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Ovarian clear cell carcinoma (OCCC) is a deadly and treatment-resistant cancer, which arises within the unique microenvironment of endometriosis. In this study, we identified a subset of endometriosis-derived mesenchymal stem cells (enMSC) characterized by loss of CD10 expression that specifically support OCCC growth. RNA sequencing identified alterations in iron export in CD10-negative enMSCs and reciprocal changes in metal transport in cocultured OCCC cells. CD10-negative enMSCs exhibited elevated expression of iron export proteins hephaestin and ferroportin and donate iron to associated OCCCs, functionally increasing the levels of labile intracellular iron. Iron is necessary for OCCC growth, and CD10-negative enMSCs prevented the growth inhibitory effects of iron chelation. In addition, enMSC-mediated increases in OCCC iron resulted in a unique sensitivity to ferroptosis. In vitro and in vivo, treatment with the ferroptosis inducer erastin resulted in significant death of cancer cells grown with CD10-negative enMSCs. Collectively, this work describes a novel mechanism of stromal-mediated tumor support via iron donation. This work also defines an important role of endometriosis-associated MSCs in supporting OCCC growth and identifies a critical therapeutic vulnerability of OCCC to ferroptosis based on stromal phenotype. Significance: Endometriosis-derived mesenchymal stem cells support ovarian clear cell carcinoma via iron donation necessary for cancer growth, which also confers sensitivity to ferroptosis-inducing therapy.

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Ovarian clear cell carcinoma (OCCC) is a deadly and treatment-resistant cancer, which arises within the unique microenvironment of endometriosis. In this study, we identified a subset of endometriosis-derived mesenchymal stem cells (enMSC) characterized by loss of CD10 expression that specifically support OCCC growth. RNA sequencing identified alterations in iron export in CD10-negative enMSCs and reciprocal changes in metal transport in cocultured OCCC cells. CD10-negative enMSCs exhibited elevated expression of iron export proteins hephaestin and ferroportin and donate iron to associated OCCCs, functionally increasing the levels of labile intracellular iron. Iron is necessary for OCCC growth, and CD10-negative enMSCs prevented the growth inhibitory effects of iron chelation. In addition, enMSC-mediated increases in OCCC iron resulted in a unique sensitivity to ferroptosis. In vitro and in vivo, treatment with the ferroptosis inducer erastin resulted in significant death of cancer cells grown with CD10-negative enMSCs. Collectively, this work describes a novel mechanism of stromal-mediated tumor support via iron donation. This work also defines an important role of endometriosis-associated MSCs in supporting OCCC growth and identifies a critical therapeutic vulnerability of OCCC to ferroptosis based on stromal phenotype.Significance:Endometriosis-derived mesenchymal stem cells support ovarian clear cell carcinoma via iron donation necessary for cancer growth, which also confers sensitivity to ferroptosis-inducing therapy.

Ovarian cancer is characterized by early, diffuse metastatic spread with most women presenting with extensive abdominal metastasis at the time of diagnosis. Prior work demonstrated carcinoma-associated mesenchymal stem cells (CA-MSCs) enhance ovarian cancer metastasis through a process of direct cellular interaction and formation of heterocellular CA-MSC and tumor cell complexes. In this study, we demonstrated that CA-MSCs enhance metastasis by increasing tumor cell heterogeneity through mitochondrial donation. We showed that CA-MSCs directly interacted with ovarian cancer cells via tunneling nanotubules (TNTs), and CA-MSCs used these TNTs to transfer live mitochondria to adjacent ovarian cancer cells. This mitochondrial donation preferentially occurred with ovarian cancer cells that had the lowest mitochondrial mass, as quantified using live, actively respiring mitochondrial labeling. These ‘mito poor’ cancer cells demonstrated decreased proliferation, increased sensitivity to chemotherapy, and decreased oxidative phosphorylation compared to ‘mito rich’ cancer cells. CA-MSCs rescued the phenotypes of mito poor cancer cells, restoring their proliferative capacity, increasing chemotherapy resistance, and increasing oxidative phosphorylation. We validated these findings in a fully autologous system using CA-MSCs and cancer cells derived from the same patient to prevent confounding effects of cellular response to foreign organelle/DNA. Using a knockdown of the mitochondrial motor protein, MIRO1, in CA-MSCs, we demonstrated that mitochondrial transfer is necessary for the CA-MSC-mediated rescue of ‘mito poor’ cancer cells. Mitochondria of CA-MSC origin persisted in tumor cells over multiple passages. Importantly, CA-MSC mitochondrial donation occurred in vivo, significantly enhanced tumor cell heterogeneity and decreased survival in an orthotopic ovarian cancer mouse model. Collectively, this work identified CA-MSC mitochondrial transfer as a critical mediator of ovarian cancer cell survival, heterogeneity, and metastasis, and blocking CA-MSC mitochondrial transfer represents a unique therapeutic target in ovarian cancer.

