Novel Paxillin-FAK Inhibitors for the Treatment of Liver Fibrosis

Abstract Chronic liver diseases like metabolic dysfunction-associated steatohepatitis (MASH), alcoholic liver disease (ALD), and viral hepatitis afflict more than 4.5 million Americans and are a major cause of death in those over 45. These diseases are associated with the development of liver fibrosis, which can lead to cirrhosis, hepatocellular carcinoma (HCC), and liver failure, necessitating liver transplantation or resulting in death. There are currently no FDA-approved therapies for modifying the progression of liver fibrosis, making this a significant, unmet medical need. Quiescent hepatic stellate cells (HSC) become activated after injury and produce excessive extracellular matrix (ECM) material. Paxillin is a key adaptor protein in recruiting multiple regulatory proteins, mediating downstream signaling, and linking the ECM to the cell cytoskeleton. Among its binding partners, focal adhesion kinase (FAK), a nonreceptor tyrosine kinase and scaffolding protein, is a highly promising target in liver fibrosis, along with paxillin, due to its key role in promoting HSC activation. Acting downstream of transforming growth factor beta-1 (TGF-β1), paxillin upregulation and FAK activation induce HSC activation, myofibroblast differentiation, the formation of stress fibers, and the upregulation of ECM proteins. Notably, paxillin downregulation and FAK inhibition attenuate liver fibrosis in animal models. Unfortunately, FAK-kinase domain inhibitors have shown only partial effects and poor selectivity in non-fibrosis indications, which is independent of its major role as a paxillin-binding partner and scaffolding protein. Furthermore, FAK-kinase inhibitors have shown significant toxicity due to the conserved ATP-binding pocket and off-target effects. Through the interaction with paxillin, FAK localizes to focal adhesions and regulates the focal adhesion complex. Therefore, peptide inhibitors targeting paxillin-FAK interaction may show greater efficacy and safety than FAK-kinase inhibitors. FAKnostics has identified a first-in-class series of peptidic inhibitors that directly target the paxillin-FAK interaction. FAKnostics’ novel peptides have 100-fold higher potency at binding the FAK than the native paxillin LD2 peptide motif. The ability of these candidates to inhibit the downstream effects of FAK activation has been confirmed in vivo in a syngeneic mouse model of melanoma. However, the effect of paxillin-FAK inhibition on the profibrogenic effects has not yet been fully evaluated. This Fast Track STTR aims to demonstrate proof-of- concept for use of these novel peptidic inhibitors in the treatment of liver fibrosis and to identify a preclinical candidate for future development. In Phase I, FAKnostics will optimize lead peptides to improve the biophysical/biochemical activity, characterize peptides in vitro, and confirm efficacy in an animal model of liver fibrosis. Phase II will include evaluation of ADMET properties, development of a long-lasting subcutaneous formulation, PK/safety studies, and comprehensive efficacy validation in liver fibrosis models. This will enable future completion of additional IND-enabling and manufacturing work to advance novel peptide drugs to the clinic.