Potency assay (off target):
Selectivity of TYRA-300 for the human kinome was assessed with a KINOMEscan (Eurofins, San Diego, CA) against a panel consisting of 468 WT human kinases and mutant kinases at a test concentration of 100 nM. TYRA-300 inhibited 3% of the kinases >90% and 6% > 65% (S(90)468 = 0.03 and S(65)468 = 0.06) under these conditions.
The WT non-FGFR human kinases that were inhibited ≥90% in the single point assay were further characterized to determine enzymatic potency in a functional kinase assay using a 10-dose IC50 curve at 100 μM ATP concentration (Reaction Biology, Malvern, PA). VEGFR2 was not inhibited ≥90% in the kinome scan. However, as a key off-target for the discovery program, the IC50 was determined to be 325 nM (>200-fold selectivity for FGFR3).
