Goat Lactoferrin: Slow hurdling

Goats rush to the marketGalina Kostina, "Expert" No. 14-2011

Scientists from the Institute of Gene Biology, having successfully solved the scientific task of obtaining valuable human protein from the milk of transgenic goats, are now trying to combine the technology created with public money with private industrial production

These goats did not even suspect that their names would be inscribed in the tablets of domestic biotechnology. Their names were Lac-1 and Lac-2, and these names denoted different genetic constructs artificially embedded in the genome of animals.

(Photos here are from the website of the Biotechnological Center of Transgenesis in the pharmaceutical industry "Transgenpharm" – VM.)

The goats were born in 2007, marking the completion of the first stage of the joint Russian-Belarusian project "Belprostransgen". Since then, their seed, carefully harvested, has manifested itself in several generations of transgenic animals. Most of the transgenic goats live in the Belarusian Zhodino, eight goats – in the Moscow region, four goats – in Stavropol, where a special storage for precious sperm carrying the gene of the human protein lactoferrin has been built. Lactoferrin is a multifunctional protein found in breast milk. With antibacterial, antiviral, antifungal, anti-inflammatory, immunomodulatory, antioxidant properties, it protects babies while they have not yet formed their own immune system.

The ideologist and one of the main executors of the project, Igor Goldman, director of the Transgenbank Institute of Gene Biology of the Russian Academy of Sciences, demonstrates test tubes with powder to everyone and waves his hand towards the window, behind which there is a large container with frozen milk of transgenic goats, where more than two tons have been stored. From this milk, scientists have learned to isolate pure protein, which is then dried, after which it is ready to become a product for the global lactoferrin market, estimated at about $ 6 billion. According to Goldman, the project participants have come close to the moment when a new technology can be introduced into industry. A private investor has already been found, but neither scientists nor a milkman investor can get an explanation of how the intellectual property of the Union state and private capital can merge.

They were given ten years and a lot of moneyIgor Goldman took up transgenesis in the distant eighties.

He colorfully tells how the first project almost failed due to the fact that he could not get the first transgenic rabbits. But he was entrusted with solving a task of national importance. I had to work day and night, and the rabbits appeared. Then came the transgenic sheep. After that, Goldman, alone and in cooperation with our and foreign scientists, participated in projects to create various transgenic animals: it turned out that he simply succeeded in embedding a gene construct into a fertilized egg, which was then returned to the female. Goldman laughed, explaining his luck with touching relationships with animals: they say, you need to look a goat in the eye to understand when to take an egg from her and when to return it with a new gene. He worked many times abroad, where he was persuaded to stay, but he stubbornly insisted that the technology for the production of valuable human proteins from the milk of transgenic animals appeared in Russia. With this idea, he trampled a lot of reception rooms – Chernomyrdin, Primakov, Kiriyenko, then went down to Luzhkov. He gave money, but not much. Goldman was really lucky when he got an appointment with the secretary of the Union state Pavel Borodin. Borodin was impressed, perhaps, not even by the scientific extreme, but by the scale of the project, and he gave the go-ahead. The project started in 2003. Goldman and the team were given five years, 50 million rubles to begin with and instructed to get the first animals with the lactoferrin gene by the end of the first stage.

The first stage of the project included genetic engineering, testing of gene designs on mice and obtaining primary goats. It took a lot of time to create and select not only optimal gene designs (at first there were 11 of them, two were left for experiments), but also goats themselves, because in Russia at that time goat breeding was completely undeveloped. "We needed goats, although it was possible to work with other animals, such as cows," explains Goldman. – Firstly, they are unpretentious; secondly, they give more milk with valuable protein per kilogram of live weight than cows; thirdly, they have a shorter reproduction cycle; fourthly, I was sure that Russia should develop goat breeding, unfairly forgotten, because goat's milk, in particular unlike cow's milk, it does not contain allergens."

