Advanced Science

Efferocytosis‐Driven Polyamine Metabolism in Macrophages Enhances Cancer Stem Cell Enrichment after Chemotherapy in Ovarian Cancer

Abstract Chemotherapy‐induced enrichment of cancer stem cells (CSCs) is a key mechanism underlying acquired chemoresistance and recurrence of epithelial ovarian cancer (OC). Although chemotherapy may enrich CSCs through selection or by inducing dedifferentiation, the dynamic changes in the tumor niche and their impact on CSCs during chemotherapy remain unclear. In this study, single‐cell sequencing and multiplex immunohistochemical analysis are used to define microenvironmental changes, and a post‐chemotherapy increase in efferocytotic macrophages that phagocytosed chemotherapy‐induced apoptotic tumor cells is identified. Efferocytotic macrophages are associated with poor prognosis and CSCs in OC. Their conditioned medium facilitates OC stemness in vitro. Meanwhile, targeting efferocytosis suppresses CSC enrichment, chemoresistance, and regrowth in vivo. Mechanistically, it is demonstrated that enhanced expression of ODC1 driven by efferocytosis increases polyamine flux, particularly putrescine, by integrating metabolomics and transcriptomics. The increase in putrescine content leads to the SPP1 and OPN overexpression in macrophages, conferring cancer stemness to OC cells through the OPN‐CD44 axis. Treatment with an ODC1 selector inhibitor mitigates CSC enrichment, sensitizes tumors to cisplatin, and restricts tumor regrowth. Together, the study shows that efferocytosis and associated polyamine metabolic reprogramming support the chemotherapy‐induced enrichment of CSCs, providing new targets for addressing chemoresistance and recurrence of OC.

Evolution of Multivalent Aptamer Corona for High‐Throughput Multiplexed Detection of Multiple Cancers

Abstract Clinical in vitro diagnosis is essential in early diagnosis and prognostic assessment, yet antibody‐based assays face limitations in multiplexity and scalability. To address the growing demand for modern molecular diagnostics, a multivalent aptamer corona platform for high‐throughput is introduced and multiplexed multi‐cancer diagnosis. With clinical serum samples, ovarian cancer‐, lung cancer‐, and colorectal cancer‐based aptamer coronas are obtained through several rounds of alternating positive and negative systematic evolution of ligands by exponential enrichment (SELEX). Leveraging a de novo ProteoFish‐SELEX strategy, which integrates nanoparticle‐protein corona technology, a multivalent aptamer corona is evolved. Proteomic profiling of protein coronas revealed cancer‐associated protein signatures, while aptomic profiling of aptamer coronas identified high‐affinity cancer‐specific aptamers. With these aptamers, high diagnostic accuracy is demonstrated in multiplexed multi‐cancer detection using clinical cohorts. Multivalent aptamer corona's outstanding diagnostic performance, multiplexed detection capability, and sequencing‐enabled high‐throughput potential position it as a promising versatile tool for novel biomarker discovery and a transformative advancement in precision oncology.

AI‐Guided SERS Defines a Pan‐Cancer Diagnostic Biomarker

Abstract Early and accurate detection of multiple cancers through a single test remains an unmet clinical need, hindered by current limitations in accuracy, throughput, automation, and multiplexing. Here, we present an AI‐powered SERS chip that combines automated exosome capture with AI‐enabled molecular fingerprinting to accurately distinguish ten common cancer types from a single serum test. The system employs a peptide‐functionalized SERS chip enabling the selective enrichment of exosomes directly from patient serum, enhancing label‐free Raman fingerprint signals. AI‐driven spectral analysis achieved 97.4% accuracy in distinguishing cancer from healthy samples, 97.08% accuracy for early‐stage cancer detection, and 93.89% accuracy in classifying ten common cancer types, including breast, thyroid, esophageal, kidney, pancreatic, duodenal, lung, colorectal, ovarian, and gastric cancers. Crucially, based on molecular profiling, we identified exosomal deoxyadenosine triphosphate as a promising pan‐cancer biomarker consistently upregulated across diverse tumor types. This discovery establishes a potential pan‐cancer diagnostic marker, while the fully automated, scalable platform offers significant promise for clinical translation in early and differential cancer diagnosis.

Exosomal CMTM4 Induces Immunosuppressive Macrophages to Promote Ovarian Cancer Progression and Attenuate Anti‐PD‐1 Immunotherapy

Abstract Exosomes shape the tumor microenvironment (TME) by modulating tumor‐associated macrophages (TAMs) and promoting ovarian cancer (OC) progression. This study reveals that exosomal CKLF Like MARVEL Transmembrane Domain Containing 4 (CMTM4) enhances OC malignancy and orchestrates immune evasion. Excessive macrophage infiltration in the TME, particularly in the presence of CMTM4, is strongly associated with poor prognosis. Within the TME, exosomal CMTM4 is actively internalized by macrophages, promoting M2 polarization and subsequently initiating immunosuppressive signaling. Exosomal CMTM4 activates the NF‐κB pathway in TAMs, suppressing immune function through enhanced secretion of cytokines, including TGF‐β1 and CXCL12, while simultaneously upregulating intercellular adhesion molecule‐1 (ICAM1) expression to further promote M2 polarization and facilitate cancer metastasis. Depletion of CMTM4 increases sensitivity to anti‐PD‐1 therapy by reversing immunosuppression. Notably, eltrombopag is identified as a CMTM4 inhibitor that attenuates OC progression in vivo and modulates the tumor immune microenvironment, synergizing with PD‐1 blockade immunotherapy to enhance therapeutic efficacy. The exosomal CMTM4—ICAM1—CD206 axis exacerbates disease risk in patients with OC. Collectively, the study highlights the critical role of tumor‐derived exosomal CMTM4 in immune suppression, emphasizing its potential as both a prognostic biomarker and a therapeutic target in OC immunotherapy.

Tumor Intrinsic METTL5 Modulates ATF4 Translation to Prevent T Cell‐Induced Ferroptosis in Ovarian Cancer

Abstract Poor clinical responses to immune checkpoint blockade (ICB) observed in ovarian cancer (OC) highlight an unmet need to understand the mechanisms driving immune evasion in this disease. To address this, an integrative analysis is conducted by combining in vitro genome‐wide immune screens, in vivo ICB screens, and clinical data mining, and METTL5 is identified as a crucial OC‐intrinsic factor that promotes immune resistance. Immunologically “cold” OC tumors and poor responders to ICB exhibit elevated METTL5 expression. Mechanistically, knocking out (KO) METTL5 in OC disrupts ATF4 translation by altering 18S rRNA m 6 A levels, leading to the downregulation of SLC7A11 and SLC3A2 , whose function is to suppress ferroptosis activity. Consequently,  METTL5 KO enhances tumor sensitivity to T cell‐mediated antitumor immunity. Notably, the immune‐sensitive phenotypes seen in METTL5 ‐KO tumors can be reversed by either ATF4 overexpression or ferroptosis inhibition. These findings underscore the central role of the METTL5/ATF4/ferroptosis axis in controlling OC responses to immunotherapy.

Fluorescence Lifetime Imaging for Quantification of Targeted Drug Delivery in Varying Tumor Microenvironments

Abstract Trastuzumab (TZM) is a monoclonal antibody that targets the human epidermal growth factor receptor 2 (HER2) and is clinically used for the treatment of HER2‐positive breast tumors. However, the tumor microenvironment can limit the access of TZM to the HER2 targets across the whole tumor and thereby compromising TZM's therapeutic efficacy. An imaging methodology that can non‐invasively quantify the binding of TZM‐HER2, which is required for therapeutic action, and distribution within tumors with varying tumor microenvironments is much needed. Near‐infrared (NIR) fluorescence lifetime (FLI) Forster Resonance Energy Transfer (FRET) is performed to measure TZM‐HER2 binding, using in vitro microscopy and in vivo widefield macroscopy, in HER2 overexpressing breast and ovarian cancer cells and tumor xenografts, respectively. Immunohistochemistry is used to validate in vivo imaging results. NIR FLI FRET in vitro microscopy data show variations in intracellular distribution of bound TZM in HER2‐positive breast AU565 and AU565 tumor‐passaged XTM cell lines in comparison to SKOV‐3 ovarian cancer cells. Macroscopy FLI (MFLI) FRET in vivo imaging data show that SKOV‐3 tumors display reduced TZM binding compared to AU565 and XTM tumors, as validated by ex vivo immunohistochemistry. Moreover, AU565/XTM and SKOV‐3 tumor xenografts display different amounts and distributions of TME components, such as collagen and vascularity. Therefore, these results suggest that SKOV‐3 tumors are refractory to TZM delivery due to their disrupted vasculature and increased collagen content. The study demonstrates that FLI is a powerful analytical tool to monitor the delivery of antibodydrugs both in cell cultures and in vivo live systems. Especially, MFLI FRET is a unique imaging modality that can directly quantify target engagement with the potential to elucidate the role of the TME in drug delivery efficacy in intact live tumor xenografts.

Splicing Factor PQBP1 Curtails BAX Expression to Promote Ovarian Cancer Progression

Abstract Splicing factor polyglutamine binding protein‐1 (PQBP1) is abundantly expressed in the central nervous system during development, and mutations in the gene cause intellectual disability. However, the roles of PQBP1 in cancer progression remain largely unknown. Here, it is shown that PQBP1 overexpression promotes tumor progression and indicates worse prognosis in ovarian cancer. Integrative analysis of spyCLIP‐seq and RNA‐seq data reveals that PQBP1 preferentially binds to exon regions and modulates exon skipping. Mechanistically, it is shown that PQBP1 regulates the splicing of genes related to the apoptotic signaling pathway, including BAX. PQBP1 promotes BAX exon 2 skipping to generate a truncated isoform that undergoes degradation by nonsense‐mediated mRNA decay, thus making cancer cells resistant to apoptosis. In contrast, PQBP1 depletion or splice‐switching antisense oligonucleotides promote exon 2 inclusion and thus increase BAX expression, leading to inhibition of tumor growth. Together, the results demonstrate an oncogenic role of PQBP1 in ovarian cancer and suggest that targeting the aberrant splicing mediated by PQBP1 has therapeutic potential in cancer treatment.

Discovery of Natural Compound α‐Hederin via Large‐Scale Screening as a Targeted JAK/STAT3 Inhibitor for Ovarian Cancer Therapy

Abstract Chemoresistance and metastasis are key obstacles to successful ovarian cancer (OC) treatment. Here, α‐Hederin, a pentacyclic triterpenoid saponin, is identified as a potent and selective dual inhibitor of JAK1/JAK2 with promising therapeutic potential in OC. Integrating transcriptomic analysis, virtual screening, molecular docking, and biochemical validation, it is shown that α‐Hederin directly binds the JH1 kinase domains of JAK1 and JAK2, suppressing their activity and downstream STAT3 phosphorylation. α‐Hederin inhibits OC cell proliferation, epithelial‐mesenchymal transition (EMT), and metastasis in vitro, and suppresses tumor growth and dissemination in multiple mouse models, with minimal systemic toxicity. Mechanistically, α‐Hederin blocks STAT3 nuclear translocation and downregulates oncogenic STAT3 targets including MYC, CCND1, and TWIST1. Rescue experiments using the STAT3 agonist Colivelin partially reversed these effects, confirming the JAK/STAT3 axis as a key target. Moreover, α‐Hederin synergizes with cisplatin to enhance antitumor efficacy and overcomes platinum resistance in OC cells. Collectively, our findings highlight α‐Hederin as a safe and effective natural JAK1/2 inhibitor that suppresses OC progression by targeting the JAK/STAT3 pathway, offering a compelling candidate for future clinical translation.

Citrullination of AKT2 Catalyzed by PAD1 Facilitates the Maintenance of Stemness Characteristics of Ovarian Cancer Stem‐Like Cells in Ovarian Cancer

Abstract The presence of ovarian cancer stem‐like cells (OCSLCs) is a crucial driving force for malignant progression, metastasis, and tumor recurrence of ovarian cancer (OC). However, the mechanisms underlying activation and maintenance of OCSLC stemness remain unclear. Here, it is identified an increased expression of peptidylarginine deiminase 1 (PAD1) in OC cells, particularly within the CD133 + subset or cisplatin‐resistant cells, which is positively associated with the upregulation of stemness‐related markers and maintenance of stemness in OCSLCs. Mechanistically, PAD1 specifically binds to the kinase domain of AKT2 and catalyzes citrullination at R202. Citrullination subsequently promotes AKT2 phosphorylation at S474 and T309, the two critical residues for AKT2 kinase activity. As a major driver of OC malignancy, the activation of AKT2 leads to an increased expression of CCAAT/Enhancer Binding Protein Beta (CEBPβ), thereby promoting CEBPβ enrichment to the promoters of a subset of stemness‐related genes. Moreover, PAD1 gene silencing, inhibition of AKT2 citrullination, and AKT2 mutation all decrease the tumor‐initiating ability of OC cells both in vitro and in vivo. Importantly, treatment with a PAD1 inhibitor can resensitize cisplatin‐resistant OC cells to cisplatin treatment, suggesting that targeting PAD1/AKT2/CEBPβ signaling axis in OCSLCs may be highly effective in preventing OC progression.

eIF4E Enriched Extracellular Vesicles Induce Immunosuppressive Macrophages through HMGCR‐Mediated Metabolic Rewiring

Abstract Tumor driven immune suppression poses a significant impediment to the success of immunotherapy in ovarian cancer. Among the various mechanisms contributing to immune suppression, intracellular communication facilitated by tumor‐derived extracellular vesicles (EVs) within the tumor microenvironment emerges as a pivotal factor influencing tumor growth. Here, it is demonstrated that extracellular vesicle‐packaged eIF4E from tumor cells alters protein translation in macrophages, contributing to antitumor immune response. Mechanistically, tumor derived EV‐packaged eIF4E significantly enhances the expression of 3‐hydroxy‐3‐methyl‐glutaryl‐coenzyme A reductase (HMGCR), driving the synthesis and secretion of cholesterol. This, in turn, activates macrophages and causes immunosuppression through the X‐box binding protein 1 and Programmed death‐ligand 1 (XBP1/PD‐L1) axis. Strikingly, both genetic and pharmacological depletion of HMGCR in macrophages effectively restores their antitumor activity. Clinically, elevated HMGCR expression in tumor‐associated macrophages is associated with poor survival outcomes in ovarian cancer patients. The pivotal role of eIF4E is underscored here as a key signaling mediator, facilitating the communication between tumor and immune cells via EVs to promote immune suppression and suggesting HMGCR as a potential therapeutic target for tumor immunotherapy.

Glucose Deprivation‐Induced Disulfidptosis via the SLC7A11‐INF2 Axis: Pan‐Cancer Prognostic Exploration and Therapeutic Validation

Abstract Disulfidptosis, a novel form of regulated cell death, involves cytoskeletal collapse due to excessive disulfide bond formation, linking metabolism and reactive oxygen species to potential cancer therapy targets. Recent multi‐omics studies highlight the prognostic value of disulfidptosis‐related gene (DRG) signatures in pan‐cancers; however, the molecular mechanisms underlying their biological functions and therapeutic relevance remain poorly defined. Herein, a DRG score model is constructed using LASSO Cox regression across 33 cancer types, and a nomogram incorporating the DRG score is developed for prognostic prediction. The tumor microenvironment, mutation profiles, and immunotherapy responses are analyzed. The DRG score serves as an independent prognostic factor across cancers, correlating with poor outcomes and malignant features. Glucose deprivation induces disulfidptosis in SLC7A11 high cells (high SLC7A11 expression), especially in cancers with a high DRG score, such as ovarian cancer. Silencing INF2 prevents disulfidptosis and decreases susceptibility to irofulven, which can be reversed by GLUT inhibitors. SLC7A11 knockdown reduces disulfidptosis, restores ATP/NADPH levels, and protects the cytoskeleton under glucose deprivation, whereas INF2 knockdown impairs cell migration. Moreover, the DRG scores predict prognosis and therapeutic responses. The SLC7A11‐INF2 axis regulates disulfidptosis, migration, and drug sensitivity, highlighting its potential as a marker of metabolic vulnerability in ovarian cancer.

