Abstract
Purpose of Study: Juvenile myelomonocytic leukemia (JMML) is a myeloproliferative disorder characterized by overproduction of myelomonocytic cells. Activating mutations of PTPN11, which encodes the protein tyrosine phosphatase, Shp-2, are found in 35% of JMML patients. Murine bone marrow low density mononuclear cells (LDMNCs) expressing activating Shp-2 mutants preferentially undergo myelomonocytic differentiation despite being subjected to conditions that select for mast cell development (Chan et al., Blood 106, Abstract #3519, 2005). Consistently, GATA-2 expression is dramatically reduced in cells expressing activating Shp-2, suggesting that Shp-2 gain-of-function mutants alter hematopoietic lineage-specific transcription factors and thus shift myeloid differentiation toward the myelomonocytic lineage. We hypothesized that ectopic GATA2 expression would normalize the aberrant myelomonocytic differentiation induced by activating Shp-2 mutants.
Methods Used: To address this hypothesis, we utilized retroviral co-transduction of LDMNCs to generate four experimental groups: 1) pMIEG3-Shp-2WT plus pCD4 (empty vector); 2) pMIEG3-Shp2D61Y plus pCD4; 3) pMIEG3-Shp-2WT plus pCD4-GATA2; and 4) pMIEG3-Shp2D61Y plus pCD4-GATA2. Cells were stained with anti-human CD4-APC, sorted for the EGFP+APC+ cells, and plated into progenitor assays. Colonies were scored for colony forming unit (CFU)-granulocyte-macrophage (GM), monocyte (M), granulocyte (G), and granulocyte-erythroid-monocyte-megakaryocyte (GEMM).
Summary of Results: As predicted, cells co-transduced with PTPN11 mutant D61Y and pCD4 produced significantly more CFU-M than cells co-transduced with WT Shp-2 and pCD4 (37.2+/-3.6 v. 14.4+/-3.4, =6, p=0.0008). However, upon co-transduction with GATA2, the number of CFU-M generated from D61Y-expressing cells was significantly reduced and was similar to that observed in cells expressing WT Shp-2. Quantitative RT-PCR verified restoration of GATA2 expression in cells co-transduced with D61Y and pCD4-GATA2.
Conclusion: These findings demonstrate that restoration of GATA2 expression normalizes the propensity toward monocytic differentiation induced by the activating PTPN11 mutant D61Y.
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Yang, Z., Voorhorst, C. & Chan, R. 24 Ectopic Expression of GATA2 Normalizes Aberrant Myelomonocytic Differentiation Induced by Activating PTPN11 Mutations.. Pediatr Res 60, 494 (2006). https://doi.org/10.1203/00006450-200610000-00046
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DOI: https://doi.org/10.1203/00006450-200610000-00046