Anti‐apoptotic protein BCL‐XL as a therapeutic vulnerability in gastric cancer

Abstract

Abstract Background New therapeutic targets are needed to improve the outcomes for gastric cancer (GC) patients with advanced disease. Evasion of programmed cell death (apoptosis) is a hallmark of cancer cells and direct induction of apoptosis by targeting the pro‐survival BCL2 family proteins represents a promising therapeutic strategy for cancer treatment. Therefore, understanding the molecular mechanisms underpinning cancer cell survival could provide a molecular basis for potential therapeutic interventions. Method Here we explored the role of BCL2L1 and the encoded anti‐apoptotic BCL‐XL in GC. Using Droplet Digital PCR (ddPCR) technology to investigate the DNA amplification of BCL2L1 in GC samples and GC cell lines, the sensitivity of GC cell lines to selective BCL‐XL inhibitors A1155463 and A1331852, pan‐inhibitor ABT‐263, and VHL‐based PROTAC‐BCL‐XL was analyzed using (CellTiter‐Glo) CTG assay in vitro. Western Blot (WB) was used to detect the protein expression of BCL2 family members in GC cell lines and the manner in which PROTAC‐BCL‐XL kills GC cells. Co‐immunoprecipitation (Co‐IP) was used to investigate the mechanism of A1331852 and ABT‐263 kills GC cell lines. DDPCR, WB, and real‐time PCR (RTPCR) were used to investigate the correlation between DNA, RNA, protein levels, and drug activity. Results The functional assay showed that a subset of GC cell lines relies on BCL‐XL for survival. In gastric cancer cell lines, BCL‐XL inhibitors A1155463 and A1331852 are more sensitive than the pan BCL2 family inhibitor ABT‐263, indicating that ABT‐263 is not an optimal inhibitor of BCL‐XL. VHL‐based PROTAC‐BCL‐XL DT2216 appears to be active in GC cells. DT2216 induces apoptosis of gastric cancer cells in a time‐ and dose‐dependent manner through the proteasome pathway. Statistical analysis showed that the BCL‐XL protein level predicts the response of GC cells to BCL‐XL targeting therapy and BCL2L1 gene CNVs do not reliably predict BCL‐XL expression. Conclusion We identified BCL‐XL as a promising therapeutic target in a subset of GC cases with high levels of BCL‐XL protein expression. Functionally, we demonstrated that both selective BCL‐XL inhibitors and VHL‐based PROTAC BCL‐XL can potently kill GC cells that are reliant on BCL‐XL for survival. However, we found that BCL2L1 copy number variations (CNVs) cannot reliably predict BCL‐XL expression, but the BCL‐XL protein level serves as a useful biomarker for predicting the sensitivity of GC cells to BCL‐XL‐targeting compounds. Taken together, our study pinpointed BCL‐XL as potential druggable target for specific subsets of GC.

Keywords

• apoptosis
• BCL2L1 (BCL‐XL)
• gastric cancer (GC)
• PROTAC‐BCL‐XL
• selective BCL‐XL inhibitors