A comprehensive analysis of loss of heterozygosity caused by hemizygous deletions in renal cell carcinoma using a subtraction library

Abstract

Several new loci were identified by a comprehensive analysis of loss of heterozygosity (LOH) using a subtraction library between matched normal and renal cell carcinoma (RCC) tissues. A total of 187 clones from the library, with a complexity of 1x10(4), were mapped, and 44 clusters of the mapped loci were subjected to LOH analysis using microsatellite markers. A total of 27 loci, which exhibited frequencies of LOH of at least 10% among 44 tumors, mostly clear-cell RCC, included several loci that were reported previously, such as, the von Hippel-Lindau gene, adenomatous polyposis coli, and interferon regulatory factor-1, as well as new loci, at 5q32-q34, 6q21-q22, 8p12, and others. These loci exhibited LOH among 11.8-93.8% of tumors, and most, if not all, were derived from the sites of hemizygous deletions. The minimum regions of LOH of chromosomes 5, 6, and 8 were 9.0, 10.3, and 0.775 Mb, respectively. The average distance between the cloned fragments on the chromosomes was 2.2 Mb in 187 clones, indicating that the minimum LOH size expected from this subtraction analysis was roughly 50 kb. Therefore, the strategy described here provides comprehensive analysis of LOH sites, which were mostly caused by hemizygous deletions.

A Novel Ankyrin Repeat-containing Gene (Kank) Located at 9p24 Is a Growth Suppressor of Renal Cell Carcinoma

“…We previously reported 27 potential tumor-related loci using a library obtained by subtraction of tumor tissue DNA from normal tissue DNA of the same patient (14). By extensive analysis of LOH in RCC tissues using microsatellite markers, one of the loci on chromosome 9 was found to show a relatively high incidence of LOH (25%, 5 of 20 cases) (14). The minimum overlapping region of LOH at this 9p24 locus was further narrowed down to a region 165 kb in size (Fig.…”

“…LOH Analysis-LOH analysis using GeneScan Version 3.1 (Applied Biosystems, Inc.) was performed as described previously (14). For LOH analysis of patient R6, genomic DNA was extracted from primary culture cells derived from normal and tumor tissues.…”

“…As a comprehensive method for searching and cloning the sites of hemizygous methylation is not available, mapping hemizygous deletions for the tumor-related loci is still an efficient strategy. We previously performed a comprehensive search for LOH sites in 44 (mostly clear cell-type) RCC cases, and a total of 27 candidate loci showing frequencies of LOH of at least 10% were found (14). Among them, LOH on chromosome 9 showed as a relatively frequent event in RCC (25%).…”

Loss of the actin regulator HSPC300 results in clear cell renal cell carcinoma protection in Von Hippel-Lindau patients

“…In renal cancer cells, it has been reported that Rac is a key regulator of cytoskeleton organization during hypoxia, and modulation of the Rac-actin pathway is essential for renal cell carcinoma development [Engers et al, 2001]. Indeed, it has been hypothesized that a candidate tumor suppressor gene in ccRCC might function by controlling the formation of the cytoskeleton through polymerization of actin [Hatano et al, 2001;Sarkar et al, 2002].…”

Genetics of Renal Tumors

“…A genome-wide association study based on more than 5,000 RCC cases revealed two loci, 2p21 and 11q13.3, to be associated with RCC susceptibility (Purdue et al, 2011). Although the authors claimed these sites to be previously unidentified, both of the loci were actually identified in 2001 (Hatano et al, 2001; Table 1). While the association is statistically significant, the frequencies among RCC cases are not very high (less than 20%), and therefore, it is doubtful that these sites alone can be used for diagnosis.…”