VP.72 Diversity of splice-acting variants in the COL6A1, COL6A2 and COL6A3 genes associated with collagen VI-related dystrophies

Collagen VI-related dystrophies (COL6-RDs) are disorders of the myomatrix that manifest with a spectrum of severity. Over 400 variants have been described in the COL6A1, COL6A2, and COL6A3 genes in association with COL6-RD, although many have unclear pathogenicity. The recent discovery of a recurrent deep intronic splice-inducing variant (COL6A1 c.930+189C>T) that is actionable with antisense therapy prompted us to systematically explore the impact of splice-modulating variants in the COL6 genes. We queried our internal database of 225 molecularly confirmed and/or suspected COL6-RD patients. We found 130 patients (57.8%) carrying variants affecting, or potentially affecting, splicing, totaling 88 unique variants (61 dominant, 27 recessive; 40 in COL6A1, 29 in COL6A2, and 19 in COL6A3). To further investigate the effect of these variants on splicing, we selected 60 patients (representing 61 unique variants) for RNA-sequencing of muscle biopsy specimen (n=3) or of cultured dermal fibroblasts (n=57). The most frequent mechanism for dominant variants was in-frame exon skipping. However, unexpectedly in select cases, variants were leaky, causing only moderate levels of exon skipping predicted to translate into lesser clinical impact. In addition to the COL6A1 c.930+189C>T variant, we found two variants leading to in-frame insertions (COL6A2 c.900+1G>A; COL6A3 c.6309+10C>G), suggesting that this mechanism might be more frequent than previously thought. A recurrent recessive variant was COL6A2 c.1970-9G>A (n=6) was also characterized and found to create a novel acceptor splice site and to insert seven nucleotides in the mRNA. This study helps to further clarify the pathogenicity of COL6 variants and resulting genotype-phenotype correlations and to identify variants that may be candidates for therapeutic splice modulation. As individualized RNA-targeting therapies are becoming more feasible, the appropriate recognition of "actionable" variants is critical. Collagen VI-related dystrophies (COL6-RDs) are disorders of the myomatrix that manifest with a spectrum of severity. Over 400 variants have been described in the COL6A1, COL6A2, and COL6A3 genes in association with COL6-RD, although many have unclear pathogenicity. The recent discovery of a recurrent deep intronic splice-inducing variant (COL6A1 c.930+189C>T) that is actionable with antisense therapy prompted us to systematically explore the impact of splice-modulating variants in the COL6 genes. We queried our internal database of 225 molecularly confirmed and/or suspected COL6-RD patients. We found 130 patients (57.8%) carrying variants affecting, or potentially affecting, splicing, totaling 88 unique variants (61 dominant, 27 recessive; 40 in COL6A1, 29 in COL6A2, and 19 in COL6A3). To further investigate the effect of these variants on splicing, we selected 60 patients (representing 61 unique variants) for RNA-sequencing of muscle biopsy specimen (n=3) or of cultured dermal fibroblasts (n=57). The most frequent mechanism for dominant variants was in-frame exon skipping. However, unexpectedly in select cases, variants were leaky, causing only moderate levels of exon skipping predicted to translate into lesser clinical impact. In addition to the COL6A1 c.930+189C>T variant, we found two variants leading to in-frame insertions (COL6A2 c.900+1G>A; COL6A3 c.6309+10C>G), suggesting that this mechanism might be more frequent than previously thought. A recurrent recessive variant was COL6A2 c.1970-9G>A (n=6) was also characterized and found to create a novel acceptor splice site and to insert seven nucleotides in the mRNA. This study helps to further clarify the pathogenicity of COL6 variants and resulting genotype-phenotype correlations and to identify variants that may be candidates for therapeutic splice modulation. As individualized RNA-targeting therapies are becoming more feasible, the appropriate recognition of "actionable" variants is critical.