Give the body to repair
Generation R
Galina Kostina, "Expert" No. 12-2013
The current generation can count on the fact that in twenty years medicine will be able to update organs and tissues, correct genome breakdowns. The beginning of this year is replete with information about achievements in the field of cellular technologies. Russian scientist Konstantin Agladze, together with Japanese colleagues, grows a heart muscle from stem cells, which is rhythmically contracting before our eyes. There, in Japan, scientists are making an approach to an incurable eye disease – macular degeneration – with the help of cells obtained by reprogramming from the skin cells of the patients themselves.
This news inspires much more optimism than the tens of thousands of news about stem cells that we have heard over the past ten years. Everyone understood that in order to obtain any healthy cells of the body, we need those that appear in the embryo's body, that is, omnipotent. They can turn into cells of the heart muscle, bones, liver, skin, brain – there are more than 200 different types of them in our tissues. The ethical problems of using omnipotent embryonic stem cells have greatly hindered scientific research. There was another problem – it was difficult to plant someone else's stem cells or cells already differentiated in the right direction: the body rejects them, and all sorts of tricks are needed to make these cells function. Their own reprogrammed cells are much closer to the body.
A qualitative leap in this area occurred thanks to the Japanese scientist Shinyo Yamanaka. He was able to return adult cells to a state of pluripotence, when they can turn into different cells of the body. They were called induced pluripotent cells (iPS). "We are not yet fully aware of the revolutionary discovery made by Shinya Yamanaka in the field of stem cells, this is really a new era," his colleagues said after he was awarded the Asian "Nobel" ShawPrize in 2008, and after the Nobel Prize in 2012, and at the end of February 2013, after presentation of the new Breakthrough Prize in Life Sciences Foundation, established by Mark Zuckerberg, Yuri Milner and Sergey Brin.
The creation of induced pluripotent cells was named by Science magazine at the end of last year as one of the most important breakthroughs of the last decade. In the same list, another landmark breakthrough is the technology of genome editing. Genetic diseases are caused by breakdowns in one or more genes. Scientists have learned how to get into the nucleus of a cell and use genome editing technology to change a damaged gene to a healthy one. Since a healthy gene must work in a certain tissue, the combination of cellular and genetic methods seems to be the most promising for researchers: for example, skin cells are taken from a patient, reprogrammed into iPS, then the genome is edited in them, and after that the corrected cells are turned, say, into neurons and injected into a certain part of the brain.
Racing from Labs to hospital bedHow wonderful that Yamanaki did not turn out to be a good surgeon, which he tried to become after graduating from Kobe University.
"They called me Dzyamanaka (from the word "dzyama" – an annoying nuisance)," the scientist recalled. – Operations that took twenty minutes for capable surgeons, lasted two hours for me." Desperate from his own lack of talent in the clinic, Yamanaka moved into science. He went to the USA, studied genetics there, returned to Japan, where he was also going to quit science, but at Nara University he was offered to take up the topic of stem cells, which he took up, although at first without much enthusiasm. However, the task of obtaining stem cells from mature adults, in particular from skin cells, fascinated the scientist so much that six years later he received the so–called induced pluripotent mouse cell for the first time in the world, and in 2007 - human iPS.
Discoveries in the field of genetics played an important role in Yamanaki's work: in particular, it was already known which genes work in embryonic omnipotent cells. The scientist's logic was simple: in an adult cell, it is necessary to include exactly those genes that work in the embryonic one. Yamanaka identified 24, in his opinion, the main ones and introduced them with the help of a special retroviral construct into the fibroblast (the precursor of the skin cell). Then he searched by iteration for the minimum number of genes that would keep the cell in a state close to embryonic. So he made a magical cocktail of four genes, which immediately began to be called the Yamanaki cocktail. Soon after, James Thomson of the University of Wisconsin-Madison created his cocktail of four genes, two of which were the same genes that Yamanaka used, but two more were different. Thomson also received induced pluripotent cells. "Similar studies are being conducted in many laboratories, including ours," says Professor Sergey Kiselyov, head of the laboratory of the Institute of General Genetics of the Russian Academy of Sciences, "because the search for the safest and most effective methods is underway. It is known that one of the genes used in Yamanaka's initial experiments is responsible for the development of tumors. However, even about those structures where this oncogene is not present, researchers are still talking with caution, they still need to be checked and checked for safety!" by American scientists from Scripps University in California under the leadership of Shen Dean, as well as a group of Robert Lanza from Advanced Cell Technology (ACT) in Santa Monica, California it seems that it was possible to reprogram mouse cells not with the help of genes, but with the help of their products – proteins, which should at least remove the problem of possible tumor formation.
