In this rare malignancy, there are limited interventional clinical trials. This trial is designed to be as accessible as possible by allowing patients to receive cisplatin and radiation locally according to accepted standard-of-care while receiving pembrolizumab and adverse event monitoring at a centralized site. A robust suite of translational correlative studies has also been built into the trial to evaluate tumor-directed immune activation. Trial registration NCT04430699

Immune-related 3-lncRNA signature with prognostic connotation in a multi-cancer setting

Abstract Background Advances in our understanding of the tumor microenvironment have radically changed the cancer field, highlighting the emerging need for biomarkers of an active, favorable tumor immune phenotype to aid treatment stratification and clinical prognostication. Numerous immune-related gene signatures have been defined; however, their prognostic value is often limited to one or few cancer types. Moreover, the area of non-coding RNA as biomarkers remains largely unexplored although their number and biological roles are rapidly expanding. Methods We developed a multi-step process to identify immune-related long non-coding RNA signatures with prognostic connotation in multiple TCGA solid cancer datasets. Results Using the breast cancer dataset as a discovery cohort we found 2988 differentially expressed lncRNAs between immune favorable and unfavorable tumors, as defined by the immunologic constant of rejection (ICR) gene signature. Mapping of the lncRNAs to a coding-non-coding network identified 127 proxy protein-coding genes that are enriched in immune-related diseases and functions. Next, we defined two distinct 20-lncRNA prognostic signatures that show a stronger effect on overall survival than the ICR signature in multiple solid cancers. Furthermore, we found a 3 lncRNA signature that demonstrated prognostic significance across 5 solid cancer types with a stronger association with clinical outcome than ICR. Moreover, this 3 lncRNA signature showed additional prognostic significance in uterine corpus endometrial carcinoma and cervical squamous cell carcinoma and endocervical adenocarcinoma as compared to ICR. Conclusion We identified an immune-related 3-lncRNA signature with prognostic connotation in multiple solid cancer types which performed equally well and in some cases better than the 20-gene ICR signature, indicating that it could be used as a minimal informative signature for clinical implementation.

Survival outcomes following treatment delays among patients with early-stage female cancers: a nationwide study

Abstract Background The coronavirus disease 2019 (COVID-19) severely hindered the timely receipt of health care for patients with cancer, especially female patients. Depression and anxiety were more pronounced in female patients than their male counterparts with cancer during treatment wait-time intervals. Herein, investigating the impact of treatment delays on the survival outcomes of female patients with early-stage cancers can enhance the rational and precise clinical decisions of physicians. Methods We analyzed five types of cancers in women from the Surveillance, Epidemiology, and End Results (SEER) program between Jan 2010 and Dec 2015. Univariate and multivariate Cox regression analyses were used to determine the impacts of treatment delays on the overall survival (OS) and cancer-specific survival (CSS) of the patients. Results A total of 241,661 females with early-stage cancer were analyzed (12,617 cases of non-small cell lung cancer (NSCLC), 166,051 cases of infiltrating breast cancer, 31,096 cases of differentiated thyroid cancer, 23,550 cases of colorectal cancer, and 8347 cases of cervical cancer). Worse OS rates were observed in patients with treatment delays ≥ 3 months in stage I NSCLC (adjustedHazard ratio (HR) = 1.11, 95% Confidence Interval (CI): 1.01–1.23, p = 0.044) and stage I infiltrating breast cancer (adjustedHR = 1.23, 95% CI 1.11–1.37, p &lt; 0.001). When the treatment delay intervals were analyzed as continuous variables, similar results were observed in patients with stage I NSCLC (adjustedHR = 1.04, 95% CI 1.01–1.06, p = 0.010) and in those with stage I breast cancer (adjustedHR = 1.03, 95% CI 1.00–1.06, p = 0.029). However, treatment delays did not reduce the OS of patients with differentiated thyroid cancer, cervical cancer, or colorectal cancer in the early-stage. Only intermediate treatment delays impaired the CSS of patients with cervical cancer in stage I (adjustedHR = 1.31, 95% CI 1.02–1.68, p = 0.032). Conclusion After adjusting for confounders, the prolonged time from diagnosis to the initiation of treatment (&lt; 6 months) showed limited negative effects on the survival of most of the patients with early-stage female cancers. Whether our findings serve as evidence supporting the treatment deferral decisions of clinicians for patients with different cancers in resource-limited situations needs further validation.

