This Article Full Text Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Dieterle, C. P. Articles by Berger, M. R. Search for Related Content PubMed PubMed Citation Articles by Dieterle, C. P. Articles by Berger, M. R.

Detection of Isolated Tumor Cells by Polymerase Chain Reaction-Restriction Fragment Length Polymorphism for K-ras Mutations in Tissue Samples of 199 Colorectal Cancer Patients

¹ Unit of Toxicology and Chemotherapy, German Cancer Research Center, Heidelberg, and ² Department of Surgery, Municipal Hospital Nürnberg Nord, Nürnberg, Germany

The aim of this study was to identify K-ras mutations as marker for isolated tumor cells in liver, lymph node, and bone marrow specimens of colorectal cancer patients. To detect these, a PCR-RFLP assay was used with a sensitivity exceeding that of routine histopathology by at least 1 order of magnitude. In addition, the ratio of mutated versus wild-type alleles was determined by an internal standard. Of 199 patients, 74 (37.5%) were found to bear a K-ras-positive tumor. Of these, 60 (81%) were mutated in codon 12 and 14 (19%) in codon 13 (P < 0.001). In addition, 14 organs were found K-ras positive, 13 of which were from 12 patients with a K-ras-positive tumor (16%) and 1 from a patient with a K-ras-negative tumor (0.8%). Eight patients exhibited liver involvement and 6 showed lymph node involvement. Remarkably, no bone marrow specimen was found K-ras positive (P < 0.017 versus liver involvement). Sequence analysis of tumor DNA revealed that GGT (Gly) was replaced by GAT (Asp; 35%), GTT (Val; 32%), AGT (Ser; 13%), GCT (Ala; 10%), TGT (Cys; 8%), and CGT (Arg; 2%) for codon 12, and by GAC (Asp) as the only type of mutation for codon 13. In colorectal carcinomas the ratio of K-ras mutated versus wild-type alleles ranged over 4 orders of magnitude (10⁰-10^-4, median: 10^-2) and was correlated with both, residual tumor load (R1/2; P = 0.028) and distant metastasis (M1; P = 0.057). These results show that detection of K-ras mutated alleles by PCR-RFLP in patients with colorectal carcinoma may aid in the identification of isolated tumor cells. High ratios of K-ras alleles were correlated with certain negative prognostic parameters (R,M). In accord with its function as a primary filter for colorectal carcinoma cells, the liver was more often contaminated with K-ras-positive cells than bone marrow.

This article has been cited by other articles:

				B. Gilje, R. Heikkila, S. Oltedal, K. Tjensvoll, and O. Nordgard High-Fidelity DNA Polymerase Enhances the Sensitivity of a Peptide Nucleic Acid Clamp PCR Assay for K-ras Mutations J. Mol. Diagn., July 1, 2008; 10(4): 325 - 331. [Abstract] [Full Text] [PDF]

				J.-D. Luo, E.-C. Chan, C.-L. Shih, T.-L. Chen, Y. Liang, T.-L. Hwang, and C.-C. Chiou Detection of rare mutant K-ras DNA in a single-tube reaction using peptide nucleic acid as both PCR clamp and sensor probe Nucleic Acids Res., January 23, 2006; 34(2): e12 - e12. [Abstract] [Full Text] [PDF]

				N. Smakman, O. Kranenburg, J. M. Vogten, A. L.A. Bloemendaal, P. van Diest, and I. H.M. Borel Rinkes Cyclooxygenase-2 Is a Target of KRASD12, Which Facilitates the Outgrowth of Murine C26 Colorectal Liver Metastases Clin. Cancer Res., January 1, 2005; 11(1): 41 - 48. [Abstract] [Full Text] [PDF]