American Journal of Reproductive Immunology

Comparative Analysis of Vaginal Inflammatory Cytokines Among HPV‐Positive and HPV‐Negative Women in Manila, Philippines

ABSTRACTProblemHuman papillomavirus (HPV) is considered the necessary cause of cervical cancer. Studies showed that disruption of the inflammatory milieu in the cervicovaginal mucosa can promote the persistence of HPV, which can result in cervical carcinogenesis. This study determined sociodemographic factors and vaginal inflammatory cytokines associated with HPV infection in Manila, Philippines.MethodCervicovaginal swab samples from 110 sexually active women, ages 21 and above, from Manila, diagnosed using real‐time PCR, were selected for comparative expression analysis using sandwich‐type ELISA of the cytokines IL‐6, IL‐8, IL‐1b, IL‐10, and TNF‐α. Independent t‐test, Wilcoxon Rank Sum, and Chi‐square test were employed. Multivariable logistic regression was conducted to determine factors associated with HPV infection.ResultsSociodemographic and behavioral factors analyzed in this study did not show a significant association with HPV infection. IL‐8 was significantly lower (p < 0.0001) while IL‐10 was significantly higher (p < 0.001) in the HPV‐positive group compared to the HPV‐negative samples. Multivariable logistic regression showed that higher IL‐1b (OR: 1.007; 95% CI: 1.003–1.012) and lower IL‐8 levels (OR: 0.979; 95% CI: 0.970–0.989) were associated with increased odds of HPV infection. IL‐8 promotes neutrophil recruitment and viral clearance, IL‐1β drives inflammation but may foster persistence via IL‐17 induction, and IL‐10 suppresses antiviral immunity, enabling HPV evasion.ConclusionThis study showed that lower IL‐8 and higher IL‐1β levels were independent predictors of HPV infection. These changes in the microenvironment may control immune cell infiltration and help establish HPV infection.

Prognostic Value of Systemic Immune‐Inflammation Index in Patients With Gynecological Tumors: A Systematic Review and Meta‐Analysis

ABSTRACT This meta‐analysis (MA) systematically evaluated the application value of systemic immune‐inflammation index (SII) in the prognosis of patients with gynecological tumors. Electronic databases including PubMed, Embase, Cochrane Library, Web of Science, Elsevier, and Wiley were searched for randomized controlled trials (RCTs) of SII according to inclusion and exclusion criteria, with the search period extending from the inception of the databases to November 2024. Clinical outcomes included overall survival (OS), disease‐free survival (DFS), progression‐free survival (PFS), and distant metastasis‐free survival (DMFS). The quality of the included articles was assessed adopting the Cochrane systematic review method, and MA was conducted adopting Stata 17.0 software. A total of eight studies (10 trials) were included, with high SII having an observable negative impact on OS [Tau 2  = 0.20, 95% confidence interval (CI): 1.30–2.58; Z  = 3.47, p  = 0.0005]. High SII had an observable negative impact on DFS and PFS (Tau 2  = 0.08, 95% CI: 1.25–1.97; Z  = 3.89, p  = 0.0001). High SII had an observable negative impact on DMFS (95% CI: 1.20–1.86; Z  = 3.54, p  = 0.0004). The MA results indicate that a high SII index predicts poor survival outcomes in patients with gynecological tumors and is an effective indicator for prognosis in clinical practice.

TCR CDR3‐antigen chemical complementarity associated with poor ovarian cancer outcomes: A vestigial immune response to early cancer antigens?

AbstractOvarian cancer continues to present significant challenges for early detection and treatment, indicating a need for novel approaches to improve disease outcomes. In this report, we applied a previously described algorithm for detecting chemical complementarity between candidate cancer antigens and complementarity determining region‐3 (CDR3) amino acid sequences from tumor resident T‐cell receptors. Current literature indicates an association between high CDR3‐cancer antigen complementarity and improved survival outcomes. For example, high CDR3‐BRAF electrostatic complementarity is associated with a better melanoma outcome. However, such CDR3‐cancer antigen chemical complementarity in ovarian cancer was largely associated with worse outcomes. Specifically, high CDR3‐MAGEB4 and CDR3‐TDRD1 electrostatic complementarity was associated with lower ovarian cancer disease free survival (DFS). Additionally, high CDR3‐MAGEB4 and CDR3‐TDRD1 electrostatic complementarity was associated with decreased MAGEB4/TDRD1 gene expression and gene copy numbers, consistent with a selection against ovarian cancer cells expressing these antigens. However, when TDRD1 was split into fragments, high CDR3‐TDRD1 hydrophobicity complementarity, for a specific TDRD1 fragment, was associated with increased DFS and higher immune marker expression levels. This dichotomy highlights the myriad of opportunities to establish risk stratifications and to identify potential, actionable cancer antigens using immunogenomic parameters.

