Cell Reports

P-cadherin mechanoactivates tumor-mesothelium metabolic coupling to promote ovarian cancer metastasis

Cancer adhesion to the mesothelium is critical for peritoneal metastasis, but how metastatic cells adapt to the biomechanical microenvironment remains unclear. Our study demonstrates that highly metastatic (HM), but not non-metastatic, ovarian cancer cells selectively activate the peritoneal mesothelium. HM cells exert a stronger adhesive force on mesothelial cells via P-cadherin, an adhesion molecule abundant in late-stage tumors. Mechanical activation of P-cadherin enhances lipogenic gene expression and lipid content in HM cells through SREBP1. P-cadherin also induces glycolysis in the interacting mesothelium without affecting lipogenic activity, with the resulting lactate serving as a substrate for lipogenesis in HM cells. Nanodelivery of small interfering RNA (siRNA) targeting P-cadherin or MCT1/4 transporters significantly suppresses metastasis in mice. Moreover, increased fatty acid synthase levels in metastatic patient samples correlate with high P-cadherin expression, supporting enhanced de novo lipogenesis in the metastatic niche. This study reveals P-cadherin-mediated mechano-metabolic coupling as a promising target to restrain metastasis.

Tim4 enables large peritoneal macrophages to cross-present tumor antigens at early stages of tumorigenesis

Receptors controlling the cross-presentation of tumor antigens by macrophage subsets in cancer tissues are poorly explored. Here, we show that TIM4

Epigenomic Reprogramming toward Mesenchymal-Epithelial Transition in Ovarian-Cancer-Associated Mesenchymal Stem Cells Drives Metastasis

A role for cancer cell epithelial-to-mesenchymal transition (EMT) in cancer is well established. Here, we show that, in addition to cancer cell EMT, ovarian cancer cell metastasis relies on an epigenomic mesenchymal-to-epithelial transition (MET) in host mesenchymal stem cells (MSCs). These reprogrammed MSCs, termed carcinoma-associated MSCs (CA-MSCs), acquire pro-tumorigenic functions and directly bind cancer cells to serve as a metastatic driver/chaperone. Cancer cells induce this epigenomic MET characterized by enhancer-enriched DNA hypermethylation, altered chromatin accessibility, and differential histone modifications. This phenomenon appears clinically relevant, as CA-MSC MET is highly correlated with patient survival. Mechanistically, mirroring MET observed in development, MET in CA-MSCs is mediated by WT1 and EZH2. Importantly, EZH2 inhibitors, which are clinically available, significantly inhibited CA-MSC-mediated metastasis in mouse models of ovarian cancer.

Patient-Derived Ovarian Cancer Organoids Mimic Clinical Response and Exhibit Heterogeneous Inter- and Intrapatient Drug Responses

There remains an unmet need for preclinical models to enable personalized therapy for ovarian cancer (OC) patients. Here we evaluate the capacity of patient-derived organoids (PDOs) to predict clinical drug response and functional consequences of tumor heterogeneity. We included 36 whole-genome-characterized PDOs from 23 OC patients with known clinical histories. OC PDOs maintain the genomic features of the original tumor lesion and recapitulate patient response to neoadjuvant carboplatin/paclitaxel combination treatment. PDOs display inter- and intrapatient drug response heterogeneity to chemotherapy and targeted drugs, which can be partially explained by genetic aberrations. PDO drug screening identifies high responsiveness to at least one drug for 88% of patients. PDOs are valuable preclinical models that can provide insights into drug response for individual patients with OC, complementary to genetic testing. Generating PDOs of multiple tumor locations can improve clinical decision making and increase our knowledge of genetic and drug response heterogeneity.

Oviduct epithelial cells constitute two developmentally distinct lineages that are spatially separated along the distal-proximal axis

Owing to technical advances in single-cell biology, the appreciation of cellular heterogeneity has increased, which has aided our understanding of organ function, homeostasis, and disease progression. The oviduct (also known as the fallopian tube) is the distalmost portion of the female reproductive tract. It is essential for reproduction and the proposed origin of high-grade serous ovarian carcinoma (HGSOC). In mammals, the oviduct is morphologically segmented along the ovary-uterus axis into four evolutionally conserved regions. It is unclear, however, if there is a diversification of epithelial cell characteristics between these regions. In this study, we identify transcriptionally distinct populations of secretory and multiciliated cells restricted to the distal and proximal regions of the oviduct. We demonstrate that distal and proximal populations are distinct lineages specified early in Müllerian duct development and are maintained separately. These results aid our understanding of epithelial development, homeostasis, and initiation of disease from the oviduct.

Molecular Analysis of Clinically Defined Subsets of High-Grade Serous Ovarian Cancer

The diversity and heterogeneity within high-grade serous ovarian cancer (HGSC), which is the most lethal gynecologic malignancy, is not well understood. Here, we perform comprehensive multi-platform omics analyses, including integrated analysis, and immune monitoring on primary and metastatic sites from highly clinically annotated HGSC samples based on a laparoscopic triage algorithm from patients who underwent complete gross resection (R0) or received neoadjuvant chemotherapy (NACT) with excellent or poor response. We identify significant distinct molecular abnormalities and cellular changes and immune cell repertoire alterations between the groups, including a higher rate of NF1 copy number loss, and reduced chromothripsis-like patterns, higher levels of strong-binding neoantigens, and a higher number of infiltrated T cells in the R0 versus the NACT groups.

