Abstract 790: Altiratinib is a potent inhibitor of TRK kinases and is efficacious in TRK-fusion driven cancer models

Abstract

Abstract Introduction: TRK kinases have been implicated in a variety of cancers, wherein TRK gene fusions, amplifications and mutations have been shown to drive tumor growth. TRK kinases have also been shown to play key roles in metastasis. Recently, large-scale sequencing efforts have identified TRK fusions at a low but significant frequency across all major cancers. Altiratinib is a potent single-digit nanomolar inhibitor of TRK, MET, TIE2, and VEGFR2 kinases. Altiratinib inhibits TRKA, TRKB, and TRKC phosphorylation in both WT TRK and TRK-fusion cell lines and inhibits cell proliferation in these cell lines. In vivo, altiratinib suppressed TRK phosphorylation for >18 hr after a single oral dose and showed significant inhibition of tumor growth in TRKA and TRKC-fusion xenograft efficacy studies. Altiratinib is currently in a Phase 1 clinical trial for patients with advanced solid tumors. Experimental procedures: Altiratinib was tested for inhibition of various recombinant kinases using a standard PK/LDH coupled spectrophotometric continuous assay. In cell assays, cells were treated with a dose response of compound. Levels of phosphorylated TRK in cell lysates were determined Western blot. Cell proliferation was measured using the fluorescent dye resazurin. Experiments were performed in triplicate. In vivo xenograft models were performed at Molecular Imaging, Inc. (Ann Arbor, MI). Summary of results: Altiratinib potently inhibited TRK kinase activity in biochemical assays (IC50 values of 0.9 nM, 4.6 nM, and 0.8 nM for TRKA, B, and C, respectively). Altiratinib inhibited phosphorylation of TRKA in the TPM3-TRKA fusion cell line KM-12 (IC50 = 1.4 nM) and of TRKC in ETV6-TRKC transformed NIH-3T3 cells (IC50 = 0.5 nM). Altiratinib inhibited NGF-stimulated phosphorylation of wild-type TRKA in K562 (IC50 = 0.7 nM) and SK-N-SH cells (IC50 = 1.2 nM) and BDNF-stimulated phosphorylation of TRKB in ATRA-transformed SK-N-SH cells (IC50 = 0.24 nM). In compound washout experiments, altiratinib inhibited TRKA phosphorylation in KM-12 cells for > 24 hr after compound washout, due to its durable Type II switch pocket binding mode to the kinase. In cell lines driven by TRK fusions, altiratinib also inhibited cell proliferation (KM-12 IC50 = 3.8 nM; NIH-3T3 ETV6/TRKC IC50 = 0.5 nM). In vivo, altiratinib inhibited phosphorylation of TRK and downstream targets in the signaling pathway for >12 hr after a single oral dose. Altiratinib treatment led to tumor regression in TRK-fusion xenograft efficacy studies. Conclusions: Altiratinib exhibits potent inhibition of oncogenic TRK fusions in vitro and in vivo. Combined with its inhibition of the tumor and microenvironment targets MET, TIE2, and VEGFR2 kinases, altiratinib provides the potential to treat cancers driven by TRK fusions. Altiratinib is currently in a Phase 1 clinical trial in patients with solid tumors. Citation Format: Bryan D. Smith, Cynthia B. Leary, Benjamin A. Turner, Michael D. Kaufman, Scott C. Wise, Maria E. R. Garcia-Rendueles, James A. Fagin, Daniel L. Flynn. Altiratinib is a potent inhibitor of TRK kinases and is efficacious in TRK-fusion driven cancer models. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 790. doi:10.1158/1538-7445.AM2015-790