p19^Arf Cooperation in Shp2-Induced Leukemogenesis.

Shp2 is a protein-tyrosine phosphatase which has both proliferation and apoptosis functions in hematopoietic cells. Gain-of-function (GOF) Shp2 mutations are found in Juvenile Myelomonocytic Leukemia (JMML) and Noonan syndrome patients predisposed to JMML, as well as in common childhood leukemias. Genetic data suggest that Shp2 alone is insufficient for leukemogenesis, and other genetic alterations are required, such as disruption of tumor suppressor pathways. We hypothesized that Arf deletion will enhance GOF Shp2-induced leukemogenesis. In colony forming cell assays (CFC), E76K Shp2 expression in arf−/− bone marrow cells led to increased colony number and size compared to E76K Shp2 expression in arf+/− cells or arf−/− cells with vector alone. To assay leukemia induction, arf+/− or arf−/− bone marrow cells transduced with E76K Shp2 mutant were transplanted into lethally irradiated WT C57BL/6 mice. We found that mice transplanted with E76K Shp2/arf−/− bone marrow cells developed leukemia/lymphoma at an accelerated rate compared to mice transplanted with E76K Shp2/arf+/− or vector/arf−/− bone marrow. While the most common hematological disease in mice transplanted with either E76KShp2/arf+/− or vector/arf−/− bone marrow was myeloproliferation, mice with E76K Shp2/arf−/− bone marrow developed predominantly B220+ leukemia. Compared to leukemia in E76K Shp2/arf+/− reconstituted mice, leukemias arising from E76K Shp2/arf−/− bone marrow were much more aggressive, effacing the bone marrow and spleen, infiltrating the liver diffusely and into surrounding tissue. Diseases arising from vector/arf−/− bone marrow were either localized lymphoma or mild myeloproliferation. Both E76K Shp2 expression and arf deletion were associated with the presence of BrdU-positive immature hematopoietic progenitors in the spleens of pre-leukemic mice, consistent with extramedullary hematopoiesis. These findings suggest that arf deletion cooperated with E76K Shp2’s proliferative function to accelerate leukemogenesis.