ABSTRACT
Single nucleotide polymorphisms (SNPs, rs12979860 e rs8099917) in the Interferon Lambda 4 gene (IFNL4, formerly IFNL3and/or IL28B) has been associated with failure in the innate immune response, sustained virological response in hepatitis C, and HTLV-1-associated myelopathy (HAM) development. To search for these polymorphisms several methodologies can be employed, such as sequencing, real-time or quantitative polymerase chain reaction (qPCR), restriction fragment length polymorphism analysis in PCR products (PCR-RFLP), and tetra-primer PCR. The present study compared the performance of the tetra-primer PCR in relation to the PCR-RFLP, both optimized in the Research HTLV Laboratory of the Center of Immunology of Instituto Adolfo Lutz in São Paulo. One hundred DNA samples obtained from patients of STD/Aids Reference Centre in São Paulo, previously analyzed for IL28B SNPs by PCR-RFLP were selected for analysis, after confirming that they represent all IL28B SNPs patterns described in the literature. The results obtained showed concordance between the PCR-RFLP and the tetra-primer PCR SNPs results, and because of the low cost, easy to perform, and minor employment of biological specimen and reagents, the tetra-primer PCR is of choice to be used in routine. (AU)
Polimorfismos de nucleotídeos únicos (single nucleotide polymorphisms, SNPs rs12979860 e rs8099917) no gene que codifica o Interferon Lambda 4 (IFNL4, antigamente IFNL3 e/ou IL28B) têm sido associados às falhas na resposta imune inata e resposta virológica sustentada na hepatite C, e a mielopatia associada ao HTLV-1 (HTLV-1-associated myelopathy, HAM). A pesquisa destes polimorfismos pode empregar diversas metodologias: sequenciamento, reação em cadeia da polimerase em tempo real ou quantitativa (quantitative polymerase chain reaction, qPCR), análise de fragmentos de restrição enzimática em produtos de PCR (restriction fragment length polymorphism in PCR products, PCR-RFLP) e a tetra-primer PCR. Este estudo comparou o desempenho da tetra-primer PCR em relação a PCR-RFLP, ambas otimizadas no Laboratório de Pesquisa em HTLV do Centro de Imunologia do Instituto Adolfo Lutz de São Paulo. Foram selecionadas 100 amostras de DNA obtidas de pacientes do Centro de Referência e Treinamento em DST/Aids de São Paulo cujos SNPs na IL28B foram anteriormente determinados por PCR-RFLP e representaram todos os perfis descritos em literatura. Os resultados obtidos mostraram concordância entre elas, e pelo fato da tetra-primer PCR ter menor custo, ser de fácil execução, empregar menos tempo, insumos e material biológico, é a técnica de escolha para uso em rotina. (AU)
Subject(s)
Polymorphism, Restriction Fragment Length , Polymerase Chain Reaction , Interleukins , Polymorphism, Single Nucleotide , Interferon LambdaABSTRACT
Hepatocellular carcinoma (HCC) is a malignancy with a leading lethality. The etiology is quite diverse, ranging from viral infections to metabolic disorders or intoxications, and associates with specific somatic mutational patterns and specific host immunological phenotypes. Particularly, hepatitis C virus (HCV)-infected liver is featured by an activation of interferon (IFN)-stimulated genes (ISGs; IFN signature), which we suppose is driven by type III IFNL4. Taking advantage of the TCGA collection of HCC patients of various different etiologies, this study aimed at validating our previous findings on hepatic IFNL4 gene activation in HCV infection in an independent and larger cohort of patients with advanced liver disease. In a cohort of n = 377 cases, the entirety of the sequencing data was used to assess the IFNL genotypes, and the cases were stratified for etiology. The number of IFNL4 transcripts within nonmalignant and malignant tissues was found to be more abundant in patients with HCV or HCV/HBV infections when compared to other risk factors. Moreover, in patients with HCV infection as a risk factor, a close, positive relationship was found between ISG activation and the number of functional IFNL4 transcripts. Data on this independent TCGA sample support the concept of an IFNL4-dependent HCV-driven activation of hepatic ISGs. In addition to that, they add to the understanding of etiology-related host immunological phenotypes in HCC.
Subject(s)
Biomarkers, Tumor/metabolism , Carcinoma, Hepatocellular/pathology , Gene Expression Regulation, Neoplastic , Hepatitis C, Chronic/complications , Interleukins/metabolism , Liver Neoplasms/pathology , Apoptosis , Biomarkers, Tumor/genetics , Carcinoma, Hepatocellular/etiology , Carcinoma, Hepatocellular/metabolism , Case-Control Studies , Cell Proliferation , Female , Hepacivirus/isolation & purification , Hepatitis C, Chronic/virology , Humans , Interleukins/genetics , Liver Neoplasms/etiology , Liver Neoplasms/metabolism , Male , Middle Aged , Tumor Cells, CulturedABSTRACT
BACKGROUND: Hepatocellular carcinoma (HCC) and pancreatic ductal adenocarcinoma (PDAC) are malignancies with a leading lethality. With reference to interferons (IFNs) known to mediate antitumor activities, this study investigated the relationship between germline genetic variations in type III IFN genes and cancer disease progression from The Cancer Genome Atlas (TCGA) data. The genetic variations under study tag a gain-or-loss-of-function dinucleotide polymorphism within the IFNL4 gene, rs368234815 [TT/ΔG]. METHODS: The entirety of the TCGA sequencing data was used to assess genotypes of 187 patients with HCC and of 162 patients with PDAC matched for ethnicity. Stratified for IFNL genotypes, both cohorts were subjected to time-to-event analyses according to Kaplan-Meier with regard to the length of the specific progression free interval (PFI) and the overall survival (OS) time as two clinical endpoints for disease progression. RESULTS: Logrank analysis revealed a significant relationship between IFNL genotypes and disease outcome for PDAC. This relationship was not found for HCC. A multiple Cox regression analysis employing patients' age, tumor grade and tumor stage as further covariates proved IFNL genotypes to be independent predictors for PDAC disease outcome. CONCLUSION: This repository-based approach unveiled clinical evidence suggestive for an impact of IFNL germline variations for PDAC progression with an IFNL haplotype predisposing for IFNL4 expression being favorable.
