Defining the Ovarian Cancer Precancerous Landscape through Modeling Fallopian Tube Epithelium Reprogramming Driven by Extracellular Vesicles.

Serous tubal intraepithelial carcinomas (lesions) in the human fallopian tube epithelium (hFTE) are theorized to give rise to high-grade serous ovarian cancers. Small extracellular vesicles (sEV) are known to mediate key signaling in both normal and cancerous tissues, but few ex vivo systems exist for studying the impact of sEV on hFTE tissue. In this study, we present a microfluidic tissue culture platform with combined spatial transcriptomic and proteomic readouts that allows us to profile dual responses in tissue exposed to sEV "messages"-capturing both short-term transcriptomic shifts in the tissue and long-term changes in protein cargo of secreted EVs (the "reply"). Using spatial transcriptomics, we show that the short-term 1-day exposure to ovarian cancer-derived sEVs alters expression of 68 transcripts in secretory cells, the progenitor of high-grade serous ovarian cancer, notably upregulating immune-related mRNA, including CXCL family chemokines, VCAM1, and pro-inflammatory mediators (NFKB1, IL1B, and IFNA7/17). Additionally, we observed that the long-term 14-day exposure to sEVs alters the expression of seven transcripts and 25 EV cargo proteins of fallopian tube-derived EVs ("secondary release EVs") following stimulus from cancer EVs. Together, tissue transcriptomics and tissue-derived EV proteomics indicate that ovarian cancer-derived sEVs rewire target cell signaling to modify the tubal immune landscape. This study provides insights into the early molecular changes associated with the pathogenesis of ovarian cancer in its tissue of origin, providing a platform to study EV-tissue interactions and identify how sEVs drive cell signaling reprogramming in hFTE. SIGNIFICANCE: We model the fallopian tube preneoplastic landscape using a microfluidic platform to study EV-induced stress and show that cancer EVs promote immune signaling changes representing the earliest stages of ovarian cancer pathogenesis.