PrP turnover in vivo and the time to effect of prion disease therapeutics

Abstract PrP lowering is effective against prion disease in animal models and is being tested clinically. Therapies in the current pipeline lower PrP production, leaving pre-existing PrP to be cleared according to its own half-life. We hypothesized that PrP’s half-life may be a rate-limiting factor for the time to effect of PrP-lowering drugs, and one reason why late treatment of prion-infected mice is not as effective as early treatment. Using isotopically labeled diet with targeted mass spectrometry, as well as antisense oligonucleotide treatment followed by timed PrP measurement, we estimate a half-life of 5-6 days for PrP in the brain. PrP turnover is not affected by over-or under-expression. Mouse PrP and human PrP have similar turnover rates measured in wild-type or humanized knock-in mice. CSF PrP appears to mirror brain PrP in real time in rats. PrP in the colon is readily quantifiable and has a half-life just slightly shorter than in brain. An under-expressed pathogenic mutant PrP, corresponding to D178N in humans, exhibits an accelerated turnover rate. Our data may inform the design of both preclinical and clinical studies of PrP-lowering drugs. Author Summary Prion disease is a fatal brain disease caused by misfolding of the prion protein (PrP). Emerging therapies for prion disease seek to reduce the amount of PrP produced in the brain in order to delay onset of disease or slow progression. Mouse studies have shown that if these therapies are initiated too late, their benefit is limited or they may not help at all. Here we measure the half-life of PrP in the mouse brain, and find that it is about 5 days. When drugs are used to lower PrP by cutting up the RNA that encodes PrP, the RNA drops rapidly while the protein lags behind, and does not reach its minimum level until 4 weeks after the drug is dosed. This half-life is about the same regardless of the species of PrP (mouse or human) and whether or not the brain is infected with prions. Cerebrospinal fluid appears to reflect the real-time levels of brain PrP with no appreciable lag. PrP can be measured in colon, which may be useful in animal studies of systemic drugs to lower PrP. PrP turns over more quickly in the presence of a pathogenic genetic variant, the equivalent of the human D178N variant. These findings suggest that clinical trials can monitor PrP in cerebrospinal fluid to look at drug activity, but should plan timepoints far enough post-dose to account for PrP’s rate of turnover, and should focus on patients who will survive long enough to benefit from the drug. Competing Interest Statement We have read the journal's policy and the authors of this manuscript have the following competing interests. BE and CB are employees of IQ Proteomics. ABS is an employee of Charles River Laboratories. NO is an employee and shareholder of Gate Bio. HTZ and BN are employees and shareholders of Ionis Pharmaceuticals. EVM has received speaking fees from Abbvie, Eli Lilly, Novartis, Vertex, and Voyager; consulting fees from Alnylam, Arrowhead, Deerfield, and Regeneron; and research support from Cenos, Eli Lilly, Gate Bio, Ionis, Regeneron, and Sangamo Therapeutics. SMV acknowledges speaking fees from Abbvie, Biogen, Eli Lilly, Illumina, Ultragenyx, and Voyager; consulting fees from Alnylam, Invitae, and Regeneron; research support from Cenos, Eli Lilly, Gate Bio, Ionis, Regeneron, and Sangamo Therapeutics. Footnotes Updates in response to reviewers round 1