Expression of HOX genes, HOX cofactors, and MLL in phenotypically and functionally defined subpopulations of leukemic and normal human hematopoietic cells

Abstract

To explore the possibility that deregulated HOX gene expression might commonly occur during leukemic hematopoiesis, we compared the relative levels of expression of these and related genes in phenotypically and functionally defined subpopulations of AML blasts and normal hematopoietic cells. Initially, a semi-quantitative RT-PCR technique was used to amplify total cDNA from total leukemic blast cell populations from 20 AML patients and light density cells from four normal bone marrows. Expression of HOX genes (A9, A10, B3 and B4), MEIS1 and MLL was easily detected in the majority of AML samples with the exception of two samples from patients with AML subtype M3 (which expressed only MLL). Low levels of HOXA9 and A10 but not B3 or B4 were seen in normal marrow while MLL was easily detected. PBX1a was difficult to detect in any AML sample but was seen in three of four normal marrows. Cells from nine AML patients and five normal bone marrows were FACS-sorted into CD34 ؉ CD38 ؊ , CD34 ؉ CD38 ؉ and CD34 ؊ subpopulations, analyzed for their functional properties in long-term culture (LTC) and colony assays, and for gene expression using RT-PCR. 93 ؎ 14% of AML LTC-initiating cells, 92 ± 14% AML colony-forming cells, and Ͼ99% of normal LTC-IC and CFC were CD34 ؉ . The relative level of expression of the four HOX genes in amplified cDNA from CD34 ؊ as compared to CD34 ؉ CD38 ؊ normal cells was reduced Ͼ10-fold. However, in AML samples this down-regulation in HOX expression in CD34 ؊ as compared to CD34 ؉ CD38 ؊ cells was not seen (P Ͻ 0.05 for comparison between AML and normal). A similar difference between normal and AML subpopulations was seen when the relative levels of expression of MEIS1, and to a lesser extent MLL, were compared in CD34 ؉ and CD34 ؊ cells (P Ͻ 0.05). In contrast, while some evidence of down-regulation of PBX1a was found in comparing CD34 ؊ to CD34 ؉ normal cells it was difficult to detect expression of this gene in any subpopulation from most AML samples. Thus, the down-regulation of HOX, MEIS1 and to some extent MLL which occurs with normal hematopoietic differentiation is not seen in AML cells with similar functional and phenotypic properties.

Identification of E74-like factor 1 (ELF1) as a transcriptional regulator of the Hox cofactor MEIS1

“…Consistent with these initial observations, subsequent studies revealed that engineered overexpresssion of MEIS1 was a potent collaborating event with overexpresssed Hox genes or NUP98-HOX fusion genes that accelerates the onset of AML in murine bone marrow transplantation models (9)(10)(11)(12). MEIS1 is frequently upregulated in human primary acute myelogenous leukemia (AML) and acute lymphocytic leukemia (ALL) samples (13)(14)(15). Recently Wong et al have shown that MEIS1 is an essential and rate-limiting regulator of MLL-leukemia stem cell (LSC) potential in murine models (16).…”

Hox genes in hematopoiesis and leukemogenesis

“…The unequivocal role of Hox genes in the pathogenesis of acute leukemia A central role for HOX genes in hematological malignancies is supported by the frequently observed elevation of HOX and MEIS1 gene expression in AML patient samples (Golub et al, 1999;Kawagoe et al, 1999;Lawrence et al, 1999;Afonja et al, 2000;Drabkin et al, 2002). Indeed, HOXA9 is the single most highly correlated gene (out of 6817) for poor prognosis in AML (Golub et al, 1999).…”

“…Gene expression analyses of both mouse and human bone marrow (BM) samples revealed that the majority of Hox genes of the A, B and C clusters are expressed in hematopoietic cells and, for the most part, are preferentially expressed in hematopoietic stem cell (HSC)-enriched subpopulations and in immature progenitor compartments and downregulated during differentiation and maturation (Giampaolo et al, 1994(Giampaolo et al, , 1995Moretti et al, 1994;Sauvageau et al, 1994;Kawagoe et al, 1999;Pineault et al, 2002). These observations led to the hypotheses that Hox genes play key functions in early hematopoietic cells, including HSCs, and that dysregulated Hox expression may impact leukemic transformation.…”

Activation of SHP2 Protein-tyrosine Phosphatase Increases HoxA10-induced Repression of the Genes Encoding gp91PHOX and p67PHOX

“…Differentiation stage-specific expression suggests that Hox proteins may regulate progression of myelopoiesis. Indeed, a number of studies indicate the importance of HoxA proteins for normal myeloid development (5)(6)(7)(8)(9).…”

“…Expression of the entire group of Abd HoxA proteins has been demonstrated in human AML bone marrow samples from subjects with translocations involving the MLL (mixed lineage leukemia) gene (8). In murine transplantation experiments, expression of such leukemia-associated MLL fusion proteins induces myeloproliferation, which progresses to blast crisis over several months (14 -19).…”