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Cape Town, South Africa

Fasinu P.,Stellenbosch University | Bouic P.J.,Synexa Life science | Bouic P.J.,Stellenbosch University | Rosenkranz B.,Stellenbosch University
Current Drug Metabolism | Year: 2012

Early understanding of the metabolic pathway and potential interaction of new drug candidates with other drugs is one of the goals of preclinical studies in the drug discovery process. Although other body organs are involved in drug biotransformation, the liver is the predominant organ of metabolism for a wide range of endogenous compounds and xenobiotics. The set of enzymes contained in the cytochrome P450 superfamily present predominantly in the liver have been identified as the single most important agent of drug metabolism and have formed the bedrock of most matured technologies for in vitro drug biotransformation studies. With the development of a number of liver-based technologies, in vitro metabolism has gained significant popularity in the past three decades. This has come in response to several demanding factors including the questionable relevance of data from animal studies; the high cost and stringent regulatory and ethical requirement, as well as safety issues involved with studies using human subjects; and the need for high throughput due to the wide range of chemical entities for routine investigations. These technologies which vary from whole liver to subcellular fractions have found ready application in generating the desired information on the substrate and inhibitor specificity of most metabolic enzymes. This paper reviews such technologies as isolated fresh liver; liver slices; primary, cultured and cryopreserved hepatocytes; microsomes; cytosolic fractions; and purified or heterologously expressed drug-metabolizing enzymes. It highlights the general principles of in vitro enzyme kinetics and the factors that determine the choice of each in vitro technology for biotransformation studies. © 2012 Bentham Science Publishers.


Fasinu P.S.,Stellenbosch University | Gutmann H.,Novartis | Schiller H.,Novartis | Bouic P.J.,Synexa Life science | And 2 more authors.
Pharmaceutical Biology | Year: 2013

Context: Aqueous decoction of Hypoxis hemerocallidea Fisch. & C.A. Mey. (Hypoxidaceae) (Hypoxis) is widely consumed in Southern Africa by people living with HIV/AIDS, some of whom are on ARV and other medications. Objective: The aim of this study was to investigate the potential of the crude aqueous extracts of Hypoxis to inhibit major forms of CYP450 and transport proteins. Materials and methods: Corms of Hypoxis were water-extracted and incubated (in graded concentrations: 1 - 100 μg/mL) with human liver microsomes (20 min) to monitor the effects on phenacetin O-deethylation, coumarin 7-hydroxylation, bupropion hydroxylation, paclitaxel 6α-hydroxylation, diclofenac 4′-hydroxylation, S-mephenytoin 4′-hydroxylation, bufuralol 1′-hydroxylation, chlorzoxazone 6-hydroxylation, midazolam 1′-hydroxylation and testosterone 6β-hydroxylation as markers for the metabolic activities of CYP1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1 and 3A4/5, respectively. The generation of metabolites were monitored and quantified with the aid of LC-MS/MS. The potential of the extracts to inhibit human ATP-binding cassette transporter activity was assessed using recombinant MDCKII and LLC-PK1 cells over-expressing human breast cancer resistant protein and human P-glycoprotein , respectively (with Ko143 and cyclosporin A as positive controls). Similar assessment was performed with human organic anion transporting polypeptide (OATP1B1 and OATP1B3) using recombinant HEK293 cells over-expressing OATP1B1 and OATP1B3, respectively (with rifamycin and 10 μM atorvastatin as positive controls). Results: Extracts of Hypoxis inhibited the production of the metabolites of the substrates of the following enzymes (as compared to controls) with the indicated IC50 values (μg/mL): CYP1A2 (120.6), CYP2A6 (210.8), CYP2B6 (98.5), CYP2C8 (195.2), CYP2C9 (156) and CYP3A4/5 (185.4). The inhibition of the uptake activity of OATP1B1 and OATP1B3 were also observed with IC50 values of 93.4 and 244.8 μg/mL, respectively. Discussion: Extract concentrations higher than the estimated IC50 values are achievable in the gastrointestinal tract when traditional doses of Hypoxis are considered. This may have profound effects on presystemic metabolism of the drug substrates. If absorbed, systemic inhibition of metabolic enzymes/transporters by Hypoxis may be expected. Conclusion: The result suggests that there is the potential for HDI between Hypoxis and the substrates of the affected enzymes/transporters, if sufficient in vivo concentration of Hypoxis extracts is attained. © 2013 Informa Healthcare USA, Inc.


