Treatment of rheumatoid arthritis with basic disease-modifying antirheumatic drugs (PDMD)

In the treatment of rheumatoid arthritis, drugs are used to slow the progression of joint erosion. These are basic disease-modifying antirheumatic drugs (DMP), which are an important component of the overall treatment program. What are these drugs and how do they work?

Disease-modifying drugs act on the immune system to slow the progression of rheumatoid arthritis, which is where their name comes from. There are many different drugs that fall into the category of DMARDs, but some are the most commonly used:

Rheumatex (Methotrexate)- the main drug in the category of BPRP. It works in the same way as other drugs and is more effective in many cases. It is also relatively inexpensive and mostly safe. Like other PDOs, methotrexate has a number of side effects: it can cause stomach upset, it can be toxic to the liver or bone marrow, and it can affect pregnancy. On rare occasions, it causes breathing difficulties. Good circulation is essential when taking methotrexate. Concurrent use of folic acid can reduce some of the side effects. The most important advantage of methotrexate is that it can be used over a long period. The drug can also be administered to children.

Biological agents: Enbrel (etanercet), Humira (adalimumab), Kineret (anakinra), Orentia (abatacet), Remikad (infliximab), and Rituxan (rituximab). These are the newest drugs for the treatment of rheumatoid arthritis, administered subcutaneously or intravenously. They neutralize the activity of the immune system that damages the joints. When combined with methotrexate, these drugs help most people overcome the symptoms of rheumatoid arthritis. Studies have shown that these drugs have fewer side effects than other PDOs. One of the complications is an increased susceptibility to acute infectious diseases. These drugs can adversely affect the liver and blood conditions and should be used with caution in the presence of chronic heart ailments. Other possible side effects may only appear after prolonged use of the drugs.

Plaquenil (hydroxychloroquine) and Azulfidine(sulfasaline ) used for moderate rheumatoid arthritis. They are not as potent as other PDOs, but they have fewer side effects. In rare cases, Plaquenil has a negative effect on the eyes. Patients taking this drug should be examined by an ophthalmologist annually.

Minocin (Minocycline)- an antibiotic that can stop the inflammatory process in RA. Its effect appears after a few months. In other cases, it takes a year for the full range of side effects to appear. With prolonged use, minocycline can cause skin pigmentation.

Arava (leflunomide) acts like methotrexate and is more effective in combination with it. The drugs have similar adverse reactions. Arava can cause diarrhea, in which case it should be discontinued. Since Arava has a negative effect on the fetus, it is contraindicated in women during pregnancy.

Neoral (azathioprine) it is used for various diseases accompanied by inflammation, including rheumatoid arthritis. However, due to its negative effect on kidney function and other side effects, it is usually used to treat exacerbations of rheumatoid arthritis if other drugs are ineffective.

Imunar (azathioprine) used for various inflammatory conditions, including rheumatoid arthritis. The most common side effects are nausea and vomiting, sometimes stomach pain and diarrhea. Long-term use of azathioprine increases the likelihood of developing cancer.

DMARDs slow the rate at which rheumatoid arthritis progresses and help many people improve their quality of life. In some cases, remission may occur. Basically, drugs provide a slowdown in the rate of progression of the disease.

The use of a single PDRP or their combination can prolong the asymptomatic course of rheumatoid arthritis and alleviate the acute manifestations of the disease. Your joints will need less time to swing in the morning. At your next check-up, your rheumatologist may advise you that there are no new lesions on your last X-rays. Also, regular use of BPRP reduces the likelihood of a long-term destructive process in the joints.

Are the BPRPs safe? All FDPs are approved by the US Food and Drug Administration. Many people take these drugs with absolutely no side effects.

However, by treating the symptoms of rheumatoid arthritis, PDBMs affect the entire body, their powerful action, as a rule, causes some side effects. There are the following typical side effects of PDBM:

Stomach upset. DMARDs often cause nausea, sometimes vomiting, and diarrhea. These symptoms can be treated with other medicines. Complications also disappear as your body gets used to the drug. If the symptoms are uncomfortable, your rheumatologist will prescribe another remedy.

Liver dysfunction. This complication is less common than indigestion. You will need to have regular blood tests to check for liver damage.

Blood condition. DMARDs can cause immune system malfunctions and increase the risk of infectious diseases. It may also decrease the level of white blood cells that protect the body from infections. Low red blood cell count (anemia) increases fatigue. A simple test done regularly will help keep your red blood cells in check.

Date of publication of the article: 08.08.2016

Date of article update: 28.01.

Arthritis is the general name for a group of joint diseases of an inflammatory nature of various origins. Inflammation of one or several joints at the same time can be both an independent disease and a manifestation of systemic pathology of the body.

What is arthritis in the simplest sense? In simple terms, this is an inflammation of the cartilage, synovial membrane, capsule, joint fluid and other elements of the joint.

There are more than 10 types of arthritis (more about them - later in the article). The mechanism of development of different types of the disease is almost the same, with the exception of some nuances.

Pathology negatively affects the patient's quality of life, its main symptoms: pain syndrome, swelling and redness of the affected area, local temperature rise, restriction of movement, joint deformation. It becomes difficult for a person to carry out everyday activities, and in case of a severe course of the disease, even elementary movements. Chronic long-term arthritis often leads to partial or complete immobilization with the registration of a disability group.

Any type of arthritis is treatable(some types are treated better and easier, some are worse), especially at the present time (the article was written in 2016), when many treatment methods have been developed and successfully applied that allow you to effectively fight not only the symptoms of the disease, but also its cause and effect.

Arthritis can be treated by doctors of the following three specialties: rheumatologist, arthrologist, orthopedic traumatologist. If the inflammation of the joints has developed against the background of tuberculosis, syphilis, brucellosis or other infection, then the emphasis is placed on the treatment of the underlying disease, which is dealt with, respectively, by a phthisiatrician, infectious disease specialist or dermatologist-venereologist.

Below I will describe in detail the types, causes and symptoms of arthritis, talk about modern methods of diagnosis and treatment of the disease.

