Quantifying mutant huntingtin protein in human cerebrospinal fluid to support the development of huntingtin-lowering therapies

BackgroundHuntingtons disease (HD) is caused by a cytosine adenine guanine-repeat expansion in the huntingtin gene. This results in the production of toxic mutant huntingtin protein (mHTT), which has an elongated polyglutamine (polyQ) stretch near the proteins N-terminal end. The pharmacological lowering of mHTT expression in the brain targets the underlying driver of HD and is one of the principal therapeutic strategies being pursued to slow or stop disease progression. This report describes the characterisation and validation of an assay designed to quantify mHTT in the cerebrospinal fluid of individuals with HD, for use in registrational clinical trials. MethodsThe assay was optimised, and its performance was characterised with recombinant huntingtin protein (HTT) varying in overall and polyQ-repeat length. ResultsThe assay was successfully validated by two independent laboratories in regulated bioanalytical environments and showed a steep signal increase as the polyQ stretch of recombinant HTTs pivoted from wild-type to mutant protein forms. Linear mixed effects modelling confirmed highly parallel dose-response curves for HTTs, with only a minor impact of individual slopes of the dose-response for different HTTs (typically <5% of the overall slope). This implies an equivalent quantitative signal behaviour for HTTs with differing polyQ-repeat lengths. ConclusionThe reported method may be a reliable biomarker tool with relevance across the spectrum of HD mutations, which can facilitate the clinical development of HTT-lowering therapies in HD. Trial registrationNot applicable. FundingF. Hoffmann-La Roche Ltd.