Preclinical Validation of Novel Gut-Restricted LRRK2 Inhibitors as Therapeutic Leads for IBD

SUMMARY Therapies to treat Crohn's disease (CD) and ulcerative colitis (UC), the types of inflammatory bowel disease (IBD) characterized by severe chronic inflammation of the gastrointestinal tract, have greatly improved over the past three decades; yet, a large proportion of IBD patients fail to achieve sustained remission. Genetic biomarkers may offer an avenue toward the dissecting heterogeneity of disease etiology, and improving personalized therapeutic approaches. Our group has recently identified a coding gain-of-function mutation in the LRRK2 (leucine-rich repeat kinase 2) gene that conferred a ~70% increased CD risk and affected CD age of onset, disease location, LRRK2 kinase activity, and autophagy flux. Importantly, LRRK2 has also attracted considerable attention for its causal link to Parkinson's disease (PD). Further analyses identified dozens of additional variants within LRRK2 that were associated with the risk of both CD and PD, suggesting shared pathogenesis between these diseases. Given extensive efforts to target LRRK2 kinase activity as a means to treat PD, we explored known LRRK2 inhibitors developed for PD as potential therapies for IBD. We showed that selective LRRK2 inhibitors have ameliorated experimental colitis and reduced inflammatory cytokine TNF-α levels, a hallmark of IBD-associated inflammation, in dendritic cells of IBD patients. However, studies of LRRK2 kinase inhibition in preclinical models suggest that brain penetration and toxicity in peripheral tissues may be a critical safety liability for these inhibitors in IBD patients. To advance this therapeutic hypothesis, we have developed structure-based approach to design novel gut-restricted LRRK2 inhibitors, synthesized prototype inhibitors, crystallized analogs in complex with kinases mutants that mimic LRRK2 binding site and tested their LRRK2 potency and efficacy, which are maintained. Thus, we are proposing, starting from known LRRK2 inhibitor structures to 1) Develop proof-of-concept gut-restricted LRRK2 inhibitors as potential IBD therapeutics using structure-based drug design and medicinal chemistry; 2) Evaluate novel compounds for LRRK2 inhibition, in vitro absorption, distribution, metabolism, excretion and pharmacokinetics, off-target selectivity, and effects on cellular substrate phosphorylation and cytokine activity using biochemical and cell-based assays, and 3) Pre- clinically validate new inhibitors for in vivo absorption, distribution, metabolism, excretion and pharmacokinetics, plasma exposures, and effects on inflammatory biomarkers and severity of experimental colitis. Our hypothesis is that gut-restricted LRRK2 inhibitors could be a safe and effective therapeutic target of IBD therapy in the presence or absence of IBD-associated LRRK2 mutations. These studies will provide a basis for future clinical trials aimed at testing LRRK2 as an effective drug target for IBD. We believe our strong preliminary data and relevant therapeutic hypothesis align well with the research goals and objectives of PAR-19-294 and will help improve the current standard of clinical care for patients with this disease of interest to the National Institute of Diabetes and Digestive and Kidney Diseases.