High-grade serous ovarian tumor cells modulate NK cell function to create an immune-tolerant microenvironment

Tubo-ovarian high-grade serous carcinoma (HGSC) is unresponsive to immune checkpoint blockade despite significant frequencies of exhausted T cells. Here we apply mass cytometry and uncover decidual-like natural killer (dl-NK) cell subpopulations (CD56+CD9+CXCR3+KIR+CD3-CD16-) in newly diagnosed HGSC samples that correlate with both tumor and transitioning epithelial-mesenchymal cell abundance. We show different combinatorial expression patterns of ligands for activating and inhibitory NK receptors within three HGSC tumor compartments: epithelial (E), transitioning epithelial-mesenchymal (EV), and mesenchymal (vimentin expressing [V]), with a more inhibitory ligand phenotype in V cells. In cocultures, NK-92 natural killer cells acquire CD9 from HGSC tumor cells by trogocytosis, resulting in reduced anti-tumor cytokine production and cytotoxicity. Cytotoxicity in these cocultures is restored with a CD9-blocking antibody or CD9 CRISPR knockout, thereby identifying mechanisms of immune suppression in HGSC. CD9 is widely expressed in HGSC tumors and so represents an important new therapeutic target with immediate relevance for NK immunotherapy.

CD63-mediated cloaking of VEGF in small extracellular vesicles contributes to anti-VEGF therapy resistance

Despite wide use of anti-vascular endothelial growth factor (VEGF) therapy for many solid cancers, most individuals become resistant to this therapy, leading to disease progression. Therefore, new biomarkers and strategies for blocking adaptive resistance of cancer to anti-VEGF therapy are needed. As described here, we demonstrate that cancer-derived small extracellular vesicles package increasing quantities of VEGF and other factors in response to anti-VEGF therapy. The packaging process of VEGF into small extracellular vesicles (EVs) is mediated by the tetraspanin CD63. Furthermore, small EV-VEGF (eVEGF) is not accessible to anti-VEGF antibodies and can trigger intracrine VEGF signaling in endothelial cells. eVEGF promotes angiogenesis and enhances tumor growth despite bevacizumab treatment. These data demonstrate a mechanism where VEGF is partitioned into small EVs and promotes tumor angiogenesis and progression. These findings have clinical implications for biomarkers and therapeutic strategies for ovarian cancer.

Multi-omics analysis reveals distinct non-reversion mechanisms of PARPi resistance in BRCA1- versus BRCA2-deficient mammary tumors

BRCA1 and BRCA2 both function in DNA double-strand break repair by homologous recombination (HR). Due to their HR defect, BRCA1/2-deficient cancers are sensitive to poly(ADP-ribose) polymerase inhibitors (PARPis), but they eventually acquire resistance. Preclinical studies yielded several PARPi resistance mechanisms that do not involve BRCA1/2 reactivation, but their relevance in the clinic remains elusive. To investigate which BRCA1/2-independent mechanisms drive spontaneous resistance in vivo, we combine molecular profiling with functional analysis of HR of matched PARPi-naive and PARPi-resistant mouse mammary tumors harboring large intragenic deletions that prevent reactivation of BRCA1/2. We observe restoration of HR in 62% of PARPi-resistant BRCA1-deficient tumors but none in the PARPi-resistant BRCA2-deficient tumors. Moreover, we find that 53BP1 loss is the prevalent resistance mechanism in HR-proficient BRCA1-deficient tumors, whereas resistance in BRCA2-deficient tumors is mainly induced by PARG loss. Furthermore, combined multi-omics analysis identifies additional genes and pathways potentially involved in modulating PARPi response.

IGF2BP2 promotes malignant progression of ovarian cancer by regulating protein synthesis through liquid-liquid phase separation

Ovarian cancer (OC) is the most lethal gynecological malignancy, and high expression of IGF2BP2 is significantly associated with poor prognosis. Intracellular macromolecules regulate diverse biological functions via liquid-liquid phase separation (LLPS). However, whether IGF2BP2 undergoes LLPS and promotes tumor progression in OC through spatiotemporal coordination remains elusive. Here, we demonstrate that IGF2BP2 forms condensates through LLPS in OC cells, which recruit target RNAs and translation initiation factors to enhance de novo protein synthesis. Both N6-methyladenosine (m

MAPK14/p38α shapes the molecular landscape of endometrial cancer and promotes tumorigenic characteristics

The molecular underpinnings of high-grade endometrial carcinoma (HGEC) metastatic growth and survival are poorly understood. Here, we show that ascites-derived and primary tumor HGEC cell lines in 3D spheroid culture faithfully recapitulate key features of malignant peritoneal effusion and exhibit fundamentally distinct transcriptomic, proteomic, and metabolomic landscapes compared with conventional 2D monolayers. Using a genetic screening platform, we identify MAPK14 (which encodes the protein kinase p38α) as a specific requirement for HGEC in spheroid culture. MAPK14/p38α has broad roles in programming the phosphoproteome, transcriptome, and metabolome of HGEC spheroids, yet has negligible impact on monolayer cultures. MAPK14 promotes tumorigenicity in vivo and is specifically required to sustain a sub-population of spheroid cells that is enriched in cancer stemness markers. Therefore, spheroid growth of HGEC activates unique biological programs, including p38α signaling, that cannot be captured using 2D culture models and are highly relevant to malignant disease pathology.

