Biomedical Research 22 (5) 257-260, 2001 Inhibition of enteropeptidase by antitrypsin drugs Masashi 1VlATSUSHIMA,1’2’4 Naohisa YAHAGI,‘ Masao lcHiNosE,"5 Kazumasa Mu<r,"6 Masao OMATA,‘ Hideshi 1NOUE,2’3 Takayuki TA1<.AHAsH1,2” and Kenji TAKAHASHI2’3 ‘Department of Gastroenterology, Faculty of Medicine and 2Department of Biophysics and Biochemistry, Graduate School of Science, The University of Tokyo, Bunkyo-ku, Tokyo 113-0033 and 3 Department of Molecular Biochemistry, School of Life Science, Tokyo Uni- versity of Pharmacy and Life Science, 1432-1 Horinouchi, Hachioji, Tokyo 192-0392, Japan (Received 20 November 2001; and accepted 29 November 2001) ABSTRACT We have investigated the inhibitory effects on porcine enteropeptidase of some typical synthetic amidino- and guanidino-compounds developed as antitrypsin drugs. The results indicated that en- teropeptidase is inhibited by these compounds almost as strongly as trypsin; their IC50 values were in the range of 0.008-4 MM, which were much lower than that of the well-known tryp- sin/enteropeptidase inhibitor benzamidine. Enteropeptidase [EC 3.4.21.9] is a key enzyme re- then activates several other zymogens including tryp- sponsible for the initial activation of trypsinogen to sinogen secreted from the pancreas into the duode- trypsin in the duodenum (8, 9). The resulting trypsin num. A genetic deficiency of this enzyme leads to malabsorption and malnutrition, especially in in- This study was supported in part by grants-in-aid for sci- fancy (3, 4). Recently, we have found that enteropep- entific research from the Ministry of Education, Sci- tidase is strongly inhibited by benzamidine and 1- ence, Sports and Culture of Japan. z‘rcms-epoxysuccinyl-L-leucylamido (4-guanidino) bu- Correspondence to: Dr Kenji Takahashi at the above ad- tane (E-64) in a reversible and competitive manner dress. (11). E-64 has been known as a cysteine proteinase Tel: +81-426-76-7146; Fax: +81-426-76-7149; inhibitor, but was found to inhibit the serine prote- E-mail: [email protected]. inase enteropeptidase, due to the presence of a gua- Present address: 4 Department of Internal Medicine, To- nidinium group. On the other hand, various orally kai University School of Medicine, Kanagavva 259-1193. active synthetic trypsin inhibitors having a guanidin- 5 Second Department of Internal Medicine, Wakayama ium or amidinium group(s), such as FOY-305 (ca- Medial University, Wakayama 641-0012. 6 First Department of Internal Medicine, Toho Univer- mostat) (5), have been developed as potential drugs sity School of Medicine, Tokyo 143-8541. for treatment of such a disease as acute and chronic i Graduate School of Science, Hokkaido University, Sap- pancreatitis, and some of them have been used clini- poro 060-0810. cally (6, 15, 16). Pancreatitis is thought to be caused Abbreviations: FO-349-01, N-al1yl-N-[4-[(4-amidinophe- by the activated trypsin and other trypsin-activated noxy)carbonyl]- oz -methylcinnamoyl]glycine methanesul- proteinases in the pancreas. The mechanism of tryp- fonate; FOY-007, ethyl-4-(6-guanidinohexanoyloxy) sinogen activation in pancreas is not clear. It may be benzoate methanesulfonate; FOY-251, 4-(4-guanidino- possible that an enteropeptidase-like enzyme is in- benzyloxy)pheny1acetate methanesulfonate; FOY-305, volved in this activation. In this connection, it is (N, N-dimethylcarbamoyl)methy14-(4-guanidinobenzy- thought to be important to examine whether those loxy)phenylacetate methanesulfonate; FUT-175, 6- amidino-2-naphthyl 4-guanidinobenzoate dimethanesul- antitrypsin drugs inhibit enteropeptidase. In this fonate; ONO-3403, N-ally1-N-[4-[(4-amidinophenoxy) study, we have examined the inhibitory effect of carbonyl]- (B-methylcinnamoyl]glycine ethyl ester me- such compounds on porcine enteropeptidase, and thanesulfonate. found that they do inhibit the enzyme almost as .