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Impact of functional studies on exome sequence variant interpretation in early-onset cardiac conduction system diseases

(2020) CARDIOVASCULAR RESEARCH. 116(13). p.2116-2130
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Abstract
Aims The genetic cause of cardiac conduction system disease (CCSD) has not been fully elucidated. Whole-exome sequencing (WES) can detect various genetic variants; however, the identification of pathogenic variants remains a challenge. We aimed to identify pathogenic or likely pathogenic variants in CCSD patients by using WES and 2015 American College of Medical Genetics and Genomics (ACMG) standards and guidelines as well as evaluating the usefulness of functional studies for determining them. Methods and Results We performed WES of 23 probands diagnosed with early-onset (<65 years) CCSD and analyzed 117 genes linked to arrhythmogenic diseases or cardiomyopathies. We focused on rare variants (minor allele frequency < 0.1%) that were absent from population databases. Five probands had protein truncating variants in EMD and LMNA which were classified as “pathogenic” by 2015 ACMG standards and guidelines. To evaluate the functional changes brought about by these variants, we generated a knock-out zebrafish with CRISPR-mediated insertions or deletions of the EMD or LMNA homologs in zebrafish. The mean heart rate and conduction velocities in the CRISPR/Cas9-injected embryos and F2 generation embryos with homozygous deletions were significantly decreased. Twenty-one variants of uncertain significance were identified in 11 probands. Cellular electrophysiological study and in vivo zebrafish cardiac assay showed that 2 variants in KCNH2 and SCN5A, 4 variants in SCN10A, and 1 variant in MYH6 damaged each gene, which resulted in the change of the clinical significance of them from “Uncertain significance” to “Likely pathogenic” in 6 probands. Conclusions Of 23 CCSD probands, we successfully identified pathogenic or likely pathogenic variants in 11 probands (48%). Functional analyses of a cellular electrophysiological study and in vivo zebrafish cardiac assay might be useful for determining the pathogenicity of rare variants in patients with CCSD. SCN10A may be one of the major genes responsible for CCSD. Translational Perspective Whole-exome sequencing (WES) may be helpful in determining the causes of cardiac conduction system disease (CCSD), however, the identification of pathogenic variants remains a challenge. We performed WES of 23 probands diagnosed with early-onset CCSD, and identified 12 pathogenic or likely pathogenic variants in 11 of these probands (48%) according to the 2015 ACMG standards and guidelines. In this context, functional analyses of a cellular electrophysiological study and in vivo zebrafish cardiac assay might be useful for determining the pathogenicity of rare variants, and SCN10A may be one of the major development factors in CCSD.
Keywords
Cardiac conduction-system disease, Whole exome sequencing, 2015 ACMG standards and guidelines, CRISPR/Cas9-mediated gene knock-out in zebrafish, Cellular electrophysiological study, LONG QT SYNDROME, HYPERTROPHIC CARDIOMYOPATHY, MUTATION, CONSEQUENCES, ASSAY, GENE

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Citation

Please use this url to cite or link to this publication:

MLA
Hayashi, Kenshi, et al. “Impact of Functional Studies on Exome Sequence Variant Interpretation in Early-Onset Cardiac Conduction System Diseases.” CARDIOVASCULAR RESEARCH, vol. 116, no. 13, 2020, pp. 2116–30, doi:10.1093/cvr/cvaa010.
APA
Hayashi, K., Teramoto, R., Nomura, A., Asano, Y., Beerens, M., Kurata, Y., … Takamura, M. (2020). Impact of functional studies on exome sequence variant interpretation in early-onset cardiac conduction system diseases. CARDIOVASCULAR RESEARCH, 116(13), 2116–2130. https://doi.org/10.1093/cvr/cvaa010
Chicago author-date
Hayashi, Kenshi, Ryota Teramoto, Akihiro Nomura, Yoshihiro Asano, Manu Beerens, Yasutaka Kurata, Isao Kobayashi, et al. 2020. “Impact of Functional Studies on Exome Sequence Variant Interpretation in Early-Onset Cardiac Conduction System Diseases.” CARDIOVASCULAR RESEARCH 116 (13): 2116–30. https://doi.org/10.1093/cvr/cvaa010.
Chicago author-date (all authors)
Hayashi, Kenshi, Ryota Teramoto, Akihiro Nomura, Yoshihiro Asano, Manu Beerens, Yasutaka Kurata, Isao Kobayashi, Noboru Fujino, Hiroshi Furusho, Kenji Sakata, Kenji Onoue, David Y Chiang, Tuomas O Kiviniemi, Eva Buys, Patrick Sips, Micah L Burch, Yanbin Zhao, Amy E Kelly, Masanobu Namura, Yoshihito Kita, Taketsugu Tsuchiya, Bunji Kaku, Kotaro Oe, Yuko Takeda, Tetsuo Konno, Masaru Inoue, Takashi Fujita, Takeshi Kato, Akira Funada, Hayato Tada, Akihiko Hodatsu, Chiaki Nakanishi, Yuichiro Sakamoto, Toyonobu Tsuda, Yoji Nagata, Yoshihiro Tanaka, Hirofumi Okada, Keiich Usuda, Shihe Cui, Yoshihiko Saito, Calum A MacRae, Seiji Takashima, Masakazu Yamagishi, Masa-aki Kawashiri, and Masayuki Takamura. 2020. “Impact of Functional Studies on Exome Sequence Variant Interpretation in Early-Onset Cardiac Conduction System Diseases.” CARDIOVASCULAR RESEARCH 116 (13): 2116–2130. doi:10.1093/cvr/cvaa010.
Vancouver
1.
Hayashi K, Teramoto R, Nomura A, Asano Y, Beerens M, Kurata Y, et al. Impact of functional studies on exome sequence variant interpretation in early-onset cardiac conduction system diseases. CARDIOVASCULAR RESEARCH. 2020;116(13):2116–30.
IEEE
[1]
K. Hayashi et al., “Impact of functional studies on exome sequence variant interpretation in early-onset cardiac conduction system diseases,” CARDIOVASCULAR RESEARCH, vol. 116, no. 13, pp. 2116–2130, 2020.
@article{8644562,
  abstract     = {Aims
The genetic cause of cardiac conduction system disease (CCSD) has not been fully elucidated. Whole-exome sequencing (WES) can detect various genetic variants; however, the identification of pathogenic variants remains a challenge. We aimed to identify pathogenic or likely pathogenic variants in CCSD patients by using WES and 2015 American College of Medical Genetics and Genomics (ACMG) standards and guidelines as well as evaluating the usefulness of functional studies for determining them.
Methods and Results
We performed WES of 23 probands diagnosed with early-onset (<65 years) CCSD and analyzed 117 genes linked to arrhythmogenic diseases or cardiomyopathies. We focused on rare variants (minor allele frequency < 0.1%) that were absent from population databases. Five probands had protein truncating variants in EMD and LMNA which were classified as “pathogenic” by 2015 ACMG standards and guidelines. To evaluate the functional changes brought about by these variants, we generated a knock-out zebrafish with CRISPR-mediated insertions or deletions of the EMD or LMNA homologs in zebrafish. The mean heart rate and conduction velocities in the CRISPR/Cas9-injected embryos and F2 generation embryos with homozygous deletions were significantly decreased. Twenty-one variants of uncertain significance were identified in 11 probands. Cellular electrophysiological study and in vivo zebrafish cardiac assay showed that 2 variants in KCNH2 and SCN5A, 4 variants in SCN10A, and 1 variant in MYH6 damaged each gene, which resulted in the change of the clinical significance of them from “Uncertain significance” to “Likely pathogenic” in 6 probands.
Conclusions
Of 23 CCSD probands, we successfully identified pathogenic or likely pathogenic variants in 11 probands (48%). Functional analyses of a cellular electrophysiological study and in vivo zebrafish cardiac assay might be useful for determining the pathogenicity of rare variants in patients with CCSD. SCN10A may be one of the major genes responsible for CCSD.

Translational Perspective
Whole-exome sequencing (WES) may be helpful in determining the causes of cardiac conduction system disease (CCSD), however, the identification of pathogenic variants remains a challenge. We performed WES of 23 probands diagnosed with early-onset CCSD, and identified 12 pathogenic or likely pathogenic variants in 11 of these probands (48%) according to the 2015 ACMG standards and guidelines. In this context, functional analyses of a cellular electrophysiological study and in vivo zebrafish cardiac assay might be useful for determining the pathogenicity of rare variants, and SCN10A may be one of the major development factors in CCSD.},
  author       = {Hayashi, Kenshi and Teramoto, Ryota and Nomura, Akihiro and Asano, Yoshihiro and Beerens, Manu and Kurata, Yasutaka and Kobayashi, Isao and Fujino, Noboru and Furusho, Hiroshi and Sakata, Kenji and Onoue, Kenji and Chiang, David Y and Kiviniemi, Tuomas O and Buys, Eva and Sips, Patrick and Burch, Micah L and Zhao, Yanbin and Kelly, Amy E and Namura, Masanobu and Kita, Yoshihito and Tsuchiya, Taketsugu and Kaku, Bunji and Oe, Kotaro and Takeda, Yuko and Konno, Tetsuo and Inoue, Masaru and Fujita, Takashi and Kato, Takeshi and Funada, Akira and Tada, Hayato and Hodatsu, Akihiko and Nakanishi, Chiaki and Sakamoto, Yuichiro and Tsuda, Toyonobu and Nagata, Yoji and Tanaka, Yoshihiro and Okada, Hirofumi and Usuda, Keiich and Cui, Shihe and Saito, Yoshihiko and MacRae, Calum A and Takashima, Seiji and Yamagishi, Masakazu and Kawashiri, Masa-aki and Takamura, Masayuki},
  issn         = {0008-6363},
  journal      = {CARDIOVASCULAR RESEARCH},
  keywords     = {Cardiac conduction-system disease,Whole exome sequencing,2015 ACMG standards and guidelines,CRISPR/Cas9-mediated gene knock-out in zebrafish,Cellular electrophysiological study,LONG QT SYNDROME,HYPERTROPHIC CARDIOMYOPATHY,MUTATION,CONSEQUENCES,ASSAY,GENE},
  language     = {eng},
  number       = {13},
  pages        = {2116--2130},
  title        = {Impact of functional studies on exome sequence variant interpretation in early-onset cardiac conduction system diseases},
  url          = {http://dx.doi.org/10.1093/cvr/cvaa010},
  volume       = {116},
  year         = {2020},
}

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