CDP840

We present the in vitro characterization of a novel phosphodiesterase type 4 inhibitor, CDP840 (R-[+]-4-[2-{3-cyclopentyloxy-4-methoxyphenyl}-2-phenylethyl]pyridine), which has shown efficacy in a phase II allergen challenge study in asthmatics without adverse effects. CDP840 potently inhibits PDE-4 isoenzymes (IC₅₀ 2–30 nM) without any effect on PDE-1, 2, 3, 5, and 7 (IC₅₀>100 μM). It exhibited no significant selectivity in inhibiting human recombinant isoenzymes PDE-4A, B, C or D and was equally active against the isoenzymes lacking UCR1 (PDE-4B2 and PDE-4D2). In contrast to rolipram, CDP840 acted as a simple competitive inhibitor of all PDE-4 isoenzymes. Studies with rolipram indicated a heterogeneity within all the preparations of PDE-4 isoenzymes, indicative of rolipram inhibiting the catalytic activity of PDE-4 with both a low or high affinity. These observations were confirmed by the use of a PDE-4A variant, PDE-4A_330–886, which rolipram inhibited with low affinity (IC₅₀=1022 nM). CDP840 in contrast inhibited this PDE-4A variant with similar potency (IC₅₀=3.9 nM), which was in good agreement with theK _d of 4.8 nM obtained from [³H]-CDP840 binding studies. Both CDP840 and rolipram inhibited the high-affinity binding of [³H]-rolipram binding to PDE-4A, B, C and D with similarK _{d app} (7–19 nM and 3–5 nM, respectively). Thus, the activity of CDP840 at the [³H]-rolipram binding site was in agreement with the inhibitor’s activity at the catalytic site. However, rolipram was ∼100-fold more potent than CDP840 at inhibiting the binding of [³H]-rolipram to mouse brain in vivo. These data clearly demonstrate that CDP840 is a potent selective inhibitor of all PDE-4 isoenzymes. In contrast to rolipram, CDP840 was well-tolerated in humans. This difference, however, cannot at present be attributed to either isoenzyme selectivity or lack of activity in vitro at the high-affinity rolipram binding site (Sr).     

Interaction of the N-(3-Methylpyridin-2-yl)amide Derivatives of Flurbiprofen and Ibuprofen with FAAH: Enantiomeric Selectivity and Binding Mode

Background

Combined fatty acid amide hydrolase (FAAH) and cyclooxygenase (COX) inhibition is a promising approach for pain-relief. The Flu-AM1 and Ibu-AM5 derivatives of flurbiprofen and ibuprofen retain similar COX-inhibitory properties and are more potent inhibitors of FAAH than the parent compounds. However, little is known as to the nature of their interaction with FAAH, or to the importance of their chirality. This has been explored here.

Methodology/Principal Findings

FAAH inhibitory activity was measured in rat brain homogenates and in lysates expressing either wild-type or FAAH^T488A-mutated enzyme. Molecular modelling was undertaken using both docking and molecular dynamics. The (R)- and (S)-enantiomers of Flu-AM1 inhibited rat FAAH with similar potencies (IC₅₀ values of 0.74 and 0.99 μM, respectively), whereas the (S)-enantiomer of Ibu-AM5 (IC₅₀ 0.59 μM) was more potent than the (R)-enantiomer (IC₅₀ 5.7 μM). Multiple inhibition experiments indicated that both (R)-Flu-AM1 and (S)-Ibu-AM5 inhibited FAAH in a manner mutually exclusive to carprofen. Computational studies indicated that the binding site for the Flu-AM1 and Ibu-AM5 enantiomers was located between the acyl chain binding channel and the membrane access channel, in a site overlapping the carprofen binding site, and showed a binding mode in line with that proposed for carprofen and other non-covalent ligands. The potency of (R)-Flu-AM1 was lower towards lysates expressing FAAH mutated at the proposed carprofen binding area than in lysates expressing wild-type FAAH.

Conclusions/Significance

The study provides kinetic and structural evidence that the enantiomers of Flu-AM1 and Ibu-AM5 bind in the substrate channel of FAAH. This information will be useful in aiding the design of novel dual-action FAAH: COX inhibitors.     