Types of arthritis

Grading arthritis by category	Views
Inflammatory arthritis	Psoriatic

	Rheumatic
	Rheumatoid
	Reactive
	Infectious
	Tuberculous
Degenerative arthritis	Traumatic
Degenerative arthritis	Osteoarthritis
Taking into account the cause and mechanism of development	Primary - ankylosing spondylitis, Still's disease, pseudogout, rheumatic, psoriatic, septic, juvenile arthritis, various types of specific infectious arthritis (viral, dysentery or gonorrheal).
Taking into account the cause and mechanism of development	Secondary - arise against the background of an underlying pathology, for example, a malignant tumor, osteomyelitis, autoimmune diseases, sarcoidosis, hepatitis, borreliosis, some diseases of the blood, lungs or gastrointestinal tract.
By the number of affected joints	Monoarthritis - isolated inflammation of only one joint, usually large
	Oligoarthritis - affection of no more than 3 joints
	Polyarthritis - inflammation of 3–6 of both large and small joints at the same time

By the nature of the changes occurring, arthritis is divided into:

inflammatory, which is characterized by the presence of inflammation,
degenerative, when at first there is a malnutrition of the cartilage, dystrophy, a change in the appearance of the affected joint, followed by its deformation.

Arthritis occurs in acute, subacute and chronic forms. An acute or subacute course is most characteristic of an inflammatory lesion, a chronic one for a degenerative-dystrophic one.

An acute inflammatory process is: serous, serous-fibrous, purulent in nature.

The most "harmless" inflammation with the formation and accumulation of serous (transparent) fluid in the synovial bag occurs with synovitis - inflammation of the joint membrane.

Particularly severe arthritis is purulent. With it, inflammation affects, in addition to the articular bag, the tissues adjacent to it, and pus appears in the articular fluid due to the active reproduction of pathogenic microorganisms. The development of a purulent process is fraught with the formation of a capsular phlegmon (when a purulent process captures the entire joint).

Causes of the disease

Common (main) reasons

Heredity;
trauma;
obesity;
metabolic disorders in the body;
frequent hypothermia;
infections;
irrational distribution of physical activity: either a long sitting position, or excessive physical activity;
acute bacterial, viral or fungal infections;
diseases of the nervous system;
autoimmune diseases.

Additional reasons

Joint surgery,
advanced age,
childbirth,
weakened immunity
vaccination,
allergy,
multiple abortions,
improper nutrition,
unfavorable ecological situation,
lack of minerals and vitamins.

Improper diet is the main cause of gouty arthritis

Causes of specific types of arthritis

(if the table is not fully visible, scroll to the right)

Types of arthritis	Causes
Traumatic	Injuries to the elements of the joint: bruises, fractures of the bones to be articulated, lacerations in the area of the joint, etc.
Vibrating	Regular excessive stress on the joints, forcing them to perform movements under heavy load
Reactive	Various infections caused by ureaplasma, chlamydia, mycoplasma, dysentery bacillus, clostridia, salmonella, influenza viruses, etc.
Rheumatoid	Not exactly established, but the likelihood of the influence of heredity is high; autoimmune diseases; herpes viruses (Epstein-Barr virus, herpes simplex, cytomegalovirus); hepatoviruses, retroviruses
Psoriatic	Infections
Psoriatic	Genetic and autoimmune mechanisms
Osteoarthritis	Insufficient nutrition of cartilage as a result of metabolic disturbances in the body
	Dysplasias - congenital anomalies in the development of joint elements
	Systemic diseases - scleroderma, lupus, etc.
	Hormonal Disorders
	Specific and nonspecific inflammation of the articular structures. The first - against the background of tuberculosis, gonorrhea, dysentery. The second - as an independent defeat without the participation of pathogens
	Defeat, destruction of joints with Perthes disease, osteochondritis
	Hemophilia is a hereditary bleeding disorder
Gouty	Heredity
	Violation of protein metabolism against the background of malnutrition with excessive consumption of food rich in special substances - purines (mackerel, herring, sardines, meat)
	Excess body weight

The development of rheumatoid arthritis is affected by the failure of the immune system. For some unknown reason, special cells of the immune system begin to "attack" the own tissues of the joints. As a result, autoimmune inflammation begins, proceeding with the growth of aggressive tissue with a tumor-like development, due to which ligaments, articular surfaces are damaged, cartilage and the bones underlying them are destroyed. This leads to the development of fibrosis, sclerosis, erosion, as a result - to contractures, subluxation, persistent immobility of the joint - ankylosis.

Typical symptoms

The leading symptom of arthritis is pain in one or more joints. At first, they are weak and practically do not affect the ordinary life of a person. Over time, the pain syndrome grows: the pain becomes wavy in nature, intensifying with movement, at night and closer to morning. The intensity of pain varies from mild to very strong, dramatically impeding any movement.

Secondary symptoms:

morning stiffness
swelling,
redness of the skin,
an increase in local temperature in the area of inflammation,
deterioration of the patient's motor activity,
limitation of his mobility,
the formation of persistent deformities of the joints.

Depending on the course of the process, the limitation of the functionality of the affected joints can be both mild and severe, with possible complete immobilization of the limb.

Let's consider the symptoms of some types of arthritis in more detail.

Traumatic arthritis

Traumatic damage to the articular elements is accompanied by an inflammatory reaction, and if pathogenic microbes have entered the cavity, then purulent inflammation of the articular fluid and bursa, gradually passing to the nearby joint tissues.

Rheumatoid Arthritis Symptoms

This type of arthritis is characterized by symmetrical lesions of the knee, wrist, elbow, ankle joints, as well as small joints of the fingers and toes. Inflammation of the hip, shoulder and spinal joints is less common, but also possible.

In an acute or subacute course of the disease, a person is disturbed by sharp pains in muscles and joints, severe weakness, fever, stiffness in small joints in the morning.

A chronic sluggish process occurs with mild pain, a gradual increase in articular changes, which are usually not accompanied by significant limitation of the functions of the limbs.

Gradually, the inflammation spreads to the muscles adjacent to the joint. As a result, their focal inflammation develops, muscle strength and their tone decrease, the patient feels muscle weakness, severe fatigue after normal physical exertion.

