Preliminary studies show transforming activity is associated with DNA from 6 cases of the advanced B cell malignancy, B cell chronic lymphocytic leukemia (B-CLL). The technique of DNA-mediated-gene-transfer was used to introduce high molecular weight DNA from B-CLL cells into mouse NIH 3T3 cells were inoculated into nude mice. Tumor formation is correlated with the transfer of DNA sequences which contain transforming activity. In this proposal we will clone the transforming gene(s) associated with advanced stages of B-CLL disease. The nude mouse tumorigenicity assay will monitor the cloning of the biologically active DNA fragment consisting of single copy human DNA will be used in Southern blot analysis to determine the pattern of gene arrangement in normal and malignant human lymphoid tissue. Northern blot analysis will determine expression of the gene(s) in mRNAs obtained from normal and malignant lymphoid tissues. To be cloned and its DNA sequence compared with the activated oncogene. The transforming DNA sequences will be analyzed by Southern blot using the active subclone to determine whether the genes are the same or different from one another. If the genes are the same, then fine restriction endonuclease mapping and DNA sequencing of the genes will determine the chromosomal change(s) involved in activation of the oncogene. The frequency of oncogene activation will be analyzed in B-CLL cases representative of different disease stages. This study will identify the transforming activity associated with B-CLL disease. Understanding the genetic mechanism of oncogene activation in B- CLL disease may have potential for improved patient diagnosis and treatment.
