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 5

Mcl-1 directly interacts with Ku via BH1 and BH3 domains, which are required for Mcl-1 dissociation of Ku/DNA complex.

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Mcl-1 directly interacts with Ku via BH1 and BH3 domains, which are requ...
(A) Schematic representation of various Mcl-1 deletion mutants. (B) GST beads coated with purified recombinant GST-tagged WT or individual Mcl-1 deletion mutants were incubated with recombinant Ku70/Ku80 heterodimer. Mcl-1–associated Ku70 or Ku80 and GST–Mcl-1 were analyzed by Western blot. (C) Mcl1–/– MEFs were transfected with FLAG-tagged Mcl-1 WT or individual Mcl-1 deletion mutants using Amaxa electroporation system. Co-IP experiments were performed using anti-FLAG M2 beads, followed by Western blot analysis of Ku70, Ku80, and FLAG–Mcl-1. (D) Mre11-Rad50 (MR) complex was expressed in Sf9 insect cells and purified using an anti-FLAG M2 affinity column. (E and F) The 5′-end-labeled overhang DNA was incubated with Ku or MR complex in the absence or presence of increasing concentrations of Mcl-1 (E) or individual Mcl-1 deletion mutant proteins (F). BSA was used as negative control. The Ku/DNA or MR/DNA complexes were analyzed by EMSA.