A typical symptom is the appearance of subcutaneous nodules of a round shape with a diameter of no more than 2 cm. They can also form on the valves of the heart and in the lungs.

This type of disease is characterized by the asymmetry of the defeat of 2 or 3 joints at the same time. And first, the small joints of the toes and hands become inflamed, then the large ones - the knees, elbows, shoulders, etc.

The development of oligoarthritis (inflammation of no more than 3 joints) is accompanied by inflammation of the membranes around the tendons, an increase in the temperature of the inflamed area and redness of the skin, swelling and soreness of the joints.

The pain syndrome is expressed at rest or at night, morning stiffness and soreness disappears during the day.

Diagnostics

The establishment of an accurate diagnosis is based on a set of clinical manifestations, data from a doctor's examination and the results of laboratory diagnostics confirming the presence of arthritis (diagnostic data also help to determine the type, stage, and degree of activity of the process).

During examination with visual examination and palpation of disturbing joints, the doctor notes swelling, redness of the skin, which is hot to the touch; with a neglected disease, there is a visible deformation of the joint.

The table below shows the specific types of tests that need to be done if arthritis is suspected:

(if the table is not fully visible, scroll to the right)

Laboratory diagnostic methods	Instrumental diagnostics methods
Clinical blood test	X-ray of the joint in 2 projections
"Biochemistry" of blood (indicators - uric acid, sialic acids, protein fraction, CRP, fibrin, haptoglobin, etc.)	Digital Microfocus Radiography is a direct magnification X-ray image, while the digital imaging system provides high-definition images. The method allows you to detect minimal changes in bone structures
Rheumatoid factor	Arthrography - taking an X-ray after a contrast agent is injected into the joint cavity
Antistreptolysin-O	Ultrasound of the affected joints
Cytological and microbiological examination of synovial fluid	Scintigraphy - obtaining a two-dimensional image of the pathological area after the introduction of a radioactive isotope into the body
If necessary, a biopsy of the articular membrane is performed and then examined	Diagnostic arthroscopy is a highly informative method of examining joint structures through an arthroscope with a miniature video camera

Treatment methods

Any type of arthritis has several stages of development. For each, certain methods of treatment are selected: for the first and second, conservative therapy is sufficient, for the third and in the presence of complications, surgical intervention may be required.

The table shows a general treatment regimen for arthritis.

(if the table is not fully visible, scroll to the right)

Treatment methods	Details

Drug therapy	Non-steroidal anti-inflammatory drugs by mouth, intramuscularly and / or intra-articularly.
Drug therapy	Oral and intra-articular corticosteroids.
Efferent therapy	Cryoapheresis is a medical technique based on the treatment with cold or special chemicals of the plasma taken from the patient. Then it is injected back to the patient.
Efferent therapy	Plasma cascade filtration (plasmapheresis) is the purification of plasma from toxins, antibodies, hormones, and other substances, the level of which in the body is sharply increased.
Physiotherapy and massage (after the acute inflammatory process subsides)	Amplipulse therapy, phonophoresis, electrophoresis, magnetic and laser therapy, applications with ozokerite and paraffin, UFO, UHF.
Physiotherapy	Exercises of exercise therapy are aimed at preventing functional disorders and the development of contractures.
Surgery	Types: arthrotomy, excision of the synovial membrane (synovectomy), arthrodesis, joint resection, medical arthroscopy, cheilectomy. When the joint is destroyed, reconstructive arthroplasty or arthroplasty (joint replacement) is indicated.

Arthritis treatments

The treatment methods for different types of arthritis are very similar, the differences are only in some specific nuances, for example:

With specific arthritis, the underlying disease is treated (with tuberculosis, the emphasis is on anti-tuberculosis drugs).
To reduce the activity of psoriatic arthritis, the above methods are supplemented with ultraviolet or laser irradiation of blood, hemosorption. And from physiotherapy, PUVA therapy is effective, combining the ingestion of a special photosensitizing drug with external exposure to long-wave ultraviolet rays.

Summary

Only by scrupulously following the doctor's recommendations can you defeat arthritis. The prognosis is usually favorable, but it completely depends on the timeliness of contacting a specialist and bringing the treatment to an end. Modern techniques make it possible to correct even the most neglected situation by performing an operation on the joint.

Owner and responsible for the site and content: Afinogenov Alexey.

Classification

Basic antirheumatic drugs are divided into several groups:

Immunosuppressive drugs:
- Infliximab ("Remicade")
- Levamisole ("Decaris")
- Leflunomide ("Arava")
- Mercaptopurine ("Puri-Netol")
- Cyclosporine (Sandimmun Neoral)
- Cyclophosphamide - rarely used due to high toxicity
Gold preparations:
- Auranofin # ("Auropan", "Ridaura")
- Sodium aurothiosulfate # ("Sanocrizin")
- Aurotioprol # ("Chrysanol")
D-Penicillamine ("Cuprenil")
Derivatives of 5-aminosalicylic acid:
4-aminoquinoline derivatives:
- Chloroquine (Delagil)
- Hydroxychloroquine ("Plaquenil")
Matrix metalloproteinase inhibitors:

# - for 2015 the drug is not registered in Russia

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Excerpt characterizing Disease-modifying antirheumatic drugs

