HOXA9 is required for survival in human MLL-rearranged acute leukemias.

Leukemias that harbor translocations involving the mixed lineage leukemia gene (MLL) possess unique biologic characteristics and often have an unfavorable prognosis. Gene expression analyses demonstrate a distinct profile for MLL-rearranged leukemias with consistent high-level expression of select Homeobox genes, including HOXA9. Here, we investigated the effects of HOXA9 suppression in MLL-rearranged and MLL-germline leukemias using RNA interference. Gene expression profiling after HOXA9 suppression demonstrated co-down-regulation of a program highly expressed in human MLL-AML and murine MLL-leukemia stem cells, including HOXA10, MEIS1, PBX3, and MEF2C. We demonstrate that HOXA9 depletion in 17 human AML/ALL cell lines (7 MLL-rearranged, 10 MLL-germline) induces proliferation arrest and apoptosis specifically in MLL-rearranged cells (P = .007). Similarly, assessment of primary AMLs demonstrated that HOXA9 suppression induces apoptosis to a greater extent in MLL-rearranged samples (P = .01). Moreover, mice transplanted with HOXA9-depleted t(4;11) SEMK2 cells revealed a significantly lower leukemia burden, thus identifying a role for HOXA9 in leukemia survival in vivo. Our data indicate an important role for HOXA9 in human MLL-rearranged leukemias and suggest that targeting HOXA9 or downstream programs may be a novel therapeutic option.

[1]  Y. Kaneko,et al.  Clinical characteristics of infant acute leukemia with or without 11q23 translocations. , 1988, Leukemia.

[2]  F. Behm,et al.  Secondary acute myeloid leukemia in children treated for acute lymphoid leukemia. , 1989, The New England journal of medicine.

[3]  S. Korsmeyer,et al.  Molecular rearrangements on chromosome 11q23 predominate in infant acute lymphoblastic leukemia and are associated with specific biologic variables and poor outcome. , 1993, Blood.

[4]  J. Rowley,et al.  Rearrangements of the MLL gene in therapy-related acute myeloid leukemia in patients previously treated with agents targeting DNA- topoisomerase II , 1993 .

[5]  S. Korsmeyer,et al.  The chromosome 4q21 gene (AF-4/FEL) is widely expressed in normal tissues and shows breakpoint diversity in t(4;11)(q21;q23) acute leukemia. , 1993, Blood.

[6]  A. Schulz,et al.  Pre-pre-B acute lymphoblastic leukemia: high frequency of alternatively spliced ALL1-AF4 transcripts and absence of minimal residual disease during complete remission. , 1994, Blood.

[7]  G. Sauvageau,et al.  Differential expression of homeobox genes in functionally distinct CD34+ subpopulations of human bone marrow cells. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[8]  N. Heerema,et al.  Cytogenetic features of infants less than 12 months of age at diagnosis of acute lymphoblastic leukemia: impact of the 11q23 breakpoint on outcome: a report of the Childrens Cancer Group. , 1994, Blood.

[9]  Frequency and prognostic significance of HRX rearrangements in infant acute lymphoblastic leukemia: a Pediatric Oncology Group study. , 1994 .

[10]  U. Thorsteinsdóttir,et al.  Overexpression of HOXB4 in hematopoietic cells causes the selective expansion of more primitive populations in vitro and in vivo. , 1995, Genes & development.

[11]  G. Sauvageau,et al.  Stage- and lineage-specific expression of the HOXA10 homeobox gene in normal and leukemic hematopoietic cells. , 1995, Experimental hematology.

[12]  M. Seto,et al.  11q23 Aberration is an additional chromosomal change in de novo acute leukemia after treatment with etoposide and mitoxantrone , 1996, American journal of hematology.

[13]  Keisuke Toyama,et al.  The t(7;11)(p15;p15) translocation in acute myeloid leukaemia fuses the genes for nucleoporin NUP96 and class I homeoprotein HOXA9 , 1996, Nature Genetics.

[14]  Takuro Nakamura,et al.  Cooperative activation of Hoxa and Pbx1-related genes in murine myeloid leukaemias , 1996, Nature Genetics.

[15]  A. Feinberg,et al.  Fusion of the nucleoporin gene NUP98 to HOXA9 by the chromosome translocation t(7;11)(p15;p15) in human myeloid leukaemia , 1996, Nature Genetics.

