Avaliação e padronização de metodologias para análise da translocação cromossômica t(11;18)(q21;q21) em portadores de linfoma MALT gástrico

Abstract

Lymphomas are a heterogeneous group of malignancies that originate in the cells of the lymphatic system, divided into two major groups: Hodgkin's and non-Hodgkin lymphomas (NHL). Considering the place of origin, it can be classified as nodal or extranodal, being the extranodal originated in any organ containing the mucosa-associated lymphoid tissue (MALT), also known as marginal zone lymphoma of B cell type. The NHLs extranodal comprise up to 1/3 of lymphomas in general, the gastrointestinal tract is the site of highest incidence being the stomach responsible for 60% of cases. Gastric lymphomas comprise 2 to 8% of gastric tumors and are the most frequent after adenocarcinoma (90 to 95%). The gastric mucosa has no lymphoid tissue, but studies have shown that infection by Helicobacter pylori (H. pylori) stimulates the development of lymphoid aggregates in 27.4 to 100% of infected individuals, which may lead to gastric MALT lymphoma (GML). GML patients present H. pylori in 85 to 92% of cases. In vitro studies demonstrated the GML low-grade B-cell proliferation by stimulation of specific strains of H. pylori. Studies correlating this infection with GML showed that eradication of the microorganism induces complete remission in 70 to 80% of premature lesions of GML, however, approximately 25% of these cases do not respond to eradication of H. pylori. The chromosomal translocation t(11;18)(q21,q21), found in a significant percentage of low-grade GML, demonstrates conditional independence of lymphoma cells to antigenic stimulus, with consequent non-regression of lymphoma after eradication of H. pylori. This translocation gene represents the most common rearrangement associated with the GML. This study aimed to implement methodologies for detection and analysis of the t(11;18) in patients with GML from techniques of reverse transcription - polymerase chain reaction (RT-PCR)and fluorescent in situ hybridization (FISH). The fresh tissue samples were collected and stored in RNAlater. The RNA was extracted with Trizol, and treated with deoxyribonuclease prior to synthesis of the complementary deoxyribonucleic acid (cDNA) by reverse transcription and PCR using specific primers for API2-MALT1. Electrophoresis was performed in 2% agarose gel. The translocation was identified in 50% (4/8) of cases, other 3/8 revealed the absence of the rearrangement and clinical complete remission of GML after the microorganism eradication, consistent with the finding and the studies described over the work; one case was identified as high-grade also not carrying the mutation. FISH was performed for paraffin samples using API2-MALT1 probe, but it was not possible to complete standardization. The results obtained so far indicate that the fluorescent probe is hybridizing other cell structures than the area of interest and further studies should be undertaken. Despite the small sample, it was possible to implement a methodology for routine identification of this mutation. The results indicated that t(11,18) might be related to the independence of the GML to the eradication of H. pylori.Additional studies with a larger number of patients and characterization by sequencing of the fusion transcripts found will contribute to better understanding of each type of transcript and elucidation regarding its pathogenicity. The molecular analysis is essential for identification and differentiation of this genetic rearrangement to intervene in early therapeutic way and benefit the Brazilian population that lacks GML molecular studies.