PDF(Pubmed)
Abstract：
OBJECTIVE: There have been needs to improve the sensitivity of liquid biopsy. This report aims to report the analytical and clinical validation of a next-generation sequencing (NGS)-based circulating tumor DNA (ctDNA) assay.
METHODS: Analytical validation was conducted in vitro by evaluating the limit of detection (LOD), precision, and specificity for various genomic aberrations. The real-world performance in non-small cell lung cancer (NSCLC) was assessed by comparing the results of AlphaLiquid100 to the tissue-based results.
RESULTS: The LODs with 30 ng input DNA were 0.11%, 0.11%, 0.06%, 0.21%, and 2.13 copies for detecting single nucleotide variants, insertions, deletions, fusions, and copy number alterations (CNA), respectively. Quantitatively, single nucleotide variants/insertions and deletions, fusions, and CNAs showed a good correlation (R2=0.91, 0.40, and 0.65; y=0.95, 1.06, and 1.19) to the manufacturer\'s values, and per-base specificities for all types of variants were near 100%. In real-world NSCLC (n=122), key actionable mutations in NSCLC were detected in 60.7% (74/122) with the ctDNA assay. Comparative analysis against the NGS-based tissue results for all key mutations showed positive percent agreement (PPA) of 85.3%. For individual genes, the PPA was as high as 95.7% for epidermal growth factor receptor (EGFR) mutations and 83.3% for ALK translocations. AlphaLiquid100 detected drug-sensitive EGFR mutation at a variant allele frequency as low as 0.02% and also identified an EGFR mutation in a case where tissue sample missed. Blood samples collected post-targeted therapies revealed additional acquired mutations.
CONCLUSIONS: The AlphaLiquid100 ctDNA assay demonstrates robust analytical validity, offering clinically important information for NSCLC patients.
METHODS: Analytical validation was conducted in vitro by evaluating the limit of detection (LOD), precision, and specificity for various genomic aberrations. The real-world performance in non-small cell lung cancer (NSCLC) was assessed by comparing the results of AlphaLiquid100 to the tissue-based results.
RESULTS: The LODs with 30 ng input DNA were 0.11%, 0.11%, 0.06%, 0.21%, and 2.13 copies for detecting single nucleotide variants, insertions, deletions, fusions, and copy number alterations (CNA), respectively. Quantitatively, single nucleotide variants/insertions and deletions, fusions, and CNAs showed a good correlation (R2=0.91, 0.40, and 0.65; y=0.95, 1.06, and 1.19) to the manufacturer\'s values, and per-base specificities for all types of variants were near 100%. In real-world NSCLC (n=122), key actionable mutations in NSCLC were detected in 60.7% (74/122) with the ctDNA assay. Comparative analysis against the NGS-based tissue results for all key mutations showed positive percent agreement (PPA) of 85.3%. For individual genes, the PPA was as high as 95.7% for epidermal growth factor receptor (EGFR) mutations and 83.3% for ALK translocations. AlphaLiquid100 detected drug-sensitive EGFR mutation at a variant allele frequency as low as 0.02% and also identified an EGFR mutation in a case where tissue sample missed. Blood samples collected post-targeted therapies revealed additional acquired mutations.
CONCLUSIONS: The AlphaLiquid100 ctDNA assay demonstrates robust analytical validity, offering clinically important information for NSCLC patients.
摘要：
■一直需要提高液体活检的灵敏度。本报告旨在报告基于下一代测序（NGS）的循环肿瘤DNA（ctDNA）测定的分析和临床验证。
■通过评估检测限（LOD）在体外进行分析验证，精度,以及对各种基因组畸变的特异性。通过将AlphaLiquid®100的结果与基于组织的结果进行比较来评估非小细胞肺癌(NSCLC)的真实世界性能。
■输入30ngDNA的LOD为0.11％，0.11%，0.06%,0.21%，和2.13个拷贝用于检测SNV，插入,删除,融合,和拷贝数改变(CNA)，分别。定量地,SNV/INDEL,融合,和CNAs显示出良好的相关性（R2=0.91、0.40和0.65；y=0.95、1.06和1.19）与制造商的值，并且所有类型的变体的每碱基特异性接近100%。在真实世界的非小细胞肺癌（n=122），用ctDNA分析检测到NSCLC中的关键可操作突变占60.7%(74/122).针对所有关键突变的基于NGS的组织结果的比较分析显示85.3%的阳性一致性百分比(PPA)。对于单个基因，EGFR突变的PPA高达95.7%,ALK易位的PPA高达83.3%.AlphaLiquid100在低至0.02%的变体等位基因频率下检测到药物敏感性EGFR突变,并且在组织样品缺失的情况下还鉴定出EGFR突变。靶向治疗后收集的血样揭示了额外的获得性突变。
■AlphaLiquid®100ctDNA分析显示出强大的分析有效性，为非小细胞肺癌患者提供临床重要信息。
■通过评估检测限（LOD）在体外进行分析验证，精度,以及对各种基因组畸变的特异性。通过将AlphaLiquid®100的结果与基于组织的结果进行比较来评估非小细胞肺癌(NSCLC)的真实世界性能。
■输入30ngDNA的LOD为0.11％，0.11%，0.06%,0.21%，和2.13个拷贝用于检测SNV，插入,删除,融合,和拷贝数改变(CNA)，分别。定量地,SNV/INDEL,融合,和CNAs显示出良好的相关性（R2=0.91、0.40和0.65；y=0.95、1.06和1.19）与制造商的值，并且所有类型的变体的每碱基特异性接近100%。在真实世界的非小细胞肺癌（n=122），用ctDNA分析检测到NSCLC中的关键可操作突变占60.7%(74/122).针对所有关键突变的基于NGS的组织结果的比较分析显示85.3%的阳性一致性百分比(PPA)。对于单个基因，EGFR突变的PPA高达95.7%,ALK易位的PPA高达83.3%.AlphaLiquid100在低至0.02%的变体等位基因频率下检测到药物敏感性EGFR突变,并且在组织样品缺失的情况下还鉴定出EGFR突变。靶向治疗后收集的血样揭示了额外的获得性突变。
■AlphaLiquid®100ctDNA分析显示出强大的分析有效性，为非小细胞肺癌患者提供临床重要信息。