Mortalin and PINK1/Parkin‐Mediated Mitophagy Represent Ovarian Cancer‐Selective Targets for Drug Development

Abstract Mortalin is an essential chaperone for the import of nuclear‐encoded proteins into mitochondria and is elevated in ovarian cancer in association with poor patient prognosis. The investigational new drug, SHetA2, interacts with mortalin releasing its client proteins. In this study, interactions of SHetA2 moieties and mortalin substrate binding domain (SBD) amino acids are demonstrated by surface plasmon resonance (SPR) and nuclear magnetic resonance (NMR) to occur at low micromolar SHetA2 concentrations that selectively kill cancer cells over noncancerous cells. In both ovarian cancer and noncancerous cells SHetA2 reduces: mitochondria import of mortalin, degradation of mortalin's mitochondrial localization sequence (MLS), mortalin/inositol 1,4,5‐trisphosphate receptors complexes and oxidative phosphorylation. In cancer cells only, SHetA2 reduces calcium levels, mitochondrial length and fusion proteins, while inducing autophagy and PTEN‐induced kinase 1 (PINK1)/PARKIN‐mediated mitophagy. Noncancerous cells exhibit increased mitochondrial branch length in response to SHetA2 and a low level of inducible autophagy that is resistant to SHetA2. Inhibition of autophagosome‐lysosome fusion reduces, or increases, SHetA2 cytotoxicity in ovarian cancer or noncancerous cells, respectively. SHetA2 inhibits mortalin and growth, and induces mitophagy in ovarian cancer xenografts and increases survival post‐surgical tumor removal. In conclusion, SHetA2 binds directly to mortalin's SBD and causes distinct responses in ovarian cancer and noncancerous cells.

Fn14 Controls the SIRT2‐Mediated Deacetylation of Slug to Inhibit the Metastasis of Epithelial Ovarian Cancer

AbstractMetastatic spread of cancer is the leading cause of death in patients with epithelial ovarian cancer (EOC), and elucidation of the molecular mechanisms underlying this process is a major focus of cancer research. Fibroblast growth factor‐inducible 14 (Fn14) has been shown to regulate wound repair, inflammation, angiogenesis, and chemoresistance, but its functional role in metastasis in EOC is still unknown. Here it is reported that Fn14 is identified as a cancer metastasis suppressor that inhibits the migratory and invasive potential of EOC cells by down‐regulating epithelial‐mesenchymal transition (EMT). Mechanistically, it is identified that Fn14 promotes acetylation‐dependent protein degradation of Slug, a key transcriptional factor associated with EMT. The deacetylase Sirtuin 2 (SIRT2) has been reported to be involved in the deacetylation of Slug protein to stabilize it and then prevent its degradation in the nucleus. The results showed that Fn14 alters the subcellular localization of (SIRT2) by interacting with SIRT2, leading to reduced SIRT2 shuttling into the nucleus and subsequently promoting the acetylated degradation of Slug. Collectively, the work has demonstrated for the first time that Fn14 inhibits EOC metastasis by regulating SIRT2‐mediated Slug deacetylation, providing a new perspective and method for the development of future novel therapeutic strategies for the treatment of EOC metastasis.

Tumor Small Extracellular Vesicle‐Transmitted LncRNA CATED Promotes Platinum‐Resistance in High‐Grade Serous Ovarian Cancer

AbstractHigh‐grade serous ovarian cancer (HGSOC) is the most lethal type of gynecological cancer, and platinum‐resistance is a serious challenge in its treatment. Long non‐coding RNAs (lncRNAs) play critical regulatory roles in the occurrence and development of cancers. Here, using RNA sequencing of tumor small extracellular vesicles (sEVs) from HGSOC patients, the lncRNA CATED is identified as significantly upregulated in both tumors and tumor‐derived sEVs in platinum‐resistant HGSOC, and low CATED levels correlate with good prognosis. Functionally, CATED enhances cisplatin resistance by promoting cell proliferation and inhibiting apoptosis in vitro and in vivo. These effects could be transferred via CATED‐overexpressing sEVs from donor cells and HGSOC tumor sEVs. Mechanistically, CATED binds to and upregulates DHX36 via PIAS1‐mediated SUMOylation at the K105 site, and elevated DHX36 levels increase downstream RAP1A protein levels by enhancing RAP1A mRNA translation, consequently activating the MAPK pathway to promote platinum‐resistance in HGSOC. Antisense oligonucleotide mediated knockdown of CATED reverse platinum‐resistance in sEV‐transmitted mouse models via the DHX36‐RAP1A‐MAPK pathway. This study newly identifies a sEV‐transmitted lncRNA CATED in driving HGSOC platinum‐resistance and elucidates the mechanism it regulates the interacting protein through SUMOylation. These findings also provide a novel strategy for improving chemotherapy in HGSOC by targeting CATED.

HPD is an RNA‐Binding Protein Sustaining Ovarian Cancer Cell Glycolysis, Tumor Growth, and Drug Resistance

Abstract 4‐hydroxyphenylpyruvate dioxygenase (HPD) is an important metabolic enzyme in the tyrosine metabolic pathway and displays aberrant expression and function in cancer. Unexpectedly, it is discovered that HPD functions as an RNA‐binding protein (RBP) to drive ovarian cancer progression. HPD is shown to bind to the RRACH motif of these target mRNAs through its two dsRNA binding domains (RBDs), resulting in increased global mRNA translation. In particular, HPD binding is demonstrated to mediate translation of glycolytic enzymes triosephosphate isomerase (TPI) and alpha‐enolase (ENO1) mRNAs, which facilitates ovarian cancer glycolysis and tumor growth. Thus, targeting the RBD domain of HPD disrupts its RNA binding ability, leading to blocking glycolysis flux, tumor growth, and enhancing drug response. HPD is a novel RNA‐binding protein, and this moonlighting function highlights the knowledge of HPD in regulating cancer development and drug response beyond only as a metabolic enzyme.

Chromatin Organization Governs Transcriptional Response and Plasticity of Cancer Stem Cells

Abstract Chromatin organization regulates transcription to influence cellular plasticity and cell fate. We explored whether chromatin nanoscale packing domains are involved in stemness and response to chemotherapy. Using an optical spectroscopic nanosensing technology we show that ovarian cancer‐derived cancer stem cells (CSCs) display upregulation of nanoscale chromatin packing domains compared to non‐CSCs. Cleavage under targets and tagmentation (CUT&Tag) sequencing with antibodies for repressive H3K27me3 and active H3K4me3 and H3K27ac marks mapped chromatin regions associated with differentially expressed genes. More poised genes marked by both H3K4me3 and H3K27me3 were identified in CSCs vs. non‐CSCs, supporting increased transcriptional plasticity of CSCs. Pathways related to Wnt signaling and cytokine‐cytokine receptor interaction were repressed in non‐CSCs, while retinol metabolism and antioxidant response were activated in CSCs. Comparative transcriptomic analyses showed higher intercellular transcriptional heterogeneity at baseline in CSCs. In response to cisplatin, genes with low baseline expression levels underwent the highest upregulation in CSCs, demonstrating transcriptional plasticity under stress. Epigenome targeting drugs downregulated chromatin packing domains and promoted cellular differentiation. A disruptor of telomeric silencing 1‐like (Dot1L) inhibitor blocked transcriptional plasticity, reversing stemness. These findings support that CSCs harbor upregulated chromatin packing domains, contributing to transcriptional and cell plasticity that epigenome modifiers can target.

The INAVA mRNA in Extracellular Vesicles Activates Normal Ovarian Fibroblasts by Phosphorylation–Ubiquitylation Crosstalk of HMGA2

Abstract Ovarian cancer is an aggressive gynecological tumor usually diagnosed with widespread metastases. Extracellular vesicles (EVs), though recognized as important mediators of tumor metastasis, have received limited attention into their specific functions via the mRNA profiling. Here it is reported elevated expression and selective enrichment of INAVA mRNA in both plasma‐ and tissue‐derived EVs from ovarian cancer patients, which is positively correlated with distant metastasis and poor prognosis. Functionally, INAVA mRNA, upon uptake and translation, activates normal ovarian fibroblasts (NOFs) and drives extensive peritoneum metastasis in the orthotopic xenograft mouse model. Mechanistically, INAVA competitively binds with high mobility group protein A2 (HMGA2) and consequently inhibit its interaction with vaccinia‐related kinase 1 (VRK1), leading to reduced HMGA2 phosphorylation on Ser105. Interestingly, this inhibitory phosphorylation stabilizes HMGA2 via blocking tripartite motif‐containing 21 (TRIM21) ‐mediated K48‐linked ubiquitylation, and ultimately enhances the transcription of STAT3 to activate NOFs. Lastly, a cell‐permeable peptide that disrupts the INAVA–HMGA2 interaction leads to attenuated NOF activation and provides a promising strategy for ovarian cancer therapy.

Radiotherapy‐Associated Cellular Senescence and EMT Alterations Contribute to Distinct Disease Relapse Patterns in Locally Advanced Cervical Cancer

AbstractA notable number of locally advanced cervical carcinoma (LACC) patients experience local or distant disease relapse following radiotherapy. The contribution of tumor microenvironment (TME) to tumor recurrence at different sites remains unclear. Here, single‐nucleus RNA sequencing data from 28 pre‐ and on‐treatment LACC samples from patients with different disease relapse patterns is analyzed. The findings revealed opposing alterations in the expression levels of the cellular senescence pathway after radiotherapy in patients with local and distant relapses. In contrast, an increase in the expression of the epithelial‐mesenchymal transition module after radiotherapy in both relapse groups is observed. Cell–cell interactions, drug‐target expression analyses in malignant cells after radiation, and multiplex immunofluorescence of tumor tissue identified interleukin‐1 receptor type I (IL1R1) as a potential therapeutic target. It is demonstrated that combining the IL1R1 inhibitor anakinra with radiation can mitigate the effects of radiation on tumor cells. This study highlights the distinct roles of cellular senescence and EMT in tumor recurrence.

Targeting Replication Fork Processing Synergizes with PARP Inhibition to Potentiate Lethality in Homologous Recombination Proficient Ovarian Cancers

AbstractSynthetic lethality in homologous recombination (HR)‐deficient cancers caused by Poly (ADP‐ribose) polymerase inhibitors (PARPi) has been classically attributed to its role in DNA repair. The mode of action of PARPi and resistance thereof are now believed to be predominantly replication associated. Therefore, effective combinatorial approaches of targeting replication fork processing along with HR‐downregulation to target HR‐proficient and possibly PARPi‐resistant tumors are warranted. Stilbenes are a privileged class of molecules, which include resveratrol, pterostilbene, piceatannol, etc, that modulate both replication processes and RAD51‐expression. In this investigation, by screening a small library of stilbenes, including in‐house synthesized molecules, trans‐4,4′‐dihydroxystilbene (DHS) was discovered as a potent natural agent, which downregulates RAD51 expression and HR repair (GFP‐reporter assay). DHS induces extensive synergistic cell death in ovarian cancers when combined with talazoparib (PARPi). Mechanistically, DHS elicits replication‐stress through severely impeding replication fork progress, speed, and inducing fork‐asymmetry. This leads to robust induction of single stranded DNA (ssDNA) gaps and poly‐ADP‐ribosylation (PARylation) in S‐phase cells, signifying issues related to lagging (Okazaki) strand synthesis. PARPi, which abrogates PARylation, potentiates DHS induced ssDNA gaps, and their conversion into lethal double strand breaks through MRE11 action. Furthermore, the combination is highly effective in mitigating ovarian tumor xenograft growth in SCID mice and exhibited a good therapeutic‐index with no/minimal tissue‐toxicity.

Targeting the SPC25/RIOK1/MYH9 Axis to Overcome Tumor Stemness and Platinum Resistance in Epithelial Ovarian Cancer

AbstractIn epithelial ovarian cancer (EOC), platinum resistance, potentially mediated by cancer stem cells (CSCs), often leads to relapse and treatment failure. Here, the role of spindle pole body component 25 (SPC25) as a key determinant promoting stemness and platinum resistance in EOC cells, with its expression being correlated with adverse clinical outcomes is delineated. Mechanistically, SPC25 acts as a scaffolding platform, orchestrating the assembly of an SPC25/RIOK1/MYH9 trimeric complex, triggering RIOK1‐mediated phosphorylation of MYH9 at Ser1943. This prompts MYH9 to disengage from the cytoskeleton, augmenting its nuclear accumulation, thus potentiating CTNNB1 transcription and subsequent activation of Wnt/β‐catenin signaling. CBP1, a competitive inhibitory peptide, can disrupt the formation of the aforementioned trimeric complex, diminishing the activity of the SPC25/RIOK1/MYH9 axis–mediated Wnt/β‐catenin signaling, and thus attenuate CSC phenotypes, thereby enhancing platinum efficacy in vitro, in vivo, and in patient‐derived organoids. Therefore, targeting the SPC25/RIOK1/MYH9 axis, which mediates the maintenance of stemness and platinum resistance in EOC cells, may enhance platinum sensitivity and increase survival in patients with EOC.

ACAT1‐Mediated ME2 Acetylation Drives Chemoresistance in Ovarian Cancer by Linking Glutaminolysis to Lactate Production

Abstract Lactate derived from aerobic glycolysis is crucial for DNA damage repair and chemoresistance. Nevertheless, it is frequently noted that cancer cells depend on glutaminolysis to replenish essential metabolites. Whether and how glutaminolysis might enhance lactate production and facilitate DNA repair in cancer cells remains unknown. Here, it is shown that malate enzyme 2 (ME2), which metabolizes glutamine‐derived malate to pyruvate, contributes to lactate production and chemotherapy resistance in ovarian cancer. Mechanistically, chemotherapy reduces the expression of glucose transporters and impairs glucose uptake in cancer cells. The resultant decrease in intracellular glucose levels triggers the acetylation of ME2 at lysine 156 by ACAT1, which in turn potentiates ME2 enzyme activity and facilitates lactate production from glutamine. ME2‐derived lactate contributes to the development of acquired chemoresistance in cancer cells subjected to prolonged chemotherapy, primarily by facilitating the lactylation of proteins involved in homologous recombination repair. Targeting ACAT1 to inhibit ME2 acetylation effectively reduced chemoresistance in both in vitro and in vivo models. These findings underscore the significance of acetylated ME2‐mediated lactate production from glutamine in chemoresistance, particularly under conditions of reduced intracellular glucose within cancer cell, thereby complementing the Warburg effect and offering new perspectives on the metabolic links to chemotherapy resistance.

CpG‐Based Nanovaccines Enhance Ovarian Cancer Immune Response by Gbp2‐Mediated Remodeling of Tumor‐Associated Macrophages

AbstractCpG oligodeoxynucleotides (CpG), as an immunoadjuvant, can facilitate the transformation of tumor‐associated macrophages (TAMs)into tumoricidal M1 macrophages. However, the accumulation of free CpG in tumor tissues remains a substantial challenge. To address this, a nanovaccine (PLGA‐CpG@ID8‐M) is engineered by encapsulating CpG within PLGA using ID8 ovarian cancer cell membranes (ID8‐M). This nanovaccine demonstrates remarkable efficacy in reprogramming TAMs in ovarian cancer and significantly extends survival in ID8‐bearing mice. Notably, these findings indicate that the nanovaccine can also mitigate chemotherapy‐induced immunosuppression by increasing the proportion of M1‐like TAMs and reducing the expression of CD47 on tumor cells, thereby achieving a synergistic effect in tumor immunotherapy. Mechanistically, through transcriptome sequencing (RNA‐seq), single‐cell RNA sequencing (scRNA‐seq), and mass spectrometry‐based proteomics, it is elucidated that the nanovaccine enhances the expression of Gbp2 and promotes the recruitment of Pin1, which activates the NFκB signaling pathway, leading to the M1 polarization of TAMs. Furthermore, macrophages with elevated Gbp2 expression significantly inhibit tumor growth in both ID8 ovarian cancer and 4T1 breast cancer models. Conversely, targeting Gbp2 diminishes the antitumor efficacy of the nanovaccine in vivo. This study offers an innovative approach to immunotherapy and elucidates a novel mechanism (Gbp2‐Pin1‐NFκB pathway) for remodeling TAMs.

