Expression of HOX genes, HOX cofactors, and MLL in phenotypically and functionally defined subpopulations of leukemic and normal human hematopoietic cells.
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 >99% 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 >10-fold. However, in AML samples this down-regulation in HOX expression in CD34- as compared to CD34+CD38- cells was not seen (P < 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 < 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.
Kawagoe
H
H
British Columbia Cancer Agency, and Department of Medicine, University of British Columbia, Vancouver, Canada.
Humphries
R K
RK
Blair
A
A
Sutherland
H J
HJ
Hogge
D E
DE
D036541:ADP-ribosyl Cyclase; D051997:ADP-ribosyl Cyclase 1; D000328:Adult; D000368:Aged; D015703:Antigens, CD; D018952:Antigens, CD34 / Q000032:analysis; D000943:Antigens, Differentiation / Q000032:analysis; D001854:Bone Marrow Cells / Q000378:metabolism*; D004268:DNA-Binding Proteins / Q000235:genetics*; D005260:Female; D015973:Gene Expression Regulation, Leukemic; D005801:Genes, Homeobox; D011495:Histone-Lysine N-Methyltransferase; D018398:Homeodomain Proteins / Q000235:genetics*; D006801:Humans; D015470:Leukemia, Myeloid, Acute / Q000235:genetics*; D008297:Male; D008562:Membrane Glycoproteins; D008875:Middle Aged; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D051788:Myeloid-Lymphoid Leukemia Protein; D009244:NAD+ Nucleosidase / Q000032:analysis; D009363:Neoplasm Proteins / Q000235:genetics*; D011519:Proto-Oncogenes; D014157:Transcription Factors / Q000235:genetics*
D015703:Antigens, CD; D018952:Antigens, CD34; D000943:Antigens, Differentiation; D004268:DNA-Binding Proteins; C090460:HOXB4 protein, human; D018398:Homeodomain Proteins; C497422:KMT2A protein, human; C000616391:MEIS1 protein, human; D008562:Membrane Glycoproteins; D000075064:Myeloid Ecotropic Viral Integration Site 1 Protein; D009363:Neoplasm Proteins; D014157:Transcription Factors; C097899:homeobox protein HOXA9; D051788:Myeloid-Lymphoid Leukemia Protein; D011495:Histone-Lysine N-Methyltransferase; D036541:ADP-ribosyl Cyclase; C498154:CD38 protein, human; D009244:NAD+ Nucleosidase; D051997:ADP-ribosyl Cyclase 1