Naturally, after such impressive successes of scientists, the race from laboratories to clinics starts: the market promises to be multibillion-dollar. Already this year, the world's first iPS clinical trials are to begin at the Center for Developmental Biology at the Riken Institute in Kobe. Masayo Takahashi, who collaborates with Yamanaka, will use them to treat age-related macular degeneration, in which retinal cells die and a person begins to go blind. This disease occurs in about 1% of the population over 50 years of age. The first study will involve six patients. A piece of skin the size of a peppercorn will be taken from their shoulder, fibroblasts will be isolated from there, reprogrammed into iPS, then with the help of specific factors they will turn into retinal cells, after which they will be transplanted into the affected area of the eye to replace the dead cells. These studies are expected all over the world: they will help determine how safe and effective such a technique can be, whether the transplanted cells will take root, whether tumors will arise. Numerous preclinical tests, according to Takahashi's assurances, showed that there were no tumors in mice and primates.
Robert Lanza from Advanced Cell Technology, commenting on this event, spoke out for caution. He does not imagine that the FDA would allow such studies to begin without a more massive evidence base of preclinical trials than the Japanese. Lanza plans to start clinical trials of platelets obtained by reprogramming, intended for the treatment of blood clotting disorders, this year. But first they will be administered to healthy people. Lanza's research is safer: platelets do not have a cell nucleus, they cannot divide, respectively, they cannot cause a tumor. Takahashi also explains that he did not accidentally choose an eye disease to begin with: the situation in the eye is easily controlled and in which case the problem is easily solved surgically. If these studies are successful, then the already developed methods for the treatment of various diseases can start. The same technology is being investigated in experiments to create not only healthy cells, but also various tissues and even organs.
It is clear that the Japanese are struggling to be pioneers in the field in which their compatriot has made a formal coup. The government allocates unprecedented investments for stem cells, this year – 21.4 billion yen. It is expected that the next ten years will be declared by the Japanese government as the "iPS decade" with a budget of about 90 billion yen.
Currently, the volume of the global regenerative medicine market is approximately 3.6 billion dollars, and by 2030, according to the forecasts of the Japanese government, it will reach more than 180 billion dollars. With the introduction of new regenerative technologies, treatment costs, according to the Japanese Ministry of Health, can be reduced by 60%, because with their help it will be possible to treat many diseases, including those that are not yet amenable to standard therapy.
You need to poke at the genes with your fingersUnderstanding the role of genes in the cell and the ability to manipulate them helped create iPS.
The same knowledge led to the idea of gene therapy. If the disease is associated with a mutation in some gene, then it is quite natural to want to replace it with a healthy one. Ideas about the possibility of introducing correct or healthy genes for the purpose of treatment have been expressed since the 1970s, after the epoch-making discovery of DNA. Since then, scientists have learned a lot: they can make constructs with the right right gene and a viral base that will "transport" the gene into the nucleus. Scientists use the property of the virus to penetrate into the nucleus, where, in fact, DNA is stored. But it did not reach the clinic. There were still too many problems. Firstly, although the virus was technologically deprived of the ability to reproduce in a cell, it could still cause unpredictable reactions of the body; secondly, the design could be embedded in any place of the genome and theoretically disrupt the work of other genes. And it could not be integrated anywhere at all and give no effect. There were numerous studies, mainly on animals. In exceptional cases, the use of not very proven gene therapy was allowed. The first such case occurred in 1990. William Andersen first used gene therapy for Ashanti de Silva, a girl who was not even five years old. She had a terrible, life–incompatible disease – congenital immunodeficiency - due to a defect in the gene encoding the enzyme adenosine deaminase (ADA). Children with such a diagnosis cannot resist infections, so for some time they live in a kind of sterile bubble. Andersen took bone marrow cells from Ashanti, implanted a healthy ADA gene in them, increased these cells in culture and introduced them into the girl's body. She had several such procedures. Now Ashanti is about thirty, she works, she has children. This experience has greatly inspired scientists, doctors, and the public. However, after a couple of deaths of patients to whom gene therapy was applied (although it was not proven that gene methods were to blame), researchers were asked not to rush to treat people.
