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The focus of this study was to determine if alterations are present within the tyrosine kinase and ligand binding domains of the epidermal growth factor receptor (EGFR) in urothelial carcinoma. In studies involving non small cell lung cancer (NSCLC), both Paez et al. and Lynch et al. found a correlation between patient response to Iressa and presence of mutations within the tyrosine kinase domain of the EGFR. These two reports focused on NSCLC patients, where 13 of the 14 patients had response to Iressa and a variety of mutations in the ATP binding pocket, comprised of exons 18–21. Based on these findings, exons 18–21 of the EGFR were sequenced in urothelial carcinomas. Samples included 11 transitional cell carcinoma (TCC) cell lines, four of which are known to have a response to Iressa, and 75 bladder cancer patient tumors. After DNA isolation and PCR amplification, the samples were subjected to automated sequencing. Our analysis failed to detect mutations within the tyrosine kinase domain of EGFR in the 11 cell lines and 75 patients tested. Therefore, a mutation in the tyrosine kinase domain cannot be attributed to the Iressa response seen in the TCC cell lines. In addition to these mutations, a rearrangement in wildtype EGFR is known to exist, consisting of a deletion of exons 2–7, which results in what is designated as EGFRvIII. This deletion causes a partial loss of the ligand-binding domain and is likely to contribute to altered EGFR targeting drug responses. In order to determine the status of the EGFRvIII in urothelial carcinoma, a TCC Tissue Microarray was utilized. Preliminary analysis revealed that approximately 50% of the patient tumors expressed the EGFRvIII. To rule out potential inconsistencies in the EGFRvIII antibody, expression of the truncated variant was verified using real-time PCR and western blot analysis. Real-time PCR did not reveal truncated EGFR expression in the 12 tumor specimens. Using an antibody targeting the cytoplasmic domain of EGFR, western blot analysis of the 9 TCC tumors also supported the Real-time PCR as no expression of the variant was seen. Taken together, we concluded that alterations within the tyrosine kinase domain and ligand binding domain of EGFR are rare events in urothelial carcinoma. The present study has clinical implications in designing tyrosine kinase inhibitors for the urothelial carcinoma therapy.

[Proc Amer Assoc Cancer Res, Volume 47, 2006]