FNA smears as a potential source of DNA for targeted next-generation sequencing of lung adenocarcinomas.
FNA涂片可作为肺腺癌靶向二代测序的潜在DNA来源
Abstract
Diff-Quik-stained fine-needle aspiration (FNA) smears and touch preparations from biopsies represent alternative specimens for molecular testing when cell block or biopsy material is insufficient. This study describes the use of these samples for targeted next-generation sequencing (NGS) of primary and metastatic lung adenocarcinoma and reports the DNA quality and success rates of FNA smears versus other specimens from 1 year of clinical use.
A validation set of 10 slides from 9 patients with prior clinical epidermal growth factor receptor (EGFR) Sanger sequencing and KRAS pyrosequencing (5 KRAS-positive/EGFR-negative and 4 KRAS-negative/EGFR-negative) underwent DNA extraction, quality assessment, and targeted NGS. Subsequently, lung adenocarcinoma specimens submitted for NGS solid tumor mutation panel testing in 1 calendar year (60 biopsies, 57 resections, 33 FNA cell blocks, 12 FNA smears, and 10 body fluid cell blocks) were reviewed for specimen adequacy, sequencing success, and DNA quality.
All 10 validation samples met the DNA quality threshold (delta Ct threshold < 8; range, -2.2 to 4.9) and yielded 0.5 to 22 μg of DNA. The KRAS and EGFR mutation status from FNA smears according to NGS was concordant with previous clinical testing for all 10 samples. In the 1-year review, FNA smears were 100% successful, and this suggested a performance equivalent to or better than the performance of established specimen types, including FNA cell blocks. DNA quality according to ΔCt was significantly better with FNA smears versus biopsies, resections, and FNA cell blocks.
FNA smears of lung adenocarcinomas are high-quality alternative specimens for a targeted NGS panel with a high success rate in clinical practice. Cancer Cytopathol 2016;124:406-14. © 2016 American Cancer Society.
A validation set of 10 slides from 9 patients with prior clinical epidermal growth factor receptor (EGFR) Sanger sequencing and KRAS pyrosequencing (5 KRAS-positive/EGFR-negative and 4 KRAS-negative/EGFR-negative) underwent DNA extraction, quality assessment, and targeted NGS. Subsequently, lung adenocarcinoma specimens submitted for NGS solid tumor mutation panel testing in 1 calendar year (60 biopsies, 57 resections, 33 FNA cell blocks, 12 FNA smears, and 10 body fluid cell blocks) were reviewed for specimen adequacy, sequencing success, and DNA quality.
All 10 validation samples met the DNA quality threshold (delta Ct threshold < 8; range, -2.2 to 4.9) and yielded 0.5 to 22 μg of DNA. The KRAS and EGFR mutation status from FNA smears according to NGS was concordant with previous clinical testing for all 10 samples. In the 1-year review, FNA smears were 100% successful, and this suggested a performance equivalent to or better than the performance of established specimen types, including FNA cell blocks. DNA quality according to ΔCt was significantly better with FNA smears versus biopsies, resections, and FNA cell blocks.
FNA smears of lung adenocarcinomas are high-quality alternative specimens for a targeted NGS panel with a high success rate in clinical practice. Cancer Cytopathol 2016;124:406-14. © 2016 American Cancer Society.
摘要
细胞块或活检标本的量不足以进行分子检测时，Diff-Quik-染色的细针穿刺活检(FNA)涂片和活检印片标本或许可以替代。本研究表明这类标本在原发性和转移性肺腺癌靶向二代测序中的应用，并通过一年来的实践将其与其它类型标本进行DNA质量和实验成功率的比较。
研究对象为9名患者的10例切片，先前临床经过表皮生长因子受体(EGFR)Sanger测序和KRAS焦磷酸测序验证(5例KRAS+ /EGFR－；4 例KRAS－/EGFR－)，进行DNA提取、质量评估和靶向二代测序。随后将一年来的肺腺癌标本行NGC实体瘤基因组突变检测，包括60例活检、57例手术标本、33例 FNA细胞块、12例 FNA涂片和10例体液细胞块，并进行了样本量、测序成功率和DNA质量的综合评估。
所有10个已验证的样本均符合DNA样本质量阈值(ΔCt值< 8;范围：-2.2 - 4.9)，DNA提取量为0.5 - 22μg。FNA涂片经NGC技术检测到的KRAS和EGFR突变状态与先前的临床检测一致。1年的病例回顾分析表明，FNA涂片NGC检测100%成功，这表明其性能相当于或优于已明确可用的标本类型，包括FNA细胞块。从ΔCt 值可以看出，FNA涂片的DNA质量显著优于活检、手术标本和FNA细胞块。
在肺腺癌靶向二代测序的临床实践中，FNA涂片标本质量更好、成功率高。
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