Can HPV Selfy be considered as a clinically validated HPV test for use in cervical cancer screening?

Gene signatures, immune infiltration, and drug sensitivity based on a comprehensive analysis of m6a RNA methylation regulators in cervical cancer

Abstract Background Cervical cancer is the fourth most common cancer in women. N6-dimethyladenosine (m6A) mRNA methylation is closely associated with cervical cancer. Methods Using TCGA database, we studied the expression and mutation of m6A-related genes in cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC) and obtained genetic characteristics based on an m6A risk model and prognostic value of m6A. We studied the effects of the m6A risk score on immune features and genomic changes of patients with CESC, evaluated the sensitivity of patients with CESC to different small-molecule drugs based on the m6A risk score, and established a clinical prediction model. Results Ten m6A-related genes were differentially expressed between CESC and normal tissues. High-risk patients had a low overall survival (OS) and significantly low immune scores but showed no significantly altered stromal scores. The tumor mutation burden (TMB) and tumor neoantigen levels significantly differed between the high- and low-risk groups. In the high-risk group, copy number variation (CNV) changes mainly led to gene amplification, while in the low-risk group, CNV changes primarily manifested as gene copy number deletions. ZC3H13 expression was low in CESC tissues. ZC3H13 knockdown promoted CESC cell proliferation, migration, and invasion, reducing the RNA methylation levels. Rapamycin suppressed the CESC cell proliferation, migration, and invasion abilities, increasing the m6A levels. Conclusion m6A mRNA methylation is closely related to the occurrence, development, immune invasion, drug sensitivity, and prognosis of cervical cancer. The prognostic m6A feature model of m6A signature genes can accurately predict the OS of patients with CESC. Drugs targeting factors regulating m6A mRNA methylation might offer a good prospect for treating cervical cancer.

MiR-1254 suppresses the proliferation and invasion of cervical cancer cells by modulating CD36

Abstract Background This study aimed to elucidate the roles of miR-1254 in cervical cancer progression and to explore the underlying mechanisms. Methods The expression levels of miR-1254 in normal-cancer cervical tissues and cells were measured using quantitive real-time polymerase chain reaction (qRT-PCR). The invasive and proliferative abilities of cervical cancer cell lines transfected with negative control (NC) mimic or miR-1254 mimic were measured using transwell, CCK-8, and colony formation assays. The binding sites between CD36 and miR-1254 were determined using luciferase reporter assays. The correlation of CD36 and miR-1254 with cervical cancer development was re-confirmed by co-transfection of miR-1254 mimic and CD36 overexpression using CCK-8, colony formation, transwell and western blot assays. Results MiR-1254 was expressed at significantly lower levels in the cervical cancer cell lines and tissues than in the controls. The functional assays revealed that upregulation of miR-1254 inhibited the invasion and proliferation of cervical cancer cells. The luciferase reporter assays demonstrated that CD36 messenger RNA and miR-1254 bound to one another. CD36 overexpression reversed the inhibitory effects of upregulated miR-1254 in the cervical cancer cells, suggesting that miR-1254 regulates cervical cancer progression by modulating CD36. Conclusion miR-1254 attenuated the invasion and proliferation of cervical cancer cells by modulating the expression levels of CD36.

