Mechanism of CALR mutants in Myeloproliferative Neoplasms

Myeloproliferative neoplasms (MPNs) are diseases characterized by abnormal expansion of myeloid cells in the hematopoietic system. Mutations in the CALR gene are seen in about 25% of the MPN patients. The mutant CALR proteins have been shown to induce activation of the Thrombopoietin receptor which is a cytokine receptor expressed on the cell surface. However, it is important to elucidate the molecular mechanisms of the CALR mutant proteins in detail for the development of any novel therapies for MPN patients. To understand these details, we have generated a novel transgenic mouse model that expresses the human mutant CALR protein, specifically within the murine hematopoietic cells, and therefore develops the MPN disease phenotype at an early age. We propose to characterize the development of the disease in this mouse model by analyzing the peripheral blood and other hematopoietic organs of the mice, at different time points. In addition, we will transplant the bone marrow of the CALR mutant mice to immuno-deficient recipient mice to study the disease causing hematopoietic stem cells with the CALR mutations. CALR mutations induce the activation of the JAK-STAT signaling pathway. We will use transgenic mice without JAK2 or STAT5, to see if the absence of JAK2 or STAT5 can rescue the mice from CALR mutant driven MPN. In addition, we will treat the mice with JAK2 inhibitor Ruxolitinib, which is currently used in clinic for MPN patients with JAK2 mutations, to evaluate the efficacy of this drug on cells with CALR mutations. To understand the molecular details of the oncogenic program of the CALR mutant proteins, we will perform a global transcriptome analysis of the hematopoietic stem cells from the CALR-transgenic mice. This will allow the identification of the genes whose expression is drastically changed in the presence of CALR mutations. We will use the CRISPR-Cas9 genome editing system to confirm the role of these genes in CALR mutant driven disease. This study will provide a novel transgenic mouse model for MPN caused by CALR mutations, which will allow us to understand the biology of these mutations at a molecular level. This will lay the foundation of the development of novel therapeutic possibilities that may prove to be clinically relevant for MPN patients in future.

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