Genetic basis of myeloproliferative disorder

Myeloproliferative disorders (MPD) are a heterogeneous group of clonal hematopoietic diseases characterized by an excessive production of myeloid cells and an inherent tendency for thrombosis, bleeding, and leukemic transformation. A major step towards understanding the pathophysiology was the discovery of a mutation (V671F) in the JAK2 gene in about 50% of MPD patients. The mutation confers factor independent growth of hematopoietic cells via a constitutive activation of the JAK/STAT pathway. The JAK2-V617F mutation associates with clinical complications such as thrombosis, secondary fibrosis, and bleeding. Recent studies from our lab and others have demonstrated that the JAK2-V617F mutation cannot account for the entire phenotype associated with MPD and rather represents a late event in the genetic evolution of MPD stem cells. We have shown that deletions on chromosome 20q (del20q) often occur before the acquisition of the JAK2-V617F mutation. In this proposal, we plan to study the genetic basis of MPD by examining the functional relevance of previously identified chromosomal defects in MPD (del20q, uniparental disomies on chromosomes 10q and 11q) and search for new aberrations using high-resolution genomic analysis. In Specific aim 1 we will examine the clinical data of patients with and without del20q. To date, a thorough analysis of clinical relevance of the del20q has not been performed. We plan to study the clinical phenotype of 580 MPD patients in prospectively and retrospectively observed cohorts and correlate the clinical variables with the presence of the del20q. Specific aim 2: In our previous work, we have shown that del20q often occurs before the acquisition of the JAK2- V617F mutation, and thus, represent an early somatic stem cell aberration. We propose experiments that aim to identify the relevant gene within the del20q chromosomal region. We will study the role of del20q candidate genes in regulating growth of hematopoietic cells and address their possible collaboration with the JAK2-V617F mutation. To identify the relevant gene within the del20q region, we will perform sequencing, gene expression, and methylation analysis of the candidate genes. We will also examine these genes using in vitro and in vivo functional assays. Specific aim 3: Two specific chromosomal defects identified previously by us in MPD patients will be further studied. These include uniparental disomy of chromosomes 10q and 11q. We plan to apply positional cloning to identify potential tumor suppressor genes or oncogenes affected by these chromosomal defects. Additionally, we will apply two approaches of genomic analysis to identify new mutations that play a functional role in the pathogenesis of MPD. Functional characterization of the target gene(s) within the common deleted region on 20q would greatly contribute to our understanding of clonal evolution of MPD and other hematopoietic malignancies in which del20q is frequently found. Results from these experiments will provide insight into the genetic mechanisms leading to MPD and potentially identify new targets for therapy also applicable to other myeloid malignancies.

Myeloproliferative disorders (MPD) are a heterogeneous group of blood disease characterized by excessive production of myeloid cells such as red blood cells, platelets, and granulocytes. MPD occurs mainly in elderly with a mean age at diagnosis between 50-60 years. Although patients typically manifest a stable chronic phase of the disease, some patients progress to acute leukemia or have thrombotic events that significantly influence mortality. As typical for cancer, the cells manifesting the disease are derived from a single cell that acquired a somatic mutation and expanded into a clone. The project described here aimed at the identification of somatic mutations in MPD that initiate clonal cellular growth. Apart from the previously described JAK2 gene mutations, we defined a number of new genetic defects in MPD by employing the latest genome analysis technologies such as SNP array genotyping and next generation sequencing. Most notably, we identified deletions of the transcription factor Ikaros and various defects of the tumor suppressor gene TP53 in patients that transformed from chronic phase MPD to post-MPD acute leukemia. We have also discovered that inherited gene variants significantly influence the occurrence of somatic mutations that cause MPD. This work demonstrated a unique interplay between inherited and acquired genetic lesions in cancer. Our work had contributed new genetic markers that can help predict detrimental disease complications such as leukemic transformation. We have also shown that MPD is a genetically heterogeneous disease emphasizing the need for individualized therapeutic approaches in the treatment of MPD.