Mass spectrometric assays for the detection of transthyretin mutations in hereditary transthyretin amyloidosis

Abstract

Hereditary transthyretin amyloidosis is a fatal autosomal dominant disorder resulting from missense single-nucleotide substitutions in the transthyretin gene, with consequent transthyretin amyloid deposition causing various serious systemic manifestations including cardiomyopathy, neuropathy, ocular and leptomeningeal amyloidosis. Laboratory diagnosis of transthyretin gene mutations in Australia has hitherto employed Sanger sequencing of the transthyretin gene. We have developed multiple reaction monitoring mass spectrometry (MRM MS) assays for the detection of variant transthyretin proteins directly from serum and plasma samples and have evaluated their performance using both research and diagnostic grade tandem mass spectrometers. We present a method that has enabled detection of eight distinct transthyretin mutations with very high sensitivity and specificity, at greatly reduced turnaround time and cost, compared to the gold standard of genomic sequencing. MRM MS methods may be readily extended to detect most non-isobaric mutations in clinically relevant peptides and proteins.