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Evaluating the analytical validity of circulating tumor DNA sequencing assays for precision oncology.

ABSTRACT: Circulating tumor DNA (ctDNA) sequencing is being rapidly adopted in precision oncology, but the accuracy, sensitivity and reproducibility of ctDNA assays is poorly understood. Here we report the findings of a multi-site, cross-platform evaluation of the analytical performance of five industry-leading ctDNA assays. We evaluated each stage of the ctDNA sequencing workflow with simulations, synthetic DNA spike-in experiments and proficiency testing on standardized, cell-line-derived reference samples. Above 0.5% variant allele frequency, ctDNA mutations were detected with high sensitivity, precision and reproducibility by all five assays, whereas, below this limit, detection became unreliable and varied widely between assays, especially when input material was limited. Missed mutations (false negatives) were more common than erroneous candidates (false positives), indicating that the reliable sampling of rare ctDNA fragments is the key challenge for ctDNA assays. This comprehensive evaluation of the analytical performance of ctDNA assays serves to inform best practice guidelines and provides a resource for precision oncology.

SUBMITTER: Deveson IW

PROVIDER: S-EPMC8434938 | biostudies-literature | 2021 Sep

REPOSITORIES: biostudies-literature

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Evaluating the analytical validity of circulating tumor DNA sequencing assays for precision oncology.

Deveson Ira W IW Gong Binsheng B Lai Kevin K LoCoco Jennifer S JS Richmond Todd A TA Schageman Jeoffrey J Zhang Zhihong Z Novoradovskaya Natalia N Willey James C JC Jones Wendell W Kusko Rebecca R Chen Guangchun G Madala Bindu Swapna BS Blackburn James J Stevanovski Igor I Bhandari Ambica A Close Devin D Conroy Jeffrey J Hubank Michael M Marella Narasimha N Mieczkowski Piotr A PA Qiu Fujun F Sebra Robert R Stetson Daniel D Sun Lihyun L Szankasi Philippe P Tan Haowen H Tang Lin-Ya LY Arib Hanane H Best Hunter H Burgher Blake B Bushel Pierre R PR Casey Fergal F Cawley Simon S Chang Chia-Jung CJ Choi Jonathan J Dinis Jorge J Duncan Daniel D Eterovic Agda Karina AK Feng Liang L Ghosal Abhisek A Giorda Kristina K Glenn Sean S Happe Scott S Haseley Nathan N Horvath Kyle K Hung Li-Yuan LY Jarosz Mirna M Kushwaha Garima G Li Dan D Li Quan-Zhen QZ Li Zhiguang Z Liu Liang-Chun LC Liu Zhichao Z Ma Charles C Mason Christopher E CE Megherbi Dalila B DB Morrison Tom T Pabón-Peña Carlos C Pirooznia Mehdi M Proszek Paula Z PZ Raymond Amelia A Rindler Paul P Ringler Rebecca R Scherer Andreas A Shaknovich Rita R Shi Tieliu T Smith Melissa M Song Ping P Strahl Maya M Thodima Venkat J VJ Tom Nikola N Verma Suman S Wang Jiashi J Wu Leihong L Xiao Wenzhong W Xu Chang C Yang Mary M Zhang Guangliang G Zhang Sa S Zhang Yilin Y Shi Leming L Tong Weida W Johann Donald J DJ Mercer Timothy R TR Xu Joshua J

Nature biotechnology 20210412 9

Circulating tumor DNA (ctDNA) sequencing is being rapidly adopted in precision oncology, but the accuracy, sensitivity and reproducibility of ctDNA assays is poorly understood. Here we report the findings of a multi-site, cross-platform evaluation of the analytical performance of five industry-leading ctDNA assays. We evaluated each stage of the ctDNA sequencing workflow with simulations, synthetic DNA spike-in experiments and proficiency testing on standardized, cell-line-derived reference samp ...[more]

PMID: 33846644

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Project description:BackgroundWith the accelerated development of next-generation sequencing (NGS), identified variants, and targeted therapies, clinicians who confront the complicated and multifarious genetic information may not effectively incorporate NGS-based circulating tumor DNA (ctDNA) analysis into routine patient care. Consequently, standardized ctDNA testing reports are of vital importance. In an effort to guarantee high-quality reporting performance, we conducted an investigation of the current detection and reporting practices for NGS-based ctDNA analysis.Materials and methodsA set of simulated ctDNA samples with known variants at known allelic frequencies and a corresponding case scenario were distributed to 66 genetic testing laboratories for ctDNA analysis. Written reports were collected to evaluate the detection accuracy, reporting integrity, and information sufficiency using 21 predefined criteria.ResultsCurrent reporting practices for NGS-based ctDNA analysis were found to be far from satisfactory, especially regarding testing interpretation and methodological details. Only 42.4% of laboratories reported the results in complete concordance with the expected results. Moreover, 74.2% of reports only listed aberrations with direct and well-known treatment consequences for the tumor type in question. Genetic aberrations for which experimental agents and/or drug access programs are available may thus be overlooked. Furthermore, methodological details for the interpretation of results were missing from the majority of reports (87.9%).ConclusionThis proof-of-principle study suggests that the capacity for accurate identification of variants, rational interpretation of genotypes, comprehensive recommendation of potential medications, and detailed description of methodologies need to be further improved before ctDNA analysis can be formally implemented in the clinic.Implications for practiceAccurate, comprehensive, and standardized clinical sequencing reports can help to translate complex genetic information into patient-centered clinical decisions, thereby shepherding precision oncology into daily practice. However, standards, guidelines, and quality requirements for clinical reports of next-generation sequencing (NGS)-based circulating tumor DNA (ctDNA) analysis are currently absent. By using a set of simulated clinical ctDNA samples and a corresponding case scenario, current practices were evaluated to identify deficiencies in clinical sequencing reports of ctDNA analysis. The recommendations provided here may serve as a roadmap for the improved implementation of NGS-based ctDNA analysis in the clinic.

Dataset Information

Evaluating the analytical validity of circulating tumor DNA sequencing assays for precision oncology.

Publications

Evaluating the analytical validity of circulating tumor DNA sequencing assays for precision oncology.

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