NEM6: From pathophysiology to therapy

Summary of research

In unraveling NEM6 pathology, our group played a role of crucial importance: 

• 2002: We described a large Dutch family with nemaline myopathy (NEM) and a peculiar kind of muscle slowness, that severely hampers daily-life activities. 
• 2010: In an international consortium, we identified the implicated gene: KBTBD13 (referred to as NEM6). 
• 2020: We discovered that the muscle slowness is caused by binding of mutant KBTBD13 to actin, thereby stiffening the actin filaments in sarcomeres. This finding was made possible by the generation of the first NEM6 mouse model, which harbours the Dutch founder mutation Kbtbd13R408C . The stage is set now to take the next step towards therapy.

Hypothesis: Knock-down of mutant KBTBD13 prevents and reverses disease development in NEM6. Two important pilot findings underlie this hypothesis:

• First, Kbtbd13R408C -knockin mice phenocopy NEM6.
• Second, Kbtbd13-KO mice, i.e. mice lacking Kbtbd13, do not develop a phenotype.

Study objectives
To test the hypothesis, we have formulated three objectives:

1. Determine the natural history of NEM6 in the Kbtbd13R408C mouse model. This will dictate the age at which knockdown of transcript should commence.
2. Test the efficacy of AAV-mediated knock-down of mutant transcript in the Kbtbd13R408C mouse model. The efficacy of intramuscular and systemic delivery is studied.
3. Develop gene-editing approaches to delete Kbtbd13 from myofibers of the Kbtbd13R408C mouse model. Longlasting knock-down of mutant transcript might require repetitive systemic AAV-delivery. Therefore, we will also determine the efficacy of single-treatment gene-editing in cultured myofibers and mice to achieve permanent knockdown

Approach
We have available: 

• State-of-the-art techniques to study muscle structure/function 
• Two unique mouse models:
  • o Kbtbd13R408C: harbours the most frequently occurring mutation in KBTBD13; phenocopies NEM6.
  • o Kbtbd13-knockout: Kbtbd13-deficient; does not develop a phenotype. 
• Techniques to generate AAV-based shRNA and gene-editing approaches. 
• Novel infrastructure to study the contractility of cultured myofibers of Kbtbd13R408C mice.

Expected results: We foresee that reducing Kbtbd13 transcript levels prevents and reverses the NEM6 phenotype, an expectation supported by adequate pilot data. The proof-of-concept thus obtained will direct us towards the testing of our approach in NEM6 patients.

Awarded grant, amount: €280.000

Funding agency: Prinses Beatrix Spierfonds; A Foundation Building Strength for Nemaline Myopathies