Vol. 297, Issue 2, 573-581, May 2001

Effect of LY287045, a Thrombin/Trypsin Inhibitor, on Thrombin and Trypsin-Induced Aortic Contraction and Relaxation

Anindya Bhattacharya, Gerald F. Smith and Marlene L. Cohen

Eli Lilly and Company, Neuroscience Drug Discovery (A.B., M.L.C.) and Cardiovascular Drug Discovery (G.F.S.), Indianapolis, Indiana

The active site tripeptide arginal inhibitor of thrombin, LY287045, was used to study thrombin-induced aortic relaxation and contraction, two responses that differ both pharmacologically and physiologically. Although thrombin (10⁷ M) and trypsin (10⁶ M) were tachyphylactic upon repeated administration, trypsin contracted the aorta following thrombin-induced contraction. LY287045 (10⁷ M) attenuated thrombin-induced vasorelaxation, but not vasoconstriction with log K_B of 8.4. LY287045 (10⁷ M) also attenuated vasorelaxation, but not vasoconstriction to trypsin, another serine-protease with a thrombin-like catalytic triad, with similar potency (log K_B = 8.6) to that for thrombin. Consistent with these vascular effects, LY287045 inhibited the protease activity of both thrombin and trypsin. To explore further the selective inhibitory effect of LY287045 on protease-induced relaxation, we examined the effect of LY287045 on the nitric oxide and prostacyclin pathways and found that LY287045 did not alter vascular responses mediated by nitric oxide or prostacyclin. Likewise, LY287045 did not exert a direct inhibitory effect on the relaxant protease-activated receptor (PAR) since relaxation to the PAR-2-activating peptide was not blocked. The selective effect of LY287045 to inhibit only protease-induced endothelial-dependent relaxation demonstrated that protease inhibition will not affect all protease responses equally. Furthermore, increases in trypsin and thrombin have been associated with inflammation and angiogenesis. To the extent that these findings suggest that LY287045 exhibit dual protease inhibition of endothelial responses, LY287045 may have specific utility in hypotensive inflammatory diseases and in cancer metastases where both trypsin and thrombin have been implicated as causative agents.