Amyloidogenicity assessment of transthyretin gene variants

• Published in: Annals of clinical and translational neurology Vol. 9; no. 8; pp. 1252 - 1263
• Main Authors: Grether, Nicolai B., Napravnik, Felix, Imhof, Thomas, Linke, Reinhold P., Bräsen, Jan H., Schmitz, Jessica, Dohrn, Maike, Schneider, Christian, Svačina, Martin K. R., Stetefeld, Jörg, Koch, Manuel, Lehmann, Helmar C.
• Format: Journal Article
• Language: English
• Published: United States John Wiley & Sons, Inc 01.08.2022; John Wiley and Sons Inc; Wiley
• Subjects: Amyloid - genetics; Amyloid - metabolism; Amyloid Neuropathies, Familial - genetics; Amyloid Neuropathies, Familial - metabolism; Amyloidosis; Cardiomyopathy; Cloning; Hemodialysis; Humans; Mutation; Prealbumin - genetics; Proteins; Sodium; Germany
• ISSN: 2328-9503; 2328-9503
• DOI: 10.1002/acn3.51626

Objective Hereditary transthyretin‐mediated amyloidosis is a treatable condition caused by amyloidogenic variants in the transthyretin‐gene resulting in severe peripheral neuropathy or cardiomyopathy. Only about a third of over 130 known variants are clearly pathogenic, most are classified as variants of uncertain significance. A clear delineation of these into pathogenic or non‐pathogenic is highly desirable but hampered by low frequency and penetrance. We thus sought to characterize their amylogenic potential by an unbiased in vitro approach. Methods Thioflavin T and turbidity assays were used to compare the potential of mammalian cell expressed wt‐transthyretin and 12 variant proteins (either variants of uncertain significance, benign, pathogenic) to aggregate and produce amyloid fibrils in vitro. As proof of principle, the assays were applied to transthyretin‐Ala65Val, a variant that was newly detected in a family with peripheral neuropathy and amyloid deposits in biopsies. In silico analysis was performed to compare the position of the benign and pathogenic variants. Results Transthyretin‐Ala65Val showed a significantly higher amyloidogenic potential than wt‐transthyretin, in both turbidity‐ and Thioflavin T‐assays, comparable to known pathogenic variants. The other eight tested variants did not show an increased amyloidogenic potential. In silico structural analysis further confirmed differences between pathogenic and benign variants in position and interactions. Interpretation We propose a biochemical approach to assess amyloidogenic potential of transthyretin variants. As exemplified by transthyretin‐Ala65Val, data of three assays together with histopathology clearly demonstrates its amyloidogenicity.