FOXM1 inhibitor, RCM-1, enhances venetoclax mediated apoptosis through downregulation of ATP2B4 in rhabdomyosarcoma

Rhabdomyosarcoma (RMS) is the most common type of soft tissue sarcoma in children. Intensifying chemotherapy has failed to improve patient survival for metastatic or relapsed RMS and RMS survivors often suffer from significant long-term toxicities. More efficient and less toxic new therapies are critically needed. RMS expresses high levels of antiapoptotic protein Bcl-2 and an oncogenic transcription factor Forkhead box protein M1 (FOXM1), which is also known to inhibit tumor cell apoptosis. The present study used a combination therapy of a recently developed non-toxic FOXM1 inhibitor, RCM-1 and the FDA-approved Bcl2 inhibitor, venetoclax, which is not effective as a mono therapy in solid tumors. Compared with venetoclax alone, the combination therapy efficiently inhibited RMS growth in the animal model by decreasing tumor cell proliferation and inducing tumor cell apoptosis. RNA-sequencing analysis demonstrated that the combination therapy uniquely decreased expression of ATPase Plasma Membrane Ca²⁺ Transporting 4 (ATP2B4), a plasma membrane calcium channel that is highly expressed in RMS compared with normal muscle cells. RCM-1, but not venetoclax treatment, inhibited ATP2B4 and enhanced the sensitivity of RMS cells to apoptosis. Knockdown of ATP2B4 decreased RMS tumor cell proliferation, migration and colony formation in vitro. Furthermore, knockdown of ATP2B4 increased tumor cell apoptosis, while overexpression of ATP2B4 decreased tumor cell apoptosis in vitro. In the animal model of RMS, depletion of ATP2B4 decreased tumor growth. In summary, combining RCM-1 with venetoclax sensitized RMS cells to apoptosis by decreasing ATP2B4. This made ATP2B4 a promising therapeutic target for RMS and provides a rationale for exploring this combination in early-stage clinical trials.