PLIN2 Promotes Lipid Accumulation in Ascites‐Associated Macrophages and Ovarian Cancer Progression by HIF1α/SPP1 Signaling

AbstractA major characteristic of ovarian cancer (OC) is its unique route of metastasis via ascites. The immune microenvironment in ascites remains understudied, leaving the mechanism of ascites‐mediated abdominal metastasis obscure. Here, a single‐cell transcriptomic landscape of CD45+ immune cells across multiple anatomical sites is depicted, including primary tumors, metastatic lesions, and ascites, from patients diagnosed with high‐grade serous ovarian carcinoma (HGSOC). A novel subset of perilipin 2 high (PLIN2hi) macrophages are identified that are enriched in ascites and positively correlated with OC progression, hence being designated as “ascites‐associated macrophages (AAMs)”. AAMs are lipid‐loaded with overexpression of the lipid droplet protein PLIN2. Overexpression or suppression of PLIN2 can enhance or inhibit tumor cell migration, invasion, and vascular permeability in vitro, which is also confirmed in vivo. Mechanistically, it is demonstrated that PLIN2 boosts HIF1α/SPP1 signaling in macrophages, thereby exerting pro‐tumor functions. Finally, a PLIN2‐targeting liposome is designed to efficiently suppress ascites production and tumor metastasis. Taken together, this work provides a comprehensive characterization of the cancer‐promoting function and lipid‐rich property of ascites‐enriched PLIN2hi macrophages, establishes a link between lipid metabolism and hypoxia within the context of the ascites microenvironment, and elucidates the pivotal role of ascites in trans‐coelomic metastasis of OC.

CircMETTL6 Suppresses Ovarian Cancer Cell Growth and Metastasis Through Inhibition of GDF15 Transcription by Disrupting the NONO‐POLR2A Complex

AbstractCircular RNAs (circRNAs) are a distinctive class of non‐coding RNAs with covalent closed‐loop structure, lacking 5′ caps and 3′ poly(A) tails. These molecules are prevalent in eukaryotes and play key roles in cancer. Here, the function of a new circRNA, circMETTL6, in ovarian cancer is identified and investigated. The prognostic significance of circMETTL6 is assessed using RNA in situ hybridization. Functional studies involving circMETTL6 overexpression are performed both in vitro and in vivo. Mechanistic investigations are performed using RNA‐seq, RNA pull‐down, RNA immunoprecipitation, co‐immunoprecipitation, chromatin immunoprecipitation, protein degradation assay and dual‐luciferase reporter assays. circMETTL6 is significantly downregulated in ovarian cancer, and its lower expression correlates with worse prognosis. Overexpression of circMETTL6 significantly inhibited proliferation, migration, and invasion of ovarian cancer cell in vitro, as well as tumor growth and metastasis in vivo. Mechanistically, circMETTL6 recruited the non‐POU domain containing octamer binding protein (NONO) by binding to its Coiled‐coil domain and disrupted its binding with RNA polymerase II subunit A (POLR2A), and consequently inhibiting growth differentiation factor 15 (GDF15) transcription, thereby suppressing ovarian cancer progression. These findings establish circMETTL6 as a novel tumor suppressor in ovarian cancer. Targeting the circMETTL6/NONO/GDF15 axis presents a potential therapeutic avenue for ovarian cancer treatment.

CDC7 Inhibition Potentiates Antitumor Efficacy of PARP Inhibitor in Advanced Ovarian Cancer

AbstractPoly (ADP‐ribose) Polymerase inhibitors (PARPi) have demonstrated remarkable clinical efficacy in treating ovarian cancer (OV) with BRCA1/2 mutations. However, drug resistance inevitably limits their clinical applications and there is an urgent need for improved therapeutic strategies to enhance the clinical utility of PARPi, such as Olaparib. Here, compelling evidence indicates that sensitivity of PARPi is associated with cell cycle dysfunction. Through high‐throughput drug screening with a cell cycle kinase inhibitor library, XL413, a potent cell division cycle 7 (CDC7) inhibitor, is identified which can synergistically enhance the anti‐tumor efficacy of Olaparib. Mechanistically, the combined administration of XL413 and Olaparib demonstrates considerable DNA damage and DNA replication stress, leading to increased sensitivity to Olaparib. Additionally, a robust type‐I interferon response is triggered through the induction of the cGAS/STING signaling pathway. Using murine syngeneic tumor models, the combination treatment further demonstrates enhanced antitumor immunity, resulting in tumor regression. Collectively, this study presents an effective treatment strategy for patients with advanced OV by combining CDC7 inhibitors (CDC7i) and PARPi, offering a promising therapeutic approach for patients with limited response to PARPi.

Tau Aggregation‐Dependent Lipid Peroxide Accumulation Driven by the hsa_circ_0001546/14‐3‐3/CAMK2D/Tau Complex Inhibits Epithelial Ovarian Cancer Peritoneal Metastasis

Abstract Intraperitoneal dissemination is the main method of epithelial ovarian cancer (EOC) metastasis, which is related to poor prognosis and a high recurrence rate. Circular RNAs (circRNAs) are a novel class of endogenous RNAs with covalently closed loop structures that are implicated in the regulation of tumor development. In this study, hsa_circ_0001546 is downregulated in EOC primary and metastatic tissues vs. control tissues and this phenotype has a favorable effect on EOC OS and DFS. hsa_circ_0001546 can directly bind with 14‐3‐3 proteins to act as a chaperone molecule and has a limited positive effect on 14‐3‐3 protein stability. This complex recruits CAMK2D to induce the Ser324 phosphorylation of Tau proteins, changing the phosphorylation status of Tau bound to 14‐3‐3 and ultimately forming the hsa_circ_0001546/14‐3‐3/CAMK2D/Tau complex. The existence of this complex stimulates the production of Tau aggregation, which then induces the accumulation of lipid peroxides (LPOs) and causes LPO‐dependent ferroptosis. In vivo, treatment with ferrostatin‐1 and TRx0237 rescued the inhibitory effect of hsa_circ_0001546 on EOC cell spreading. Therefore, based on this results, ferroptosis caused by Tau aggregation occurs in EOC cells, which is not only in Alzheimer's disease‐ or Parkinson's disease‐related cells and this kind of ferroptosis driven by the hsa_circ_0001546/14‐3‐3/CAMK2D/Tau complex is LPO‐dependent rather than GPX4‐dependent is hypothesized.

Mitochondrial‐Derived Peptide MOTS‐c Suppresses Ovarian Cancer Progression by Attenuating USP7‐Mediated LARS1 Deubiquitination

AbstractMitochondrial‐nuclear communication plays a vital role in maintaining cellular homeostasis. MOTS‐c, a short peptide derived from the 12S rRNA of mitochondrial DNA, has been suggested as a retrograde mitochondrial signal. Although recent clinical studies have suggested a possible link between MOTS‐c and human cancer, the role of MOTS‐c in tumorigenesis has yet to be investigated. Here, MOTS‐c levels are found to be reduced in both serum and tumor tissues from ovarian cancer (OC) patients, which are associated with poor patients’ prognosis. Exogenous MOTS‐c inhibits the proliferation, migration and invasion of OC cells, and induces cell cycle arrest and apoptosis. Mechanistically, MOTS‐c interacts with LARS1 and promotes its ubiquitination and proteasomal degradation. In addition, USP7 was identified as a deubiquitinase of LARS1, and MOTS‐c can attenuates USP7‐mediated LARS1 deubiquitination by competing with USP7 for binding to LARS1. Besides, LARS1 was found to be increased and play an important oncogenic function in OC. More importantly, MOTS‐c displays a marked anti‐tumor effect on OC growth without systemic toxicity in vivo. In conclusion, this study reveals a crucial role of MOTS‐c in OC and provides a possibility for MOTS‐c as a therapeutic target for the treatment of this manlignacy.

Highly Efficient Synergistic Chemotherapy and Magnetic Resonance Imaging for Targeted Ovarian Cancer Therapy Using Hyaluronic Acid‐Coated Coordination Polymer Nanoparticles

AbstractThe diagnosis and treatment of ovarian cancer (OC) are still a grand challenge, more than 70% of patients are diagnosed at an advanced stage with a dismal prognosis. Magnetic resonance imaging (MRI) has shown superior results to other examinations in preoperative assessment, while cisplatin‐based chemotherapy is the first‐line treatment for OC. However, few previous studies have brought together the two rapidly expanding fields. Here a technique is presented using cisplatin prodrug (Pt‐COOH), Fe3+, and natural polyphenols (Gossypol) to construct the nanoparticles (HA@PFG NPs) that have a stable structure, controllable drug release behavior, and high drug loading capacity. The acidic pH values in tumor sites facilitate the release of Fe3+, Pt‐COOH, and Gossypol from HA@PFG NPs. Pt‐COOH with GSH consumption and cisplatin‐based chemotherapy plus Gossypol with pro‐apoptotic effects displays a synergistic effect for killing tumor cells. Furthermore, the release of Fe3+ at the tumor sites promotes ferroptosis and enables MRI imaging of OC. In the patient‐derived tumor xenograft (PDX) model, HA@PFG NPs alleviate the tumor activity. RNA sequencing analysis reveals that HA@PFG NPs ameliorate OC symptoms mainly through IL‐6 signal pathways. This work combines MRI imaging with cisplatin‐based chemotherapy, which holds great promise for OC diagnosis and synergistic therapy.

Dissecting the Distinct Tumor Microenvironments of HRD and HRP Ovarian Cancer: Implications for Targeted Therapies to Overcome PARPi Resistance in HRD Tumors and Refractoriness in HRP Tumors

AbstractHigh‐grade serous tubo‐ovarian cancer (HGSTOC) is an aggressive gynecological malignancy including homologous recombination deficient (HRD) and homologous recombination proficient (HRP) groups. Despite the therapeutic potential of poly (ADP‐ribose) polymerase inhibitors (PARPis) and anti‐PDCD1 antibodies, acquired resistance in HRD and suboptimal response in HRP patients necessitate more precise treatment. Herein, single‐cell RNA and single‐cell T‐cell receptor sequencing on 5 HRD and 3 HRP tumors are performed to decipher the heterogeneous tumor immune microenvironment (TIME), along with multiplex immunohistochemistry staining and animal experiments for validation. HRD tumors are enriched with immunogenic epithelial cells, FGFR1+PDGFRβ+ myCAFs, M1 macrophages, tumor reactive CD8+/CD4+ Tregs, whereas HRP tumors are enriched with HDAC1‐expressing epithelial cells, indolent CAFs, M2 macrophages, and bystander CD4+/CD8+ T cells. Significantly, customized therapies are proposed. For HRD patients, targeting FGFR1+PDGFRβ+ myCAFs via tyrosine kinase inhibitors, targeting Tregs via anti‐CCR8 antibodies/TNFRSF4 stimulation, and targeting CXCL13+ exhausted T cells by blocking PDCD1/CTLA‐4/LAG‐3/TIGIT are proposed. For HRP patients, targeting indolent CAFs, targeting M2 macrophages via CSF‐1/CSF‐1R inhibitors, targeting bystander T cells via tumor vaccines, and targeting epithelial cells via HDAC inhibitors. The study provides comprehensive insights into HRD and HRP TIME and tailored therapeutic approaches, addressing the challenges of PARPi‐resistant HRD and refractory HRP tumors.

KAT6A Condensates Impair PARP1 Trapping of PARP Inhibitors in Ovarian Cancer

AbstractMost clinical PARP inhibitors (PARPis) trap PARP1 in a chromatin‐bound state, leading to PARPi‐mediated cytotoxicity. PARPi resistance impedes the treatment of ovarian cancer in clinical practice. However, the mechanism by which cancer cells overcome PARP1 trapping to develop PARPi resistance remains unclear. Here, it is shown that high levels of KAT6A promote PARPi resistance in ovarian cancer, regardless of its catalytic activity. Mechanistically, the liquid‐liquid phase separation (LLPS) of KAT6A, facilitated by APEX1, inhibits the cytotoxic effects of PARP1 trapping during PARPi treatment. The stable KAT6A‐PARP1‐APEX1 complex reduces the amount of PARP1 trapped at the DNA break sites. In addition, inhibition of KAT6A LLPS, rather than its catalytic activity, impairs DNA damage repair and restores PARPi sensitivity in ovarian cancer both in vivo and in vitro. In conclusion, the findings demonstrate the role of KAT6A LLPS in fostering PARPi resistance and suggest that repressing KAT6A LLPS can be a potential therapeutic strategy for PARPi‐resistant ovarian cancer.

An Antibody‐CRISPR/Cas Conjugate Platform for Target‐Specific Delivery and Gene Editing in Cancer

AbstractThe CRISPR/Cas system has been introduced as an innovative tool for therapy, however achieving specific delivery to the target has been a major challenge. Here, an antibody‐CRISPR/Cas conjugate platform that enables specific delivery and target gene editing in HER2‐positive cancer is introduced. The CRISPR/Cas system by replacing specific residues of Cas9 with an unnatural amino acid is engineered, that can be complexed with a nanocarrier and bioorthogonally functionalized with a monoclonal antibody targeting HER2. The resultant antibody‐conjugated CRISPR/Cas nanocomplexes can be specifically delivered and induce gene editing in HER2‐positive cancer cells in vitro. It is demonstrated that the in vivo delivery of the antibody‐CRISPR/Cas nanocomplexes can effectively disrupt the plk1 gene in HER2‐positive ovarian cancer, resulting in substantial suppression of tumor growth. The current study presents a useful therapeutic platform for antibody‐mediated delivery of CRISPR/Cas for the treatment of various cancers and genetic diseases.

Hierarchical Self‐Assembly Molecular Building Blocks as Intelligent Nanoplatforms for Ovarian Cancer Theranostics

AbstractHierarchical self‐assembly from simple building blocks to complex polymers is a feasible approach to constructing multi‐functional smart materials. However, the polymerization process of polymers often involves challenges such as the design of building blocks and the drive of external energy. Here, a hierarchical self‐assembly with self‐driven and energy conversion capabilities based on p‐aminophenol and diethylenetriamine building blocks is reported. Through β‐galactosidase (β‐Gal) specific activation to the self‐assembly, the intelligent assemblies (oligomer and superpolymer) with excellent photothermal and fluorescent properties are dynamically formed in situ, and thus the sensitive multi‐mode detection of β‐Gal activity is realized. Based on the overexpression of β‐Gal in ovarian cancer cells, the self‐assembly superpolymer is specifically generated in SKOV‐3 cells to achieve fluorescence imaging. The photothermal therapeutic ability of the self‐assembly oligomer (synthesized in vitro) is evaluated by a subcutaneous ovarian cancer model, showing satisfactory anti‐tumor effects. This work expands the construction of intelligent assemblies through the self‐driven cascade assembly of small molecules and provides new methods for the diagnosis and treatment of ovarian cancer.

