TY - JOUR
T1 - Binding of released bim to Mcl-1 is a mechanism of intrinsic resistance to ABT-199 which can be overcome by combination with daunorubicin or cytarabine in AML cells
AU - Niu, Xiaojia
AU - Zhao, Jianyun
AU - Ma, Jun
AU - Xie, Chengzhi
AU - Edwards, Holly
AU - Wang, Guan
AU - Caldwell, J. Timothy
AU - Xiang, Shengyan
AU - Zhang, Xiaohong
AU - Chu, Roland
AU - Wang, Zhihong J.
AU - Lin, Hai
AU - Taub, Jeffrey W.
AU - Ge, Yubin
N1 - Funding Information:
This study was supported by Start-up Funds from Jilin University, Changchun, China The Decerchio/Guisewite Family, and the Barbara Ann Karmanos Cancer Institute, grants from the National Natural Science Foundation of China, NSFC 31271477 and NSFC 31471295, the Graduate Innovation Fund of Jilin University (to X. Niu), the Ring Screw Textron Endowed Chair for Pediatric Cancer Research, Hyundai Hope On Wheels, and the Christoph A.L.L. Star Foundation. J.T. Caldwell is a predoctoral trainee supported by T32 CA009531 from the NCI. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
Publisher Copyright:
©2016 American Association for Cancer Research.
PY - 2016/9/1
Y1 - 2016/9/1
N2 - Purpose: To investigate the molecular mechanism underlying intrinsic resistance to ABT-199. Experimental Design: Western blots and real-time RT-PCR were used to determine levels of Mcl-1 after ABT-199 treatment alone or in combination with cytarabine or daunorubicin. Immunoprecipitation of Bim and Mcl-1 were used to determine the effect of ABT-199 treatment on their interactions with Bcl-2 family members. Lentiviral short hairpin RNA knockdown of Bim and CRISPR knockdown of Mcl-1 were used to confirm their role in resistance to ABT-199. JC-1 assays and flow cytometry were used to determine drug-induced apoptosis. Results: Immunoprecipitation of Bim from ABT-199-treated cell lines and a primary patient sample demonstrated decreased association with Bcl-2, but increased association with Mcl-1 without corresponding change in mitochondrial outer membrane potential. ABT-199 treatment resulted in increased levels of Mcl-1 protein, unchanged or decreased Mcl-1 transcript levels, and increased Mcl-1 protein half-life, suggesting that the association with Bim plays a role in stabilizing Mcl-1 protein. Combining conventional chemotherapeutic agent cytarabine or daunorubicin with ABT-199 resulted in increased DNA damage along with decreased Mcl-1 protein levels, compared with ABT-199 alone, and synergistic induction of cell death in both AML cell lines and primary patient samples obtained from AML patients at diagnosis. Conclusions: Our results demonstrate that sequestration of Bim by Mcl-1 is a mechanism of intrinsic ABT-199 resistance and supports the clinical development of ABT-199 in combination with cytarabine or daunorubicin for the treatment of AML.
AB - Purpose: To investigate the molecular mechanism underlying intrinsic resistance to ABT-199. Experimental Design: Western blots and real-time RT-PCR were used to determine levels of Mcl-1 after ABT-199 treatment alone or in combination with cytarabine or daunorubicin. Immunoprecipitation of Bim and Mcl-1 were used to determine the effect of ABT-199 treatment on their interactions with Bcl-2 family members. Lentiviral short hairpin RNA knockdown of Bim and CRISPR knockdown of Mcl-1 were used to confirm their role in resistance to ABT-199. JC-1 assays and flow cytometry were used to determine drug-induced apoptosis. Results: Immunoprecipitation of Bim from ABT-199-treated cell lines and a primary patient sample demonstrated decreased association with Bcl-2, but increased association with Mcl-1 without corresponding change in mitochondrial outer membrane potential. ABT-199 treatment resulted in increased levels of Mcl-1 protein, unchanged or decreased Mcl-1 transcript levels, and increased Mcl-1 protein half-life, suggesting that the association with Bim plays a role in stabilizing Mcl-1 protein. Combining conventional chemotherapeutic agent cytarabine or daunorubicin with ABT-199 resulted in increased DNA damage along with decreased Mcl-1 protein levels, compared with ABT-199 alone, and synergistic induction of cell death in both AML cell lines and primary patient samples obtained from AML patients at diagnosis. Conclusions: Our results demonstrate that sequestration of Bim by Mcl-1 is a mechanism of intrinsic ABT-199 resistance and supports the clinical development of ABT-199 in combination with cytarabine or daunorubicin for the treatment of AML.
UR - http://www.scopus.com/inward/record.url?scp=84986892344&partnerID=8YFLogxK
U2 - 10.1158/1078-0432.CCR-15-3057
DO - 10.1158/1078-0432.CCR-15-3057
M3 - Article
C2 - 27103402
AN - SCOPUS:84986892344
SN - 1078-0432
VL - 22
SP - 4440
EP - 4451
JO - Clinical Cancer Research
JF - Clinical Cancer Research
IS - 17
ER -