Magdalene knew that in order to fulfill Radomir's order, she had to feel confident, collected and strong. But while she only lived, locked in her deepest sorrow, and was madly lonely ...
Without Radomir, her life became empty, worthless and bitter ... He lived now somewhere far away, in an unfamiliar and wondrous World, where her soul could not reach ... And she missed him so insanely as a human, as a woman ! .. And no one, unfortunately, could help her in any way.
Then we saw her again ...
Magdalena sat alone on a high cliff, completely overgrown with wildflowers, pressing her knees to her chest ... She, as has already become customary, saw off the sunset - another regular day lived without Radomir ... She knew that there would be many more days like this ... so many. And she knew she would have to get used to it. Despite all the bitterness and emptiness, Magdalena understood well - a long, difficult life awaited her ahead of her, and she would have to live it alone ... Without Radomir. What so far she could not imagine, because he lived everywhere - in every cell of her, in her dreams and wakefulness, in every object that he once touched. It seemed that the entire surrounding space was saturated with Radomir's presence ... And even if she wished, there was no salvation from this.
The evening was calm, calm and warm. The nature reviving after the heat of the day raged with the smells of warmed up flowering meadows and pine needles ... Magdalene listened to the monotonous sounds of the ordinary forest world - it was surprisingly so simple, and so calm! .. Swollen by the summer heat, bees were loudly buzzing in the nearby bushes. Even they, hardworking, preferred to get away from the burning rays of the day, and now they were happily absorbing the invigorating evening chill. Feeling human goodness, the tiny colored bird fearlessly sat down on Magdalena's warm shoulder and in gratitude burst into sonorous silvery trills ... But Magdalene did not notice this. She again flew into the familiar world of her dreams, in which Radomir still lived ...
And she remembered him again ...
His incredible kindness ... His exuberant thirst for Life ... His bright affectionate smile and piercing gaze of his blue eyes ... And his firm confidence in the rightness of his chosen path. I remembered a wonderful, strong man who, being still a child, already subjugated whole crowds to himself! ..
I remembered his affection ... The warmth and loyalty of his big heart ... All this now lived only in her memory, not succumbing to time, not going into oblivion. It all lived and ... hurt. Sometimes it even seemed to her - just a little more, and she would stop breathing ... But the days passed. And life still went on. She was obliged by the DUTY left by Radomir. Therefore, as far as she could, she did not take into account her feelings and desires.
Her son, Svetodar, whom she missed madly, was in distant Spain with Radan. Magdalene knew - it was harder for him ... He was still too young to come to terms with such a loss. But she also knew that even in the deepest grief, he would never show his weakness to strangers.
He was the son of Radomir ...
And this obliged him to be strong.
Several months passed again.
And now, little by little, as happens even with the most terrible loss, Magdalene began to revive. Apparently, the right time has come to return to the living ...

Having chosen tiny Montsegur, which was the most magical castle in the Valley (as it stood at the "transition point" to other worlds), Magdalene and her daughter soon began to move there slowly. They began to settle in their new, still unfamiliar, House ...
And, finally, remembering Radomir's persistent desire, Magdalena gradually began to recruit her first students ... This was probably one of the easiest tasks, since every person on this wondrous piece of land was more or less gifted. And almost everyone was thirsty for knowledge. Therefore, very soon Magdalene already had several hundred very diligent disciples. Then this figure grew into a thousand ... And very soon the entire Valley of the Mages was covered by her teachings. And she took as many people as possible in order to escape from her bitter thoughts, and was unspeakably glad that the Occitanians were eagerly reaching for Knowledge! She knew that Radomir would have been happy about this from the bottom of his heart ... and recruited even more applicants.
- Sorry, Sever, but how did the Magi agree with this ?! After all, they are so carefully guarding their Knowledge from everyone? How did Vladyka allow this? Magdalene taught everyone, didn't she choose only initiates?
- Vladyka never agreed with this, Isidora ... Magdalene and Radomir went against his will, revealing this knowledge to people. And I still don't know which of them was really right ...

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Inflammatory rheumatic diseases, the main forms of which are rheumatoid arthritis (RA), diffuse connective tissue diseases (DBTD), systemic vasculitis, seronegative and microcrystalline arthropathies, are among the most severe forms of chronic human pathology. Pharmacotherapy of these diseases continues to be one of the most difficult problems of modern clinical medicine.

The etiology of many diseases is unknown, which makes it impossible to conduct effective etiotropic therapy. However, in deciphering their pathogenesis in recent years, there has been obvious progress, which is primarily due to the expansion of knowledge about the structural and functional features of the immune system, the mechanisms of development of the immune response and inflammation.

Currently, for the treatment of rheumatic diseases, a large number of drugs with different chemical structures and pharmacological mechanisms of action are used, the common property of which is the ability to suppress the development of inflammation. These include non-steroidal anti-inflammatory drugs, glucocorticoids with anti-inflammatory activity and the so-called basic antirheumatic drugs (gold salts, antimalarial drugs, cytotoxic drugs, etc.), which are believed to have a deeper effect on the immune system and the inflammatory processes underlying rheumatic diseases. New approaches to treatment based on the use of immunotherapeutic methods are being intensively developed.

In our country, several monographs have been published on the pharmacotherapy of rheumatic diseases (V.A.Nasonova, Ya. A. Sigidin. Pathogenetic therapy of rheumatic diseases, 1985; V.A. A. Sigidin, N. G. Guseva, M. M. Ivanova. Diffuse connective tissue diseases, 1994). However, in recent years, a very large amount of new clinical and experimental data has appeared concerning the mechanisms of action, tactics of use and effectiveness of both previously known antirheumatic drugs and new drugs and treatment methods.

The book systematically presents modern information about the most important anti-inflammatory drugs, but the main task was to get acquainted with new trends in the development of pharmacotherapy of inflammatory rheumatic diseases.

We hope that the book will be useful for practitioners in the treatment of patients with rheumatic diseases and stimulates interest in the pharmacological aspects of rheumatology among specialists involved in the development of theoretical problems of medicine, immunologists, biochemists, pharmacologists.

One of the most common and severe rheumatic diseases is RA, for the treatment of which the entire arsenal of antirheumatic drugs and methods of therapy is used (V.A.Nasonova and M.G. Astapenko, 1989). That is why classifications of antirheumatic drugs are being developed in terms of their place in the treatment of RA.

Based on differences in pharmacological properties, antirheumatic drugs are classified into anti-inflammatory analgesics (NSAIDs); anti-inflammatory glucocorticoids (GC), immunomodulatory / immunosuppressive agents (gold salts, antimalarial drugs, cytotoxic drugs, etc.). According to another classification, NSAIDs are considered symptomatic, not affecting the mechanisms of development of the disease, as opposed to disease-modifying or slow-acting antirheumatic drugs, which were believed to affect the etiopathogenesis of the disease.