Circ_0031027 adjusts the advancement of cervical cancer by miR‐587/SOCS5 axis

AbstractBackgroundCircular RNAs (circRNAs) might participate in the growth of cervical cancer (CC). In this research, we reconnoitered the characters of circ_0031027 in CC.MethodsCirc_0031027, microRNA‐587 (miR‐587), and suppressor of cytokine signaling 5 (SOCS5) abundances were distinguished by qRT‐PCR and western blot. The cell functions were examined by colony formation assay, EdU assay, and transwell assay. The combined relationship of miR‐587 and circ_0031027 or SOCS5 was identified by dual‐luciferase reporter assay. Ultimately, the mice test was utilized to assess the part of circ_0031027.ResultsCirc_0031027 and SOCS5 were downregulated, and the miR‐587 was upregulated in CC. For functional analysis, circ_0031027 overexpression repressed cell proliferation, cell migration, and invasion in CC cells. Circ_0031027 as a miR‐587 sponge to adjust SOCS5. MiR‐587 facilitated the advancement of CC cells by overturning SOCS5. In addition, circ_0031027 overexpression subdued tumor growth.ConclusionCirc_0031027 inhibited the enlargement of CC by miR‐587/SOCS5.

Circ_0060551 promotes the migration and invasion of cervical cancer by upregulating TPD52

AbstractProblemCervical cancer has been recognized as the second most common cancer in women worldwide. Previous studies have reported that some circular RNAs (circRNAs) can influence the progression of cervical cancer. However, more researches are still required to unveil the underlying mechanism of how circRNAs regulate the progression of such cancer. We aimed at unveiling the mechanism of how circ_0060551 effected the progression of cervical cancer.Method of studyRT‐qPCR and western blot assays were used to detect the expression and protein levels. Mechanism experiments were conducted to investigate the relationship among circ_0060551, TPD52, miR‐520a‐5p and ELAVL1. Rescue assays were mainly carried out to verify how circ_0060551 influenced the migration and invasion of cervical cancer cells.ResultsAccording to the results, circ_0060551 was upregulated in cervical cancer cells and could promote the migration and invasion of cells via TPD52. In addition, circ_0060551 could up‐regulate TPD52 expression through a ceRNA model to target miR‐520a‐5p. Moreover, circ_0060551 could stabilize the mRNA expression of TPD52 via recruiting ELAVL1.ConclusionOur study proved that circ_0060551 could promote the migration and invasion of cervical cancer cells.

SMURF2 Inhibits Autophagy and Growth in Ovarian Cancer by Regulating the RACK1/AKT/mTOR Pathway

ABSTRACTBackgroundOvarian cancer (OC) is a common malignancy characterized by disseminated peritoneal metastases. Smad ubiquitin regulatory factor 2 (SMURF2) is involved in OC progression by stabilizing receptor for activated C kinase 1 (RACK1). However, the functions and mechanisms of action of SMURF2 in OC remain unclear. This biological function of SMURF2 in OC and its potential mechanisms of action were investigated in this study.MethodsThe expression of SMURF2 in ovarian tumor tissues, patient serum, and OC cell lines was determined using reverse transcription‐quantitative polymerase chain reaction (RT‐qPCR) and/or western blotting. 3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyl‐2H‐tetrazolium bromide (MTT) assays, flow cytometry, and terminal deoxynucleotidyl transferase dUTP nick‐end labeling (TUNEL) were used for detecting cell proliferation and apoptosis. Autophagosomes in SKOV3 cells were observed using transmission electron microscopy. Immunohistochemistry and RT‐qPCR were performed to evaluate SMURF2 expression. The levels of proteins related to autophagy and RACK1 were measured using western blotting and RT‐qPCR, respectively. Western blotting was performed to assess the expression of AKT/mTOR pathway‐related proteins.ResultsSMURF2 was underexpressed in OC tissues and cell lines compared with that in adjacent normal tissues or normal ovarian epithelial cells. RT‐qPCR results suggested that SMURF2 was downregulated in the serum of patients with OC. SMURF2 overexpression inhibited SKOV3 cell growth and autophagy, and induced apoptosis both in vitro and in vivo. Moreover, SMURF2 overexpression suppressed RACK1 expression in SKOV3 cells. The AKT/mTOR pathway was activated by SMURF2 overexpression in SKOV3, and OC cells and tissues.ConclusionsSMURF2 plays a key role in OC by inhibiting cell autophagy and growth via activation of the RACK1/AKT/mTOR pathway, which might potentially be a new biomarker for OC diagnosis and therapy.