Leader cells promote immunosuppression to drive ovarian cancer progression in vivo

Over 75% of patients with ovarian cancer present with late-stage disease, often accompanied by extensive metastasis. The metastatic cascade is driven by a sub-population of transcriptionally plastic cells known as "leader cells" (LCs), which play a critical role in collective invasion yet remain poorly understood. LCs are marked by the expression of keratin-14 (KRT14), which determines their migratory and invasive capacity in ovarian cancer. This study demonstrates that KRT14+ LCs promote tumor progression through immunosuppression and immune privilege in vivo. In the ID8 syngeneic epithelial ovarian cancer mouse model, tumor-specific loss of KRT14+ LCs impairs tumor progression and metastatic spread without affecting cellular proliferation. Immune profiling shows reduced immunosuppressive regulatory T cells (Tregs) and M2 macrophages and improved CD8

Targeting the GPI transamidase subunit GPAA1 abrogates the CD24 immune checkpoint in ovarian cancer

CD24 is frequently overexpressed in ovarian cancer and promotes immune evasion by interacting with its receptor Siglec10, present on tumor-associated macrophages, providing a "don't eat me" signal that prevents targeting and phagocytosis by macrophages. Factors promoting CD24 expression could represent novel immunotherapeutic targets for ovarian cancer. Here, using a genome-wide CRISPR knockout screen, we identify GPAA1 (glycosylphosphatidylinositol anchor attachment 1), a factor that catalyzes the attachment of a glycosylphosphatidylinositol (GPI) lipid anchor to substrate proteins, as a positive regulator of CD24 cell surface expression. Genetic ablation of GPAA1 abolishes CD24 cell surface expression, enhances macrophage-mediated phagocytosis, and inhibits ovarian tumor growth in mice. GPAA1 shares structural similarities with aminopeptidases. Consequently, we show that bestatin, a clinically advanced aminopeptidase inhibitor, binds to GPAA1 and blocks GPI attachment, resulting in reduced CD24 cell surface expression, increased macrophage-mediated phagocytosis, and suppressed growth of ovarian tumors. Our study highlights the potential of targeting GPAA1 as an immunotherapeutic approach for CD24

Cell-autonomous inflammation of BRCA1-deficient ovarian cancers drives both tumor-intrinsic immunoreactivity and immune resistance via STING

In this study, we investigate mechanisms leading to inflammation and immunoreactivity in ovarian tumors with homologous recombination deficiency (HRD). BRCA1 loss is found to lead to transcriptional reprogramming in tumor cells and cell-intrinsic inflammation involving type I interferon (IFN) and stimulator of IFN genes (STING). BRCA1-mutated (BRCA1

Endothelial p130cas confers resistance to anti-angiogenesis therapy

Anti-angiogenic therapies, such as anti-VEGF antibodies (AVAs), have shown promise in clinical settings. However, adaptive resistance to such therapies occurs frequently. We use orthotopic ovarian cancer models with AVA-adaptive resistance to investigate the underlying mechanisms. Genomic profiling of AVA-resistant tumors guides us to endothelial p130cas. We find that bevacizumab induces cleavage of VEGFR2 in endothelial cells by caspase-10 and that VEGFR2 fragments internalize into the nucleus and autophagosomes. Nuclear VEGFR2 and p130cas fragments, together with TNKS1BP1 (tankyrase-1-binding protein), initiate endothelial cell death. Blockade of autophagy in AVA-resistant endothelial cells retains VEGFR2 at the membrane with bevacizumab treatment. Targeting host p130cas with RGD (Arg-Gly-Asp)-tagged nanoparticles or genomic ablation of vascular p130cas in p130cas

Quantitative differences between cyclin-dependent kinases underlie the unique functions of CDK1 in human cells

One of the most intriguing features of cell-cycle control is that, although there are multiple cyclin-dependent kinases (CDKs) in higher eukaryotes, a single CDK is responsible for both G

Single-cell resolution of an open chromatin signature in persister tumor cells

PRMT5-mediated methylation of METTL3 promotes cisplatin resistance in ovarian cancer by facilitating DNA repair mechanisms

Cisplatin (CDDP) is a widely used chemotherapy drug for treating various solid tumors. However, resistance to CDDP significantly hampers patient outcomes. This study reveals that protein arginine methyltransferase (PRMT)5 methylates METTL3 at the R36 residue (METTL3-R36me2), which is crucial for CDDP resistance in ovarian cancer (OC) cells. Following CDDP exposure, MST4 is transactivated by nuclear factor-erythroid 2-related factor 2 (NRF2), a key regulator of antioxidant responses. MST4 stimulates PRMT5's methyltransferase activity and promotes its interaction with METTL3 via phosphorylation at Ser439 and Ser463, resulting in increased levels of METTL3-R36me2 and mRNA methylation at the N6 position of adenosine (m

