Design, Synthesis And Activity Evaluation Of α-Halogen Substituted Succinate-Based Hydroxamic Acids As Matrix Metalloproteases Inhibitors

Matrix metalloproteinases (MMPs) are a family of Zn-dependent endopeptidases involved in the degradation and repair of the major components of the extracellular matrix of connective tissues. Zn2+ plays an importat role in the hydrolysis process. The overexpression of these proteinases is associated with a number of pathological processes such as arthritis, cancer and emphysema. Thus, matrix metalloprotease inhibitors (MMPIs) are expected to be useful for the treatment of these disorders. Now MMPIs have become one of the focuses in the field of drug research and development.Succinyl hydroxamate is the most important category of small molecule MMPIs, which has been studied widely. Hydroxamic acid is called zinc-binding group, which could chalate Zn2+ in catalytic domain of MMPs and devitalize MMPs. Modification of the positionαto hydroxamic acid is the common strategy for designing succinyl hydroxamates as small molecule inhibitors of MMPs. Most of the previous studies in this aspect were characterized with improving molecule's pharmacokinetics.This thesis was focused on the design, syntheses and activity evaluation of novelα-halogen substituted succinate-based hydroxamic acid compouds as MMP inhibitors. The principal design idea is introducing halogen to theα-position of hydroxamic acid, which is expected to increase hydroxamic acid's acidity through electron-withdrawing effect of halogen, so strengthen it's affinity with proteinases'catalytic Zn2+ and potentialize compoud'inhibitory activity. Moreover halogen can also improves molecule'lipophilicify. This idea was first drew, which hasn't occurred in any published literatures up to now.Our strategy to discover new inhibitors is as follows: succinyl hydroxamate is selected as parent structure, the R1 substituent is fixed to an isobutyl group. In the prerequisite of introducingα-halogen, modifications of the R2 substituent and R3 substituent are examined. R2 substituent is made with natural or unnatural aromatic amino acids, while various hydrophilic or hydrophobic groups are used as R3 substituents. General formula of target compouds is as follow: The target componds were synthesized from amino acid derivates and 4-methylpentanoyl chloride by seven steps of acylation, asymmetric alkylation, hydrolysis, halonation, condensation, deprotection of tert-butyl ester and condensation with hydroxylamine. Among them, the key step is introducing halogen, which was achieved by treating succinic acid monoester derivate with LDA and electrophilic halonation reagents (CCl4, CBr4, N-fluorobenzenesulfonimide). There produced a new chiral centre in theα-position of ester after introducing one halogen atom.Reasoning from NMR spectrums of intermediates, it is very likely that the chlorination and fluorination reactions are stereoselective and could produce the products with single configuration. But the absolute configuration of new chiral carbon needs to be determined by further reaserch. While bromination reaction is non-stereoselective and the products are mixture of two diastereoisomers. 44α-halogen substituted succinate-based hydroxamic acid compouds were obtained, which were all new compouds and included 15α-fluoride, 23α-chloride, 4α-bromide (two pairs of epimers atα-carbon of hydroxamic acid), 2α-disfluoride substituted compouds. The structures of them were confirmed via MS and 1H NMR. Purities were determined by HPLC.All of new compounds'inhibitory activities against MMP-2 and MMP-9 were tested in vitro. Ilomastat, a broad and potent succinate hydroxamate MMPI, was selected as the positive control. Among these compounds, seven of them exhibited similar inhibitory activities against MMP-2 and seven of them showed similar inhibitory activities against MMP-9 comparing respectively with Ilomastat. The activity results indicated the following initial SAR: (1) It's likely that introducing halogen hasn't significant effect on inhibitory activity. Compouds with two fluoride atoms atα-position showed weaker inhibitory activities than their single fluoride substitute counterparts, so whether introducing electron-withdrawing groups to the neighboring region of the hydroxamate facilitates improving compouds'inhibitory activity should be further examined; (2)According to available activity data, configuration of the new chiral centre arising from introducing halogen perhaps has nothing to do with the inhibitory activity; (3)Compouds, possessing hydrophilic groups, methyl or mehyl analogs as the R3 substituents, appeared to have improved potency as compared to those with hydrophobic groups as the R3 substituents, when R3 is hydrogen, the inhibitory activity became weaker; (4)Planar fused-ring aryl groups as the R2 substituents perhaps facilitate improving compouds'inhibitory activity; (5)The two amide groups in the molecule are essential for maintaining inhibitory activity, replacement by other functional groups could cause compouds'inhibitory activity decreasing.