A Novel Whole-Cell Biocatalyst with NAD+ Regeneration for Production of Chiral Chemicals

Background

The high costs of pyridine nucleotide cofactors have limited the applications of NAD(P)-dependent oxidoreductases on an industrial scale. Although NAD(P)H regeneration systems have been widely studied, NAD(P)⁺ regeneration, which is required in reactions where the oxidized form of the cofactor is used, has been less well explored, particularly in whole-cell biocatalytic processes.

Methodology/Principal Findings

Simultaneous overexpression of an NAD⁺ dependent enzyme and an NAD⁺ regenerating enzyme (H₂O producing NADH oxidase from Lactobacillus brevis) in a whole-cell biocatalyst was studied for application in the NAD⁺-dependent oxidation system. The whole-cell biocatalyst with (2R,3R)-2,3-butanediol dehydrogenase as the catalyzing enzyme was used to produce (3R)-acetoin, (3S)-acetoin and (2S,3S)-2,3-butanediol.

Conclusions/Significance

A recombinant strain, in which an NAD⁺ regeneration enzyme was coexpressed, displayed significantly higher biocatalytic efficiency in terms of the production of chiral acetoin and (2S,3S)-2,3-butanediol. The application of this coexpression system to the production of other chiral chemicals could be extended by using different NAD(P)-dependent dehydrogenases that require NAD(P)⁺ for catalysis.     

Characterization and Distribution of a Cloned Rat μ-Opioid Receptor

Abstract: We have cloned and expressed a rat brain cDNA, TS11, that encodes a μ-opioid receptor based on pharmacological, physiological, and anatomical criteria. Membranes were prepared from COS-7 cells transiently expressing TS11 bound [³H]diprenorphine with high affinity (K_D = 0.23 ± 0.04 nM). The rank order potency of drugs competing with [³H]diprenorphine was as follows: levorphanol (K_i = 0.6 ± 0.2 nM) ≈β-endorphin (K_i = 0.7 ± 0.5 nM) ≈ morphine (K_i = 0.8 ± 0.5 nM) ≈ [d -Ala², N-Me-Phe⁴,Gly-ol⁵]-enkephalin (DAMGO; K_i = 1.6 ± 0.5 nM) ? U50,488 (K_i = 910 ± 0.78 nM) > [d -Pen^2,5]-enkephalin (K_i = 3,170 ± 98 nM) > dextrorphan (K_i = 4,100 ± 68 nM). The rank order potencies of these ligands, the stereospecificity of levorphanol, and morphine's subnanomolar K_i are consistent with a μ-opioid binding site. Two additional experiments provided evidence that this opioid-binding site is functionally coupled to G proteins: (a) In COS-7 cells 50 µM 5′-guanylylimidodiphosphate shifted a fraction of receptors with high affinity for DAMGO (IC₅₀ = 3.4 ± 0.5 nM) to a lower-affinity state (IC₅₀ = 89.0 ± 19.0 nM), and (b) exposure of Chinese hamster ovary cells stably expressing the cloned μ-opioid receptor to DAMGO resulted in a dose-dependent, naloxone-sensitive inhibition of forskolin-stimulated cyclic AMP production. The distribution of mRNA corresponding to the μ-opioid receptor encoded by TS11 was determined by in situ hybridization to brain sections prepared from adult female rats. The highest levels of μ-receptor mRNA were detected in the thalamus, medial habenula, and the caudate putamen; however, significant hybridization was also observed in many other brain regions, including the hypothalamus.     

The oncometabolite 2-hydroxyglutarate inhibits histone lysine demethylases

Mutations in isocitrate dehydrogenases (IDHs) have a gain-of-function effect leading to R(−)-2-hydroxyglutarate (R-2HG) accumulation. By using biochemical, structural and cellular assays, we show that either or both R- and S-2HG inhibit 2-oxoglutarate (2OG)-dependent oxygenases with varying potencies. Half-maximal inhibitory concentration (IC₅₀) values for the R-form of 2HG varied from approximately 25 μM for the histone N^ɛ-lysine demethylase JMJD2A to more than 5 mM for the hypoxia-inducible factor (HIF) prolyl hydroxylase. The results indicate that candidate oncogenic pathways in IDH-associated malignancy should include those that are regulated by other 2OG oxygenases than HIF hydroxylases, in particular those involving the regulation of histone methylation.     

