Poster Board III-897
The NUP98-HOXA9 fusion oncogene results from a t(7;11)(p15;p15) chromosomal translocation and is associated with inferior prognosis in de novo and treatment-related acute myeloid leukemia (AML), as well as blast crisis in chronic myeloid leukemia (CML). HOXA9 belongs to the highly-conserved HOX gene family of developmental transcription factors and is critical for vertebrate hematopoiesis. Upregulation of HOXA9 is a frequent phenomenon in human AML, implicating a central role in myeloid disease. Thus, elucidating the activity of oncogenic NUP98-HOXA9 may reveal universal mechanisms of leukemogenesis and lead to design of targeted therapies. The zebrafish is a robust model for studying vertebrate hematopoiesis and leukemogenesis, by virtue of its ex utero development, and conserved genetics and cell biology. We have engineered a Cre-lox inducible transgenic zebrafish harboring the human NUP98-HOXA9 fusion oncogene downstream of the zebrafish pu.1 promoter. NUP98-HOXA9 perturbed zebrafish embryonic hematopoiesis, leading to upregulated expression of myeloid-specific genes, pu.1, lysC, and l-plastin, and downregulation of the erythroid-specific gene, gata1. These changes in gene transcription affect both ‘primitive’ and ‘definitive’ waves of zebrafish hematopoiesis (including the recently-described erythro-myeloid progenitors [EMPs]), suggesting that NUP98-HOXA9 reprograms blood cell precursors to a predominantly myeloid fate. Preferential upregulation of pu.1 during ‘primitive’ hematopoiesis further suggests an impaired potential for terminal myeloid differentiation. These effects appear developmentally mediated, as cell proliferation was not increased as measured by phosphorylated histone-H3 (pH3) staining. However, following ionizing irradiation (IR), NUP98-HOXA9 expression preserved cell proliferation and suppressed apoptosis in the presence of DNA damage. Irradiated transgenic embryos displayed wild-type levels of pH3, suggesting inhibition of cell cycle checkpoints. Irradiated embryos also displayed reduced levels of the conserved apoptotic marker, activated caspase-3, as well as downregulation of the puma pro-apoptotic gene, and upregulation of bcl2 and bcl-xL anti-apoptotic genes. These data postulate suppression of caspase-3-dependent apoptosis as a mechanism for NUP98-HOXA9 -mediated oncogenesis in vivo. Incomplete phenotypic ‘rescue’ with bcl-xL morpholino suggests insufficiency of single-target therapy, with a need for multi-target interruptions to restore radiosensitivity. Between 20-24 months post fertilization, a proportion of these NUP98-HOXA9 transgenic zebrafish (n=9/19; 47%) presented with gross abdominal masses and/or abnormal swimming behavior. Thus far, histological sectioning has demonstrated enlarged kidneys and livers, with evidence of infiltrative, pleiomorphic, mitotically-active cells with myeloid morphology. This pathology is reminiscent of the myeloproliferative disease (MPD) observed in NUP98-HOXA9 transgenic mice. Taken together, these data provide insights into the mechanisms underlying NUP98-HOXA9-mediated leukemogenesis and provide an unprecedented opportunity for in vivo chemical modifier screens to identify promising therapeutic agents that restore a normal phenotype.
No relevant conflicts of interest to declare.
Asterisk with author names denotes non-ASH members.