Análisis de nuevas alteraciones genéticas en neoplasias mieloproliferativas crónicas BCR-ABL1 negativas. Desarrollo de un modelo experimental de mutaciones de tipo 1 y tipo 2 en CALR en Caenorhabditis elegans
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Materias Investigacion::Ciencias de la Salud::Oncología
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EDER AZANZA, Laura. “Análisis de nuevas alteraciones genéticas en neoplasias mieloproliferativas crónicas BCR-ABL1 negativas. Desarrollo de un modelo experimental de mutaciones de tipo 1 y tipo 2 en CALR en Caenorhabditis elegans”. Vizmanos, J.L. (dir.). Tesis doctoral. Universidad de Navarra, Pamplona, 2017.
The genetic events that caused the BCR-ABL1 negative myeloproliferative neoplasms (MPNs) were elusive until 2005. That year, a JAK2 somatic mutation (p.V617F) was described in a significant proportion of patients. Shortly after, mutations in JAK2 exon 12 and in MPL were also described. All of them are considered primary events that promote an aberrant activation of the canonical JAK/STAT pathway and seem to be mutually exclusive. In recent years, many other mutated genes affecting different cell processes have been described. Pointing to that, these diseases are more complex than initially thought. Specifically, recent findings suggest that mutations in genes encoding epigenetic regulators are more frequent than expected. One of the last genes described as mutated is SETBP1. In silico tools show that there are several human genes paralogs to SETBP1 such as NSD1, NSD2 and NSD3 that are also involved in the development of other hematologic malignancies. For this reason, we analysed the mutational profile of NSD1, NSD2, NSD3 and SETBP1 in a selected group of patients with BCR-ABL1 negative CMPNs with or without p.V617F JAK2 mutation. No missense changes have been detected in NSD1, NSD2 and NSD3 in any group analysed suggesting that these genes are not frequently mutated in these diseases. However, although the coexistence of JAK2 and SETPBP1 mutations in MNPs has not been described before, we have identified a novel p.S867G SETBP1 mutation in a patient with p.V617F JAK2 positive primary myelofibrosis (PMF). In line with this, our group previously found new base substitution changes in JAK2 exon 8 (p.R340Q and p.Y317H) in a minor proportion of p.V617F JAK2 negative PMF patients. These changes affect FERM domain of this protein that could have an oncogenic role as predicted by in silico tools. Recent studies have also reported oncogenic mutations in FERM domains of JAK1 and JAK3 and also an oncogenic mutation in this domain was previously described in Drosophila melanogaster in hop (a JAK2 ortholog). All of this led us to hypothesize that p.R340Q and p.Y317H could also have a role in malignant transformation. Functional analyses show that p.Y317H, but not p.R340Q, has different transforming features to p.V617F which may represent an important event in the progression of the disease. Although important advances have been made to understand the genetic aberrations that cause CMPNs, until 2013 there were a significant proportion of patients in which the molecular basis of the disease was still unknown. That year some groups reported that most of p.V617F JAK2/MPL-negative essential thrombocythemia (ET) and PMF patients have mutations in CALR, the gene that encodes calreticulin. All of these mutations lead to a new C-terminal tail of the protein and most of them are classified as either type 1 (52-bp deletion) or type 2 (5-bp insertion). Emerging evidences suggest that both types have different biological features associated with distinct clinical outcomes. Recently, it has been reported that CALR mutations promote the aberrant activation of the JAK/STAT pathway through MPL although it seems that the differences observed between mutants are not directly related to the activation of this pathway. We wanted to functionally characterize the pathogenic mechanisms of CALR mutations in a Caernorhabditis elegans model through the generation of stable mutant organisms by CRISPR-Cas9. We have found differences between both types of mutations that could be related to the differences in clinical features. In our model, type 1 mutation seems to lead to a loss of function of the protein but the type 2 could be a gain-of-function mutation with an additional pathogenic mechanism possibly related to the regulation of transcription through nhr-17.

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