Inhibition of DNA Methyltransferases By Volasertib in HMA Resistant Cell Lines

Introduction: Resistance to hypomethylating agents (HMA) in myelodysplastic syndrome (MDS) and acute myeloid leukemia (AML) is a concerning problem. We established HMA resistant cell lines with overexpression of DNA methyltransferases (DNMTs), which were not suppressed by azacitidine or decitabine. Polo-like kinase 1 (PLK1) is a key cell cycle modulator and is associated with regulation of the PIK3/Akt pathway, which was reported to stabilize DNMTs. We hypothesized that volasertib, a PLK inhibitor, might overcome HMA resistance through regulation of PIK3/Akt pathway and DNMTs.

Methods: Cell viability was determined in HMA resistant cell lines (MOLM/AZA-1 and MOLM/DEC-5) and MOLM-13. Specific antibodies including phosphorylated-histone H3 (PHH3), caspase 3, PARP, and XIAP were used to evaluate apoptosis or DNA damage. We also analyzed phosphorylated-Akt (p-Akt), Akt, and DNMTs (DNMT1, 3A, and 3B) by immunoblot assay.

Results: Volasertib effectively inhibit proliferation of MOLM/AZA-1 and MOLM/DEC-5 cells as well as MOLM-13 cells in a dose dependent manner. It also showed superior anti-leukemic effects compared to cytarabine, azicitidine or decitiabine. Volasertib showed a dose dependent increased expression of PHH3, caspase 3, and PARP, and decrease of XIAP. Treatment of volasertib caused decreased expression of p-AKT and all DNMTs (DNMT1, 3A, and 3B). GDC-0941, a PI3K inhibitor, suppressed DNMT1 and DNMT3A as well as p-Akt, but it could not decrease DNMT3B. A combination of volasertib and decitabine showed synergistic effects in MOLM/DEC-5. Anti-proliferative effects of volasertib were sustained in ex vivo AML or MDS samples.

Conclusion: In our HMA resistant models, volasertib effectively inhibited all DNMTs. DNMT1 and 3A seemed to be suppressed via regulation of PI3K/Akt, but suppression of DNMT3B by volasertib might have separate mechanisms. Our data suggest that volasertib with or without decitabine has potential role in overcoming the HMA resistance in patients with AML and MDS.