Molecular interactions important for Axl activation

AXL, a receptor tyrosine kinase, is found at high levels in many human cancers, and the AXL signal modulates cancer cell survival, proliferation, and migration. The AXL signal also affects how the immune system responds to cancer. Given its role in many cellular processes across various malignant tissue types, it is critical to better understand AXL signaling in tumor cells and surrounding tissue. This project aims to improves the understanding of the molecular mechanism of interaction of AXL with GAS6, PtdSer and the therapeutic antibody tilvestamab using a structural biology approach. Cryo-electron microscopy and X-ray crystallography are used for studying the interactions. Following onset of the global pandemic, AXL was identified as a possible interaction partner of the Covid spike protein. A collaboration with Professor Wendy Maury at the University of Iowa confirmed that AXL mediates enhancement of Covid-19 infection, and this project showed that the interaction is not through a direct interaction with the virus spike protein but rather through a lipid in the viral membrane. This finding is currently in press in the scientific journal PLoS Pathogens.

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The receptor tyrosine kinase Axl regulates vital physiological processes including cell survival, cell proliferation, migration, motility, epithelial to mesenchymal transition, and immunologic response. Axl activation is associated with invasiveness and resistance to therapy in many cancers and has recently been shown to play a key role in a range of fibrotic diseases. Axl is activated by its ligand, growth arrest-specific protein 6 (Gas6). Optimal activation of Axl requires clustering of Gas6 through binding to the lipid phosphatidylserine (PtdSer), found on the membranes of dying cells. This interaction depends on the Vitamin K-dependent post-translational modification of several glutamate residues near the N-terminus of Gas6 to ?-carboxyglutamate. In this project, in order to better understand extracellular events in Axl signaling, the structure of the entire extracellular domain of Axl receptor in complex with Gas6 and phosphatidylserine will be determined. BerGenBio is focussed on the pre-clinical and clinical development of Axl inhibitors, including a high affinity function-blocking antibody tilvestamab that targets the N-terminal domain of Axl. To provide insight into the mode of action of tilvestamab, the crystal structure of Axl extracellular domain in complex with tilvestamab will also be determined. Key R&D challenges with this project will be the production of sufficient quantities of the individual protein components and finding appropriate conditions for the growth of well-diffracting crystals for X-ray structure determination. Gas6 is expected to be especially challenging due to the difficulty in ensuring complete ?-carboxylation. The anticipated results will provide us with a clearer understanding of the molecular details of Gas6 activation of Axl, and how this activation can most effectively be inhibited therapeutically.