Activating rearranged during transfection (RET) proto-oncogene alterations can be identified using next-generation sequencing (NGS) of tumor DNA/RNA. We assessed factors associated with NGS (Oncomine Dx Target Test [ODxTT]) success for resected thyroid cancer (TC) specimens, including sample age, processing conditions, and DNA/RNA quality. TC samples were from three Japanese hospitals, with sample age <1–<10 years, fixative 10%/15% neutralized buffered formalin (NBF), and fixation time ≤48 h/>48 h–≤72 h. NGS success rate was defined as the percentage of samples returning validated NGS results (RET fusion-positive/negative [RNA] or RET mutation-positive/negative [DNA], detected using ODxTT). DNA/RNA quality was assessed with indexes based on electrophoresis (DNA/RNA integrity number, DV₂₀₀) and quantitative polymerase chain reaction (DNA/RNA integrity score [ddCq/ΔCq]). NGS success rate (N = 202) was 90%/93% (DNA/RNA) overall, 98%–100% (DNA and RNA) for samples <3 years old, and 91% (DNA and RNA) for samples ≥3–<5 years old fixed in 10% NBF for ≤48 h. Multivariate logistic regression analysis identified ddCq and ΔCq as significant predictors of DNA and RNA NGS success rates, respectively. Quality assessment of nucleic acid extracted from archival tissue samples is important for achieving high NGS success rates in clinical practice, especially for samples ≥3 years old.

中文翻译：

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甲状腺癌切除组织样本质量对二代测序检测的影响

使用肿瘤 DNA/RNA 的下一代测序 (NGS) 可以识别转染期间激活重排 (RET) 原癌基因的改变。我们评估了与切除的甲状腺癌 (TC) 标本的 NGS（Oncomine Dx 目标测试 [ODxTT]）成功相关的因素，包括样本年龄、处理条件和 DNA/RNA 质量。 TC样本来自日本3家医院，样本年龄<1-<10年，固定液10%/15%中和缓冲福尔马林（NBF），固定时间≤48 h/>48 h-≤72 h。 NGS 成功率定义为返回经过验证的 NGS 结果的样本百分比（RET 融合阳性/阴性 [RNA] 或 RET 突变阳性/阴性 [DNA]，使用 ODxTT 检测）。 DNA/RNA质量通过基于电泳（DNA/RNA完整性值，DV ₂₀₀）和定量聚合酶链反应（DNA/RNA完整性得分[ddCq/ΔCq]）的指标进行评估。 NGS 成功率 ( N  = 202) 总体为 90%/93% (DNA/RNA)，<3 岁样本为 98%–100%（DNA 和 RNA），≥3 岁样本为 91%（DNA 和 RNA） –<5岁儿童在10% NBF中固定≤48小时。多变量逻辑回归分析确定 ddCq 和 ΔCq 分别是 DNA 和 RNA NGS 成功率的重要预测因子。从档案组织样本中提取的核酸的质量评估对于在临床实践中实现高 NGS 成功率非常重要，特别是对于 3 年以上的样本。