Nanoparticle Targeting in Chemo‐Resistant Ovarian Cancer Reveals Dual Axis of Therapeutic Vulnerability Involving Cholesterol Uptake and Cell Redox Balance

Abstract Platinum (Pt)‐based chemotherapy is the main treatment for ovarian cancer (OC); however, most patients develop Pt resistance (Pt‐R). This work shows that Pt‐R OC cells increase intracellular cholesterol through uptake via the HDL receptor, scavenger receptor type B‐1 (SR‐B1). SR‐B1 blockade using synthetic cholesterol‐poor HDL‐like nanoparticles (HDL NPs) diminished cholesterol uptake leading to cell death and inhibition of tumor growth. Reduced cholesterol accumulation in cancer cells induces lipid oxidative stress through the reduction of glutathione peroxidase 4 (GPx4) leading to ferroptosis. In turn, GPx4 depletion induces decreased cholesterol uptake through SR‐B1 and re‐sensitizes OC cells to Pt. Mechanistically, GPx4 knockdown causes lower expression of the histone acetyltransferase EP300, leading to reduced deposition of histone H3 lysine 27 acetylation (H3K27Ac) on the sterol regulatory element binding transcription factor 2 (SREBF2) promoter and suppressing expression of this key transcription factor involved in the regulation of cholesterol metabolism. SREBF2 downregulation leads to decreased SR‐B1 expression and diminished cholesterol uptake. Thus, chemoresistance and cancer cell survival under high ROS burden obligates high GPx4 and SR‐B1 expression through SREBF2. Targeting SR‐B1 to modulate cholesterol uptake inhibits this axis and causes ferroptosis in vitro and in vivo in Pt‐R OC.

Reshaping Intratumoral Mononuclear Phagocytes with Antibody‐Opsonized Immunometabolic Nanoparticles

AbstractMononuclear phagocytes (MPs) are vital components of host immune defenses against cancer. However, tumor‐infiltrating MPs often present tolerogenic and pro‐tumorigenic phenotypes via metabolic switching triggered by excessive lipid accumulation in solid tumors. Inspired by viral infection‐mediated MP modulation, here enveloped immunometabolic nanoparticles (immeNPs) are designed to co‐deliver a viral RNA analog and a fatty acid oxidation regulator for synergistic reshaping of intratumoral MPs. These immeNPs are camouflaged with cancer cell membranes for tumor homing and opsonized with anti‐CD163 antibodies for specific MP recognition and uptake. It is found that internalized immeNPs coordinate lipid metabolic reprogramming with innate immune stimulation, inducing M2‐to‐M1 macrophage repolarization and tolerogenic‐to‐immunogenic dendritic cell differentiation for cytotoxic T cell infiltration. The authors further demonstrate that the use of immeNPs confers susceptibility to anti‐PD‐1 therapy in immune checkpoint blockade‐resistant breast and ovarian tumors, and thereby provide a promising strategy to expand the potential of conventional immunotherapy.

N6‐Methyladenosine Regulates Cilia Elongation in Cancer Cells by Modulating HDAC6 Expression

AbstractPrimary cilia are microtubule‐based organelles that function as cellular antennae to address multiple metabolic and extracellular cues. The past decade has seen significant advances in understanding the pro‐tumorigenic role of N6‐methyladenosine (m6A) modification in tumorigenesis. Nevertheless, whether m6A modification modulates the cilia dynamics during cancer progression remains unclear. Here, the results show that m6A methyltransferase METTL3 regulates cilia length in cancer cells via HDAC6‐dependent deacetylation of axonemal α‐tubulin, thereby controlling cancer development. Mechanically, METTL3 positively regulates the translation of HDAC6 in an m6A‐dependent manner, while m6A methylation of A3678 in the coding sequence (CDS) of HDAC6 ameliorates its translation efficiency via facilitating the binding with YTHDF3. The upregulation of HDAC6 induced by METTL3 over‐expression is capable of inhibiting cilia elongation and acetylation of α‐tubulin, thereby shortening cilia length and accelerating the progression of cervical cancer both in vitro and in vivo. Collectively, depletion of METTL3‐mediated m6A modification leads to abnormally elongated cilia via suppressing HDAC6‐dependent deacetylation of axonemal α‐tubulin, ultimately attenuating cell growth and cervical cancer development.

Phosphorylation of IDH1 Facilitates Progestin Resistance in Endometrial Cancer

Abstract The progestin regimen is one of the main therapeutic strategies for women with endometrial cancer who undergo conservative management. Although many patients respond well to initial therapy, progestin‐refractory disease inevitably emerges, and the molecular basis underlying progestin resistance has not been comprehensively elucidated. Herein, they demonstrated progestin results in p38‐dependent IDH1 Thr 77 phosphorylation (pT77‐IDH1). pT77‐IDH1 translocates into the nucleus and is recruited to chromatin through its interaction with OCT6. IDH1‐produced α‐ketoglutarate (αKG) then facilitates the activity of OCT6 to promote focal adhesion related target gene transcription to confer progestin resistance. Pharmacological inhibition of p38 or focal adhesion signaling sensitizes endometrial cancer cells to progestin in vivo. The study reveals p38‐dependent pT77‐IDH1 as a key mediator of progestin resistance and a promising target for improving the efficacy of progestin therapy.

EIF1AX Nucleolar Condensates Enhance Susceptibilities for the Management of Endometrial Cancer

Abstract Endometrial cancer harboring TP53 aberrations presents a significant therapeutic challenge due to the lack of druggable targets. A promising strategy involves inducing senescence in cancer cells followed by targeted elimination using senolytic agents. The preliminary findings indicated that the aberrant subcellular localization of EIF1AX in endometrial cancer is significantly correlated with a poor prognosis. In this study, a compound library is employed to screen for therapeutic agents that induce the nuclear localization of EIF1AX in endometrial cancer cells, followed by a CRISPR library screen to identify senolytic compounds. The results demonstrated that the combination of 2,5‐MeC and dacinostat effectively inhibited tumor growth. Mechanistically, co‐immunoprecipitation mass spectrometry and cleavage under targets and tagmentation sequencing analyses demonstrated that 2,5‐MeC acts as a potent inducer of EIF1AX nucleolar translocation. This translocation promoted senescence by recruiting DDX21 to form nucleolar aggregates, which suppressed rDNA transcription. Additionally, RNA sequencing and antibody array analyses revealed that the synthetic lethality of 2,5‐MeC and dacinostat is mediated through the activation of the JNK/MAPK signaling pathway. Collectively, these findings highlight a novel therapeutic strategy for TP53‐aberrant endometrial cancer.

Lipid‐Driven OLR1/FOXM1/FGF19 Axis Orchestrates Crosstalk in an Epithelial‐Fibroblast Positive Feedback Promoting Progesterone Resistance in Endometrial Cancer

Abstract Progesterone resistance (ProR) remains a major obstacle in the conservative management of endometrial cancer (EC). Here, a metabolic‐stromal signaling loop centered on the OLR1/FOXM1/FGF19 axis is identified that drives progesterone resistance in EC. Single‐cell transcriptomic profiling first revealed a striking correlation between epithelial cells and fibroblasts in EC tissues with ProR. Tumor epithelial cells display profound alterations in lipid metabolism, whereas fibroblasts exhibited enhanced oxidative stress signatures. Clinical samples analyses indicated that oxidized low density lipoprotein (oxLDL), a product of LDL oxidation, is associated with adverse outcomes. The binding of oxLDL to its receptor OLR1 promoted the expression of FOXM1, a transcription factor that directly upregulates fibroblast growth factor 19 (FGF19). Immunofluorescence confirmed not only the spatial co‐localization of epithelial cells and fibroblasts but also the enrichment of OLR1 within epithelial compartments. Furthermore, treatment with the antioxidant resveratrol (RSV) and its nanoformulation (RSV‐NPs) markedly inhibited tumor growth in mice with lipid metabolic disorders, highlighting their potential to counteract progesterone resistance by disrupting this OLR1/FOXM1/FGF19 axis. This work highlights the therapeutic potential of targeting the tumor–stroma metabolic axis to increase progesterone sensitivity and improve outcomes in EC patients with fertility‐preserving demands.

SF3A3 Drives Tumorigenesis in Endometrial Cancer by Enhancing c‐FOS Expression and Represents a Potential Therapeutic Target

AbstractAberrant alternative splicing plays a crucial role in tumorigenesis. Here, splicing factor 3A subunit 3 (SF3A3) is   significantly upregulated in endometrial cancer (EC) tissues and associated with poor prognosis. Functionally, SF3A3 drives tumor progression by promoting cell proliferation, suppressing apoptosis, and enhancing cisplatin resistance in vitro and in vivo. Mechanistically, SF3A3 regulates alternative splicing of fos proto‐oncogene, AP‐1 transcription factor subunit (c‐FOS), an oncogene linked to chemoresistance, resulting in a ≈2‐fold increase in full‐length c‐FOS expression and activation of downstream anti‐apoptotic pathways. Notably, phenylethyl isothiocyanate (PEITC) as a direct inhibitor of SF3A3 through database screening and biophysical validation via surface plasmon resonance and mass spectrometry is identified. PEITC reduces c‐FOS expression and induces apoptosis in EC cells. Moreover, encapsulating PEITC in a hydrogel delivery system significantly enhances its therapeutic efficacy by enabling controlled release, reducing dosing frequency, and improving clinical applicability. The therapeutic potential of SF3A3 inhibition is further validated using patient‐derived tumor‐like cell clusters (PTCs), where PEITC and the c‐FOS inhibitor T‐5224 exhibit synergistic effects in suppressing EC. Collectively, our findings establish SF3A3 as a novel oncogenic regulator in EC and highlight PEITC, particularly in its hydrogel formulation, as a promising therapeutic strategy for improving clinical outcomes in EC patients.

Fatty Acid Oxidation Supports Lymph Node Metastasis of Cervical Cancer via Acetyl‐CoA‐Mediated Stemness

AbstractAccumulating evidence indicates that metabolic reprogramming of cancer cells supports the energy and metabolic demands during tumor metastasis. However, the metabolic alterations underlying lymph node metastasis (LNM) of cervical cancer (CCa) have not been well recognized. In the present study, it is found that lymphatic metastatic CCa cells have reduced dependency on glucose and glycolysis but increased fatty acid oxidation (FAO). Inhibition of carnitine palmitoyl transferase 1A (CPT1A) significantly compromises palmitate‐induced cell stemness. Mechanistically, FAO‐derived acetyl‐CoA enhances H3K27 acetylation (H3K27Ac) modification level in the promoter of stemness genes, increasing stemness and nodal metastasis in the lipid‐rich nodal environment. Genetic and pharmacological loss of CPT1A function markedly suppresses the metastatic colonization of CCa cells in tumor‐draining lymph nodes. Together, these findings propose an effective method of cancer therapy by targeting FAO in patients with CCa and lymph node metastasis.

PRMT3‐Mediated Arginine Methylation of METTL14 Promotes Malignant Progression and Treatment Resistance in Endometrial Carcinoma

AbstractProtein arginine methyltransferase (PRMT) plays essential roles in tumor initiation and progression, but its underlying mechanisms in the treatment sensitivity of endometrial cancer (EC) remain unclear and warrant further investigation. Here, a comprehensive analysis of the Cancer Genome Atlas database and Clinical Proteomic Tumor Analysis Consortium database identifies that PRMT3 plays an important role in EC. Specifically, further experiments show that PRMT3 inhibition enhances the susceptibility of EC cells to ferroptosis. Mechanistically, PRMT3 interacts with Methyltransferase 14 (METTL14) and is involved in its arginine methylation. In addition, PRMT3 inhibition‐mediated METTL14 overexpression promotes methylation modification via an m6A‐YTHDF2‐dependent mechanism, reducing Glutathione peroxidase 4 (GPX4) mRNA stability, increasing lipid peroxidation levels, and accelerating ferroptosis. Notably, combined PRMT3 blockade and anti‐PD‐1 therapy display more potent antitumor effects by accelerating ferroptosis in cell‐derived xenograft models. The specific PRMT3 inhibitor SGC707 exerts the same immunotherapeutic sensitizing effect in a patient‐derived xenograft model. Notably, blocking PRMT3 improves tumor suppression in response to cisplatin and radiation therapy. Altogether, this work demonstrates that PRMT3 depletion is a promising target for EC.

MGAT4A/Galectin9‐Driven N‐Glycosylation Aberration as a Promoting Mechanism for Poor Prognosis of Endometrial Cancer with TP53 Mutation

AbstractEmerging evidence recognizes aberrant glycosylation as the malignant characteristics of cancer cells, but little is known about glycogenes’ roles in endometrial carcinoma (EC), especially the most aggressive subtype carrying TP53 mutations. Using unsupervised hierarchical clustering, an 11‐glycogene cluster is identified to distinguish an EC subtype associated with frequent TP53 mutation and worse prognosis. Among them, MGAT4A (alpha‐1,3‐mannosyl‐glycoprotein 4‐β‐N‐acetylglucosaminyltransferase A) emerges as the most consistently overexpressed glycogene, contributing to EC aggressiveness. In the presence of galectin‐9, MGAT4A increases EC cell proliferation and invasion via promoting glucose metabolism. N‐glycoproteomics further revealed GLUT1, a glucose transporter, as a glycoprotein modified by MGAT4A. Binding of galectin‐9 to the MGAT4A‐branched N‐glycan on GLUT1 enhances its cell membrane distribution, leading to glucose uptake increase. In addition, oncogenic mutations of TP53 gene in EC cells upregulate MGAT4A expression by disrupting the regulatory oversight exerted by wild‐type p53 on tumor‐suppressive miRNAs, including miR‐34a and miR‐449a/b. The findings highlight a new molecular mechanism involving MGAT4A‐regulated N‐glycosylation on the key regulator of glucose metabolism in p53 mutants‐driven EC aggressiveness, which may provide a strategic avenue to combat advanced EC.

Halofuginone Disrupted Collagen Deposition via mTOR‐eIF2α‐ATF4 Axis to Enhance Chemosensitivity in Ovarian Cancer

AbstractThe interplay between cancer‐associated fibroblasts (CAFs) and extracellular matrix (ECM) mediates progress, metastasis, and therapy resistance. However, strategy of targeting ECM remodeling to enhance chemosensitivity in ovarian cancer remains elusive. Here, a 22‐gene matrisome signature predicts chemotherapy response and survival in ovarian cancer. The dense, collagen‐rich ECM secreted by CAFs harbors more M2 tumor‐associated macrophages (TAMs) than the looser ECM based on single cell RNA‐seq (scRNA‐seq) of ovarian cancer, suggesting the promising approach of targeting collagen to remodel ECM. An integrated analysis identifies collagen type I alpha 1 chain (COL1A1) as a major component of the ECM that contributes to chemoresistance and poor prognosis, highlighting its potential as a therapeutic target. Halofuginone (HF), a clinically active derivative of febrifugine, is identified as a COL1A1‐targeting natural compound by screening the Encyclopedia of Traditional Chinese Medicine (ETCM). Mechanistically, HF inhibits COL1A1 production via the mTOR‐eIF2α‐ATF4 axis in CAFs. Notably, HF disrupts collagen deposition and promotes CD8+ T cell infiltration, partially via M2‐M1 macrophage polarization to enhance chemosensitivity. Overall, the findings suggest that HF combined with chemotherapy is a promising and effective treatment for ovarian cancer.

A New Type of Endometrial Cancer Models in Mice Revealing the Functional Roles of Genetic Drivers and Exploring their Susceptibilities

AbstractEndometrial cancer (EC) is the most common female reproductive tract cancer and its incidence has been continuously increasing in recent years. The underlying mechanisms of EC tumorigenesis remain unclear, and efficient target therapies are lacking, for both of which feasible endometrial cancer animal models are essential but currently limited. Here, an organoid and genome editing‐based strategy to generate primary, orthotopic, and driver‐defined ECs in mice is reported. These models faithfully recapitulate the molecular and pathohistological characteristics of human diseases. The authors names these models and similar models for other cancers as organoid‐initiated precision cancer models (OPCMs). Importantly, this approach can conveniently introduce any driver mutation or a combination of driver mutations. Using these models,it is shown that the mutations in Pik3ca and Pik3r1 cooperate with Pten loss to promote endometrial adenocarcinoma in mice. In contrast, the Kras G12D mutati led to endometrial squamous cell carcinoma. Then, tumor organoids are derived from these mouse EC models and performed high‐throughput drug screening and validation. The results reveal distinct vulnerabilities of ECs with different mutations. Taken together, this study develops a multiplexing approach to model EC in mice and demonstrates its value for understanding the pathology of and exploring the potential treatments for this malignancy.

