Targeting of the AXL receptor tyrosine kinase by small molecule inhibitor leads to AXL cell surface accumulation by impairing the ubiquitin-dependent receptor degradation

AXL TKI BMS777607 treatment increased AXL protein levels after 24 h as observed by Western blot and flow cytometry analysis. Mechanistically, this inhibition-induced AXL cell surface accumulation was neither associated with epigenetic modifications, nor altered transcriptional and translational regulation. Further, we saw no impact on glycosylation and receptor shedding by α-secretases. However, we observed that BMS777607 increased the glycosylated 140 kDa AXL protein abundance, which was impaired in the kinase dead mutant AXL (K567R). We demonstrated that AXL kinase activity and subsequent kinase phosphorylation is necessary for GAS6-dependent receptor internalization and degradation. Blocking of kinase function by BMS777607 resulted in ubiquitination prohibition, impaired internalization and subsequent cell surface accumulation. Subsequently, AXL cell surface accumulation was accompanied by increased proliferation of 3D-Speroids induced by low μM levels of BMS777607 treatment.