[16]  G. Sauvageau,et al.  Mice bearing a targeted interruption of the homeobox gene HOXA9 have defects in myeloid, erythroid, and lymphoid hematopoiesis. , 1997, Blood.

[17]  D. Izon,et al.  Loss of function of the homeobox gene Hoxa-9 perturbs early T-cell development and induces apoptosis in primitive thymocytes. , 1998, Blood.

[18]  J. Mesirov,et al.  Molecular classification of cancer: class discovery and class prediction by gene expression monitoring. , 1999, Science.

[19]  F. Frassoni,et al.  The retroviral transduction of HOXC4 into human CD34(+) cells induces an in vitro expansion of clonogenic and early progenitors. , 2000, Experimental hematology.

[20]  C. Lavau,et al.  Retrovirus-mediated gene transfer of MLL-ELL transforms primary myeloid progenitors and causes acute myeloid leukemias in mice. , 2000, Proceedings of the National Academy of Sciences of the United States of America.

[21]  C. Croce,et al.  Upregulation of Meis1 and HoxA9 in acute lymphocytic leukemias with the t(4 : 11) abnormality , 2001, Oncogene.

[22]  Unnur Thorsteinsdottir,et al.  Defining Roles for HOX and MEIS1 Genes in Induction of Acute Myeloid Leukemia , 2001, Molecular and Cellular Biology.

[23]  B. Smith,et al.  A FLT3 tyrosine kinase inhibitor is selectively cytotoxic to acute myeloid leukemia blasts harboring FLT3 internal tandem duplication mutations. , 2001, Blood.

[24]  M. Cleary,et al.  Molecular mechanisms of leukemogenesis mediated by MLL fusion proteins , 2001, Oncogene.

[25]  U. Thorsteinsdóttir,et al.  NUP98–HOXA9 expression in hemopoietic stem cells induces chronic and acute myeloid leukemias in mice , 2001, The EMBO journal.

[26]  R. Humphries,et al.  Differential expression of Hox, Meis1, and Pbx1 genes in primitive cells throughout murine hematopoietic ontogeny. , 2002, Experimental hematology.

[27]  C. Peschle,et al.  Expression pattern of HOXB6 homeobox gene in myelomonocytic differentiation and acute myeloid leukemia , 2002, Leukemia.

[28]  A. Borkhardt,et al.  Infant acute lymphoblastic leukemia – combined cytogenetic, immunophenotypical and molecular analysis of 77 cases , 2002, Leukemia.

[29]  Thomas A Milne,et al.  MLL targets SET domain methyltransferase activity to Hox gene promoters. , 2002, Molecular cell.

[30]  G. Sauvageau,et al.  Deregulated expression of HOXB4 enhances the primitive growth activity of human hematopoietic cells. , 2002, Blood.

[31]  A. Baron,et al.  Quantitative HOX expression in chromosomally defined subsets of acute myelogenous leukemia , 2002, Leukemia.

[32]  J. Downing,et al.  Classification, subtype discovery, and prediction of outcome in pediatric acute lymphoblastic leukemia by gene expression profiling. , 2002, Cancer cell.

[33]  C. Pui,et al.  Outcome of treatment in childhood acute lymphoblastic leukaemia with rearrangements of the 11q23 chromosomal region , 2002, The Lancet.

[34]  U. Thorsteinsdóttir,et al.  marrow cells induces stem cell expansion gene in bone Hoxa 9 associated − Overexpression of the myeloid leukemia , 2001 .

[35]  Toshiki Mori,et al.  ALL-1 is a histone methyltransferase that assembles a supercomplex of proteins involved in transcriptional regulation. , 2002, Molecular cell.

[36]  E. Lander,et al.  MLL translocations specify a distinct gene expression profile that distinguishes a unique leukemia , 2002, Nature Genetics.

[37]  K. Calvo,et al.  Nup98-HoxA9 immortalizes myeloid progenitors, enforces expression of Hoxa9, Hoxa7 and Meis1, and alters cytokine-specific responses in a manner similar to that induced by retroviral co-expression of Hoxa9 and Meis1 , 2002, Oncogene.

[38]  I. Weissman,et al.  MLL-GAS7 transforms multipotent hematopoietic progenitors and induces mixed lineage leukemias in mice. , 2003, Cancer cell.

