An extensive cocktail approach for rapid risk assessment of in vitro CYP450 direct reversible inhibition by xenobiotic exposure

Spaggiari, Dany; Daali, Youssef; Rudaz, Serge

doi:10.1016/j.taap.2016.04.013

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Title		An extensive cocktail approach for rapid risk assessment of in vitro CYP450 direct reversible inhibition by xenobiotic exposure
Authors		Spaggiari, Dany Daali, Youssef Rudaz, Serge
Published in		Toxicology and applied pharmacology. 2016, vol. 302, p. 41-51
Abstract		Acute exposure to environmental factors strongly affects the metabolic activity of cytochrome P450 (P450). As a consequence, the risk of interaction could be increased, modifying the clinical outcomes of a medication. Because toxic agents cannot be administered to humans for ethical reasons, in vitro approaches are therefore essential to evaluate their impact on P450 activities. In thiswork, an extensive cocktail mixturewas developed and validated for in vitro P450 inhibition studies using human liver microsomes (HLM). The cocktail comprised eleven P450-specific probe substrates to simultaneously assess the activities of the following isoforms: 1A2, 2A6, 2B6, 2C8, 2C9, 2C19, 2D6, 2E1, 2J2 and subfamily 3A. The high selectivity and sensitivity of the developed UHPLC-MS/MS methodwere critical for the success of this methodology, whose main advantages are: (i) the use of eleven probe substrates with minimized interactions, (ii) a low HLMconcentration, (iii) fast incubation (5 min) and (iv) the use of metabolic ratios as microsomal P450 activities markers. This cocktail approach was successfully validated by comparing the obtained IC50 values for model inhibitorswith those generatedwith the conventional single probe methods. Accordingly, reliable inhibition values could be generated 10-fold faster using a 10-fold smaller amount of HLM compared to individual assays. This approach was applied to assess the P450 inhibition potential of widespread insecticides, namely, chlorpyrifos, fenitrothion, methylparathion and profenofos. In all cases, P450 2B6 was the most affected with IC50 values in the nanomolar range. For the first time, mixtures of these four insecticides incubated at lowconcentrations showed a cumulative inhibitory in vitro effect on P450 2B6.
Keywords		Risk assessment — Cytochrome P450 — UHPLC-MS/MS — IC50 — Human liver microsomes — Pesticides
Identifiers		DOI: 10.1016/j.taap.2016.04.013
Full text		Article (Published version) (3.9 MB) - Limited access to UNIGE
Structures		Faculté des sciences / Section des sciences pharmaceutiques
Research groups		Groupe Desmeules Jules (pharmacologie/toxicologie) (567) Pharmaco-omiques et médecine de précision (1003) Sciences analytiques
Citation (ISO format)		SPAGGIARI, Dany, DAALI, Youssef, RUDAZ, Serge. An extensive cocktail approach for rapid risk assessment of in vitro CYP450 direct reversible inhibition by xenobiotic exposure. In: Toxicology and applied pharmacology, 2016, vol. 302, p. 41-51. doi: 10.1016/j.taap.2016.04.013 https://archive-ouverte.unige.ch/unige:85973