Abstract 5655: AP26113 possesses pan-inhibitory activity versus crizotinib-resistant ALK mutants and oncogenic ROS1 fusions.
Authors: Squillace Rachel M. , Anjum Rana , Miller David , Vodala Sadanand , Moran Lauren , Wang Frank , Clackson Tim , Garner Andrew P. , Rivera Victor M. 
Published: August 2014

AP26113 is a potent, reversible inhibitor of ALK fusions and mutant (but not native) EGFR. To overcome mutation-based resistance, AP26113 was designed to maintain activity against crizotinib-resistant ALK variants such as the gatekeeper ALK mutant L1196M. In an ongoing phase 1 dose-escalation study, AP26113 has achieved steady-state trough concentrations in excess of 1 μM and demonstrated promising clinical activity in both crizotinib-resistant and naïve ALK-positive NSCLC patients and preliminary evidence of activity in patients with mutant EGFR (#439O, ESMO 2012). To further assess the activity of AP26113 against crizotinib-resistant ALK mutants, and the structurally related ROS1 fusions recently identified in NSCLC, we engineered Ba/F3 cells to express the appropriate oncogenic drivers and evaluated their sensitivities to crizotinib and AP26113.

Ba/F3 cell lines expressing clinically-identified EML4-ALK mutants (T1151T insertion, S1206Y, D1203N, L1196M, G1202R, F1174C and C1156Y) had substantially reduced sensitivity to crizotinib (viability IC50= 363-1296 nM) compared with native EML4-ALK (IC50= 137 nM). These data are consistent with the identification of these mutants in patients with acquired crizotinib resistance. AP26113 potently inhibited both native (IC50= 21 nM) and crizotinib-resistant ALK mutants (IC50= 26-254 nM) at concentrations substantially below the clinically achievable trough levels of AP26113 (1 μM), suggesting it may possess a pan-ALK inhibitory profile.

AP26113 also effectively inhibited the viability of Ba/F3 cells expressing CD74-ROS1 (IC50= 18 nM), FIG-ROS1 (IC50= 31 nM), SDC4-ROS1 (IC50= 16 nM) and EZR-ROS1 (IC50= 41 nM) thus demonstrating that it is an equipotent inhibitor of ALK and ROS1. In a Ba/F3 CD74-ROS1 xenograft model, AP26113 inhibited tumor growth in a dose-dependent manner, with 50 mg/kg AP26113 inducing substantial tumor regression. Since ROS1 and ALK are structurally related and similarly sensitive to crizotinib, we reasoned that acquired drug resistance in ROS1-driven tumors may also occur via mutation. We therefore generated Ba/F3 cells driven by L2026M gatekeeper mutant forms of CD74-ROS1 and FIG-ROS1 and tested their drug sensitivity. Interestingly, the inhibitory capacity of AP26113 was unaffected by L2026M. In contrast, crizotinib potencies were reduced approximately 4-fold.

In conclusion, we have demonstrated that AP26113 inhibits clinically relevant crizotinib- resistant ALK mutants and oncogenic ROS1 fusions recently identified in NSCLC irrespective of the fusion partner. Importantly, since AP26113 maintains potent activity against mutant forms of ALK and ROS1 at concentrations substantially below its human trough concentrations these data suggest that AP26113 may be able to prevent the emergence of drug-resistant mutants in NSCLC patients.

Citation Format: Rachel M. Squillace, Rana Anjum, David Miller, Sadanand Vodala, Lauren Moran, Frank Wang, Tim Clackson, Andrew P. Garner, Victor M. Rivera. AP26113 possesses pan-inhibitory activity versus crizotinib-resistant ALK mutants and oncogenic ROS1 fusions. [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 5655. doi:10.1158/1538-7445.AM2013-5655