Gerald M. Segal

Effects of a selective inhibitor of the Abl tyrosine kinase on the growth of Bcr–Abl positive cells

The bcr–abl oncogene, present in 95% of patients with chronic myelogenous leukemia (CML), has been implicated as the cause of this disease. A compound, designed to inhibit the Abl protein tyrosine kinase, was evaluated for its effects on cells containing the Bcr–Abl fusion protein. Cellular proliferation and tumor formation by Bcr–Abl–expressing cells were specifically inhibited by this compound. In colony–forming assays of peripheral blood or bone marrow from patients with CML, there was a 92–98% decrease in the number of bcr–abl colonies formed but no inhibition of normal colony formation. This compound may be useful in the treatment of bcr–abl–positive leukemias.

Repression of Fanconi anemia gene (FACC) expression inhibits growth of hematopoietic progenitor cells.

Bone marrow failure is a consistent feature of Fanconi anemia (FA) but it is not known whether the bone marrow failure is a direct and specific result of the inherited mutation or a consequence of accumulated stem cell losses resulting from nonspecific DNA damage. We tested the hypothesis that the protein encoded by the FA group C complementing gene (FACC) plays a regulatory role in hematopoiesis. We exposed normal human lymphocytes, bone marrow cells, endothelial cells, and fibroblasts to an antisense oligodeoxynucleotide (ODN) complementary to bases -4 to +14 of FACC mRNA. The mitomycin C assay demonstrated that the antisense ODN, but not missense or sense ODNs, repressed FACC gene expression in lymphocytes. Treatment with the antisense ODN substantially reduced, in a sequence-specific fashion, cytoplasmic levels of FACC mRNA in bone marrow cells and lymphocytes. Escalating doses of antisense ODN increasingly inhibited clonal growth of erythroid and granulocyte-macrophage progenitor cells but did not inhibit growth of fibroblasts or endothelial cells. The antisense ODN did not inhibit growth factor gene expression by low density bone marrow cells or marrow-derived fibroblasts. We conclude that, while the FACC gene product plays a role in defining cellular tolerance to cross-linking agents, it also functions to regulate growth, differentiation, and/or survival of normal hematopoietic progenitor cells.

Studies of Murine Megakaryocyte Colony Size and Ploidization

Megakaryocytes are the product of a series of proliferative and differentiative steps involving a complex cellular hierarchy. Pluripotent stem cells undergo a progressive narrowing in their differentiative potential eventually giving rise to unipotent progenitor cells committed to the megakaryocytic lineage. These cells, which are operationally termed colony-forming units — megakaryocyte (CFU-Meg), undergo varying numbers of mitotic divisions, giving rise to a class of cells often referred to as promegakaryoblasts. These diploid cells cease cell division but, instead, undergo a series of synchronous nuclear endoreduplications or endomitoses, giving rise to the unique polyploid cells megakaryocytes. Megakaryocyte colony formation represents an in vitro model of these processes in which colony size reflects the mitotic activity of megakaryocytic progenitor cells while the ploidy of colony cells is determined by the endomitotic activity of promegakaryoblasts and their descendants. Therefore, it might be expected that the analysis of megakaryocytic colony size and the ploidy of colony cells could provide insight into the cellular responses which underlie megakaryocytopoietic regulation in vivo.