Identifying novel ovarian tumor biomarkers through mining of the transcriptome of circulating immune cells: A proof‐of‐concept study

AbstractObjectiveTreatment of high‐grade serous ovarian cancer (HGSOC) will benefit from early detection of cancer. Here, we provide proof‐of‐concept data supporting the hypothesis that circulating immune cells, because of their early recognition of tumors and the tumor microenvironment, can be considered for biomarker discovery.MethodsLongitudinal blood samples from C57BL/6 mice bearing syngeneic ovarian tumors and peripheral blood mononuclear cells (PBMC) from healthy postmenopausal women and newly diagnosed for HGSOC patients were subjected to RNASeq. The results from human immune cells were validated using Affymetrix microarrays. Differentially expressed transcripts in immune cells from tumor‐bearing mice and HGSOC patients were compared to matching controls.ResultsA total of 1282 transcripts (798 and 484, up‐ and downregulated, respectively) were differentially expressed in the tumor‐bearing mice as compared with controls. Top 100 genes showing longitudinal changes in gene expression 2, 4, 7, and 18 days after tumor implantation were identified. Analysis of the PBMC from healthy post‐menopausal women and HGSOC patients identified 4382 differentially expressed genes and 519 of these were validated through Affymetrix microarray analysis. A total of 384 genes, including IL‐1R2, CH3L1, Infitm1, FP42, CXC42, Hdc, Spib, and Sema6b, were differentially expressed in the human and mouse datasets.ConclusionThe PBMC transcriptome shows longitudinal changes in response to the progressing tumor. Several potential biomarker transcripts were identified in HGSOC patients and mouse models. Monitoring their expression in individual PBMC subsets can serve as additional discriminator for the diagnosis of HGSOC.

Triaging abnormal cervical cancer screening tests using p16INK4a detection by ELISA on fresh cervical samples

AbstractProblemCervical cancer screening strategies in the United States include cotesting (human papillomavirus (HPV) with cytology), primary HPV with genotyping and reflex cytology, and cytology alone. An ongoing challenge is the appropriate triage of patients to colposcopy to those at highest risk. We investigated whether incorporation of p16INK4a immunodetection by enzyme‐linked immunosorbent assay (ELISA) on fresh cervical samples obtained at the time of screening could improve appropriate referral to colposcopy.Method of StudyA derivation group comprised of cervical swabs collected from subjects with high‐grade dysplasia or cancer (positive control) and from subjects with negative screening history (negative control). Samples collected from colposcopy were used to evaluate the existing screening strategies individually and with incorporation of p16INK4a ELISA. Histology was used as the gold standard.ResultsAmong 163 subjects recruited, 138 were included. In the derivation group, mean p16INK4a level was 2.86 ng/mL (n = 31) and 0.58 ng/mL (n = 20) among positive and negative controls respectively (p = 0.002) with an area under the receiver operator characteristic curve of 0.79 (p < 0.001). Among colposcopy subjects, sensitivity/specificity for cotesting, primary HPV, and cytology were 94%/42%, 88%/45%, and 88%/49%, respectively. Incorporation of p16INK4a resulted in similar sensitivity and improved specificity (cotesting+p16 88%/58%, primary HPV+p16 88%/57%, cytology+p16 81%/62%; p = 0.23/p = 0.008) with decrease in colposcopy referrals by 15% to 22% (p = 0.01).ConclusionsThese results demonstrate the feasibility of quantifying p16INK4a by ELISA in fresh cervical samples, and its potential as an adjunct to existing screening strategies in the identification of high grade‐dysplasia while reducing the number of colposcopic referrals.

Expression of MHC Class I Molecules (HLA‐A, ‐B, ‐C, ‐E, ‐F, ‐G, and ‐J) Decreases From Early to Late Stage in Ovarian Cancer

ABSTRACTProblemInterferon‐ε (IFNε), which is highly abundant in the epithelium of the female reproductive tract (FRT), is a recently identified tumor suppressor for ovarian cancer. IFNε induces the expression of certain HLA class I family members in HGSOC (high‐grade serous ovarian cancer), and its expression is lost during ovarian tumorigenesis. However, tumor stage–dependent expression of HLA class I family members in ovarian cancer has not been previously studied.Method of StudyData analysis and visualization were performed using various gene expression and transcriptomics datasets in the R statistical programming environment.ResultsWe found that the expression of HLA‐A, ‐B, ‐C, ‐E, ‐F, ‐G, and ‐J is lower in late stage ovarian tumors compared to early‐stage tumors. The total expression of HLA class I family members decreases with age in ovarian cancer. Furthermore, we showed that the expression of some IFN‐regulated genes, which were shown to be upregulated by IFNε, decreases from early to late stage in ovarian cancer, in parallel to the loss of IFNε expression in ovarian tumorigenesis and possibly in tumor progression. We also found that breast tumors (another hormonally driven cancer) with positive progesterone receptor status have lower IFNε mRNA expression compared to those with negative PR status. Besides, we reported that breast tumors with positive estrogen receptor (ER) status have lower expression of IFNε compared to those with negative ER status.ConclusionsCombined, this study points that the decrease in the expression of IFNε, HLAs, or some other IFNε‐regulated genes during ovarian cancer progression might contribute to worse prognosis in advanced disease.