Targeting ZDHHC12-mediated PARP1 palmitoylation potentiates PARP inhibitor cytotoxicity

Poly(ADP-ribose) polymerase inhibitors (PARPi) elicit cytotoxicity by trapping PARP1 at DNA lesions, but clinical resistance remains a major challenge. Here, we identify reversible S-palmitoylation as a negative regulator of PARP1 chromatin engagement. Mass spectrometry reveals PARP1 palmitoylation at conserved cysteines within its DNA-binding domains. DNA damage reduces PARP1 palmitoylation, enhancing DNA binding, whereas inhibition of depalmitoylases APT1/2 elevates palmitoylation and suppresses DNA binding. The palmitoyltransferase ZDHHC12 catalyzes PARP1 palmitoylation, and its inhibition, along with the blockade of palmitate synthesis or global palmitoylation, augments PARP1 trapping and sensitizes high-grade ovarian cancer (HGSOC) cells to the PARPi Niraparib. Patient-derived PARP1 variants R138C and R591C display hyper-palmitoylation, impaired trapping, and PARPi resistance through an indirect mechanism independent of palmitoylation at the mutation sites. ZDHHC12 knockdown restores PARP1 trapping and resensitizes resistant cells and xenografts to Niraparib. These findings establish ZDHHC12-mediated PARP1 palmitoylation as a targetable vulnerability to overcome PARPi resistance.

PAI-1-Dependent Inactivation of SMAD4-Modulated Junction and Adhesion Complex in Obese Endometrial Cancer

While plasminogen activator inhibitor-1 (PAI-1) is known to potentiate cellular migration via proteolytic regulation, this adipokine is implicated as an oncogenic ligand in the tumor microenvironment. To understand the underlying paracrine mechanism, here, we conduct transcriptomic analysis of 1,898 endometrial epithelial cells (EECs) exposed and unexposed to PAI-1-secreting adipose stromal cells. The PAI-1-dependent action deregulates crosstalk among tumor-promoting and tumor-repressing pathways, including transforming growth factor β (TGF-β). When PAI-1 is tethered to lipoprotein receptor-related protein 1 (LRP1), the internalized signaling causes downregulation of SMAD4 at the transcriptional and post-translational levels that attenuates TGF-β-related transcription programs. Repression of genes encoding the junction and adhesion complex preferentially occurs in SMAD4-underexpressed EECs of persons with obesity. The findings highlight a role of PAI-1 signaling that renders ineffective intercellular communication for the development of adiposity-associated endometrial cancer.

MAP4K Interactome Reveals STRN4 as a Key STRIPAK Complex Component in Hippo Pathway Regulation

Mitogen-activated protein kinase kinase kinase kinases (MAP4Ks) constitute a mammalian STE20-like serine/threonine kinase subfamily. Recent studies provide substantial evidence for MAP4K family kinases in the Hippo pathway regulation, suggesting a broad role of MAP4Ks in human physiology and diseases. However, a comprehensive analysis of the regulators and effectors for this key kinase family has not been fully achieved. Using a proteomic approach, we define the protein-protein interaction network for human MAP4K family kinases and reveal diverse cellular signaling events involving this important kinase family. Through it, we identify a STRIPAK complex component, STRN4, as a generic binding partner for MAP4Ks and a key regulator of the Hippo pathway in endometrial cancer development. Taken together, the results of our study not only generate a rich resource for further characterizing human MAP4K family kinases in numerous biological processes but also dissect the STRIPAK-mediated regulation of MAP4Ks in the Hippo pathway.

ESM1 SUMOylation mediates bevacizumab resistance in ovarian cancer through ITGB1-FAK-driven angiogenesis

Bevacizumab (Bev) resistance limits therapeutic efficacy in ovarian cancer (OC) patients. We identified ESM1 as a key gene in Bev-resistant OC. ESM1 secreted by OC-resistant cell lines activates the ITGB1/FAK axis to induce neovascularization and Bev resistance. Additionally, ESM1 overexpression promoted the growth and Bev resistance of OC, lung, intestinal, and hepatocellular carcinoma tumors. Then, we identified TRIM28 as an upstream regulator that stabilizes ESM1 by promoting SUMOylation, inhibiting its proteasomal degradation. In OC mice, TRIM28 overexpression promotes angiogenesis and Bev resistance via ESM1-mediated ITGB1/FAK activation. This work unveils a new molecular pathway underlying Bev resistance in OC and proposes TRIM28 and ESM1 as potential therapeutic targets.

