Background <p>Tumor microenvironment is well known to have a key role in tumor development. Extracellular vesicles are capable in cell signaling transduction and important in regulation of tumor microenvironment.</p> Objective <p>To investigate whether peritoneal fluid–derived extracellular vesicles help regulate the tumor microenvironment in the malignant transformation of endometriosis.</p> Methods <p>Samples of peritoneal fluid were taken from women with benign gynecological disease, endometriosis, or endometriosis-associated ovarian cancer. Small extracellular vesicles in the samples were isolated via ultracentrifugation and characterized by western blotting, transmission electron microscopy, and nanoparticle tracking. A global microRNA (miRNA) expression profile array was used to analyze miRNA abundance in peritoneal fluid–derived small extracellular vesicles. Candidate miRNAs were quantified by reverse transcription PCR to assess their potential role in cell migration.</p> Results <p>A total of 22 miRNAs were identified from the analysis of miRNAs in peritoneal fluid–derived small extracellular vesicles from patients with endometriosis or endometriosis-associated ovarian cancer. We confirmed that each miRNA was expressed in various ovarian cell lines. The miRNA miR-302f was consistently highly expressed in both clinical specimens and ovarian cell lines from patients with endometriosis, yet expression was relatively low in specimens and cell lines from patients with endometriosis-associated ovarian cancer. A bio-functional assay revealed that miR-302f regulates cell migration. Finally, the possible target mRNAs of miR-302f were identified in the Gene Expression Omnibus database.</p> Conclusions <p>These data may provide a basis for the development of novel therapeutic strategies for endometriosis-associated ovarian cancer patients by downregulating PDGFRA abundance in cancer cells via overexpression of miR-302f.</p>