Kbtbd13 knockdown restores muscle function in a clinically relevant mouse model of nemaline myopathy type 6.

Nemaline myopathy type 6 (NEM6) is a rare neuromuscular disorder caused by pathogenic variants in the Kelch repeat and BTB domain-containing 13 (KBTBD13) gene. Patients experience muscle weakness, excessive fatigue, and impaired muscle relaxation that affect their daily activities related to abnormal protein aggregation in muscle cells and a predominance of slow-twitch myofibers, and no specific therapies are available. Here, we studied a clinically relevant Kbtbd13:p.Arg408Cys knockin mouse model to elucidate the molecular changes driving disease pathogenesis through measurements of muscle contractility, single myofiber assays, super-resolution microscopy, and x-ray diffraction. Kbtbd13:p.Arg408Cys knockin mice closely phenocopied human NEM6 pathology at the morphological, functional, and transcriptional levels, with the p.Arg408Cys variant causing mislocalization of KBTBD13 in the muscle sarcomere, associated with disease onset between 1 and 3 months of age, plateauing by 9 months, and with little progression at 18 months. As a potential therapeutic approach, we knocked down Kbtbd13 using short hairpin RNA against Kbtbd13 delivered intramuscularly via an adeno-associated virus 9 vector at either the prephenotype (1-month-old) or peak-phenotype (3- or 7-month-old) stages. A single treatment at the prephenotype stage prevented the development of impaired relaxation kinetics, nemaline rod aggregation, and slow-twitch myofiber predominance at 3 months of age. A single treatment at 3 months of age, after onset of disease, restored muscle morphology, contractility, and muscle-relaxation kinetics over 6 months. These data provide insights into NEM6 pathogenesis and suggest that Kbtbd13 knockdown might be a promising therapeutic strategy for patients with NEM6.