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Abstract Cancer progression is enabled by the accumulation of somatic mutations and genomic aberrations, leading to metastasis, treatment resistance, and poor clinical outcomes. The analysis of cancer genomes has identified APOBEC3A (A3A) cytidine deaminase activity as a cause of somatic mutagenesis in over 20 tumor types. In particular, A3A mutational signatures are found in a large proportion of ovarian tumors, ranging from 2-26% depending on subtype. Furthermore, genomic analyses of ovarian tumors suggest that A3A mutagenesis occurred during early metastatic progression, indicating the potential for A3A activity to promote clonal evolution. In non-ovarian tumors, A3A activity is associated with metastatic progression through the identification of A3A mutational patterns enriched in or unique to metastatic sites. Therefore, we hypothesized that aberrant A3A activity on OC genomes would increase mutagenesis, promote clonal diversity, and enable tumor progression. To examine the impact of aberrant A3A activity during OC progression, we developed a model system of inducible A3A expression in OC cell lines constitutively expressing CBR-GFP. We treated cells for 45 days with doxycycline to induce A3A expression (long-term, LT-A3A). LT-A3A cells demonstrated increased colony formation relative to NT cells, indicating improved cell survival. We assessed cell migration by scratch-wound assays and found that LT-A3A cells exhibited accelerated wound closure rates compared to NT controls, despite similar proliferation rates. In addition, cellular invasion was assessed by spheroid Matrigel invasion assays in which the invasive distance was significantly greater in LT-A3A cells relative to NT controls. By generating cell lines with a catalytically inactive doxycycline-inducible A3A transgene, we determined that the observed A3A-mediated cellular phenotypes were deaminase-dependent. Next, to determine the impact of LT-A3A on tumor behavior, we engrafted LT-A3A or NT cells into the peritoneal cavity of NSG mice. Tumor burden was monitored weekly using bioluminescent imaging (BLI). At d165, the mice were sacrificed and tumor burden was assessed via rapid ex vivo BLI, total tumor weight, and count of metastatic nodules in the lungs. From this, we identified that LT-A3A promotes metastatic spread to the lungs in vivo. To assess the molecular alterations driving the phenotypic differences, we performed RNA-seq on LT-A3A and NT cell lines. Gene set enrichment analysis of differentially expressed genes identified significant overlap between the Epithelial-Mesenchymal Transition (EMT) and WNT/Beta Catenin, Hedgehog, and TGFβ signaling gene sets. These related signaling pathways converge on the promotion of metastatic cancer progression. These data suggest that aberrant A3A activity promotes the survival, migration, and invasion of OC cells by altering intracellular signaling pathways that promote EMT. Our ongoing experimental investigation will assess how long-term A3A mutagenesis impacts genomic heterogeneity and will define the mechanisms of A3A-mediated OC metastasis. Citation Format: Jessica Merritt Devenport, Abby Green. Long-term episodic APOBEC3A activity promotes metastatic ovarian cancer progression abstract. In: Proceedings of the AACR Special Conference on Ovarian Cancer; 2023 Oct 5-7; Boston, Massachusetts. Philadelphia (PA): AACR; Cancer Res 2024;84 (5 Suppl₂): Abstract nr B105.
Devenport et al. (Mon,) studied this question.