593 Modulation of the F1F0-ATPase Induces Apoptosis of Mouse and Human Lamina Propria T-cells and Is Efficacious in Models of IBD

Abstract

Background: There is an increasing recognition that targeting immune metabolism may represent a new approach for therapeutic intervention in autoimmune diseases such as inflammatory bowel disease (IBD). Small molecules that allosterically modulate the activity of the mitochondrial F1F0-ATPase induce apoptosis in susceptible cells and have shown robust efficacy in several models of immune-mediated disease. Chronically activated lymphocytes have a distinctive metabolic phenotype that sensitizes these cells to ATPase Modulatorinduced apoptosis compared to acutely activated cells. This may provide an advantage over current less specific immunosuppressive treatments. In IBD, lamina propria (LP) lymphocytes are chronically activated, overexpress anti-apoptotic proteins and are resistant to activationinduced cell death (AICD). Restoring AICD of lymphocytes by inhibiting ATPase activity is a potential therapeutic strategy. Here, three ATPase Modulators were evaluated in murine IBD models and in LP cells from IBD patient biopsies.Methods: A relapsing-remitting model of chronic murine IBD was induced by intracolonic administration of 4-5 escalating doses of TNBS. ATPase modulators were dosed orally after 2-3 TNBS challenges. Mucosal LP mononuclear cells were isolated from mice treated with vehicle or LYC-51194 and analyzed for viability and inflammatory molecules by flow cytometry and RT-PCR. LP cells isolated from IBD patient biopsies were treated with LYC-51194 or anti-Fas mAb and viability assessed by flow cytometry. Results: Bz-423, LYC-51661 and LYC-51194 inhibited the F1F0-ATPase and induced lymphocyte apoptosis in vitro. In a relapsing-remitting model of chronic murine IBD, ATPase modulators did not show any overt toxicity and produced benefits including attenuated weight loss, normalized histology, decreased inflammatory cytokine production (IFNγ, IL-6, TNF) and increased the percentage of cell death in LP T cells (vehicle 22±6 vs LYC-51194 53±10 p<0.0009). To translate these findings to human biology, LP cells isolated from IBD patient biopsies were treated with LYC-51194 or antiFas mAb. In agreement with previously published results, LP T cells from inflamed areas from CD subjects showed resistance to Fas-induced cell death (% cell death: vehicle 22.5±7.7 n=10 vs anti-Fas 22±12 n=7). However, similar to what was observed in rodent disease models, LYC-51194 treatment was able to induce apoptosis in LP T cells from inflamed IBD patient samples (% cell death: vehicle 22.5±7.7 n=10 vs LYC-51194 49.3±8.1 n=9; p= 0.0003). Conclusions: Taken together, these results suggest that induction of apoptosis by application of ATPase Modulator compounds may be a promising approach for treating IBD.