Poor outcome for patients with acute myelogenous leukemia is primarily due to treatment refractory disease. The identification of mechanisms that regulate leukemic cell survival and drug resistance are essential for defining new therapeutic targets. Our laboratory identified two novel activating mutations of c-Kit at codon 816 (Asp816) from MO7e AML cells and de novo AML that cause cell transformation, cytokine independent survival, increased proliferation and resistance to chemotherapeutic drugs and radiation. Expression of constitutively activated STAT3 restores the mutant receptor's transforming ability, partially reconstitutes cytokine independent survival and increased proliferation, but does not confer the drug resistant phenotype. However, when PI-3K was inhibited with the specific blocker, LY29004, MO7e transduced with Asp816 mutant c-Kit encoding cDNA showed increased cytotoxic drug sensitivity. These phenotypic changes suggest that Asp816 c-Kit mutations play a role in the poor clinical outcome of some patients with AML. The proposed studies will build on these observations and test the hypothesis that activating Asp816 mutations of the c-Kit receptor lead to altered ability to activate signal transduction pathways, including STAT3, PI-3K and downstream effectors, resulting in a phenotype that predicts a poor prognosis in patients with AML.
Specific aims i nclude:
Aim 1) To test the hypothesis that Asp816 mutant c-Kit receptor activates STAT3 directly. An alternative hypothesis is that the mutant receptor referentially recruits of other specific kinases to activate STAT3.
Aim 2) To test the hypothesis that Asp816 mutant c-Kit mediates a survival advantage and drug resistance phenotype through activation of STAT3 and PI-3K pathways along with downstream targets. Methods will include biochemical and gene expression analyses.
Aim 3) To test the hypothesis that the phenotypic changes mediated by Asp816 c-kit mutations lead to poor outcome in patients with AML. The proposed studies will provide additional cellular targets for the development of novel therapeutic approaches in patients with AML characterized by activating mutations of c-Kit and, possibly, other receptor tyrosine kinases. These data may also be useful for stratification of patients according to cytokine receptor mutations and subsequent treatment with non-genotoxic therapies that target specific biological pathways.
