Inhibition Of Isozymes Research Articles

Inhibition of human carbonic anhydrase isozymes I, II, IX and XII with a new series of sulfonamides incorporating aroylhydrazone-, [1,2,4]triazolo[3,4-b][1,3,4]thiadiazinyl- or 2-(cyanophenylmethylene)-1,3,4-thiadiazol-3(2H)-yl moieties

A series of benzenesulfonamides incorporating aroylhydrazone, piperidinyl, sulfone, [1,2,4]triazolo[3,4-b][1,3,4]thiadiazinyl- or 2-(cyanophenyl-methylene)-1,3,4-thiadiazol-3(2H)-yl moieties was investigated as inhibitors of four α-carbonic anhydrases (CAs, EC 4.2.1.1), the human (h) isoforms hCA I, II (cytosolic, offtarget enzymes) and hCA IX and XII (transmembrane, tumor-associated isoforms). Low nanomolar activity was observed against hCA II (KIs of 0.56–17.1 nM) with these sulfonamides, whereas the slow cytosolic isoform hCA I was less inhibited by these compounds (KIs of 86.4 nM–32.8 µM). Most of these sulfonamides significantly inhibited CA IX, with KIs in the range of 4.5–47.0 nM, although some of the derivatives incorporating bulkier bicyclic moieties, as well as 2-thienyl fragments, showed a weaker activity against this isoform (KIs in the range 50.1–553 nM). All the investigated compounds also inhibited CA XII with KIs in the range 0.85–376 nM. The best inhibitors were those incorporating bulky [1,2,4]triazolo[3,4-b][1,3,4]thiadiazinyl moieties and 1,3,4-thiadiazol-3(2H)-yl groups.

Dual PDE3/4 and PDE4 Inhibitors: Novel Treatments For COPD and Other Inflammatory Airway Diseases

Selective phosphodiesterase (PDE) 4 and dual PDE3/4 inhibitors have attracted considerable interest as potential therapeutic agents for the treatment of respiratory diseases, largely by virtue of their anti-inflammatory (PDE4) and bifunctional bronchodilator/anti-inflammatory (PDE3/4) effects. Many of these agents have, however, failed in early development for various reasons, including dose-limiting side effects when administered orally and lack of sufficient activity when inhaled. Indeed, only one selective PDE4 inhibitor, the orally active roflumilast-n-oxide, has to date received marketing authorization. The majority of the compounds that have failed were, however, orally administered and non-selective for either PDE3 (A,B) or PDE4 (A,B,C,D) subtypes. Developing an inhaled dual PDE3/4 inhibitor that is rapidly cleared from the systemic circulation, potentially with subtype specificity, may represent one strategy to improve the therapeutic index and also exhibit enhanced efficacy versus inhibition of either PDE3 or PDE4 alone, given the potential positive interactions with regard to anti-inflammatory and bronchodilator effects that have been observed pre-clinically with dual inhibition of PDE3 and PDE4 compared with inhibition of either isozyme alone. This MiniReview will summarize recent clinical data obtained with PDE inhibitors and the potential for these drugs to treat COPD and other inflammatory airways diseases such as asthma and cystic fibrosis.

Nonsteroidal Anti-Inflammatory Drugs and the Heart

Aspirin has been on the market for 115 years. Beginning with the marketing of indomethacin for the treatment of rheumatoid arthritis in 1963, at least 20 other nonsteroidal anti-inflammatory drugs (NSAIDs) with aspirin-like actions have been developed over the past 50 years,1 culminating with the introduction of a new class of selective inhibitors of cyclooxygenase (COX)-2, the coxibs, approximately 15 years ago.2 The NSAIDs represent the single most crowded family of drugs sharing the same therapeutic activities and mechanism of action, perhaps reflecting the unmet therapeutic need in the area of pain management and the large interindividual variability in response to these agents. NSAIDs provide symptomatic relief of pain and inflammation associated with a variety of human disorders, including the rheumatic diseases. Their shared therapeutic actions (ie, analgesic, anti-inflammatory, and antipyretic) are usually accompanied by mechanism-based adverse effects that can, at least in part, be attenuated as a function of individual pharmacokinetic or pharmacodynamic properties.1 Nuances in the tolerability of different NSAIDs had been described in the precoxib era on the basis of clinical trials of a few hundred patients treated for up to a few months with soft end points. More recently, however, important differences in safety have been demonstrated in head-to-head randomized comparisons of individual coxibs and 1 or more traditional NSAIDs that involved tens of thousands of patients treated for up to a few years with hard end points. Even more significantly, the interpretation of the effects of NSAIDs has been greatly enhanced by the availability, for the first time, of large, placebo-controlled trials that aimed to assess the potential for chemoprevention of colorectal cancer by rofecoxib or celecoxib. As a result of these developments, the whole field of NSAIDs has been illuminated during the past 15 years. Although epidemiological studies had previously associated …