Ovarian cancer is characterized by early, diffuse metastasis with 70% of women having metastatic disease at the time of diagnosis. While elegant transgenic mouse models of ovarian cancer exist, these mice are expensive and take a long time to develop tumors. Intraperitoneal injection xenograft models lack human stroma and do not accurately model ovarian cancer metastasis. Even patient derived xenografts (PDX) do not fully recapitulate the human stromal microenvironment as serial PDX passages demonstrate significant loss of human stroma. The ability to easily model human ovarian cancer within a physiologically relevant stromal microenvironment is an unmet need. Here, the protocol presents an orthotopic ovarian cancer mouse model using human ovarian cancer cells combined with patient-derived carcinoma-associated mesenchymal stem cells (CA-MSCs). CA-MSCs are stromal progenitor cells, which drive the formation of the stromal microenvironment and support ovarian cancer growth and metastasis. This model develops early and diffuses metastasis mimicking clinical presentation. In this model, luciferase expressing ovarian cancer cells are mixed in a 1:1 ratio with CA-MSCs and injected into the ovarian bursa of NSG mice. Tumor growth and metastasis are followed serially over time using bioluminescence imaging. The resulting tumors grow aggressively and form abdominal metastases by 14 days post injection. Mice experienced significant decreases in body weight as a marker of systemic illness and increased disease burden. By day 30 post injection, mice met endpoint criteria of >10% body weight loss and necropsy confirmed intra-abdominal metastasis in 100% of mice and 60%-80% lung and parenchymal liver metastasis. Collectively, orthotopic engraftment of ovarian cancer cells and stroma cells generates tumors that closely mimic the early and diffuse metastatic behavior of human ovarian cancer. Furthermore, this model provides a tool to study the role of ovarian cancer cell: stroma cell interactions in metastatic progression.

SummaryOvarian cancer develops early intra-peritoneal metastasis establishing a supportive tumor microenvironment (TME) through reprogramming normal mesenchymal stem cells into carcinoma-associated mesenchymal stem cells (CA-MSCs). CA-MSCs are the stromal stem cell of the TME, supporting cancer growth, increasing desmoplasia, angiogenesis and chemotherapy resistance. We demonstrate epigenetic reprogramming drives CA-MSC formation via enhancer-enriched DNA hypermethylation, altered chromatin accessibility and differential histone modifications inducing a partial mesenchymal to epithelial transition (MET) increasing adhesion to tumor cells. Direct CA-MSC:tumor cell interactions, confirmed in patient ascites, facilitate ovarian cancer metastasis through co-migration. WT1, a developmental mediator of MET, and EZH2, mediate CA-MSC epigenetic reprogramming.WT1overexpression induces CA-MSC conversion whileWT1knock-down, along with EZH2 inhibition, blocks CA-MSC formation. EZH2 inhibition subsequently decreases intra-abdominal metastasis.SignificanceThis work presents a new paradigm of stromal reprogramming involving a partial mesenchymal to epithelial transition. Rather than a classic tumor cell epithelial to mesenchymal transition, metastasis relies on epigenetic rewiring of a CA-MSC allowing enhanced tumor cell binding and co-migration with tumor cells to form metastasis. Indeed, CA-MSCs in complex with tumor cells are abundant in patient ascites. Reversion of CA-MSCs to normal MSCs is observed in patients achieving complete response with neoadjuvant therapy. Identification of WT1 and EZH2 as mediators of the epigenetic reprogramming of CA-MSCs present potential targets to block the formation of CA-MSCs thus disrupting the TME and limiting ovarian cancer metastasis.

