Lomeguatrib, a O⁶-Methylguanine-DNA-Methyltransferase (MGMT) Inhibitor, Induces DNA Damage Induced Apoptosis By Targeting Double-Strand DNA Repair in Multiple Myeloma

Background/Aim:Multiple Myeloma (MM), characterized by aberrant accumulation of terminally differentiated plasma cells in the bone marrow, is promoted by ongoing DNA damage and genomic instability. O⁶-methylguanine-DNA-methyltransferase (MGMT), a DNA repair protein, transfers the alkyl group from guanine to its cysteine residue and is implicated as an important chemoresistance factor particularly in chemotherapy-induced DNA alkylation. A growing body of evidence has shown that enhanced DNA repair capacity stimulates cancer cell survival and facilitates genomic instability. Thus, targeting DNA repair activities including MGMT seems to be a promising strategy for the treatment of MM.

Materials and Methods: In our study, MGMT expression was evaluated in MGUS, SMM and MM patients compared to normal plasma cells using publically available gene expression data sets (Gene Expression Omnibus; GSE47552). Furthermore, MGMT mRNA and protein levels were determined in human MM cell lines (NCI H929, RPMI 8226 and U266), three samples of control PBMCs, matched CD138+ myeloma and CD138- non-tumorigenic cells obtained from bone marrow aspirates of same MM patients [newly diagnosed (ND, n=12) and relapsed / refractory (RR, n=8)] by RealTime Ready PCR Assay and Western blot, respectively. The experiments with human cells were approved by the Ethical Committee of Ankara University School of Medicine. Following treatment of MM cells with Lomeguatrib for 48 hours, cell viability, cell cycle progression and apoptosis were assessed by MTT and flow cytometry. DNA damage levels were examined by alkaline comet assay and immunoblotting of ΥH2AX phosphorylation.

Results: Comparable MGMT expression was detected between MM cells and normal plasma cells, and between ND CD138+ and RR CD138+ myeloma cells. Inhibition of MGMT activity by Lomeguatrib reduced MM cell viability and induced apoptosis in MGMT proficient but not in MGMT deficient MM cells in a dose-dependent manner. In contrast, Lomeguatrib did not affect the cell viability of PBMCs from three healthy donors and CD138- non-tumorigenic cells from three MM patients. Furthermore, Lomeguatrib perturbed cell cycle and decreased phosphorylation of G1/S specific CyclinE1 in these cells and led to an increase in DNA damage. Western blot analysis revealed a reduction in protein levels of Fen1, Ku-70 and Rad51 which are involved in double-strand DNA break repair.

Conclusion: These results revealed that MGMT promotes repair of DNA double-strand breaks in MGMT proficient MM cells and MGMT inhibition enhances DNA damage-induced apoptosis, suggesting that inhibition of MGMT by Lomeguatrib might be a useful strategy to overcome unfaithful DNA repair in MM. This research has been supported by The Scientific and Technological Research Council of Turkey (No:113Z383).