Microplate-based method to screen inhibitors of isozymes of cyclic nucleotide phosphodiesterase fused to SUMO

The feasibility for microplate-based screening of inhibitors of isozymes of cyclic nucleotide phosphodiesterase (PDE) was tested via the coupled action of a phosphatase on adenosine-5′-monophosphate and an improved malachite green assay of phosphate. Human full-length PDE4B2 and truncated mutant (152–528aa) were expressed in Escherichia coli via fusion to SUMO, which after purification through Ni-NTA column exhibited specific activities >0.017 U mg−1. In the presence of proteins <30 mg L−1, absorbance for 10 µΜ phosphate was measurable; a PDE isozyme of specific activity over 0.008 U mg−1 after reaction for 20 min thus suited for microplate-based screening of inhibitors. By using Biotek ELX 800 microplate reader, affinities of two forms of PEDE4B2 for cAMP, rolipram and papaverine varied over three magnitudes and were consistent with those by routine assay, respectively. Hence, the proposed method was promising for high-throughput-screening of inhibitors of phosphate-releasing enzymes bearing specific activities over 0.008 U mg−1.

Dietary Flavonoids Modulate CYP2C to Improve Drug Oral Bioavailability and Their Qualitative/Quantitative Structure–Activity Relationship

This study aims to improve the drug oral bioavailability by co-administration with flavonoid inhibitors of the CYP2C isozyme and to establish qualitative and quantitative (QSAR) structure-activity relationships (SAR) between flavonoids and CYP2C. A total of 40 naturally occurring flavonoids were screened in vitro for CYP2C inhibition. Enzyme activity was determined by measuring conversion of tolbutamide to 4-hydroxytolbutamide by rat liver microsomes. The percent inhibition and IC50 of each flavonoid were calculated and used to develop SAR and QSAR. The most effective flavonoid was orally co-administered in vivo with a cholesterol-reducing drug, fluvastatin, which is normally metabolized by CYP2C. The most potent CYP2C inhibitor identified in vitro was tamarixetin (IC50 = 1.4 μM). This flavonoid enhanced the oral bioavailability of fluvastatin in vivo, producing a >2-fold increase in the area under the concentration-time curve and in the peak plasma concentration. SAR analysis indicated that the presence of a 2,3-double bond in the C ring, hydroxylation at positions 5, 6, and 7, and glycosylation had important effects on flavonoid-CYP2C interactions. These findings should prove useful for predicting the inhibition of CYP2C activity by other untested flavonoid-like compounds. In the present study, tamarixetin significantly inhibited CYP2C activity in vitro and in vivo. Thus, the use of tamarixetin could improve the therapeutic efficacy of drugs with low bioavailability.

Design, synthesis and biological evaluation of 5-oxo-1,4,5,6,7,8 hexahydroquinoline derivatives as selective cyclooxygenase-2 inhibitors.

