Sequential combination of bortezomib and WEE1 inhibitor, MK-1775, induced apoptosis in multiple myeloma cell lines.

Abstract

INTRODUCTION\nMultiple myeloma (MM) remains incurable due to high rates of relapse after various treatment regimens. WEE1 is a cell cycle related gene that regulates the G2/M checkpoint and promotes cell cycle suspension for consequent DNA repair. To date, there are clinical studies for the evaluation of WEE1 inhibitors in the treatment of solid tumors and studies on cell lines of non-MM hematological tumors.\n\n\nOBJECTIVES\nTo perform in\xa0vitro functional studies to verify the effect of the inhibition of WEE1 on MM cell lines viability and its potential as therapeutic target.\n\n\nMATERIAL AND METHODS\nWEE1 expression was evaluated in 22 newly diagnosed MM patients and in four MM cell lines, RPMI-8226, U266 and SKO-007 and SK-MM2, by quantitative real-time PCR (qPCR). After treatment with the WEE1 inhibitor (MK-1775), with or without proteasome inhibitor (bortezomib) pretreatment, we assessed cell viability through Prestoblue functional test, microspheres formation in soft agar, and induction of apoptosis and cell cycle alterations by flow cytometry.\n\n\nRESULTS\nAll MM cell lines showed WEE1 expression by qPCR. RPMI-8226 and U266 showed a 50% reduction in cell viability after 24\u202fh of incubation with MK-1775, at concentrations of 5\u202fμM and 20\u202fμM, respectively. SKO-007 showed dose and time dependence to this drug. Combination therapy with bortezomib and MK-1775 abolished the formation of soft agar microspheres in the RPMI-8226\u202fcell line (also responsive to the use of both drugs) and U266, but SKO-007 was resistant to all drugs, isolated and combined. However, treatment of bortezomib followed by MK-1775 (sequential treatment) versus bortezomib alone showed statistically significant impact on cell lines total apoptosis: 88.8% vs 74.1% in RPMI-8222 (confirmed by cell cycle experiments); 92.5% vs 86.6% in U266; and 60.2% 30.9% on SKO-007 (p\u202f<\u202f0.05).\n\n\nCONCLUSION\nThe sequential combination of bortezomib and WEE1 inhibitor, MK-1775, induced apoptosis in RPMI-8226, U266, and especially SKO-007\u202fcell lines, more efficiently than the use of the same isolated drugs, highlighting its effect in inhibition of proliferation of tumor cells in MM cell lines. Our data suggest that WEE1 can figure as a MM target and that the sequential combination of bortezomib and MK-1775 may be explored in future clinical trials.