Structure–cytotoxic activity relationship of sesquilignan,morinol A

The cytotoxic activities of sesquilignans, (7S,8S,7′R,8′R)- and (7R,8R,7′S,8′S)-morinol A and (7S,8S,7′S,8′S)- and (7R,8R,7′R,8′R)-morinol B were compared, showing no significant difference between stereoisomers (IC₅₀ = 24–35 μM). As a next stage, the effect of substituents at 7, 7′, and 7″-aromatic ring on the activity was evaluated to find out the higher activity of (7S,8S,7′R,8′R)-7,7′,7″-phenyl derivative 18 (IC₅₀ = 6–7 μM). In the research on the structure–activity relationship of 7″-position of (7S,8S,7′R,8′R)-7,7′,7″-phenyl derivative 18, the most potent compounds were 7,7′,7″-phenyl derivative 18 (IC₅₀ = 6 μM) against HeLa cells. Against HL-60 cells, 7″-(4-nitrophenyl)-7,7′-phenyl derivative 33 and 7″-hexyl-7,7′-phenyl derivative 37 (IC₅₀ = 5 μM) showed highest activity. We discovered the compounds showed four to sevenfold potent activity than that of natural (7S,8S,7′R,8′R)-morinol A. It was also confirmed that the 7′-benzylic hydroxy group have an important role for exhibiting activity, on the other hand, the resonance system of cinnamyl structure is not crucial for the potent activity.     

Two new cytotoxic eudesmane sesquiterpenoids from Artemisia anomala

Two new eudesmane sesquiterpenoids artanoate (1) and eudesmanomolide (2) were isolated from the aerial parts of Artemisia anomala S. Moore. Their structures were elucidated as methyl (4R, 5S, 6S, 7S, 10R)-1-oxo-4, 6-dihydroxy-eudesma-2, 11 (13)-dien-12-oate (1) and (1R, 5R, 6R, 10R)-3, 13-diacetoxy-1-hydroxy-3, 7(11)-diene-12, 6-olide (2) on the basis of extensive spectroscopic analyses. Compound 1 showed cytotoxicity against HCT-8 cell lines with IC₅₀ value of 9.13 μM, and compound 2 exhibited inhibitory activities against HCT-8 and A549 cell lines with IC₅₀ values of 3.76 and 5.49 μM, respectively.     

Synthesis,NMR characterization and divergent biological actions of 2′-hydroxy-ceramide/dihydroceramide stereoisomers in MCF7 cells

A straightforward method for the simultaneous preparation of (2S,3R,2′R)- and (2S,3R,2′S)-2′-hydroxy-ceramides (2′-OHCer) from (2S,3R)-sphingosine acetonide precursors and racemic mixtures of 2-hydroxy fatty acids (2-OHFAs) is described. The obtained 2′-OH-C4-, -C6-, -C12-, -C16-Cer and 2′-OH-C6-dhCer pairs of diastereoisomers were characterized thoroughly by TLC, MS, NMR, and optical rotation. Dynamic and multidimensional NMR studies provided evidence that polar interfaces of 2′-OHCers are extended and more rigid than observed for the corresponding non-hydroxylated analogs. Stereospecific profile on growth suppression of MCF7 cells was observed for (2′R)- and (2′S)-2′-OH-C6-Cers and their dihydro analogs. The (2′R)-isomers were more active than the (2′S)-isomers (IC₅₀ ～3 μM/8 μM and IC₅₀ ～8 μM/12 μM, respectively), surpassing activity of the ordinary C6-Cer (IC₅₀ ～12 μM) and C6-dhCer (IC₅₀ ～38 μM). Neither isomer of 2′-OH-C6-Cers and 2′-OH-C6-dhCers was metabolized to their cellular long chain 2′-OH-homologs. Surprisingly, the most active (2′R)-isomers did not influence the levels of the cellular Cers nor dhCers. Contrary to this, the (2′S)-isomers generated cellular Cers and dhCers efficiently. In comparison, the ordinary C6-Cer and C6-dhCer also significantly increased the levels of their cellular long chain homologs. These peculiar anabolic responses and SAR data suggest that (2′R)-2′-OHCers/dhCers may interact with some distinct cellular regulatory targets in a specific and more effective manner than their non-hydroxylated analogs. Thus, stereoisomers of 2′-OHCers can be potentially utilized as novel molecular tools to study lipid–protein interactions, cell signaling phenomena and to understand the role of hydroxylated sphingolipids in cancer biology, pathogenesis and therapy.     

