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Benchmarking bioinformatics approaches for tumour mutational burden evaluation from a large cancer panel against whole-exome sequencing
  1. Jiuhong Pang,
  2. Hongai Xia,
  3. Shijun Mi,
  4. Wen Zhang,
  5. Danielle Pendrick,
  6. Christopher Freeman,
  7. Helen Fernandes,
  8. Mahesh Mansukhani,
  9. Susan J Hsiao
  1. Pathology and Cell Biology, Columbia University Irving Medical Center, New York City, New York, USA
  1. Correspondence to Dr Susan J Hsiao, Department of Pathology and Cell Biology, Columbia University Medical Center, New York, NY 10032, USA; sjh2155{at}


Tumour mutational burden (TMB) is used to predict response to immunotherapies. Although several groups have proposed calculation methods for TMB, a clear consensus has not yet emerged. In this study, we explored TMB calculation approaches with a 586-gene cancer panel (1.75 Mb) benchmarked to TMB measured by whole-exome sequencing (WES), using 30 samples across a range of tumour types. We explored variant allelic fraction (VAF) cut-offs of 5% and 10%, population database filtering at 0.001, 0.0001 and 0.000025, as well as different combinations of synonymous, insertion/deletion and intronic (splice site) variants, as well as exclusion of hotspot mutations, and examined the effect on TMB correlation. Good correlation (Spearman, range 0.66–0.78) between WES and panel TMB was seen across all methods evaluated. Each method of TMB calculation evaluated showed good positive per cent agreement and negative per cent agreement using 10 mutations/Mb as a cut-off, suggesting that multiple TMB calculation approaches may yield comparable results.

  • Biomarkers, Tumor
  • Pathology, Molecular
  • Diagnostic Techniques and Procedures

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  • Handling editor Runjan Chetty.

  • Contributors JP—conceptualisation, methodology, formal analysis and investigation. HX, SM and WZ—conceptualisation and investigation. DP and CF—conceptualisation, methodology and project administration. HF and MM—conceptualisation, methodology and writing (review and editing). SJH—conceptualisation, methodology, investigation and writing (original draft).

  • Funding This study was funded by Bristol Myers Squibb.

  • Competing interests SJH has received consulting fees from Opentrons Labworks and Loxo Oncology and honoraria from Illumina and Medscape.

  • Provenance and peer review Not commissioned; externally peer reviewed.

  • Supplemental material This content has been supplied by the author(s). It has not been vetted by BMJ Publishing Group Limited (BMJ) and may not have been peer-reviewed. Any opinions or recommendations discussed are solely those of the author(s) and are not endorsed by BMJ. BMJ disclaims all liability and responsibility arising from any reliance placed on the content. Where the content includes any translated material, BMJ does not warrant the accuracy and reliability of the translations (including but not limited to local regulations, clinical guidelines, terminology, drug names and drug dosages), and is not responsible for any error and/or omissions arising from translation and adaptation or otherwise.