Abstract 198: Ovary to the moon: proximity ligation assay catches APoE in the act

Abstract Within different subtypes of ovarian cancer, the most common histotype is high-grade serous ovarian cancer (HGSC), which typically presents as aggressive advanced-stage disease and accounts for over 70% of all ovarian cancer deaths. The current standard of care includes primary debulking surgery followed by at least 6 cycles of platinum- and taxane-based combination chemotherapy, or neoadjuvant chemotherapy followed by interval debulking surgery and adjuvant chemotherapy. At MDACC, we have incorporated second look laparoscopy (SLL) to assess minimal residual disease (MRD) after frontline therapy. During this procedure peritoneal washings are obtained, and subsequent analysis pre- and post-treatment has demonstrated a significant difference in protein concentration between MRD+/- . Our recent investigations using multiplex immunofluorescent (mIF) and spatial transcriptomics analyses revealed that a novel cross-talk signaling network between HGSC cells (expressing LRP5 receptor) and cancer-associated fibroblasts (CAFs, expressing APoE ligand) at the stromal-tumor interface is associated with poor patient survival rates. CAFs heterogeneity may also play essential roles in modulating tumor growth, but the molecular mechanisms by which CAFs modulate the immune suppressive TME and HGSC patient survival rates are unclear. Preliminary data suggests this is linked to metabolism, which could foster MRD+ and analysis of MRD+ fluid has indicated increased secreted protein concentrations that may play a significant role in the upregulation of APoE-LRP5 interactions. To quantify APoE-LRP5 modulation by MRD +/- washes we utilized two human cell lines: OV90 tumor cells and ovarian CAF, which don’t express endogenous levels of APoE in vitro. CAFs were primed in media containing MRD +/- washes for 24 hours in the presence or absence of two APoE inhibiting drugs (CN-105, a modified five amino acid ApoE mimetic; and HJ6.3 anti-ApoE monoclonal antibody) prior to coculture in the Samplix X-Drop double emulsion droplets and a Naveni proximity ligation assay to detect LRP5 and ApoE interactions. Droplets were imaged by confocal microscopy to quantify interactions based on presence of puncta. In parallel, CAFs and OV90 cells were co-cultured in a tissue culture plate and their growth kinetics quantified. Priming of CAFs with MRD+ surgical fluids increased the expression of APoE by these CAFs and APoE-LRP5 interactions. We demonstrated that the inhibition of APoE signaling during the priming process through both the mimetic and antibody led to decreased expression of APoE and APoE-LRP5 interactions in co-culture. Growth kinetic analysis of OV90 cells demonstrated that the increased expression of APoE led to more aggressive tumor growth that could be inhibited with the presence of APoE-blocking drugs during chemotherapy. Citation Format: Danielle L. Stolley, Erin Seeley, Anna K. Casasent, Amir A. Jazaeri, Jared K. Burks, Sammy Ferri-Borgogno. Ovary to the moon: proximity ligation assay catches APoE in the act [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 198.