Abstract
Genotyping of the highly polymorphic cytochrome P450 2D6 (CYP2D6) permits a gross classification of individual phenotype (viz. ultra-rapid, extensive, and poor metabolizers). It does not, however, provide a precise prediction of CYP2D6 activity, particularly in an individual possessing at least one functional CYP2D6 allele. It has been suggested that the level of mRNA expression or enzyme activity in lymphocytes, isolated from blood, could potentially provide a better quantitative estimate of in vivo hepatic enzymatic activity in human subjects. Although short sequences of CYP2D6 mRNA have been detected in human lymphocytes by reverse transcriptase-polymerase chain reaction (RT-PCR) that suggests the potential use of lymphocyte RNA as a readily accessible biomarker, it is not known whether a functional enzyme is expressed in human lymphocytes. In this study, human lymphocyte activity was assessed with a CYP2D6-specific, high-turnover probe substrate that is severalfold more sensitive than traditional markers of CYP2D6 (e.g., dextromethorphan). CYP2D6 catalytic activity could not be detected in homogenates of human lymphocytes, even at high protein concentrations and with supplementation of enzyme cofactors. Further RT-PCR analysis of lymphocytes collected from eight human donors revealed the presence of only a fragment, but not the complete transcript, of CYP2D6 mRNA. Northern blot RNA transcript analysis also failed to indicate the presence of the full-length transcript in lymphocytes. Collectively, these data indicate that human lymphocytes express neither the full-length CYP2D6 mRNA transcript nor functional enzyme activity. Therefore, the utility of lymphocytes as a functional biomarker for CYP2D6 enzyme activity is not clear at present.
Footnotes
-
↵1 Abbreviations used are: P450, cytochrome P450; bp, base pair(s); RT-PCR, reverse transcriptase-polymerase chain reaction; kb, kilobase(s); NPS-1378, HNF4, hepatocyte nuclear factor 4; R-568, (R)-N-(3-methoxy-α-phenylethyl)-3-(2′-chlorophenyl)-1-(propylamine hydrocholoride).
-
Supported in part by National Institutes of Health Grants AI31854, HL 56548, and GM62883, and by Amgen Inc. L.A.M. is also supported in part by National Institutes of Health Pharmacological Sciences Training Grant GM07750.
- Received March 13, 2003.
- Accepted May 16, 2003.
- The American Society for Pharmacology and Experimental Therapeutics
DMD articles become freely available 12 months after publication, and remain freely available for 5 years.Non-open access articles that fall outside this five year window are available only to institutional subscribers and current ASPET members, or through the article purchase feature at the bottom of the page.
|