Gene therapy partially restored the rats' eyesight
Until now, genetic engineering methods, such as the CRISPR-Cas9 system, based on the use of the cell's own DNA copying mechanisms, have been most effective against dividing cells, such as skin and intestinal cells.
However, recently, researchers at the Salk Institute, working under the guidance of Professor Juan Carlos Izpisua Belmonte, managed to change the situation and realize the cherished dream of specialists in the field of genetic engineering. They have developed an approach that not only increases the efficiency of embedding DNA fragments into the chromosomes of dividing cells by 10 times, but also allows for highly accurate correction of the genome of non-dividing cells, which mainly consist of many organs of an adult organism. This technology, with the help of which the authors managed to partially restore the vision of blind rodents, opens up new prospects for both fundamental research and the development of new therapeutic approaches, including the treatment of diseases of the retina, heart and nervous system.
Image of a fragment of the brain of an adult mouse. The cell nuclei are colored blue,
and neurons with an edited genome are green.
The secret of the new approach is to combine the CRISPR-Cas9 system, which allows inserting DNA fragments into the genome with very high accuracy, with a DNA damage repair mechanism known as non-homologous end joining (from the English NHEJ or non-homologous end-joining). This mechanism ensures the restoration of routine DNA breaks by connecting the ends formed during the rupture. To optimize the mechanism of non-homologous connection of the ends and to ensure the possibility of its joint action with CRISPR-Cas9, the authors developed a special cocktail of nucleic acids, called "homology-independent targeted integration" (from the English HITI or homology-independent targeted integration).
After that, they used an inert virus to deliver the HITI cocktail to neurons derived from human embryonic stem cells. The success of the procedure was the first evidence that the new approach could be effective against non-dividing cells.
The next stage of the work was the successful delivery of the genetic construct to the brain cells of adult mice. Finally, to test the possibility of using HITI technology in gene therapy, the researchers tested it on a rat model of retinitis pigmentosa, a hereditary disease characterized by retinal degeneration and blindness. As part of this experiment, 3-month-old animals were injected with a functional copy of the Mertk gene, usually damaged in retinitis pigmentosa. The examination carried out after the animals reached the age of 8 months showed that they reacted to light and passed a series of tests with results indicating partial restoration of the functionality of retinal cells.
The next stage of the work will be to increase the efficiency of delivery of the HITI construct to non-dividing cells. The authors note that, as with any other approaches of genetic engineering, in this case the difficulty also lies in introducing new DNA into a large number of cells at once. At the same time, the advantage of HITI technology is that it can be adapted to any genome editing system, and not only to the CRISPR-Cas9 system. Therefore, as the safety and efficiency of these systems increase, the scope of promising applications of HITI technology will expand.
Article by Keiichiro Suzuki et al. In vivo genome editing via CRISPR/Cas9 mediated homology-independent targeted integration is published in the journal Nature.
Evgeniya Ryabtseva
Portal "Eternal youth" http://vechnayamolodost.ru based on the materials of the Salk Institute: New gene-editing technology partially restores vision in blind animals.
18.11.2016