Calibrating a functional assay for variant classification in RYR1-related malignant hyperthermia susceptibility

Authors

Evan Z. Ying, Center for Precision Health Research, National Human Genome Research Institute, National Institutes of Health, 50 South Drive, Bethesda, MD 20892, United States.
Amber Douglass, Center for Precision Health Research, National Human Genome Research Institute, National Institutes of Health, 50 South Drive, Bethesda, MD 20892, United States.
Marwan A. Hawari, Center for Precision Health Research, National Human Genome Research Institute, National Institutes of Health, 50 South Drive, Bethesda, MD 20892, United States.
Linda Groom, Department of Pharmacology and Physiology, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, United States.
Kathryn Stowell, School of Food Technology and Natural Sciences, Massey University, Tennent Drive, Palmerston North 4410, New Zealand.
Robert T. Dirksen, Department of Pharmacology and Physiology, University of Rochester Medical Center, 601 Elmwood Avenue, Rochester, NY 14642, United States.
Jennifer J. Johnston, Center for Precision Health Research, National Human Genome Research Institute, National Institutes of Health, 50 South Drive, Bethesda, MD 20892, United States.
Leslie G. Biesecker, Center for Precision Health Research, National Human Genome Research Institute, National Institutes of Health, 50 South Drive, Bethesda, MD 20892, United States.

Document Type

Journal Article

Publication Date

4-7-2025

Journal

Human molecular genetics

DOI

10.1093/hmg/ddaf038

Keywords

Calcium Release Assay; Malignant Hyperthermia Susceptibility; RYR1; Variant Classification

Abstract

PURPOSE: Identifying individuals with pathogenic variants for RYR1-related Malignant Hyperthermia Susceptibility (MHS) could reduce morbidity and mortality due to MH reactions. Realization of this goal requires knowledge of variant pathogenicity, and proper weighting of functional assays is important for accurate variant classification. Caffeine-induced Ca2+ release assays can be used to support pathogenicity per the ClinGen Variant Curation Expert Panel (VCEP). However, the caffeine-induced Ca2+ release assay lacks formal validation with known pathogenic and benign variants. METHODS: Fifteen benign/likely benign and six pathogenic/likely pathogenic RYR1 variants were used to calibrate the caffeine-induced Ca2+ release assay using a multi-mode microplate reader. Five variants of unknown significance (VUS) were assayed for possible reclassification. RESULTS: Our data support use of the caffeine-induced Ca2+ release assay at a moderate weight per the American College of Medical Genetics and Genomics pathogenicity criteria schema with a positive likelihood ratio of 12.14:1 (pathogenicity) and a negative likelihood ratio of 0.22:1 (4.5:1 benignity). Using this validated assay, two of five VUS were reclassified as likely benign. CONCLUSION: Formal validation of the caffeine-induced Ca2+ release assay supports the VCEP functional criteria weighting at moderate strength based on these data. Additional variants should be assayed to shift more from VUS to benign or pathogenic classifications.

APA Citation

Ying, Evan Z.; Douglass, Amber; Hawari, Marwan A.; Groom, Linda; Stowell, Kathryn; Dirksen, Robert T.; Johnston, Jennifer J.; and Biesecker, Leslie G., "Calibrating a functional assay for variant classification in RYR1-related malignant hyperthermia susceptibility" (2025). GW Authored Works. Paper 7090.
https://hsrc.himmelfarb.gwu.edu/gwhpubs/7090

Department

School of Medicine and Health Sciences Student Works