Imidazole-pyrazole hybrids: Synthesis,characterization and in-vitro bioevaluation against α-glucosidase enzyme with molecular docking studies

Herein, substituted imidazole-pyrazole hybrids (2a-2n) were prepared via a multi component reaction employing pyrazole-4-carbaldehydes (1a-1d), ammonium acetate, benzil and arylamines as reactants. All the new compounds were characterized through their spectral and elemental analyses. Further these compounds were tested against α-glucosidase enzyme. The compounds 2k, 2l and 2n possessed good inhibition potencies, however, compounds 2f (IC₅₀ value: 25.19 ± 0.004 μM) and 2m (IC₅₀ value: 33.62 ± 0.03 μM) were the most effective compounds of the series. Furthermore, molecular docking helped to understand the binding interactions of 2f and 2m with the understudy yeast’s α-glucosidase enzyme.     

Diterpenoids from the needles and twigs of the cultivated endangered pine Pinus kwangtungensis and their PTP1B inhibitory effects

Pinus kwangtungensis is an endangered pine species native to China. In the present study, 15 diterpenoids including three new labdane-type analogs were isolated and characterized during a pioneer phytochemical investigation on a mass-limited sample of the needles and twigs of this plant, which is growing in a Cantonese garden. The new structures, (4S,5R,9S,10R)-6-oxo-labd-7,13-dien-16,15- olid-19-oic acid (1), 15(S)-n-butoxypinusolidic acid (2), and β-d-glucopyranosyl- (4S,5R,9S,10R)-labda-8(17),13-dien-15,16-olid-19-oate (3), were established by extensive spectroscopic methods and some chemical transformations. Among the isolates, lambertianic acid (10) and cassipourol (15) showed inhibitory activities against human protein tyrosine phosphatase 1 B (PTP1B), a target for the treatment of type-II diabetes and obesity, with IC₅₀ values of 25.5 and 11.2 μM, respectively.     

Anti-hepatitis B virus constituents from the stem bark of Streblus asper

Seven compounds, (7′S,8′S)-trans-streblusol A, (7′R,8′S)-erythro-streblusol B, (7′S,8′S)-threo-streblusol B, 8′R-streblusol C, streblusquinone, (8R,8′R)-streblusol D, and streblusol E, along with 15 known compounds (8–22) were isolated from the n-butanol-soluble part of the MeOH extract of stem bark of Streblus asper. Their structures were elucidated through application of extensive spectroscopic methods, including ESI-MS and 2D NMR spectroscopy (HMQC and HMBC). The stereochemistry at the chiral centers was determined using CD spectra, as well as analyses of coupling constants and optical rotation data. The isolated lignans and allylbenzene derivatives were evaluated for their anti-HBV activities in vitro using the HBV transfected Hep G2.2.15 cell line. The most active compounds, magnolol and 9-β-xylopyranosyl-isolariciresinol, exhibited significant anti-HBV activities with IC₅₀ values of 2.03 and 6.58 μM for secretion of HBsAg, with no cytotoxicity, and of 3.76 and 24.86 μM for secretion of HBeAg, with no cytotoxicity.     

