Validation of Ultrasensitive Mutant Huntingtin Detection in Human Cerebrospinal Fluid by Single Molecule Counting Immunoassay.

By Fodale, Valentina; Boggio, Roberto; Daldin, Manuel; Cariulo, Cristina; Spiezia, Maria Carolina; Byrne, Lauren M.; Leavitt, Blair R.; Wild, Edward J.; Macdonald, Douglas; Weiss, Andreas; et al
From Journal of Huntington’s Disease (2017), 6(4), 349-361. Language: English, Database: CAPLUS, DOI:10.3233/jhd-170269

The measurement of disease-relevant biomarkers has become a major component of clin. trial design, but in the absence of rigorous clin. and anal. validation of detection methodol., interpretation of results may be misleading. In Huntington’s disease (HD), measurement of the concn. of mutant huntingtin protein (mHTT) in cerebrospinal fluid (CSF) of patients may serve as both a disease progression biomarker and a pharmacodynamic readout for HTT-lowering therapeutic approaches. We recently published the quantification of mHTT levels in HD patient CSF by a novel ultrasensitive immunoassay-based technol. and here anal. validate it for use. Objective: : This work aims to anal. and clin. validate our ultrasensitive assay for mHTT measurement in human HD CSF, for application as a pharmacodynamic biomarker of CNS mHTT lowering in clin. trials. Methods: : The single mol. counting (SMC) assay is an ultrasensitive bead-based immunoassay where upon specific recognition, dye-labeled antibodies are excited by a confocal laser and emit fluorescent light as a readout. The detection of mHTT by this technol. was clin. validated following established Food and Drug Administration and European Medicine Agency guidelines. Results: : The SMC assay was demonstrated to be accurate, precise, specific, and reproducible. While no matrix influence was detected, a list of interfering substances was compiled as a guideline for proper collection and storage of patient CSF samples. In addn., a set of recommendations on result interpretation is provided. Conclusions: : This SMC assay is a robust and ultrasensitive method for the relative quantification of mHTT in human CSF.