Division of Gastrointestinal Oncology, Shizuoka Cancer Center, 1007 Shimonagakubo Nagaizumi-cho Sunto-gun, Shizuoka, 411-8777, Japan.
Drug Discovery and Development Division, Shizuoka Cancer Center Research Institute, 1007 Shimonagakubo Nagaizumi-cho Sunto-gun, Shizuoka, 411-8777, Japan.
BMC Cancer. 2018 Aug 16;18(1):826. doi: 10.1186/s12885-018-4733-7.
Practical and reliable genotyping procedures with a considerable number of samples are required not only for risk-adapted therapeutic strategies, but also for stratifying patients into future clinical trials for molecular-targeting drugs. Recent advances in mutation testing, including next-generation sequencing, have led to the increased use of formalin-fixed paraffin-embedded tissue. We evaluated gene alteration profiles of cancer-related genes in esophageal cancer patients and correlated them with clinicopathological features, such as smoking status and survival outcomes.
Surgically resected formalin-fixed, paraffin-embedded tissue was collected from 135 consecutive patients with esophageal cancer who underwent esophagectomy. Based on the assessment of DNA quality with a quantitative PCR-based assay, uracil DNA glycosylase pretreatment was performed to ensure quality and accuracy of amplicon-based massively parallel sequencing. Amplicon-based massively parallel sequencing was performed using the Illumina TruSeq® Amplicon Cancer Panel. Gene amplification was detected by quantitative PCR-based assay. Protein expression was determined by automated quantitative fluorescent immunohistochemistry.
Data on genetic alterations were available for 126 patients. The median follow-up time was 1570 days. Amplicon-based massively parallel sequencing identified frequent gene alterations in TP53 (66.7%), PIK3CA (13.5%), APC (10.3%), ERBB4 (7.9%), and FBXW7 (7.9%). There was no association between clinicopathological features or prognosis with smoking status. Multivariate analyses revealed that the PIK3CA mutation and clinical T stage were independent favorable prognostic factors (hazard ratio 0.34, 95% confidence interval: 0.12-0.96, p = 0.042). PIK3CA mutations were significantly associated with APC alterations (p = 0.0007) and BRAF mutations (p = 0.0090).
Our study provided profiles of cancer-related genes in Japanese patients with esophageal cancer by next-generation sequencing using surgically resected formalin-fixed, paraffin-embedded tissue, and identified the PIK3CA mutation as a favorable prognosis biomarker.
不仅需要能够对大量样本进行操作且可靠的基因分型程序，还需要将患者分层以便进行未来的分子靶向药物临床试验，以此来制定风险适应的治疗策略。包括下一代测序在内的突变检测技术的进步，导致了对福尔马林固定石蜡包埋组织的更多使用。我们评估了癌症相关基因在食管癌患者中的基因改变谱，并将其与临床病理特征（如吸烟状态和生存结果）相关联。
对 135 例接受食管癌切除术的连续食管癌患者的手术切除福尔马林固定、石蜡包埋组织进行了收集。基于基于定量 PCR 检测的 DNA 质量评估，进行尿嘧啶 DNA 糖基化酶预处理，以确保基于扩增子的大规模平行测序的质量和准确性。使用 Illumina TruSeq®Amplicon Cancer Panel 进行基于扩增子的大规模平行测序。通过基于定量 PCR 的检测方法检测基因扩增，通过自动化定量荧光免疫组织化学检测蛋白质表达。
126 例患者的遗传改变数据可用。中位随访时间为 1570 天。基于扩增子的大规模平行测序确定了 TP53（66.7%）、PIK3CA（13.5%）、APC（10.3%）、ERBB4（7.9%）和 FBXW7（7.9%）的高频基因改变。临床病理特征或预后与吸烟状态之间没有关联。多变量分析显示 PIK3CA 突变和临床 T 期是独立的有利预后因素（危险比 0.34，95%置信区间：0.12-0.96，p=0.042）。PIK3CA 突变与 APC 改变（p=0.0007）和 BRAF 突变（p=0.0090）显著相关。
我们的研究通过使用手术切除的福尔马林固定石蜡包埋组织进行下一代测序，提供了日本食管癌患者癌症相关基因的图谱，并确定了 PIK3CA 突变是有利的预后生物标志物。