To classify antirheumatic drugs, an approach was also used that takes into account, first of all, their toxicity, according to which they are subdivided into drugs of the first, second and third line. It was proposed to classify antirheumatic drugs based on the rapidity of the onset of the therapeutic effect and its duration after the cessation of treatment. NSAIDs and GCs, unlike disease-modifying / slow-acting antirheumatic drugs, show their effect very quickly (within a few hours or days). In addition, it was assumed that if, after the withdrawal of NSAIDs and GCs, the exacerbation develops rather quickly, then the effect of slow-acting antirheumatic drugs persists for a longer time.

However, it has now become apparent that traditional classifications do not meet modern requirements in terms of both terminology and subdivision into pharmacological categories. In fact, only NSAIDs and GCs are relatively homogeneous in terms of pharmacological and therapeutic activity of the group of drugs.

Since 1991, under the auspices of the WHO and the International League Against Rheumatic Diseases, a new classification of antirheumatic drugs has been created (H. E. Paulus et al., 1992; J. P. Edmonds et al., 1993), according to which these drugs are divided into two main categories:

I. Symptom-modifying antirheumatic drugs that have a positive effect on the symptoms and clinical manifestations of inflammatory synovitis:
1) non-steroidal anti-inflammatory drugs
2) glucocorticoids
3) slow-acting drugs: antimalarials, gold salts, antimetabolites, cytotoxic agents
II. Disease-controlling antirheumatic drugs that affect the course of RA, which must meet the following requirements:
a. improve and maintain the functional ability of joints in combination with a decrease in the intensity of inflammatory synovitis;
b. prevent or significantly reduce the rate of progression of structural changes in the joints.

In this case, the listed effects should appear for at least 1 year from the start of therapy; in the process of drug classification, the period should be indicated (at least 2 years) during which its therapeutic effect meets the listed criteria.

This classification differs from the previous ones in a more realistic approach to assessing the therapeutic efficacy of drugs in RA. It has now become obvious that the common proven property of all existing antirheumatic drugs is the ability to cause clinical improvement, while their ability to influence the progression and outcomes of the rheumatoid process cannot be considered strictly proven. Therefore, no antirheumatic drug currently can be classified as "disease control".

This, however, does not exclude the possibility of transferring certain drugs from the first group to the second in the process of further research. This position seems to be fundamental, since it should contribute to the expansion of pharmacological and clinical research in rheumatology in terms of the development of criteria for the effectiveness of treatment, as well as the creation of new, more effective antirheumatic drugs or their rational combinations.

E.L. Nasonov

Genetic engineering and drugs

Microbiological production of medicines

Before the advent of recombinant DNA technology, many drugs based on human proteins could be obtained only in small quantities, their production was very expensive, and the mechanism of biological action was sometimes poorly understood. With the help of new technology, a full range of such drugs is obtained in quantities sufficient both for their effective testing and for use in the clinic. To date, more than 400 genes have been cloned (mostly in the form of cDNA) of various human proteins that can become drugs. Most of these genes are already expressed in host cells, and their products are now being used to treat various human diseases. As usual, they are first tested on animals and then followed by rigorous clinical trials. The annual volume of the world market for medicines based on human proteins is about $ 150 billion and is constantly growing. The volume of the world market for drugs based on recombinant proteins is increasing by 12-14% per year and in 2000 amounted to approximately $ 20 billion.

On the other hand, the use of specific antibodies as therapeutic agents is promising. They are used to neutralize toxins, fight bacteria, viruses, and treat cancer. The antibody either neutralizes the "intruder" - a foreign agent, or destroys a specific target cell. Despite promising opportunities, antibodies are still rarely used to prevent and treat disease. And only with the development of recombinant DNA technology and the development of methods for producing monoclonal antibodies and with the deciphering of the molecular structure and function of immunoglobulins, commercial interest in the use of specific antibodies for the treatment of various diseases arose again.

The development of new methods for the prevention and treatment of many human diseases made a huge contribution to the growth of human well-being in the 20th century. However, this process cannot be considered complete. The so-called "old" diseases, for example, malaria, tuberculosis, etc., can make themselves felt again, as soon as preventive measures are weakened, or resistant strains appear. A typical situation in this respect is in Ukraine and Russia.

The first GMO products are antibiotics

Antibiotics include low molecular weight substances that differ in chemical structure. What these compounds have in common is that, being the products of the vital activity of microorganisms, in negligible concentrations they specifically disrupt the growth of other microorganisms.

Most antibiotics are secondary metabolites. They, like toxins and alkaloids, cannot be classified as substances strictly necessary for the growth and development of microorganisms. On this basis, secondary metabolites differ from primary ones, in the presence of which the death of the microorganism occurs.

The biosynthesis of antibiotics, like other secondary metabolites, usually occurs in cells that have stopped growing (idiophase). Their biological role in ensuring the vital activity of producer cells remains not fully investigated. Experts studying the prospects of biotechnology in the field of microbiological production of antibiotics believe that in unfavorable conditions they inhibit the growth of competing microorganisms, thereby providing more favorable conditions for the survival of the microbe-producer of one or another antibiotic. The importance of the process of antibiotic production in the life of a microbial cell is confirmed by the fact that in streptomycetes, about 1% of genomic DNA is accounted for by genes encoding antibiotic biosynthesis enzymes, which may not be expressed for a long time. The producers of known antibiotics are mainly six genera of filamentous fungi, three genera of actinomycetes (almost 4000 different antibiotics) and two genera of true bacteria (about 500 antibiotics). Of the filamentous fungi, special attention should be paid to the molds of the genera Cephalosporium and Penicillium, which are producers of the so-called beta-lactam antibiotics - penicillins and cephalosporins. Most actinomycetes that synthesize antibiotic substances, including tetracyclines, belong to the genus Streptomyces.

Of the known 5000-6000 natural antibiotic substances, only about 1000 are produced for sale to consumers. At the time when the antibacterial effect of penicillin and the possibility of its use as a drug were established (H.W. Flory, E.B. Chain et al., 1941 ), the productivity of the laboratory strain of mold - 2 mg of the preparation per 1 liter of culture liquid - was clearly insufficient for the industrial production of the antibiotic. Repeated systematic exposure of the original strain of Penicillium chrisogenum to such mutagens as X-ray and ultraviolet irradiation, nitrogen mustard gas in combination with spontaneous mutations and selection of the best producers, it was possible to increase the productivity of the fungus by 10,000 times and bring the concentration of penicillin in the culture liquid to 2%.