The interactions between tumor cells and the non-malignant stromal and immune cells that make up the tumor microenvironment (TME) are critical to the pathophysiology of cancer. Mesenchymal stem cells (MSCs) are multipotent stromal stem cells found within most cancers and play a critical role influencing the formation and function of the TME. MSCs have been reported to support tumor growth through a variety of mechanisms including (i) differentiation into other pro-tumorigenic stromal components, (ii) suppression of the immune response, (iii) promotion of angiogenesis, (iv) enhancement of an epithelial-mesenchymal transition (EMT), (v) enrichment of cancer stem-like cells (CSC), (vi) increase in tumor cell survival, and (vii) promotion of tumor metastasis. In contrast, MSCs have also been reported to have antitumorigenic functions including (i) enhancement of the immune response, (ii) inhibition of angiogenesis, (iii) regulation of cellular signaling, and (iv) induction of tumor cell apoptosis. Although literature supporting both arguments exists, most studies point to MSCs acting in a cancer supporting role within the confines of the TME. Tumor-suppressive effects are observed when MSCs are used in higher ratios to tumor cells. Additionally, MSC function appears to be tissue type dependent and may rely on cancer education to reprogram a naïve MSC with antitumor effects into a cancer-educated or cancer-associated MSC (CA-MSC) which develops pro-tumorigenic function. Further work is required to delineate the complex crosstalk between MSCs and other components of the TME to accurately assess the impact of MSCs on cancer initiation, growth, and spread.

The interactions between tumor cells and the non-malignant stromal and immune cells that make up the tumor microenvironment (TME) are critical to the pathophysiology of cancer. Mesenchymal stem cells (MSCs) are multipotent stromal stem cells found within most cancers and play a critical role influencing the formation and function of the TME. MSCs have been reported to support tumor growth through a variety of mechanisms including (i) differentiation into other pro-tumorigenic stromal components, (ii) suppression of the immune response, (iii) promotion of angiogenesis, (iv) enhancement of an epithelial-mesenchymal transition (EMT), (v) enrichment of cancer stem-like cells (CSC), (vi) increase in tumor cell survival, and (vii) promotion of tumor metastasis. In contrast, MSCs have also been reported to have antitumorigenic functions including (i) enhancement of the immune response, (ii) inhibition of angiogenesis, (iii) regulation of cellular signaling, and (iv) induction of tumor cell apoptosis. Although literature supporting both arguments exists, most studies point to MSCs acting in a cancer supporting role within the confines of the TME. Tumor-suppressive effects are observed when MSCs are used in higher ratios to tumor cells. Additionally, MSC function appears to be tissue type dependent and may rely on cancer education to reprogram a naive MSC with antitumor effects into a cancer-educated or cancer-associated MSC (CA-MSC) which develops pro-tumorigenic function. Further work is required to delineate the complex crosstalk between MSCs and other components of the TME to accurately assess the impact of MSCs on cancer initiation, growth, and spread.

Carcinoma-associated mesenchymal stem cells (CA-MSCs) are critical stromal progenitor cells within the tumor microenvironment (TME). We previously demonstrated that CA-MSCs differentially express bone morphogenetic protein family members, promote tumor cell growth, increase cancer “stemness,” and chemotherapy resistance. Here, we use RNA sequencing of normal omental MSCs and ovarian CA-MSCs to demonstrate global changes in CA-MSC gene expression. Using these expression profiles, we create a unique predictive algorithm to classify CA-MSCs. Our classifier accurately distinguishes normal omental, ovary, and bone marrow MSCs from ovarian cancer CA-MSCs. Suggesting broad applicability, the model correctly classifies pancreatic and endometrial cancer CA-MSCs and distinguishes cancer associated fibroblasts from CA-MSCs. Using this classifier, we definitively demonstrate ovarian CA-MSCs arise from tumor mediated reprograming of local tissue MSCs. Although cancer cells alone cannot induce a CA-MSC phenotype, the in vivo ovarian TME can reprogram omental or ovary MSCs to protumorigenic CA-MSCs (classifier score of >0.96). In vitro studies suggest that both tumor secreted factors and hypoxia are critical to induce the CA-MSC phenotype. Interestingly, although the breast cancer TME can reprogram bone marrow MSCs into CA-MSCs, the ovarian TME cannot, demonstrating for the first time that tumor mediated CA-MSC conversion is tissue and cancer type dependent. Together these findings (a) provide a critical tool to define CA-MSCs and (b) highlight cancer cell influence on distinct normal tissues providing powerful insights into the mechanisms underlying cancer specific metastatic niche formation. Stem Cells 2019;37:257–269

