How the medicine is created

Nitrogen, sterility and Chinese hamsters

Darina Gribova, "Nanometer"

The program "Innovation Workshops" of FIOP RUSNANO and Lomonosov Moscow State University summed up the results of the competition "My Laboratory". We are happy to publish the best works.

It is not easy to get into the production building of the biopharmaceutical company "Biocad" if you are not a doctor, a graduate student or the best employee of the month on an excursion. We were lucky: in dressing gowns and shoe covers, after a noisy air shower, the photographer and I find ourselves at the beginning of the production cycle – the cell bank preparation room.

Biocad is one of the few full–cycle companies in Russia. This means that innovative medicines are independently developed here from the molecule to the market. In St. Petersburg (more precisely, in the village of Strelna in the south of the city), in addition to production and sales office, there is a "brain" of the company – a number of laboratories. These are laboratories of molecular genetics, high-performance biotechnological methods, cellular technologies, as well as analytical methods where drugs are developed and tested.

"Now we have put on sale a new drug – the Russian biosimilar rituximab for the treatment of blood and lymph node cancer," says Dmitry Morozov, CEO of Biocada. – All the steps for this were planned in advance. Four years ago, we had to adopt a program, build a plant, and get a large number of approvals." This drug in combination with chemotherapy helps patients cope with a terrible cancer.

Irina Kovaleva, a specialist in the quality assurance department at production, is proud of her work and can tell you about all the stages of creating a medicine. "It is the substances of monoclonal antibodies that are used to treat oncological and autoimmune diseases. The producer of our monoclonal antibodies are the ovarian cells of Chinese hamsters. Why? Because mammalian cells are closest to human cells, and the antibodies they produce are as effective and safe as possible when administered to a patient," she explains in the cell bank preparation room. The bank is divided into two parts by cryo-storage; this way you can avoid the loss of all cells in emergency circumstances.

Up to a certain point, the cells are stored in frozen nitrogen, then the cryovials (frozen test tubes) are thawed: "Special boxes are opened, where there is one milliliter of cells in tiny test tubes. The whole procedure should take from two to five minutes, otherwise their viability will be lost." The tubes are heated at a temperature of 37 ° C, preserving the viability of more than 90% of the cells. The next stage of production begins.

I must say that all this happens in a room invisible to the audience: we watched the process on video. Moreover, we are separated from the rooms of class B cleanliness, which we cannot enter even after an air shower, by glass, a corridor for staff and again by glass. Cleanliness is taken seriously here: weekly samples are removed from the clothes and hands of the staff, flushes of water and air are checked to maintain constant sterility. Sterility is when there is not a single microorganism in the room that should not be there.

Further down the corridor is the seed preparation room. Here, in addition to a laminar flow cabinet for manual work with cells in an atmosphere of complete sterility, there is an incubator – an orange cabinet where nutritional supplements are added to one milliliter of cells and it is increased to two liters. Gradually – first up to 125 ml, then up to 500 ml and so on. A certain temperature, humidity and mixing speed are maintained in the incubator shaker.

The next stage is the fermentation hall. "This is the most important room, a large–scale production of the final substance takes place here," says Irina. "It all starts with a wave bioreactor, it's a platform that swings – along with a disposable sterile bag." The nutrient medium and the contents of the two-liter flask that was received earlier are pumped into the bag, and a gas mixture of air and carbon dioxide is also fed there to maintain the pH of the medium.

For about four days, the build-up takes place to the required concentration, and then everything is sent to a 250-liter bioreactor. In the same room there are five reactors for 1000 liters each, where, in turn, the cells are transplanted further. The scale and flexibility of this technology make it possible to produce several drugs at once.

After the cultivation cycle, the bag with the substance travels further into the room of pre-viral purification: it is also hidden from our eyes. There are two stages of purification of the target protein through special chromatographic columns; the first purification is on a sorbent with protein A, the second is on a multimodal sorbent CaptoAdhere. A culture liquid or protein solution is passed through the columns, selectively binding the product or impurities (for example, aggregates of antibodies, proteins of producer cells, DNA).

All the equipment here is dazzlingly clean, like in a surgical room. "Imagine that the bag for the wave bioreactor came with a hole – this is unacceptable," Irina shakes her head. – Samples are taken from us every day, and if we see that the batch does not fit, it is immediately rejected. We try not to work with such suppliers anymore." This is not surprising: the risks are too great, the cost of one such bag at the end is 175 million rubles.

"Our finished products are stored here. I have access... no," Irina laughs, putting the pass card to the warehouse door without any effect. She says that the Strelna plant is responsible only for the production of the substance, and the bottling line for vials is located in Petrovo-Dalny in the Moscow region, from where ready-made medicines come out.

In the other wing of the production building there is a central factory laboratory, in which the parameters of the cell culture process are controlled. Here a special automatic counter of Cedex cells allows you to figure out which of them are initially non-viable. Another device, a high chromatographic column, calculates how much protein is in the substance, the most important element for the drug.

We pass through the weighing room into the room for the preparation of nutrient media and additives. All of them are tested for bacterial endotoxins – these are substances that can cause a toxic reaction in humans, fever, fever, and which need to be disposed of at the early stages of inspections. "The yellow label means that the additives have arrived to us and are still in quarantine," Irina lists, "the blue label: the sample has been selected, it is taken from each jar. Green – can be used in production. And while there are no these three colors, you can't work with them. Everything is very strict with us."

After making a full circle around the perimeter of the production shop, we return to the air shower. "You can leave the shoe covers, hand over the dressing gowns," Irina commands. She, a specialist in the quality assurance department, is only 23 years old. A very young staff is a distinctive feature of Biocad and, according to Dmitry Morozov, one of the factors of the success of the enterprise at the junction of science and production. "The future is being created now, so I, as a manager, should have a young gang creating a future in which it will live in 5-10 years. We must invest in science, in young people and in modern equipment," he believes.

Finally, I ask the CEO which of the dozens of developments of the company he considers the most important at the moment. "Now we have passed the laboratory stage of creating a new product based on the PD-1 blocker – this is an antibody with which you can fight melanoma, a very aggressive tumor. This is one of the drugs that will be used in complex therapy, – Dmitry Morozov thinks. – I am also very concerned about breast cancer in women, and we are working a lot on this. It's not fair when young girls suffer and die from it."

About the author:
Darina Gribova is a freelance journalist, a graduate of St. Petersburg State University, Higher School of Journalism and Mass Communications (Faculty of Journalism), majoring in international journalism.

Now he is studying under the joint Master's program of St. Petersburg State University & Freie Universitat Berlin (Master's program "Global Communication and International Journalism", 1st year).

Portal "Eternal youth" http://vechnayamolodost.ru16.03.2015