PARP inhibitor combinations with Olaparib for synergistic efficacy in Olaparib-resistant triple-negative breast cancer cells

Triple-negative breast cancer (TNBC) is associated with aggressive and heterogenous tumour phenotype, early tumour relapse and poor prognosis than other types of invasive breast cancer. TNBC does not respond to endocrine therapy due to the lack of oestrogen receptor (ER), progesterone receptor (PR) and human epidermal growth factor receptor type 2 (HER2) expression. Although poly(ADP-ribose) polymerase (PARP) inhibitors (PARPi) have been successfully developed to treat breast cancer; however, these PARPi as monotherapies are limited to TNBC patients with defective gene where BRCAdeficient TNBC accounts for a small subset (< 30%) of TNBC patients. With few exceptions, monotherapy remains ineffective due to insufficient tumour suppression, requires high dose of drug, dose-limiting toxicity and acquired drug resistance is common. Thus, combining PARPi with other therapeutics to increase efficacy, reduce drug dosage, and overcome drug resistance is a promising strategy to treat TNBC. Moreover, drug combination screening of a PARPi is a relatively new concept and having a compound library with diverse chemical space to be screened alongside PARPi holds potential to new drug combination discovery. Therefore, this study aims to discover new combinations of Olaparib, the most successful PARPi to date, alongside other therapeutic in MDA-MB-231 TNBC cells without BRCA mutation. The library used in this screen comprises of a total of > 100 molecules including DNA-binding ruthenium compounds and commercially-available drugs. In this study, Olaparib-resistant (OlaR) MDA-MB-231R cells were developed from the parental MDA-MB-231 cells after prolonged treatment (~8 months) with Olaparib. Olaparib resistance was assessed by clonogenic survival assay in which 28-fold level of Olaparib resistance was observed in MDA-MB-231R cells in comparison to the parental cells. The acquisition of Olaparib resistance is associated with upregulation of drug efflux pumps of P-glycoprotein (P-gp), loss of poly(ADP-ribose) glycohydrolase (PARG) and the activation of ATR/Chk1 signalling pathway. Synergistic drug combinations were identified using Chou and Talalay combination index (CI) method in which CI < 0.9 demonstrated synergy. This study identified four hits including a natural compound, curcumin; two ruthenium(II)-rhenium(I) metallomacrocyles, Ru(bpy)Re and Ru(dppz)Re; and FDA-approved Exemestane (CI < 0.9 across the tested concentrations). Synergy was confirmed using long-term clonogenic survival assay in which single agents showed survival fractions (S.F.) of > 45%, and upon co-treatments, significant (P < 0.01) loss in clonogenic potential of cells was observed. Moreover, synergy was retained in the acquired OlaR MDA-MB-231R cells. Mechanistic studies indicated synergy was achieved via significant (P < 0.05) enhancement of DNA damage and the resultant apoptosis. Most importantly, all identified combinations showed low cytotoxicity in MCF10A normal breast cells with substantial cancer versus normal cell selectivity (selectivity index (SI) > 50). These identified combinations were also able to inhibit the growth of 3D breast cancer spheroids. Thus, this study supports the concept that PARPi Olaparib combination strategy represents a promising approach towards aggressive strains of BRCA-proficient TNBC and can overcome acquired PARPi resistance. Additionally, co-treatment with the synergistic pairing of Olaparib and Ru-PIP, a ruthenium compound, in non-small cell lung cancer (NSCLC) cells showed enhanced (P < 0.05) DNA damages and reactive oxygen species (ROS) levels, and was able to inhibit the lung cancer spheroids growth, providing further affirmation of expanding the benefits of these identified combinations to other cancer types beyond TNBC. Moreover, exposure of zebrafish embryos to these identified synergistic pairings over 96 h did not lead to any noticeable signs of toxicity, making these newly identified combinations as suitable candidates for future in vivo investigations.

Text 118100.pdf Download (1MB) Official URL or Download Paper: http://ethesis.upm.edu.my/id/eprint/18349

Actions (login required)

View Item