Ultra-High Affinity Engineered Protein Therapeutics for Treating Metastatic Disease

We create engineered growth factor ligands and receptors that possess altered biochemical and biophysical properties with several goals in mind. For example, these engineered proteins are being used as tools for studying the relationship between properties such as binding kinetics and affinity and biological effects including cell signaling, proliferation, and migration. We also use growth factor ligands and receptors as starting points for the development of engineered protein therapeutics. Recent results on an engineered soluble receptor ‘decoy’, based on the Axl receptor tyrosine kinase extracellular domain, will be presented.  Aberrant signaling through the Axl receptor tyrosine kinase has been associated with a myriad of human diseases, most notably metastatic cancer, identifying Axl and its ligand Gas6 as important therapeutic targets. Using rational and combinatorial approaches, we engineered an Axl ‘decoy receptor’ that binds Gas6 with high affinity and inhibits its function, offering an alternative approach from drug discovery efforts that directly target Axl. Four mutations within this high-affinity Axl variant caused structural alterations in side chains across the Gas6-Axl binding interface, stabilizing a conformational change on Gas6. When reformatted as an Fc fusion, the engineered decoy receptor bound Gas6 with femtomolar affinity, an 80-fold improvement compared to binding of the wild-type Axl receptor, allowing effective sequestration of Gas6 and specific abrogation of Axl signaling. Moreover, increased Gas6 binding affinity was critical and correlative with the ability of decoy receptors to potently inhibit metastasis and disease progression in vivo.