Murat Senturk

In vitro inhibition of salicylic acid derivatives on human cytosolic carbonic anhydrase isozymes I and II

The inhibition of two human cytosolic carbonic anhydrase (hCA, EC 4.2.1.1) isozymes, hCA I and II, with a series of salicylic acid derivatives was investigated by using the esterase method with 4-nitrophenyl acetate as substrate. IC(50) values for sulfasalazine, diflunisal, 5-chlorosalicylic acid, dinitrosalicylic acid, 4-aminosalicylic acid, 4-sulfosalicylic acid, 5-sulfosalicylic acid, salicylic acid, acetylsalicylic acid (aspirin) and 3-metylsalicylic acid were of 3.04 microM, 3.38 microM, 4.07 microM, 7.64 microM, 0.13 mM, 0.29 mM, 0.42 mM, 0.56 mM, 2.71 mM and 3.07 mM for hCA I and of 4.49 microM, 2.70 microM, 0.72 microM, 2.80 microM, 0.75 mM, 0.72 mM, 0.29 mM, 0.68 mM, 1.16 mM and 4.70 mM for hCA II, respectively. Lineweaver-Burk plots were also used for the determination of the inhibition mechanism of these substituted phenols, most of which were noncompetitive inhibitors with this substrate. Some salicylic acid derivatives investigated here showed effective hCA I and II inhibitory activity, and might be used as leads for generating enzyme inhibitors eventually targeting other isoforms which have not been assayed yet for their interactions with such agents.

Carbonic anhydrase inhibitors: in vitro inhibition of α isoforms (hCA I, hCA II, bCA III, hCA IV) by flavonoids

A series of flavonoids, such as quercetin, catechin, apigenin, luteolin, morin, were investigated for their inhibitory effects against the metalloenzyme carbonic anhydrase (CA). The compounds were tested against four α-CA isozymes purified from human and bovine (hCA I, hCA II, bCA III, hCA IV) tissues. The four isozymes showed quite diverse inhibition profiles with these compounds. The flavonoids inhibited hCA I with KI-s in the range of 2.2–12.8 μM, hCA II with KI-s in the range of 0.74–6.2 μM, bCA III with KI-s in the range of 2.2–21.3 μM, and hCA IV with inhibition constants in the range of 4.4–15.7, with an esterase assay using 4-nitrophenyl acetate as substrate. Some simple phenols/sulfonamides were also investigated as standard inhibitors. The flavonoids incorporate phenol moieties which inhibit these CAs through a diverse, not yet determined inhibition mechanism, compared to classic inhibitors such as the sulfonamide/sulfamate ones.

NO-releasing esters show carbonic anhydrase inhibitory action against human isoforms I and II

Carbonic anhydrase (CA, EC 4.2.1.1) inhibitors (CAIs) are a class of pharmaceuticals used as antiglaucoma agents, diuretics, antiepileptics, in the management of mountain sickness, gastric and duodenal ulcers, neurological disorders, or osteoporosis. We report here the inhibitory capacities of some organic nitrates against two human (hCA) isozymes, hCA I and hCA II. The IC(50) values of compounds 1-12 against hCA I ranged between 7.13mM and 124mM, and against hCA II between 65.1microM and 0.79mM. Nitrate esters are thus interesting hCA I and II inhibitors, and might be used as leads for generating enzyme inhibitors eventually targeting other isoforms which have not been assayed yet for their interactions with such agents.

Synthesis and biological activity of novel thiourea derivatives as carbonic anhydrase inhibitors

Abstract A new series of chiral thiourea derivatives (5a–5c) and thiourea containing benzimidazole moieties (9b–9e) were synthesized from different amino acids (l-valine, l-isoleucine, l-methionine, l-phenylalanine, and d-phenylglycine). The compounds were characterized and tested against the two most studied members of the pH regulatory enzyme family, carbonic anhydrase (CA, EC 4.2.1.1). KI values of the novel compounds were measured in the range of 3.4–73.6 μM for hCA I isozyme and 8.7–1.44.2 μM for hCA II isozyme, respectively. Phenol was also tested as standard in order to understand the structure activity relationship and the clinically used sulfonamide acetazolamide was tested for comparison reasons. All of the compounds exhibited competitive inhibition with 4-nitrophenylacetate as substrate.

Effects of some antibiotics on human erythrocyte glutathione reductase: an in vitro study

Inhibitory effects of some antibiotics on purified human erythrocyte glutathione reductase were investigated. Human erythrocyte glutathione reductase was purified 2800-fold (29% yield) at 4°C using 2′, 5′-ADP Sepharose 4B affinity gel and Sephadex G-200 gel filtration chromatography. SDS polyacrylamide gel electrophoresis showed a single band for the enzyme. Imipenem, rifamycin, sulfanylacetamide, ceftazidime, chloramphenicol, seftriaxon, vancomycin, cefuroxime and ornidazole exhibited inhibitory effects but clindamycin, lincomycin, amoxicillin, amikacin exhibited activatory effects on the enzyme in vitro. The IC50 values of imipenem, rifamycin, sulfanylacetamide, ceftazidime, chloramphenicol, seftriaxon, vancomycin, cefuroxime and ornidazole were 0.030, 0.146, 0.59, 2.476, 2.36, 2.88, 4.83, 15.43 and 19.632 mM, respectively, and the Ki constants were 0.06 ± 0.01, 0.275 ± 0.10, 0.85 ± 0.05, 3.59 ± 0.51, 3.85 ± 0.40, 3.71 ± 0.60, 15.11 ± 2.50, 23.50 ± 2.94 and 28.49 ± 6.50 mM, respectively. While imipenem, rifamycin, sulfanylacetamide, ceftazidime, chloramphenicol and seftriaxon cefuroxime and ornidazole showed competitive inhibition, vankomycine displayed noncompetitive inhibition.

