Whole exome sequencing for determination of tumor mutation load in liquid biopsy from advanced cancer patients

Tumor mutation load (TML) has been proposed as a biomarker of patient response to immunotherapy in several studies. TML is usually determined by tumor biopsy DNA (tDNA) whole exome sequencing (WES), therefore TML evaluation is limited by informative biopsy availability. Circulating cell free DNA (cfDNA) provided by liquid biopsy is a surrogate specimen to biopsy for molecular profiling. Nevertheless performing WES on DNA from plasma is technically challenging and the ability to determine tumor mutation load from liquid biopsies remains to be demonstrated. In the current study, WES was performed on cfDNA from 32 metastatic patients of various cancer types included into MOSCATO 01 (NCT01566019) and/or MATCHR (NCT02517892) molecular triage trials. Results from targeted gene sequencing (TGS) and WES performed on cfDNA were compared to results from tumor tissue biopsy. In cfDNA samples, WES mutation detection sensitivity was 92% compared to targeted sequencing (TGS). When comparing cfDNA-WES to tDNA-WES, mutation detection sensitivity was 53%, consistent with previously published prospective study comparing cfDNA-TGS to tDNA-TGS. For samples in which presence of tumor DNA was confirmed in cfDNA, tumor mutation load from liquid biopsy was correlated with tumor biopsy. Taken together, this study demonstrated that liquid biopsy may be applied to determine tumor mutation load. Qualification of liquid biopsy for interpretation is a crucial point to use cfDNA for mutational load estimation.

多项研究已将肿瘤突变负荷（Tumor Mutation Load, TML）提出作为评估患者免疫治疗响应的生物标志物。通常，TML需通过肿瘤活检DNA（tumor biopsy DNA, tDNA）的全外显子组测序（Whole Exome Sequencing, WES）进行测定，因此其评估受到活检样本可及性的限制。液体活检所提供的循环游离DNA（Circulating Cell Free DNA, cfDNA）可作为活检样本的替代标本用于分子分型。然而，对血浆来源DNA开展WES在技术上仍存在挑战，通过液体活检测定肿瘤突变负荷的可行性仍有待验证。本研究对纳入MOSCATO 01（NCT01566019）和/或MATCHR（NCT02517892）分子分诊试验的32例不同癌症类型转移性患者的cfDNA开展WES检测。将cfDNA的靶向基因测序（Targeted Gene Sequencing, TGS）结果与WES结果，一同与肿瘤组织活检的检测结果进行对比。在cfDNA样本中，与靶向测序（TGS）相比，WES的突变检测灵敏度达92%。将cfDNA-WES与tDNA-WES进行对比时，突变检测灵敏度为53%，这与此前已发表的对比cfDNA-TGS与tDNA-TGS的前瞻性研究结果一致。对于经确认存在肿瘤DNA的cfDNA样本，液体活检所得的肿瘤突变负荷与肿瘤活检结果具有相关性。综上，本研究证实液体活检可用于测定肿瘤突变负荷。对液体活检的合格性进行验证，是利用cfDNA进行突变负荷估算的关键要点。