Use of uracil-DNA glycosylase enzyme to reduce DNA-related artifacts from formalin-fixed and paraffin-embedded tissues in diagnostic routine
Appl. cancer res
; 39: 1-6, 2019. ilus, tab
Artigo
em Inglês
| LILACS, Inca
| ID: biblio-1006568
Biblioteca responsável:
BR30.1
Localização: BR30.1
ABSTRACT
Background:
Detection of somatic mutations is a mandatory practice for therapeutic definition in precision oncology. However, somatic mutation detection protocols use DNA from formalin-fixed and paraffin-embedded (FFPE) tumor tissues, which can result in detection of nonreproducible sequence artifacts, especially CG > TA transitions, in DNA. In recent studies, DNA pretreatment with uracil DNA glycosylase (UDG), an enzyme involved in base excision repair, significantly reduced the number of DNA artifacts after mutation detection by next-generation sequencing (NGS) and other methods, without affecting the capacity to detect real mutations. This study aimed to evaluate the effects of UDG enzymatic pretreatment in reducing the number of DNA sequencing artifacts from FFPE tumor samples, to improve the accuracy of genetic testing in the molecular diagnostic routine.Methods:
We selected 12 FFPE tumor samples (10 melanoma, 1 lung, and 1 colorectal tumor sample) with different storage times. We compared sequencing results of a 16-hotspot gene panel of NGS libraries prepared with UDG-treated and untreated samples.Results:
All UDG-treated samples showed large reductions in the total number of transitions (medium reduction of 80%) and the transition/transversion ratio (medium reduction of 75%). In addition, most sequence artifacts presented a low variant allele frequency (VAF < 10%) which are eliminated with UDG treatment.Conclusion:
Including UDG enzymatic treatment before multiplex amplification in the NGS workflow significantly decreased the number of artifactual variants detected in FFPE samples. Thus, including this additional step in the current methodology should improve the rate of true mutation detection in the molecular diagnostic routine.
Texto completo:
Disponível
Coleções:
Bases de dados internacionais
Temas:
Geral
Base de dados:
LILACS
/
PrevCan
Assunto principal:
Medição da Dor
/
Inclusão em Parafina
/
Testes Diagnósticos de Rotina
/
Uracila-DNA Glicosidase
/
Sequenciamento Completo do Genoma
Tipo de estudo:
Diagnostic_studies
/
Guia de prática clínica
Limite:
Humanos
Idioma:
Inglês
Revista:
Appl. cancer res
Assunto da revista:
Neoplasias
Ano de publicação:
2019
Tipo de documento:
Artigo
País de afiliação:
Brasil
Instituição/País de afiliação:
A.C.Camargo Cancer Center/BR
/
University of São Paulo/BR
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