Hypha present poster on PolyCYPs™ cytochrome P450 kit at ISSX meeting in Cologne June 2017
A cell-free kit of cytochrome P450 enzymes and ferredoxin /ferredoxin reductase partners, termed PolyCYPsTM, is being developed for generating scalable quantities of oxidised metabolites. Cytochromes in the kit have been derived from some of Hypha’s talented biotransforming bacteria and are capable of generating human and other mammalian metabolites of drug compounds. The hydroxylating abilities of one P450 enzyme in the kit, which has been cloned from an actinomycete species into E.coli together with redox partners, is illustrated using various drugs including tivantinib. Tivantinib is an experimental anti-cancer drug which is extensively metabolised in humans, with two major circulating metabolites M4 and M51. M4 and M5 are stereoisomers hydroxylated at the benzyl position of the tricyclic ring of Tivantinib and comprise 19% and 12% of the total area under the concentration-time curve (AUC) respectively2, implicating these metabolites under the FDA MIST guideline. Recombinant enzyme PolyCYPsTM-6.1 was able to produce all four of the hydroxylated human metabolites originally observed in the wild-type microbial biotransformation, from which the cytochrome was derived. As well as utility for providing sufficient material for MetID, reactions can be scaled to produce milligram to gram quantities of metabolites and novel derivatives for further evaluation.
1 Murai et al., 2014. Metabolism and disposition of [14C] tivantinib after oral administration to humans, dogs and rats. Clinical Pharmacokinetics and Metabolism 44 (11), 996-1008.
2 Nishiya et al., 2016. Stereoselective hydroxylation by CYP2C19 and oxidation by ADH4 in the in vitro metabolism of tivantinib. Xenobiotica 46 (11), 967-976.