RGS1 and related genes as potential targets for immunotherapy in cervical cancer: computational biology and experimental validation

Abstract Background Effective treatment is needed for advanced, inoperable, or chemotherapy-resistant cervical cancer patients. Immunotherapy has become a new treatment modality for cervical cancer patients, and there is an urgent need to identify additional targets for cervical cancer immunotherapy. Methods In this study the core gene, RGS1, which affects immune status and the FIGO stage of cervical cancer patients was identified by WGCNA analysis and differential analysis using TCGA database. 10 related genes interacting with RGS1 were identified using PPI network, and the functional and immune correlations were analyzed. Based on the expression of RGS1 and related genes, the consensus clustering method was used to divide CESC patients into two groups (group 1, high expression of RGS1; group 2, low expression of RGS1). Then, the functional enrichment analysis was used to search for the functional differences in differentially expressed genes (DEGs) between group 1 and group 2. Immune infiltration analysis was performed using ESTIMATE, CIBERSORT, and ssGSEA, and the differences in expression of immune checkpoint inhibitors (ICIs) targets were assessed between the two groups. We investigated the effect of RGS1 on the clinical relevance of CESC patients, and experimentally verified the differences in RGS1 expression between cervical cancer patient tissues and normal cervical tissues, the role of RGS1 in cell function, and the effect on tumor growth in tumor-bearing mice. Results We found that RGS1 was associated with CD4, GNAI3, RGS2, GNAO1, GNAI2, RGS20, GNAZ, GNAI1, HLA-DRA and HLA-DRB1, especially CD4 and RGS2. Functional enrichment of DEGs was associated with T cell activation. Compared with group 2, group 1 had stronger immune infiltration and higher ICI target expression. RGS1 had higher expression in cervical cancer tissues than normal tissues, especially in HPV-E6 positive cancer tissues. In cervical cancer cell lines, knockdown of RGS1 can inhibited cell proliferation, migration, invasion, and tumor growth in nude mice and promoted apoptosis. Conclusions RGS1, as an oncogenic gene of cervical cancer, affects the immune microenvironment of patients with cervical cancer and may be a target of immunotherapy.

Metabolic interplay between endometrial cancer and tumor-associated macrophages: lactate-induced M2 polarization enhances tumor progression

Tumor-associated macrophages (TAMs), abundant within the tumor microenvironment, are key mediators of immunosuppression and represent promising therapeutic targets. Metabolic crosstalk between tumor cells and TAMs is a critical regulator of immune phenotype switching. However, the interactions between endometrial cancer (EC) cells and TAMs remain incompletely understood. Here, we demonstrate that EC cells exhibit increased aerobic glycolysis, as confirmed by bulk transcriptomics, extracellular lactate measurements, RT-PCR, and immunohistochemistry. M2-polarized TAMs were significantly more prevalent in EC tissues compared to normal endometrium, and this prevalence correlated with deep myometrial invasion and advanced stages. In vitro assays revealed that EC cell-derived lactate promotes M2 polarization of macrophages, enhancing epithelial-mesenchymal transition and angiogenesis, thereby increasing EC cell invasiveness and metastasis. Cytokine profiling and functional assays further demonstrated that lactate-stimulated M2 TAMs secrete elevated IL-6, which promotes tumor progression. Importantly, blocking IL-6 signaling significantly reduced the M2 TAM-mediated effects on EC cells in vitro and inhibited tumor growth and metastasis in vivo. Our study underscores the pivotal role of aerobic glycolysis-derived lactate in inducing TAM M2 polarization and highlights the IL-6 axis as a therapeutic target, offering new strategies for treating EC by disrupting metabolic-immune crosstalk.

