Development and characterization of a monoclonal antibody with cross-reactivity towards uracil and thymine, and its potential use in screening patients treated with 5-fluorouracil for possible risks.

BACKGROUND Dihydropyrimidine dehydrogenase (DPD) is the initial and rate-limiting enzyme in catabolism of pyrimidines including 5-fluorouracil. There have been efforts to isolate a monoclonal antibody that will bind selectively to pyrimidine and can be used to measure the concentration of pyrimidine in blood and/or in urine that may reflect the activity of dihydropyrimidine dehydrogenase. However, the monoclonal antibodies selective to pyrimidine have not been available. METHODS Using 1-carboxymethyl-uracil as a hapten, in which steric conformation of uracil is thought to be well maintained, extensive screening was done to isolate a monoclonal antibody specific to uracil. RESULTS We established the first monoclonal antibody that reacted with uracil and thymine but not with pseudouridine, dihydrouracil, dihydrothymine, cytosine, uridine, or N-carbamyl-beta-alanine at the concentration of 100 microg/ml. CONCLUSIONS The monoclonal antibody can be used to develop a simple screening assay for patients with dihydropyrimidine dehydrogenase deficiency. This may increase the safety of 5-fluorouracil treatment.

[1]  Y. Wada,et al.  Urinary Screening for Pyrimidine Metabolism Disorders , 1998 .

[2]  Y. Wada,et al.  Automated screening system for purine and pyrimidine metabolism disorders using high-performance liquid chromatography. , 1995, Journal of chromatography. B, Biomedical applications.

[3]  K. Higashino,et al.  Monoclonal antibody to pseudouridine used to develop a radioimmunoassay. , 1990, Journal of immunoassay.

[4]  T. Tsuruo,et al.  Characterization of dihydropyrimidine dehydrogenase on immunohistochemistry in colon carcinoma, and correlation between immunohistochemical score and protein level or messenger RNA expression. , 2000, Annals of oncology : official journal of the European Society for Medical Oncology.

[5]  P. Vreken,et al.  Clinical implications of dihydropyrimidine dehydrogenase (DPD) deficiency in patients with severe 5-fluorouracil-associated toxicity: identification of new mutations in the DPD gene. , 2000, Clinical cancer research : an official journal of the American Association for Cancer Research.

[6]  B. B. Mishell,et al.  Selected Methods in Cellular Immunology , 1980 .

[7]  M. Durán,et al.  New defects of pyrimidine metabolism. , 1984, Advances in Experimental Medicine and Biology.

[8]  R. Danesi,et al.  Relationship between 5-fluorouracil disposition, toxicity and dihydropyrimidine dehydrogenase activity in cancer patients. , 2001, Annals of oncology : official journal of the European Society for Medical Oncology.

[9]  G. Milano,et al.  Potential importance of dihydropyrimidine dehydrogenase (DPD) in cancer chemotherapy. , 1994, Pharmacogenetics.

[10]  H. McLeod,et al.  Molecular basis of the human dihydropyrimidine dehydrogenase deficiency and 5-fluorouracil toxicity. , 1996, The Journal of clinical investigation.

[11]  J. Sludden,et al.  Dihydropyrimidine dehydrogenase pharmacogenetics in patients with colorectal cancer. , 1998, British Journal of Cancer.

[12]  M. Ratain,et al.  Pharmacogenetics and cancer chemotherapy. , 1998, European journal of cancer.

[13]  A. Cruchten,et al.  Comparative study of thymine and uracil metabolism in healthy persons and in a patient with dihydropyrimidine dehydrogenase deficiency. , 1989 .

[14]  C. Wasternack,et al.  Degradation of pyrimidines and pyrimidine analogs--pathways and mutual influences. , 1980, Pharmacology & therapeutics.

[15]  A. V. van Kuilenburg,et al.  Prevalence of a common point mutation in the dihydropyrimidine dehydrogenase (DPD) gene within the 5'-splice donor site of intron 14 in patients with severe 5-fluorouracil (5-FU)- related toxicity compared with controls. , 2001, Clinical cancer research : an official journal of the American Association for Cancer Research.

[16]  F. Baas,et al.  Lethal outcome of a patient with a complete dihydropyrimidine dehydrogenase (DPD) deficiency after administration of 5-fluorouracil: frequency of the common IVS14+1G>A mutation causing DPD deficiency. , 2001, Clinical cancer research : an official journal of the American Association for Cancer Research.

[17]  R. Diasio,et al.  Dihydropyrimidine dehydrogenase activity in human peripheral blood mononuclear cells and liver: population characteristics, newly identified deficient patients, and clinical implication in 5-fluorouracil chemotherapy. , 1993, Cancer research.

[18]  J. Mrochek,et al.  Urinary excretion by cancer patients of the nucleosides N-dimethylguanosine, 1-methylinosine, and pseudouridine. , 1973, Journal of the National Cancer Institute.

[19]  R. Diasio,et al.  Dihydropyrimidine dehydrogenase activity and fluorouracil chemotherapy. , 1994, Journal of clinical oncology : official journal of the American Society of Clinical Oncology.

[20]  Christopher D. Richards,et al.  Human Physiology, The Basis of Medicine , 1999 .

[21]  M. Durán,et al.  Dihydropyrimidine dehydrogenase deficiency leading to thymine-uraciluria. An inborn error of pyrimidine metabolism. , 1984, Clinica chimica acta; international journal of clinical chemistry.

[22]  H. Liebich,et al.  Quantitation of urinary nucleosides by high-performance liquid chromatography. , 1997, Journal of chromatography. A.