Plasmids to help gene therapy
Researchers at Stanford University, working under the guidance of Professor Lawrence Steinman, have demonstrated that gene therapy can be effective without the risk of developing a dangerous side effect characteristic of all gene therapy approaches – an autoimmune reaction to proteins that the patient's immune system encounters for the first time.
According to Professor Steinman, at the present stage, gene therapy is approaching a turning point, after which it will become a first-line therapy for monogenic (caused by a defect in one gene) diseases. However, there is a rather serious problem: if a gene encoding a normal protein is introduced into the human body, whose cells synthesize an abnormal version of this protein, its immune system will respond to a normal, but unfamiliar version of the protein with the development of an immune response. In some cases, this can even lead to the death of the patient. In addition, an immune response can develop to the proteins of the viral vector used as a carrier of the therapeutic gene.
Steinman and his colleagues believe that they have managed to overcome this problem. They demonstrated this in a mouse model of Duchenne muscular dystrophy, a genetic disease affecting 1 in 5,000 boys. It causes gradual muscle atrophy, so such patients are usually confined to a wheelchair already in adolescence, and about 20 years die due to weakening of the respiratory muscles. This pathology is caused by a genetic defect that deprives skeletal and cardiac muscles of a functional version of the protein dystrophin.
The monogenic nature of the disease makes it an ideal candidate for treatment with gene therapy by replacing a defective gene with its functional variant. This can be done with the help of viral vectors that penetrate into cells and start copying the genes recorded in them in each infected cell. To obtain such vectors, unwanted genes are removed from the DNA of viruses, leaving only the genes responsible for the ability to infect cells, and a therapeutic gene is introduced into it, which is necessary to correct the patient's genetic defect.
The gene encoding the complete variant of dystrophin is too large to be embedded in the viral vector. However, the researchers managed to isolate a fragment of a gene that provides the synthesis of a sufficiently functional version of the protein, called microdystrophin. This fragment is successfully embedded in an adenovirus-based vector, but their use as a gene therapy drug does not exclude the development of serious immune reactions.
To solve this problem, the authors suggest using plasmids – ring DNA, with which bacteria exchange important properties for them, such as resistance to antibiotics. The plasmid used by the authors usually contains several short DNA sequences or "motifs" that the immune system recognizes as suspicious and triggers a strong immune response. However, the authors managed to replace these fragments with other DNA sequences that not only do not cause an immune response, but also suppress it. Such immune tolerance-inducing plasmids have already demonstrated promising results in clinical trials of therapy for two autoimmune diseases.
As part of the study, mice without functional myodystrophin were injected with an adenoviral vector carrying the microdystrophin gene upon reaching 6 weeks of age, which roughly corresponds to early human childhood. A week later, the animals were divided into 3 groups, after which they were injected once a week for 32 weeks. One group was injected with an empty placebo solution, the second with a solution containing immune tolerance–inducing plasmids, and the third with the same plasmids, but with the microdystrophin gene embedded in them.
At the end of the 32-week period, the mice reached an age approximately equivalent to the age of majority of a human. A comparison of the muscle strength of animals from different experimental groups, as well as the number of dystrophin-producing muscle fibers in their musculature demonstrated significantly higher rates for the group receiving injections of plasmids carrying the microdystrophin gene. In addition, the blood of animals of this group contained fewer pro-inflammatory cytokines, as well as antibodies to immunogenic fragments of myodystrophin.
The authors note that they are at the very beginning of the path, but the results of their work indicate the possibility of inducing tolerance to a wide range of immunogenic proteins using plasmids carrying copies of the genes of these proteins. The possibility of using this approach to form tolerance to the insulin precursor in patients with type 1 diabetes mellitus and to myelin in patients with multiple sclerosis is already being studied in clinical studies. Apparently, the underlying concept is also effective in relation to gene therapy.
Article by Peggy P. Ho et al. Engineered DNA plasmid reduces immunity to dystrophin while improving muscle force in a model of gene therapy of Duchenne dystrophy published in the journal Proceedings of the National Academy.
Evgenia Ryabtseva, portal "Eternal Youth" http://vechnayamolodost.ru based on Stanford University School of Medicine: Scientists engineer way to prevent immune response to gene therapy in mice.