Scientists were thinking about how to embed the right gene into the genome. For some time now it has become known that in nature such inserts occur, for example, at the conception of a child: by mixing, the chromosomes of mom and dad can exchange parts of DNA. But how to achieve such a directed exchange in adulthood? The discovery of certain proteins, which were called "zinc fingers", helped. "By the way, one of the authors of this discovery was our compatriot from the biotech company Sangamo BioSciences Fedor Urnov," says Sergey Kiselyov. "Scientists have shown that "zinc fingers" can easily stick to the corresponding sections of DNA." Using these qualities, the researchers synthesized many constructs with such proteins that could not only moor in certain places of the genome, but also cut out, for example, a damaged gene so that a healthy gene specially launched into the cell could be embedded in its place. However, this technology was quickly bought by another American company that sells such proteins for decent money. However, the researchers found a way out. "There are similar proteins of plant origin, the so–called talens," Kiselyov continues. – In addition, it turned out that their capabilities are much wider than those of “zinc fingers”, it is easier to work with them, and they can be more effective. Therefore, designs with talens allow for almost jewel-like editing of the genome in cells." By the way, Science magazine named this technology one of the ten scientific breakthroughs along with the creation of iPS.
In parallel, other methods were being searched for that could correct the work of broken genes. For example, Mitsuo Oshimura from Tottori University in 2011 proposed his technology of gene therapy for the treatment of a genetic disease – Duchenne myodystrophy, as a result of which the work of the muscular apparatus is disrupted. Oshimura makes an additional artificial chromosome containing the desired gene without mutation, inserts the chromosome into a stem cell and then launches it into the body. Oshimura published the results of his successful experiments on mice. "This technology can be good when you need to correct the work of some very large gene, for example, dystrophin. It is difficult to match "fingers" or talens to its colossal size," says Sergey Kiselyov. However, such methods are still aimed at replacing one gene, and this applies to about 10% of all genetic diseases. "The treatment of so–called multigenic diseases, which, in particular, include cancer, is a much more difficult task. Perhaps, to begin with, it can be solved not by replacing genes in the DNA itself, but by using genes that, without being embedded in the genome, will produce the proteins the body needs." Other diseases, not only genetic, can be treated with the same method. For example, the Neovasculgen drug for lower limb ischemia, created by Kiselyov's group and the Human Stem Cell Institute (HSC), is based on a design with an included gene encoding the synthesis of vascular endothelial growth factor. "Imagine water pipes that have flattened over time, have broken through somewhere," explains Sergey Kiselyov, "and they need to be repaired. Otherwise, houses will be left without water. So the legs are left without a good blood supply when the vessels are damaged." The images obtained during clinical trials clearly show how "Neovasculgen" stimulated vascular growth, which literally saved some patients from amputation. According to the ISHR, in Russia, the diagnosis of "chronic ischemia of the lower extremities" is made annually by about 140 thousand people, of whom 30-40 thousand are sentenced by the disease to the deprivation of limbs.
Neovasculgen was registered at the end of 2012 and became the third approved gene drug in the world. The first two – anticancer drugs created in China, work on the same principle. At the end of last year, Europe also "gave up": the gene therapy drug "Glibera" was registered there for the treatment of a rare and severe disease associated with a deficiency of lipoprotein lipase, as a result of which patients with this genetic disease are unable to digest fats. "Glibera" has become the world's fourth means of gene correction. There are about a hundred drugs for gene therapy in the phase of clinical trials, so far these are mainly drugs based on constructs with viruses, or drugs with genes that will not be embedded in the genome, but will, like "Neovasculgen", produce proteins.
"The first decade of the new century can really be called a turning point for cellular and gene technologies that allow treating both genetic and numerous other diseases associated with a lack of certain proteins in the body," Sergey Kiselyov believes. "And as for genetic diseases, the future is most likely for combined methods, when the genome will be corrected in induced pluripotent cells, and then these cells will differentiate into the desired type."
Portal "Eternal youth" http://vechnayamolodost.ru25.03.2013