Oxime‐Based Click Chemistry in the Development of 3‐Isoxazolecarboxylic Acid Containing Inhibitors of Yersinia pestis Protein Tyrosine Phosphatase, YopH

Abstract

Pathogenicity of Yersinia pestis (Y. pestis) relies on several effector proteins, including YopH, a protein-tyrosine phosphatase. Previously, we screened a library of analogues based on the ubiquitous PTP substrate, p-nitrophenylphosphate (pNPP) and found that incorporation of a 3-phenyl substituent (6-nitro-[1,1'-biphenyl]-3-yl dihydrogen phosphate (1)) enhanced affinity. The current study reports the conversion of 1 from a substrate to inhibitor by replacing the hydrolysable phosphoryl group with a 3-isoxazolecarboxylic acid moiety and by introduction of an aminooxy group and subsequent diversification using oxime-based click chemistry. As reported herein, this approach led to the identification of non-promiscuous low micromolar affinity bidentate YopH inhibitors.

Finding the Smoking Gun: Protein Tyrosine Phosphatases as Tools and Targets of Unicellular Microorganisms and Viruses

“…As potent inhibitors of YopH were reported isothiazolidinone heterocycles (known also as PTP1B inhibitors); the YopH crystal structure in presence of isothiazolidinone derivatives (18)(19) as phosphoryl mimetics was reported [145]. Bidentate inhibitors acting in the low micromolar range were reported to be developed based on 3-isoxazolecarboxylic acid using oxime-based click chemistry [146]. The same approach applied on nitrophenylphosphate-containing substrates was reported to yield a selective noncytotoxic YopH inhibitor with IC 50 190 nM (20) [147].…”

Yersinia pestis and Approaches to Targeting its Outer Protein H Protein-Tyrosine Phosphatase (YopH)

“…Screening the oximes with a small library of aldehydes resulted in the identification of bidentate non-promiscuous inhibitors 27 and 28, which showed YopH IC 50 values of 190 nm and 3.3 M, respectively Fig. (10) [79,81]. Binding modes of these inhibitors were studied using in-situ docking experiments.…”

“…10) was first identified using an SAS protocol and then converted to an inhibitor by the replacement of its hydrolyzable phosphoryl group with either a difluorophosphonomethyl group (25) [79] or 3-isoxazolecarboxylic acid group (26) Fig. (10) [81]. These served as non-hydrolyzable phosphoryl mimetics.…”

“…These reactions typically proceed quantitatively to yield oxime products of sufficient purity for direct biological evaluation. The methodology has been used to prepare multi-valent YopH inhibitors designed to combine binding interactions both inside and outside of the catalytic pocket [78][79][80][81].…”

“…The PDB entry 2Y2F was used as the starting structure to dock inhibitor 27 and the carboxyfuranyl-oxime side chain interacts at the peripheral sites of the catalytic pocket by hydrogen bonding with Arg230 and Asp231 [79]. The 5-phenyl-3-isocarboxylic acid group of inhibitor 28 occupies mainly the pTyr binding site, and the 5-carboxyindolyl part of the inhibitor is predicted to bind in either auxiliary pockets P2 or P3 [81]. Inhibitor 27 showed better selectivity for YopH relative to other panel of phosphatases.…”