A group of regioisomeric 5-oxo-1,4,5,6,7,8 hexahydroquinoline derivatives possessing a COX-2 SO2Me pharmacophore at the para position of the C-2 or C-4 phenyl ring, in conjunction with a C-4 or C-2 phenyl (4-H) or substituted-phenyl ring (4-F,4-Cl,4-Br,4-OMe,4-Me, 4-NO2), were designed for evaluation as selective cyclooxygenase-2 (COX-2) inhibitors. These target 5-oxo-1,4,5,6,7,8 hexahydroquinolines were synthesized via a Hansch condensation reaction. In vitro COX-1/COX-2 isozyme inhibition structure-activity studies identified 7,8-dihydro- 7,7-dimethyl-2-(4-methoxyphenyl)-4-(4-(methylsulfonyl)phenyl)quinolin-5(1H,4H,6H)- one (9c) as a potent COX-2 inhibitor (IC50 = 0.17 M) with a high COX-2 selectivity index (S.I. = 97.6) comparable to the reference drug celecoxib (COX-2 IC50 = 0.05 mM; COX-2 S.I= 405). A molecular modeling study where 9c was docked in active site of COX-2 showed that the p-SO2Me substituent on the C-2 phenyl ring is inserted into the secondary COX-2 binding site. The structure activity data acquired indicate that the position of the COX-2 SO2Me pharmacophore and type of substituent are important for COX-2 inhibitory activity.

Carbonic anhydrase inhibitors. Synthesis, and molecular structure of novel series N-substituted N′-(2-arylmethylthio-4-chloro-5-methylbenzenesulfonyl)guanidines and their inhibition of human cytosolic isozymes I and II and the transmembrane tumor-associated isozymes IX and XII

A series of novel N-substituted N'-(2-arylmethylthio-4-chloro-5-methylbenzenesulfonyl)guanidines 9-41 have been synthesized and investigated as inhibitors of four isoforms of zinc enzyme carbonic anhydrase (CA.EC 4.2.1.1), that is the cytosolic CA I and II, and cancer-associated isozymes CA IX and XII. Against the human CA I investigated compounds showed KI in the range of 87-6506 nM, toward hCA II ranging from 7.8 to 4500 nM, against hCA IX in the range of 4.7-416 nM and against hCA XII at range of 0.96-540 nM. Compounds 10, 12-14, 16, 18-20, 24-26, 31 and 32 exhibited a powerful inhibitory potency toward hCA IX (K(I) = 4.7-21 nM) in comparison to the reference sulfonamides AAZ, MZA, EZA, DCP and IND (K(I) = 24-50 nM). Compound 14 was the most potent inhibitor of hCA I (K(I) = 87 nM), hCA IX (K(I) = 4.7 nM) and hCA XII (K(I) = 0.96 nM), while 26 was the most effective inhibitor of hCA II (K(I) = 7.8 nM). The most promising compound 32 exerted the highest selectivity ratios toward hCA IX versus hCA I (hCA I/hCA IX = 261) and hCA II (hCA II/hCA IX = 26). The in vitro antitumor activity of compounds 10, 13, 14, 21, 22, 25, 32, 38 and 41 was evaluated at the US National Cancer Institute (NCI) against a panel of 60 human tumor cell lines. The most active antitumor agents 21 and 25, inhibiting 32-35 human tumor cell lines with GI₅₀ in the range of 2.1-5.0 μM also showed relatively high inhibitory activity toward hCA IX and XII with KI from 18 to 40 nM.

Resonant-Mie-scattering of aggregates of phosphomolybdate and papaverine for measuring activities and screening inhibitors of cyclic nucleotide phosphodiesterase isozymes

A resonant-light-scattering (RLS) method was proposed to quantify phosphate for screening inhibitors of isozymes of cyclic nucleotide phosphodiesterase (PDE). In acidified mixtures of phosphate, papaverine and molybdate, there were aggregates exhibiting micrometre sizes, no absorbance peaks over 360 nm but strong RLS peaks at 392 nm; Mie scattering thus accounted for the RLS signals. When papaverine was added before molybdate to acidified samples of phosphate, RLS signals at 392 nm were stable from 5 to 25 min since the addition of molybdate; after optimization, phosphate from 0.40 to 3.60 μM was quantifiable. This RLS method tolerated 60 mg L(-1) proteins besides common PDE inhibitors and dimethyl sulfoxide in acidified samples of phosphate; the integration of this RLS method with the coupled action of a phosphomonoesterase on PDE product was thus rational to measure PDE activities without the removal of proteins in samples. By quantifying activities of a truncated mutant of human PDE4B2 via this RLS method, Michaelis-Menten constant, inhibition constants of rolipram, papaverine and theophylline varied over three magnitudes and were consistent with those estimated by an improved malachite green assay of phosphate, respectively. Hence, this novel RLS method was promising for screening inhibitors of PDE isozymes.