Targeting Cancer Metabolism Plasticity with JX06 Nanoparticles via Inhibiting PDK1 Combined with Metformin for Endometrial Cancer Patients with Diabetes

AbstractDiabetes is closely related to the occurrence of endometrial cancer (EC) and its poor prognosis. However, there is no effective clinical treatment for EC patients with diabetes (patientEC+/dia+). To explore new therapeutic targets, Ishikawa is cultured with high glucose (IshikawaHG) mimicking hyperglycemia in patientEC+/dia+. Subsequently, it is discovered that IshikawaHG exhibits glucose metabolic reprogramming characterized by increased glycolysis and decreased oxidative phosphorylation. Further, pyruvate dehydrogenase kinase 1 (PDK1) is identified to promote glycolysis of IshikawaHG by proteomics. Most importantly, JX06, a novel PDK1 inhibitor combined metformin (Met) significantly inhibits IshikawaHG proliferation though IshikawaHG is resistant to Met. Furthermore, a reduction‐sensitive biodegradable polymer is adopted to encapsulate JX06 to form nanoparticles (JX06‐NPs) for drug delivery. It is found that in vitro JX06‐NPs have better inhibitory effect on the growth of IshikawaHG as well as patient‐derived EC cells (PDC) than JX06. Additionally, it is found that JX06‐NPs can accumulate to the tumor of EC‐bearing mouse with diabetes (miceEC+/dia+) after intravenous injection, and JX06‐NPs combined Met can significantly inhibit tumor growth of miceEC+/dia+. Taken together, the study demonstrates that the combination of JX06‐NPs and Met can target the cancer metabolism plasticity, which significantly inhibits the growth of EC, thereby provides a new adjuvant therapy for patientsEC+/dia+.

Immune Predictors of Radiotherapy Outcomes in Cervical Cancer

ABSTRACT The immune microenvironment influences the sensitivity of patients to radiotherapy (RT), yet determinants of therapeutic resistance remain elusive. This study integrates single‐cell transcriptomics and machine learning to delineate immune predictors of RT outcomes. Comprehensive analysis reveals reduced epithelial cell numbers, accompanied by enhanced apoptosis, complement activation, and inflammatory responses. RT triggers macrophage accumulation, particularly an RT‐responsive M1‐like HSPA1B+ subset with elevated antigen‐presenting capacity. While T and NK cell cytotoxicity increases, their exhaustion markers (e.g., PDCD1, TIGIT) are exacerbated. CellChat analysis identifies robust epithelial‐myeloid crosstalk mediated by the C3/C3AR1 axis. In murine models, C3AR1 antagonism diminishes RT efficacy, impairing macrophage infiltration and M1 polarization. Leveraging 25 single‐cell‐derived immune features, an 8‐feature multilayer perceptron model: Cervical Cancer Radiotherapy Immune‐Response Model (CCRTIM) is developed. CCRTIM robustly predicts prognosis (AUC = 0.76) and exhibits risk stratification. These findings unveil dynamic immune remodeling post‐RT and establish actionable biomarkers for precision radiotherapy strategies.

Reaction Pathway Differentiation Enabled Fingerprinting Signal for Single Nucleotide Variant Detection

AbstractAccurate identification of single‐nucleotide variants (SNVs) is paramount for disease diagnosis. Despite the facile design of DNA hybridization probes, their limited specificity poses challenges in clinical applications. Here, a differential reaction pathway probe (DRPP) based on a dynamic DNA reaction network is presented. DRPP leverages differences in reaction intermediate concentrations between SNV and WT groups, directing them into distinct reaction pathways. This generates a strong pulse‐like signal for SNV and a weak unidirectional increase signal for wild‐type (WT). Through the application of machine learning to fluorescence kinetic data analysis, the classification of SNV and WT signals is automated with an accuracy of 99.6%, significantly exceeding the 80.7% accuracy of conventional methods. Additionally, sensitivity for variant allele frequency (VAF) is enhanced down to 0.1%, representing a ten‐fold improvement over conventional approaches. DRPP accurately identified D614G and N501Y SNVs in the S gene of SARS‐CoV‐2 variants in patient swab samples with accuracy over 99% (n = 82). It determined the VAF of ovarian cancer‐related mutations KRAS‐G12R, NRAS‐G12C, and BRAF‐V600E in both tissue and blood samples (n = 77), discriminating cancer patients and healthy individuals with significant difference (p < 0.001). The potential integration of DRPP into clinical diagnostics, along with rapid amplification techniques, holds promise for early disease diagnostics and personalized diagnostics.

Erucic Acid, Derived by Lactobacillus Crispatus , Induces Ferroptosis in Cervical Cancer Organoids Through the PPAR‐δ Signaling Pathway

Abstract The microbiome present throughout the human body serves a variety of functions. In this study, 16S rRNA sequencing is employed to uncover differences in the abundance of Lactobacillus within the vaginal microbiota between individuals with cervical cancer and those with healthy cervixes. The research further identifies that the metabolite of Lactobacillus crispatus can induce ferroptosis in cervical cancer cells. This conclusion is reached through targeted bacterial culture, patient‐derived organoids (PDO) and single‐cell RNA sequencing. Erucic acid, identified as a primary metabolite via untargeted metabolomics, acts as a ligand for PPARδ receptor. It has the capacity to activate PPARδ pathway and subsequently trigger downstream fatty acid oxidation (FAO). Excessive enhancement of FAO can generate large amounts of H 2 O 2 and O 2 ‐, known as ROS. Utilizing PDO, cell lines and cervical cancer xenograft (CDX) models, the study demonstrate both in vitro and in vivo that the metabolite of L. crispatus , erucic acid, can modulate the proliferation, migration and invasion of cervical cancer by activating the PPAR‐δ pathway. This activation leads to fatty acid oxidation, release ROS, and ultimately induces ferroptosis. Therefore, L. crispatus and erucic acid show potential as novel adjuvant therapeutic agents in the treatment of cervical cancer.

A Field‐Deployable RotEx‐LAMP‐LFA Platform for Molecular Triage of HPV‐Driven Oncogenesis

Abstract Persisting as a major global health disparity, HPV16/18‐associated cervical carcinogenesis continues to disproportionately affect populations in resource‐limited regions with compromised screening infrastructure. While molecular detection of E6/E7 oncogenic transcripts surpasses conventional cytology in clinical specificity, current nucleic acid amplification platforms remain impeded by technical complexity, prolonged turnaround times (>4 h), and substantial per‐test costs (>$50) that hinder scale‐up in low‐income settings. Here, RotEx‐LAMP‐LFA is presented: an innovative point‐of‐care system integrating microfluidic nucleic acid extraction, rapid isothermal amplification (30 min at 63 °C), and combined lateral flow detection in a single disposable cartridge. Validated with 69 clinical specimens (19 histology‐confirmed SCC, 14 HPV+ precancerous lesions, 15 HPV+ infections, and 21 healthy controls), this sample‐to‐answer platform achieved 94.74% sensitivity (18/19; 95% CI: 84.72–100%) for invasive carcinoma detection with 100% (36/36; 95% CI: 90.44‐99.93%) specificity for non‐cancer. In addition, a subset of samples is collected through paired self‐sampling to validate consistency with clinician‐collected specimens, indicating that this innovative platform shows promise for community‐level molecular stratification of persistent and progressive HPV infections. It paves the way for accessible, patient‐centered cervical cancer risk reporting, with the potential to reduce psychological burden on patients.

Targeting Glutamine Metabolism Transporter SLC25A22 Enhances CD8+ T‐Cell Function and Anti‐PD‐1 Therapy Efficacy in Cervical Squamous Cell Carcinoma: Integrated Metabolomics, Transcriptomics and T‐Cell‐Incorporated Tumor Organoid Studies

Abstract Cervical squamous cell carcinoma(CSCC) represents formidable challenge in clinical oncology, exacerbated by poor prognosis and resistance to current treatments, including anti‐PD‐1 therapy, highlighting the urgent need for alternative therapeuties. Metabolic characteristics have emerged as potential drivers of treatment resistance and immune evasion. Herein, 1) based on metabolomic and transcriptomic analyses of 44 CSCC and 18 normal tissues, glutamine‐enriched and immunosuppressive microenvironment is identified in CSCC. 2) Integrative metabolomic and transcriptomic analyses revealed the glutamine metabolism transporter SLC25A22 as a key mediator in high glutamine metabolism, immune checkpoint activation and CD8+T‐cell cytotoxicity. 3) Immunohistochemistry(IHC), multiplex IHC, and flow cytometry validation with clinical CSCC samples revealed not only increased SLC25A22, PD‐1 expression and reduced CD8+T‐cell cytotoxicity in CSCC but also increased SLC25A22 expression in high PD‐L1 expressed CSCC patients, suggesting the potential of targeting SLC25A22 for enhancing CD8+T‐cell cytotoxicity and improving anti‐PD‐1 efficacy, especially in high PD‐L1 expressed patients. 4) Novelly, 3D‐CSCC organoids are constructed, replicating parental tumor features, and 3D‐T‐cell‐incorporated CSCC organoid models, replicating the interaction between tumor cells and CD8+T cells, for in vitro experiments. 5) Importantly, it is validated through in vitro 3D T‐cell‐incorporated CSCC organoid models and in vivo animal experiments that targeting the glutamine metabolism transporter SLC25A22, showed promise in enhancing CD8+T‐cell cytotoxicity and sensitizing anti‐PD‐1 therapy. These findings provided insights for future clinical trials exploring metabolic modulation to improve immunotherapy responses in CSCC patients.

BAG2 Inhibits Cervical Cancer Progression by Modulating Type I Interferon Signaling through Stabilizing STING

Abstract Cervical cancer possesses high morbidity and mortality rates, and a comprehensive understanding of its molecular underpinnings is essential for advancing clinical management strategies. The innate immune sensor STING, which activates type I interferon signaling, plays a pivotal role in enhancing anti‐tumor activity. Despite increased attention to STING's involvement in cervical cancer, the regulatory mechanisms governing its protein homeostasis remain poorly understood. In this study, it is found that the BAG2‐STUB1 complex regulates ubiquitin proteasomal degradation of STING, which affects the development of cervical cancer. Mechanistically, BAG2 inhibits ubiquitination of STING and stabilizes it by interacting with STING. Specifically, BAG2 inhibits STUB1 from attaching the K48‐linked ubiquitin chains at K338 and K370 of STING by forming a complex with STUB1. Functionally, enhanced BAG2 expression suppresses cervical cancer progression by activating the type I interferon pathway in a STING‐dependent manner. Notably, clinical cervical cancer samples revealed a positive correlation between BAG2 and STING levels, with low BAG2 expression is strongly linked to advanced disease and poor prognosis in cervical cancer. Collectively, these findings elucidate the molecular mechanism by which the BAG2‐STUB1 complex regulates STING homeostasis, underscoring BAG2's potential as a diagnostic biomarker and therapeutic target in cervical cancer.

Deep Learning‐Enhanced Hand‐Driven Microfluidic Chip for Multiplexed Nucleic Acid Detection Based on RPA/CRISPR

AbstractThe early detection of high‐risk human papillomavirus (HR‐HPV) is crucial for the assessment and improvement of prognosis in cervical cancer. However, existing PCR‐based screening methods suffer from inadequate accessibility, which dampens the enthusiasm for screening among grassroots populations, especially in resource‐limited areas, and contributes to the persistently high mortality rate of cervical cancer. Here, a portable system is proposed for multiplexed nucleic acid detection, termed R‐CHIP, that integrates Recombinase polymerase amplification (RPA), CRISPR detection, Hand‐driven microfluidics, and an artificial Intelligence Platform. The system can go from sample pre‐processing to results readout in less than an hour with simple manual operation. Optimized for sensitivity of 10−17 M for HPV‐16 and 10−18 M for HPV‐18, R‐CHIP has an accuracy of over 95% in 300 tests on clinical samples. In addition, a smartphone microimaging system combined with the ResNet‐18 deep learning model is used to improve the readout efficiency and convenience of the detection system, with initial prediction accuracies of 96.0% and 98.0% for HPV‐16 and HPV‐18, respectively. R‐CHIP, as a user‐friendly and intelligent detection platform, has great potential for community‐level HR‐HPV screening in resource‐constrained settings, and contributes to the prevention and early diagnosis of other diseases.

Autologous Peripheral Vγ9Vδ2 T Cell Synergizes with αβ T Cell Through Antigen Presentation and BTN3A1 Blockade in Immunotherapy of Cervical Cancer

AbstractNew treatment strategies are urgently needed for patients with advanced cervical cancer (CC). Here, a synergistic anti‐CC effect of a novel combinatorial immunotherapy with adoptively transferred autologous Vγ9Vδ2 T cells and αβ T cells is shown. The pivotal role of both circulating and tumor‐infiltrating Vγ9Vδ2 T cells in anti‐CC immunity is uncovered. Importantly, autologous Vγ9Vδ2 T cells show a synergistic anti‐CC effect with αβ T cells not only through killing tumor directly, but also by promoting the activation and tumoricidal activity of syngeneic αβ T cells through antigen presentation, which can be further boosted by conventional chemotherapy. Moreover, Vγ9Vδ2 T cells can restore the tumoricidal function of αβ T cell through competitively binding to BTN3A1, a TCR‐Vγ9Vδ2 ligand on CC cells upregulated by IFN‐γ derived from activated αβ T cell. These findings uncover a critical synergistic effect of autologous Vγ9Vδ2 T cells and αβ T cells in immunotherapy of CC and reveal the underlying mechanisms.

Programmable Multiplexed Nucleic Acid Detection by Harnessing Specificity Defect of CRISPR‐Cas12a

AbstractCRISPR‐Cas12a works like a sophisticated algorithm in nucleic acid detection, yet its challenge lies in sometimes failing to distinguish targets with mismatches due to its specificity limitations. Here, the mismatch profiles, including the quantity, location, and type of mismatches in the CRISPR‐Cas12a reaction, are investigated and its various tolerances to mismatches are discovered. By harnessing the specificity defect of the CRISPR‐Cas12a enzyme, a dual‐mode detection strategy is designed, which includes approximate matching and precise querying of target sequences and develop a programmable multiplexed nucleic acid assay. With the assay, 14 high‐risk human papillomavirus (HPV) subtypes are simultaneously detected, collectively responsible for 99% of cervical cancer cases, with attomolar sensitivity. Specifically, the assay not only distinguishes HPV16 and HPV18, the two most common subtypes but also detects 12 other high‐risk pooled HPV subtypes. To enable low‐cost point‐of‐care testing, the assay is incorporated into a paper‐based microfluidic chip. Furthermore, the clinical performance of the paper‐based microfluidic chip is validated by testing 75 clinical swab samples, achieving performance comparable to that of PCR. This programmable multiplexed nucleic acid assay has the potential to be widely applied for sensitive, specific, and simultaneous detection of different pathogens.

HPV E6/E7‐Induced Acetylation of a Peptide Encoded by a Long Non‐Coding RNA Inhibits Ferroptosis to Promote the Malignancy of Cervical Cancer

AbstractAlthough a fraction of functional peptides concealed within long non‐coding RNAs (lncRNAs) is identified, it remains unclear whether lncRNA‐encoded peptides are involved in the malignancy of cervical cancer (CC). Here, a 92‐amino acid peptide is discovered, which is named TUBORF, encoded by lncRNA TUBA3FP and highly expressed in CC tissues. TUBORF inhibits ferroptosis to promote the malignant proliferation of CC cells. Mechanistically, human papillomavirus (HPV) oncogenes E6 and E7 upregulate TUBORF through CREB‐binding protein (CBP)/E1A‐binding protein p300 (p300)‐mediated histone H3 lysine 27 acetylation (H3K27ac) of lncTUBA3FP enhancer. Furthermore, E6 and E7 elevate and recruit acetyltransferase establishment of sister chromatid cohesion N‐acetyltransferase 1 (ESCO1) to bind to and acetylate TUBORF, which facilitates the degradation of immunity‐related GTPase Q (IRGQ) via a ubiquitin‐proteasome pathway, resulting in the inhibition of ferroptosis and promotion of the malignant proliferation of CC cells. Importantly, silencing ESCO1 or TURORF amplifies anticancer effects by paclitaxel both in CC cells and in vivo. These novel findings reveal oncopeptide TUBORF and its acetyltransferase ESCO1 as important regulators of ferroptosis and tumorigenesis during cervical cancer pathogenesis and establish the scientific basis for targeting these molecules for treating CC.

