A team of researchers from Barcelona, Spain, led by Dr. Mark Isalan and Dr. Mireia Garraga-Canut, have developed artificial zinc finger protein strands which bind the longer CAG repeat stretches of the HD protein while leaving the normal huntingtin protein gene, neighboring genes, and other genes with CAG repeat stretches unaffected. They achieved a 60 percent reduction of the HD protein in a mouse model and improved symptoms.
Zinc fingers were first discovered by researchers looking at a transcription factor in the clawed African frog. They found that the transcription factor was able to bind to DNA so well because it had finger like structures consisting of ions of zinc bound in a coordination complex with molecules called ligands. The zinc fingers stabilized the fold of the protein. Transcription factors are proteins which bind to DNA and control the flow of genetic information to RNA, either repressing or activating the gene. Zinc fingers are widely found in biology.
Researchers have been looking for ways to artificially engineer them for medical applications. Dr. Isalan and colleagues were able to develop a zinc finger protein (ZFP) which binds to the longer CAG repeat stretches found in the HD gene and tested them in HD patient and mouse model cell lines. The most effective one reduced the messenger RNA by 80 percent and the actual HD protein by 95 percent.
This ZFP was then administered to the 6/2 mice at 4 weeks through stereotaxic injections into the striatum using an adeno-associated viral vector for delivery (as has been used for RNA interference). The levels of the HD transgene mRNA were reduced in the striatum by 40 percent while mRNA from the normal huntingtin protein gene remained unaffected. Protein aggregation was reduced by 40 percent. Clasping behavior, a symptom of HD in the mice, was reduced with only one mouse out of eight showing any clasping behavior by seven weeks, compared to half of the untreated HD mice. Performance on the rotarod (maintaining balance on a rotating rod) is often used to access motor ability in HD mice. While the performance of the untreated mice was significantly impaired, treated mice performed as well as normal mice. The researchers were encouraged by these good results and recommend that the technology be pursued as a therapeutic strategy.
Sangamo Biosciences, in collaboration with Shire AG, is also working on zinc finger technology as a potential treatment for Huntington’s Disease. Their researchers presented data at the 2012 Annual Meeting of the Society for Neuroscience in October, 2012 which showed that their own zinc finger technology could repress the HD gene while allowing the normal huntingtin protein gene to express itself in cells derived from HD patients.
"The data presented at Neuroscience 2012 highlight our ability, using zinc finger technology, to intervene in HD specifically and directly at the level of the mutated DNA," stated Philip Gregory, D. Phil., Sangamo's vice president of research and chief scientific officer. "We are delighted with the potency, selectivity and unique specificity of our HD-targeted ZFP TFs and we are working with Shire to accelerate the development of our Huntington’s disease ZFP Therapeutics.”
Mireia Garraga-Canut, Carmen Agustin-Pavon, Frank Herrmann, Aurora Sanchez, Mara Dierssen, Cristina Fillat, and Mark Isalan . “Synthetic zinc finger repressors reduce mutant huntingtin expression in the brain of R6/2 mice.” Proceedings of the American Academy of Political and Social Sciences of the USA 2012 Nov 6;109(45):E3136-45.
Sangamo Biosciences press release