Discovery of Inhibitors of 4′-Phosphopantetheine Adenylyltransferase (PPAT) To Validate PPAT as a Target for Antibacterial Therapy

Inhibitors of 4'-phosphopantetheine adenylyltransferase (PPAT) were identified through high-throughput screening of the AstraZeneca compound library. One series, cycloalkyl pyrimidines, showed inhibition of PPAT isozymes from several species, with the most potent inhibition of enzymes from Gram-positive species. Mode-of-inhibition studies with Streptococcus pneumoniae and Staphylococcus aureus PPAT demonstrated representatives of this series to be reversible inhibitors competitive with phosphopantetheine and uncompetitive with ATP, binding to the enzyme-ATP complex. The potency of this series was optimized using structure-based design, and inhibition of cell growth of Gram-positive species was achieved. Mode-of-action studies, using generation of resistant mutants with targeted sequencing as well as constructs that overexpress PPAT, demonstrated that growth suppression was due to inhibition of PPAT. An effect on bacterial burden was demonstrated in mouse lung and thigh infection models, but further optimization of dosing requirements and compound properties is needed before these compounds can be considered for progress into clinical development. These studies validated PPAT as a novel target for antibacterial therapy.

Variability in the Response to Non‐Steroidal Anti‐Inflammatory Drugs: Mechanisms and Perspectives

Non-steroidal anti-inflammatory drugs (NSAIDs) are a chemically heterogeneous group of compounds that provide unmistakable and significant health benefits in the treatment of pain and inflammation. They include traditional NSAIDs (tNSAIDs), which act by inhibiting both cyclooxygenase (COX)-1 and COX-2 and selective COX-2 inhibitors (coxibs). The development of biomarkers predictive of the impact of NSAIDs on COX-1 and COX-2 activities in vitro, ex vivo and in vivo has been essential to read out the clinical consequences of selective and non-selective inhibition of COX isozymes in human beings. The analgesic and anti-inflammatory effects of NSAIDs are COX-2-dependent effects, unrelated to COX-2 selectivity. The intensity and duration of these effects are influenced by dose and half-life of the NSAID. However, the inhibition of COX-1 in cells of the gastrointestinal (GI) system and COX-2 in vascular cells translates into increased risk of serious GI adverse events and atherothrombosis and hypertension, respectively. The COX-2 selectivity of NSAIDs can predict, at least in part, the GI toxicity. In contrast, the CV effects are largely COX-2-dependent effects, unrelated to COX-2 selectivity but are dose dependent. The reduction in the dose is recommended and presumably will limit the number of patients exposed to a CV or a GI hazard by NSAIDs and coxibs. It will not, however, eliminate the risk on an individual level because there is a marked variability in how different people react to these drugs, based on their genetic background. The challenge of the next future will be to develop biomarkers useful to identify the individuals who react abnormally to COX inhibition.

Inhibition of tumor-associated human carbonic anhydrase isozymes IX and XII by a new class of substituted-phenylacetamido aromatic sulfonamides

Here, we investigate 28 structurally new sulfonamides and their subsequent testing for enzyme inhibition of cytosolic and tumor-associated carbonic anhydrases (CAs, EC 4.2.1.1). The compounds showed very potent inhibition of four physiologically relevant human (h) CA isoforms, namely hCA I, II, IX and XII. Interestingly, the KI values were in the nanomolar range for the tumor-associated hCA IX and hCA XII. Docking studies have revealed details regarding the very favorable interactions between the scaffolds of this new class of inhibitors and the active sites of the investigated CA isoforms. As there are reported cases of tumors overexpressing both CA II and IX, such potent inhibitors for the two isoforms as those detected in this work, may have applications for targeting more than one CA present in tumors.

