Six-month partial suppression of Huntingtin is well tolerated in the adult rhesus striatum

Abstract

Huntington's disease is caused by expression of a mutant form of Huntingtin protein containing an expanded polyglutamine repeat. One possible treatment for Huntington's disease may be to reduce expression of mutant Huntingtin in the brain via RNA interference. Unless the therapeutic molecule is designed to be allele-specific, both wild-type and mutant protein will be suppressed by an RNA interference treatment. A key question is whether suppression of wild-type as well as mutant Huntingtin in targeted brain regions can be tolerated and result in a net benefit to patients with Huntington's disease. Whether Huntingtin performs essential functions in the adult brain is unclear. Here, we tested the hypothesis that the adult primate brain can tolerate moderately reduced levels of wild-type Huntingtin protein for an extended period of time. A serotype 2 adeno-associated viral vector encoding for a short hairpin RNA targeting rhesus huntingtin messenger RNA (active vector) was bilaterally injected into the striatum of four adult rhesus monkeys. Four additional animals received a comparable vector encoding a scrambled control short hairpin RNA (control vector). General health and motor behaviour were monitored for 6 months. Upon termination, brain tissues were sampled and assessed blindly for (i) huntingtin messenger RNA knockdown; (ii) Huntingtin protein expression; and (iii) neuropathological changes. Reduction in wild-type huntingtin messenger RNA levels averaging ∼30% was measured in the striatum of active vector recipients 6 months post-injection. A widespread reduction in Huntingtin protein levels was also observed by immunohistochemistry in these animals, with an average protein reduction of ∼45% relative to controls measured by western blot analysis in the putamen of active vector recipients. As with control vector recipients, no adverse effects were observed behaviourally, and no neurodegeneration was found on histological examination of active vector recipients. Our results suggest that long-term partial suppression of wild-type Huntingtin may be safe, and thus if a comparable level of suppression of mutant Huntingtin is beneficial, then partial suppression of both wild-type and mutant Huntingtin may result in a net benefit in patients with heterozygous Huntington's disease.

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).…”

First use of creatine hydrochloride in premanifest Huntington disease

“…Intense research is being conducted into the genetic aspects of the disease with a view to modifying or correcting the genetic mutation, either by gene silencing/interference, replacement of the expanded CAG repeat or by use of molecular chaperones to alter downstream effects [50][51][52][53][54][55] . Whilst these strategies are promising, and are moving closer to clinical trials 53,56 , they currently represent strategies for the future. Thus, there is an urgent need to identify alternate treatments capable of impacting on disease progression with a clear intent to improve quality of life whilst the search for a cure continues.…”

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].…”

“…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.…”