RESUMO
PURPOSE: There is a quest for novel noninvasive diagnostic markers for the detection of breast cancer. The goal of this study is to identify circulating microRNA (miRNA) signatures using a cohort of Asian Chinese patients with breast cancer, and to compare miRNA profiles between tumor and serum samples. EXPERIMENTAL DESIGN: miRNA from paired breast cancer tumors, normal tissue, and serum samples derived from 32 patients were comprehensively profiled using microarrays or locked nucleic acid real-time PCR panels. Serum samples from healthy individuals (n = 22) were also used as normal controls. Significant serum miRNAs, identified by logistic regression, were validated in an independent set of serum samples from patients (n = 132) and healthy controls (n = 101). RESULTS: The 20 most significant miRNAs differentially expressed in breast cancer tumors included miRNA (miR)-21, miR-10b, and miR-145, previously shown to be dysregulated in breast cancer. Only 7 miRNAs were overexpressed in both tumors and serum, suggesting that miRNAs may be released into the serum selectively. Interestingly, 16 of the 20 most significant miRNAs differentially expressed in serum samples were novel. MiR-1, miR-92a, miR-133a, and miR-133b were identified as the most important diagnostic markers, and were successfully validated; receiver operating characteristic curves derived from combinations of these miRNAs exhibited areas under the curves of 0.90 to 0.91. CONCLUSION: The clinical use of miRNA signatures as a noninvasive diagnostic strategy is promising, but should be further validated for different subtypes of breast cancers.
Assuntos
Neoplasias da Mama/sangue , Neoplasias da Mama/genética , MicroRNAs/genética , Transcriptoma , Adulto , Idoso , Neoplasias da Mama/diagnóstico , Estudos de Casos e Controles , Análise por Conglomerados , Feminino , Perfilação da Expressão Gênica , Regulação Neoplásica da Expressão Gênica , Humanos , MicroRNAs/sangue , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Razão de Chances , Curva ROC , Receptor ErbB-2/metabolismo , Receptores de Estrogênio/metabolismo , Reprodutibilidade dos TestesRESUMO
In a clinical setting, next-generation sequencing (NGS) approaches for the enrichment and resequencing of DNA targets may have limitations in throughput, cost, or accuracy. We evaluated an NGS workflow for targeted DNA sequencing for mutation detection. Targeted sequence data of the BRCA1 and BRCA2 genes, generated using a PCR-based, multiplexed NGS approach using the SOLiD 4 (n = 24) and Ion Torrent PGM (n = 20) next-generation sequencers, were evaluated against sequence data obtained by Sanger sequencing. The overall sensitivity for SOLiD and PGM were 97.8% (95% CI = 94.7 to 100.0) and 98.9% (95% CI = 96.8 to 100.0) respectively. The specificity for the SOLiD platform was high, at 100.0% (95% CI = 99.3 to 100.0). PGM correctly identified all 3 indels, but 68 false-positive indels were also called. Equimolar normalization of amplicons was not necessary for successful NGS. Both platforms are highly amenable to scale-up, potentially reducing the reagent cost for BRCA testing to Assuntos
Proteína BRCA2/genética
, Análise Mutacional de DNA/métodos
, Sequenciamento de Nucleotídeos em Larga Escala/métodos
, Mutação
, Análise Mutacional de DNA/economia
, Genes BRCA2
, Sequenciamento de Nucleotídeos em Larga Escala/economia
, Humanos
, Sensibilidade e Especificidade