Leukaemogenesis, gene interplay, and the role of the haemopoietic environment

Malignant initiation, leukaemic transformation, and disease progression in haematological malignancies involves a series of mutational events in genes involved in normal housekeeping functions of the cell. These acquired genetic changes can lead to either increased proliferation or a decreased rate of apoptosis, thus allowing expansion of the malignant clone. Although leukaemia can arise as a de novo disease, it has become increasingly clear that therapies, including the use of irradiation and/or chemotherapy, can give rise to malignancy. Therapy-associated myelodysplasia (t-MDS) and therapy-associated acute myeloid leukaemia (t-AML) account for 10-20% of new cases of these diseases. Although these secondary malignancies have been recognised as a clinical entity for nearly 30 years, molecular studies are now pinpointing various regions of the genome that are susceptible to DNA damage by these chemotherapeutic/radiotherapeutic strategies. The detection of new malignancies (both solid tumours and haematological tumours) following allogeneic bone marrow transplantation (BMT) is also providing us with some clues to the nature of leukaemogenesis, particularly with the observation that leukaemia can occur in donor cells postallogeneic BMT.

Continuous MLL-ENL expression is necessary to establish a "Hox Code" and maintain immortalization of hematopoietic progenitor cells

The t[(11;19)(p22;q23)] translocation, which gives rise to the MLL-ENL fusion protein, is commonly found in infant acute leukemias of both the myeloid and lymphoid lineage. To investigate the molecular mechanism of immortalization by MLL-ENL we established a Tet-regulatable system of MLL-ENL expression in primary hematopoietic progenitor cells. Immortalized myeloid cell lines were generated, which are dependent on continued MLL-ENL expression for their survival and proliferation. These cells either terminally differentiate or die when MLL-ENL expression is turned off with doxycycline. The expression profile of all 39 murine Hox genes was analyzed in these cells by real-time quantitative PCR. This analysis showed that loss of MLL-ENL was accompanied by a reduction in the expression of multiple Hoxa genes. By comparing these changes with Hox gene expression in cells induced to differentiate with granulocyte colony-stimulating factor, we show for the first time that reduced Hox gene expression is specific to loss of MLL-ENL and is not a consequence of differentiation. Our data also suggest that the Hox cofactor Meis-2 can substitute for Meis-1 function. Thus, MLL-ENL is required to initiate and maintain immortalization of myeloid progenitors and may contribute to leukemogenesis by aberrantly sustaining the expression of a "Hox code" consisting of Hoxa4 to Hoxa11.

A practical miniature long-term bone marrow culture system for investigating early myelodysplasia

A method was devised to grow haemopoietic cells in long-term bone marrow culture (LTBMC) which requires only 1 x 10(6) cells/culture. Such miniature cultures were used to study growth patterns of marrow from patients with myelodysplastic syndromes (MDS). Consistent differences in LTBMC cellularity and cellular composition were noted between MDS and normal marrow. These differences were accentuated by rGM-CSF. The criteria which distinguished between and MDS marrows were: cell count at weeks 1 and 4, % neutrophils and % blasts. In 10 patients with unexplained macrocytosis or pancytopenia miniature LTBMC results clearly segregated into either 'normal' or 'MDS' growth patterns. Miniature LTBMC with rGM-CSF may therefore be a useful diagnostic test for early MDS.