methyl 3,5-dichloro-2-ethyl-4,6-dihydroxybenzoate

The genes from the oxygenase cluster nagAaGHAbAcAd of naphthalene-degrading Ralstonia sp. strain U2 were cloned and overexpressed. Salicylate 5-hydroxylase (S5H) activity, converting salicylate to gentisate, was present in vitro only in the single extract of cells with overexpressed nagAaGHAb or in a mixture of three cell extracts containing, respectively, NagGH (the oxygenase components), NagAa (ferredoxin reductase), and NagAb (ferredoxin). Each of the three extracts required for S5H activity was rate limiting in the presence of excess of the others but, when in excess, did not affect the rate of catalysis. S5H catalyzed the 5-hydroxylation of the aromatic rings of 3- and 4-substituted salicylates. However, the methyl group of 5-methylsalicylate was hydroxylated to produce the 5-hydroxymethyl derivative and the 6-position on the ring of 5-chlorosalicylate was hydroxylated, producing 5-chloro-2,6-dihydroxybenzoate. In an assay for the nag naphthalene dioxygenase (NDO) based on the indole-linked oxidation of NADH, three extracts were essential for activity (NagAcAd, NagAa, and NagAb). NDO and S5H were assayed in the presence of all possible combinations of the nag proteins and the corresponding nah NDO proteins from the "classical" naphthalene degrader P. putida NCIMB9816. All three oxygenase components functioned with mixed combinations of the electron transport proteins from either strain. The S5H from strain U2 is a unique monooxygenase which shares sequence similarity with dioxygenases such as NDO but is also sufficiently similar in structure to interact with the same electron transport chain and probably does so in vivo during naphthalene catabolism in strain U2.

Glucose regulated protein 94 (Grp94) is the endoplasmic reticulum resident of the heat shock protein 90 kDa (Hsp90) family of molecular chaperones. Grp94 associates with many proteins involved in cell adhesion and signaling, including integrins, Toll-like receptors, immunoglobulins, and mutant myocilin. Grp94 has been implicated as a target for several therapeutic areas including glaucoma, cancer metastasis, and multiple myeloma. While 85% identical to other Hsp90 isoforms, the N-terminal ATP-binding site of Grp94 possesses a unique hydrophobic pocket that was used to design isoform-selective inhibitors. Incorporation of a cis-amide bioisostere into the radamide scaffold led to development of the original Grp94-selective inhibitor, BnIm. Structure-activity relationship studies have now been performed on the aryl side chain of BnIm, which resulted in improved analogues that exhibit better potency and selectivity for Grp94. These analogues also manifest superior antimigratory activity in a metastasis model as well as enhanced mutant myocilin degradation in a glaucoma model compared to BnIm.

The use of supramolecular synthons as a strategy to control crystalline structure is a crucial factor in developing new solid forms with physicochemical properties optimized by design. However, to achieve this objective, it is necessary to understand the intermolecular interactions in the context of crystal packing. The feasibility of a given synthon depends on its flexibility to combine the drug with a variety of coformers. In the present work, the imidazole-hydroxy synthon is investigated using as the target molecule benzoylmetronidazole [BZMD; systematic name 2-(2-methyl-5-nitro-1H-imidazol-1-yl)ethyl benzoate], whose imidazole group seems to be a suitable acceptor for hydrogen bonds. Thus, coformers with carboxylic acid and phenol groups were chosen. According to the availability of binding sites presented in the coformer, and considering the proposed synthon and hydrogen-bond complementarity as major factors, different drug-coformer stoichiometric ratios were explored (1:1, 2:1 and 3:1). Thirteen new solid forms (two salts and eleven cocrystals) were produced, namely BZMD-benzoic acid (1/1), C13H13N3O4·C7H6O2, BZMD-β-naphthol (1/1), C13H13N3O4·C10H8O, BZMD-4-methoxybenzoic acid (1/1), C13H13N3O4·C8H8O3, BZMD-3,5-dinitrobenzoic acid (1/1), C13H13N3O4·C7H4N2O6, BZMD-3-aminobenzoic acid (1/1), C13H13N3O4·C7H7NO2, BZMD-salicylic acid (1/1), C13H13N3O4·C7H6O3, BZMD-maleic acid (1/1) {as the salt 1-[2-(benzoyloxy)ethyl]-2-methyl-5-nitro-1H-imidazol-3-ium 3-carboxyprop-2-enoate}, C13H14N3O4+·C4H3O4-, BZMD-isophthalic acid (1/1), C13H13N3O4·C8H6O4, BZMD-resorcinol (2/1), 2C13H13N3O4·C6H6O2, BZMD-fumaric acid (2/1), C13H13N3O4·0.5C4H4O4, BZMD-malonic acid (2/1), 2C13H13N3O4·C3H2O4, BZMD-2,6-dihydroxybenzoic acid (1/1) {as the salt 1-[2-(benzoyloxy)ethyl]-2-methyl-5-nitro-1H-imidazol-3-ium 2,6-dihydroxybenzoate}, C13H14N3O4+·C7H5O4-, and BZMD-3,5-dihydroxybenzoic acid (3/1), 3C13H13N3O4·C7H6O4, and their crystalline structures elucidated, confirming the robustness of the selected synthon.