Deletion mapping at 12p12-13 in metastatic prostate cancer

The identification of homozygous deletions in malignant tissue is a powerfu l tool for the localization of tumor suppressor genes. Representational dif ference analysis (RDA) uses selective hybridization and the polymerase chai n reaction (PCR) to isolate regions of chromosomal loss and has facilitated the identification of tumor suppressor genes, such as BRCA2 and PTEN, We h ave recently identified a 1-5-cM homozygous deletion on 12p12-13 in a prost ate cancer xenograft and found that 47% of patients who died of prostate ca rcinoma demonstrate focal loss of heterozygosity (LOH) in this region in me tastatic deposits. We have now characterized the region of interest by asse mbling a yeast artificial chromosome (YAC) contig spanning the homozygous d eletion and identifying which Known genes and expressed sequence tags (EST) lie within the homozygous deletion. A rib metastasis was harvested at auto psy and placed subcutaneously in a male SCID mouse. Genomic DNA from this x enograft and from the patient's normal renal tissue was extracted. Multiple x PCR, with the xenograft and normal DNA used as template, was performed us ing primers for loci on the Whitehead contig 12.1 believed to be near our r egion of interest. We found that our deletion lay in a 1-2-Mb interval betw een WI-664 and D12S358. We then used the same primers to construct a YAC co ntig across the homozygous deletion. PCR amplification of YAC DNA, using pr imers for the genomic sequences of known genes and ESTs reported to lie on 12p12-13, was used to identify candidate genes that lay within the deletion . Duplex PCR, with control primers known not to be deleted in the xenograft , was used to confirm that both the CDKNIB and ETV6 genes were homozygously deleted in the xenograft. Mutations in either or both of these genes may p lay an important role in metastatic prostate carcinoma. (C) 1999 Wiley-Liss , Inc.