Tese de Doutoramento, Medicina, Genética, Oncologia e Toxicologia Humana, Universidade Nova de Lisboa

Myeloproliferative neoplasms are classically divided into BCR-ABL1 (Philadelphia chromosome) positive chronic myeloid leukemia and BCR-ABL1 negative myeloproliferative neoplasms, including polycythemia vera, essential thrombocythemia and primary myelofibrosis. These disorders have origin from the malignant transformation of a hematopoietic stem-cell, leading to abnormal amplification and proliferation of myeloid lineages. The phenotypic diversity of Philadelphia chromosome negative myeloproliferative neoplasms results from the combination of somatic mutations already identified and characterized, inherited genetic variability, postgenetic regulation, and individual conditioners. According to the literature, several genetic polymorphisms have been identified, which may influence the apoptosis mechanisms and the DNA repair capacity, compromising genetic transcription and/or cell function, conferring genetic predisposition to disease, including myeloproliferative disorders, and conditioning the therapeutic response, the clinical outcome and prognosis. The present work had the purpose of characterize the Portuguese population according to the type of Philadelphia chromosome negative myeloproliferative neoplasm, its prevalence and the presence of JAK2 mutation, with special emphasis on some particular patients; investigate the role of polymorphisms in genes involved in apoptosis and base excision repair pathways in these disorders susceptibility and, along with other risk factors, in survival, clinical outcome and prognosis. Case-control studies were carried out in a Caucasian Portuguese sample of 133 patients and 281 matched control subjects, nine and eight polymorphisms of genes involved in apoptosis and base excision repair pathways, respectively, were selected and genotyped using real time polymerase chain reaction technique and statistical analysis was performed with SPSS version 22.0. The results from this study revealed a pathology distribution of 60.2% of patients with essential thrombocythemia, 29.3% with polycythemia vera and 10.5% with primary myelofibrosis, with a discrete predominance in females, and a total of 75.0% of patients positive for the presence of the JAK2 V617F mutation, with an increased incidence in essential thrombocythemia and a decreased incidence in polycythemia vera patients than expected. Among the patients, some particular cases were highlighted and presented in this thesis, namely the case of a polycythemia vera patient JAK2 exon 12 positive, presenting a novel mutation – c.1605GT (p.Met535Ile) - associated with another mutation previously described, evidencing an atypical clinical phenotype; two patients with essential thrombocythemia, with suspicion of the rare coexistence of JAK2 V617F mutation and BCR-ABL1 translocation; and a patient with chronic myeloid leukemia, presenting a new BCR-ABL1 mutation kinase domain mutation – c.839TG (p.Val280Gly) – which might be associated with resistance to imatinib. Concerning the polymorphisms investigated, our results revealed potential associations between some polymorphisms and individual susceptibility to these disorders, suggesting the potential involvement of CASP9 (Phe136Phe) gene and XRCC1_399 (Gln399Arg) and MUTYH (Gln335His) genes polymorphisms after stratification by pathology diagnosis for essential thrombocythemia patients and when they are women. When patients are JAK2 positive an association with CASP9 (Phe136Phe) gene and XRCC1_399 (Gln399Arg) gene polymorphisms was found. In base excision repair genes, combination of alleles also demonstrated an association with the disease for one specific haplogroup. Regarding the clinical outcome in patients under treatment, the majority of them with hydroxyurea, according to our results: 17 patients showed progression to secondary myelofibrosis/acute myeloid leukemia, influencing survival and being associated with exposure to cytoreductive agents. In these patients, there was also global evidence of association with CASP8 (3’UTR) gene and XRCC1_194 (Arg194Trp) gene polymorphisms, and after stratification for essential thrombocythemia with CASP9 (Arg173His) gene, and APEX1 (Asp148Glu) and XRCC1_194 (Arg194Trp) genes polymorphisms; 11 patients developed a new primary nonmyeloid neoplasm, evidencing a global association with CASP8 (Asp270His) gene and XRCC1_399 (Gln399Arg) gene polymorphisms; 22 patients presented thrombotic events and a global association with XRCC1_399 (Gln399Arg) gene polymorphisms. The results shown in the present dissertation helped to characterize a population of Philadelphia negative myeloproliferative neoplasms patients that could reflect Portuguese reality, and have revealed the potential role of polymorphisms and other factors in disease susceptibility and clinical outcome of patients under treatment, with prognostic impact. Although larger studies are required to confirm these results, these new data may contribute to a best knowledge of the pathophysiology and, in the future, to a more rational and efficient choice of therapeutic strategies to be adopted in these disorders.