Targeting Mcl-1 enhances DNA replication stress sensitivity to cancer therapy
Guo Chen, … , Paul W. Doetsch, Xingming Deng
Guo Chen, … , Paul W. Doetsch, Xingming Deng
Published December 11, 2017
Citation Information: J Clin Invest. 2018;128(1):500-516. https://doi.org/10.1172/JCI92742.
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Research Article Cell biology

Targeting Mcl-1 enhances DNA replication stress sensitivity to cancer therapy

Abstract

DNA double-strand breaks (DSBs) are mainly repaired either by homologous recombination (HR) or by nonhomologous end-joining (NHEJ) pathways. Here, we showed that myeloid cell leukemia sequence 1 (Mcl-1) acts as a functional switch in selecting between HR and NHEJ pathways. Mcl-1 was cell cycle–regulated during HR, with its expression peaking in S/G2 phase. While endogenous Mcl-1 depletion reduced HR and enhanced NHEJ, Mcl-1 overexpression resulted in a net increase in HR over NHEJ. Mcl-1 directly interacted with the dimeric Ku protein complex via its Bcl-2 homology 1 and 3 (BH1 and BH3) domains, which are required for Mcl-1 to inhibit Ku-mediated NHEJ. Mcl-1 also promoted DNA resection mediated by the Mre11 complex and HR-dependent DSB repair. Using the Mcl-1 BH1 domain as a docking site, we identified a small molecule, MI-223, that directly bound to BH1 and blocked Mcl-1–stimulated HR DNA repair, leading to sensitization of cancer cells to hydroxyurea- or olaparib-induced DNA replication stress. Combined treatment with MI-223 and hydroxyurea or olaparib exhibited a strong synergy against lung cancer in vivo. This mechanism-driven combination of agents provides a highly attractive therapeutic strategy to improve lung cancer outcomes.

Authors

Guo Chen, Andrew T. Magis, Ke Xu, Dongkyoo Park, David S. Yu, Taofeek K. Owonikoko, Gabriel L. Sica, Sarah W. Satola, Suresh S. Ramalingam, Walter J. Curran, Paul W. Doetsch, Xingming Deng

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Figure 1

Mcl-1 is cell cycle–regulated with expression peaking in S/G2 phase.

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Mcl-1 is cell cycle–regulated with expression peaking in S/G2 phase. (A)...
(A) CENP-F and cyclin A as S/G2 markers were coimmunostained with Mcl-1 in H1299 cells. DAPI was used as nuclear marker. Scale bars: 25 μm. (B) H1299 cells were synchronized by double-thymidine block. After thymidine was washed off, cells were released to normal medium for a time course up to 24 hours. Mcl-1, Bcl-2, Bcl-xL, cyclin A, and cyclin F in total cell lysates were analyzed by Western blot. As, asynchronous cells. (C and D) Relative protein and mRNA levels of Mcl-1, Bcl-2, and Bcl-xL were quantified during cell cycle progression. (E) Mcl-1 ubiquitination was analyzed by anti–Mcl-1 co-IP and Western blot using anti-HA antibody following transfection with HA-tagged ubiquitin (HA-Ub) constructs during cell cycle progression. (F) Mitochondrial and nuclear fractions were isolated from cells at various time points during cell cycle progression. Mcl-1 was analyzed by Western blot. Prohibitin or PCNA was used as mitochondrial or nuclear marker, respectively.