Experiments were going on in Zhodino, goats regularly brought offspring, but scientists did not find transgenics among the goats. Goldman has already been reproached for spending money on a mythical project. But the stubborn Goldman was sure that everything would work out. And finally, in October 2007, two baby goats were born, in which human lactoferrin genes were identified. The Russian-Belarusian team could not breathe enough on them, Varnish-1 and Varnish-2 were looked after like crowned babies. No one was allowed to approach them – neither strangers, nor brothers and sisters with the usual genome. It was necessary to hold out for a year – until their puberty. Well, then run them to the goats and again anxiously wait for the offspring to appear, among which there should have been transgenics.

2008, despite the success, turned out to be nervous, it was necessary to run again – to extend the project, to guess whether there would be funding. To the delight of scientists, the project was extended for another five years and $ 20 million was subscribed to it. The second stage involved the reproduction of primary producing animals, the development of technology for the isolation of human lactoferrin from goat milk and the development of a formulation of medicines based on lactoferrin.

At the end of 2008, semen from Belarusian transgenics was brought to Moscow in a thermos. "In the Shakhovsky district, 120 specially trained queens were inseminated by a private farmer with whom we have an agreement," says Elena Sadchikova, Deputy director for the Belprostransgen project at the Institute of Gene Biology of the Russian Academy of Sciences. – We waited five months of pregnancy and at the beginning of 2009 we checked all the offspring for the presence of the lactoferrin gene. Seven goats and four goats were selected. The goats were sent to a sperm receptacle in Stavropol, and the goats were grown to a sexually mature state and covered. However, only four of the seven goats were covered. Two transgenic goats were again selected from their offspring, and the mothers' milk turned out to have a very good lactoferrin content – about 10 grams per liter. By the way, in the colostrum of a woman who gave birth to lactoferrin 5-7 grams per liter, and in milk – 1-2 grams."

In dietary supplements and medicinesIn newborns, lactoferrin of mother's milk is a key protein responsible for immunity in the first year of life, when the baby's own immune system has not yet formed.

Children who do not receive breast milk are much more likely to get sick than their infant peers. From one and a half to two million newborns in the world die a year from various infections. Many of them did not receive breast milk or were born prematurely. "In Italy, with the consent of mothers, studies of premature babies weighing from one to one and a half kilograms were conducted," says Igor Goldman. – Lactoferrin obtained from cows was added to their food. Even though cow lactoferrin is almost 30 percent different from human lactoferrin, the results were encouraging. According to statistics, about 20 percent of premature babies die from sepsis. All infants who took cow lactoferrin survived."

Lactoferrin from cow's milk is already widely used in infant formulas, various beverages, healthy food products, dietary supplements – mainly in Japan and South Korea. In these countries, dairy products are consumed very little and they try to restore valuable proteins in another way.

In Europe, cow lactoferrin is produced by several industrial enterprises, but not for its own market. This is explained by the phobias of Europeans regarding cow diseases and frequent cases of allergy to cow's milk. In particular, cow lactoferrin is one of those milk proteins that can cause an undesirable reaction in allergy sufferers. In addition, it is not identical to human lactoferrin: studies have shown that it binds worse to receptors, therefore, it is less effective. Lactoferrin in cow's milk is much less than in the milk of transgenic goats: 0.5 grams per liter versus 10 grams. Therefore, in many laboratories around the world, work is underway to obtain human lactoferrin. Human lactoferrin from the milk of transgenic cows and goats, from genetically engineered rice, as well as genetically engineered aspergillus mushrooms is currently being tested. According to some reports, cows still have a very low yield, and the mushroom has a slightly different form in which human protein is folded, so its functions are not so extensive. In general, no one has yet entered the industrial production of human lactoferrin.

Our scientists believe that they have already come to the moment when the technology can be put into production. "We are already cramped within the framework of this program," says Elena Sadchikova. – We have practically done everything we had to do by the end of 2013. Now, together with other scientists and physicians, we are investigating various properties of lactoferrin in order to then prepare recommendations for its use in medicine and formulations." A study was conducted with the Institute of Immunology, which showed high activity of lactoferrin against those strains of bacteria that are already insensitive to a huge number of antibiotics. And research by the Institute of Carcinogenesis of the Russian Cancer Research Center showed that lactoferrin, which was given to mice with vaccinated tumors, actively works against these tumors. Another study was conducted with periodontists for antibacterial activity. "This is a complex protein,– says Elena Sadchikova. "Its antimicrobial, antiviral, antioxidant, and immunomodulatory properties have already been proven, so demand may be high." Scientists plan to move in two directions of using lactoferrin – as food additives to children's, dietary, healthy nutrition, as well as as part of medicines against various infections. "The Americans, by the way, have recently discovered that lactoferrin removes radionuclides very well," Sadchikova adds.

