Synergistic Anti-Myeloma Effects of the Lyn Kinase Inhibitor INNO-406 In Combination with Doxorubicin, Melphalan and Bortezomib

Abstract 5015

Lyn is a member of the Src kinase (SFK) family and controls the activation threshold of multiple signaling pathways in different types of cells, including B cells (Yamanashi et al., 1989, PNAS). Multiple myeloma (MM) is a hematological malignancy of plasma cells. In MM, Lyn is involved in IL-6-induced cell proliferation (Hallek et al., 1997, Exp Hematol). As one of the most important growth and anti-apoptotic factors for MM cells, both the IL-6/STAT3 and IL-6/ERK pathways have been shown to promote MM cell proliferation and prevent apoptosis. However, STAT3 and ERK activation is not sufficient for IL-6-induced proliferation, which further requires the activation of SFK (Ishikawa et al., 2002, Blood). IL-6-induced proliferation of U266 myeloma cells has been reported to be significantly suppressed following exposure to Lyn-specific antisense oligonucleotides or a Src kinase inhibitor (Ishikawa et al., 2002, Blood). Therefore, a Lyn specific inhibitor, either alone or in combination with other anti-MM drugs, may be a more effective regimen for the treatment of MM. INNO-406 is a novel inhibitor of ABL as well as Lyn. In laboratory-based studies, it has shown anti-tumor activity in hematological malignancies, especially in leukemia cells harboring BCR-ABL mutations (Kimura et al., 2005, Blood). Since Lyn is involved in MM cell proliferation, targeting Lyn may be an alternative therapeutic approach and improve the efficacy of other anti-MM agents. Thus, we conducted this study in order to determine the anti-MM effects of INNO-406. First, Western blot analyses demonstrated that Lyn was expressed in all MM cells analyzed, including bone marrow (BM) mononuclear cells derived from MM patients and MM cells from LAGκ1A, LAGκ1B and LAGκ2 xenografts, three of our unique in vivo SCID-hu models of human MM, which were originally developed from MM patient BM biopsies and have been maintained in SCID mice. However, Lyn activation, as demonstrated by phosphorylation of its downstream molecule HS1, was infrequently found in MM cells. INNO-406 inhibited the growth of the three MM cell lines tested including RPMI8226, U266 and MM1S. Interestingly, the U266 cell line is most resistant to INNO-406 among the three MM cell lines tested although its Lyn activation and expression level is the highest. Apoptosis analysis using flow cytometric analysis following Annexin V staining demonstrated that INNO-406 induced apoptosis in MM cells. When combined with melphalan, doxorubicin or bortezomib, INNO-406 synergistically inhibited MM cell growth. We are currently studying the molecular mechanisms of INNO-406's anti-MM effect, either alone or in combination treatment, and validating our in vitro findings using our unique in vivo mouse models of human MM. Updated data from these additional studies will be reported at the meeting.