Preclinical Safety of RNAi-Mediated HTT Suppression in the Rhesus Macaque as a Potential Therapy for Huntington's Disease

Abstract

To date, a therapy for Huntington's disease (HD), a genetic, neurodegenerative disorder, remains elusive. HD is characterized by cell loss in the basal ganglia, with particular damage to the putamen, an area of the brain responsible for initiating and refining motor movements. Consequently, patients exhibit a hyperkinetic movement disorder. RNA interference (RNAi) offers therapeutic potential for this disorder by reducing the expression of HTT, the disease-causing gene. We have previously demonstrated that partial suppression of both wild-type and mutant HTT in the striatum prevents behavioral and neuropathological abnormalities in rodent models of HD. However, given the role of HTT in various cellular processes, it remains unknown whether a partial suppression of both alleles will be safe in mammals whose neurophysiology, basal ganglia anatomy, and behavioral repertoire more closely resembles that of a human. Here, we investigate whether a partial reduction of HTT in the normal non-human primate putamen is safe. We demonstrate that a 45% reduction of rhesus HTT expression in the mid- and caudal putamen does not induce motor deficits, neuronal degeneration, astrogliosis, or an immune response. Together, these data suggest that partial suppression of wild-type HTT expression is well tolerated in the primate putamen and further supports RNAi as a therapy for HD.

Synthetic zinc finger repressors reduce mutant huntingtin expression in the brain of R6/2 mice

“…Although this technique could be very powerful, mutant-selective RNAi depends on targeting single nucleotide or deletion polymorphisms that differentiate between alleles, and these often differ from patient to patient. However, there is evidence that partial repression of wt HTT can be tolerated (14,15), suggesting that generic approaches that repress both alleles should also be pursued. Peptide nucleic acids and locked nucleic acids are generic; yet, some promising partially selective inhibition of expanded CAG repeats of the ataxin-3 and HTT genes has been reported (16,17).…”

Recent Advances in RNA Interference Therapeutics for CNS Diseases

“…Both nonallele-specific (targeting the mutant and wild type alleles) and allele-specific (targeting only the mutant allele) approaches for HD therapy are under development. It has been shown recently that nonallele-specific silencing, using AAVmediated delivery of RNAi, provides benefits in a mouse model of HD and 2 studies assessing the effect of knockdown of endogenous HTT in rhesus found no adverse effects up to 6 months post-injection [81,82]. However, HTT is necessary for embryonic development and is involved in cellular pathways in differentiated neurons [83][84][85][86][87][88].…”

“…For example, SCA1, SCA2, and SCA3 knockout mice are viable and fertile, indicating that knockdown of the wild type allele function may be tolerable [96,166,167]. Nonallele-specific silencing of HTT in HD mice resulted in a significant rescue of the HD phenotype, and 2 studies have shown that reducing levels of wild type HTT in the adult rhesus macaque striatum is safe and well tolerated for at least 6 months [80][81][82]. However, as the HD null mice are embryonic lethal, and the levels of HTT required for cell viability of adult neurons is unknown, researchers are also investigating allele-specific silencing options [83].…”

“…To detect siRNAs after delivery, Stiles et al [168] used radiolalebeled siRNAs, which allowed comparison of the volume of brain for which there was target suppression, as a function of dose and spread. For viral vectors, a common strategy to determine the distribution of transduced cells takes advantage of reporter genes, such as eGFP [80,82]. In the future, targeting the brain after systemic delivery may be possible.…”

“…Thus, it is important to understand what kind of dose may be appropriate and how long the RNAi efficacy is retained in the human brain. A number of studies have focused on determining an appropriate dose of RNAi that is efficacious and safe in the primate brain using viral and nonviral methods [81,82,168,169]. Nonviral delivery systems used cannulas implanted in the brain or convection enhanced delivery systems that use flow pressure to increase the volume of distribution.…”