Ann Lab Med.  2019 Nov;39(6):515-523. 10.3343/alm.2019.39.6.515.

Korean Society for Genetic Diagnostics Guidelines for Validation of Next-Generation Sequencing-Based Somatic Variant Detection in Hematologic Malignancies

• Kim H
• Yun JW
• Lee ST
• Kim HJ
• Kim SH
• Kim JW
• The Korean Society for Genetic Diagnostics Clinical Guidelines Committee

• KMID: 2450945
• DOI: http://doi.org/10.3343/alm.2019.39.6.515

Abstract

Next-generation sequencing (NGS) is currently used in the clinical setting for targeted therapies and diagnosis of hematologic malignancies. Accurate detection of somatic variants is challenging because of tumor purity, heterogeneity, and the complexity of genetic alterations, with various issues ranging from high detection design to test implementation. This article presents guidelines developed through consensus among a panel of experts from the Korean Society for Genetic Diagnostics. They are based on experiences with the validation processes of NGS-based somatic panels for hematologic malignancies, with reference to previous international recommendations. These guidelines describe basic parameters with emphasis on the design of a validation protocol for NGS-based somatic panels to be used in practice. In addition, they suggest thresholds of key metrics, including minimum coverage, mean coverage with uniformity index, and minimum variant allele frequency, for the initial diagnosis of hematologic malignancies.

Keyword

Next-generation sequencing; Validation; Somatic variant; Guidelines; Panels; Hematologic malignancies

MeSH Terms

Clothing
Consensus
Diagnosis
Gene Frequency
Hematologic Neoplasms*
Population Characteristics

Figure

• 

  Fig. 1 Overview of validation process for somatic variants in hematologic malignancies using NGS testing.*Samples include patient samples, validated cell lines, and/or commercial controls.Abbreviations: NGS, next-generation sequencing; LoD, limit of detection; PPA, positive percentage agreement; PPV, positive predictive value; AF, allele frequency.

• 

  Fig. 2 An example of Step 1 validation (pilot test).*Pooled samples can comprise one short deletion, one short insertion, and one long insertion in different regions of the patient samples.Abbreviations: AF, allele frequency; RM, reference material; SNV, single-nucleotide variant; indel, insertion and/or deletion.

• 

  Fig. 3 An example of Step 2 validation (formal validation) with 64 samples (including separately bar-coded samples from the same patient/RMs/commercial positive controls). For LoD validation, samples with various VAF were guaranteed by dilution with RM.*Pooled samples can be comprised of one short deletion, one short insertion, and one long insertion in different regions of the patient samples.Abbreviations: indels, insertions and/or deletions; RM, reference material; VAF, variant allele frequency; SNV, single nucleotide variant; LoD, limit of detection.

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