Variant-specific deep phenotyping as a tool to develop precision therapies

Gene-modifying therapies represent a transformative advance in rare genetic disease, enabling personalized interventions that target the underlying genetic causes with unprecedented accuracy.1 Antisense oligonucleotides (ASOs) are short synthetic strands of nucleic acids designed to specifically bind to a target mRNA sequence and modulate gene expression. By binding to a specific region of mRNA, ASOs can block or alter the splicing process, prevent the production of harmful proteins, or promote the production of functional proteins. ASOs opened new therapeutic avenues in neuromuscular diseases by enabling tailored interventions in spinal muscular atrophy (SMA) and Duchenne muscular dystrophy (DMD).2,3 In SMA, ASOs can compensate for the lack of SMN1 by increasing the expression of the SMN2 gene via altered splicing, and have been used worldwide in clinical practice through intrathecal administration (nusinersen).4 Exon skipping using ASOs is also a promising therapeutic strategy that enables the bypassing of defective exons in the dystrophin gene, restoring the production of a truncated but functional dystrophin protein in patients with specific mutations in DMD.2 However, targeting muscle with ASOs through intravenous administration has been less effective. Novel ASOs with improved muscle targeting are currently undergoing clinical trials for a variety of muscular dystrophies including DMD, facioscapulohumeral muscular dystrophy (FSHD) and myotonic dystrophy.3 The paper by Foley et al.5 in this issue of Brain highlights that targeting a single intronic variant with ASOs would be feasible to treat COL6-related dystrophy (COL6-RD) due to the recurrent variant COL6A1 c.930+189C>T. The resulting protein is expected to be full-length without a mutation, which is a particularly favourable outcome of exon skipping, unlike the truncated in-frame minidystrophin proteins resulting from exon skipping in DMD. Collagen VI-related dystrophies (COL6-RDs) encompass a broad spectrum of clinical phenotypes, ranging from the severe presentation of Ullrich congenital muscular dystrophy (UCMD), characterized by early-onset progressive muscle weakness, joint contractures and respiratory insufficiency, to the milder and later-onset Bethlem muscular dystrophy, which often resembles limb girdle muscular dystrophy.5 Intermediate phenotypes exhibit a clinical course between UCMD and Bethlem muscular dystrophy. Distinct clinical symptoms and muscle pathology are characteristic for COL6-RD. However, a subset of patients with typical presentation has remained without a causative variant in the canonical genes—COL6A1, COL6A2 or COL6A3. The combination of muscle RNA-sequencing and whole-genome sequencing in these individuals identified a recurrent, de novo deep intronic variant in intron 11 of COL6A1 (c.930+189C>T) that leads to a dominantly acting in-frame pseudo-exon insertion.6 Screening for this variant in additional patients with typical COL6-RD presentation has detected 44 individuals from 15 different countries, making this variant one of the most common recurrent causative variants in the collagen 6 genes.5 Detailed analysis of the clinical presentation and progression of individuals with this variant in a natural history study demonstrates a distinct phenotype—bridging between UCMD and Bethlem muscular dystrophy.5 While the symptoms are less prominent at birth, they show an accelerated progression to a severe form of UCMD and affected children lose independent ambulation at a mean age of 8.0 ± 3.0 years and require non-invasive ventilation at the mean age at 11.9 ± 4.4 years. One patient with somatic mosaicism for the COL6A1 intron 11 variant manifested a milder phenotype consistent with Bethlem muscular dystrophy, suggesting that a slight reduction in the pseudo-exon insertion in COL6A1 can result in partial rescue of UCMD into Bethlem muscular dystrophy, providing a strong rationale for the development of a splice-modifying ASO therapy.6,7 Previous studies have shown that splice modulating ASOs applied in vitro effectively decreased the mutant pseudoexon-containing COL6A1 transcripts to levels comparable to the somatic mosaicism found in one individual.7 Performing an international natural history study of this single variant and directly relating it to a ‘comparison cohort’ of patients with any other variants in the same gene causing UCMD has been a crucial step to determine how patients with this variant progress without a therapy and highlighted a precise baseline dataset enabling detection of treatment effect in a single or in a few patients in future treatment trials. The study also highlighted that developmental factors may affect splicing, which delayed the first manifestation of disease (almost no symptoms at birth), but resulted in rapid progression in childhood, which may be relevant for other genetic diseases with a similar mechanism. These findings highlight that using ASOs to treat patients with COL6-RD caused by COL6A1 c.930+189C>T may be a feasible approach. Genetic technologies have already shifted medicine from population-based therapeutics to stratified cohorts, and more recently this is further moving to truly individualized medicines in the rare disease space.8 These personalized treatments hold the potential to correct or modulate defective gene expression, offering hope where traditional approaches fail. However, the application of ASOs in rare diseases also presents substantial challenges.9 Small patient populations and variable phenotypes lead to difficulties in validating efficacy and safety. The high cost of developing and manufacturing personalized ASOs and the regulatory hurdles can also restrict clinical trials. Additionally, the long-term effects of such treatments remain uncertain, highlighting the need for continuous monitoring and refinement as the field evolves. Despite these difficulties, the field of individualized, or N-of-1, therapy development is growing and increasingly gaining attention as a novel option for people with severe diseases, caused by unique genetic variants for whom approved therapies are not available. The roadmap for developing such treatments highlights what needs to be established for N-of-1 therapies.8,9 The paper by Foley et al.5 closes a translational gap paving the way to develop an ‘N-of-few’ ASO therapy for a single COL6A1 mutation, which can be applicable for other rare genetic conditions.

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