The way to increase the efficiency of antibiotic-producing strains, based on random mutations and which has become a classic, despite the colossal labor costs, is still used. This situation is due to the fact that an antibiotic, unlike a protein, is not a product of a specific gene; antibiotic biosynthesis occurs as a result of the combined action of 10-30 different enzymes encoded by the corresponding number of different genes. In addition, for many antibiotics, the microbiological production of which has been established, the molecular mechanisms of their biosynthesis have not yet been studied. The polygenic mechanism underlying the biosynthesis of antibiotics is the reason why changes in individual genes do not lead to success. Automation of routine techniques for analyzing the productivity of mutants makes it possible to study tens of thousands of functioning strains and thereby accelerates the selection procedure using the classical genetic technique.

The new biotechnology, based on the use of antibiotic superproducer strains, presupposes the improvement of the mechanisms of protection of the producer from the antibiotic synthesized by him.

Strains resistant to high concentrations of antibiotics in the culture medium show high productivity. This property is also taken into account when designing superproducer cells. Since the discovery of penicillin in the late 1920s, more than 6,000 antibiotics have been isolated from various microorganisms with different specificities and different mechanisms of action. Their widespread use in the treatment of infectious diseases has helped save millions of lives. The vast majority of major antibiotics have been isolated from the gram-positive soil bacterium Streptomyces, although fungi and other gram-positive and gram-negative bacteria also produce them. Every year, 100,000 tonnes of antibiotics are produced worldwide, worth an estimated $ S billion, including more than $ 100 million in antibiotics added to livestock feed as additives or growth accelerators.

Scientists are estimated to discover between 100 and 200 new antibiotics every year, primarily through extensive research programs to find thousands of different microorganisms that would synthesize unique antibiotics. The production and clinical trials of new drugs are very expensive, and only those of great therapeutic value and economic interest are marketed. They account for 1-2% of all detected antibiotics. The technology of recombinant DNA has a great effect here. First, it can be used to create new antibiotics with a unique structure that have a more powerful effect on certain microorganisms and have minimal side effects. Secondly, genetic engineering approaches can be used to increase the yield of antibiotics and, accordingly, to reduce the cost of their production.

We can assume that clinical biotechnology originated with the beginning of the industrial production of penicillin in the 40s. and its use in therapy. Apparently, the use of this first natural penicillin influenced the reduction of morbidity and mortality more than any other drug, but, on the other hand, posed a number of new problems that could be solved again with the help of biotechnology.

First, the successful use of penicillin caused a great need for this drug, and to satisfy it, it was necessary to dramatically increase the yield of penicillin during its production. Secondly, the first penicillin - C (benzylpenicillin) - acted mainly on gram-positive bacteria (for example, Streptococci and Staphylococci), and it was necessary to obtain antibiotics with a broader spectrum of action and / or activity, infecting both gram-negative bacteria such as E. coli and Pseudomonas. Thirdly, since antibiotics caused allergic reactions (most often minor, such as a skin rash, but sometimes more severe, life-threatening manifestations of anaphylaxis), it was necessary to have a whole set of antibacterial agents so that one could choose from equally effective drugs one that would not cause an allergy in the patient. Fourth, penicillin is unstable in the acidic environment of the stomach and cannot be administered orally. Finally, many bacteria acquire antibiotic resistance. A classic example of this is the formation by staphylococci of the enzyme penicillinase (more correctly, beta-lactamase), which hydrolyzes the amide bond in the beta-lactam ring of penicillin with the formation of pharmacologically inactive penicillinic acid. It was possible to increase the yield of penicillin during its production mainly due to the consistent use of a series of mutants of the original strain of Penicillium chrysogenum, as well as by changing the growing conditions.

The process of biosynthesis of one antibiotic can consist of dozens of enzymatic reactions, so cloning all genes of its biosynthesis is not an easy task. One of the approaches to isolating a complete set of such genes is based on the transformation of one or several mutant strains that are unable to synthesize this antibiotic with a bank of clones created from the chromosomal DNA of a wild-type strain. After the introduction of the clone bank into the mutant cells, the selection of transformants capable of synthesizing the antibiotic is carried out. Then, plasmid DNA of a clone containing a functional ex-premixing antibiotic gene (i.e., a gene that restores the function lost by the mutant strain) is isolated and used as a probe for screening another bank of chromosomal DNA clones of the wild-type strain, from which clones containing nucleotide sequences that overlap with the probe sequence. Thus, DNA elements adjacent to the complementing sequence are identified and then cloned, and the complete cluster of antibiotic biosynthesis genes is recreated. The described procedure refers to the case when these genes are grouped in one site of chromosomal DNA. If the genes for biosynthesis are scattered in the form of small clusters at different sites, then you need to have at least one mutant per cluster in order to obtain DNA clones, which can be used to identify the remaining genes of the clusters.

Using genetic or biochemical experiments, one can identify and then isolate one or several key biosynthetic enzymes, determine their N-terminal amino acid sequences, and, based on these data, synthesize oligonucleotide probes. This approach was used to isolate the isopenicillin N synthetase gene from Penicillium chrysogenum. This enzyme catalyzes the oxidative condensation of 5- (1_-a-aminoadipylN-cysteinyl-P-valine to isopenicillin N, a key intermediate in the biosynthesis of penicillins, cephalosporins, and cephalosporins.

New antibiotics with unique properties and specificity can be obtained by carrying out genetically engineered manipulations with genes involved in the biosynthesis of already known antibiotics. One of the first experiments, during which a new antibiotic was obtained, consisted of combining two slightly different pathways of antibiotic biosynthesis in one microorganism.

One of the Streptomyces plasmids, plJ2303, carrying a 32.5 kb fragment of S.coelicoior chromosomal DNA contains all the genes of the enzymes responsible for the biosynthesis of the antibiotic actinorodin from acetate, a member of the isochromanquinone antibiotic family. The whole plasmid and various subclones carrying parts of the 32.5 kb fragment (for example, plJ2315) were introduced either into the strain AM-7161 of Streptomyces sp.T synthesizing the related antibiotic medermicin, or into the strain B1140 or Tu22 S. violaceoruber synthesizing related antibiotics granaticin and dihydrogranaticin.