FAK inhibition in ovarian cancer releases omega-3 fatty acids to program CXCL13-producing anti-tumor resident peritoneal macrophages

High-grade serous ovarian cancer (HGSOC) is a lethal malignancy characterized by therapy resistance. Focal adhesion kinase (FAK) is highly expressed in HGSOC, yet its impact on tumor-immune communication remains incompletely defined. Using three syngeneic ovarian cancer models, we show that FAK inhibition (FAKi) increased macrophage CXCL13 expression and promoted peritoneal B cell infiltration. Combining FAKi with low-dose pegylated doxorubicin and anti-T cell immunoreceptor with Ig and ITIM domains (TIGIT) checkpoint blockade suppressed orthotopic ovarian tumor growth, extended survival, and induced tertiary lymphoid structures. Macrophage lineage factor GATA6 inactivation reduced CXCL13 expression, enhanced FAK-knockout tumor growth, and limited ascites B cell accumulation. Mechanistically, FAKi-treated or FAK-deficient tumor cells release exosomes enriched in omega-3 fatty acids that stimulated macrophage CXCL13 production. Exposure of macrophages to tumor-derived omega-3 lipids or eicosapentaenoic acid induced anti-tumor reprogramming and CXCL13 expression. Together, these findings reveal a tumor lipid-macrophage signaling axis activated by FAKi that supports B cell recruitment and anti-TIGIT immunotherapy.

Allosterically inhibited PFKL via prostaglandin E2 withholds glucose metabolism and ovarian cancer invasiveness

Metastasis is the leading cause of high ovarian-cancer-related mortality worldwide. Three major processes constitute the whole metastatic cascade: invasion, intravasation, and extravasation. Tumor cells often reprogram their metabolism to gain advantages in proliferation and survival. However, whether and how those metabolic alterations contribute to the invasiveness of tumor cells has yet to be fully understood. Here we performed a genome-wide CRISPR-Cas9 screening to identify genes participating in tumor cell dissemination and revealed that PTGES3 acts as an invasion suppressor in ovarian cancer. Mechanistically, PTGES3 binds to phosphofructokinase, liver type (PFKL) and generates a local source of prostaglandin E2 (PGE2) to allosterically inhibit the enzymatic activity of PFKL. Repressed PFKL leads to downgraded glycolysis and the subsequent TCA cycle for glucose metabolism. However, ovarian cancer suppresses the expression of PTGES3 and disrupts the PTGES3-PGE2-PFKL inhibitory axis, leading to hyperactivation of glucose oxidation, eventually facilitating ovarian cancer cell motility and invasiveness.

Integrated Proteomic and Glycoproteomic Characterization of Human High-Grade Serous Ovarian Carcinoma

Many gene products exhibit great structural heterogeneity because of an array of modifications. These modifications are not directly encoded in the genomic template but often affect the functionality of proteins. Protein glycosylation plays a vital role in proper protein functions. However, the analysis of glycoproteins has been challenging compared with other protein modifications, such as phosphorylation. Here, we perform an integrated proteomic and glycoproteomic analysis of 83 prospectively collected high-grade serous ovarian carcinoma (HGSC) and 23 non-tumor tissues. Integration of the expression data from global proteomics and glycoproteomics reveals tumor-specific glycosylation, uncovers different glycosylation associated with three tumor clusters, and identifies glycosylation enzymes that were correlated with the altered glycosylation. In addition to providing a valuable resource, these results provide insights into the potential roles of glycosylation in the pathogenesis of HGSC, with the possibility of distinguishing pathological outcomes of ovarian tumors from non-tumors, as well as classifying tumor clusters.

Decoding the epigenetics of persister states in ovarian cancer

Dumbrava et al. map a unified epigenetically primed persister state in high-grade serous ovarian carcinoma using single-nucleus multi-omics. Their work uncovers a chromatin-encoded tolerance program that precedes transcriptional change, revealing therapeutic options and motivating deeper spatiotemporal mapping of persister cells.

Targeting TCF19 sensitizes MSI endometrial cancer to anti-PD-1 therapy by alleviating CD8+ T cell exhaustion via TRIM14-IFN-β axis

Immune checkpoint blockade (ICB) therapies display clinical efficacy in microsatellite instable (MSI) endometrial cancer (EC) treatment, the key mechanism of which is reversing T cell exhaustion and restoration of anti-tumor immunity. Here, we demonstrate that transcription factor 19 (TCF19), one of the most significantly differentially expressed genes between MSI and microsatellite stable (MSS) patients in The Cancer Genome Atlas (TCGA)-EC cohort, is associated with poor prognosis and immune exhaustion signature. Specifically, TCF19 is significantly elevated in MSI EC, which in turn promotes tripartite motif-containing 14 (TRIM14) transcription and correlates with hyperactive signaling of the TANK-binding kinase 1 (TBK1)-interferon regulatory factor 3 (IRF3)-interferon β (IFN-β) pathway. The TCF19-TRIM14 axis promotes tumorigenicity under non-immunological background, and the enhanced downstream secretion of IFN-β facilitates CD8

A tumor-resident bacterium promotes cervical cancer progression through lysoPC-mediated c-Jun/c-Fos activation

Tumor-resident microbiota are identified as a critical modulator in the carcinogenesis and progression of cervical malignancies. However, the mechanisms linking these microbiota to cervical cancer remain poorly understood. In this study, we find that patients with cervical cancer exhibit increased microbiota diversity, characterized by eight distinct bacterial species that distinguish them from control subjects. Intratumoral or intravenous administration of an isolated strain, Gordonia polyisoprenivorans GP-2, derived from tumor-resident microbiota, significantly promotes the growth and metastasis of cervical cancer in mice. Further investigation reveals that GP-2 metabolizes host-derived choline to produce the bacterial metabolite lysophosphatidylcholine (lysoPC). We propose and validate the biosynthetic pathway for lysoPC in strain GP-2 and demonstrate that lysoPC activates the classical tumorigenic genes c-Jun/c-Fos using cervical tumor organoids derived from patients. These findings elucidate the mechanisms underlying microbiota-host crosstalk in cervical cancer and highlight a potential therapeutic target for intervening in tumor-resident bacteria.

