ABSTRACT Small-cell lung cancer (SCLC) is an aggressive malignancy with limited therapeutic options. The dismal prognosis in SCLC is in part associated with an upregulation of BCL-2 family anti-apoptotic proteins, including BCL-X L and MCL-1. Unfortunately, the currently available inhibitors of BCL-2 family anti-apoptotic proteins, except BCL-2 inhibitors, are not clinically relevant because of various on-target toxicities. We, therefore, aimed to develop an effective and safe strategy targeting these anti-apoptotic proteins with DT2216 (our platelet-sparing BCL-X L degrader) and AZD8055 (an mTOR inhibitor) to avoid associated on-target toxicities while synergistically optimizing tumor response. Through BH3 mimetic screening, we identified a subset of SCLC cell lines that is co-dependent on BCL-X L and MCL-1. After screening inhibitors of selected tumorigenic pathways, we found that AZD8055 selectively downregulates MCL-1 in SCLC cells and its combination with DT2216 synergistically killed BCL-X L /MCL-1 co-dependent SCLC cells, but not normal cells. Mechanistically, the combination caused BCL-X L degradation and suppression of MCL1 expression, and thus disrupted MCL-1 interaction with BIM leading to an enhanced apoptotic induction. In vivo , DT2216+AZD8055 combination significantly inhibited the growth of cell line-derived and patient-derived xenografts and reduced tumor burden accompanied with extended survival in a genetically-engineered mouse (GEM) model of SCLC without causing significant thrombocytopenia or other normal tissue injury. Thus, these preclinical findings lay a strong foundation for future clinical studies to test DT2216+mTOR inhibitor combination in a subset of SCLC patients whose tumors are co-driven by BCL-X L and MCL-1.

Co-targeting BCL-X<sub>L</sub> and MCL-1 with DT2216 and AZD8055 synergistically inhibits small-cell lung cancer growth without causing on-target toxicities in mice

Co-targeting BCL-XL and MCL-1 with DT2216 and AZD8055 synergistically inhibits small-cell lung cancer growth without causing on-target toxicities in mice