Figure 4.

Serial EV profiling with an HGSOC animal model. A) Study design. Oncogenic mFT cells were implanted into ovary fat mass in mice. Blood samples were collected from the animals before engraftment and up to 3 months thereafter. For each sample, EVs were collected and profiled by SAViA for nine HGSOC candidate markers. B) Survival analysis of mFT3635 (Brca1 ^−/−) and mFT3666 (Brca2 ^−/−) implanted animals. No significant difference (P = 0.989; log‐rank test) in survival was observed. The median survival was 107 (mFT3635, n = 6) and 114 days (mFT3666, n = 6). C) Immunohistochemical staining confirmed the expression of the FT epithelial (PAX8) and tumor markers (p53, WT1, STMN1) in a late‐stage (day 90) tumor. D) Longitudinal EV profiling in tumor‐bearing animals (n = 6). Nine HGSOC markers were measured in plasma EVs. The heatmap shows the z‐score of each marker. E) The expression of HGSOC markers increased after tumor initiation (day 9) and peaked 30 days after the mFT cell implant. Each data point is the average of fold changes from six animals. Data are displayed as mean ± s.d. (n = 6). F) Single EVs were imaged in plasma samples collected before the mFT cell engraftment (day 0) and during disease progression (day 30). EVs were stained for tetraspanins (CD63, CD9), PAX8 (FT epithelial marker), and CA125 marker (see Figure S8, Supporting Information for other markers). G) Tetraspanin‐positive EVs were present in both samples (day 0 and day 30) with no significant difference in numbers (P = 0.290; non‐paired, two‐sided t‐test). PAX8‐positive EVs, however, significantly increased in the tumor sample (P = 0.019; non‐paired, two‐sided t‐test). Data are displayed as mean ± s.d. (n = 15 field of views). H) EV imaging revealed that more EVs were both PAX8 and HGSOC‐marker positive in tumor samples. Data are displayed as mean ± s.d. (n = 3). The P‐values (non‐paired, two‐sided t‐test) are 0.002 (CA125), 0.0009 (VCAN), 0.014 (TNC), 0.105 (FOLR1), and 0.018 (HE4).