Cancer-associated mesothelial cells are regulated by the anti-Müllerian hormone axis

Cancer-associated mesothelial cells (CAMCs) in the tumor microenvironment are thought to promote growth and immune evasion. We find that, in mouse and human ovarian tumors, cancer cells express anti-Müllerian hormone (AMH) while CAMCs express its receptor AMHR2, suggesting a paracrine axis. Factors secreted by cancer cells induce AMHR2 expression during their reprogramming into CAMCs in mouse and human in vitro models. Overexpression of AMHR2 in the Met5a mesothelial cell line is sufficient to induce expression of immunosuppressive cytokines and growth factors that stimulate ovarian cancer cell growth in an AMH-dependent way. Finally, syngeneic cancer cells implanted in transgenic mice with Amhr2

Mouse Ovarian Cancer Models Recapitulate the Human Tumor Microenvironment and Patient Response to Treatment

Although there are many prospective targets in the tumor microenvironment (TME) of high-grade serous ovarian cancer (HGSOC), pre-clinical testing is challenging, especially as there is limited information on the murine TME. Here, we characterize the TME of six orthotopic, transplantable syngeneic murine HGSOC lines established from genetic models and compare these to patient biopsies. We identify significant correlations between the transcriptome, host cell infiltrates, matrisome, vasculature, and tissue modulus of mouse and human TMEs, with several stromal and malignant targets in common. However, each model shows distinct differences and potential vulnerabilities that enabled us to test predictions about response to chemotherapy and an anti-IL-6 antibody. Using machine learning, the transcriptional profiles of the mouse tumors that differed in chemotherapy response are able to classify chemotherapy-sensitive and -refractory patient tumors. These models provide useful pre-clinical tools and may help identify subgroups of HGSOC patients who are most likely to respond to specific therapies.

Landscape of extrachromosomal DNA characteristics in high-grade serous ovarian cancer via long-read sequencing

The non-Mendelian inheritance of extrachromosomal DNA (ecDNA) exacerbates tumor genomic heterogeneity and evolution. Due to short-read sequencing's reconstruction deficiencies, ecDNA abundance in clinical tumor tissues requires further estimation, and the mechanisms driving tumor evolution remain underexplored. Here, we perform long-read whole-genome sequencing on primary and paired metastatic tumor tissues from 12 patients with high-grade serous ovarian cancer (HGSOC) and 6 normal tubal tissues, constructing a comprehensive ecDNA profile. In HGSOC, ecDNA exhibits significantly greater genomic instability and lower methylation than chromosomal DNA. Beyond oncogene and immunomodulatory gene amplification, ecDNA amplifies extensive enhancers to promote gene expression via mechanisms including enhancer hijacking and mobile enhancers. Notably, we observe activation of transposable elements on hypomethylated ecDNA, which is strongly correlated with epithelial-mesenchymal transition gene expression and poor clinical outcomes. These findings reveal ecDNA as a multifunctional driver of genomic variations and tumor progression in HGSOC, highlighting its therapeutic potential.

GAGE mediates radio resistance in cervical cancers via the regulation of chromatin accessibility

Radiotherapy (RT) resistance is a major cause of treatment failure in cancers that use definitive RT as their primary treatment modality. This study identifies the cancer/testis (CT) antigen G antigen (GAGE) as a mediator of radio resistance in cervical cancers. Elevated GAGE expression positively associates with de novo RT resistance in clinical samples. GAGE, specifically the GAGE12 protein variant, confers RT resistance through synemin-dependent chromatin localization, promoting the association of histone deacetylase 1/2 (HDAC1/2) to its inhibitor actin. This cumulates to elevated histone 3 lysine 56 acetylation (H3K56Ac) levels, increased chromatin accessibility, and improved DNA repair efficiency. Molecular or pharmacological disruption of the GAGE-associated complex restores radiosensitivity. Molecularly, this study demonstrates the role of GAGE in the regulation of chromatin dynamics. Clinically, this study puts forward the utility of GAGE as a pre-screening biomarker to identify poor responders at initial diagnosis and the therapeutic potential of agents that target GAGE and its associated complex in combination with radiotherapy to improve outcomes.