Better Outcomes for Ovarian Cancer Associated With the Detection of Anti‐EBV TCR CDR3s: Potential Relevance to Diffuse Large B‐Cell Lymphoma

ABSTRACTObjectivesGiven the ongoing challenges regarding the specific roles of viral infections in cancer etiology, or as cancer co‐morbidities, this study assessed potential associations between anti‐viral, T‐cell receptor (TCR) complementarity domain region‐3 (CDR3s), and clinical outcomes for ovarian cancer.MethodsTCR CDR3s were isolated from ovarian cancer specimens for a determination of which patients had anti‐viral CDR3s and whether those patients had better or worse outcomes.ResultsAnalyses revealed that patients with exact matches of anti‐Epstein–Barr virus (EBV) CDR3 amino acid sequences exhibited better outcomes for both overall and disease‐specific survival. However, better outcomes were not observed when assessing anti‐viral CDR3s representing cytomegalovirus, influenza A, or Sars‐CoV‐2. Due to previous occurrences of the occasional misdiagnoses of lymphoma as ovarian cancer, the frequency of anti‐EBV CDR3s in lymphoma patients was determined. These frequencies were relatively high, particularly for diffuse large B‐cell lymphoma.ConclusionsThese findings (i) underscore the potential value of anti‐EBV immune responses in terms of patient outcomes; (ii) raise questions about the potential value of anti‐EBV immunotherapies; and (iii) support further inquiry into the relationship between EBV infection and previously reported cases of ovary‐resident lymphoma.

Long non‐coding RNA TPT1‐AS1 inhibits ferroptosis in ovarian cancer by regulating GPX4 via CREB1 regulation

AbstractBackgroundLong non‐coding RNAs (lncRNAs) play crucial roles in cellular processes, with dysregulation implicated in various diseases, including cancers. The lncRNA TPT1‐AS1 (TPT1 Antisense RNA 1) promotes tumor progression in several cancers, including ovarian cancer (OC), but its influence on ferroptosis and interaction with other proteins remains underexplored.MethodsIn this study, we employed a multi‐faceted approach to investigate the functional significance of TPT1‐AS1 in OC. We assessed TPT1‐AS1 expression in OC specimens and cell lines using RT‐qPCR, in situ hybridization (ISH), and fluorescence in situ hybridization (FISH) assays. Functional assays included evaluating the impact of TPT1‐AS1 knockdown on OC cell proliferation, migration, invasiveness, and cell cycle progression. Further, we explored and validated the interaction of TPT1‐AS1 with other proteins using bioinformatics. Finally, we investigated TPT1‐AS1 involvement in erastin‐induced ferroptosis using Iron Assay, Malondialdehyde (MDA) assay, and reactive oxygen species (ROS) detection.ResultsOur findings revealed that TPT1‐AS1 overexpression in OC correlated with an unfavorable prognosis. TPT1‐AS1 knockdown suppressed cell proliferation, migration, and invasiveness. Additionally, TPT1‐AS1 inhibited erastin‐induced ferroptosis, and in vivo experiments confirmed its oncogenic impact on tumor development. Mechanistically, TPT1‐AS1 was found to regulate Glutathione Peroxidase 4 (GPX4) transcription via CREB1 (cAMP response element‐binding protein 1) and interact with RNA‐binding protein (RBP) KHDRBS3 (KH RNA Binding Domain Containing, Signal Transduction Associated 3) to regulate CREB1.ConclusionTPT1‐AS1 promotes OC progression by inhibiting ferroptosis and upregulating CREB1, forming a regulatory axis with KHDRBS3. These findings highlight the regulatory network involving lncRNAs, RBPs, and transcription factors in cancer progression.

“Identification of Malignancy Risk Factors in Endometrial Pathologies: The Role of Clinical, Laboratory Parameters, and Peripheral Blood Inflammatory Indices”

ABSTRACT Objective Postmenopausal bleeding (PMB) and increased endometrial thickness are key clinical indicators that may suggest underlying malignancies. While endometrial biopsy remains the diagnostic gold standard, its invasiveness underscores the need for alternative, noninvasive biomarkers. This study evaluates the potential of clinical, laboratory, and peripheral blood inflammatory indices (PBII) in distinguishing malignant from benign endometrial pathologies. Methods This retrospective study included 162 patients who underwent endometrial biopsy due to PMB and/or increased endometrial thickness between January 2023 and January 2024. Patients were categorized into benign ( n = 134) and malignant ( n = 28) groups. Demographic, clinical, and laboratory parameters were collected, PBII parameters were calculated, and comparisons were performed. Logistic regression analyses were conducted to identify independent predictors of malignancy. Results Malignant cases were significantly associated with older age ( p < 0.001), longer postmenopausal duration ( p = 0.002), higher body mass index (BMI) ( p = 0.018), and greater endometrial thickness ( p = 0.042) compared to benign cases. Hemoglobin levels were significantly lower ( p = 0.022), while neutrophil ( p < 0.001) and monocyte ( p = 0.042) counts were notably higher in malignant cases. Among PBII parameters, neutrophil‐to‐lymphocyte ratio (NLR), systemic immune‐inflammation index (SII), pan‐immune‐inflammation value (PIV), and systemic inflammation response index (SIRI) were significantly elevated ( p < 0.001 for all). Multivariate analysis identified older age ( p < 0.001), lower hemoglobin ( p = 0.016), higher neutrophil count ( p = 0.030), and increased PIV ( p = 0.022) as independent predictors of malignancy. Conclusion Integrating clinical and laboratory parameters with PBII, particularly PIV, may be a valuable, noninvasive tool for the early detection and risk stratification of endometrial malignancies. This approach could enhance diagnostic accuracy, reduce the need for invasive biopsies, and improve patient management.

