European Journal of Clinical Pharmacology

, Volume 61, Issue 12, pp 887–892

Rapid genotyping for relevant CYP1A2 alleles by pyrosequencing

  • Carsten Skarke
  • Anja Kirchhof
  • Gerd Geisslinger
  • Jörn Lötsch



To develop a rapid and reliable screening method for identifying the relevant cytochrome P450 (CYP) 1A2 alleles CYP1A2*1D (−2467Tdel), *1F (−163A>C), and *1K (−739T>G, −729C>T, −163A>C) that are in linkage disequilibrium with the functionally relevant CYP1A2 polymorphisms and therefore are considered to be predictive for the CYP1A2 phenotype.


CYP1A2 single nucleotide polymorphisms (SNPs) −2467Tdel, −739T>G, −729C>T, and −163A>C were screened for in 495 healthy Caucasian volunteers using newly developed pyrosequencing duplex and simplex assays. Conventional sequencing of randomly selected samples served as quality control.


Frequencies were 7.9% for CYP1A2*1D, 31.8% for *1F, and 0.4% for *1K. The observed distribution of homozygous and heterozygous carriers of the alleles corresponded to the predicted one according to the Hardy-Weinberg law. It also corresponded to reported allelic frequencies from Caucasians but differed significantly from the distribution seen in other ethnicities. The most frequent haplotype was −2467T/−739T/−729C/−163A (allelic frequency 61.6%), followed by −2467T/−739T/−729C/−163C (30.5%), −2467Tdel/−739T/−729C/−163A (5.1%), −2467Tdel/−739G/−729C/−163A (1.2%), and −2467Tdel/−739T/−729C/−163C (1.1%). Complete linkage disequilibrium (value of D’ nearly 1) existed between −2467Tdel, −739T>G, and −729C>T and between −729T>G and −163A>C.


Pyrosequencing facilitates rapid and reliable detection of those CYP1A2 alleles that, based on current knowledge, can be considered predictive for the CYP1A2 phenotype.