Ovarian cancer is the most deadly gynecologic cancer largely due to early, diffuse metastatic spread. The ovarian cancer tumor microenvironment (TME) significantly impacts the metastatic capacity of ovarian cancer. We recently demonstrated that a critical stromal progenitor cell within the TME, termed carcinoma-associated mesenchymal stem/stromal cells (CA-MSCs), dramatically enhance the metastasis of ovarian cancer cells. CA-MSCs directly bind to cancer cells forming heterocellular units which co-metastasize. The goal of this work is to understand the mechanism driving CA-MSC-mediated ovarian cancer metastasis. We hypothesize that CA-MSCs increase the metastatic potential of ovarian cancer cells during the process of co-metastasis. Ovarian cancer uses both hematogenous and transcoelomic routes of metastasis therefore we used two ovarian cancer murine models to evaluate both modes of metastasis. We used a genomic barcode system to assess the clonal patterns of metastasis. Ovarian cancer cells with one unique barcode per cell were grown with or without CA-MSCs. Sites of metastasis were quantified and cancer cells were harvested from each site. Barcodes were sequenced to determined clonal representation at each metastatic site. We found CA-MSCs increased the number of metastatic sites in both the hematogenous and transcoelomic mouse models. Interestingly, CA-MSCs also increased the clonal heterogeneity at all metastatic sites (2.5 - 5 fold increase in blood, lung and liver). To investigate the mechanism enabling CA-MSC-mediated enhancement of clonal heterogeneity, we studied primary patient derived CA-MSCs and ovarian cancer cells. The process of metastasis poses unique metabolic stresses on cancer cells and normal mesenchymal stem cells are known to donate mitochondria to damaged epithelial cells. We therefore assessed if CA-MSCs use a similar mechanism to enhance survival of metastatic cancer cells. We stably labeled CA-MSC mitochondria with COX8-GFP via lentiviral transduction to enable the visualization and quantification of potential mitochondrial transfer. We demonstrate that CA-MSCs actively transfer mitochondria to cancer cells. This transfer is dependent on the physical interaction of CA-MSCs with cancer cells and is increased 2-4 fold when CA-MSCs and cancer cells form metastatic heterocellular units under non-adherent conditions. We tested the functional consequences of CA-MSC to cancer cell mitochondrial transfer and demonstrate cancer cells which receive mitochondria have increased chemotherapy resistance, increased sphere forming capacity and stem-like properties. Collectively, our results indicate CA-MSCs are a significant driver of ovarian cancer metastasis by forming heterocellular metastatic units which enable CA-MSC to cancer cell mitochondrial transfer. This leads to increased cancer cell survival and tumor initiation properties and represents a novel method of maintaining cancer cell heterogeneity during metastasis. Citation Format: Catherine A. Pressimone, Leonard G. Frisbie, Alexander Pearson, Lan G. Coffman. Carcinoma-associated mesenchymal stem/stromal cells enhance ovarian cancer metastasis and increase cancer cell clonal heterogeneity through direct mitochondrial transfer [abstract]. In: Proceedings of the AACR Virtual Special Conference on the Evolving Tumor Microenvironment in Cancer Progression: Mechanisms and Emerging Therapeutic Opportunities; in association with the Tumor Microenvironment (TME) Working Group; 2021 Jan 11-12. Philadelphia (PA): AACR; Cancer Res 2021;81(5 Suppl):Abstract nr PO021.

