Abstract PR007: Identifying targeted therapies for muscle invasive bladder cancer via STAG2 expression
Authors: Athans Sarah R. , Withers Henry , Kazmierczak Zara , Gurova Katerina , Woloszynska Anna 
Published: May 2024

Abstract
High stromal antigen 2 (STAG2) protein expression is significantly associated with shorter overall and progression free survival of muscle invasive bladder cancer (MIBC) patients. Unfortunately, STAG2 itself is difficult to target as it functions as a transcriptional regulator and has no intrinsic enzymatic activity. Therefore, it becomes essential to identify downstream therapeutic vulnerabilities based on STAG2 expression. In this study, we accomplish this by employing a drug screen in isogenic STAG2 wild-type (WT) and knock-out (KO) MIBC cell lines, analyzing drug sensitivity of DepMap bladder cancer cell lines, and performing in vitro dose-response experiments based on our drug screen findings. To delineate changes in drug sensitivity due specifically to altered STAG2 expression, we used CRISPR-Cas9 to knock out STAG2 expression in T24 MIBC cells and generated two clonal STAG2 KO cell lines to compare to control. Then, we performed a drug screen with 312 drugs targeting oncogenic signaling pathways. We treated cells with drugs at 534 nM and read viability after 72 hours. To expand our drug screen findings to a larger group of cell lines and drugs, we further queried the DepMap database which includes drug sensitivity data for 4,868 drugs and approximately 2,000 cancer cell lines. We selected drug screened bladder cancer cell lines from DepMap and grouped them into ‘STAG2-high’ or ‘STAG2-low’ cohorts based on STAG2 mRNA expression. We then compared these two groups to identify if either group was more sensitive to drugs in the DepMap database. From these drug screen and DepMap analyses, we identified TAK-733 (MEKi), Berzosertib (ATRi), Prexasertib (CHKi), PI-103 (PI3Ki), and Rigosertib (PLKi) as candidate drugs that are differentially effective based on STAG2 expression level. We performed validation studies with these candidate drugs by treating matched STAG2 WT and KO T24 cells with each drug at concentrations ranging from 0.001 to 100 µM, then computationally modeled EC50 values in R. Cells with STAG2 KO were more sensitive to Berzosertib (T24 Control EC50: 1.78 µM, T24 KO EC50s: 1.03, 0.77 µM) compared to cells with WT STAG2 expression. Interestingly, cells with WT STAG2 expression were more sensitive to PI-103 (T24 Control EC50: 0.33 µM, T24 KO EC50s: 0.97, 1.41 µM) compared to cells with STAG2 KO. This suggests that STAG2 expressing cells may have a unique dependency on PI3K signaling, which has previously been reported in sarcoma cell lines. These findings lay the foundation for further investigation of Berzosertib as a targeted therapy for MIBC patients with mutated or low STAG2, and PI-103 for MIBC patients with high STAG2 who currently suffer worse clinical outcomes.
Citation Format: Sarah R. Athans, Henry Withers, Zara Kazmierczak, Katerina Gurova, Anna Woloszynska. Identifying targeted therapies for muscle invasive bladder cancer via STAG2 expression [abstract]. In: Proceedings of the AACR Special Conference on Bladder Cancer: Transforming the Field; 2024 May 17-20; Charlotte, NC. Philadelphia (PA): AACR; Clin Cancer Res 2024;30(10_Suppl):Abstract nr PR007.