Therapy for сollagen VI-CMD

Collagen VI congenital muscular dystrophy (COL VI-CMD), previously also known as Bethlem or Ullrich myopathy, is caused by mutations in the three collagen VI genes Col6A1, Col6A2, and Col6A3.

The disease is one of more than 8,000 rare diseases with a frequency of 1-9/1,000,000. However, among the different muscle diseases, COL VI-CMD is one of the most common disorder accounting for about 30% of cases.

So far, no therapy for collagen VI-CMD is known.


Prof. Dr. Jeanette Erdmann

Franziska Haarich, MSc

Svenja Kohler

Projects

Franziska Haarich, MSc

Svenja Kohler

RNA-based therapies attack even before the “blueprint” read from the DNA has been converted into a protein. In principle, this usually involves the suppression of mutated gene copies at the RNA level by means of so-called small interfering RNA (siRNA) or antisense oligonucleotides (ASO).

Also, in the case of collagen VI myopathy, suppression of the mutated allele should lead to therapeutic success. In the literature, first steps towards the establishment of RNA-based therapies can be found for individual mutations in the Col6A2 or Col6A3 gene. However, the first and only approach to target Col6A2 dates back to 2012 and targeted a variant leading to Ullrich CMD located in intron 9 (c.954 + 17_954 + 22del28).

We have started – together with Prof. Dr. Georg Sczakiel, director of the Institute for Molecular Medicine at the University of Lübeck – to establish an RNA-based therapy concept for the mutation p.Gly283Glu in the COL6A2 gene in the sense of individualized medicine. The goal is to generally establish this therapeutic approach targeting other dominant-negative acting glycine mutations in Col6A2 gene as well.

Furthermore, we want to develop another therapeutic approach for Col VI-CMD. Here, the aim is to use CRISPR interference (CRISPRi) to deactivate the activity of regulatory elements of Col6A2 in order to prevent transcription of the mutated allele. For CRISPRi, a Cas9 is used that no longer has a functional nuclease domain due to specific mutations (dCas9) and thus cannot cause double-strand breaks in the target sequence. Coupled with transcriptional repressors and in combination with specific guide RNAs (gRNA), dCas9 binds DNA and thus prevents transcription of the targeted gene. We will use frequent variants that are located in regulatory elements as targets to treat different Col6A2 mutations with the same gRNA. Important is that these common variants are in phase with (on the same allele as) the pathogenic variant to repress the pathogenic allele specifically. For the phasing, we enrich a 60kb region (including the gene and regulatory elements) via a method called Cas9 enrichment and sequence it using an Oxford Nanopore MinION (portable Nanopore Sequencer). The subsequent data analysis pipeline contains different command line tools. 

Funded by Deutsche Gesellschaft für Muskelkranke e.V. & Cure CMD

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