I. Goal. Identification of the genetic defects in B-cell chronic lymphocytic leukemia (B-CLL). II. Approach. A. Linkage study. Using the mouse model for CLL (the NZB inbred mouse) we are using the recently developed simple sequence length polymorphism (SSLP) technology to define the number and location of genes linked to the NZB proliferative disorder. We are currently typing some 600 F1 backcross mice for this study. Using the human SSLP's we propose to map molecular defects in human CLL families to correlate with the locations implicated by the mouse linkage study. We are actively seeking to extend our current work on CLL family pedigrees to allow more precise mapping. B. Loss of heterozygosity (LOH). We are gathering normal and tumor DNA samples of familial and common CLL to use to map deletions through an analysis of SSLP's. The rationale for this approach is that most, if not all, cancer causing genes that have contained molecular defects have been paired with deletions. We propose to extend this study to include the other B-cell malignancies based upon the hypothesis that B-cell malignancies will be expected to contain common pathways. C. We are in the process of identifying CLL genes that are up or down- regulated by the differential display method. In this method message levels are compared in normal and tumor by comparing band intensity of reverse PCR products after electrophoresis. After sequencing and cloning, the PCR fragment acts as a probe for the isolation of the parent gene. These genes can then be correlated with areas of linkage and deletion. There would be a high index of suspicion that an abnormally expressed gene located in an area of deletion or linkage is on the site of a primary genetic defect and thus should be treated as a candidate gene. D. Candidate genes. Genes that have been found to contain molecular defects in one or more malignancies become candidate genes for CLL. We propose to assay these genes for the presence of molecular defects in common CLL and familial CLL. With regards to the differential display, autologous leukemic and normal EBV transformed cell lines will be compared.
