Fragments vs AXL, with help from an ELF

The receptor tyrosine kinase AXL has been implicated in multiple cancers, and more recently as a possible target for COVID-19. The protein has been pursued by many groups, mostly starting from high-throughput screens or existing inhibitors. In a recent Bioorg. Med. Chem. paper Pearly Ng, Alvin Hung, and collaborators at Experimental Drug Development Centre Singapore, University of Sussex, and LigatureTX describe a fragment-based approach.

 

The researchers started with a biochemical screen of 1700 fragments at 500 µM. Subsequent dose-response experiments yielded 16 fragment hits with IC₅₀ < 400 µM, all of which are shown in the paper. Four of these were based on 7-azaindole, a kinase hinge-binding motif that has previously been used in AXL inhibitors.

 

Seeking novelty, the researchers focused on the remaining 12 fragments and turned to their “expanded library of fragments (ELF),” which they have been building for several years. Though not described in depth, this collection was designed such that near-neighbors of their primary library could be rapidly screened to establish SAR and identify productive growth vectors. Compound 11, an indazole, led to several analogs from the ELF such as compound 24. This was docked into a publicly available crystal structure of AXL, and structure-based design led to compound 32. Further optimization ultimately led to molecules such as compound 52, the most potent in the paper.

 

Although compounds in this series were potent in the biochemical assay, they were typically 100-1000 less potent in a cell-based NanoBRET target-engagement assay, an annoying but not uncommon phenomenon. Several of the compounds showed high nanomolar activity in an ovarian cancer cell line with AXL overexpression. Interestingly, two compounds showed considerably more potent antiproliferative activity than target-engagement, and the researchers speculate they may be hitting other kinases. In support of this hypothesis, the researchers found that one of their compounds inhibited 12 of 97 kinases by >90% at 1 µM. This is not surprising given that indazoles are privileged fragments for kinases. Mouse pharmacokinetic studies revealed poor oral exposure for most of the compounds, though this could be improved by decreasing the basicity of the molecules.

 

A nice aspect of this paper is that most of the work was done without the benefit of structural information. Crystallography attempts with AXL and any of the ligands were unsuccessful, though the researchers were eventually able to obtain a structure of one of their more potent molecules bound to a related kinase. And just in time for the holidays, the paper also illustrates the utility of having an ELF on your shelf.