Placenta-derived trophoblast extracellular vesicles promote CD169+ macrophage migration and suppress ovarian tumour growth

A lower survival rate due to a later stage of diagnosis requires a more effective therapy for ovarian cancer. Extracellular vesicles (EVs) have shown therapeutic potential in various diseases, partly due to their ability to modulate immune responses. We previously showed that placental-derived trophoblast EVs suppressed ovarian tumour growth, accompanied by increased infiltration of CD169 SKOV-3 ovarian cancer cell xenografts were intraperitoneally injected with placental-derived trophoblast EVs. Tumour growth was monitored, and EV biodistribution was tracked at four time points. Additionally, the dynamics of CD169 A reduction of tumour growth after EV treatment was observed. At early time points (within 24 hours), trophoblast EVs preferentially accumulated in immune organs, including the inguinal lymph nodes, but not in tumours. The significantly higher intensity of CD169 Placental-derived trophoblast EVs may act as an immune modulator, specifically in the presence of tumours, through promoting CD169

piRNA-14633 promotes cervical cancer cell malignancy in a METTL14-dependent m6A RNA methylation manner

Abstract Background Cervical cancer (CC) is one of the most common gynecological tumors that threatens women's health and lives. Aberrant expression of PIWI-interacting RNA (piRNA) is closely related with a range of cancers and can serve as a tumor promoter or suppressor in proliferation, migration and invasion. In this study, the aim was not only to discover differential expression of piRNA in CC tissue (CC cells) and normal cervical tissue (normal cervical epithelium cells), but also to investigate the biological function and action mechanism of piRNA in CC. Methods The DESeq2 approach was used to estimate fold change in piRNA between CC tissue and normal cervical tissue. The relative expressions of piRNAs (piRNA-20657, piRNA-20497, piRNA-14633 and piRNA-13350) and RNA m6A methyltransferases/demethylases were detected using RT-qPCR. After intervention with piRNA-14633 and METTL14 expression, the viability of CaSki cells and SiHa cells was detected by CCK8. CC cell proliferation was detected by colony formation assay. Apoptosis rate and cell cycle were detected by flow cytometry. Transwell assay was performed to detect cell migration and invasion. EpiQuik m6A RNA Methylation Quantification Kit was used to evaluate m6A RNA methylation levels. Expression of methyltransferase-like protein 14 (METTL14), PIWIL-proteins and CYP1B1 were detected by RT-qPCR and western blot. The effect of piRNA-14633 on METTL14 was evaluated by a dual-luciferase reporter assay. The in vivo effects of piRNA-14633 on CC was assessed by nude mice experiments. Results piRNA-14633 showed high expression in CC tissues and cells, piRNA-14633 mimic (piRNA-14633 overexpression) promoted viability, proliferation, migration and invasion of CaSki cells and SiHa cells. Besides, piRNA-14633 mimic increased m6A RNA methylation levels and METTL14 mRNA stability. Results of dual luciferase reporter assays indicated that METTL14 was a directed target gene of piRNA-14633. Knockdown of METTL14 with siRNA attenuated proliferation, migration and invasion of CC cells. piRNA-14633 increased CYP1B1 expression, while silencing of METTL14 impaired its expression. The effect of piRNA overexpression on METTL14 expression has concentration-dependent characteristics. Results from in vivo experiment indicated that piRNA-14633 promoted cervical tumor growth. Conclusion piRNA-14633 promotes proliferation, migration and invasion of CC cells by METTL14/CYP1B1 signaling axis, highlighting the important role of piRNA-14633 in CC.

Identification of COPA as a potential prognostic biomarker and pharmacological intervention target of cervical cancer by quantitative proteomics and experimental verification

AbstractBackgroundCervical cancer is the most fatal gynecological carcinoma in the world. It is urgent to explore novel prognostic biomarkers and intervention targets for cervical cancer.MethodsThrough integrated quantitative proteomic strategy,we investigated the protein expression profiles of cervical cancer; 28 fresh frozen tissue samples (11 adenocarcinoma (AC), 12 squamous cell carcinoma (SCC) and 5 normal cervixes (HC)) were included in discover cohort; 45 fresh frozen tissue samples (19 AC, 18 SCC and 8 HC) were included in verification cohort; 140 paraffin-embedded tissues samples of cervical cancer (85 AC and 55 SCC) were used for immunohistochemical evaluation (IHC) of coatomer protein subunit alpha (COPA) as a prognostic biomarker for cervical cancer; how deficiency of COPA affects cell viability and tumorigenic ability of cervical cancer cells (SiHa cells and HeLa cells) were evaluated by cell counting kit-8 and clone formation in vitro.ResultsWe identified COPA is a potential prognostic biomarker for cervical cancer in quantitative proteomics analysis. By retrospective IHC analysis, we additionally verified the proteomics results and demonstrated moderate or strong IHC staining for COPA is an unfavourable independent prognostic factor for cervical cancer. We also identified COPA is a potential pharmacological intervention target of cervical cancer by a series of in vitro experiments.ConclusionThis study is the first to demonstrate that COPA may contribute to progression of cervical cancer. It can serve as a potential prognostic biomarker and promising intervention target for cervical cancer.

