Intrauterine gene therapy
Genetic editing caught up with the cubs in the womb
And it allowed to cure mice with congenital lung disease
Polina Loseva, "The Attic"
Molecular "scissors" CRISPR-Cas9 continue to be tried on to different targets. While some scientists are testing gene therapy on adults, and others are arguing about whether it is acceptable to apply it to single-celled embryos, others have chosen an intermediate goal. They tried to edit the genes of "adult" mouse embryos in the womb and rid them of a deadly lung disease. Despite the fact that they managed to repair only 20% of the cells and save only 6% of the mice, it is hardly worth judging strictly the first experiment of its kind that marked the way for the next attempts.
The scientific community is just beginning to explore the limits of the CRISPR-Cas9 system's capabilities for "fixing" people. So far, we have come closest to correcting monogenetic diseases – hereditary pathologies caused by a mutation of a single gene. If only one of the two copies of the gene is mutant in the embryo, then it can be cut out, leaving the second intact. According to this principle, scientists have recently saved mice from premature aging and muscular dystrophy.
In previous experiments, researchers have worked with either adult animals or single-celled embryos. The first way is inconvenient because it is impossible to deliver molecular "scissors" to all tissues of the body at the same time, so the nervous tissue is practically not "repaired". In addition, many organs of adult mice (and humans, too) are protected from outside intrusion, such as the skin or lungs, and the effectiveness of their editing is low. As for single-celled embryos, we still do not know whether such experiments on humans will be allowed in the near future. In addition, at the stage of a single cell, it is impossible to determine whether the embryo really inherited the mutant gene from its parents and whether it needs editing.
Therefore, American researchers went the third way. They experimented with "adult", 16-day-old mouse embryos (pregnancy in these animals lasts about 20 days). The target for them was a congenital lung disease caused by a mutation in the Sftpc gene. It is responsible for the release of surfactant – mucus lining the lungs from the inside. Mutation in Sftpc leads to the fact that substances – precursors of surfactant – do not come out of lung cells, but accumulate inside. As a result, the cells die, the lungs lose elasticity, the efficiency of gas exchange decreases and the carrier of the mutation dies shortly after birth. Only a lung transplant can help him.
The researchers worked with heterozygous mice carrying only one of the mutant copies of the gene. "Neutralized" viruses carrying CRISPR-Cas9 genes were injected directly into the amniotic fluid of a pregnant mouse mother. Then the embryos "inhaled" viruses along with the liquid, and to stimulate this process, the mother was briefly placed in a chamber with a high content of carbon dioxide. The embryos stopped having enough oxygen, and they reflexively "inhaled" deeper. Then the effectiveness of editing was evaluated at different stages of development: a few days after the injection, in the first days after childbirth and further, up to six months of independent life.
It turned out that the technique allows you to edit about 20% of lung cells – they stop accumulating excess products and secrete surfactant on the surface. This figure remains stable for at least six months after birth. At the same time, all types of lung epithelial cells can be edited, and the underlying vascular and connective tissue cells remain intact. Other organs of mice also did not change at the genetic level, with the exception of some cells of the digestive system, where part of the swallowed amniotic fluid got. This means that the new technology not only acts on those cells that are inaccessible in adulthood, but also bypasses other organ systems and therefore does not cause side effects.
Unedited lung epithelial cells in the images are marked in blue, and those that scientists were able to successfully edit are green. Figure from the article by Alapati et al. In utero gene editing for monogenic lung disease, published in the journal Science Translational Medicine.
However, the survival rate of mice still leaves much to be desired. The editing technique itself leads to high mortality – only 25% of control (healthy) mice that were injected with CRISPR-Cas9 viruses remained alive. Among the sick and edited mice, only 5.7% overcame the threshold of six months. The figure seems small, but it is worth remembering that sick mice die without treatment in the first hours after birth.
Thus, the first attempt to apply the new technology has borne some fruit, and the researchers hope to make it more effective and expand the field for its application. They add that selective editing of individual organ systems saves us from the risk of genetically modifying the germ cells of the embryo, thus removing one of the most acute ethical problems of modern science.
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