Replication stress and defective checkpoints make fallopian tube epithelial cells putative drivers of high-grade serous ovarian cancer

Clinical and molecular evidence indicates that high-grade serous ovarian cancer (HGSOC) primarily originates from the fallopian tube, not the ovarian surface. However, the reasons for this preference remain unclear. Our study highlights significant differences between fallopian tube epithelial (FTE) and ovarian surface epithelial (OSE) cells, providing the molecular basis for FTEs as site of origin of HGSOC. FTEs, unlike OSEs, exhibit heightened replication stress (RS), impaired repair of stalled forks, ineffective G2/M checkpoint, and increased tumorigenicity. BRCA1 heterozygosity exacerbates these defects, resulting in RS suppression haploinsufficiency and an aggressive tumor phenotype. Examination of human and mouse sections reveals buildup of the RS marker 53BP1 primarily in the fallopian tubes, particularly at the fimbrial ends. Furthermore, menopausal status influences RS levels. Our study provides a mechanistic rationale for FTE as the site of origin for HGSOC, investigates the impact of BRCA1 heterozygosity, and lays the groundwork for targeting early HGSOC drivers.

Characterizing the tumor microenvironment of metastatic ovarian cancer by single-cell transcriptomics

Understanding the cellular composition of the tumor microenvironment and the interactions of the cells is essential to the development of successful immunotherapies in cancer. We perform single-cell RNA sequencing (scRNA-seq) of 9,885 cells isolated from the omentum in 6 patients with ovarian cancer and identify 9 major cell types, including cancer, stromal, and immune cells. Transcriptional analysis of immune cells stratifies our patient samples into 2 groups: (1) high T cell infiltration (high T

PAI-1 uncouples integrin-β1 from restrain by membrane-bound β-catenin to promote collagen fibril remodeling in obesity-related neoplasms

The paracrine actions of adipokine plasminogen activator inhibitor-1 (PAI-1) are implicated in obesity-associated tumorigenesis. Here, we show that PAI-1 mediates extracellular matrix (ECM) signaling via epigenetic repression of DKK1 in endometrial epithelial cells (EECs). While the loss of DKK1 is known to increase β-catenin accumulation for WNT signaling activation, this epigenetic repression causes β-catenin release from transmembrane integrins. Furthermore, PAI-1 elicits the disengagement of TIMP2 and SPARC from integrin-β1 on the cell surface, lifting an integrin-β1-ECM signaling constraint. The heightened interaction of integrin-β1 with type 1 collagen (COL1) remodels extracellular fibrillar structures in the ECM. Consequently, the enhanced nanomechanical stiffness of this microenvironment is conducive to EEC motility and neoplastic transformation. The formation of extensively branched COL1 fibrils is also observed in endometrial tumors of patients with obesity. The findings highlight PAI-1 as a contributor to enhanced integrin-COL1 engagement and extensive ECM remodeling during obesity-associated neoplastic development.

Pharmacological depletion of RNA splicing factor RBM39 by indisulam synergizes with PARP inhibitors in high-grade serous ovarian carcinoma

Ovarian high-grade serous carcinoma (HGSC) is the most common subtype of ovarian cancer with limited therapeutic options and a poor prognosis. In recent years, poly-ADP ribose polymerase (PARP) inhibitors have demonstrated significant clinical benefits, especially in patients with BRCA1/2 mutations. However, acquired drug resistance and relapse is a major challenge. Indisulam (E7070) has been identified as a molecular glue that brings together splicing factor RBM39 and DCAF15 E3 ubiquitin ligase resulting in polyubiquitination, degradation, and subsequent RNA splicing defects. In this work, we demonstrate that the loss of RBM39 induces splicing defects in key DNA damage repair genes in ovarian cancer, leading to increased sensitivity to cisplatin and various PARP inhibitors. The addition of indisulam also improved olaparib response in mice bearing PARP inhibitor-resistant tumors. These findings demonstrate that combining RBM39 degraders and PARP inhibitors is a promising therapeutic approach to improve PARP inhibitor response in ovarian HGSC.

Human iPSC-derived fallopian tube organoids with BRCA1 mutation recapitulate early-stage carcinogenesis

Germline pathogenic mutations in BReast CAncer (BRCA1) genes are thought to drive normal fallopian tube epithelial (FTE) cell transformation to high-grade serous ovarian cancer. No human models capture the sequence of events for disease initiation and progression. Here, we generate induced pluripotent stem cells (iPSCs) from healthy individuals and young ovarian cancer patients with germline pathogenic BRCA1 mutations (BRCA1

RNA-binding protein FXR1 drives cMYC translation by recruiting eIF4F complex to the translation start site

Fragile X-related protein-1 (FXR1) gene is highly amplified in patients with ovarian cancer, and this amplification is associated with increased expression of both FXR1 mRNA and protein. FXR1 expression directly associates with the survival and proliferation of cancer cells. Surface sensing of translation (SUnSET) assay demonstrates that FXR1 enhances the overall translation in cancer cells. Reverse-phase protein array (RPPA) reveals that cMYC is the key target of FXR1. Mechanistically, FXR1 binds to the AU-rich elements (ARE) present within the 3' untranslated region (3'UTR) of cMYC and stabilizes its expression. In addition, the RGG domain in FXR1 interacts with eIF4A1 and eIF4E proteins. These two interactions of FXR1 result in the circularization of cMYC mRNA and facilitate the recruitment of eukaryotic translation initiation factors to the translation start site. In brief, we uncover a mechanism by which FXR1 promotes cMYC levels in cancer cells.

