P135 ORALLY ADMINISTERED FOLH1/GCPII INHIBITORS AS NOVEL THERAPEUTICS IN INFLAMMATORY BOWEL DISEASE (IBD)

Abstract

The folate hydrolase gene (FOLH1), which encodes for the enzyme glutamate carboxypeptidase II (GCPII), has been implicated in the pathogenesis of inflammatory bowel disease (IBD) where it exhibits a robust 300-1000% increase in enzymatic activity in intestinal biopsies collected from Crohn’s Disease and Ulcerative Colitis patients. We have previously shown that systemic administration of the GCPII inhibitor 2-PMPA (IC50=300 pM) led to significant improvement in disease activity index in three commonly used murine models of IBD: DSS-colitis, TNBS-colitis and spontaneous colitis of IL10-/- mice. While systemic administration of 2-PMPA was effective in preclinical models, in the clinic an oral dosing regimen would be preferred. 2-PMPA is a highly polar compound with minimal oral bioavailability and tissue penetration. Despite displaying systemic restriction in normal animals, we hypothesized that oral administration might provide direct local therapy to inflamed colonic epithelia and be equally effective to systemic dosing in murine colitis models. Age-, gender- and weight-matched wild-type C57Bl/6NHsd mice were exposed to 2.5% DSS in drinking water for 5 days to induce colitis followed by a 2-day exposure to fresh water. Body weight loss, stool consistency and presence of rectal bleeding were scored daily and summated to generate a disease activity index. Daily oral 2-PMPA (0.01 – 100 mg/kg) or vehicle (HEPES buffered saline) was administered. Mice were euthanized at multiple time points (study days 0, 3,5 and 7) and tissues were harvested for pharmacokinetic analysis. Oral 2-PMPA significantly reduced disease activity index in DSS-colitis at doses as low as 1 mg/kg, which is 100-fold lower than the dose utilized for systemic treatment (p<0.01, n=20/group). However, DSS treatment resulted in loss of epithelial barrier integrity and caused increased systemic 2-PMPA plasma exposures, thus we cannot fully determine whether the mechanism of action of oral 2-PMPA was locally- versus systemically-mediated in this model. This is relevant as the barrier disruption after acute DSS treatment in mice is likely to be more severe than what is observed clinically in IBD patients. Therefore, we embarked upon further studies to explore whether 2-PMPA has a direct activity on colonic epithelium. Pilot studies in Caco2 monolayers show that 2-PMPA protects against LPS-induced alterations in epithelial barrier permeability as measured by TEER (p<0.05) and attenuates RNA expression of TNFα (p<0.05), providing support that 2-PMPA has direct effects on colon epithelial cells. The discovery that orally administered 2-PMPA displays robust and potent efficacy in the murine DSS-model, with preliminary evidence of direct epithelial actions, provides strong support that FOLH1/GCPII is a clinically relevant, druggable, target in IBD.