Development of payload-resistant human ovarian cancer cell lines to better understand antibody-drug conjugate sequencing and resistance mechanisms.

Abstract

5549 Background: Within gynecologic oncology, limited data exist regarding mechanisms of resistance to antibody-drug-conjugates (ADC) or the impact on response to future ADC treatment. The purpose of this study was to develop and characterize payload-resistant human ovarian cancer cell lines, and to determine the effect of payload resistance on subsequent ADC exposure. Methods: Three commercially available human high grade serous ovarian cancer cell lines (PEA1, PEA2, and OVCAR8) were each exposed separately to three commercially available payloads with corresponding, clinically approved ADCs (deruextecan/trastuzumab deruxetecan, SN38/sacituzumab govitecan, and DM4/mirvetuximab soravtansine). Cell viability assays were performed to confirm and quantify resistance after completion of treatment for payload-resistant cell lines, and western blot analyses performed to evaluate changes in HER2, TROP2, and FOLR1 expression between payload-resistant cells and their controls. Lastly, paired payload-resistant and -sensitive cell lines were then exposed to ADCs with different payloads for 48 hours, and cell viability assays were performed. Results: The development of payload-resistant cell lines demonstrates notable variability in timelines for attaining resistance, both across the three cell lines and within individual cell lines across different payload exposures. Compared to payload-sensitive controls, payload-resistant cells displayed similar morphologic changes including increased surface area; however, demonstrated variable differences in HER2, TROP2, and FOLR1 protein expression. Additionally, different payload-resistant cell lines responded differently to subsequent ADC exposure. As a specific example, DM4-resistant PEA1 cells were more sensitive to subsequent exposure to sacituzumab govetecan compared to DM4-senstive controls at various doses (p < 0.0001, see Table). Conclusions: This study illustrates challenges in developing payload-resistant cell lines, and highlights variability in molecular changes with payload resistance. The data demonstrate subsequent ADC susceptibility in some payload-resistant cell lines, highlighting the complexities of changes that occur with drug treatment. Our data suggests that HGSOC patients that received mirvetuximab soravtansine may be more susceptible to future treatment with sacituzumab govitecan. Future work is underway to expand these findings to determine ideal sequencing of these different ADCs, and to determine other off-target effects that may influence non-ADC treatment response. Viability of DM4-resistant versus sensitive PEA1 human ovarian cancer cells after exposure to sacituzumab govitecan (SG). DM4 resistant Control Average % viability SEM Average % viability SEM p value SG 400nM 52.01 1.63 98.98 1.01 < 0.000001 SG 600nM 45.43 0.65 96.87 0.49 < 0.000001