Sesquiterpenoids from the Chinese endangered plant Manglietia aromatica

One new bourbonane-type (1) and one new cadinane-type (2) sesquiterpenoids, along with one known aromodendrane-type (3) and five known megastigmane-type (4–8) compounds, were isolated from the leaves and twigs of Manglietia aromatica, a Chinese endangered plant that has not been previously phytochemically investigated. The structures and absolute configurations of the new isolates, (1R,4S,5S,6S,7S,10S)-4-hydroxy-bourbon-8-one (1) and (1R,6S,7S)-1-hydroxy- cadin-4,9-dien-8-one (2), were established by means of spectroscopic methods and a combination of experimental and calculated electronic circular dichroism (ECD). Among the isolates, compound 2 was found to show a moderate inhibitory effect against the human protein tyrosine phosphatase 1 B (PTP1B) enzyme, a target for the treatment of type-II diabetes and obesity, with an IC₅₀ value of 83.5 μM.     

Characterization of (2S,2'R,3'R)-2-(2',3'-[3H]-Dicarboxycyclopropyl)glycine Binding in Rat Brain

Abstract: [(2S,2′R,3′R)-2-(2′,3′-[³H]Dicarboxycyclopropyl)glycine ([³H]DCG IV) binding was characterized in vitro in rat brain cortex homogenates and rat brain sections. In cortex homogenates, the binding was saturable and the saturation isotherm indicated the presence of a single binding site with a K_D value of 180 ± 33 nM and a B_max of 780 ± 70 fmol/mg of protein. The nonspecific binding, measured using 100 µM LY354740, was <30%. NMDA, AMPA, kainate, l (?)-threo-3-hydroxyaspartic acid, and (S)-3,5-dihydroxyphenylglycine were all inactive in [³H]DCG IV binding up to 1 mM. However, several compounds inhibited [³H]DCG IV binding in a concentration-dependent manner with the following rank order of potency: LY341495 = LY354740 > DCG IV = (2S,1′S,2′S)-2-(2-carboxycyclopropyl)glycine > (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid > (2S,1′S,2′S)-2-methyl-2-(2-carboxycyclopropyl)glycine > l -glutamate = ibotenate > quisqualate > (RS)-α-methyl-4-phosphonophenylglycine = l (+)-2-amino-3-phosphonopropionic acid > (S)-α-methyl-4-carboxyphenylglycine > (2S)-α-ethylglutamic acid > l (+)-2-amino-4-phosphonobutyric acid. N-Acetyl-l -aspartyl-l -glutamic acid inhibited the binding in a biphasic manner with an IC₅₀ of 0.2 µM for the high-affinity component. The binding was also affected by GTPγS, reducing agents, and CdCl₂. In parasagittal sections of rat brain, a high density of specific binding was observed in the accessory olfactory bulb, cortical regions (layers 1, 3, and 4 > 2, 5, and 6), caudate putamen, molecular layers of the hippocampus and dentate gyrus, subiculum, presubiculum, retrosplenial cortex, anteroventral thalamic nuclei, and cerebellar granular layer, reflecting its preferential (perhaps not exclusive) affinity for pre- and postsynaptic metabotropic glutamate mGlu2 receptors. Thus, the pharmacology, tissue distribution, and sensitivity to GTPγS show that [³H]DCG IV binding is probably to group II metabotropic glutamate receptors in rat brain.     

High-performance liquid chromatography separation and quantitation of ofloxacin enantiomers in rat microsomes

A sensitive, simple and accurate method for determination of enantiomers of ofloxacin in microsomal incubates was developed by chiral ligand-exchange RP-HPLC with fluorescence detection to examine stereoselective metabolism of ofloxacin in the glucuronidation process. The C₁₈ stationary phase was used as analytical column. The solution of chiral mobile phase additive was made up of 6 mM l-phenylalamine mixed with 3 mM CuSO₄ in water. Mobile phase consisted of the solution of chiral mobile phase additive–methanol (86:14).The fluorescence detector was operated at λ_ex 330 nm and λ_em 505 nm. The flow-rate of mobile phase was set at 1.0 ml/min. The achiral ODS column offers good separation of the two enantiomers in less than 25 min. The recovery of the assay was 97.9±6.1% (n=10) for S-ofloxacin and 99.6±6.0% (n=10) for R-ofloxacin. The method provides a high sensitivity and good precision (RSD<10%). The LOD was 0.6 μM for both enantiomers and the LOQ was 5.70±0.45 μM (n=8) for S-ofloxacin and 5.66±0.47 μM (n=8) for R-ofloxacin. The standard curves showed excellent linearity over the concentration range 5.5–2078 μM for S-(−)-ofloxacin and R-(+)-ofloxacin. The enantioselective method developed has been applied to determine the stereoselectivity of glucuronidation metabolism of ofloxacin optical isomers in rat liver microsomes.     

