ARTICLE IN BRIEF
In a mouse model of spinocerebellar ataxia 7, scientists observed that treatment with an antisense oligonucleotide (ASO) led to a significant reduction in aggregated pathogenic protein ataxin-7, as well as normal-length protein, in retinal neurons. Electroretinograms of treated versus control-injected eyes indicated that ASO treatment led to improved retention of visual function at four weeks and six weeks.
When injected directly into the eye, antisense oligonucleotides (ASOs) that target the gene for spinocerebellar ataxia type 7 (SCA7) reduce levels of the pathogenic protein ataxin-7 and restore visual function in mice with visual impairments due to the mutant protein, according to an October 31 study in Science Translational Medicine.
While SCA7 is rare and visual loss is only one aspect of this multifaceted disorder, the results of the study support the growing interest in using ASOs to treat neurodegenerative disease in general and SCA7 in particular.
Because the retina is easily accessible, and because there are objective means of monitoring response to therapy, “ASOs that reduce the dosage of the mutant protein may represent a viable treatment strategy for SCA7 retinal degeneration,” the principal investigator Albert La Spada, MD, PhD, professor of neurology at Duke University School of Medicine in Durham, NC, told Neurology Today.
Retinal degeneration is an uncommon feature in other neurodegenerative diseases, but it can be one of the most debilitating features of SCA7. SCA7 is a polyglutamine repeat disorder, which shows very strong anticipation, Dr. La Spada explained, such that the number of CAG repeats in the coding region of the ataxin-7 gene tends to grow between generations so much that children may present with symptoms before their parents.
Vision loss occurs earliest and becomes most severe in those with longer CAG repeats. “Patients who have 56 or more repeats typically present with vision loss before they develop cerebellar abnormalities,” he said, and the longer the repeat, the more significant and rapidly progressive that vision loss tends to be. Those with shorter repeats tend to develop neurologic symptoms before vision loss; the vision loss may not develop until a decade or more after cerebellar symptoms begin and is usually less troublesome. The normal repeat number is ten or fewer.
In SCA7, cones degenerate before rods, leading initially to blue-yellow color blindness, followed by central scotomas that gradually expand, resulting in blindness. Dr. La Spada pointed out that, apart from ataxin-7, there are relatively few cone-rod dystrophy genes, and that one of them, called CRX is, like the ataxin-7 protein, a transcription factor. That has led to the prevailing theory that interaction between mutant ataxin-7 and CRX contributes to retinal degeneration.
The ASOs used in the current study were designed to reduce the amount of both mutant and normal protein by binding to messenger RNA and triggering its degradation. While targeting the polyglutamine repeat section of the mRNA seemed an obvious choice, since it is not shared by the normal allele, Dr. La Spada chose not to do so, because of concerns about the potential for off-target effects.
“There are many genes with considerable CAG tracts that serve normal functions,” he said, including other transcription factors, and reducing the expression of these may be problematic.
Instead, the ASO targeted a region shared by both the mutant and normal allele. He noted that even knocking out ataxin-7 entirely doesn't cause visual problems, and that in most ASO therapeutic applications, protein reduction is at best 60- to 80-percent, suggesting the choice was likely to be safe.
Dr. La Spada first showed that injection of ataxin-7-specific ASOs into the mouse vitreous humor was safe and effectively suppressed production of the ataxin-7 protein. At the dose selected, the team achieved more than 50 percent knockdown of ataxin-7 mRNA at four weeks post-injection; it fell to more than 40 percent at six weeks. Immunostaining indicated that the ASO was well distributed throughout the retina.
In a mouse model of SCA7 that includes progressive electrophysiological dysfunction and retinal degeneration, they observed a significant reduction in aggregated protein, as well as normal-length protein, in retinal neurons. Electroretinograms of treated versus control-injected eyes indicated that ASO treatment led to improved retention of visual function at four weeks and six weeks.
In non-transgenic mice, ASO treatment did not lead to measurable change in visual function, strengthening the case that reducing the level of the normal protein is safe.
In the transgenic mice, treatment also improved retinal histopathology, changes in expression of other genes, and chromatin structural changes caused by the disease gene.
“The reduction of misfolded protein is encouraging,” Dr. La Spada said, “because it is a biomarker for the disease. But the most important thing in the final analysis is visual function, so the fact that treatment led to retention of visual function in the treated eye is the most important result.”
In mice with a slower disease course, Dr. La Spada showed that treatment after the development of significant visual loss improved visual function in the ASO-treated eye compared to the control-treated eye, although not to the same extent as earlier treatment in the more rapidly progressing model had.
“Our results suggest that ataxin-7 ASO treatment might ameliorate [visual loss] progression even when administered after symptoms develop,” Dr. La Spada said, although determining the point at which treatment might no longer offer clinically significant benefit will remain an important question for future trials.
Human trials are being considered, Dr. La Spada said, with development of human-specific ASOs currently underway at Ionis Pharmaceuticals. Intravitreal injections are well-tolerated, and the first ASO to receive US Food and Drug Administration approval, fomiversen, is delivered to the vitreous to treat cytomegalovirus retinitis. Assuming clinical development doesn't hit a roadblock, treatment would likely be three times per year, to replenish ASOs as they are degraded.
“If we can show that dosage reduction of the mutant protein is effective in the eye in SCA7 patients, it should embolden us to move forward with dosage reduction in more common protein misfolding disorders, such as amyotrophic lateral sclerosis, Parkinson's disease, and Alzheimer's disease,” Dr. La Spada said.
“What I found most striking in this study was the degree of restoration of physiological retinal function,” said Harry Orr, PhD, professor and chair in genetics at the University of Minnesota Medical School in Minneapolis. Dr. Orr is developing ASO therapy to treat SCA1, in collaboration with Ionis Pharmaceuticals, which created the ASOs for the SCA7 study as well. “It would be important to be cautious, since one can't directly test mouse vision, but the authors very nicely showed they had an impact on the underlying pathology and altered physiology of the photoreceptor cells.”
“This study adds to the growing number of investigations showing that ASO-based therapies have promise for the treatment of neurodegeneration,” said Stefan M. Pulst, MD, DrMed, FAAN, professor and chair of neurology at the University of Utah in Salt Lake City. “Although SCA7 is a disease affecting multiple cell types in the central nervous system, the retinal disease can be extremely debilitating to patients. It is important to note that the authors studied symptomatic rather than pre-symptomatic mice, as this more closely reflects the situation in the real-world practice setting.”
The study was supported by the National Ataxia Foundation, Harrington Discovery Initiative, Foundation Fighting Blindness, and grants from the National Institutes of Health. Drs. La Spada, Orr, and Pulst reported no disclosures relevant to this study.