ESPE Yearbook of Paediatric Endocrinology

To read the full abstract: Science 2019;363(6424).

A wide range of human diseases result from haploinsufficiency, where gene expression is decreased as compared to normal conditions. Haploinsufficiency is a typical mechanism in autosomal dominant disorders. Rare mutations in Sim1 or Mc4r are models of haploinsufficiency causing severe early-onset obesity with hyperphagia. Gene therapy by adenovirus vectors has been developed to rescue such disorders, by expressing the normal allele in place of the mutant. However, this strategy is limited by the size of the DNA that can be packaged in this virus, and also by the ectopic insertion of the transgene leading to off-target side-effects.

Another way to circumvent these problems is to increase the expression level of the normal allele. The authors did this by coming up with a derivative of the CRISPR-Cas9 genome editing tool. ‘CRISPR-mediated activation’ (CRISPRa) is a method which combines a deficient Cas9 enzyme (dCas9) fused to an activator protein. When this protein complex is guided to a specific regulatory region, for instance, a promoter, which controls gene expression, the genome is not edited but rather the expression level of the targeted gene is increased.

Using this novel approach, they increased the expression levels of Sim1 and Mc4r specifically in the hypothalamus, by targeting hypothalamus-specific enhancers. They first confirmed that CRISPRa was able to regulate Sim1 expression in cells. They then adapted the method in CRISPRa transgenic mice or by adenovirus administration of dCas9 specifically to the hypothalamus. By both approaches, the severe obesity phenotype was rescued in Sim1^−/+ obese with very few off-target effects. Similar results were observed for Mc4r.

One great potential of this strategy is the possibility to rescue a complex phenotype, such as obesity, by increasing the expression of the normal allele. It is therefore unnecessary to define specific approaches for each genetic variant. The authors propose that up-regulation of endogenous genes could be a potential strategy to treat various altered gene dosage diseases.