K-ras codon 12 and 61 point mutations in bromodeoxyuridine- and N-nitrosomethylurea-induced rat renal mesenchymal tumors

Cancer Lett. 1996 Dec 3;109(1-2):1-7. doi: 10.1016/s0304-3835(96)04350-9.


The mutagenic thymidine analog bromodeoxyuridine (BrdUrd) may incorrectly incorporate opposite deoxyguanine in DNA, then pair with deoxyadenosine during subsequent replication. It appears to preferentially target the 3'-G of 5'-NGGN-3' sequences in mammalian cells in culture to induce G-->A transitions. Ras genes should therefore be vulnerable to activation by mutation at glycine codons 12 (GGT) and/or 13 (GGC) by misincorporation of BrdUrd. There is limited evidence that BrdUrd may be carcinogenic or co-carcinogenic in rats: three renal mesenchymal tumors, a tumor known to be associated with activating mutations in the c-K-ras-2 oncogene, were reported in 87 rats treated with BrdUrd alone, while N-nitrosomethylurea (NMU) alone or NMU + BrdUrd resulted in incidences of 12/52 and 26/76, respectively, against a zero incidence in untreated rats. We analyzed renal mesenchymal tumors from rats treated with BrdUrd for mutations in K-ras exons 1 and 2 and compared the prevalence and spectrum of mutations with those found in comparable tumors induced with NMU. DNAs from 22 paraffin-embedded renal mesenchymal tumors from rats treated 12-15 months earlier with BrdUrd (three specimens) or NMU (11 specimens) or both agents sequentially (eight specimens) were amplified by PCR. The base sequence of codons 12-13 and 59-63 of K-ras was determined by the dideoxynucleotide method. Sequencing results were confirmed by allele-specific oligonucleotide hybridization. Two of three tumors that appeared in rats given BrdUrd alone contained both a codon 12 GGT-->GAT transition and a codon 61 CAA-->CTA transversion. One tumor induced by NMU alone also showed a codon 12 GGT-->GAT mutation, while only wild type sequence could be demonstrated in the codon 12-13 region in the remaining ten such tumors. Three NMU-induced tumors also showed codon 61 CAA-->CTA mutations, while the remaining tumors had wild type sequence. While the GGT-->GAT transitions identified in tumors from BrdUrd-treated rats are consistent with BrdUrd mutagenesis by misincorporation, the co-occurrence of CAA-->CTA transversions, the overall low prevalence of mutations, and the lack of any difference in mutation spectrum between tumors induced by NMU and those that occurred in BrdUrd-treated rats suggests that in both groups the mutations that did occur did not result from a direct effect of either agent.

MeSH terms

  • Adenine
  • Animals
  • Bromodeoxyuridine*
  • Carcinogens*
  • Codon / genetics*
  • Female
  • Genes, ras / genetics*
  • Guanine
  • Kidney Neoplasms / chemically induced
  • Kidney Neoplasms / genetics*
  • Male
  • Mesenchymoma / chemically induced
  • Mesenchymoma / genetics*
  • Methylnitrosourea
  • Neoplasms, Experimental / chemically induced
  • Neoplasms, Experimental / genetics
  • Point Mutation / genetics*
  • Polymerase Chain Reaction
  • Rats


  • Carcinogens
  • Codon
  • Guanine
  • Methylnitrosourea
  • Bromodeoxyuridine
  • Adenine