All of these antibiotics are acid-base indicators that give the growing culture a characteristic color, depending on the pH of the medium. In turn, the pH (and color) of the medium depends on which compound is being synthesized. Mutants of the parental strain S.coelicoior, which are unable to synthesize actino rodinum, are colorless. The appearance of color after transformation of the strain AM-7161 Streptomyces sp. or strains B1J40 or Tu22 S. violaceoruber plasmid carrying all or several genes encoding enzymes of actinorodin biosynthesis, indicates the synthesis of a new antibiotic Transformants strain AM-7161 Streptomyces sp. and strain-6 1140 S. violaceoruber containing plasmid pM2303 synthesize antibiotics encoded by both the plasmid and chromosomal DNA.

However, upon transformation of S. violaceoruber strain Tu22 with plasmid plJ2303, along with actinorodin, a new antibiotic, dihydrogranatirodine, is synthesized, and upon transformation of strain AM-7161 Streptomyces sp. PlJ2315 plasmid synthesizes another new antibiotic - mederrodin A.

Structurally, these new antibiotics differ little from actinorodin, medermicin, granaticin, and hydrogranaticin and are probably formed when an intermediate product of one biosynthetic pathway serves as a substrate for an enzyme in another pathway. When the biochemical properties of various biosynthetic pathways of antibiotics are studied in detail, it will be possible to create new unique highly specific antibiotics by manipulating the genes that encode the corresponding enzymes.

Development of new methods for obtaining modern polyketide antibiotics.

The term "polyketide" refers to a class of antibiotics that result from the sequential enzymatic condensation of carboxylic acids such as acetate, propionate, and butyrate. Some polyketide antibiotics are synthesized by plants and fungi, but most of them are formed by actinomycetes in the form of secondary metabolites. Before carrying out manipulations with genes encoding enzymes of biosynthesis of polyketide antibiotics, it was necessary to find out the mechanism of action of these enzymes.

Having studied in detail the genetic and biochemical components of erythromycin biosynthesis in Saccharopolyspora erythraea cells, it was possible to introduce specific changes in the genes associated with the biosynthesis of this antibiotic, and to synthesize erythromycin derivatives with other properties. First, the primary structure of the S. erythraea DNA fragment was determined. 56 kbp, containing the ery gene cluster, then erythromycin polyketide synthase was modified in two different ways. To do this, 1) the DNA region encoding beta-ketoreductase was removed, or 2) a change was made in the DNA region encoding enoyl reductase. These experiments made it possible to show experimentally that if a cluster of genes encoding enzymes of biosynthesis of a certain polyketide antibiotic is identified and characterized, then, by making specific changes in them, it will be possible to directionally change the structure of the antibiotic.

In addition, by cutting out and joining certain sections of DNA, it is possible to move polyketide synthase domains and obtain new polyketide antibiotics.

DNA technology to improve antibiotic production

With the help of genetic engineering, it is possible not only to create new antibiotics, but also to increase the efficiency of the synthesis of already known ones. The limiting factor in the industrial production of antibiotics using Streptomyces spp. often is the amount of oxygen available to cells. Due to the poor solubility of oxygen in water and the high density of the culture of Streptomyces, it is often insufficient, cell growth slows down, and the yield of the antibiotic is reduced. To solve this problem, it is possible, firstly, to change the design of bioreactors in which the culture of Streptomyces is grown, and secondly, using genetic engineering methods, to create strains of Streptomyces that use the available oxygen more efficiently. These two approaches are not mutually exclusive.

One of the strategies used by some aerobic microorganisms to survive in a lack of oxygen is to synthesize a hemoglobin-like product that can accumulate oxygen and deliver it to cells. For example, the aerobic bacterium Vitreoscilla sp. synthesizes a homodimeric heme-containing protein functionally similar to eukaryotic hemoglobin. The Vitreoscilla “hemoglobin” gene was isolated, inserted into the Streptomyces plasmid vector and introduced into the cells of this microorganism. After its expression, Vitreoscilla hemoglobin accounted for approximately 0.1% of all cellular proteins of S.coelicoior even when expression was carried out under the control of Vitreoscilla's own hemoglobin gene promoter, and not the Streptomyces promoter. The transformed S.coelicoior cells growing at a low content of dissolved oxygen (about 5% of the saturating concentration) synthesized 10 times more actinorodin per 1 g of dry cell mass and had a higher growth rate than nontransformed cells. This approach can also be used to provide oxygen to other microorganisms growing under oxygen-deficient conditions.

The starting material for the chemical synthesis of some cephalosporins - antibiotics that have a slight side effect and are active against many bacteria - is 7-aminocephalosporic acid (7ASA), which in turn is synthesized from the antibiotic cephalosporin C. Unfortunately, natural microorganisms capable of synthesizing 7ASA , has not yet been identified.

A new biosynthetic pathway for 7ACA was constructed by inserting specific genes into the plasmid of the fungus Acremonium chrysogenum, which normally synthesizes only cephalosporin-C. One of these genes was represented by the cDNA of the fungus Fusarium solani, encoding D-amino acid oxidase, and the other originated from the genomic DNA of Pseudomonas diminuta and encoded a cephalosporin acylase. In the plasmid, the genes were under the control of the A. chrysogenum promoter. At the first stage of the new biosynthetic pathway, cephalosporin-C is converted to 7-p- (5-carboxy-5-oxopentanamide) cephalosporic acid (keto-AO-7ACA) by amino acid oxidase. Part of this product, by reacting with hydrogen peroxide, one of the by-products, is converted to 7-beta- (4-carboxybutanamide) -cephalosporic acid (GL-7ACA). Both cephalosporin-C, keto-A0-7ACA, and GL-7ACA can be hydrolyzed by cephalosporinacylase to form 7ACA, but only 5% of cephalosporin-C is directly hydrolyzed to 7ACA. Therefore, both enzymes are required for the formation of 7ACA in high yield.

