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. Author manuscript; available in PMC: 2017 Nov 22.
Published in final edited form as: Cell Rep. 2016 Nov 22;17(9):2367–2381. doi: 10.1016/j.celrep.2016.10.077

Figure 3. Disruption of HR by dinaciclib and sensitization to PARP inhibition of BRCA wild type and BRCA-mutated cells with acquired PARP inhibitor resistance.

Figure 3

(A) Cells were treated with vehicle or 10 nM dinaciclib for 18 hrs prior to treatment with 10 Gy γ-irradiation (IR). Six hrs post-IR, cells were analyzed for BRCA1, RAD51 and γ-H2AX focus formation. (B) Quantification of cells with > 5 foci in irradiated cells pretreated with vehicle or dinaciclib at the indicated concentrations. (C) USOS DR-GFP cells were transfected I-SceI in the presence of vehicle or 15 nM dinaciclib. The percentage of GFP-positive cells is significantly reduced (P < 0.0001) in the presence of dinaciclib, consistent with direct inhibition of HR repair (See also Figure S3). (D) BRCA-proficient TNBC cell lines were treated with veliparib in the absence and presence of dinaciclib, demonstrating reduced IC50 values in the presence of dinaciclib. (E) (Left) An MDA-MB-436 PARP inhibitor-resistant derivative (MDA-MB-436-RR2) (Johnson et al., 2013) was treated with veliparib in the absence or presence of dinaciclib, demonstrating reduced IC50 in the presence of dinaciclib. (Middle) Reduced expression of the mutant BRCA1 protein and RAD51 in response to dinaciclib. (Right) MDA-MB-436-RR2 cells were treated with vehicle or dinaciclib at the indicated concentration for 18 hours, subjected to 10 Gy IR and assessed for RAD51 focus formation 6 hours later. (P < 0.0001) (F) Treatment history of the BRCA2 carrier; PDX 12-58 was procured after progression on cisplatin and olaparib (see also Figure S4A) . SD, stable disease. (G) (Left) Mice bearing 12–58 xenografts were treated with vehicle (n = 3) or cisplatin (n = 3) on days 1 and 22 (arrows), with tumor volume measured over 36 days. (Right) Mice were treated with vehicle (n = 4), veliparib (n = 8), dinaciclib (n = 8) or the combination (n = 8). Combination treatment produced significant tumor growth inhibition at day 42 compared to vehicle (P < 0.0001) or monotherapies (P < 0.0001 for both veliparib and dinaciclib). * P < 0.01, ** P < 0.001, ***P < 0.0001 for experimental value vs. control.