1,4-Diaryl-substituted triazoles as cyclooxygenase-2 inhibitors: Synthesis, biological evaluation and molecular modeling studies

A novel group of 1,4-diaryl-substituted triazoles was designed and synthesized by introducing the cyclooxygenase-2 (COX-2) pharmacophore SO2NH2 attached to one aryl ring and various substituents (H, F, Cl, CH3 or OCH3) attached to the other aryl ring. The effects of size and flexibility of the compounds upon COX-1/COX-2 inhibitory potency and selectivity was studied by increasing the size of an alkyl linker chain [(-CH2)n, where n=0, 1, 2]. In vitro COX-1/COX-2 inhibition studies showed that all compounds (14-18, 21-25 and 28-32) are more potent inhibitors of COX-2 isozyme (IC50=0.17-28.0μM range) compared to COX-1 isozyme (IC50=21.0 to >100μM range). Within the group of 1,4 diaryl-substituted triazoles, 4-{2-[4-(4-chloro-phenyl)-[1,2,3]triazol-1-yl]-ethyl}-benzenesulfonamide (compound 30) displayed highest COX-2 inhibitory potency and selectivity (COX-1: IC50=>100μM, COX-2: IC50=0.17μM, SI >588). Molecular docking studies using the catalytic site of COX-1 and COX-2, respectively, provided complementary theoretical support for the obtained experimental biological structure-activity relationship data. Results of molecular docking studies revealed that COX-2 pharmacophore SO2NH2 in compound 30 is positioned in the secondary pocket of COX-2 active site; with the nitrogen atom of the SO2NH2 group being hydrogen bonded to Q192 (N⋯OC=2.85Å), and one of the oxygen atoms of SO2NH2 group forming a hydrogen bond to H90 (SO⋯N=2.38Å).

Evaluation of vanillic acid as inhibitor of carbonic anhydrase isozyme III by using a modified Hummel–Dreyer method: approach for drug discovery

α-3 carbonic anhydrase isozyme (CAIII) is the most abundant protein in adipocytes and considered insensitive to sulfonamide inhibitors. It was reported recently that the knock-down of CAIII is attributed with controlling lipogenesis. Thus inhibition of this target may lead to the discovery of new therapies against obesity and insulin resistance. Vanillic acid as a small molecule with coordinating groups and has a potential to bind zinc atoms in CA binding sites. Inhibition of CAIII by vanillic acid was evaluated by Hummel-Dreyer chromatography because it provides free interaction between ligand and macromolecule and introduces solution for faulty results obtained by current colorimetric assays. HPLC system of vanillic acid produces vacancy (negative) peak representing the amount of attached vanillic acid with CAIII. It was found that vanillic acid is able to bind with CAIII through two equilibria, one at equimolar ratio and another at 2:1 (vanillic acid-CAIII) ratio. The affinity constant of equimolar binding between CAIII and vanillic acid was found to be 14,400 m(-1) . It was found that vanillic acid binding with CAIII is much stronger than phenol and acetazolamide (positive controls).

A novel anti-HIV active integrase inhibitor with a favorable in vitro cytochrome P450 and uridine 5′-diphospho-glucuronosyltransferase metabolism profile

Research efforts on the human immunodeficiency virus (HIV) integrase have resulted in two approved drugs. However, co-infection of HIV with Mycobacterium tuberculosis and other microbial and viral agents has introduced added complications to this pandemic, requiring favorable drug-drug interaction profiles for antiviral therapeutics targeting HIV. Cytochrome P450 (CYP) and uridine 5'-diphospho-glucuronosyltransferase (UGT) are pivotal determining factors in the occurrence of adverse drug-drug interactions. For this reason, it is important that anti-HIV agents, such as integrase inhibitors, possess favorable profiles with respect to CYP and UGT. We have discovered a novel HIV integrase inhibitor (compound 1) that exhibits low nM antiviral activity against a diverse set of HIV-1 isolates, and against HIV-2 and the simian immunodeficiency virus (SIV). Compound 1 displays low in vitro cytotoxicity and its resistance and related drug susceptibility profiles are favorable. Data from in vitro studies revealed that compound 1 was not a substrate for UGT isoforms and that it was not an inhibitor or activator of key CYP isozymes.

Abstract 4241: Isozyme-selective intracellular binding assay for histone deacetylases .