PIWIL2/PDK1 Axis Promotes the Progression of Cervical Epithelial Lesions via Metabolic Reprogramming to Maintain Tumor‐Initiating Cell Stemness

AbstractWhen PIWIL2 expression is restored via heterogeneous integration of human papillomavirus, cellular reprogramming is initiated to form tumor‐initiating cells (TICs), which triggers cervical squamous intraepithelial lesions (SIL). TIC stemness is critical for the prognosis of SIL. However, the mechanisms underlying TIC stemness maintenance and tumorigenicity remain unclear. Here, it is revealed that aberrant pyruvate dehydrogenase kinase 1 (PDK1) expression is closely related to aerobic glycolysis in SIL and poor survival in patients with cervical cancer. Mechanistically, that PIWIL2, which induced by stable transfection of either PIWIL2 or HPV16 oncogene E6 in human primary cervical basal epithelial cells and keratinocyte cell line HaCaT, upregulates PDK1 expression via the LIN28/let‐7 axis, hence reprogramming metabolism to activate glycolysis and synchronize with TIC formation. It is further demonstrate that PDK1 is critical for TIC stemness maintenance and tumorigenicity via the PI3K/AKT/mTOR pathway both in vitro and in vivo, revealing a previously unclear mechanism for SIL progression, regression or relapse. Therefore, this findings suggest a potential rationale for prognostic predictions and selecting targeted therapy for cervical lesions.

An Injection Molded SlipChip with Self‐Sampling for Integrated Point‐of‐Care Testing of Human Papilloma Virus

AbstractHigh‐risk human papillomavirus (HPV) screening is crucial for cervical cancer prevention. However, laboratory‐based nucleic acid amplification tests (NAATs) require costly equipment, designated lab space, and skilled personnel. Additionally, cervical swabs collected by healthcare professionals can be inconvenient, uncomfortable, and reduce privacy, limiting broader application and patient compliance. A SlipChip‐based Integrated Point‐of‐Care (SIPOC) system featuring an injection‐molded SlipChip is presented with preloaded reagents for nucleic acid extraction and a portable four‐channel real‐time quantitative PCR instrument for detection. This system incorporates a self‐sampling method that allows participants to collect their own vaginal swabs, with the β‐Globin gene as a control. After testing 130 participants for HPV‐16 and HPV‐18, 97.7% of the self‐collected samples are valid. Among valid samples, 25 tested positive for HPV‐16 and 9 for HPV‐18. Compared to Roche's standard HPV PCR test, the SIPOC system shows 100% positive predictive value (PPV) for both HPV‐16 and HPV‐18 and negative predictive values (NPVs) of 99.0% and 99.1%, respectively. This system is promising for HPV screening in resource‐limited settings and adaptable for other point‐of‐care NAAT applications, including home testing.

Plasma Cell‐Free DNA Concentration and Fragmentomes Predict Neoadjuvant Chemotherapy Response in Cervical Cancer Patients

AbstractCervical cancer remains one of the most lethal gynecological malignancies. However, biomarkers for more precise patient care are an unmet need. Herein, the concentration of 285 plasma cell‐free DNA (cfDNA) samples are analyzed from 84 cervical patients and the clinical significance of cfDNA fragmentomic characteristics across the neoadjuvant chemotherapy (NACT) treatment. Patients with poor NACT response exhibit a significantly greater escalation in cfDNA levels following the initial cycle of treatment, in comparison to patients with a favorable response. Distinctive end motif profiles and promoter coverages of cfDNA in initial plasma are observed between patients with differing NACT responses. Notably, the DNASE1L3 analysis further demonstrates the intrinsic association between cfDNA characteristics and chemotherapy resistance. The cfDNA and motif ratios show a good discriminative capacity for predicting non‐responders from responders (area under the curve (AUC) > 0.8). In addition, transcriptional start sites (TSS) coverages around promoters discern the alteration of biological processes associated with chemotherapy resistance and reflect the potential value in predicting chemotherapy response. These findings in predictive biomarkers may optimize treatment selection, minimize unnecessary treatment, and assist in establishing personalized treatment strategies for cervical cancer patients.

LNMAC Promotes Cervical Squamous Cell Carcinoma Lymphatic Metastasis via Epigenetic Regulation of FGF2‐Induced Lymphangiogenesis

AbstractThe lymph node is the most common site of distant metastasis of cervical squamous cell carcinoma (CSCC), which elicits dismal prognosis and limited efficiency for treatment. Elucidation of the mechanisms underlying CSCC lymphatic metastasis would provide potential therapeutic strategies for nodal metastatic of CSCC. Here, based on in vivo lymphatic metastasis screening model, a circular RNA is identified that is termed as lymph node metastasis associated circRNA (LNMAC), is markedly upregulated in lymphatic metastatic CSCC and correlated with lymph node metastasis. Overexpression of LNMAC dramatically augments the metastatic capability of CSCC cells to the lymph node via inducing lymphangiogenesis. Mechanistically, LNMAC epigenetically upregulates fibroblast growth factor 2 (FGF2) expression by directly associating with histone acacetylase 1 (HDAC1), preventing Importin α6/8‐mediated nuclear translocation of HDAC1 and eliciting histone H3K27ac‐induced FGF2 transcriptional activation. Treatment with 3F12E7, an anti‐FGF2 monoclonal antibody, effectively inhibits LNMAC‐induced CSCC lymphatic metastasis. Taken together, these findings indicate that LNMAC plays a crucial role in FGF2‐mediated lymphangiogenesis and lymphatic metastasis, highlighting that LNMAC might be a therapeutic target for lymph node metastasis in CSCC patients.

In Vivo Intra‐Uterine Delivery of TAT‐Fused Cre Recombinase and CRISPR/Cas9 Editing System in Mice Unveil Histopathology of Pten/p53‐Deficient Endometrial Cancers

AbstractPhosphatase and TENsin homolog (Pten) and p53 are two of the most frequently mutated tumor suppressor genes in endometrial cancer. However, the functional consequences and histopathological manifestation of concomitant p53 and Pten loss of function alterations in the development of endometrial cancer is still controversial. Here, it is demonstrated that simultaneous Pten and p53 deletion is sufficient to cause epithelial to mesenchymal transition phenotype in endometrial organoids. By a novel intravaginal delivery method using HIV1 trans‐activator of transcription cell penetrating peptide fused with a Cre recombinase protein (TAT‐Cre), local ablation of both p53 and Pten is achieved specifically in the uterus. These mice developed high‐grade endometrial carcinomas and a high percentage of uterine carcinosarcomas resembling those found in humans. To further demonstrate that carcinosarcomas arise from epithelium, double Pten/p53 deficient epithelial cells are mixed with wild type stromal and myometrial cells and subcutaneously transplanted to Scid mice. All xenotransplants resulted in the development of uterine carcinosarcomas displaying high nuclear pleomorphism and metastatic potential. Accordingly, in vivo CRISPR/Cas9 disruption of Pten and p53 also triggered the development of metastatic carcinosarcomas. The results unfadingly demonstrate that simultaneous deletion of p53 and Pten in endometrial epithelial cells is enough to trigger epithelial to mesenchymal transition that is consistently translated to the formation of uterine carcinosarcomas in vivo.

Inhibiting the IRE1α Axis of the Unfolded Protein Response Enhances the Antitumor Effect of AZD1775 in TP53 Mutant Ovarian Cancer

AbstractTargeting the G2/M checkpoint mediator WEE1 has been explored as a novel treatment strategy in ovarian cancer, but mechanisms underlying its efficacy and resistance remains to be understood. Here, it is demonstrated that the WEE1 inhibitor AZD1775 induces endoplasmic reticulum stress and activates the protein kinase RNA‐like ER kinase (PERK) and inositol‐required enzyme 1α (IRE1α) branches of the unfolded protein response (UPR) in TP53 mutant (mtTP53) ovarian cancer models. This is facilitated through NF‐κB mediated senescence‐associated secretory phenotype. Upon AZD1775 treatment, activated PERK promotes apoptotic signaling via C/EBP‐homologous protein (CHOP), while IRE1α‐induced splicing of XBP1 (XBP1s) maintains cell survival by repressing apoptosis. This leads to an encouraging synergistic antitumor effect of combining AZD1775 and an IRE1α inhibitor MKC8866 in multiple cell lines and preclinical models of ovarian cancers. Taken together, the data reveal an important dual role of the UPR signaling network in mtTP53 ovarian cancer models in response to AZD1775 and suggest that inhibition of the IRE1α‐XBP1s pathway may enhance the efficacy of AZD1775 in the clinics.

Targeting GRP75 with a Chlorpromazine Derivative Inhibits Endometrial Cancer Progression Through GRP75–IP3R‐Ca 2+ ‐AMPK Axis

Abstract Tumors often overexpress glucose‐regulated proteins, and agents that interfere with the production or activity of these proteins may represent novel cancer treatments. The chlorpromazine derivative JX57 exhibits promising effects against endometrial cancer with minimal extrapyramidal side effects; however, its mechanisms of action are currently unknown. Here, glucose‐regulated protein 75 kD (GRP75) is identified as a direct target of JX57 using activity‐based protein profiling and loss‐of‐function experiments. The findings show that GRP75 is necessary for the biological activity of JX57, as JX57 exhibits moderate anticancer properties in GRP75‐deficient cancer cells, both in vitro and in vivo. High GRP75 expression is correlated with poor differentiation and poor survival in patients with endometrial cancer, whereas the knockdown of GRP75 can significantly suppress tumor growth. Mechanistically, the direct binding of JX57 to GRP75 impairs the structure of the mitochondria‐associated endoplasmic reticulum membrane and disrupts the endoplasmic reticulum–mitochondrial calcium homeostasis, resulting in a mitochondrial energy crisis and AMP‐activated protein kinase activation. Taken together, these findings highlight GRP75 as a potential prognostic biomarker and direct therapeutic target in endometrial cancer and suggest that the chlorpromazine derivative JX57 can potentially be a new therapeutic option for endometrial cancer.

Mapping CSC‐Mediated Ovarian Cancer Chemoresistance via CXCR4‐PET to Guide Precision Cisplatin Re‐Sensitization Therapy

ABSTRACT Therapy targeting cancer stem cells (CSCs) has been proposed as a promising strategy to reduce chemoresistance and relapse risks in ovarian cancer (OC) patients. However, the lack of targetable markers impedes research progress. Here, we demonstrate that CXC motif chemokine receptor 4 (CXCR4) may be a targetable functional marker of ovarian CSCs and propose a new translational model incorporating targeted imaging and CXCR4 blockade in CXCR4 + tumors. Expression profile analysis of chemoresistant CSC‐like ovarian cancer cells highlighted that CXCR4 functions as a potential stemness marker. CXCR4 + ovarian cancer cells exhibited high self‐renewal capacity in vitro and in vivo, and an association with chemoresistance. CXCR4 inhibitor AMD3100 significantly impaired the self‐renewal ability of CSC‐like ovarian cancer cells and enhanced their sensitivity to cisplatin. CXCR4‐targeted [ 68 Ga]Ga‐Pentixafor was highly specific in delineating CXCR4‐high cell line‐derived xenografts and patient‐derived xenografts (PDXs) via positron emission tomography (PET) imaging, with precise tumor‐targeting and persistent retention. A combination of AMD3100 and CDDP exerted an excellent antitumor effect in CXCR4‐high PDXs, but not in CXCR4‐low PDXs. These results suggest that CXCR4 may represent a functional CSC marker associated with chemoresistance. Moreover, [ 68 Ga]Ga‐Pentixafor PET imaging can guide decision‐making for AMD3100 therapy, paving the way for further clinical translation.

Retracted: MYH10 Combines with MYH9 to Recruit USP45 by Deubiquitinating Snail and Promotes Serous Ovarian Cancer Carcinogenesis, Progression, and Cisplatin Resistance

Abstract The poor prognosis of serous ovarian cancer (SOC) is due to its high invasive capacity and cisplatin resistance of SOC cells, whereas the molecular mechanisms remain poorly understood. In the present study, the expression and function of non‐muscle myosin heavy chain IIB (MYH10) in SOC are identified by immunohistochemistry, in vitro, and in vivo studies, respectively. The mechanism of MYH10 is demonstrated by co‐immunoprecipitation, GST pull‐down, confocal laser assays, and so on. The results show that the knockdown of MYH10 suppressed SOC cell proliferation, migration, invasion, metastasis, and cisplatin resistance both in vivo and in vitro. Further studies confirm that the MYH10 protein functional domain combines with non‐muscle myosin heavy chain IIA (MYH9) to recruit the deubiquitinating enzyme Ubiquitin‐specific proteases 45 and deubiquitinates snail to inhibit snail degradation, eventually promoting tumorigenesis, progression, and cisplatin resistance in SOC. In clinical samples, MYH10 expression is significantly elevated in SOC samples compared to the paratumor samples. And the expression of MYH10 is positively correlated with MYH9 expression. MYH10+/MYH9+ co‐expression is an independent prognostic factor for predicting SOC patient survival. These findings uncover a key role of the MYH10‐MYH9‐snail axis in SOC carcinogenesis, progression, and cisplatin resistance, and provide potential novel therapeutic targets for SOC intervention.

Engineering Bifunctional Calcium Alendronate Gene‐Delivery Nanoneedle for Synergistic Chemo/Immuno‐Therapy Against HER2 Positive Ovarian Cancer

Abstract Ovarian cancer is the most lethal gynecological malignancy. Most patients are diagnosed at an advanced stage with widespread peritoneal dissemination and ascites. Bispecific T‐cell engagers (BiTEs) have demonstrated impressive antitumor efficacy in hematological malignancies, but the clinical potency is limited by their short half‐life, inconvenient continuous intravenous infusion, and severe toxicity at relevant therapeutic levels in solid tumors. To address these critical issues, the design and engineering of alendronate calcium (CaALN) based gene‐delivery system is reported to express therapeutic level of BiTE (HER2×CD3) for efficient ovarian cancer immunotherapy. Controllable construction of CaALN nanosphere and nanoneedle is achieved by the simple and green coordination reactions that the distinct nanoneedle‐like alendronate calcium (CaALN‐N) with a high aspect ratio enabled efficient gene delivery to the peritoneum without system in vivo toxicity. Especially, CaALN‐N induced apoptosis of SKOV3‐luc cell via down‐regulation of HER2 signaling pathway and synergized with HER2×CD3 to generate high antitumor response. In vivo administration of CaALN‐N/minicircle DNA encoding HER2×CD3 (MC‐HER2×CD3) produces sustained therapeutic levels of BiTE and suppresses tumor growth in a human ovarian cancer xenograft model. Collectively, the engineered alendronate calcium nanoneedle represents a bifunctional gene delivery platform for the efficient and synergistic treatment of ovarian cancer.

A Promising New Model: Establishment of Patient‐Derived Organoid Models Covering HPV‐Related Cervical Pre‐Cancerous Lesions and Their Cancers

AbstractThe lack of human‐derived in vitro models that recapitulate cervical pre‐cancerous lesions has been the bottleneck in researching human papillomavirus (HPV) infection‐associated pre‐cancerous lesions and cancers for a long time. Here, a long‐term 3D organoid culture protocol for high‐grade squamous intraepithelial lesions and cervical squamous cell carcinoma that stably recapitulates the two tissues of origin is described. Originating from human‐derived samples, a small biobank of cervical pre‐tumoroids and tumoroids that faithfully retains genomic and transcriptomic characteristics as well as the causative HPV genome is established. Cervical pre‐tumoroids and tumoroids show differential responses to common chemotherapeutic agents and grow differently as xenografts in mice. By coculture organoid models with peripheral blood immune cells (PBMCs) stimulated by HPV antigenic peptides, it is illustrated that both organoid models respond differently to immunized PBMCs, supporting organoids as reliable and powerful tools for studying virus‐specific T‐cell responses and screening therapeutic HPV vaccines. In this study, a model of cervical pre‐cancerous lesions containing HPV is established for the first time, overcoming the bottleneck of the current model of human cervical pre‐cancerous lesions. This study establishes an experimental platform and biobanks for in vitro mechanistic research, therapeutic vaccine screening, and personalized treatment for HPV‐related cervical diseases.

