Repotrectinib Alone and in Combination with Immunomodulatory Drugs Exhibits Potent Anti-Multiple Myeloma Activity

Introduction: Repotrectinib (Turning Point Therapeutics) is a novel next-generation ROS1/TRK/ALK-tyrosine kinase inhibitor (TKI). Repotrectinib also inhibits the Janus kinase (JAK) 2 and indirectly down-regulates STAT5 phosphorylation in the SET2 cancer cell line harboring a JAK2 V617F tyrosine kinase mutation (Drilon et al 2018). We have previously evaluated the preclinical anti-MM effects of other JAK1/2 inhibitors in combination with immunomodulatory drugs, such as lenalidomide (LEN) and corticosteroids both in vitro and in vivo. Ruxolitinib (RUX) in combination with lenalidomide and corticosteroids showed anti-MM effects. We have reported that RUX overcomes resistance of MM patients to LEN in an ongoing clinical trial (Berenson et al, 2018). Although activation of JAK2 and PI3K signaling is often detected in hematological malignancies, there is no data regarding the potential anti-MM effects of repotrectinib alone or in combination with immunomodulatory drugs. Thus, we evaluated the anti-myeloma effects of repotrectinib alone or in combination with LEN or pomalidomide (POM) in vitro using MM cell lines and ex-vivo using the human MM xenograft model LAG- λ1. Method: The human MM cell lines U266, RPMI8226, and MM1s were obtained from ATCC. Mononuclear cells (MCs) were isolated form MM patients' bone marrow (BM) aspirates. Cell viability was quantified using the MTS cell proliferation assay. For the ex-vivo studies, cells were isolated from the human myeloma xenograft LAG- λ1 after it was established in SCID mice. We determined the IC curves for the MM cell lines RPMI8226, U266, MM1S and fresh BMMCs from the MM patients and examined the anti-proliferative effects of repotrectinib alone using these 3 MM cell lines and 6 patients' fresh MM BMMC samples (3 patients with progressive disease [PD] and 3 patients in complete remission [CR]). Next, we combined repotrectinib with other active anti-MM drugs using these same three MM cell lines and the BMMCs from the 6 MM patients to evaluate the anti-proliferative effects of repotrectinib in combination with LEN or POM. Results MM tumor cells of three myeloma cell lines were exposed to increasing concentrations of repotrectinib from 0.39 to 100 µM. The results of anti-MM effects on cell viability showed that repotrectinib alone markedly inhibited MM tumor cell proliferation in all three MM cell lines in a concentration dependent manner. The MM1 cell line was the most sensitive to repotrectinib (with an IC50 of 1.5µM). We further determined the anti-myeloma effects of repotrectinib alone and in combination with other anti-myeloma agents by using fresh BMMCs from 3 patients with PD and 3 patients in CR using a cell proliferation assay. The results showed that repotrectinib alone minimally inhibited MM tumor cell proliferation. Repotrectinib combined with LEN or POM decreased cells from PD patients but not CR patients. We also determined the anti-MM effects of repotrectinib alone and in combination with LEN or POM using tumor cells that were isolated from the human LAG- λ1 human MM xenograft. The results showed repotrectinib alone inhibited tumor cell growing in a concentration dependent manner. Repotrectinib combined with either LEN or POM significantly enhanced inhibition of LAG- λ1 MM tumor cell growth when compared to repotrectinib alone. Summary These studies suggest that the combination of repotrectinib and immunomodulatory drugs may be effective for treatment of MM. We will further evaluate the anti-MM effects of Repotrectinib in combination with immunomodulatory drugs in vivo using our human MM xenograft models.