WIF1 promoter hypermethylation induce endometrial carcinogenesis through the Wnt/β‐catenin signaling pathway

AbstractThis study explores the mechanism underlying WIF1 promoter methylation and its relationship with the pathogenesis of endometrial carcinoma. WIF1 promoter methylation was detected using methylation‐specific polymerase chain reaction (MSP). WIF1 expression was examined through qRT‐PCR and western blotting. Furthermore, 5‐aza‐2′‐deoxycytidine (5‐Aza) was used to demethylate the WIF1 promoter. The roles of WIF1 were investigated using in vitro loss‐ and gain‐of‐function assays. Xenograft models were used to analyze WIF1 expression and downstream genes, and results were confirmed using immunofluorescence and western blotting. WIF1 promoter methylation in endometrial cancer cells was significantly higher than that in normal cells, but the WIF1 mRNA and protein levels were reduced. The expression of WIF1 increased significantly after 5‐Aza treatment (p < .05). Thus, 5‐Aza treatment can inhibit the proliferation of endometrial cancer cells and induce apoptosis, while knockdown of WIF1 significantly inhibits the effects of 5‐Aza. 5‐Aza treatment can also inhibit Wnt pathway genes, including phosphorylation of β‐catenin protein, c‐Myc, and CyclinD1, inhibit downstream functional genes, and activate the tumor suppressor gene APC, which can be blocked by WIF1 knockdown in endometrial carcinoma cells. Finally, 5‐Aza inhibited the proliferation of subcutaneous tumor‐bearing nude mice with endometrial cancer cells, but the effect was weaker than that of WIF1 overexpression. Our research shows that WIF1 promoter hypermethylation may promote the progression of endometrial cancer by downregulating WIF1 expression, activating the Wnt/β‐catenin pathway, and promoting proliferation and inhibiting apoptosis. WIF1 may be a potential biological target for gene therapy and drug development for the treatment of endometrial cancer.

Active Heme Metabolism Suppresses Macrophage Phagocytosis via the TLR4/Type I IFN Signaling/CD36 in Uterine Endometrial Cancer

ABSTRACTBackgroundUterine endometrial cancer (UEC) is a common gynecological estrogen‐dependent carcinoma, usually accompanied by intermenstrual bleeding. Active heme metabolism frequently plays an increasingly important role in many diseases, especially in cancers. Tumor‐associated macrophages (TAMs) are the major population in the immune microenvironment of UEC. However, the roles of heme metabolisms in the crosstalk between UEC cells (UECCs) and macrophages are unclear.Materials and MethodsIn our study, by using TCGA database analysis, integration analysis of the protein–protein interaction (PPI) network and sample RNA transcriptome sequencing were done. The expression level of both heme‐associated molecules and iron metabolism‐related molecules were measured by quantitative real‐time polymerase chain reaction. Heme level detection was done through dehydrohorseradish peroxidase assay. In addition to immunohistochemistry, phagocytosis assay of macrophages, immunofluorescence staining, intracellular ferrous iron staining, as well as enzyme‐linked immune sorbent assay were performed.ResultsIn the study, we verified that heme accumulation in UECCs is apparently higher than in endometrial epithelium cells. Low expression of succinate dehydrogenase B under the regulation of estrogen contributes to over‐production of succinate and heme accumulation in UECC. More importantly, excessive heme in UECCs impaired macrophage phagocytosis by regulation of CD36. Mechanistically, this process is dependent on toll‐like receptor (TLR4)/type I interferons alpha (IFN Iα) regulatory axis in macrophage.ConclusionCollectively, these findings elucidate that active heme metabolism of UECCs directly decreases phagocytosis by controlling the secretion of TLR4‐mediated IFN Iα and the expression of CD36, and further contributing to the immune escape of UEC.

