Abstract P6-08-03: Dynamics of cancer stem cell surface marker, CD44 and CD44v6 in the antitumor efficacy of PARP inhibitor in combination with PI3K pathway inhibitor in TNBC xenograft model
Authors: Lin X , Sun Y , Carlson JH , Williams C , Krie A , Friedman L , De P , Dey N , Leyland-Jones B 
Published: March 2017

Introduction: Triple Negative (TN) BC has a limited benefit from conventional therapy and a few options for targeted therapy like PARP inhibitor. We reported that doubling down on the PI3K-mTOR pathway enhances the antitumor efficacy of PARP inhibitor in TNBC (Neoplasia, 2014). Tumor cells with distinctive stem cell (CSC) like properties exist in carcinomas including breast and CD44+CSC marker expression correlates with decreased survival (Fillmore and Kuperwasser, 2008). In TNBC the intratumoral presence of stem cell population is associated with increased aggressiveness. Among many CSC markers, CD44 is best known to have a prognostic role (Collina et al., 2015) and the PI3K/E2F1 pathway has been identified as a potential signaling link to HA/CD44 activation. Hypothesis: We hypothesized that xenografts of TNBC cell lines bearing stem cell property will respond in a characteristic way to GDC0980 in combination with ABT888 plus carboplatin. Methods:Athymic mice bearing BRCA-competent TNBC xenograft tumors were used to test the combination of GDC-0980 with ABT888 plus carboplatin. Mechanism-based in vitro studies were conducted to understand the mode of action of the drugs. IHC for CD44 and CD44v6 was standardized in positive tumor-controls and identified in (1) BC TMAs, (2) TNBC cell lines and (3) patients from our Avera cohort. Finally, the CD44-expression in tumors of different arms of the xenograft study (MDA-MB231 and MDA-MB468) was evaluated independently by a pathologist who was unaware of the study design. Results: CD44 was identified primarily in the membrane and to a lesser extent in the cytosol of tumor cells of BC patients (TMA and Avera cohort), TNBC cell lines and xenograft-tumors. GDC-0980 in combination with ABT888 plus carboplatin blocked the growth of established xenograft tumors by 80-90% with a concomitant decrease in Ki67 IHC-levels. Membrane expression of CD44 inversely correlated to tumor sizes which significantly reduced in response to drug combinations. Mechanistically, GDC-0980 treatment led to DNA damage (increased pgH2AX), gain in PAR and a subsequent sensitization of BRCA-competent TNBC cells to ABT888 plus carboplatin with a time-dependent (1) decrease in proliferation signals (pAKT), PAR/PARP ratios, PAR/pgH2AX ratios, live/dead cell ratios, and clonogenic 3D growth & (2) increase in apoptosis markers (cleaved-caspase3, cleaved-PARP and annexinV positivity). The relationship between expression of CD44 and CD44v6 in the xenograft tumors the following treatment is being worked out which will be presented at the meeting. Significance: Our findings demonstrated that PARP inhibitor plus carboplatin in combination with dual PI3K-mTOR inhibition effectively reduced the CD44+CSC population in TNBC xenograft tumors. In a BRCA-competent model, PI3K-pathway inhibition not only enhanced the antitumor activity of ABT888 plus carboplatin by inhibiting proliferation and tumor-induced angiogenesis but also by decreasing CD44+CSC. We present the first mechanism-based study to demonstrate the integral role of PI3K-mTOR pathway and DDR pathway in the control of CSC surface marker in orchestrating antitumor actions of PARP inhibitor in TNBC.

Citation Format: Lin X, Sun Y, Carlson JH, Williams C, Krie A, Friedman L, De P, Dey N, Leyland-Jones B. Dynamics of cancer stem cell surface marker, CD44 and CD44v6 in the antitumor efficacy of PARP inhibitor in combination with PI3K pathway inhibitor in TNBC xenograft model [abstract]. In: Proceedings of the 2016 San Antonio Breast Cancer Symposium; 2016 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2017;77(4 Suppl):Abstract nr P6-08-03.