Common Regions of Deletion on Chromosomes 5q, 6q, and lOq in Renal Cell Carcinoma

Relatively frequent losses of heterozygosity on chromosomes 5q, 6q, and lOq, in addition to loss of heterozygosity on the short arm of chromosome 3, have been observed in renal cell carcinomas. As the first step toward isolation of tumor suppressor genes on these three chromo somal arms, we used six restriction fragment length polymorphism markers for 5q, nine for 6q, and eight for lOq to identify regions commonlydeleted in a panel of 64 renal cell carcinomas. Allelic losses were common at chromosome 5q21, the region where the MCC (mutated in colorectal cancer) gene was recently identified; at chromosome 6q27; and at chro mosome 10q21-23. Furthermore, an association was observed between accumulation of allelic losses on these three chromosomal arms and progression of tumors. Loss of heterozygosity on chromosome S showed a correlation with the histopathological grade of a given tumor and the incidence of distant metastasis. INTRODUCTION Numerous studies using RFLP2 markers to investigate the molecular biology of carcinogenesis have suggested that an accumulation of genetic alterations at specific chromosomal loci contributes to tumor development and/or progression. Several genes, including oncogenes and tumor suppressor genes, appear to be involved in tumorigenesis of colon and breast cancers (1-3). In RCC, several lines of evidence point to the existence of a tumor suppressor gene(s) on the short arm of chromosome 3: somatic chromosomal aberrations involving this chromosome (4, 5); t(3;8) balanced translocations in germline cells of some patients with hereditary renal cell carcinoma (6); and frequent LOH on chromosome 3p (7-10). Our earlier study suggested that, in addition to the gene on chromosome 3p, tumor suppressor genes associated with RCC may exist on chromosomes 5q, 6q, lOq, llq, and 17p (11). However, the frequency of LOH on these chromosomal arms was not as high as that on chromosome 3p. To examine further whether these chromosomal arms contain tumor suppressor genes, and if they do, to map the loci precisely, we have analyzed these candidate loci in detail with a large number of RFLP markers. Addition ally, we have expanded the number of tumors investigated to 64 and correlated the accumulation of events causing LOH on various chromosomal arms with clinical and histopathological parameters. MATERIALS AND METHODS Materials. Normal and cancerous tissues from 64 patients with primary sporadic RCC were obtained during radical nephrectomy at Received 6/19/91 ; accepted 8/13/91. The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact. ' To whom requests for reprints should be addressed, at Department of Biochemistry, Cancer Institute, 1-37-1, Kami-ikebukuro. Toshima-ku, Tokyo 170, Japan. 2The abbreviations used are: RFLP, restriction fragment length polymor phism; RCC, renal cell carcinoma: LOH, loss of heterozygosity. the Kobe National Hospital, the School of Medicine, Kobe University, or the Department of Urology, Kyoto University. The tissues were frozen immediately after nephrectomy and stored at -70°C until ex traction of DNA. DNA Extraction and Probes. The extraction of DNA from tissues was carried out according to methods described previously (3). All probes used in this study are listed in Table 1. The probes in the cCI6 series were isolated from a human/mouse hybrid cell line which con tained only chromosome 6 as human-derived material (12) and were mapped by fluorescent in situ hybridization.' Southern Blotting and Hybridization. All DNA samples were trans ferred to nylon membrane (Pall Biodyne) with 0.1 N NaOH/0.1 M NaCl. Neutralization, fixation, and hybridization were done according to methods described previously (3).