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Quantitative evaluation of MSI testing using NGS detects the imperceptible microsatellite changed caused by MSH6 deficiency

Familial Cancer volume 20, pages 137–143 (2021)Cite this article

Abstract

Microsatellite instability (MSI) is an effective biomarker for diagnosing Lynch syndrome (LS) and predicting the responsiveness of cancer therapy. MSI testing is conventionally performed by capillary electrophoresis, and MSI status is judged by visual assessment of allele size change. Here, we attempted to develop a quantitative evaluation model of MSI using next-generation sequencing (NGS). Microsatellite markers were analyzed in tumor and non-tumor tissues of colorectal cancer patients by NGS after a single multiplex polymerase chain reaction amplification. The read counts corresponding to microsatellite loci lengths were calculated independently of mapping against a reference genome, and their distribution was digitized by weighted mean. Weighted mean differences between tumor and non-tumor samples with different MSI status were assessed, and cut-off values for each marker in the discovery cohort were determined. Each microsatellite maker was defined as unstable if the weighted mean difference was greater than the cut-off value. In the discovery cohort, the evaluation model demonstrated sensitivity and specificity of 100% for all markers. In the validation cohort, MSI status determined by the new model was consistent with the outcome of the conventional method in 29/30 cases (97%). The single inconsistent case was classified as low-frequency MSI by the conventional method but considered MSI-high by NGS. Genetic testing for mismatch repair genes revealed a pathogenic variant in MSH6 in the discordant case. We successfully developed a quantitative evaluation method for determining MSI status using NGS. This is a robust and sensitive method and could improve LS diagnosis.

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Data availability

The datasets generated during and/or analyzed during the current study are available from the corresponding author on reasonable request.

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Acknowledgements

We thank Edanz Group (https://en-author-services.edanzgroup.com/) for editing a draft of this manuscript.

Funding

This research was supported by Japan Agency for Medical Research and Development (AMED) under Grant JP18kk0205004, JSPS KAKENHI Grant Number JP18K07339 and National Cancer Center Research and Development Fund Grant Number 31-A-2.

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Author notes

  1. Gou Yamamoto and Takashi Takenoya have contributed equally to this work.

Affiliations

  1. Department of Molecular Diagnosis and Cancer Prevention, Saitama Cancer Center, 780 Komuro, Ina-machi, Kitaadachi-gun, Saitama, 362-0806, Japan

    Gou Yamamoto, Takashi Takenoya, Akemi Takahashi, Yukiko Osanai, Miho Kakuta, Tetsuhiko Tachikawa & Kiwamu Akagi

  2. Bioinformatics Department, Riken Genesis Co., Ltd., 1-2-2 Osaki, Shinagawa-ku, Tokyo, 141-0032, Japan

    Takanori Washio & Masato Kamiyama

  3. Information Technology Center, Saitama Medical University, 38 Moro-hongo, Moroyama-machi, Iruma-gun, Saitama, 350-0495, Japan

    Michio Shiibashi

  4. Department of Gastroenterological Surgery, Saitama Medical University International Medical Center, 1397-1 Yamane, Hidaka-shi, Saitama, 350-1241, Japan

    Shigeki Yamaguchi

Authors
  1. Gou Yamamoto
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  2. Takashi Takenoya
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  3. Akemi Takahashi
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  4. Yukiko Osanai
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  5. Miho Kakuta
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  6. Tetsuhiko Tachikawa
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  7. Takanori Washio
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  8. Masato Kamiyama
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  9. Michio Shiibashi
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  10. Shigeki Yamaguchi
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  11. Kiwamu Akagi
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Contributions

Conceptualization: GY, TT, KA. Methodology: GY, TT, TW, MK, MS, KA. Formal analysis and investigation: GY, TT, AT, YO, MK, TT, TW, MK, MS. Writing—original draft: GY, TT, KA. Writing—review & editing: GY, MS, SY, KA. Funding acquisition: GY, KA. Resources: GY, KA. Supervision: SY, KA.

Corresponding author

Correspondence to Kiwamu Akagi.

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Conflicts of interest

The authors declare that they have no conflict of interest.

Ethical approval

All procedures were performed in accordance with the ethical standards of the responsible committee on human experimentation and with the 1964 Helsinki Declaration, as revised in 2013, as well as the Japanese ethical guidelines for human genome/gene analysis research. This study was approved by the Institutional Review Boards of Saitama Cancer Center (No. 476 and 729). Written consent was obtained from the patient before inclusion in the study.

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Yamamoto, G., Takenoya, T., Takahashi, A. et al. Quantitative evaluation of MSI testing using NGS detects the imperceptible microsatellite changed caused by MSH6 deficiency. Familial Cancer 20, 137–143 (2021). https://doi.org/10.1007/s10689-020-00203-3

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  • DOI: https://doi.org/10.1007/s10689-020-00203-3

Keywords

  • Microsatellite instability
  • Lynch syndrome
  • Next generation sequencing
  • MSH6
  • Colorectal cancer