Effects of some analgesic anaesthetic drugs on human erythrocyte glutathione reductase: an in vitro study

Inhibitory effects of some analgesic and anaesthetic drugs on human erythrocyte glutathione reductase were investigated. For this purpose, human erythrocyte glutathione reductase was initially purified 2139-fold in a yield of 29% by using 2′, 5′-ADP Sepharose 4B affinity gel and Sephadex G-200 gel filtration chromatography. SDS polyacrylamide gel electrophoresis confirmed the purity of the enzyme by sharing a single band. A constant temperature (+4°C) was maintained during the purification process. Diclofenac sodium, ketoprofen, lornoxicam, tenoxicam, etomidate, morphine and propofol exhibited inhibitory effects on the enzyme in vitro using the Beutler assay method. Ki constants and IC50 values for drugs were determined from Lineweaver-Burk graphs and plotting activity % versus [I] graphs, respectively. The IC50 values of diclofenac sodium, ketoprofen, lornoxicam, propofol, tenoxicam, etomidate and morphine were 7.265, 6.278, 0.3, 0.242, 0.082, 0.0523 and 0.0128 mM and the Ki constants were 23.97 ± 2.1, 22.14 ± 7.6, 0.42 ± 0.18, 0.418 ± 0.056, 0.13 ± 0.025, 0.0725 ± 0.0029 and 0.0165 ± 0.0013 mM, respectively. While diclofenac sodium, ketoprofen, lornoxicam, tenoxicam etomidate and morphine showed competitive inhibition, propofol displayed noncompetitive inhibition.

Pyridazinone substituted benzenesulfonamides as potent carbonic anhydrase inhibitors.

A series of sulfonamide derivatives (2a-l) incorporating substituted pyridazinone moieties were investigated for the inhibition of two human cytosolic carbonic anhydrase isoforms, hCA I and hCA II. All these compounds, together with the clinically used sulfonamide acetazolamide were investigated as inhibitors of the physiologically relevant isozymes I and II. These sulfonamides showed very strong inhibition against all these isoforms with K(I)s in the range of 0.98-8.5 nM which makes such molecules possible to be used as leads for discovery of novel effective CA inhibitors targeting other isoforms with medicinal chemistry applications.

Discovering novel carbonic anhydrase type IX (CA IX) inhibitors from seven million compounds using virtual screening and in vitro analysis.

Abstract Carbonic anhydrase type IX (CA IX) enzyme is mostly over expressed in different cancer cell lines and tumor tissues. Potent CA IX inhibitors can be effective for adjusting the pH imbalance in tumor cells. In the present work, we represented the successful application of high throughput virtual screening (HTVS) of large dataset from ZINC database included of ∼7 million compounds to discover novel inhibitors of CA IX. HTVS and molecular docking were performed using consequence Glide/standard precision (SP), extra precision (XP) and induced fit docking (IFD) molecular docking protocols. For each compound, docking code calculates a set of low-energy poses and then exhaustively scans the binding pocket of the target with small compounds. Novel CA IX inhibitor candidates were suggested based on molecular modeling studies and a few of them were tested using in vitro analysis. These compounds were determined as good inhibitors against human CA IX target with Ki in the range of 0.85–1.58 μM. In order to predict the pharmaceutical properties of the selected compounds, ADME (absorption, distribution, metabolism and excretion) analysis was also carried out.

Kinetic and in silico studies of hydroxy-based inhibitors of carbonic anhydrase isoforms I and II.

Abstract A series of hydroxy and phenolic compounds have been assayed for the inhibition of two physiologically relevant carbonic anhydrase (CA, EC 4.2.1.1) isozymes, the cytosolic human isozymes I and II. The investigated molecules showed inhibition constants in the range of 1.07–4003 and 0.09–31.5 μM at the hCA I and hCA II enzymes, respectively. In order to investigate the binding mechanisms of these inhibitors, in silico studies were also applied. Molecular docking scores of the studied compounds are compared using three different scoring algorithms, namely Glide/SP, Glide/XP and Glide/IFD. In addition, different ADME (absorption, distribution, metabolism and excretion) analysis was performed. All the examined compounds were found within the acceptable range of pharmacokinetic profiles.

Kinetic and docking studies of cytosolic/tumor-associated carbonic anhydrase isozymes I, II and IX with some hydroxylic compounds

Abstract A series of hydroxylic compounds (1–10, NK-154 and NK-168) have been assayed for the inhibition of three physiologically relevant carbonic anhydrase isozymes, the cytosolic isozymes I, II and tumor-associated isozyme IX. The investigated compounds showed inhibition constants in the range of 0.068–4003, 0.012–9.9 and 0.025–115 μm at the hCA I, hCA II and hCA IX enzymes, respectively. In order to investigate the binding mechanisms of these inhibitors, in silico studies were also applied. Molecular docking scores of the studied compounds are calculated using scoring algorithms, namely Glide/induced fit docking. The inhibitory potencies of the novel compounds were analyzed at the human isoforms hCA I, hCA II and hCA IX as targets and the KI values were calculated.