Characterization of Leukemia-Inducing Genes Using the Retroviral cDNA Library

Acute myeloid leukemia is a disease caused by oncogenic change(s) of hematopoietic stem cells or progenitor cells. The purpose of this research is to characterize the gene involved in the leukemia induction processes by screening the retroviral cDNA libraries of cellular oncogenes, homeobox genes. The cDNA libraries were constructed by cloning the individual genes to a MSCV retroviral vector backbone. For convenience of detecting the transduced cells and their protein products, the MSCV retroviral vector was modified to include HA tag and GFP marker and ninety cellular oncogenes and 30 homeobox genes were individually cloned and the structural integrity was verified. To screen for the leukemia-inducing genes, 5-FU treated mouse bone marrow cells were transduced with retroviral mixtures of oncogenes, and the cells were transplanted into ten lethally irradiated mice. All ten mice developed acute leukemia between the eight and ten weeks post-transplantation. The oncogenes that were responsible for the leukemia induction were characterized by genomic DNA PCR of the leukemic cells of each mouse. Surprisingly, all ten mice had c-myc genes in their leukemic cells. However, except for three mice, all the mice have additional oncogenes within the leukemic cells. The list of the additional genes include; RAB3D, RAB7B, PDGF-beta, CRK, and PIM-2 and Ras. Theses results show that the c-myc is the major leukemia-inducing oncogenes in our system. In addition, since the initial transduction rate of 5-FU-treated bone marrow just prior to in vivo transplantation was 10.56%, it is highly unlikely that all these additional oncogenes were present in the transplantable leukemic cells just by chance. Therefore, the retroviral cDNA library-mediated leukemia induction system we developed may be an useful system in systematically screening the cooperating oncogenes in leukemia induction. We are currently verifying the cooperating potential of the genes co-transduced with c-myc in inducing leukemia in the same animal model.