Akkermansia supplementation reverses the tumor-promoting effect of the fecal microbiota transplantation in ovarian cancer

Ovarian cancer (OC) remains a clinical challenge for its difficulty in early diagnosis and insensitivity to treatments. Gut microbiota modulate multiple carcinoma progression through immunoregulation. The relationship between OC and gut microbiota has not been fully characterized. We find that the feces of patients with OC demonstrate different characteristics from benign controls. After fecal microbiota transplantation (FMT) from patients with OC into OC-bearing mice, the tumor development accelerates. Further, an Akkermansia supplementation with FMT significantly suppresses OC progression in mice. RNA sequencing of tumors shows that T cell activation pathways are upregulated after Akkermansia supplementation with FMT. Moreover, acetate accumulation accompanies Akkermansia abundance elevation, which is associated with enhanced interferon γ (IFNγ) secretion of CD8

Endothelin-1 drives invadopodia and interaction with mesothelial cells through ILK

Cancer cells use actin-based membrane protrusions, invadopodia, to degrade stroma and invade. In serous ovarian cancer (SOC), the endothelin A receptor (ET

Most Commonly Mutated Genes in High-Grade Serous Ovarian Carcinoma Are Nonessential for Ovarian Surface Epithelial Stem Cell Transformation

High-grade serous ovarian carcinoma (HGSOC) is the fifth leading cause of cancer-related deaths of women in the United States. Disease-associated mutations have been identified by the Cancer Genome Atlas Research Network. However, aside from mutations in TP53 or the RB1 pathway that are common in HGSOC, the contributions of mutation combinations are unclear. Here, we report CRISPR mutagenesis of 20 putative HGSOC driver genes to identify combinatorial disruptions of genes that transform either ovarian surface epithelium stem cells (OSE-SCs) or non-stem cells (OSE-NSs). Our results support the OSE-SC theory of HGSOC initiation and suggest that most commonly mutated genes in HGSOC have no effect on OSE-SC transformation initiation. Our results indicate that disruption of TP53 and PTEN, combined with RB1 disruption, constitutes a core set of mutations driving efficient transformation in vitro. The combined data may contribute to more accurate modeling of HGSOC development.

Glycosphingolipids are mediators of cancer plasticity through independent signaling pathways

The molecular repertoire promoting cancer cell plasticity is not fully elucidated. Here, we propose that glycosphingolipids (GSLs), specifically the globo and ganglio series, correlate and promote the transition between epithelial and mesenchymal cells. The epithelial character of ovarian cancer remains stable throughout disease progression, and spatial glycosphingolipidomics reveals elevated globosides in the tumor compartment compared with the ganglioside-rich stroma. CRISPR-Cas9 knockin mediated truncation of endogenous E-cadherin induces epithelial-to-mesenchymal transition (EMT) and decreases globosides. The transcriptomics analysis identifies the ganglioside-synthesizing enzyme ST8SIA1 to be consistently elevated in mesenchymal-like samples, predicting poor outcome. Subsequent deletion of ST8SIA1 induces epithelial cell features through mTOR

G6PD inhibition sensitizes ovarian cancer cells to oxidative stress in the metastatic omental microenvironment

Ovarian cancer (OC) is the most lethal gynecological malignancy, with aggressive metastatic disease responsible for the majority of OC-related deaths. In particular, OC tumors preferentially metastasize to and proliferate rapidly in the omentum. Here, we show that metastatic OC cells experience increased oxidative stress in the omental microenvironment. Metabolic reprogramming, including upregulation of the pentose phosphate pathway (PPP), a key cellular redox homeostasis mechanism, allows OC cells to compensate for this challenge. Inhibition of glucose-6-phosphate dehydrogenase (G6PD), the rate-limiting enzyme of the PPP, reduces tumor burden in pre-clinical models of OC, suggesting that this adaptive metabolic dependency is important for OC omental metastasis.

Carcinoma-associated mesenchymal stem cells promote ovarian cancer heterogeneity and metastasis through mitochondrial transfer

Ovarian cancer is characterized by early metastatic spread. This study demonstrates that carcinoma-associated mesenchymal stromal cells (CA-MSCs) enhance metastasis by increasing tumor cell heterogeneity through mitochondrial donation. CA-MSC mitochondrial donation preferentially occurs in ovarian cancer cells with low levels of mitochondria ("mito poor"). CA-MSC mitochondrial donation rescues the phenotype of mito poor cells, restoring their proliferative capacity, resistance to chemotherapy, and cellular respiration. Receipt of CA-MSC-derived mitochondria induces tumor cell transcriptional changes leading to the secretion of ANGPTL3, which enhances the proliferation of tumor cells without CA-MSC mitochondria, thus amplifying the impact of mitochondrial transfer. Donated CA-MSC mitochondrial DNA persisted in recipient tumor cells for at least 14 days. CA-MSC mitochondrial donation occurs in vivo, enhancing tumor cell heterogeneity and decreasing mouse survival. Collectively, this work identifies CA-MSC mitochondrial transfer as a critical mediator of ovarian cancer cell survival, heterogeneity, and metastasis and presents a unique therapeutic target in ovarian cancer.