Abstract P4-14-25: Single agent and combined targeting of PI3K, mTOR, HER2 and ER signaling in a panel of HER2+/ER+ versus HER2+/ER- breast cancer cell lines
Authors: O'Brien NA , Nichols CM , Thomas J , Conklin D , Kalous O , Linnartz R , Di Tomasso E , Hurvitz SA , Hirawat S , Slamon DJ 
Published: May 2016

Abstract
Background: Altered PI3K/mTOR signaling, through activating mutations in PIK3CA and/or PTEN loss, has been implicated in resistance to both hormonal and HER2-directed therapeutics for breast cancer. Recent data from the BOLERO-1 phase III clinical trials demonstrate an improvement in progression free survival (PFS) of 7.2 months in patients with HER2+/estrogen receptor negative (HER2+/ER-) advanced breast cancer when treated with trastuzumab and paclitaxel plus the mTORC1 inhibitor, everolimus (Afinitor ®). However, improvement in PFS was not observed in patients with HER2+/ER+ positive disease. These data suggest that hormone receptor levels may influence the response of HER2-amplifed cells to both HER2 and PI3K/mTOR directed therapies in the absence of hormonally directed therapies. In this study, we investigated the role of ER in HER2+ breast cancer by screening a large panel of HER2+/ER- and HER2+/ER+ breast cancer cell lines for responses to single agent and combined treatment with PI3K, mTOR, HER2 and ER-directed therapeutics.
Materials and Methods: The anti-proliferative activity of BKM120 (pan-PI3K inhibitor), BYL719 (p110α-specific) and everolimus were assessed as single agents or in combination with trastuzumab and/or tamoxifen in a panel of six HER2+/ER+ versus six HER2+/ER- breast cancer cell lines in two-dimensional culture. Drug response IC50s were generated from actual cell counts as measured by Z2-particle counting. Biomarker analyses were conducted using baseline mRNA microarray (Agilent) and reverse phase protein array (RPPA) profiling of each of the cell lines to determine associations with response or resistance data.
Results: RPPA analysis confirmed the presence of higher levels of ER protein in the cell lines designated as ER+ and significantly higher levels of PTEN protein were detected in those cell lines. Interestingly, each of the PI3K/mTOR pathway inhibitors tended to have increased single agent activity in the ER+ relative to the ER- lines. Combined activity with trastuzumab and either BKM120 or BYL719 was observed in 6 of the 12 cell lines tested and occurred independent of ER status. Similarly, combined activity of everolimus and trastuzumab was also observed in both HER2+/ER+ and HER2+/ER- cell lines. In 3/6 HER2+/ER+ cell lines the addition of tamoxifen provided no benefit to the combination of trastuzumab and PI3K/mTOR pathway inhibitor, whereas in two cell lines mild antagonism was observed with the triple combination. Finally, one cell line did show significant potentiation from the addition of endocrine therapy on top of HER2/PI3K/mTOR targeting.
Discussion: These data confirm levels of estrogen receptor are likely playing a role in response to single agent PI3K/mTOR pathway inhibition and highlight the potential utility of combining endocrine therapy with HER2/PI3K/mTOR-directed therapeutics in a sub-group of HER2-amplified breast cancers. Further in depth biomarker analyses may reveal additional molecular alterations responsible for this differential sensitivity to the double and triple combinations and is underway.
Citation Format: O'Brien NA, Nichols CM, Thomas J, Conklin D, Kalous O, Linnartz R, Di Tomasso E, Hurvitz SA, Hirawat S, Slamon DJ. Single agent and combined targeting of PI3K, mTOR, HER2 and ER signaling in a panel of HER2+/ER+ versus HER2+/ER- breast cancer cell lines. [abstract]. In: Proceedings of the Thirty-Eighth Annual CTRC-AACR San Antonio Breast Cancer Symposium: 2015 Dec 8-12; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2016;76(4 Suppl):Abstract nr P4-14-25.