Xenograft mouse model in vivo study and tumor tissue analysis

To generate xenograft tumors, 15 million MDA-MB231 breast cancer cells were subcutaneously implanted in the flanks of female SCID mice 8 wk of age, followed by tumor growth monitoring. Mice were orally dosed with either vehicle, 80 mg/kg niraparib daily, IP dosed 100 mg/kg cysmethynil every other day, or the combination of same doses of niraparib and cysmethynil for the treatment, as specified by each study, starting when the tumors reached the stable volume of 100–500 mm³. Tumor growth was followed until termination of experiment, which was when the fastest growing group of tumors, in this case the vehicle-treated control group, reached the mean size of 1,500 mm³. The study protocol was approved by the institutional Animal Care and Use Committee (IACUC). The tumors were isolated after mouse euthanization for imaging and sample preservation, which is by the standard fixation (HT501128; Sigma-Aldrich) and paraffin embedding method. Histology slide preparation and H&E staining were performed by Duke-NUS Histology Service. TUNEL assay to visualize apoptotic cells was per protocol of the manufacture of TUNEL Assay Kit (ab206386; Abcam).