Functional cytochrome p450 in vitro assay
AssigneePharmGenetix GmbH
InventorsCharity NOFZIGER, Markus PAULMICHL, Simone VANONI
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Abstract
An in vitro method for determining the functionality of a cytochrome P450 enzyme variant or an orthologue thereof to metabolize at least one substrate. In particular, the cytochrome P450 enzyme variant is a CYP2D6 variant. Further, a kit including the required reagents and biological material, i.e. cells, to carry out the method.
Claims20
-19. (canceled)
An in vitro method for determining the functionality of a CYP450 variant or an orthologue thereof to metabolize at least one substrate comprising the steps of:
- i) providing human target cells that do not express an enzyme of the cytochrome P450 family (CYP450) or express a non-functional variant of a CYP450 enzyme or express a CYP450 that is not involved in the metabolism of the substrate
- ii) transiently transfecting the target cells with at least one nucleic acid encoding at least one CYP450 variant or an orthologue thereof,
- iii) culturing the transfected target cells in the presence of the at least one substrate under conditions that allow for expression of the CYP450 variant or orthologue thereof and metabolism of the substrate;
- and
- iv) determining the level of CYP450 induced metabolism of the substrate, wherein detection of CYP450 induced metabolism of the substrate is indicative of CYP450 activity.
A kit for determining the functionality of at least one CYP450 variant or an orthologue thereof comprising:
- a) cells that do not express an enzyme of the cytochrome P450 family (CYP450) or express a non-functional variant of a CYP450 enzyme;
- and
- b) vectors encoding CYP450 control variants.
Dependencies are inferred from the claim text.