Tyro-3, Axl, and Mer (abbreviated TAMs) are a family of three homologous receptor tyrosine kinases (RTKs) expressed predominantly on myeloid-derived hematopoietic cells and epithelial cells that function as inhibitory receptors that dampen inflammatory responses and maintain tissue tolerance. Both TAMs and their ligands (vitamin K activated proteins that consist of Gas6 and Protein S) are overexpressed in many human malignancies and expression is associated with more aggressive cancer staging, drug resistance, and poorer patient survival. Based on their clinical importance, there has been considerable effort to generate anti-TAM cancer therapeutics in the form of small molecule tyrosine kinase inhibitors, monoclonal antibodies, and soluble receptor traps that capture TAM ligands and prevent receptor activation. However, whereas several tyrosine kinase inhibitors have been reported, the high degree of sequence conservation in the TAM kinase domain is associated with lack of specificity and off target effects. To generate more selective inhibitors of TAMs, we have targeted the Ig1 domain of Axl, a ligand-binding region in the extracellular domain that gives TAMs their unique structural features. From the X-ray crystallographic structure of Gas6 in complex with the human Axl ectodomain (PDB 2c5d), two unique sites were targeted that include (i) the major binding pocket around Glu59 in Axl that interacts with Arg310 and Lys312 in Gas6, and (ii) the main hydrophobic interface between the Ig1 and the Lg1 that is required for Gas6-inducible dimerization. Virtual screening of the ZINC molecular data base was performed, and several drug-like compounds were selected based on ligand-Gas6 docking. We report on the characterization of two novel types of Axl inhibitors that include a triazolo pyridazine compound that targets the Ig1 binding pocket, and a 3-cyanocoumarin compound that targets the Ig1-Lg1 interface. Both compounds show activity on human Axl reporter lines, and therefore may represent a new class of Axl inhibitors with potential applications for anticancer therapy.
Citation Format: William J. Welsh, Vladyslav Kholodovych, Raymond Birge, Tom Comollo, Stanley Kimani, Kamal Singh. Novel Ig1 inhibitors of the Axl tyrosine kinase that block Gas6-inducible receptor activation. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 1748. doi:10.1158/1538-7445.AM2014-1748