Trisomies of chromosomes 7 and 17 are specific genetic alterations in papillary RCTs irrespective of their size, grade and cellular differentiation.

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Key words: renal cell cancer; BRAF; mutation; allelic changesSporadic and hereditary papillary renal cell tumors are characterized by a combined trisomy of the chromosome 7 and 17q regions. 1,2 Selective duplication of chromosome 7q in a hereditary papillary RCC called attention to this chromosomal region. 3 Subsequently, Schmidt et al. 4 detected activating mutations of the tyrosine kinase domain of the MET gene at chromosome 7q31 in the germline of patients with hereditary papillary RCCs. It was also shown that duplication and overexpression of the mutant MET allele is necessary before a tumor develops. 2 Despite the frequent duplication of this chromosomal region only 5-13% of sporadic papillary RCCs carry a MET mutation. 4,5 This finding suggest that another gene at chromosome 7q might be involved in the development of sporadic papillary RCCs.Recently, mutations of the BRAF gene (mapped to chromosome 7q34) have been detected in 66 -80% of nevi and malignant melanomas and also at lower frequency in colorectal and other types of cancers. 6 -11 A single base substitution mutation was detected in exon 15 in the overwhelming majority of tumors and also in exon 11 in some cases. Although Davies et al. 7 have analyzed 10 renal cell tumor cell lines for BRAF mutations, they included only one papillary RCC cell line. To establish the possible involvement of BRAF in the development of sporadic papillary RCCs, we have sequenced exons 11 and 15 of the BRAF in 50 primary RCCs including 20 papillary ones. Although, allelotyping indicated a frequent duplication of the chromosome 7q34 region, our study failed to detect mutation of the BRAF. MATERIAL AND METHODS Tumor samples and DNA extractionFresh tumor and normal kidney parenchymal tissues were obtained by nephrectomy at the Departments of Urology, Hannover Medical School and the Heidelberg University, Germany between 1986 -88 and 1993-97, respectively. A piece of tumor was immediately snap-frozen in liquid nitrogen and stored at Ϫ80°C whereas the remaining tissue was fixed in 4% buffered formaldehyde for histological report. The diagnosis was established according to the Heidelberg Classification. 12 A frozen tumor sample was placed in a plastic Petri dish, covered with 2 ml TE9 buffer, and allowed to thaw. The tumor cells were then carefully scraped or pushed out under an inverted microscope by a pathologist (GK) experienced in this technique to separate them from stromal tissue. The stromal tissue rests were then discarded. The tumor cells were suspended in 3-5 ml TE9 buffer with 1% SDS and 0.2 mg/ml proteinase K and were incubated for 5 hr at 55°C. DNA was extracted by phenol-chloroform and dissolved in TE buffer after ethanol precipitation. Normal control DNA was extracted from corresponding kidney parenchymal specimens by the same method. The concentration of DNA samples was adjusted to 50 ng/ l. Microsatellite analysisMicrosatellites flanking the BRAF were selected from the sequence segment NT_ 007914.10 (http://www.ncbi.nlm.nih.gov/ mapview). The dinucleotide rep...

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