Bi-allelic loss-of-function mutations in the collagen XII-encoding gene, COL12A1, are known to cause congenital hypotonia, respiratory insufficiency, joint hyperlaxity and joint contractures. Monoallelic, dominantly acting mutations in this gene cause a similar but milder phenotype and have been reported only in a handful of patients. We report detailed clinical characteristics of 6 patients (age 3-62 years) from 4 unrelated families with novel dominant mutations in COL12A1. All patients developed weakness at birth or during early childhood. Adult patients reported improvement of weakness and had little-to-no limitation of motor activities in young adulthood and mild weakness reemerged during 4th decade of life or later. Three pediatric patients had generalized weakness, while three adult patients had predominant distal upper and lower limb weakness. Joint contractures or hyperlaxity was observed in 5 patients. Three families carried dominant missense mutations affecting glycine residues, one in a laminin G-like domain and two in the Gly-X-Y repeats of the triple helix domain, while one family carried heterozygous in-frame deletion of exon 52. All of the mutations resulted in increased intracellular retention of collagen XII and loss of fibrillar pattern of staining in patients' fibroblasts. Since haploinsufficiency is not known to cause disease, we designed and screened small interfering RNAs (siRNAs) that specifically target the exon 52 mutant allele in patient-derived skin fibroblasts, thus converting the genetic abnormality to a haploinsufficient state. Immunostaining of the patient's fibroblasts treated with targeting siRNA corrected the intracellular retention and restored the fibrillar pattern of collagen XII staining. This study characterizes a distal myopathy phenotype in adults with dominant COL12A1 mutations, further defines the phenotypic spectrum and natural history of COL12A1-related myopathies, and provides proof of concept of a precision medicine treatment approach by proposing and validating allele-specific knockdown using siRNAs specifically designed to target a patient's COL12A1 mutation.

The exosome is essential for RNA metabolism via degradation of unstable AU-rich elements containing mRNAs. Mutations in EXOSC3 and EXOSC8, coding for core exosome subunits, and RBM7, coding for an exosomal-related protein, result in abnormal RNA metabolism, manifesting as spinal muscular atrophy, demyelination of the central nervous system and pontocerebellar hypoplasia (PCH1). Through whole exome sequencing (WES) we identified rare, predicted to be damaging, recessive mutations in a novel exosomal gene EXOSC9, in 2 independent patients with severe progressive neurodegeneration. Patient 1 (P1) is a 21-month-old female with normal tone at birth, followed by severe progressive weakness, feeding and respiratory difficulties by 8 months of age. MRI showed cerebellar atrophy, and EMG was suggestive of an axonal motor neuronopathy. WES identified a homozygous mutation p.L14P in EXOSC9. Patient 2 was born with fractures, arthrogryposis, severe hypotonia and respiratory insufficiency. MRI revealed generalized volume loss with cerebellum hypoplasia and delayed myelination. Muscle biopsy showed neurogenic changes. The same p.L14P mutation, in compound heterozygosity with a p.R161X mutation, was identified in EXOSC9. In vitro studies in P1's fibroblasts revealed a reduction of exosomal EXOSC8 and EXOSC3 protein levels, indicating secondary destabilization of the complex. Morpholino knockdown of Exosc9 in zebrafish resulted in embryos with phenotypes ranging from mildly affected to severely abnormal. CRISPR/Cas9-induced mutations in zebrafish Exosc9 resulted in milder, but similar defects with failure of motor neuron development and migration. Our data suggest that mutations in EXOSC9 may cause defective RNA metabolism due to abnormal function of the exosome, resulting in a severe clinical presentation related to PCH1. We establish EXOSC9 as a new gene for this phenotype, while further elucidating the role of exosome-mediated mRNA degradation in neurodegenerative disease.