Abstract P4-08-01: PI3K/mTOR inhibition overcomes in vitro and in vivo trastuzumab resistance independent of feedback activation of pAKT
Authors: O'Brien NA , McDonald K , Tong L , Von Euw E , Conklin D , Kalous O , Di Tomaso E , Schnell C , Linnartz R , Hurvitz SA , Finn RS , Hirawat S , Slamon DJ 
Published: August 2014

Background: Aberrant activity of the PI3K/mTOR pathway has been implicated in resistance to trastuzumab and anti-hormonal therapy for HER2-amplified and ER-positive breast cancer, respectively. Previous studies in our laboratory and others have shown that increased PI3K/mTOR signaling, either through PTEN loss or activating PIK3CA mutations, can confer resistance to trastuzumab therapy. In this study, we assessed the potential of targeting the PI3K/mTOR pathway in overcoming both de novo and acquired trastuzumab resistance.

Materials and Methods: The in vitro activity of the pan-PI3K inhibitor BKM120, the mTORC1 inhibitor RAD001 and the dual PI3K/mTORC1/2 inhibitor BEZ235 were evaluated in a panel of 49 human breast cancer and immortalized cell lines. The in vivo activity of these molecules was assessed in six cell line xenografts models representing, ER+/HER2− (KPL-1, ZR75-1), ER+/HER2+ (UACC812, MDA361), ER−/HER2+ (SUM190), PIK3CA mutant (SUM190, MDA361), PTEN-null (ZR75-1) and trastuzumab resistant (BT-TR) breast cancer. Finally, the effect of PI3K/mTOR inhibition on feedback activation of PI3K signaling and compensatory pathways was measured by Western blot, immunohistochemistry (IHC) and reverse phase protein analysis (RPPA) of control and treated cell lysates/tumors.

Results: Using a sensitivity cut-off of an IC50 of &lt; 1 µmol/L, 16 of the 18 HER2-amplified breast cancer cell lines and 8 of 10 cell lines with activating mutations in PIK3CA were sensitive to the pan-PI3K inhibitor BKM120. BEZ235 showed the most potent efficacy across the panel with IC50s &lt; 100 nmol/L for each of the 49 cell lines tested. The HER2-amplified/PIK3CA mutant cell lines were also unexpectedly sensitive to the mTORC1 inhibitor RAD001, this was despite the silencing of mTORC1 signaling being followed by a feedback increase in phospho-AKT signaling. Furthermore, each of these molecules showed remarkable in vivo activity across the panel of xenografts models. BKM120, RAD001 and BEZ235 induced both tumor stabilization and regression independent of PTEN, PI3K, ER and HER2 status. Pharmacodynamic analysis of tumor tissue revealed that BEZ235 and RAD001 both inhibited mTORC1 signaling as indicated by a reduction in the levels of phosphorylated ribosomal protein S6 (pS6). However, in contrast to BKM120 and BEZ235, RAD001 did not induce a reduction in the levels of pAKT (S473 or TH308) yet showed comparable in vivo efficacy to each of these molecules. Finally, combined targeting of HER2 and PI3K/mTOR in vitro increased the anti-proliferative activity of the molecules and led to an increased induction of apoptosis. The efficacy of these molecules (alone or in combination with trastuzumab) was assessed in a model of in vivo trastuzumab resistance generated through long-term treatment of the trastuzumab sensitive BT474 cells (BT-TR). All PI3Ki induced complete inhibition of tumor proliferation in monotherapy, while the combination of trastuzumab and each of these molecules induced tumor regression in the trastuzumab resistant tumors.

Discussion: These pre-clinical data indicate that targeting the PI3K/mTOR pathway either alone or in combination with trastuzumab is effective strategy for overcoming trastuzumab resistance.

Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P4-08-01.