Introduction: Ovarian cancer is the deadliest gynecologic malignancy in the USA, and many patients incur diffuse metastases before diagnosis. In order to improve prognoses and reveal novel therapeutics, we must understand the mechanisms of ovarian tumor metastasis. Previous studies demonstrate that interactions between ovarian tumor cells (TCs) and Carcinoma-Associated Mesenchymal Stem Cells (CA-MSCs) are important drivers of metastasis. In addition, preliminary data exposes mitochondrial transfer between CA-MSCs and ovarian TCs as a potential contributor to cancer stem cell (CSC) development. Since CSCs contribute to cancer persistence, these new findings motivate current research exploring whether mitochondrial gain enhances TC metastatic fitness and CSC formation. Methods: CA-MSC mitochondria were transiently stained with MitoTracker Green, and GFP+ CA-MSCs were co-cultured with RFP+ TCs for various durations. Immunofluorescence visualized mitochondrial transfer following co-culture. Mitochondrial transfer from CA-MSCs to TCs was quantified with flow cytometry, separating GFP+ TCs from GFP-TCs. To examine whether mitochondrial transfer requires direct cell-cell contact, CA-MSC were separated from TCs with 0.4um Transwell membranes during co-culture. To elucidate functional phenotypic differences associated with mitochondrial gain, TCs and GFP+ CA-MSCs were directly co-cultured and TCs that acquired GFP+ mitochondria were sorted out via FACS. The TC populations that did or did not gain mitochondria were compared for cell proliferation rates, chemotherapy resistance, and stem-like phenotypes via cell viability and sphere formation assays. Results: We obtained fluorescent images illustrating that CA-MSC mitochondria transferred to adjacent TCs within 4 hours of direct co-culture. When CA-MSCs and TCs were indirectly co-cultured using Transwell membranes, mitochondria were not transferred as robustly as they were via direct co-culture. Interestingly, TCs that gained mitochondria appeared to grow more slowly and were less resistant to cisplatin treatment when compared to TCs that did not gain mitochondria. However, the TCs that gained mitochondria appeared to form more spheres and thus demonstrate a more CSC-like phenotype than TCs that did not gain mitochondria. Conclusions: These data suggest that ovarian TCs receive mitochondria from adjacent CA-MSCs via direct interaction. The TCs that obtain mitochondria may functionally diverge from TCs that don't gain mitochondria, exhibiting phenotypes consistent with CSCs. Future experiments will further delineate the functional difference and translational significance attributable to TC mitochondrial gain, potentially demonstrating that mitochondrial transfer in ovarian tumors can be therapeutically exploited to reduce metastases. Citation Format: Catherine Pressimone, Huda Issa Atiya, Leonard Frisbie, Lan Coffman. Mitochondrial transfer in the ovarian tumor microenvironment contributes to tumor stem cell phenotypes [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 1446.

Ovarian cancer, the most deadly gynecologic cancer, resides within a complex tumor microenvironment that supports cancer growth, survival, and spread. Carcinoma-associated mesenchymal stem cells (CA-MSCs) are stromal stem cells within the ovarian TME that strongly support tumor growth promoting cancer initiation, increasing the cancer stem cell pool and enhancing chemotherapy resistance. We recently demonstrated that ovarian cancer stimulation converts normal tissue-derived MSCs into CA-MSCs. The goal of this work is to investigate the mechanism of cancer-mediated CA-MSC formation. We hypothesize that ovarian cancer stimulation induces the epigenetic reprogramming of MSCs creating CA-MSCs. EPIC array DNA methylation analysis, histone mass spectrometry, and Assay for Transposase-Accessible Chromatin using sequencing (ATACseq) were performed on CA-MSCs derived from ovarian cancer patient samples and normal MSCs derived from benign fallopian tube and omentum. We demonstrate that CA-MSCs have large-scale DNA methylation differences vs. normal tissue MSCs. This DNA methylation pattern is distinct from cancer cells; however, the CA-MSC methylation pattern is represented in the DNA methylation pattern of bulk tumor from clear-cell, endometrioid, and high-grade serous cancers, indicating the importance of stromal contribution to bulk tumor analysis. DNA methylation changes were acquired during the conversion of a normal MSC into a CA-MSC and retained after CA-MSC adipocyte and fibroblast differentiation. Histone mass spectrometry and ATACseq demonstrated enrichment in repressive histone marks and overall decreased ATACseq peaks consistent with altered chromosome accessibility. Integration of DNA methylation, ATACseq, and RNAseq data demonstrates strong correlation between epigenetic changes and transcriptomic changes. Interestingly, cell:cell adhesion pathways were consistently upregulated consistent with a partial mesenchymal-to-epithelial transition (MET). This partial MET engenders CA-MSCs with enhanced tumor cell-binding capacity. Direct tumor cell:CA-MSC binding enhanced ovarian cancer migration, extravasation, and nonadherent survival in vitro and increased ovarian cancer metastasis in vivo. This partial MET may be mediated via WT1, a mediator of MET during development, as knockdown of WT1 prevented the formation of a CA-MSCs while overexpression of WT1 enhances the formation of a CA-MSC. Collectively, we demonstrate ovarian cancer stimulates the epigenetic reprogramming of normal MSCs into CA-MSCs, thus enhancing direct tumor cell binding, which facilitates ovarian cancer metastasis. Further investigation is needed to identify the molecular drivers of this reprogramming to allow the disruption of the protumorigenic TME, which may represent a powerful approach to the treatment of ovarian cancer. Citation Format: Leonard Frisbie, HuiHui Fan, Thomas Pisanic, Hui Shen, Lan Coffman. Epigenetic reprogramming of mesenchymal stem cells by ovarian carcinoma to facilitate metastasis [abstract]. In: Proceedings of the AACR Special Conference on Advances in Ovarian Cancer Research; 2019 Sep 13-16, 2019; Atlanta, GA. Philadelphia (PA): AACR; Clin Cancer Res 2020;26(13_Suppl):Abstract nr B55.

