Glutamic acid analogues as potent dipeptidyl peptidase IV and 8 inhibitors I-Lin Lu, a,b,  Shiow-Ju Lee, a,  Hsu Tsu, a,  Su-Ying Wu, a Kuo-His Kao, a Chia-Hui Chien, a Ying-Ying Chang, a Yuan-Shou Chen, a Jai-Hong Cheng, a Chung-Nien Chang, a Tung-Wei Chen, a Sheng-Ping Chang, c Xin Chen a, * and Weir-Torn Jiaang a, * a Division of Biotechnology and Pharmaceutical Research, National Health Research Institutes, National Health Research Institutes, No. 35, Keyan Rd., Zhunan Town, Miaoli Country 350, Taiwan, ROC b Graduate Institute of Life Sciences, National Defense Medical Center, No.161, Section 6, Min-Chuan East Road, Taipei 114, Taiwan, ROC c Department of Obstetrics and Gynecology, Taipei Veterans General Hospital, No. 201, Sec. 2, Shih-Pai Road, Taipei 112, Taiwan, ROC Received 1 April 2005; revised 15 April 2005; accepted 22 April 2005 Available online 31 May 2005 Abstract—To find potent and selective inhibitors of dipeptidyl peptidase IV (DPP-IV), we synthesized a series of 2-cyanopyrrolidine with P2-site 4-substituted glutamic acid derivatives and tested their activities against DPP-IV, DPP8, and DPP-II. Analogues that incorporated a bulky substituent at the first carbon position of benzylamine or isoquinoline showed over 30-fold selectivity for DPP- IV over both DPP8 and DPP-II. From structure–activity relationship studies, we speculate that the S2 site of DPP8 might be similar to that of DPP-IV, while DPP-IV inhibitor with N-substituted glycine in the P2 site and/or with a moiety involving in hydrophobic interaction with the side chain of Phe357 might provide a better selectivity for DPP-IV over DPP8. Ó 2005 Elsevier Ltd. All rights reserved. Dipeptidyl peptidase IV (DPP-IV, also known as CD26) (EC 3.4.14.5) is a drug target for type II diabetes. It is a prolyl dipeptidase involved in the in vivo degradation of two insulin-sensing hormones, glucagon-like peptide 1 (GLP-1) and glucose-dependent insulinotropic polypep- tide (GIP), by cleaving at the peptide bond of the penul- timate position. 1,2 DPP-IV knockout mice and rats have consistently displayed healthy phenotypes. 3 When chal- lenged with a high concentration of glucose, these ani- mals show improved glucose tolerance, enhanced insulin secretions, and increased circulating active GLP-1 peptide. 3 By extending the duration of action of GLP-1, DPP-IV inhibitors stimulate insulin secretion, inhibit glucagon releases, and slow gastric emptying in animal models, each action a benefit in the control of glucose homeostasis. 4 Recent successes in clinical trials of several DPP-IV inhibitors have demonstrated that selective and potent inhibitors of DPP-IV protease are effective for the treatment of type II diabetes. 5 DPP-IV belongs to the prolyl dipeptidase family, which includes DPP-II, DPP-IV, DPP8, and DPP9. 6 The in vivo functions of these prolyl dipeptidases, other than DPP-IV, are largely unknown. DPP-II is known as qui- escent cell proline dipeptidase or DPP-VII (DPP7). 6,7 The inhibition of DPP-II has been shown to result in the apoptosis of quiescent T cells. 8 DPP8 is a cytoplas- mic protease with a 51% homology with DPP-IV at the amino acid level. 9 The administration of selective DPP8/DPP9 inhibitors in animals results in severe toxic reactions, including alopecia, thrombocytopenia, ane- mia, enlarged spleen, multiple histological pathologies, and increased mortality. 10 In light of the physiological importance of prolyl cleaving enzymes and the toxicity associated with DPP8/DPP9, it is important to have selective inhibitors targeting DPP-IV for the treatment of type II diabetes. In the current drug design of inhibitors, the selectivity of inhibitor over other closely related enzymes must be 0960-894X/$ - see front matter Ó 2005 Elsevier Ltd. All rights reserved. doi:10.1016/j.bmcl.2005.04.051 Keywords: DPP-IV; DPP8. * Corresponding authors. Tel.: +886 37 246166x35712; fax: +886 37 586456; e-mail addresses: xchen@nhri.org.tw; wtjiaang@nhri.org.tw   These authors contributed equally. Bioorganic & Medicinal Chemistry Letters 15 (2005) 3271–3275