(137d) Silica Gel Encapsulation of Ovarian Cancer Cells for Selection of Cells Exhibiting Both a Higher Propensity to Enter Quiescence and Enhanced Chemoresistance

Despite advances in standard of care treatment, 60-80% of ovarian cancer patients will exhibit disease recurrence, either due to the presence of quiescent or chemoresistant cancer cells. As chemotherapies typically target rapidly dividing cells, it is thought that quiescent cells evade treatment. However, the precise connection between chemoresistance and the ability to enter quiescence remains unclear, in part because it has been challenging to induce and maintain quiescence in vitro. A silica gel encapsulation platform that uses physical confinement to inhibit proliferation has previously shown the ability to distinguish between breast cancer cells lines with differential propensities to exhibit quiescence in vivo. In this work, we sought to apply this platform to the OVCAR-3 ovarian adenocarcinoma cell line and determine if it could be used to select for cells with a greater propensity to exit the cell cycle. Upon encapsulation in silica gels, surviving OVCAR-3 cells demonstrated hallmarks of quiescence, including a high P38:ERK activity ratio and a significant reduction in cycling marker Ki67 expression over time, with the live cell population consisting of 72.4%, 37.4%, and 11.9% Ki67-positive cells at Day 0, 3, and 5 of encapsulation, respectively. Furthermore, encapsulated cells showed downregulation of key cell cycle regulatory genes (CDK2, CCNE1, and CCND1) and upregulation of CDKN1A, which was consistent with cells becoming quiescent under hypoxic or serum starvation conditions. Upon extraction from the silica gels, the cells were able to resume proliferation and regained Ki67 expression, indicating that the growth-arrested cells were quiescent and not senescent. Interestingly, being in a proliferative state once extracted, the cells that survived a round of encapsulation demonstrated 95.8% viability after cisplatin drug treatment in the extracted state, while only 48.5% of control cells that had never been encapsulated remained viable. Furthermore, this same population of extracted cells also showed increased survival when re-encapsulated into silica gels, suggesting that this unique subset of cells is both more able to enter a quiescent state when needed and less susceptible to platinum chemotherapy. Conversely, when chemoresistant cells were generated by selecting for surviving cells after a round of cisplatin treatment, the chemoresistant cells demonstrated significantly higher viability upon encapsulation in silica gels relative to untreated cells, further demonstrating the interplay between augmented ability to enter quiescence and enhanced chemoresistance. Lastly, RNA sequencing analysis of encapsulated, extracted, and cisplatin-treated cells demonstrated that the population of cells selected for the ability to survive growth arrest in the silica gels and the chemoresistant population selected for the ability to survive cisplatin treatment shared similar regulation of genes and signaling pathways at the transcriptome level via Gene Ontology enrichment analysis and Ingenuity Pathway Analysis. These results indicate that cells with an enhanced ability to enter a quiescent state are also better able to evade chemotherapy, even if they are not quiescent at the time of treatment. This unique subset of cells is now being studied further to identify mechanistic links between the propensity for quiescence and chemoresistance to provide novel targets for improving ovarian cancer treatment.