e18072 Background: Carcinoma-associated mesenchymal stem cells (CA-MSCs) are mesenchymal stem cells (MSCs) within the tumor microenvironment (TME). We demonstrated that CA-MSCs promote ovarian cancer chemotherapy resistance through paracrine signaling with ovarian cancer cells, interact with ovarian cancer cells to form mixed-cellularity complexes which enhance metastasis, and arise from cancer cell and TME reprogramming of normal tissue MSCs. To identify mediators of the CA-MSC:tumor cell interaction, we investigated the role of tetraspanins, which are membrane-spanning proteins that have been implicated in cancer development and metastasis by influencing cell adhesion and cell-cell interactions. The tetraspanins CD9, CD81, CD151, and CD63 were identified as potential mediators of cell surface interactions between ovarian cancer cells and CA-MSCs through homo- and heterodimerization. Methods: Td-labeled OVCAR3 cancer cells were co-cultured with 3 patient-derived MSC (derived from normal tissue) and 3 patient-derived CA-MSC (derived from malignant tissue) cell lines. Flow cytometry was performed to measure surface protein expression of the tetraspanins CD9, CD81, CD151, and CD63, and was compared to control cells (not co-cultured) and their median fluorescence intensity (MFI). We next separated OVCAR3 cells co-cultured with MSCs or CA-MSCs using fluorescent activated cell sorting (FACS) based on Td expression. Following FACS, tetraspanin expression in OVCAR3 cells, MSCs, and CA-MSCs was assessed via qRT-PCR and western blotting, and compared to cell lines that were not co-cultured. Results: Flow cytometric analysis revealed an increase in MFI of CD9 and CD151 in co-cultured OVCAR3 cells, as well as CD81 and CD63 in co-cultured CA-MSCs and MSCs when compared to non-co-cultured matched cells. Increased RNA expression of CD9, CD151, and CD63 was seen in OVCAR3 cells that were co-cultured with CA-MSCs or MSCs when compared to non-co-cultured OVCAR3 cells. Increased RNA expression of CD81 and CD63 was noted in both co-cultured CA-MSCs and MSCs compared to non-co-cultured cells. Lastly, western blotting demonstrated increased protein expression of CD9 and CD151 in co-cultured OVCAR3 cells, as well as CD81 and CD63 in co-cultured CA-MSCs and MSCs compared to non-co-cultured matched cells. Conclusions: These results indicate that direct interactions between OVCAR3 cells and CA-MSCs or MSCs lead to overexpression of specific tetraspanins at the RNA and protein levels, implying that they may be facilitating tumor cell:CA-MSC and tumor cell:MSC binding.

SUMMARY 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., Graphical Abstract, In Brief Fan et al. demonstrate that ovarian cancer reprograms the epigenome of stromal cells, inducing a mesenchymal-to-epithelial transition (MET) to form carcinoma-associated mesenchymal stem cells (CA-MSCs). This MET, mediated by WT1 and EZH2, enables CA-MSC:cancer cell co-metastasis. EZH2 inhibition decreases CA-MSC-mediated metastasis, presenting a potential therapeutic opportunity in ovarian cancer.