The thorny wayIndustrial production, of course, is not included in the program of scientists.

Nevertheless, having spent almost ten years creating a lactoferrin production technology that can be used to produce other human proteins, they are afraid that at the end of the project a "tick" will simply be put. They also want to see a product that will come to market. Scientists have found an investor – the owner of a large Novgorod dairy company "Lactis" Ravil Dautov. He became interested in the project because, according to him, the competition in the traditional dairy market is tough enough and it would be nice to come out not only with a new product based on goat's milk, but also to develop an innovative lactoferrin production line. The farm "Nefedovskoe", owned by "Laktis", has almost completed the construction of a farm, where about a thousand goats will be imported. Considerable investments are required in this. A good breeding Russian goat costs about 20 thousand rubles, but Dautov wants to add French women to the herd, who are twice as large and, accordingly, give almost twice as much milk as Russian slim girls. But French women are almost twice as expensive. So, according to Dautov, about 230 million rubles may go to Nefedovskoye and another neighboring farm that joins it. Estimates of investments in the production of goat's milk and lactoferrin isolated from it are still the most approximate, since there is no ready-made equipment for the isolation of lactoferrin from the milk of transgenic goats. According to some estimates, it will cost at least 1 million euros. Ravil Dautov believes that lactoferrin will have a market. "Recently, Vladimir Vladimirovich Putin complained that more than half of our newborns do not receive breast milk," he says. "Having such a valuable technology that allows us to obtain human lactoferrin and add it to infant formula, it's somehow stupid to delay the production of this product." By the way, after hearing that Russia is going to produce human lactoferrin, Igor Goldman was immediately sent two applications for 2.5 tons from Europe and Asia.

However, in order to get an industrial product, legal issues need to be resolved. "While Dautov is buying ordinary goats, there are no questions for him," says Igor Goldman. – However, as soon as he launches a transgenic goat there, they will arise, because the transgenic goat is the intellectual property of the state. And when the milk goes, whose will it be?" When the joint Russian-Belarusian project began, it was decided that commercial issues would no longer be handled by the Union state, but by Russia and Belarus separately. But the program also says that the issue of the distribution of intellectual property will be decided after the end of the project at a meeting of the Union state, that is, in two years. And they want to move forward. Now scientists are busy preparing the registration of patents. But in order to issue a full package of patents, including for industrial production, it is necessary to create this industrial production. "In our case, there are too many subtleties associated, in particular, with the fact that the project was funded by both the Russian and the Union state,– Sadchikova explains. – As the lawyers explained to us, we may have obstacles to creating a small enterprise, including because the investor and I need to create a large production right away. According to lawyers, the most favorable way for us would be to create a public-private partnership, but it seems that this form is not very developed in the country yet. They explained to us that this requires a special decision of the Russian government." And while there is no clarity with legal issues, we have prepared an industrial production project to create a scientific and technological base for products based on human medicinal proteins obtained from the milk of producing animals for Skolkovo. "We have already received the status of a Skolkovo resident, which means that the project has passed a serious international examination," Sadchikova says. – In parallel with the lactoferrin project, we are engaged in new scientific developments: we aimed at regulatory proteins, monoclonal antibodies to fight cancer, which can be obtained from transgenic goats. It is known that a person needs about 200 proteins, currently 30 at most are in development. So there is an untilled field here." If this project is approved, researchers and investors will have not only new opportunities for scientific research and excellent benefits for production, but also a chance to deal with the legal difficulties of the lactoferrin project.

Portal "Eternal youth" http://vechnayamolodost.ru13.04.2011