Abstract The precise modulation of histone acetylation is crucial to cellular survival and control of reproduction.Misregulation of acetylation is often correlated with uncontrolled cellular proliferation, and general inhibition of histone deacetylases has proven to be an effective strategy in the treatment of some cancers. The underlying biology of histone deacetylases is however quite complex, with four classes of isozymes and a large number of interacting proteins that modulate the activity of each. The general scarcity of potent isozyme-selective HDAC inhibitors further complicates the task of deciphering individual isozyme activity in a cellular context. We now report a novel approach to monitoring the intracellular inhibition of single isozymes based on bioluminescence resonance energy transfer from NanoLucTM, a small and exceedingly bright bioluminescent reporter. In our approach, a NanoLucTM fusion of the isozyme of interest is used in combination with an optimized fluorescent tracer. When bound, energy transfer occurs from the bioluminescent enzyme to the fluorescent probe. Displacement of the probe by a competitive inhibitor is then measured as a decrease in the energy transfer ratio, yielding IC50 data specific to the chosen isozyme. We demonstrate the ability to measure intracellular IC50 values of inhibitors against individual isozymes of class I and class IIb based on choice of the NanoLucTM fusion. Citation Format: Carolyn C. Woodroofe, Matthew B. Robers, Thomas A. Kirkland, Kevin Kupcho, Andrew L. Niles, Mei Cong, Keith V. Wood. Isozyme-selective intracellular binding assay for histone deacetylases . [abstract]. In: Proceedings of the 104th Annual Meeting of the American Association for Cancer Research; 2013 Apr 6-10; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2013;73(8 Suppl):Abstract nr 4241. doi:10.1158/1538-7445.AM2013-4241

Antibacterial oxazolidinone analogues having a N-hydroxyacetyl-substituted seven-membered [1,2,5]triazepane or [1,2,5]oxadiazepane C-ring unit

We synthesized a series of oxazolidinone analogues bearing a N-hydroxyacetyl-substituted [1,2,5]triazepane or [1,2,5]oxadiazepane C-ring unit as homologues of an earlier drug candidate, eperezolid. Several of these compounds exhibited potent invitro antibacterial activities towards not only Gram-positive, but also Gram-negative and linezolid-resistant pathogens. Compounds 21a and 21b, bearing a thiocarbamate side chain, showed high invivo activity against methicillin-resistant Staphylococcus aureus SR3637, together with a promising safety profile in terms of weak inhibition of monoamine oxidase and cytochrome P450 isozymes.

4-Substituted-2,3,5,6-tetrafluorobenzenesulfonamides as inhibitors of carbonic anhydrases I, II, VII, XII, and XIII

A series of 4-substituted-2,3,5,6-tetrafluorobenezenesulfonamides were synthesized and their binding potencies as inhibitors of recombinant human carbonic anhydrase isozymes I, II, VII, XII, and XIII were determined by the thermal shift assay, isothermal titration calorimetry, and stop-flow CO2 hydration assay. All fluorinated benzenesulfonamides exhibited nanomolar binding potency toward tested CAs and fluorinated benzenesulfonamides posessed higher binding potency than non-fluorinated compounds. The crystal structures of 4-[(4,6-dimethylpyrimidin-2-yl)thio]-2,3,5,6-tetrafluorobenzenesulfonamide in complex with CA II and CA XII, and 2,3,5,6-tetrafluoro-4-[(2-hydroxyethyl)sulfonyl]benzenesulfonamide in complex with CA XIII were determined. The observed dissociation constants for several fluorinated compounds reached subnanomolar range for CA I isozyme. The affinity and the selectivity of the compounds towards tested isozymes are presented.

Modulation of paracetamol-induced hepatotoxicity by phosphodiesterase isozyme inhibition in rats: a preliminary study

