High-throughput genotyping by DHPLC of the dihydropyrimidine dehydrogenase gene implicated in (fluoro)pyrimidine catabolism

  • Authors:
    • Eva Gross
    • Katharina Seck
    • Steffi Neubauer
    • Jutta Mayr
    • Heide Hellebrand
    • Adisorn Ratanaphan
    • Verena Lutz
    • Hubertus Stockinger
    • Marion Kiechle
  • View Affiliations

  • Published online on: February 1, 2003     https://doi.org/10.3892/ijo.22.2.325
  • Pages: 325-332
Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )
Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )


Abstract

Dihydropyrimidine dehydrogenase (DPD) is the first and rate-limiting enzyme in the degradation of pyrimidines and pyrimidine base analogs including the anticancer drugs 5-fluorouracil (5-FU) and Xeloda®. A decreased DPD enzyme activity has been described in cancer patients experiencing severe and life-threatening toxicity after 5-FU treatment and distinct sequence variants in the DPD gene (DPYD) have been associated with impaired enzyme function. The most prominent mutation in the DPD deficient patient group, a mutation in the splicing donor consensus sequence of intron 14, IVS14+1g>a, resulting in a truncated protein, has been observed in the Caucasian population at frequencies as high as 0.91%-0.94%. This underlines the need for a test system for DPYD mutations in patients undergoing chemotherapy with 5-FU or with Xeloda. To set up a fast and sensitive method to identify variant DPYD alleles, we analyzed 50 healthy individuals by denaturing high performance liquid chromatography (DHPLC). A primer set spanning the whole coding region and the exon-intron boundaries of DPYD was used. In addition, a cDNA-based assay was developed to rapidly identify the 165 base pair deletion in the corresponding RNA of IVS14+1g>a mutation carriers. The optimal mutation detection was elaborated for each of the PCR fragments. DHPLC analysis detected 5 different genetic alterations occurring in the coding region of the gene, as well as 10 intronic sequence variants, respectively. In conclusion, high-throughput screening for DPYD variants by DHPLC may be a reliable tool in the investigation of the molecular basis of DPD deficiency. Furthermore, it will help to identify patients at risk for toxic side effects upon chemotherapy using 5-FU and will facilitate individual treatment of patients.

Related Articles

Journal Cover

February 2003
Volume 22 Issue 2

Print ISSN: 1019-6439
Online ISSN:1791-2423

Sign up for eToc alerts

Recommend to Library

Copy and paste a formatted citation
x
Spandidos Publications style
Gross E, Seck K, Neubauer S, Mayr J, Hellebrand H, Ratanaphan A, Lutz V, Stockinger H and Kiechle M: High-throughput genotyping by DHPLC of the dihydropyrimidine dehydrogenase gene implicated in (fluoro)pyrimidine catabolism. Int J Oncol 22: 325-332, 2003.
APA
Gross, E., Seck, K., Neubauer, S., Mayr, J., Hellebrand, H., Ratanaphan, A. ... Kiechle, M. (2003). High-throughput genotyping by DHPLC of the dihydropyrimidine dehydrogenase gene implicated in (fluoro)pyrimidine catabolism. International Journal of Oncology, 22, 325-332. https://doi.org/10.3892/ijo.22.2.325
MLA
Gross, E., Seck, K., Neubauer, S., Mayr, J., Hellebrand, H., Ratanaphan, A., Lutz, V., Stockinger, H., Kiechle, M."High-throughput genotyping by DHPLC of the dihydropyrimidine dehydrogenase gene implicated in (fluoro)pyrimidine catabolism". International Journal of Oncology 22.2 (2003): 325-332.
Chicago
Gross, E., Seck, K., Neubauer, S., Mayr, J., Hellebrand, H., Ratanaphan, A., Lutz, V., Stockinger, H., Kiechle, M."High-throughput genotyping by DHPLC of the dihydropyrimidine dehydrogenase gene implicated in (fluoro)pyrimidine catabolism". International Journal of Oncology 22, no. 2 (2003): 325-332. https://doi.org/10.3892/ijo.22.2.325