Abstract
The rhesus monkey (Macaca mulatta) is a primate species used extensively as a preclinical safety species in drug development. In this report, we describe the cloning, expression, and characterization of CYP3A64 (AY334551), a CYP3A4 homolog expressed in rhesus liver. The deduced amino acid sequence was found to be 93% similar to human CYP3A4, 83% similar to human CYP3A5, and identical to the previously reported cynomolgus monkey CYP3A8 (Komori et al., 1992). The substrate specificity of CYP3A64 for testosterone (0–250 μM), midazolam (0–200 μM), nifedipine (0–200 μM), and 7-benzoxy-4-trifluoromethylcoumarin (0–200 μM) were compared with recombinant enzymes from rat (CYP3A1, CYP3A2), dog (CYP3A12, CYP3A26), rabbit (CYP3A6), and human (CYP3A4, CYP3A5). Immunoinhibition and chemical inhibition of CYP3A64 was demonstrated using the inhibitory monoclonal antibody (MAb) 10-1-1 (anti-3A4) and ketoconazole (0–10 μM). The utility of CYP3A64 to be used as a standard in monkey induction assays was shown and the concentration of CYP3A64 protein in rhesus liver microsomes was estimated to be 72 pmol/mg protein. In summary, these results support the utilization of rhesus monkey CYP3A64 for in vitro drug metabolism studies and provide a more complete understanding of CYP3A substrate specificities and species differences in metabolic capabilities.
Footnotes
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Article, publication date, and citation information can be found at http://dmd.aspetjournals.org.
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doi:10.1124/dmd.106.009977.
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ABBREVIATIONS: P450, cytochrome P450; bp, base pair(s); BFC, 7-benzoxy-4-trifluoromethylcoumarin; HFC, 7-hydroxy-4-trifluoromethylcoumarin; OR, oxidoreductase; MAb, monoclonal antibody; PCR, polymerase chain reaction; KPi, potassium phosphate; OH, hydroxy; LC-MS/MS, liquid chromatography-tandem mass spectrometry; APCI, atmospheric pressure chemical ionization; MDZ, midazolam; RSS, residual sum of squares.
- Received February 23, 2006.
- Accepted June 27, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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