LAPTM5 drives omental metastasis in high-grade serous ovarian cancer via TGF-β/Smad-mediated epithelial plasticity

Tumor marker–guided precision BNCT for CA19-9–positive cancers: a new paradigm in molecularly targeted chemoradiation therapy

Abstract Background Boron neutron capture therapy (BNCT) is a molecularly targeted chemoradiation modality that relies on boron delivery agents such as p-borophenylalanine (BPA), which require LAT1 (L-type amino acid transporter 1) for tumor uptake. However, the limited efficacy of BPA in LAT1-low tumors restricts its therapeutic scope. To address this limitation, we developed a tumor marker–guided BNCT strategy targeting cancers overexpressing the clinically validated glycan biomarker CA19-9. Methods We conducted transcriptomic analyses using The Cancer Genome Atlas (TCGA) datasets to identify LAT1-low cancers with high CA19-9 expression. These analyses revealed elevated expression of fucosyltransferase 3 (FUT3), which underlies CA19-9 biosynthesis, in pancreatic, biliary, and ovarian malignancies. Based on this, we synthesized a novel boron compound, fucose-BSH, designed to selectively accumulate in CA19-9–positive tumors. We evaluated its physicochemical properties, pharmacokinetics, biodistribution, and antitumor efficacy in cell lines and xenograft models, comparing its performance to that of BPA. Results Fucose-BSH demonstrated significantly greater boron uptake in CA19-9–positive cell lines (AsPC-1, Panc 04.03, HuCCT-1, HSKTC, OVISE) compared to CA19-9–negative PANC-1. In HuCCT-1 xenografts, boron accumulation reached 36.2 ppm with a tumor/normal tissue ratio of 2.1, outperforming BPA. Upon neutron irradiation, fucose-BSH–mediated BNCT achieved &gt; 80% tumor growth inhibition. Notably, fucose-BSH retained therapeutic efficacy in LAT1-deficient models where BPA was ineffective, confirming LAT1-independent targeting. Conclusions This study establishes a novel precision BNCT approach by leveraging CA19-9 as a tumor-selective glycan marker for boron delivery. Fucose-BSH offers a promising platform for expanding BNCT to previously inaccessible LAT1-low malignancies, including pancreatic, biliary, and ovarian cancers. These findings provide a clinically actionable strategy for tumor marker–driven chemoradiation and lay the foundation for translational application in BNCT. This strategy has the potential to support companion diagnostic development and precision stratification in ongoing and future BNCT clinical trials. Translational Relevance Malignancies with elevated CA19-9 expression, such as pancreatic, biliary, and ovarian cancers, are associated with poor prognosis and limited response to current therapies. This study presents a tumor marker–guided strategy for boron neutron capture therapy (BNCT) by leveraging CA19-9 glycan biology to enable selective tumor targeting via fucose-BSH, a novel boron compound. Through transcriptomic data mining and preclinical validation, fucose-BSH demonstrated LAT1-independent boron delivery, potent BNCT-mediated cytotoxicity, and tumor-specific accumulation in CA19-9–positive models. These findings support a precision chemoradiation approach that addresses a critical gap in BNCT applicability, offering a clinically actionable pathway for patient stratification and therapeutic development in CA19-9–expressing cancers. Graphical Abstract