A Fifteen‐Gene Classifier to Predict Neoadjuvant Chemotherapy Responses in Patients with Stage IB to IIB Squamous Cervical Cancer

AbstractNeoadjuvant chemotherapy (NACT) remains an attractive alternative for controlling locally advanced cervical cancer. However, approximately 15–34% of women do not respond to induction therapy. To develop a risk stratification tool, 56 patients with stage IB‐IIB cervical cancer are included in 2 research centers from the discovery cohort. Patient‐specific somatic mutations led to NACT non‐responsiveness are identified by whole‐exome sequencing. Next, CRISPR/Cas9‐based library screenings are performed based on these genes to confirm their biological contribution to drug resistance. A 15‐gene classifier is developed by generalized linear regression analysis combined with the logistic regression model. In an independent validation cohort of 102 patients, the classifier showed good predictive ability with an area under the curve of 0.80 (95% confidence interval (CI), 0.69–0.91). Furthermore, the 15‐gene classifier is significantly associated with patient responsiveness to NACT in both univariate (odds ratio, 10.8; 95% CI, 3.55–32.86; p = 2.8 × 10−5) and multivariate analysis (odds ratio, 17.34; 95% CI, 4.04–74.40; p = 1.23 × 10−4) in the validation set. In conclusion, the 15‐gene classifier can accurately predict the clinical response to NACT before treatment, representing a promising approach for guiding the selection of appropriate treatment strategies for locally advanced cervical cancer.

NAT10/ac4C/FOXP1 Promotes Malignant Progression and Facilitates Immunosuppression by Reprogramming Glycolytic Metabolism in Cervical Cancer

AbstractImmunotherapy has recently emerged as the predominant therapeutic approach for cervical cancer (CCa), driven by the groundbreaking clinical achievements of immune checkpoint inhibitors (ICIs), such as anti‐PD‐1/PD‐L1 antibodies. N4‐acetylcytidine (ac4C) modification, catalyzed by NAT10, is an important posttranscriptional modification of mRNA in cancers. However, its impact on immunological dysregulation and the tumor immunotherapy response in CCa remains enigmatic. Here, a significant increase in NAT10 expression in CCa tissues is initially observed that is clinically associated with poor prognosis. Subsequently, it is found that HOXC8 activated NAT10 by binding to its promoter, thereby stimulating ac4C modification of FOXP1 mRNA and enhancing its translation efficiency, eventually leading to induction of GLUT4 and KHK expression. Moreover, NAT10/ac4C/FOXP1 axis activity resulted in increased glycolysis and a continuous increase in lactic acid secretion by CCa cells. The lactic acid‐enriched tumor microenvironment (TME) further contributed to amplifying the immunosuppressive properties of tumor‐infiltrating regulatory T cells (Tregs). Impressively, NAT10 knockdown enhanced the efficacy of PD‐L1 blockade‐mediated tumor regression in vivo. Taken together, the findings revealed the oncogenic role of NAT10 in initiating crosstalk between cancer cell glycolysis and immunosuppression, which can be a target for synergistic PD‐1/PD‐L1 blockade immunotherapy in CCa.

Performance of Liquid Biopsy‐Based Multi‐Omics Biomarkers for Early Detection of Gynecological Malignancies: A Prospective Study (PERCEIVE‐I)

Abstract Liquid biopsy is a promising approach for early detection of gynecological malignancies. In the PERCEIVE‐I study, gynecological Cancer cases ( n  = 249) and age‐matched non‐cancer controls ( n  = 249) are randomly divided into training and test sets at a 1:1 ratio. Data derived from multi‐omics assays are obtained including a cell‐free DNA methylation panel targeting ≈490 000 CpG sites, a mutation panel comprising 168 genes, and eight tumor protein markers. The results showed that the methylation model outperformed the protein and mutation models, demonstrating higher sensitivity (77.2%) while maintaining similar specificity. The multi‐omics model combining methylation and protein markers achieved improved sensitivity (81.9%) with a good specificity (96.9%). The sensitivity varied across different stages, ranging from 66.7% to 100%. The model accurately identified the tissue of origin in 72.1% of cases. The superior performance of the methylation model highlights the potential of integrating multi‐omics for non‐invasive early detection of gynecological malignancies.

Machine Learning‐Enhanced Analysis of Exosomal Surface Sialic Acid Using Surface‐Enhanced Raman Spectroscopy for Ovarian Cancer Diagnosis and Therapeutic Monitoring

ABSTRACT Currently, the absence of ovarian cancer (OC)‐specific biomarkers impedes the development of precise noninvasive diagnostic and monitoring strategies. Exosomal surface sialic acid (SA), a key mediator of intercellular communication and disease progression, emerges as a promising biomarker, though its role in OC remains unclear. Conventional exosome isolation and detection methods exhibit limited clinical utility. Herein, we developed a CD63 aptamer‐functionalized gold array chip integrated with a surface‐enhanced Raman scattering (SERS) nanosensor for sensitive SA analysis. The chip efficiently isolated exosomes from clinical serum, while the nanosensor selectively bound exosomal SA via molecular recognition, thereby altering the SERS intensity ratio of the nanosensor. More importantly, machine learning can discern SA signatures from SERS spectra, achieving 93% accuracy in OC diagnosis. The longitudinal monitoring of SA throughout the entire treatment period (preoperative, postoperative, and chemotherapy) revealed a potential correlation with treatment response as indicated by clinical markers (CA125, HE4), demonstrating the utility of exosomal SA in precision treatment evaluation. This provides a powerful tool for the diagnosis and treatment monitoring of OC and plays a critical role in precision medicine.

Selenium‐Doped Nanoheterojunctions for Highly Efficient Cancer Radiosensitization

AbstractExploring efficient and low‐toxicity radiosensitizers to break through the bottleneck of radiation tolerance, immunosuppression and poor prognosis remains one of the critical developmental challenges in radiotherapy. Nanoheterojunctions, due to their unique physicochemical properties, have demonstrated excellent radiosensitization effects in radiation energy deposition and in lifting tumor radiotherapy inhibition. Herein, they doped selenium (Se) into prussian blue (PB) to construct a nano‐heterojunction (Se@PB), which could promote the increase of Fe2+/Fe3+ ratio and conversion of Se to a high valence state with Se introduction. The Fe2+‐Se‐Fe3+ electron transfer chain accelerates the rate of electron transfer on the surface of the nanoparticles, which in turn endows it with efficient X‐ray energy transfer and electron transport capability, and enhances radiotherapy physical sensitivity. Furthermore, Se@PB induces glutathione (GSH) depletion and Fe2+ accumulation through pro‐Fenton reaction, thereby disturbs the redox balance in tumor cells and enhances biochemical sensitivity of radiotherapy. As an excellent radiosensitizer, Se@PB effectively enhances X‐ray induced mitochondrial dysfunction and DNA damage, thereby promotes cell apoptosis and synergistic cervical cancer radiotherapy. This study elucidates the radiosensitization mechanism of Se‐doped nanoheterojunction from the perspective of the electron transfer chain and biochemistry reaction, which provides an efficient and low‐toxic strategy in radiotherapy.

Inaugurating High‐Throughput Profiling of Extracellular Vesicles for Earlier Ovarian Cancer Detection

AbstractDetecting early cancer through liquid biopsy is challenging due to the lack of specific biomarkers for early lesions and potentially low levels of these markers. The current study systematically develops an extracellular‐vesicle (EV)‐based test for early detection, specifically focusing on high‐grade serous ovarian carcinoma (HGSOC). The marker selection is based on emerging insights into HGSOC pathogenesis, notably that it arises from precursor lesions within the fallopian tube. This work thus establishes murine fallopian tube (mFT) cells with oncogenic mutations and performs proteomic analyses on mFT‐derived EVs. The identified markers are then evaluated with an orthotopic HGSOC animal model. In serially‐drawn blood of tumor‐bearing mice, mFT‐EV markers increase with tumor initiation, supporting their potential use in early cancer detection. A pilot clinical study (n = 51) further narrows EV markers to five candidates, EpCAM, CD24, VCAN, HE4, and TNC. The combined expression of these markers distinguishes HGSOC from non‐cancer with 89% sensitivity and 93% specificity. The same markers are also effective in classifying three groups (non‐cancer, early‐stage HGSOC, and late‐stage HGSOC). The developed approach, for the first time inaugurated in fallopian tube‐derived EVs, could be a minimally invasive tool to monitor women at high risk of ovarian cancer for timely intervention.

PRDX6 Prevents NNMT Ubiquitination and Degradation as a Nonenzymatic Mechanism to Promote Ovarian Cancer Progression

AbstractCancer cells cope with oxidative stress for their proliferation and metastasis by equipping antioxidant systems, among which the antioxidant enzymes peroxiredoxins (PRDXs) play crucial roles. However, whether PRDXs exhibit nonenzymatic functions remains unclear. Here, it is shown that the 1‐cysteine PRDX (PRDX6) upregulates nicotinamide N‐methyltransferase (NNMT) to promote the growth and metastasis of ovarian cancer cells, independently of PRDX6's enzymatic activities. Mechanistically, PRDX6 interacts with NNMT to prevent its binding to the E3 ubiquitin ligase tripartite‐motif protein 56 (TRIM56), leading to the inhibition of NNMT ubiquitination at lysine 23 and 210 and suppression of subsequent proteasomal degradation. In addition, PRDX6‐mediated NNMT upregulation activates mitogen‐activated protein kinase (MAPK) signaling, thereby promoting the growth and metastasis of ovarian cancer cells. Notably, PRDX6 overexpression is associated with higher NNMT protein levels in human ovarian cancer tissues and is predictive of poor prognosis of ovarian cancer patients. Overall, the findings illustrate a critical oncogenic mechanism of the antioxidant enzyme PRDX6 in promoting ovarian cancer progression beyond its enzymatic mechanisms.

High Spatiotemporal Near‐Infrared II Fluorescence Lifetime Imaging for Quantitative Detection of Clinical Tumor Margins

AbstractAccurate detection of tumor margins is essential for successful cancer surgery. While indocyanine green (ICG)‐based near‐infrared (NIR) fluorescence (FL) surgical navigation enhances the visual identification of tumor margins, its accuracy remains subjective, underscoring the need for quantitative approaches. In this study, a high spatiotemporal fluorescence lifetime (FLT) imaging technology is developed in the second near‐infrared window (NIR‐II, 1000–1700 nm) for quantitative tumor margin detection, utilizing folate receptor‐targeted ICG nanoprobes (FPH‐ICG). FPH‐ICG exhibits a significantly prolonged NIR‐II FLT (750 ± 7 ps vs 260 ± 3 ps) and increased NIR‐II FL brightness (FPH‐ICG/ICG = 3.8). In vitro and in vivo studies confirm that FPH‐ICG specifically targets folate receptor‐α (FRα) on SK‐OV‐3 ovarian cancer cells, achieving high‐contrast NIR‐II FL imaging with a signal‐to‐background ratio of 10.8. Notably, NIR‐II FLT imaging demonstrates superior accuracy (90%) and consistency in defining tumor margins compared to NIR‐II FL imaging (58%) in both SK‐OV‐3 tumor‐bearing mice and clinical tumor samples. These findings underscore the potential of NIR‐II FLT imaging as a quantitative tool for guiding surgical tumor margin detection.

Single‐Nucleus Transcriptome Profiling of Locally Advanced Cervical Squamous Cell Cancer Identifies Neural‐Like Progenitor Program Associated with the Efficacy of Radiotherapy

AbstractRadiotherapy is the first‐line treatment for locally advanced cervical squamous cell cancer (CSCC). However, ≈50% of patients fail to respond to therapy and, in some cases, tumors progress after radical radiotherapy. Here, single‐nucleus RNA‐seq is performed to construct high‐resolution molecular landscapes of various cell types in CSCC before and during radiotherapy, to better understand radiotherapy related molecular responses within tumor microenvironment. The results show that expression levels of a neural‐like progenitor (NRP) program in tumor cells are significantly higher after radiotherapy and these are enriched in the tumors of nonresponding patients. The enrichment of the NRP program in malignant cells from the tumors of nonresponders in an independent cohort analyzed by bulk RNA‐seq is validated. In addition, an analysis of The Cancer Genome Atlas dataset shows that NRP expression is associated with poor prognosis in CSCC patients. In vitro experiments on the CSCC cell line demonstrate that downregulation of neuregulin 1 (NRG1), a key gene from NRP program, is associated with decreased cell growth and increased sensitivity to radiation. Immunohistochemistry staining in cohort 3 validated key genes, NRG1 and immediate early response 3 from immunomodulatory program, as radiosensitivity regulators. The findings reveal that the expression of NRP in CSCC can be used to predict the efficacy of radiotherapy.

Cancer Patient Tissueoid with Self‐Homing Nano‐Targeting of Metabolic Inhibitor

Abstract The current paradigm of cancer medicine focuses on patient‐ and/or cancer‐specific treatments, which has led to continuous progress in the development of patient representatives (e.g., organoids) and cancer‐targeting carriers for drug screening. As breakthrough concepts, i) living cancer tissues convey intact profiles of patient‐specific microenvironmental signatures. ii) The growth mechanisms of cancer mass with intense cell‐cell interactions can be harnessed to develop self‐homing nano‐targeting by using cancer cell‐derived nanovesicles (CaNVs). Hence, a tissueoid model of ovarian cancer (OC) is developed by culturing OC patient tissues in a 3D gel chip, whose microchannel networks enable perfusion to maintain tissue viability. A novel model of systemic cancer responses is approached by xenografting OC tissueoids into ischaemic hindlimbs in nude mice. CaNVs are produced to carry general chemotherapeutics or new drugs under pre/clinical studies that target the BRCA mutation or energy metabolism, thereby increasing the test scope. This pioneer study cross‐validates drug responses from the OC clinic, tissueoid, and animal model by demonstrating the alignment of results in drug type‐specific efficiency, BRCA mutation‐dependent drug efficiency, and metabolism inhibition‐based anti‐cancer effects. Hence, this study provides a directional foundation to accelerate the discovery of patient‐specific drugs with CaNV application towards future precision medicine.

Gold Nanoparticles Disrupt the IGFBP2/mTOR/PTEN Axis to Inhibit Ovarian Cancer Growth

AbstractBy exploiting the self‐therapeutic properties of gold nanoparticles (GNPs) a molecular axis that promotes the growth of high‐grade serous ovarian cancer (HGSOC), one of the deadliest gynecologic malignancies with poorly understood underlying molecular mechanisms, has been identified. The biodistribution and toxicity of GNPs administered by intravenous or intraperitoneal injection, both as a single dose or by repeated dosing over two weeks are first assessed; no biochemical or histological toxicity to vital organs is found. Using an orthotopic patient‐derived xenograft (PDX) model of HGSOC, the authors then show that GNP treatment robustly inhibits tumor growth. Investigating the molecular mechanisms underlying the GNP efficacy reveals that GNPs downregulate insulin growth factor binding protein 2 (IGFBP2) by disrupting its autoregulation via the IGFBP2/mTOR/PTEN axis. This mechanism is validated by treating a cell line‐based human xenograft tumor with GNPs and an mTOR dual‐kinase inhibitor (PI‐103), either individually or in combination with GNPs; GNP and PI‐103 combination therapy inhibit ovarian tumor growth similarly to GNPs alone. This report illustrates how the self‐therapeutic properties of GNPs can be exploited as a discovery tool to identify a critical signaling axis responsible for poor prognosis in ovarian cancer and provides an opportunity to interrogate the axis to improve patient outcomes.

