Chemokines coordinate many aspects of leukocyte migration. As chemoattractants they play an important role in the innate and acquired immune response. There is good experimental evidence that N-terminal truncation by secreted or cell surface proteases is a way of modulating chemokine action. The localization of CD26/ dipeptidyl peptidase IV on cell surfaces and in biological fluids, its primary specificity, and the type of naturally occurring truncated chemokines are consistent with such a function.We determined the steady-state catalytic parameters for a relevant selection of chemokines (CCL3b, CCL5, CCL11, CCL22, CXCL9, CXCL10, CXCL11, and CXCL12) previously reported to alter their chemotactic behavior due to CD26/dipeptidyl peptidase IV-catalyzed truncation. The results reveal a striking selectivity for stromal cell-derived factor-1␣ (CXCL12) and macrophage-derived chemokine (CCL22). The kinetic parameters support the hypothesis that CD26/dipeptidyl peptidase IV contributes to the degradation of certain chemokines in vivo. The data not only provide insight into the selectivity of the enzyme for specific chemokines, but they also contribute to the general understanding of CD26/dipeptidyl peptidase IV secondary substrate specificity.

The feasibility of the fluoro-olefin function as a peptidomimetic group in inhibitors for dipeptidyl peptidase IV and II (DPP IV and DPP II) is investigated by evaluation of N-substituted Gly-Psi[CF=C]pyrrolidines, Gly-Psi[CF=C]piperidines, and Gly-Psi[CF=C](2-cyano)pyrrolidines. Of this later class, the (Z)- and (E)-fluoro-olefin analogues were prepared and chemical stability in comparison with the parent amide was checked. Most of these compounds exhibited a strong binding preference toward DPP II with IC(50) values in the low micromolar range, while only low DPP IV inhibitory potential is seen.

In this paper we report the systematic search for new, potent, and selective DPP II inhibitors. A study of the structure-activity relationship was conducted starting from aminoacyl pyrrolidides as lead compounds. Rational exploration of the P(1) and P(2) building blocks led to the discovery of some very potent DPP II inhibitors which can be characterized by their high selectivity for DPP II with regard to DPP IV. Dab-Pip and Dab-Pip-2-CN were selected as the most promising inhibitors (IC(50) nM range) and will enable us to study the physiological role of DPP II and to differentiate between DPP II and DPP IV in biological systems.

In this review the structural and functional aspects of dipeptidyl peptidase IV (DPP IV) will be described, and the therapeutic potential of DPP IV inhibitors will be highlighted. DPP IV will be situated in clan SC, a group of serine proteases that contains several proline specific peptidases. Structural aspects of DPP IV and its interaction with different types of inhibitors are recently revealed by the publication of several crystal structures. Especially the design and development of new DPP IV inhibitors based on the three-dimensional structure, substrate specificity and catalytic mechanism will be discussed. In the last years there was an important development of new pyrrolidine-2-nitriles with very promising therapeutic properties for the treatment of type 2 diabetes. The role of DPP IV in peptide metabolism of members of the PACAP/glucagon peptide family, neuropeptides and chemokines has been thoroughly investigated during recent years. This is directly related to the promising therapeutic potential of DPP IV inhibitors in the treatment of type 2 diabetes and in the treatment of immunological disorders. Several inhibitors are currently under investigation in clinical trials for the treatment of type 2 diabetes and represent a new class of drugs for the treatment of this disease.

The presence of DPPII (dipeptidyl peptidase II; E.C. 3.4.14.2) has been demonstrated in various mammalian tissues. However, a profound molecular and catalytic characterization, including substrate selectivity, kinetics and pH-dependence, has not been conducted. In the present study, DPPII was purified from human seminal plasma to apparent homogeneity with a high yield (40%) purification scheme, including an inhibitor-based affinity chromatographic step. The inhibitor lysyl-piperidide (K(i) approximately 0.9 microM at pH 5.5) was chosen, as it provided a favourable affinity/recovery ratio. The human enzyme appeared as a 120 kDa homodimer. Mass spectrometric analysis after tryptic digestion together with a kinetic comparison indicate strongly its identity with QPP (quiescent cell proline dipeptidase), also called dipeptidyl peptidase 7. pH profiles of both kcat and kcat/K(m) clearly demonstrated that DPPII/QPP possesses an acidic and not a neutral optimum as was reported for QPP. Kinetic parameters of the human natural DPPII for dipeptide-derived chromogenic [pNA (p-nitroanilide)] and fluorogenic [4Me2NA (4-methoxy-2-naphthylamide)] substrates were determined under different assay conditions. DPPII preferred the chromogenic pNA-derived substrates over the fluorogenic 4Me2NA-derived substrates. Natural human DPPII showed high efficiency towards synthetic substrates containing proline at the P1 position and lysine at P2. The importance of the P1' group for P2 and P1 selectivity was revealed, explaining many discrepancies in the literature. Furthermore, substrate preferences of human DPPII and dipeptidyl peptidase IV were compared based on their selectivity constants (kcat/K(m)). Lys-Pro-pNA (k(cat)/K(m) 4.1x10(6) s(-1) x M(-1)) and Ala-Pro-pNA (kcat/K(m) 2.6x10(6) s(-1) x M(-1)) were found to be the most sensitive chromogenic substrates for human DPPII, but were less selective than Lys-Ala-pNA (kcat/K(m) 0.4x10(6) s(-1) x M(-1)).