5-Azacytidine, a DNA methyltransferase inhibitor, induces ATR-mediated DNA double-strand break responses, apoptosis, and synergistic cytotoxicity with doxorubicin and bortezomib against multiple myeloma cells.
In this study, we investigated the cytotoxicity of 5-azacytidine, a DNA methyltransferase inhibitor, against multiple myeloma (MM) cells, and characterized DNA damage-related mechanisms of cell death. 5-Azacytidine showed significant cytotoxicity against both conventional therapy-sensitive and therapy-resistant MM cell lines, as well as multidrug-resistant patient-derived MM cells, with IC(50) of approximately 0.8-3 micromol/L. Conversely, 5-azacytidine was not cytotoxic to peripheral blood mononuclear cells or patient-derived bone marrow stromal cells (BMSC) at these doses. Importantly, 5-azacytidine overcame the survival and growth advantages conferred by exogenous interleukin-6 (IL-6), insulin-like growth factor-I (IGF-I), or by adherence of MM cells to BMSCs. 5-Azacytidine treatment induced DNA double-strand break (DSB) responses, as evidenced by H2AX, Chk2, and p53 phosphorylations, and apoptosis of MM cells. 5-Azacytidine-induced apoptosis was both caspase dependent and independent, with caspase 8 and caspase 9 cleavage; Mcl-1 cleavage; Bax, Puma, and Noxa up-regulation; as well as release of AIF and EndoG from the mitochondria. Finally, we show that 5-azacytidine-induced DNA DSB responses were mediated predominantly by ATR, and that doxorubicin, as well as bortezomib, synergistically enhanced 5-azacytidine-induced MM cell death. Taken together, these data provide the preclinical rationale for the clinical evaluation of 5-azacytidine, alone and in combination with doxorubicin and bortezomib, to improve patient outcome in MM.
Kiziltepe
Tanyel
T
Jerome Lipper Multiple Myeloma Center, Department of Medical Oncology, Dana-Farber Cancer Institute and Harvard Medical School, Boston, Massachusetts 02115, USA. kenneth_anderson@dfci.harvard.edu
Hideshima
Teru
T
Catley
Laurence
L
Raje
Noopur
N
Yasui
Hiroshi
H
Shiraishi
Norihiko
N
Okawa
Yutaka
Y
Ikeda
Hiroshi
H
Vallet
Sonia
S
Pozzi
Samantha
S
Ishitsuka
Kenji
K
Ocio
Enrique M
EM
Chauhan
Dharminder
D
Anderson
Kenneth C
KC
D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001374:Azacitidine / Q000494:pharmacology*; D001897:Boronic Acids / Q000494:pharmacology*; D000069286:Bortezomib; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D004249:DNA Damage; D004273:DNA, Neoplasm / Q000187:drug effects*; D004317:Doxorubicin / Q000494:pharmacology*; D004357:Drug Synergism; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D006801:Humans; D009101:Multiple Myeloma / Q000473:pathology*; D011719:Pyrazines / Q000494:pharmacology*
D000970:Antineoplastic Agents; D001897:Boronic Acids; D004273:DNA, Neoplasm; D004791:Enzyme Inhibitors; D011719:Pyrazines; D000069286:Bortezomib; D004317:Doxorubicin; D001374:Azacitidine