CRISPR/Cas9-based functional genomics strategy to decipher the pathogenicity of genetic variants in inherited metabolic disorders
Por:
Muñoz-Pujol G, Ugarteburu O, Segur-Bailach E, Moliner S, Jurado S, Garrabou G, Guitart-Mampel M, García-Villoria J, Artuch-Iriberri R, Fons-Estupina C, Ribes A and Tort F
Publicada:
1 nov 2023
Ahead of Print:
1 oct 2023
Resumen:
The determination of the functional impact of variants of uncertain significance (VUS) is one of the major bottlenecks in the diagnostic workflow of inherited genetic diseases. To face this problem, we set up a CRISPR/Cas9-based strategy for knock-in cellular model generation, focusing on inherited metabolic disorders (IMDs). We selected variants in seven IMD-associated genes, including seven reported disease-causing variants and four benign/likely benign variants. Overall, 11 knock-in cell models were generated via homology-directed repair in HAP1 haploid cells using CRISPR/Cas9. The functional impact of the variants was determined by analyzing the characteristic biochemical alterations of each disorder. Functional studies performed in knock-in cell models showed that our approach accurately distinguished the functional effect of pathogenic from non-pathogenic variants in a reliable manner in a wide range of IMDs. Our study provides a generic approach to assess the functional impact of genetic variants to improve IMD diagnosis and this tool could emerge as a promising alternative to invasive tests, such as muscular or skin biopsies. Although the study has been performed only in IMDs, this strategy is generic and could be applied to other genetic disorders.
Filiaciones:
Muñoz-Pujol G:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Ugarteburu O:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Segur-Bailach E:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Moliner S:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Jurado S:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Garrabou G:
Inherited Metabolic diseases and Muscle Disorder's lab, Cellex-IDIBAPS, Faculty of Medicine and Health Sciences - University of Barcelona, Internal Medicine Service- Hospital Clinic of Barcelona, Barcelona, Spain and CIBERER, Madrid, Spain
Guitart-Mampel M:
Inherited Metabolic diseases and Muscle Disorder's lab, Cellex-IDIBAPS, Faculty of Medicine and Health Sciences - University of Barcelona, Internal Medicine Service- Hospital Clinic of Barcelona, Barcelona, Spain and CIBERER, Madrid, Spain
García-Villoria J:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Artuch-Iriberri R:
Clinical Biochemistry and Molecular Medicine and Genetics Departments, Institut de Recerca Sant Joan de Déu, Hospital Sant Joan de Déu, and CIBERER, Esplúgues de Llobregat, Barcelona, Spain
Fons-Estupina C:
Neurology Department, Fetal, Neonatal Neurology and Early Epilepsy Unit, Institut de Recerca, Hospital Sant Joan de Déu, Barcelona, Spain
Ribes A:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Tort F:
Secció d'Errors Congènits del Metabolisme-IBC, Servei de Bioquímica i Genètica Molecular, Hospital Clínic de Barcelona, IDIBAPS, CIBERER, Barcelona, Spain
Hosp Clin Barcelona, Seccio Errors Congenits Metab IBC, Serv Bioquim &
Genet Mol, IDIBAPS,CIBERER, Barcelona, Spain
Univ Barcelona, Hosp Clin Barcelona, Fac Med & Hlth Sci, Internal Med
Serv,Inherited Metab Dis & Muscle Dis, Barcelona, Spain
CIBERER, Barcelona, Spain
Hosp St Joan Deu, Inst Recerca St Joan Deu, Clin Biochem Dept,
Barcelona, Spain
Hosp St Joan Deu, Inst Recerca St Joan Deu, Mol Med & Genet Dept,
Hosp St Joan Deu, Inst Recerca, Neurol Dept, Fetal Neonatal Neurol &
Early Epilepsy Unit, Barcelona, Spain
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