Cytokine mRNA and protein expression by cell cultures of epithelial ovarian cancer—Methodological considerations on the choice of analytical method for cytokine analyses

ProblemTo get a comprehensive picture of cytokine expression in health and disease is difficult, cytokines are transiently and locally expressed, and protein analyses are burdened by biological modifications, technical issues, and sensitivity to handling of samples. Thus, alternative methods, based on molecular techniques for cytokine mRNA analyses, are often used. We compared cytokine mRNA and protein expression to evaluate whether cytokine mRNA profiles can be used instead of protein analyses.Method of studyIn kinetic experiments, cytokine mRNA and protein expression of IL‐1β, IL‐6, IL‐8, TNF‐α, and TNF‐β/LTA were studied using real‐time RT‐qPCR and Luminex® microarrays in the ovarian cancer cell lines OVCAR‐3, SKOV‐3 and the T‐cell line Jurkat, after activation of transcription by thermal stress. In addition, we analyzed IL‐6 and IL‐8 mRNA and protein in a small number of ovarian cancer patients.ResultsOvarian cancer cells can express cytokines on both mRNA and protein level, with 1‐4 hours’ time delay between the mRNA and protein peak and a negative Spearman correlation. The mRNA and protein expression in patient samples was poorly correlated, reflecting previous studies.ConclusionCytokine mRNA and protein expression levels show diverging results, depending on the material analyzed and the method used. Considering the high sensitivity and reproducibility of real‐time RT‐qPCR, we suggest that cytokine mRNA profiles could be used as a proxy for protein expression for some specific purposes, such as comparisons between different patient groups, and in defining mechanistic pathways involved in the pathogenesis of cancer and other pathological conditions.

NKG2D‐mediated cytotoxicity improves after primary surgery for high‐grade serous ovarian cancer

AbstractProblemTumors compromise the patients’ immune system to promote their own survival. We have previously reported that HGSC exosomes play a central role, downregulating NKG2D cytotoxicity. Primary surgery's effect on tumor exosomes and NKG2D cytotoxicity in HGSC patients has not been studied before. The overall objective of this study was to explore the effect of surgery on the exosome‐induced impairment of NKG2D cytotoxicity in HGSC.Method of studyPaired pre‐ and post‐operative blood samples were subjected to cell and exosome analyses regarding the NKG2D receptor and ligands, and NKG2D‐mediated cytotoxicity. Lymphocytes were phenotyped by immunoflow cytometry. Exosomes, isolated by ultracentrifugation, and characterized by nanoparticle tracking analysis, transmission and immune electron microscopy and western blot were used in functional cytotoxic experiments. HGSC explant culture‐derived exosomes, previously studied by us, were used for comparison.ResultsHGSC exosomes from patients’ sera downregulated NKG2D‐mediated cytotoxicity in NK cells of healthy donors. In a subgroup of subjects, NKG2D expression on CTLs and NK cells was upregulated after surgery, correlating to a decrease in the concentration of exosomes in postoperative sera. An overall significantly improved NKG2D‐mediated cytotoxic response of the HGSC patients’ own NK cells in postoperative compared to preoperative samples was noted.ConclusionsSurgical removal of the primary tumor has a beneficial effect, relieving the exosome‐mediated suppression of NKG2D cytotoxicity in HGSC patients, thus boostering their ability to combat cancer.

Knockdown of TRIM47 Overcomes Paclitaxel Resistance in Ovarian Cancer by Suppressing the TGF‐β Pathway via PPM1A

ABSTRACTObjectiveWe investigated the role of tripartite motif 47 (TRIM47) in paclitaxel resistance in ovarian cancer, focusing on its regulation of protein phosphatase magnesium‐dependent 1A (PPM1A), transforming growth factor‐β (TGF‐β) pathway activation, and methyltransferase‐like 3 (METTL3)‐mediated N6‐methyladenosine (m6A) modification.MethodsBioinformatics analysis using Gene Expression Profiling Interactive Analysis (GEPIA), Kaplan–Meier plotter (KM plot), Linkedomics, and sequence‐based RNA adenosine methylation site predictor (SRAMP) databases identified TRIM47 and PPM1A expression patterns, prognostic significance, co‐expression networks, and m6A modification sites. Paclitaxel‐resistant ovarian cancer cell lines were generated. Quantitative reverse transcriptase polymerase chain reaction (qRT‐PCR) and western blot analyzed gene and protein expression. Co‐immunoprecipitation (Co‐IP) and GST pull‐down assays assessed TRIM47‐PPM1A interaction, while cycloheximide (CHX) chase, and IP assays examined PPM1A stability and ubiquitination. RNA immunoprecipitation (RIP) and dual‐luciferase assays determined METTL3’s effect on TRIM47 m6A modification. Functional assays (3‐(4,5‐dimethylthiazol‐2‐yl)‐2,5‐diphenyltetrazolium bromide (MTT), colony formation, and flow cytometry) evaluated proliferation, apoptosis, and drug response. An in vivo xenograft model confirmed TRIM47’s role in chemoresistance.ResultsBioinformatics analysis showed that TRIM47 was overexpressed in ovarian cancer and negatively correlated with the expression of PPM1A. Kaplan–Meier analysis showed that high TRIM47 and low PPM1A expression were correlated with poor prognosis. TRIM47 was upregulated in paclitaxel‐resistant ovarian cancer cells. The knockdown of TRIM47 restored drug sensitivity, inhibited cell proliferation, and induced cell apoptosis. Mechanistically, TRIM47 functioned as an E3 ubiquitin ligase, targeting PPM1A for degradation, leading to sustained TGF‐β signaling and enhanced chemoresistance. CHX chase assays demonstrated reduced PPM1A stability in the presence of TRIM47, while IP‐WB confirmed increased PPM1A ubiquitination. METTL3‐mediated m6A modification enhanced TRIM47 mRNA stability, further promoting its oncogenic role. In vivo, TRIM47 knockdown reduced tumor growth and improved paclitaxel efficacy, reinforcing its role in resistance.ConclusionTRIM47 promoted paclitaxel resistance in ovarian cancer by inducing PPM1A degradation and activating the TGF‐β pathway.

