Guinea pig whole blood 5-lipoxygenase assay: utility in the assessment of potential 5-lipoxygenase inhibitors

Spaethe, Stephen M.; Snyder, David W.; Pechous, Penelope A.; Clarke, Trafford; VanAlstyne, Eldwin L.

doi:10.1016/0006-2952(92)90302-y

Cited by 14 publications

(5 citation statements)

References 10 publications

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“…Samples were then centrifuged at 1,500 rpm (300 g) for 10 min at 4°C and the supernatant diluted between 20 and 50-fold (batch dependent) for LTB4 detection, using an ELISA detection kit (Cayman Chemicals Inc.). Inhibitors were added at 10 µM concentrations [36]–[38] and the IC 50 values were generated using a one point IC 50 estimation equation.…”

Section: Methodsmentioning

confidence: 99%

Discovery of a Novel Dual Fungal CYP51/Human 5-Lipoxygenase Inhibitor: Implications for Anti-Fungal Therapy

et al. 2013

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We report the discovery of a novel dual inhibitor targeting fungal sterol 14α-demethylase (CYP51 or Erg11) and human 5-lipoxygenase (5-LOX) with improved potency against 5-LOX due to its reduction of the iron center by its phenylenediamine core. A series of potent 5-LOX inhibitors containing a phenylenediamine core, were synthesized that exhibit nanomolar potency and >30-fold selectivity against the LOX paralogs, platelet-type 12-human lipoxygenase, reticulocyte 15-human lipoxygenase type-1, and epithelial 15-human lipoxygenase type-2, and >100-fold selectivity against ovine cyclooxygenase-1 and human cyclooxygnease-2. The phenylenediamine core was then translated into the structure of ketoconazole, a highly effective anti-fungal medication for seborrheic dermatitis, to generate a novel compound, ketaminazole. Ketaminazole was found to be a potent dual inhibitor against human 5-LOX (IC50 = 700 nM) and CYP51 (IC50 = 43 nM) in vitro. It was tested in whole blood and found to down-regulate LTB4 synthesis, displaying 45% inhibition at 10 µM. In addition, ketaminazole selectively inhibited yeast CYP51 relative to human CYP51 by 17-fold, which is greater selectivity than that of ketoconazole and could confer a therapeutic advantage. This novel dual anti-fungal/anti-inflammatory inhibitor could potentially have therapeutic uses against fungal infections that have an anti-inflammatory component.

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Section: Methodsmentioning

confidence: 99%

Discovery of a Novel Dual Fungal CYP51/Human 5-Lipoxygenase Inhibitor: Implications for Anti-Fungal Therapy

et al. 2013

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“…It is one of the best models in studies on the simultaneous effects of drugs on cyclooxygenase and 5-lipoxygenase both in vitro (Patrignani et al 1990;Spaethe et al 1992) and ex vivo (Sirois et al 1991;Surette et al 1994) taking into account the complex interaction between the different cell types capable of modulating arachidonic acid metabolism. The cyclooxygenase and 5-lipoxygenase pathways of arachidonic acid metabolism are shown in fig.…”

Section: Introductionmentioning

confidence: 99%

Nicotinic Acid and Pyridoxine Modulate Arachidonic Acid Metabolism in vitro and ex vivo in Man

Saareks

Muchá²,

Sievi

et al. 1999

Pharmacology & Toxicology

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The in vitro effects of nicotinic acid (10-1000 microM), pyridoxine (0.1-500 microM) and pyridoxal-5'-phosphate (0.1-500 microM) and the ex vivo effects of nicotinic acid (2500 mg orally during 12 h) and pyridoxine (600 mg orally daily for seven days) on arachidonic acid metabolism were investigated in calcium ionophore A23187 (calcimycin)-stimulated human whole blood. In vitro nicotinic acid stimulated prostaglandin E2, thromboxane B2 and leukotriene E4 synthesis. Pyridoxine at all concentrations and pyridoxal-5'-phosphate at the highest concentration stimulated prostaglandin E2 and thromboxane B2 production, but had no effect on leukotriene E4 synthesis. Nicotinic acid treatment increased ex vivo prostaglandin E2, thromboxane B2 and leukotriene E4 synthesis to 185%, 165% and 175% of the initial values, respectively. In the pyridoxine-treated subjects, ex vivo prostaglandin E2, thromboxane B2 and leukotriene E4 synthesis was decreased after seven days to 75%, 65% and 45% of the initial values, respectively. In the present study the effects of nicotinic acid on the 5-lipoxygenase pathway in arachidonic acid metabolism were studied for the first time and the drug was found to stimulate this pathway in vitro and ex vivo. In vitro pyridoxine and pyridoxal-5'-phosphate had no effect on the 5-lipoxygenase pathway. The inhibition of leukotriene synthesis by pyridoxine ex vivo might be of therapeutic importance.

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“…Inhibitors were added at 10 or 15 μ m concentrations (0.5 μ m for control compound setileuton), and IC 50 values were generated using a one point IC 50 estimation equation. Drug efficacy was determined using a minimum of two different donors .…”

Section: Methodsmentioning

confidence: 99%

Enzymatic Studies of Isoflavonoids as Selective and Potent Inhibitors of Human Leukocyte 5‐Lipo‐Oxygenase

et al. 2014

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Continuing our search to find more potent and selective 5-LOX inhibitors, we present now the enzymatic evaluation of seventeen isoflavones (IR) and nine isoflavans (HIR), and their in vitro and in cellulo potency against human leukocyte 5-LOX. Of the 26 compounds tested, 10 isoflavones and 9 isoflavans possessed micromolar potency, but only three were selective against 5-LOX (IR-2, HIR-303 and HIR-309), with IC 50 values at least 10 times lower than those of 12-LOX, 15-LOX-1 and 15-LOX-2. Of these three, IR-2 (6,7-dihydroxy-4-methoxy-isoflavone, known as texasin) was the most selective 5-LOX inhibitor, with over 80-fold potency difference, SMD studies supported these findings. The presence of the catechol group on ring A (6,7-dihydroxy versus 7,8-dihydroxy) correlated with their biological activity, but the reduction of ring C, converting the isoflavones to isoflavans, and the substituent positions on ring B did not affect their potency against 5-LOX. Computer docking of the three most selective inhibitors to the crystal structure of human leukocyte 5-LOX confirmed these SAR experimental results. Two of the most potent/selective inhibitors (HIR-303 and HIR-309) were reductive inhibitors and were potent against 5-LOX in human whole blood, indicating that isoflavans can be potent and selective inhibitors against human leukocyte 5-LOX in vitro and in cellulo.

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Guinea pig whole blood 5-lipoxygenase assay: utility in the assessment of potential 5-lipoxygenase inhibitors

Cited by 14 publications

References 10 publications

Discovery of a Novel Dual Fungal CYP51/Human 5-Lipoxygenase Inhibitor: Implications for Anti-Fungal Therapy

Discovery of a Novel Dual Fungal CYP51/Human 5-Lipoxygenase Inhibitor: Implications for Anti-Fungal Therapy

Nicotinic Acid and Pyridoxine Modulate Arachidonic Acid Metabolism in vitro and ex vivo in Man

Enzymatic Studies of Isoflavonoids as Selective and Potent Inhibitors of Human Leukocyte 5‐Lipo‐Oxygenase

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