Dynamin impacts homology-directed repair and breast cancer response to chemotherapy
Sophia B. Chernikova, … , Balázs Győrffy, J. Martin Brown
Sophia B. Chernikova, … , Balázs Győrffy, J. Martin Brown
Published October 29, 2018
Citation Information: J Clin Invest. 2018;128(12):5307-5321. https://doi.org/10.1172/JCI87191.
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Dynamin impacts homology-directed repair and breast cancer response to chemotherapy

Abstract

After the initial responsiveness of triple-negative breast cancers (TNBCs) to chemotherapy, they often recur as chemotherapy-resistant tumors, and this has been associated with upregulated homology-directed repair (HDR). Thus, inhibitors of HDR could be a useful adjunct to chemotherapy treatment of these cancers. We performed a high-throughput chemical screen for inhibitors of HDR from which we obtained a number of hits that disrupted microtubule dynamics. We postulated that high levels of the target molecules of our screen in tumors would correlate with poor chemotherapy response. We found that inhibition or knockdown of dynamin 2 (DNM2), known for its role in endocytic cell trafficking and microtubule dynamics, impaired HDR and improved response to chemotherapy of cells and of tumors in mice. In a retrospective analysis, levels of DNM2 at the time of treatment strongly predicted chemotherapy outcome for estrogen receptor–negative and especially for TNBC patients. We propose that DNM2-associated DNA repair enzyme trafficking is important for HDR efficiency and is a powerful predictor of sensitivity to breast cancer chemotherapy and an important target for therapy.

Authors

Sophia B. Chernikova, Rochelle B. Nguyen, Jessica T. Truong, Stephano S. Mello, Jason H. Stafford, Michael P. Hay, Andrew Olson, David E. Solow-Cordero, Douglas J. Wood, Solomon Henry, Rie von Eyben, Lei Deng, Melanie Hayden Gephart, Asaithamby Aroumougame, Claudia Wiese, John C. Game, Balázs Győrffy, J. Martin Brown

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Figure 5

Inhibition of DNM2 impairs HDR.

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Inhibition of DNM2 impairs HDR. (A) Dynasore reduces gene conversion in ...
(A) Dynasore reduces gene conversion in the U2OS-DR-GFP cells. (B) Dynasore does not inhibit NHEJ, as measured by the appearance of CD8+GFP– cells after induction of DSB by the I-SceI enzyme in 293-1040 cells. (C) Dynasore reduces numbers of cells with Rad51-positive/γH2AX-positive foci after IR. CHO AA8 cells were fixed 2 hours after 3 Gy. (D) Left: Rad51 foci in the CHO AA8 cells at 2 hours after 3 Gy are positive for γH2AX, showing that HR repair occurs at the sites of DSBs. Right: Treatment with dynasore reduces the accumulation of Rad51 at the γH2AX foci. Insets show higher magnification of selected areas. Scale bars: 10 μm. (E) Treatment with dynasore increases sensitivity of HDR-proficient CHO AA8 cells to chlorambucil, while it does not alter the sensitivity to chlorambucil in HDR-deficient XRCC3-mutant irs1SF cells. See Supplemental Figure 6A for the survival of irs1SF cells at lower concentrations of chlorambucil. (F) DNM2 knockdown results in increased sensitivity to chlorambucil in the human TNBC cell line MDA-MB-231-BR3. DNM2 knockdown was induced by the addition of doxycycline, as shown in the Western blot (right). Cells were treated with chlorambucil on day 5 after the addition of doxycycline. Shown are means ± SDs from n ≥ 5 experiments. (G) Correction of HDR defect in the irs1SF cells by introduction of XRCC3 cDNA restores both the resistance to chlorambucil and the potentiation of chlorambucil effect by inhibition of DNM2. (A–C, E, and G) Shown are means ± SDs (ranges) from n ≥ 2 MTS experiments. (A–C and E–G) Significance analysis: ANOVA. Dynasore-treated groups were compared with no-drug group (A–C, E, and G). *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.