Mechanism of Endometrial Receptivity Affected by Fibroids

ABSTRACT Fibroids are the most common benign tumors of the female reproductive system. Most patients with fibroids are asymptomatic, but the presence of fibroids can still cause some abnormal clinical symptoms, such as increased menstrual volume, abnormal uterine bleeding, pelvic pain, urinary tract and gastrointestinal tract compression symptoms, etc. The impact of fibroids on pregnancy is worth discussing. At present, it is believed that submucosal myoma and intramural myoma affecting uterine cavity shape affect the pregnancy outcome of patients, while the impact of type III intramural myoma on pregnancy is still controversial. A number of studies have found that in addition to direct contact with the endometrial compression, uterine myoma also affects the endometrial flexibility through other ways. In this review, we summarized the effects of fibroids on endometrial receptivity and discussed in depth the mechanisms of such effects, including secretion of cytokines, changes in endometrial blood flow and angiogenesis, effects on endometrial peristalsis and mechanical stress conduction, changes in uterine microecological environment, and abnormal signal transduction pathways. Understanding the mechanism of endometrial receptivity affected by fibroids is significant for exploring the treatment of fibroids, improving the pregnancy outcome of patients with fibroids and increasing the clinical pregnancy rate.

Circulating CD14+HLA‐DRlo/− monocytic cells as a biomarker for epithelial ovarian cancer progression

AbstractProblemPrevious studies identified circulating CD14+HLA‐DRlo/− monocytic cells as an immune suppressive subset in solid malignancies, such as prostate, renal cell carcinoma, and pancreatic cancer. Such monocytic cells have been implicated not only in tumour progression but also as a potential barrier for immunotherapy. This study examined the relationship between the frequency of circulating monocytic cells and epithelial ovarian cancer (EOC) progression pre‐ and post‐frontline chemotherapy, defined by disease stage, which is a leading prognostic factor for this malignancy.Method of studyIncident cases of 236 women with EOC were recruited and comprehensive flow cytometry was utilized to assess the frequency of peripheral blood CD33+CD11b+HLA‐DR−/lowCD14+CD15− monocytic cells, henceforth termed CD14+HLA‐DRlo/− monocytic cells, prior to and after completion of frontline chemotherapy. Multivariable odds ratios (OR) were used to estimate the association between CD14+HLA‐DRlo/− monocytic cell percentages and disease stage. Wilcoxon signed‐rank tests evaluated changes in these monocytic cell levels pre‐ and post‐chemotherapy in a patient subset (n = 70).ResultsPatients with elevated frequencies of circulating CD14+HLA‐DRlo/− monocytic cells at diagnosis were at 3.33‐fold greater odds of having advanced stage (III/IV) EOC (CI: 1.04‐10.64), with a significant trend in increasing CD14+HLA‐DRlo/−monocytic cell levels (P = .04). There was a 2.02% median decrease of these monocytic cells post‐chemotherapy among a subset of patients with advanced stage disease (P < .0001).ConclusionThese findings support the potential clinical relevance of CD14+HLA‐DRlo/−monocytic cells in EOC for prognosis and may indicate a non‐invasive biomarker to measure disease progression.

IL6, IL8, and IL10 in the distinction of malignant ovarian neoplasms and endometriomas