Nanomedicine Strategies for Heating “Cold” Ovarian Cancer (OC): Next Evolution in Immunotherapy of OC

AbstractImmunotherapy has revolutionized cancer treatment, dramatically improving survival rates of melanoma and lung cancer patients. Nevertheless, immunotherapy is almost ineffective against ovarian cancer (OC) due to its cold tumor immune microenvironment (TIM). Many traditional medications aimed at remodeling TIM are often associated with severe systemic toxicity, require frequent dosing, and show only modest clinical efficacy. In recent years, emerging nanomedicines have demonstrated extraordinary immunotherapeutic effects for OC by reversing the TIM because the physical and biochemical features of nanomedicines can all be harnessed to obtain optimal and expected tissue distribution and cellular uptake. However, nanomedicines are far from being widely explored in the field of OC immunotherapy due to the lack of appreciation for the professional barriers of nanomedicine and pathology, limiting the horizons of biomedical researchers and materials scientists. Herein, a typical cold tumor‐OC is adopted as a paradigm to introduce the classification of TIM, the TIM characteristics of OC, and the advantages of nanomedicines for immunotherapy. Subsequently, current nanomedicines are comprehensively summarized through five general strategies to substantially enhance the efficacy of immunotherapy by heating the cold OC. Finally, the challenges and perspectives of this expanding field for improved development of clinical applications are also discussed.

Salt‐Inducible Kinase 2‐Triggered Release of Its Inhibitor from Hydrogel to Suppress Ovarian Cancer Metastasis

AbstractSalt‐inducible kinase 2 (SIK2) is a promising target for ovarian cancer therapy due to its critical role in tumorigenesis and progression. Currently available SIK2 inhibitors have shown remarkable therapeutic effects on ovarian cancers in preclinical studies. However, direct administration of the SIK2 inhibitors may bring significant off‐target effect, limiting their clinical applications. In this work, by rational design of a hydrogelator Nap‐Phe‐Phe‐Glu‐Glu‐Leu‐Tyr‐Arg‐Thr‐Gln‐Ser‐Ser‐Ser‐Asn‐Leu‐OH (Nap‐S) to coassemble a SIK2 inhibitor HG‐9‐91‐01 (HG), a SIK2‐responsive supramolecular hydrogel (Gel Nap‐S+HG) for local administration and SIK2‐responsive release of HG is reported to efficiently suppress ovarian cancer metastasis. Under the activation of SIK2 overexpressed in ovarian cancers, Nap‐S in the hydrogel is phosphorylated to yield hydrophilic Nap‐Phe‐Phe‐Glu‐Glu‐Leu‐Tyr‐Arg‐Thr‐Gln‐Ser(H2PO3)‐Ser‐Ser‐Asn‐Leu (Nap‐Sp), triggering the disassembly of the hydrogel and a responsive release of the inhibitor. Cell experiments indicate that sustained release of HG from Gel Nap‐S+HG induce a prominent therapeutic effect on cancer cells by inhibiting SIK2 and phosphorylation of their downstream signaling molecules. Animal experiments demonstrate that, compared with those tumor model mice treated with free HG, Gel Nap‐S+HG‐treatment mice show an enhanced inhibition on ovarian tumor growth and metastasis. It is anticipated that the Gel Nap‐S+HG can be applied for ovarian cancer therapy in clinic in the near future.

Topical pH Sensing NIR Fluorophores for Intraoperative Imaging and Surgery of Disseminated Ovarian Cancer

AbstractFluorescence‐guided surgery (FGS) aids surgeons with real‐time visualization of small cancer foci and borders, which improves surgical and prognostic efficacy of cancer. Despite the steady advances in imaging devices, there is a scarcity of fluorophores available to achieve optimal FGS. Here, 1) a pH‐sensitive near‐infrared fluorophore that exhibits rapid signal changes in acidic tumor microenvironments (TME) caused by the attenuation of intramolecular quenching, 2) the inherent targeting for cancer based on chemical structure (structure inherent targeting, SIT), and 3) mitochondrial and lysosomal retention are reported. After topical application of PH08 on peritoneal tumor regions in ovarian cancer‐bearing mice, a rapid fluorescence increase (< 10 min), and extended preservation of signals (> 4 h post‐topical application) are observed, which together allow for the visualization of submillimeter tumors with a high tumor‐to‐background ratio (TBR > 5.0). In addition, PH08 is preferentially transported to cancer cells via organic anion transporter peptides (OATPs) and colocalizes in the mitochondria and lysosomes due to the positive charges, enabling a long retention time during FGS. PH08 not only has a significant impact on surgical and diagnostic applications but also provides an effective and scalable strategy to design therapeutic agents for a wide array of cancers.

Tumor Derived Extracellular Vesicles Drive T Cell Exhaustion in Tumor Microenvironment through Sphingosine Mediated Signaling and Impacting Immunotherapy Outcomes in Ovarian Cancer

AbstractSPHK1 (sphingosine kinase‐1) catalyzes the phosphorylation of sphingosine to sphingosine‐1‐phosphate (S1P), is found to be highly expressed in solid tumors. Here, extracellular vesicles (EVs) are identified as the key transporters of SPHK1 to the tumor microenvironment. Consequently, SPHK1‐packaged EVs elevate S1P levels in the tumor microenvironment, where S1P appears as an immunosuppressive agent. However, the exact mechanism of how S1P mediates its immunosuppressive effects in cancer is not understood. It is investigated that S1P can induce T cell exhaustion. S1P can also upregulate programmed death ligand‐1 (PDL‐1) expression through E2F1‐mediated transcription. Notably, an SPHK1 inhibitor PF543 improves T cell‐mediated cytotoxicity. Furthermore, combining PF543 with an anti‐PD‐1 antibody reduces tumor burden and metastasis more effectively than PF543 alone in vivo. These data demonstrate a previously unrecognized mechanism of how SPHK1‐packaged EVs contribute to the progression of ovarian cancer and thus present the potential clinical application of inhibiting SPHK1/S1P signaling to improve immune checkpoint blockage (anti‐PD‐1 antibody) therapy in ovarian cancer.

Lemon‐Derived Extracellular Vesicles Nanodrugs Enable to Efficiently Overcome Cancer Multidrug Resistance by Endocytosis‐Triggered Energy Dissipation and Energy Production Reduction

AbstractMultidrug resistance remains a great challenge for cancer chemotherapy. Herein, a biomimetic drug delivery system based on lemon‐derived extracellular vesicles (EVs) nanodrugs (marked with heparin‐cRGD‐EVs‐doxorubicin (HRED)) is demonstrated, achieving highly efficient overcoming cancer multidrug resistance. The HRED is fabricated by modifying functional heparin‐cRGD (HR) onto the surface of EVs and then by loading with doxorubicin (DOX). The obtained HRED enable to effectively enter DOX‐resistant cancer cells by caveolin‐mediated endocytosis (main), macropinocytosis (secondary), and clathrin‐mediated endocytosis (last), exhibiting excellent cellular uptake capacity. The diversified endocytosis capacity of HRED can efficiently dissipate intracellular energy and meanwhile trigger downstream production reduction of adenosine triphosphate (ATP), leading to a significant reduction of drug efflux. Consequently, they show excellent anti‐proliferation capacities to DOX‐resistant ovarian cancer, ensuring the efficiently overcoming ovarian cancer multidrug resistance in vivo. The authors believe this strategy provides a new strategy by endocytosis triggered‐energy dissipation and ATP production reduction to design drug delivery system for overcoming cancer multidrug resistance.

KLF5 Promotes Tumor Progression and Parp Inhibitor Resistance in Ovarian Cancer

AbstractOne major characteristic of tumor cells is the aberrant activation of epigenetic regulatory elements, which remodel the tumor transcriptome and ultimately promote cancer progression and drug resistance. However, the oncogenic functions and mechanisms of ovarian cancer (OC) remain elusive. Here, super‐enhancer (SE) regulatory elements that are aberrantly activated in OC are identified and it is found that SEs drive the relative specific expression of the transcription factor KLF5 in OC patients and poly(ADP‐ribose) polymerase inhibitor (PARPi)‐resistant patients. KLF5 expression is associated with poor outcomes in OC patients and can drive tumor progression in vitro and in vivo. Mechanistically, KLF5 forms a transcriptional complex with EHF and ELF3 and binds to the promoter region of RAD51 to enhance its transcription, strengthening the homologous recombination repair (HRR) pathway. Notably, the combination of suberoylanilide hydroxamic acid (SAHA) and olaparib significantly inhibits tumor growth and metastasis of PARPi‐resistant OC cells with high KLF5. In conclusion, it is discovered that SEs‐driven KLF5 is a key regulatory factor in OC progression and PARPi resistance; and potential therapeutic strategies for OC patients with PARPi resistance and high KLF5 are identified.

NAD+ Metabolism Reprogramming Drives SIRT1‐Dependent Deacetylation Inducing PD‐L1 Nuclear Localization in Cervical Cancer

AbstractCervical cancer (CC) is a major health threat to women, with immunotherapies targeting the programmed death receptor 1/programmed death ligand 1(PD‐1/PD‐L1) axis showing promise but encountering resistance in a significant patient population. This resistance has driven a critical quest to uncover the underlying mechanisms. This study uncovers a novel metabolic axis involving the nicotinamide adenine dinucleotide (NAD+) salvage pathway enzyme nicotinamide phosphoribosyltransferase (NAMPT) and the deacetylase Sirtuin 1 (SIRT1), which regulates PD‐L1 expression and nuclear localization in CC. This axis may be a key factor contributing to the resistance observed in immunotherapy. This study reveals that PD‐L1 overexpression in cancers is regulated by both transcriptional and post‐transcriptional processes. Acetyl‐proteomic analysis pinpoints SIRT1 as a central regulator in the deacetylation of histone H3 at lysines 27, which may influence PD‐L1 subcellular distribution. This finding reveals the epigenetic control of immune checkpoint proteins by metabolic pathways, offering a new perspective on the regulation of PD‐L1. The identification of the NAMPT/SIRT1 metabolic axis as a critical factor suggests that targeting this axis may enhance therapeutic responses.

Single‐Nucleus RNA Sequencing and Spatial Transcriptomics Reveal the Immunological Microenvironment of Cervical Squamous Cell Carcinoma

Abstract The effective treatment of advanced cervical cancer remains challenging. Herein, single‐nucleus RNA sequencing (snRNA‐seq) and SpaTial enhanced resolution omics‐sequencing (Stereo‐seq) are used to investigate the immunological microenvironment of cervical squamous cell carcinoma (CSCC). The expression levels of most immune suppressive genes in the tumor and inflammation areas of CSCC are not significantly higher than those in the non‐cancer samples, except for LGALS9 and IDO1 . Stronger signals of CD56 + NK cells and immature dendritic cells are found in the hypermetabolic tumor areas, whereas more eosinophils, immature B cells, and Treg cells are found in the hypometabolic tumor areas. Moreover, a cluster of pro‐tumorigenic cancer‐associated myofibroblasts (myCAFs) are identified. The myCAFs may support the growth and metastasis of tumors by inhibiting lymphocyte infiltration and remodeling of the tumor extracellular matrix. Furthermore, these myCAFs are associated with poorer survival probability in patients with CSCC, predict resistance to immunotherapy, and might be present in a small fraction (< 30%) of patients with advanced cancer. Immunohistochemistry and multiplex immunofluorescence staining are conducted to validate the spatial distribution and potential function of myCAFs. Collectively, these findings enhance the understanding of the immunological microenvironment of CSCC and shed light on the treatment of advanced CSCC.

Pressure Drives Rapid Burst‐Like Coordinated Cellular Motion from 3D Cancer Aggregates

AbstractA key behavior observed during morphogenesis, wound healing, and cancer invasion is that of collective and coordinated cellular motion. Hence, understanding the different aspects of such coordinated migration is fundamental for describing and treating cancer and other pathological defects. In general, individual cells exert forces on their environment in order to move, and collective motion is coordinated by cell–cell adhesion‐based forces. However, this notion ignores other mechanisms that encourage cellular movement, such as pressure differences. Here, using model tumors, it is found that increased pressure drove coordinated cellular motion independent of cell–cell adhesion by triggering cell swelling in a soft extracellular matrix (ECM). In the resulting phenotype, a rapid burst‐like stream of cervical cancer cells emerged from 3D aggregates embedded in soft collagen matrices (0.5 mg mL−1). This fluid‐like pushing mechanism, recorded within 8 h after embedding, shows high cell velocities and super‐diffusive motion. Because the swelling in this model system critically depends on integrin‐mediated cell–ECM adhesions and cellular contractility, the swelling is likely triggered by unsustained mechanotransduction, providing new evidence that pressure‐driven effects must be considered to more completely understand the mechanical forces involved in cell and tissue movement as well as invasion.

PARPi Combining Nanoparticle LIN28B siRNA for the Management of Malignant Ascites

ABSTRACT Malignant serous effusion (MSE), including malignant pleural effusion (MPE) and malignant ascites (MA), is a common and severe complication in advanced malignancies, associated with poor prognosis and high recurrence rates. Currently, no standardized treatments are available for MSE management, posing significant clinical challenges. Here, we identify elevated LIN28B expression and dysregulation of DNA repair pathways as two major features associated with MSE from patient and preclinical samples. We develop a targeted siRNA nanoparticle delivery system (siLin28B/DSSP@lip‐PEG‐FA) in combination with the PARP inhibitor BMN673, providing a synergistic therapeutic strategy against MSE. This combination significantly alleviated MA accumulation and prolonged survival in a preclinical ovarian cancer (OC) model without causing systemic cytotoxicity. Mechanistically, single‐cell RNA sequencing (scRNA‐seq) revealed that this combination therapy markedly remodeled the immune microenvironment by decreasing M2 macrophages and neutrophil populations with altered subtypes. Notably, Arg1‐positive neutrophils, producing pro‐inflammatory cytokines to increase vascular permeability, were diminished after the combination treatment. Furthermore, in vitro and in vivo experiments demonstrated that suppression of PARP and LIN28B inhibited vascular leakage and reinforced tight junction integrity. Collectively, our findings highlight dual targeting of PARP and LIN28B as a promising MA management approach in patients with advanced cancers, with the potential to improve patient quality of life.

Dual Targeting of Mutant p53 and SNRPD2 via Engineered Exosomes Modulates Alternative Splicing to Suppress Ovarian Cancer

ABSTRACT Mutation of the tumor suppressor gene TP53 promotes ovarian cancer progression and therapeutic resistance. Whether mutant p53 (mtp53) regulates alternative splicing and how this regulation can be exploited for cancer therapy remain unclear. Here, small nuclear ribonucleoprotein D2 polypeptide (SNRPD2) as a binding partner of mtp53 is identified. SNRPD2 is highly expressed in ovarian cancer and associated with an unfavorable prognosis. The overexpression of SNRPD2 promotes, whereas its depletion inhibits, the growth and migration of ovarian cancer cells. Mechanistically, mtp53 cooperates with SNRPD2 to facilitate the assembly of the Sm/SMN protein complex, an essential component of the spliceosome, modulating alternative splicing of pre‐mRNAs. Specifically, the co‐depletion of mtp53 and SNRPD2 reduces the level of OTUD3 oncogenic transcripts while increasing its tumor suppressor counterparts through an exon‐skipping event. Moreover, therapeutic engineered exosomes are developed with their surfaces decorated with iRGD and their interiors loaded with siRNAs targeting mtp53 and SNRPD2. These exosomes effectively suppress the growth of ovarian cancer cells and enhance their sensitivity to chemotherapy in vivo. Collectively, this study uncovers that mtp53 and SNRPD2 cooperatively regulate alternative splicing to drive ovarian cancer progression, and co‐targeting these two molecules via engineered exosomes represents a potential therapeutic strategy for ovarian cancer.