Awortwe C.,Stellenbosch University | Bouic P.J.,Synexa Life science | Bouic P.J.,Stellenbosch University | Masimirembwa C.M.,Harare Institute of Technology | Rosenkranz B.,Stellenbosch University
Drug Metabolism Letters | Year: 2013

The purpose of this study was to evaluate the potential risk of common herbal medicines used by HIV-infected patients in Africa for herb-drug interactions (HDI). High throughput screening assays consisting of recombinant Cytochrome P450 enzymes (CYPs) and fluorescent probes, and parallel artificial membrane permeability assays (PAMPA) were used. The potential of herbal medicines to cause HDI was ranked according to FDA guidelines for reversible inhibition and categorization of time dependent inhibition was based on the normalized ratio. CYPs 1A2 and 3A4 were most inhibited by the herbal extracts. H. hemerocallidea (IC50 = 0.63 μg/mL and 58 μg/mL) and E. purpurea (IC50 = 20 μg/mL and 12 μg/mL) were the potent inhibitors of CYPs 1A2 and 3A4 respectively. L. frutescens and H. hemerocallidea showed clear time dependent inhibition on CYP3A4. Furthermore, the inhibitory effect of both H. hemerocallidea and L. frutescens before and after PAMPA were identical. The results indicate potential HDI of H. hemerocallidea, L. frutescens and E. purpurea with substrates of the affected enzymes if maximum in vivo concentration is achieved. © 2013 Bentham Science Publishers.


Awortwe C.,Stellenbosch University | Manda V.K.,University of Mississippi | Avonto C.,University of Mississippi | Khan S.I.,University of Mississippi | And 5 more authors.
Xenobiotica | Year: 2015

1.This study investigated the mechanism underlying Echinacea-mediated induction of CYP1A2, CYP3A4 and MDR1 in terms of human pregnane X receptor (PXR) activation. 2.Crude extracts and fractions of Echinacea purpurea were tested for PXR activation in HepG2 cells by a reporter gene assay. Quantitative real-time PCR was carried out to determine their effects on CYP1A2 and CYP3A4 mRNA expressions. Capsules and fractions were risk ranked as high, intermediate and remote risk of drug-metabolizing enzymes induction based on EC50 values determined for respective CYPs. 3.Fractions F1, F2 and capsule (2660) strongly activated PXR with 5-, 4- and 3.5-fold increase in activity, respectively. Echinacea preparations potentiated up-regulation of CYP1A2, CYP3A4 and MDR1 via PXR activation. 4.Thus E. purpurea preparations cause herb-drug interaction by up-regulating CYP1A2, CYP3A4 and P-gp via PXR activation. © 2014 Informa UK Ltd. All rights reserved: reproduction in whole or part not permitted.


Fasinu P.S.,Stellenbosch University | Fasinu P.S.,University of Mississippi | Bouic P.J.,Synexa Life science | Bouic P.J.,Stellenbosch University | Rosenkranz B.,Stellenbosch University
African Journal of Traditional, Complementary and Alternative Medicines | Year: 2014

Background: Studies have suggested an increasing practice of concurrent herb-drug consumption. One of the major clinical risks of such concomitant herb-drug use is pharmacokinetic herb-drug interaction (HDI). This is brought about by the ability of phytochemicals to inhibit or induce the activity of metabolic enzymes. The aim of this study was to investigate the potential of the crude aqueous extracts of three popular medicinal herbs used in South Africa to inhibit major cytochrome P450 (CYP) enzymes. Materials and Methods: The extracts of Bowiea volubilis, Spirostachys africana and Tulbaghia violacea were incubated with human liver microsomes (HLM) to monitor the phenacetin O-deethylation, diclofenac 4’-hydroxylation, S-mephenytoin 4’-hydroxylation and testosterone 6β-hydroxylation as respective probe reactions for CYP1A2, CYP2C9, CYP2C19 and CYP3A4. The inhibitory activity, where observed, was profiled against the extract concentration. Results: Extracts of Bowiea volubilis inhibited the metabolic activity of CYP1A2 and CYP3A4 with IC50 values of 92.3 ± 5.5 μg/mL and 8.1 ±0.6 μg/mL respectively. Similar observation with Spirostachys africana showed inhibitory activity against CYP1A2 and CYP3A4 with respective IC50 values of 14.3 ± 0.6 μg/mL and 47.4 ± 2.4 μg/mL. Tulbaghia violacea demonstrated relatively weak inhibitory activity against CYP1A2 (767.4 ± 10.8 μg/mL) and CYP2C9 (921 ± 15.3 μg/mL). Conclusion: The results suggest the potential for HDI between the herbs and the substrates of the affected enzymes, if sufficient in vivo concentration is attained. © 2014, African Ethnomedicines Network. All rights reserved.

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