Selective Inhibition of the Expression of Signal Transduction Proteins by Lithium in Nerve Growth Factor-Differentiated PC12 Cells

Abstract: Muscarinic receptor in human neuroblastoma SK-N-BE(2)C cells was identified and characterized. Treatment of the cells with carbachol evoked the generation of inositol 1,4,5-trisphosphate (IP₃) with a peak level reached at 1 min after stimulation. Carbachol increased intracellular Ca²⁺ ([Ca²⁺]_i) with an EC₅₀ value of 35 µM. In addition, carbachol produced a 1.3–3-fold increase in the cyclic AMP (cAMP) level compared with untreated control and elevated synergistically the cAMP level in the treatment with prostaglandin E₂ (PGE₂). The M₃ antagonist p-fluorohexahydrosiladifenidol (IC₅₀ = 0.5–0.8 µM) inhibited the increases in [Ca²⁺]_i, IP₃, and cAMP more effectively than the M₁ antagonist pirenzepine (IC₅₀ = 5–9 µM) and the M₂ antagonist methoctramine (IC₅₀ = 20–30 µM). The involvements of [Ca²⁺]_i elevation and protein kinase C activation induced by phospholipase C activation were tested in the carbachol-induced cAMP production. The calcium chelator BAPTA/AM (75 µM) inhibited significantly the synergistic effects of carbachol and PGE₂ on the production of cAMP, whereas the Ca²⁺ ionophore ionomycin (1 µM) clearly enhanced PGE₂-induced cAMP production. However, phorbol 12-myristate 13-acetate did not enhance PGE₂-stimulated cAMP production. These data suggest that phospholipase C-linked M₃ receptors are present and that stimulation of the receptors activates adenylyl cyclase, at least in part, by the Ca²⁺-dependent system in the neuronal cells.     

Inhibition of serotonin and norepinephrine reuptake and inhibition of phosphodiesterase by multi-target inhibitors as potential agents for depression

Compounds possessing more than one functional activity incorporated into the same molecule may have advantages in treating complex disease states. Balanced serotonin/norepinephrine reuptake inhibitors (SNRIs) (i.e., (R)- and (S)-norduloxetine) were chemically linked to a PDE4 inhibitor via a five carbon bridge. The new dual SNRI/PDE4 inhibitors (i.e., (R)-15 and (S)-15) showed moderately potent serotonin reuptake inhibition (IC₅₀ values of 442 and 404 nM, respectively) but low reuptake inhibition of norepinephrine (IC₅₀ values of 2097 and 2190 nM, respectively) in vitro. The dual SNRI/PDE4 inhibitors (i.e., (R)-15 and (S)-15) also inhibited PDE4D2 (i.e., K_i values of 23 and 45 nM, respectively). Due to their synergistic functional activity, SNRI/PDE4 inhibitors may be effective in treating diseases such as depression.     

Stereochemical Control in the Still-Wittig Rearrangement Synthesis of Cyclohexyl (Z)-Alkene Inhibitors of Pin1

Three stereoisomeric inhibitors of Pin1: (2R,5S)-, (2S,5R)- and (2S,5S)-Ac–pSer–Ψ[(Z)CH = C]–pipecolyl(Pip)–2-(2-naphthyl)ethylamine 1, that mimic L-pSer–D-Pro, D-pSer–L-Pro, and D-pSer–D-Pro amides respectively, were synthesized by a 13-step route. The newly formed stereogenic centers in the pipecolyl ring were introduced by Luche reduction, followed by stereospecific [2,3]-Still-Wittig rearrangement. The (Z)- to (E)-alkene ratio in the rearrangements were consistently 5.5 to 1. The stereochemistry at the original Ser α-carbon controlled the stereochemistry of the Luche reduction, but it did not affect the stereochemical outcome of the rearrangement, which consistently gave the (Z)-alkene. The epimerized by-product, (2S,5S)-10, resulting from the work-up after Na/NH₃ debenzylation of (2S,5R)-9, was carried on to the (2S,5S)-1 isomer. Compound (2S,5S)-10 was resynthesized from the Luche reduction by-product, (2R,3R)-3, and the stereochemistry was confirmed by comparison of the optical rotations. The IC₅₀ values for (2R,5S)-1, (2S,5R)-1 and (2S,5S)-1 Pin1 inhibition were: 52, 85, and 140 μM, respectively.     