[39]  R. Iggo,et al.  Induction of an interferon response by RNAi vectors in mammalian cells , 2003, Nature Genetics.

[40]  S. Armstrong,et al.  Gene expression signatures in MLL-rearranged T-lineage and B-precursor acute leukemias: dominance of HOX dysregulation. , 2003, Blood.

[41]  M. Cleary,et al.  Transformation of myeloid progenitors by MLL oncoproteins is dependent on Hoxa7 and Hoxa9. , 2003, Genes & development.

[42]  Rob Pieters,et al.  Inhibition of FLT3 in MLL. Validation of a therapeutic target identified by gene expression based classification. , 2003, Cancer cell.

[43]  John H Kersey,et al.  Hoxa9 influences the phenotype but not the incidence of Mll-AF9 fusion gene leukemia. , 2004, Blood.

[44]  J. Downing,et al.  Gene Expression Profiling of Pediatric Acute Myelogenous Leukemia Materials and Methods , 2022 .

[45]  Arndt Borkhardt,et al.  Hoxa9 and Meis1 Are Key Targets for MLL-ENL-Mediated Cellular Immortalization , 2004, Molecular and Cellular Biology.

[46]  Todd R Golub,et al.  Gene expression–based high-throughput screening(GE-HTS) and application to leukemia differentiation , 2004, Nature Genetics.

[47]  Chi Wai So,et al.  Leukemic transformation of hematopoietic progenitors by MLL-GAS7 in the absence of Hoxa7 or Hoxa9. , 2004, Blood.

[48]  T. Golub,et al.  Conditional MLL‐CBP targets GMP and models therapy‐related myeloproliferative disease , 2005, The EMBO journal.

[49]  A. Buchberg,et al.  Meis1-mediated apoptosis is caspase dependent and can be suppressed by coexpression of HoxA9 in murine and human cell lines. , 2005, Blood.

[50]  T. Golub,et al.  Gefitinib induces myeloid differentiation of acute myeloid leukemia. , 2005, Blood.

[51]  Richard A Young,et al.  Global and Hox-specific roles for the MLL1 methyltransferase. , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[52]  Judy Lieberman,et al.  Antibody mediated in vivo delivery of small interfering RNAs via cell-surface receptors , 2005, Nature Biotechnology.

[53]  Pablo Tamayo,et al.  Gene set enrichment analysis: A knowledge-based approach for interpreting genome-wide expression profiles , 2005, Proceedings of the National Academy of Sciences of the United States of America.

[54]  J. Hess,et al.  Leukemogenic MLL fusion proteins bind across a broad region of the Hox a9 locus, promoting transcription and multiple histone modifications. , 2005, Cancer research.

[55]  I. Weissman,et al.  Loss of expression of the Hoxa-9 homeobox gene impairs the proliferation and repopulating ability of hematopoietic stem cells. , 2005, Blood.

[56]  Paul A Clemons,et al.  The Connectivity Map: Using Gene-Expression Signatures to Connect Small Molecules, Genes, and Disease , 2006, Science.

[57]  T. Golub,et al.  Transformation from committed progenitor to leukaemia stem cell initiated by MLL–AF9 , 2006, Nature.

[58]  M. Cleary,et al.  Identification and characterization of leukemia stem cells in murine MLL-AF9 acute myeloid leukemia. , 2006, Cancer cell.

[59]  Anne E Carpenter,et al.  A Lentiviral RNAi Library for Human and Mouse Genes Applied to an Arrayed Viral High-Content Screen , 2006, Cell.

[60]  J. Mesirov,et al.  GenePattern 2.0 , 2006, Nature Genetics.

[61]  B. Cullen.  Enhancing and confirming the specificity of RNAi experiments , 2006, Nature Methods.

[62]  D. Campana,et al.  Age-related differences in leukemia biology and prognosis: the paradigm of MLL-AF4-positive acute lymphoblastic leukemia , 2007, Leukemia.

[63]  Scott A. Armstrong,et al.  MLL translocations, histone modifications and leukaemia stem-cell development , 2007, Nature Reviews Cancer.

[64]  M. Cleary,et al.  Protein arginine-methyltransferase-dependent oncogenesis , 2007, Nature Cell Biology.

[65]  Xiaobo Xia,et al.  H3K79 methylation profiles define murine and human MLL-AF4 leukemias. , 2008, Cancer cell.