Altered regulation of nitric oxide-cyclic guanosine monophosphate (NO-cGMP) is present in liver cirrhosis. Several experimental studies have shown that selective modulation of NO metabolism in the liver reduces intrahepatic resistance and portal pressure in cirrhosis. This preliminary study investigated whether selective inhibition of phosphodiesterase-5 (PDE-5), which prevents the conversion of cGMP to 5'-GMP, as well as non-selective inhibition of PDE isozymes could ameliorate hepatic toxicity induced by paracetamol (PCM). PCM (250 mg/kg, i.p.) was administered to induce hepatotoxicity. Control rats received physiological saline (10 mL/kg, p.o.), while sildenafil (a selective PDE-5 inhibitor) and aminophylline (a non-selective PDE inhibitor) were administered separately at 10 mg/kg p.o. to PCM-treated rats. PCM hepatotoxicity, characterized by elevation of aspartate and alanine aminotransferases, hepatic degeneration, and centrilobular necrosis, was attenuated by both PDE inhibitors. Sildenafil and aminophylline significantly (p<0.05) reduced plasma aspartate aminotransferase activity by 49.6% and 39.8%, respectively, with moderate increase in alanine aminotransferase activity by 26.1% and 20.4%, respectively, in PCM-treated rats. Decreases in total protein and albumin induced by PCM were significantly (p<0.05) prevented by 30.0% and 22.2%, respectively, following sildenafil administration, while aminophylline decreased these proteins by 14.0% and 25.9%, respectively. Sildenafil and aminophylline significantly (p<0.05) reduced lipid peroxidation by 30.7% and 19.7%, respectively, while moderately increasing glutathione (GSH) in the PCM-treated rats. Both drugs did not significantly alter the total cholesterol and triglyceride levels. These preliminary data suggest that pharmacological inhibition of PDE isozymes may be a useful strategy in protecting against PCM hepatic toxicity.

Synthesis and biologic evaluation of new 3-phenoxyazetidin-2-one derivatives as selective cyclooxygenase-2 inhibitors

A new series of 3-phenoxyazetidin-2-ones (β-lactams) were designed and synthesized for the evaluation as selective cyclooxygenase-2 (COX-2) inhibitors. In vitro COX-1 and COX-2 inhibition studies showed that all compounds were selective inhibitors of the COX-2 isozyme with IC50 values in the 0.054–0.095 μM range, and COX-2 selectivity indexes in the 228.47–355.6 range. Among the synthesized compounds, 1-(4-methoxyphenyl)-4-(4-(methylsulfonyl)phenyl)-3-phenoxyazetidin-2-one (4j) possessing methoxy group at the para position of N-1 phenyl ring exhibited the highest COX-2 inhibitory selectivity and potency even more potent than the reference drug celecoxib. Molecular modeling studies indicated that the methylsulfonyl pharmacophore group can be inserted into the secondary pocket of COX-2 active site.

QSAR studies of sulfamate and sulfamide inhibitors targeting human carbonic anhydrase isozymes I, II, IX and XII

The last version of the PRECLAV algorithm was used to investigate a series of sulfamate/sulfamide carbonic anhydrase (CA, EC 4.2.1.1) inhibitors. PRECLAV allows identification of outliers for lead hopping, significant molecular fragments and similarity computation of a calibration set vs. a prediction set of compounds, from the viewpoint of computed QSAR. In the current study the database included 65 sulfamates and sulfamides as calibration set and 51 not yet synthesized sulfamates and sulfamides as prediction set. The dependent property was inhibitory activity for human (h) CA isozymes I, II, IX and XII. There were no outliers for lead hopping in the calibration set. In some cases, replacing of the O-SO2-NH2 by the NH-SO2-NH2 moiety, led to modifications in the structure of molecular fragments in another regions of the molecule. According to the descriptors in the obtained models, the inhibition of all CA isozymes was strongly influenced by the molecular shape and size of the sulfamates/sulfamides (which incorporate a substituted aryl-ureido scaffold). The lipophilicity of the inhibitors seemed to be a minor factor influencing CA inhibitory action. The presence of specific PRECLAV molecular fragments such as a C6 (totally substituted benzene), F, O and NO2 increases the activity of inhibitors against certain isoforms. The presence of molecular fragments such as C, CH, CxHy (substituted benzene/naphthalene) and NH decreased the activity of inhibitors against certain isoforms. In the prediction set, we evidenced seven compounds estimated to be highly active inhibitors for at least three of the investigated isozymes and eight molecules estimated to possess low activity against at least three CAs. The paper suggests the structure of some possible interesting inhibitors incorporating F, O and NO2 fragments that may have affinity for certain human CAs.