The Design, Synthesis And Preliminary Activity Assay In Vitro Of L-iso-Glutamine Peptidomimetic Derivatives

Aminopeptidases are a group of exopeptidase which are widely distributed throughout the animal and plant kingdoms.There are a number of mammalian aminopeptidases which include aminopeptidase A,aminopeptidase B and aminopeptidase N.Aminopeptidases N(APN,CD13),one of membrane-bound zinc-dependent exopeptidase,is widely expressed in a wide variety of human tissues and cell types(endothelial,epithelial,fibroblast,leukocyte).In vivo,this enzyme is able to modulate bioactive peptide responses and to influence immune functions,at same time APN has been also reported to play an important role in the cell proliferation,secretion,invasion and angiogenesis of malignant tumor cells.It is also reported that APN is a receptor for corona-viruses TEGV and 229E in humans.All these findings make this enzyme as an interesting target for possible anti-tumor drugs research.Based on the 3D structures and binding models of some novel APN inhibitors in complex with APN,two series of L-iso-glutamine derivatives are designed with the aid of Chem3D Ultra 7.0 Program.All compounds are synthesized starting from the acylation reaction of 3,4-dichlorobenzoic acid with L-glutamine to form the basic scaffold,followed by the cyclic reaction in acetic anhydride and the key intermediate cyclic anhydride(compound 3)was obtained.The intermediate compound is condensed with different amino acids or primary amines,and produces the target compounds.The chemical structures of target compounds are identified by IR,ESI-MS and NMR(¹H-NMR,¹³C-NMR and HMBC)spectra.Preliminary bioactivity assays are carried out in vitro.Inhibitory activity of compounds against APN is measured with L-leucine p-nitroanilide as substrate.As a result,most of these newly synthesized compounds show good inhibitory activity on APN.Structure-activity relationships of fifteen tested compounds are elucidated and compounds containing aromatic heterocyclic substituents have higher activity than straight side chain;compounds para-substituted by electron withdrawing group such as chlorine and nitro group had weak inhibiting activity.To determine the selectivity of these inhibitors towards the targeted enzyme,inhibiting activity on gelatinase(MMP-2,9),which is one of endo-peptidases of Zn²⁺metallopeptidases, are measured by using succinylated gelatin as substrate.Data of the comparative studies exhibits the selectivity towards APN is higher than towards MMP-2,9.All the data strongly support that compounds containing carbonyl and hydroxamates show higher inhibiting activity than others significantly,and support the hypothesis that compounds inhibit APN by chelating to the zinc ion of this enzyme.In this thesis,APN inhibitors of L-iso-glutamate peptidomimetic derivatives have good inhibiting activity both on APN in vitro.A new and powerful APN inhibitor is found in preliminary screening and predicted to have good activity in vivo.