afimoxifene and Leukemia

MLL fusion genes are a predominant feature of acute leukemias in infants and in secondary acute myeloid leukemia (AML) associated with prior chemotherapy with topo-II poisons. The former is considered to possibly arise in utero via transplacental chemical exposure. A striking feature of these leukemias is their malignancy and remarkably brief latencies implying the rapid acquisition of any necessary additional mutations. We have suggested that these coupled features might be explained if MLL fusion gene encoded proteins rendered cells more vulnerable to further DNA damage and mutation in the presence of chronic exposure to the agent(s) that induced the MLL fusion itself. We have tested this idea by exploiting a hormone regulated MLL-ENL (MLLT1) activation system and show that MLL-ENL function in normal murine progenitor cells substantially increases the incidence of chromosomal abnormalities in proliferating cells that survive exposure to etoposide VP-16. This phenotype is associated with an altered pattern of cell cycle arrest and/or apoptosis.

Topics: Animals; Cell Line, Transformed; Chromosome Aberrations; DNA Damage; Estrogen Antagonists; Humans; In Situ Hybridization, Fluorescence; Infant; Leukemia; Mice; Myeloid-Lymphoid Leukemia Protein; Oncogene Proteins, Fusion; Reaction Time; Stem Cells; Tamoxifen

M1 mouse myeloid leukemia cells exhibit growth arrest and differentiation to monocytes/macrophages in response to leukemia inhibitory factor (LIF) stimulation. Although recent studies have demonstrated that STAT3 plays a central role in this process, it is unknown whether STAT3 activation alone is sufficient. To address this issue, we have established M1/STAT3ER cells, where STAT3 is selectively activated by 4-hydroxytamoxifen (4HT). 4HT stimulation did not have any effect on growth and morphology of M1/ STAT3ER cells, and did not induce the down-regulation of mRNA of c-myc and c-myb, which is necessary for M1 cell differentiation. On the other hand, mRNA of jun-B, IRF1 and p19 was increased by 4HT. DNA precipitation assay indicated that both stimulation of LIF and 4HT similarly activated STAT3ER. Introduction of a constitutive active MAP kinase kinase (MEK1) into M1/STAT3ER cells did not induce differentiation either. Together, our present data suggest that signaling other than the activation of STAT3 and MEK1 may be necessary for M1 cell-growth arrest and differentiation, while a set of early genes of LIF are induced by only STAT3 activation.

Topics: Animals; Apoptosis; Blotting, Northern; Cell Cycle; Cell Cycle Proteins; Cell Differentiation; Cell Division; Cell Line, Tumor; Cell Proliferation; Cyclin-Dependent Kinase Inhibitor p19; DNA; DNA-Binding Proteins; Down-Regulation; Enzyme Activation; Estrogen Antagonists; Immunoblotting; Interferon Regulatory Factor-1; Interleukin-6; Leukemia; Leukemia Inhibitory Factor; MAP Kinase Kinase 1; Mice; Oncogene Protein p65(gag-jun); Phosphoproteins; Phosphorylation; RNA, Messenger; Signal Transduction; STAT3 Transcription Factor; Tamoxifen; Time Factors; Trans-Activators

MLL fusion proteins are oncogenic transcription factors that are associated with aggressive lymphoid and myeloid leukemias. We constructed an inducible MLL fusion, MLL-ENL-ERtm, that rendered the transcriptional and transforming properties of MLL-ENL strictly dependent on the presence of 4-hydroxy-tamoxifen. MLL-ENL-ERtm-immortalized hematopoietic cells required 4-hydroxy-tamoxifen for continuous growth and differentiated terminally upon tamoxifen withdrawal. Microarray analysis performed on these conditionally transformed cells revealed Hoxa9 and Hoxa7 as well as the Hox coregulators Meis1 and Pbx3 among the targets upregulated by MLL-ENL-ERtm. Overexpression of the Hox repressor Bmi-1 inhibited the growth-transforming activity of MLL-ENL. Moreover, the enforced expression of Hoxa9 in combination with Meis1 was sufficient to substitute for MLL-ENL-ERtm function and to maintain a state of continuous proliferation and differentiation arrest. These results suggest that MLL fusion proteins impose a reversible block on myeloid differentiation through aberrant activation of a limited set of homeobox genes and Hox coregulators that are consistently expressed in MLL-associated leukemias.

Topics: Animals; Cell Line; Cell Transformation, Neoplastic; Down-Regulation; Genes, Homeobox; Homeodomain Proteins; Humans; Leukemia; Mice; Myeloid Ecotropic Viral Integration Site 1 Protein; Myeloid-Lymphoid Leukemia Protein; Neoplasm Proteins; Oncogene Proteins, Fusion; Receptors, Estrogen; Tamoxifen; Up-Regulation

Presence of a collagenolytic activity has been demonstrated in the human leukemic cell line K562. Among various effectors studied, tamoxifen, a well-known antiestrogenic compound, exhibited a strong inhibitory effect. After 3 days of culture in the presence of 10(-7) M of tamoxifen, 75% of the collagenolytic activity was inhibited. Hydroxytamoxifen and N-desmethyltamoxifen were equally potent inhibitors though devoid of the direct cytotoxic effect. Cis-tamoxifen was less efficient. K562 cells have no binding sites for estrogens but they possess high affinity binding sites for 3H-tamoxifen (295 fmol/mg of proteins, KD = 0.25 x 10(-9) M). Tamoxifen had no effect on cellular differentiation or enzyme secretion. Anticollagenolytic activity of tamoxifen (10(-7)-10(-6) M) could be related to its inhibitory action on plasmin and plasminogen activator.

Topics: Binding Sites; Collagen; Estrogen Antagonists; Fibrinolysin; Humans; Leukemia; Plasminogen Inactivators; Tamoxifen; Tumor Cells, Cultured