Vol. 297, Issue 2, 811-818, May 2001

Characterization of the Caspase Inhibitor IDN-1965 in a Model of Apoptosis-Associated Liver Injury

Niel C. Hoglen, Brad P. Hirakawa, Craig D. Fisher, Suzanne Weeks, Anu Srinivasan, Angela M. Wong, Karen L. Valentino, Kevin J. Tomaselli, Xu Bai, Don S. Karanewsky and Patricia C. Contreras

Idun Pharmaceuticals, Inc., La Jolla, California

Previous studies have shown that caspase inhibitors are effective at protecting against anti-Fas antibody (-Fas)-mediated liver injury/lethality. The purpose of these experiments was to characterize more fully the efficacy of a broad-spectrum, irreversible caspase inhibitor, IDN-1965 (N-[(1,3-dimethylindole-2-carbonyl)valinyl]-3-amino-4-oxo-5-fluoropentanoic acid), in this model and the role of caspase inhibition in long-term protection. The ED₅₀ for IDN-1965 by i.p. administration, based on alanine aminotransferase activities, was 0.14 mg/kg. The caspase inhibitor was also efficacious when administered intravenously and orally (ED₅₀ values of 0.04 and 1.2 mg/kg, respectively). Histologically, marked reduction in Fas-induced apoptosis with IDN-1965 (1 mg/kg, i.p.) was apparent at 6 h. Also, caspase 3-like activities were decreased in a dose-dependent manner, but the inhibition of caspase activity was transient. Immunohistochemical studies demonstrated that IDN-1965 greatly reduced the activation of caspase 3. In survival studies, a single i.p. treatment of 1 mg/kg IDN-1965 or continuous i.p. infusion via osmotic pumps completely blocked lethality measured up to 7 days after -Fas administration. IDN-1965 was also effective in inhibiting liver injury when administered as long as 3 h after or 1 h before -Fas administration. Lastly, Western blot analysis demonstrated that processing of caspases 3, 6, and 8, as well as Bid (a protein responsible for the release of mitochondrial cytochrome C and amplification of the apoptotic cascade) was inhibited by IDN-1965. In conclusion, the broad-spectrum caspase inhibitor IDN-1965 is markedly effective at inhibiting Fas-mediated apoptosis by multiple routes of administration. The therapeutic potential of caspase inhibitors appears promising for the treatment of apoptosis-mediated liver injury based on potency and postinsult efficacy.