Role of K-ras Mutations and Activation of the EGFR-Signalling Pathway in Colorectal Carcinoma
M Dantes, CW Hann von Weyhern, R Rosenberg, H Hoefler, F Fend. University of Tuebingen, Tuebingen, Germany; University Hospital Technical University, Munich, Germany; Technical University Munich, Munich, Germany
Background: Mutations in the Kirsten-ras (K-ras) gene have been known as early events in carcinogenesis of colorectal cancer since the hypothesis of linear cancerogenesis by Vogelstein. Recently a monoclonal antibody Cetuximab against the Epidermal growth factor receptor (EGFR) was introduced in treatment of advanced colorectal cancer. In this context molecular analysis of the K-ras became important as only wild-type carriers have shown to benefit from targeted therapy. Still the rate of non-responders to EGFR-antibodies is approximately 45% among wild-type carriers.
Design: 274 primarily resected colorectal cancers were arranged to tissue microarrays (TMA). FFPE material (1993-2003) were retrieved from the archive of the Institute of Pathology Technical University Munich. With the small flaps of tissue, Light-cycler-analysis of K-ras-mutations in exon 2, codons 12 and 13, were accomplished. Results were compared with results of regular slides. Additionally the TMA-material of a subgroup of 53 patients was tested for gene copy number of EGFR by silver-enhanced in-situ hybridization (SISH) and compared with the immunohistochemical staining.
Results: For mutation analysis, 207 cases from TMA-slides were evaluable. 102 (49%) presented mutation in K-ras exon 2. Most common ones were the mutations G13D (27=26.5%), G12D (26=25.5%) and G12V (17=16.7%). As control of effectiveness and accuracy of results from the TMA material, selected cases were compatred with whole original slides demonstrating identical findings of mutations. Immunohistochemical evaluation of EGFR shows even distribution for staining intensities: 80 tumors (33.1%) recieved score 1+, 69 cases (28.5%) score 2+, 31 (12.8%) score 3+. 62 cases (25.6%) were negative. Ratio of EGFR gene copy number/CEP was below 1.5 in 53 selected cases.
Conclusions: Small amounts of tissue from a TMA comparable to small biopsy specimens in routine diagnostics provide exact information about K-ras mutations in comparison to whole tumor preparations. This demonstrates that the K-ras analysis is possible and reliable even in very small amounts of tissue. Frequency and distribution of mutations is similar to previously published data. No EGFR gene amplification could be observed. EGFR-expression by immunohistochemistry is very heterogenious, even in between needle-biopsies.
Tuesday, March 23, 2010 9:30 AM
Poster Session III # 113, Tuesday Morning