AOH1996, an Inhibitor of PCNA, Sensitises Breast Cancer Cells to Cytotoxic Chemotherapy.

Purpose: AOH1996 is an inhibitor of proliferating cell nuclear antigen (PCNA). We have assessed efficacies of AOH1996 as a therapeutic, alone and in combination with representative chemotherapies using breast cancer cells of luminal, HER2 positive, and triple negative subtypes. Methods: MCF-7, AU-565, MDA-MB-231, MDA-MB-468, and HCC1143 breast cancer cells were used. Effects of AOH1996, epirubicin, and/or docetaxel on cell proliferation/survival were assessed using MTT and clonogenic assays. DNA damage was evaluated by γ-H2AX immunofluorescence. Results: Single agent AOH1996 treatment reduced proliferation/survival in all cells tested in a dose- and time-dependent manner. Use of AOH1996 in combination with either epirubicin or docetaxel caused significant, consistent and substantial reductions in proliferation/survival that exceeded additive effects for the agents alone. For example, in clonogenic assays using MCF-7 cells, the lowest doses tested of epirubicin and AOH1996 caused inhibition of only 4% and 3% individually, while the combination caused significant inhibition by 34%, representing a 4.8-fold increase on the additive effect. Similarly, using AU-565 cells, docetaxel and AOH1996 caused 6% and 9% inhibition individually, while the combination caused 48%, representing a 3.2-fold increase on the additive effect. Nuclear γ-H2AX foci, indicative of double‑strand DNA damage, were quantified to investigate mechanisms. Combinations of AOH1996 with epirubicin or docetaxel consistently caused significant increases in foci that exceeded additive effects in every cell line, demonstrating that combinations were highly DNA-damaging. Conclusion: Based on our in vitro data, AOH1996 has potential as a breast cancer therapy and causes chemo-sensitisation in combination with chemotherapies standardly used clinically in primary breast cancer. Future in vivo studies are warranted.