Análise do perfil global de metilação do DNA de pacientes com Síndrome Mielodisplásica

Abstract

Myelodysplastic syndrome (MDS) is characterized by a heterogeneous group of hematopoietic progenitor cell clonal diseases, identified by insufficient bone marrow and increased apoptosis, which leads to inefficient hematopoiesis and cytopenia in the peripheral blood of one or more strains. The pathogenesis of MDS involves epigenetic, cytogenetic and / or gene mutations in up to 80% of cases and it is believed that these changes are due to DNA damage, after exposure to endogenous, exogenous / environmental, physical, chemical and biological agents, that cause genomic instability. The altered DNA methylation appears as a participant in the molecular pathophysiology of the disease, activating or inactivating the genes involved in growth, differentiation and apoptosis. The objectives of this study were to evaluate the global profile of DNA methylation and hydroxymethylation (5-methylcytokine and 5-hydroxymethylcytosine) in patients with myelodysplastic syndrome (MDS) and to correlate the levels of 5-hydroxymethylcytosine and 5-methylcytosine with the levels of expression of genes related to the cell cycle and mitotic spindle (AURKA, AURKB, MAD2, CDC20, TPX2), DNA repair (ATM, RAD51, LIG 4, BRCA1, BRCA2 XRCC5, XRCC6, XPA, XPC, XPG, CSA and CSB ) and polymerases with translesion activity (REV3L, REV1, POLI, POLH, POLL, POLK, POLQ, POLN and PCNA). 73 patients with MDS diagnosed at a tertiary referral hospital and 10 controls were evaluated. For the cytogenetic study, the karyotype exam was performed by G-Band and for the global DNA methylation analysis, immunohistochemistry of the global methylation markers 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) was performed, in biopsy of marrow (BM). The scoring of marked bone marrow hematopoietic cells was performed and the association of global DNA methylation with the findings of chromosomal changes, clinical and laboratory variables, gene expressions and patient survival was made, looking for new biomarkers and / or possible targets treatments for MDS. We identified an increase in levels of 5mC in patients with altered karyotype (p = 0.022) and a decrease of 5hmC in patients older than 60 years (p = 0.044) with an increase in the 5mC / 5hmC ratio (p = 0.017). Comparing patients with MDS and healthy controls, we observed an increase in the ratio of 5mC / 5mC in the group with MDS (p = 0.039). Additionally, patients with MDS were divided into initial (SMD-CRDU, SMD-CRDM and SMD-SA) and advanced (SMD-EB 1 and 2) forms based on the 2016 WHO classification and we observed that the ratio of 5mC / 5hmC was higher in patients classified as advanced forms (p = 0.040). Regarding bone marrow cellularity, we found that patients with hypercellular marrow had an increase in the ratio of 5hmC / 5mC (p = 0.011). When carrying out gene expression correlations and global methylation levels, we found that the polymerase genes with traslesion activity (TLS) were found to have moderate correlations for the REV1 (p = 0.049), POLK (p = 0.036), POLQ ( p = 0.001) and PCNA (p = 0.020) where we found that when methylcytosine standards were high when expression was reduced. These results support the importance of the regulation of global DNA methylation as a participant in maintaining the genomic stability of hematopoietic stem cells, promoting a better understanding of the etiology, diagnostic and prognostic stratification of the Myelodysplastic Syndrome, demonstrating that deregulation in this process may be involved in the pathogenesis and evolution of the disease.