Syntheses of 6-(2-hydroxy-6-phenylhexyl)-5,6-dihydro-2H-pyran-2-one derivatives and their cytotoxicities

The cytotoxicities against cancer cells (HL-60, HeLa) and insect cells (Sf9) of four stereoisomers of 6-(2-hydroxy-6-phenylhexyl)− 5,6-dihydro-2H-pyran-2-one (1) were evaluated, and then their structure-activity relationships examined. The 2′-dehydroxy derivative 5 of (6 R,2′R)- and (6 R,2′S)-1 showed the highest activity against HeLa cells (IC₅₀ = 1.4 μM). To evaluate the effect of the 2′-hydroxy group of 1, 6R-and 6S-oxetane derivatives were also synthesized and their activities examined. Against HeLa and HL-60 cells, the activities of the less potent stereoisomers were enhanced 3–4-fold by the introduction of the oxetane moieties at the 2′-position. Against the insect cell line (Sf9), phenyl derivative 7 showed the highest activity with an IC₅₀ value of 8.0 μM.     

Synthesis and antibacterial activity of alaremycin derivatives for the porphobilinogen synthase

The preparation and the antibacterial activity of alaremycin derivatives such as their CF₃-derivatives and (R)- and (S)-4-oxo-5-acetylaminohexanoic acid for the porphobilinogen synthase (PBGS), were described. The IC₅₀ values of the antibacterial activity of the prepared materials for the inhibitor of PBGS, were determined using PBGS assay.     

The interaction of cytosolic epoxide hydrolase with chiral epoxides

• 1.1. The kinetic parameters of the cytosolic epoxide hydrolase were examined with two sets of spectrophotometric substrates. The (2S,3S)- and (2R,3R)-enantiomers of 4-nitrophenyl trans-2,3-epoxy-3-phenylpropyl carbonate had a K_mof 33 and 68 μm and a V_max of 16 and 27 μmol/min/mg, respectively. With the (2S,3S)- and (2R,3R)- enantiomers of 4-nitrophenyl trans-2,3-epoxy-3-(4-nitrophenyl)propyl carbonate, cytosolic epoxide hydrolase had a K_M of 8.0 and 15 μM and a V_max of 7.8 and 5.0 μmol/min/mg, respectively.
• 2.2. Glycidyl 4-nitrobenzoate had the lowest I₅₀ of the compounds tested in the glycidyl 4-nitrobenzoate series (I₅₀= 140 μM). The I₅₀ of the (2R)-enantiomer was 3.7-fold higher. The inhibitor with the lowest i₅₀ in the glycidol series, and the lowest I₅₀ of any compound tested, was (2S,3S)-3-(4-nitrophenyl)glycidol (I₅₀ = 13.0μM). It also showed the greatest difference in I₅₀ between the enantiomers (330-fold).
• 3.3. All enantiomers of glycidyl 4-nitrobenzoates and _trans-3-phenylglycidols gave differential inhibition of cytosolic epoxide hydrolase. However, neither the (S,S)-/(S)- or (R,R)-/(R)-enantiomer always had the lower I₅₀.
• 4.4. Addition of one or more methyl groups to either enantiomer of glycidyl 4-nitrobenzoate resulted in increased I₅₀. However, addition of a methyl group to C₂ of either enantiomer of 3-phenylglycidol resulted in a decreased I₅₀. Finally, when the hydroxyl group of trans-3-(4-nitrophenyl)glycidol was esterified the I₅₀ of the (2S,3S)- but not the (2R,3R)-enantiomer increased.