CYP1A2 Pyrosequencing Pharmacogenetics 


  1. 1.
    Brosen K (1995) Drug interactions and the cytochrome P450 system. The role of cytochrome P450 1A2. Clin Pharmacokinet 29 (Suppl 1):20–25PubMedCrossRefGoogle Scholar
  2. 2.
    Sachse C, Brockmoller J, Bauer S, Roots I (1999) Functional significance of a C->A polymorphism in intron 1 of the cytochrome P450 CYP1A2 gene tested with caffeine. Br J Clin Pharmacol 47(4):445–449CrossRefPubMedGoogle Scholar
  3. 3.
    Nordmark A, Lundgren S, Ask B, Granath F, Rane A (2002) The effect of the CYP1A2 *1F mutation on CYP1A2 inducibility in pregnant women. Br J Clin Pharmacol 54(5):504–510PubMedCrossRefGoogle Scholar
  4. 4.
    Aklillu E, Carrillo JA, Makonnen E, et al (2003) Genetic polymorphism of CYP1A2 in Ethiopians affecting induction and expression: characterization of novel haplotypes with single-nucleotide polymorphisms in intron 1. Mol Pharmacol 64(3):659–669CrossRefPubMedGoogle Scholar
  5. 5.
    Shimoda K, Someya T, Morita S, et al. (2002) Lack of impact of CYP1A2 genetic polymorphism (C/A polymorphism at position 734 in intron 1 and G/A polymorphism at position −2964 in the 5′–flanking region of CYP1A2) on the plasma concentration of haloperidol in smoking male Japanese with schizophrenia. Prog Neuropsychopharmacol Biol Psychiatry 26(2):261–265CrossRefPubMedGoogle Scholar
  6. 6.
    Nakajima M, Yokoi T, Mizutani M, Kinoshita M, Funayama M, Kamataki T (1999) Genetic polymorphism in the 5′–flanking region of human CYP1A2 gene: effect on the CYP1A2 inducibility in humans. J Biochem (Tokyo) 125(4):803–808Google Scholar
  7. 7.
    Allorge D, Chevalier D, Lo–Guidice JM, et al (2003) Identification of a novel splice–site mutation in the CYP1A2 gene. Br J Clin Pharmacol 56(3):341–344CrossRefPubMedGoogle Scholar
  8. 8.
    Murayama N, Soyama A, Saito Y, et al (2004) Six novel nonsynonymous CYP1A2 gene polymorphisms: catalytic activities of the naturally occurring variant enzymes. J Pharmacol Exp Ther 308(1):300–306CrossRefPubMedGoogle Scholar
  9. 9.
    Sachse C, Bhambra U, Smith G, et al (2003) Polymorphisms in the cytochrome P450 CYP1A2 gene (CYP1A2) in colorectal cancer patients and controls: allele frequencies, linkage disequilibrium and influence on caffeine metabolism. Br J Clin Pharmacol 55(1):68–76PubMedCrossRefGoogle Scholar
  10. 10.
    Ronaghi M (2001) Pyrosequencing sheds light on DNA sequencing. Genome Res 11(1):3–11CrossRefPubMedGoogle Scholar
  11. 11.
    Lewontin RC (1964) The interaction of selection and linkage. II. Optimum models. Genetics 50:757–782Google Scholar
  12. 12.
    Gaut BS, Long AD (2003) The lowdown on linkage disequilibrium. Plant Cell 15(7):1502–1506CrossRefPubMedGoogle Scholar
  13. 13.
    Stephens M, Donnelly P (2003) A comparison of Bayesian methods for haplotype reconstruction from population genotype data. Am J Hum Genet 73(5):1162–1169CrossRefPubMedGoogle Scholar
  14. 14.
    Stephens M, Smith NJ, Donnelly P (2001) A new statistical method for haplotype reconstruction from population data. Am J Hum Genet 68(4):978–989CrossRefPubMedGoogle Scholar
  15. 15.
    Soyama A, Saito Y, Hanioka N, et al (2005) Single nucleotide polymorphisms and haplotypes of CYP1A2 in a Japanese population. Drug Metab Pharmacokinet 20(1):24–33Google Scholar
  16. 16.
    Dandara C, Basvi PT, Bapiro TE, Sayi J, Hasler JA (2004) Frequency of −163 C>A and 63 C>G single nucleotide polymorphism of cytochrome P450 1A2 in two African populations. Clin Chem Lab Med 42(8):939–941CrossRefPubMedGoogle Scholar
  17. 17.
    Chida M, Yokoi T, Fukui T, Kinoshita M, Yokota J, Kamataki T (1999) Detection of three genetic polymorphisms in the 5′–flanking region and intron 1 of human CYP1A2 in the Japanese population. Jpn J Cancer Res 90(9):899–902PubMedGoogle Scholar
  18. 18.
    Basile VS, Ozdemir V, Masellis M, et al (2000) A functional polymorphism of the cytochrome P450 1A2 (CYP1A2) gene: association with tardive dyskinesia in schizophrenia. Mol Psychiatry 5(4):410–417CrossRefPubMedGoogle Scholar
  19. 19.
    Christiansen L, Bygum A, Jensen A, et al (2000) Association between CYP1A2 polymorphism and susceptibility to porphyria cutanea tarda. Hum Genet 107(6):612–614PubMedCrossRefGoogle Scholar
  20. 20.
    Aitchison KJ, Gonzalez FJ, Quattrochi LC, et al (2000) Identification of novel polymorphisms in the 5′ flanking region of CYP1A2, characterization of interethnic variability, and investigation of their functional significance. Pharmacogenetics 10(8):695–704PubMedCrossRefGoogle Scholar
  21. 21.
    Han XM, Ou–Yang DS, Lu PX, et al (2001) Plasma caffeine metabolite ratio (17X/137X) in vivo associated with G–2964A and C734A polymorphisms of human CYP1A2. Pharmacogenetics 11(5):429–435CrossRefPubMedGoogle Scholar
  22. 22.
    Goodman MT, McDuffie K, Kolonel LN, et al (2001) Case-control study of ovarian cancer and polymorphisms in genes involved in catecholestrogen formation and metabolism. Cancer Epidemiol Biomarkers Prev 10(3):209–216Google Scholar
  23. 23.
    Cornelis MC, El–Sohemy A, Campos H (2004) Genetic polymorphism of CYP1A2 increases the risk of myocardial infarction. J Med Genet 41(10):758–762CrossRefPubMedGoogle Scholar
  24. 24.
    Schrenk D, Brockmeier D, Morike K, Bock KW, Eichelbaum M (1998) A distribution study of CYP1A2 phenotypes among smokers and non-smokers in a cohort of healthy Caucasian volunteers. Eur J Clin Pharmacol 53(5):361–367CrossRefPubMedGoogle Scholar
  25. 25.
    Rasmussen BB, Brosen K (1996) Determination of urinary metabolites of caffeine for the assessment of cytochrome P4501A2, xanthine oxidase, and N-acetyltransferase activity in humans. Ther Drug Monit 18(3):254–262CrossRefPubMedGoogle Scholar
  26. 26.
    Hamdy SI, Hiratsuka M, Narahara K, et al (2003) Genotyping of four genetic polymorphisms in the CYP1A2 gene in the Egyptian population. Br J Clin Pharmacol 55(3):321–324CrossRefPubMedGoogle Scholar
  27. 27.
    Todesco L, Torok M, Krahenbuhl S, Wenk M (2003) Determination of −3858G−>A and −164C−>A genetic polymorphisms of CYP1A2 in blood and saliva by rapid allelic discrimination: large difference in the prevalence of the −3858G−>A mutation between Caucasians and Asians. Eur J Clin Pharmacol 59(4):343–346CrossRefPubMedGoogle Scholar
  28. 28.
    Han XM, Chen XP, Wu QN, Jiang CH, Zhou HH (2000) G–2964A and C734A genetic polymorphisms of CYP1A2 in Chinese population. Acta Pharmacol Sin 21(11):1031–1034PubMedGoogle Scholar
  29. 29.
    Han XM, Ouyang DS, Chen XP, et al (2002) Inducibility of CYP1A2 by omeprazole in vivo related to the genetic polymorphism of CYP1A2. Br J Clin Pharmacol 54(5):540–543Google Scholar
  30. 30.
    Goodman MT, Tung KH, McDuffie K, Wilkens LR, Donlon TA (2003) Association of caffeine intake and CYP1A2 genotype with ovarian cancer. Nutr Cancer 46(1):23–29CrossRefPubMedGoogle Scholar
  31. 31.
    Schulze TG, Schumacher J, Muller DJ, et al (2001) Lack of association between a functional polymorphism of the cytochrome P450 1A2 (CYP1A2) gene and tardive dyskinesia in schizophrenia. Am J Med Genet 105(6):498–501CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag 2005

Authors and Affiliations

  • Carsten Skarke
    • 1
  • Anja Kirchhof
    • 1
  • Gerd Geisslinger
    • 1
  • Jörn Lötsch
    • 1
  1. 1.pharmazentrum frankfurt/ZAFES, Institute of Clinical PharmacologyJohann Wolfgang Goethe-UniversityFrankfurtGermany

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