AbstractProblemStudies have shown a relationship between endometriosis and ovarian cancer. Our aims were to evaluate and compare the dosages of cytokines IL‐2, IL‐5, IL‐6, IL‐8, IL‐10, and TNF‐α in serum, intracystic fluid, and peritoneal fluid of patients with ovarian endometrioma, malignant and benign ovarian neoplasms, and non‐neoplastic ovarian tumors; to verify if there is a correlation between the values of these cytokines between ovarian endometrioma and ovarian malignancy; and to determine the best cut‐off point for serum cytokines that can be used to differentiate patients with ovarian malignancy and endometrioma.Method of studyThe concentrations of cytokines were quantified by enzyme‐linked immunosorbent assay (ELISA), analyzed by Kruskal‐Wallis test with the Dunn post‐test. Receiver operating feature (ROC) curve was used to obtain the area under the curve (AUC) and to determine the best cut‐off values that could be used in the diagnosis of ovarian malignancy. Correlations of cytokine concentrations were performed by the Spearman test.ResultsIL‐6, IL‐8, and IL‐10 concentrations were higher in patients with malignant neoplasia. When evaluating the area under the curve (AUC) of serum cytokine levels comparing patients with malignant neoplasia and endometriomas, there was statistical significance for IL‐6, IL‐8, and IL‐10.ConclusionOur results showed utility in serum concentrations of IL‐6, IL‐10, and IL‐8 as parameters that differentiate endometriomas from ovarian malignancies.

Identification of unique clusters of T, dendritic, and innate lymphoid cells in the peritoneal fluid of ovarian cancer patients

AbstractProblemWe hypothesize that activated peritoneal immune cells can be redirected to target ovarian tumors. Here, we obtain fundamental knowledge of the peritoneal immune environment through deep immunophenotyping of T cells, dendritic cells (DC), and innate lymphoid cells (ILC) of ovarian cancer patients.Method of studyT cells, DC, and ILC from ascites of ovarian cancer patients (n = 15) and peripheral blood of post‐menopausal healthy donors (n = 6) were immunophenotyped on a BD Fortessa cytometer using three panels—each composed of 16 antibodies. The data were analyzed manually and by t‐SNE/DensVM. CA125 levels were obtained from patient charts.ResultsWe observed decreased CD3+ T cells and a higher proportion of activated CD4+ and effector memory CD4+/CD8+ T cells, plasmacytoid DC, CD1c+ and CD141+ myeloid DC and CD56Hi NK cells in ascites. t‐SNE/DensVM identified eight T cell, 17 DC, and 17 ILC clusters that were unique in the ascites compared to controls. Hierarchical clustering of cell frequency distinctly segregated the T‐cell and ILC clusters from controls. Increased CA125 levels were associated with decreased CD8+/CD45RA+/CD45RO−/CCR7− T cells.ConclusionThe identified immune clusters serve as the basis for interrogation of the peritoneal immune environment and the development of novel immunologic modalities against ovarian cancer.

Myeloid‐derived suppressor cells in obstetrical and gynecological diseases

AbstractMyeloid‐derived suppressor cells (MDSCs) are a heterogeneous group of myeloid‐origin cells which have immunosuppressive activities in several conditions, such as cancer and inflammation. Recent research has also associated MDSCs with numerous obstetrical and gynecological diseases. During pregnancy, MDSCs accumulate to ensure maternal‐fetal immune tolerance, whereas they are decreased in patients who suffer from early miscarriage or pre‐eclampsia. While the etiology of endometriosis is still unknown, abnormal accumulation of MDSCs in the peripheral blood and peritoneal fluid, alongside an increased level of reactive oxygen species (ROS), has been observed in these patients, which is central to the cellular immune regulations by MDSCs. Additionally, the regulation of MDSCs observed in tumours is also applicable to gynecologic neoplasms, including ovarian cancer and cervical cancer. More recently, emerging evidence has shown that there are high levels of MDSCs in premature ovarian failure (POF) and in vitro fertilization (IVF), but the underlying mechanisms are unknown. In this review, the generation and mechanisms of MDSCs are summarized. In particular, the modulation of these cells in immune‐related obstetrical and gynecological diseases is discussed, including potential treatment options targeting MDSCs.

A gene signature for immune subtyping of desert, excluded, and inflamed ovarian tumors

AbstractProblemThe current tumor immunology paradigm emphasizes the role of the immune tumor microenvironment and distinguishes several histologically and transcriptionally different immune tumor subtypes. However, the experimental validation of such classification is so far limited to selected cancer types. Here, we aimed to explore the existence of inflamed, excluded, and desert immune subtypes in ovarian cancer, as well as investigate their association with the disease outcome.Method of studyWe used the publicly available ovarian cancer dataset from The Cancer Genome Atlas for developing subtype assignment algorithm, which was next verified in a cohort of 32 real‐world patients of a known tumor subtype.ResultsUsing clinical and gene expression data of 489 ovarian cancer patients in the publicly available dataset, we identified three transcriptionally distinct clusters, representing inflamed, excluded, and desert subtypes. We developed a two‐step subtyping algorithm with COL5A2 serving as a marker for separating excluded tumors, and CD2, TAP1, and ICOS for distinguishing between inflamed and desert tumors. The accuracy of gene expression–based subtyping algorithm in a real‐world cohort was 75%. Additionally, we confirmed that patients bearing inflamed tumors are more likely to survive longer.ConclusionOur results highlight the presence of transcriptionally and histologically distinct immune subtypes among ovarian tumors and emphasize the potential benefit of immune subtyping as a clinical tool for treatment tailoring.