Interferons

In the late 70s - early 80s. XX century DNA technology for the first time began to attract the attention of the public and large investors. One promising biotech product was interferon, which was hoped at the time as a miracle cure against a variety of viral diseases and cancer. The isolation of human interferon cDNA and its subsequent expression in Escherichia coll was reported by all interested publications in the world.

Different approaches are used to isolate human genes or proteins. Typically, the desired protein is isolated and the amino acid sequence of the corresponding portion of the molecule determined. Based on this, the nucleotide sequence encoding it is found, the corresponding oligonucleotide is synthesized and used as a hybridization probe to isolate the desired gene or cDNA from genomic or cDNA libraries. Another approach is to generate antibodies to a purified protein and use them to screen libraries in which certain genes are expressed. For human proteins synthesized predominantly in one tissue, a cDNA library based on mRNA isolated from this tissue will be enriched in the target DNA sequence. For example, the main protein synthesized by the cells of the islets of Langerhans in the pancreas is insulin, and 70% of the mRNA isolated from these cells encode it.

However, the principle of cDNA enrichment is inapplicable for those human proteins, the amount of which is very small or the place of synthesis of which is unknown. In this case, other experimental approaches may be needed. For example, human interferons (IFs), including alpha, beta and gamma interferons, are natural proteins, each of which can find its own therapeutic use. The first interferon gene was isolated in the early 1980s. XX century. Since then, several different interferons have been discovered. A polypeptide with the action of human leukocyte interferon is synthesized in E. coli.

Several features of interferon made the isolation of its cDNA particularly difficult. First, despite the fact that interferon was purified more than 80,000 times, it could only be obtained in very small quantities. its exact molecular weight was not known at the time. Secondly, unlike many other proteins, interferon does not have an easily identifiable chemical or biological activity: it was assessed only by the decrease in the cytopathic effect of the animal virus on the cell culture, and this is a complex and lengthy process. Thirdly, unlike insulin, it was not known whether there are human cells capable of producing interferon in sufficiently large quantities, i.e. whether there is a source of interferon mRNA. Despite all these difficulties, the cDNA encoding interferon was eventually isolated and characterized. When isolating their cDNA, a special approach had to be developed to overcome the difficulties associated with the insufficient content of the corresponding mRNA and proteins. Now this DNA extraction procedure is common and standard and for interferons is as follows.

1. From human leukocytes isolated mRNA and fractionated by size; performed reverse transcription and inserted into the Psti site of the pBR322 plasmid.

2. The resulting product was transformed into Escherichia coli. The resulting clones were divided into groups. Testing was carried out on clones, which made it possible to speed up the process of their identification.

3. Each clone clone was hybridized with a crude IF-mRNA preparation.

4. From the resulting hybrids containing cloned DNA and mRNA, mRNA was isolated and translated into a cell-free protein synthesis system.

5. Determined the interferoic antiviral activity of each mixture obtained as a result of translation. The groups that showed interferon activity contained a clone with cDNA hybridized with IF-mRNA.

6. Positive groups were divided into subgroups containing several clones and tested again. The subgrouping was repeated until a clone containing full-length human IF-cDNA was identified.

Since then, several different types of interferons have been discovered. The genes of several interferons were isolated and their effectiveness was shown in the treatment of various viral diseases, but, unfortunately, interferon did not become a panacea.

Based on the chemical and biological properties of interferon, three groups can be distinguished: IF-alpha, IF-beta and IF-gamma. IF-alpha and IF-beta are synthesized by cells treated with drugs of viruses or viral RNA, and IF-gamma is produced in response to the action of substances that stimulate cell growth. IF-alpha is encoded by a gene family of at least 15 non-allelic genes, while IF-beta and IF-gamma are encoded by one gene each. The IF-alpha subtypes have different specificities. For example, when testing the effectiveness of IF-elf-1 and IF-alpha-2 on a virus-treated bovine cell line, these interferons exhibit similar antiviral activity, while in the case of virus-treated human cells, IF-alpha-2 is seven times more active than IF- alpha 1. If the antiviral activity is tested on mouse cells, then IF-alpha-2 is 30 times less effective than IF-alpha-1.

Due to the fact that there is a family of interferons, several attempts have been made to create IFs with combined properties, using the fact that different members of the IF-alpha family differ in the degree and specificity of their antiviral activity. In theory, this can be achieved by connecting parts of the gene sequences of different IF-alphas. This will lead to the formation of a fusion protein with different properties than each of the original proteins. Comparison of the cDNA sequences of IF-alpha-1 and IF-alpha-2 showed that they contain the same restriction sites at positions 60, 92 and 150. After cleavage of both cDNAs at these sites and subsequent ligation of the fragments, several hybrid genes were obtained. These genes were expressed in E. coli, the synthesized proteins were purified and their biological functions were investigated. Testing the protective properties of hybrid IFs in mammalian cell culture showed that some of them are more active than parental molecules. In addition, many hybrid IFs induced the formation of 2 "-5" -oligoisoadenylate synthetase in control cells. This enzyme is involved in the synthesis of 2 "-5" -linked oligonucleotides, which in turn activate latent cellular endoribonuclease, which cleaves viral mRNA. Other hybrid IFs exhibited greater antiproliferative activity than the parent molecules in cultures of various human cancer cells.

A growth hormone

The strategy of constructing new proteins by changing functional domains or by directed mutagenesis can be used to enhance or weaken the biological property of a protein. For example, native human growth hormone (HGH) binds to both the growth hormone receptor and the prolactin receptor in different cell types. To avoid unwanted side effects during treatment, it is necessary to exclude the attachment of hGH to the prolactin receptor. Since the region of the growth hormone molecule that binds to this receptor only partially coincides in its amino acid sequence with the region of the molecule that interacts with the prolactin receptor, it was possible to selectively reduce the binding of the hormone to the latter. For this, site-specific mutagenesis was used, as a result of which certain changes occurred in the side groups of some amino acids (His-18, His-21, and Glu-174) - ligands for Zn 2+ ions required for high-affinity binding of hGH to the prolactin receptor. The modified growth hormone binds only to "its" receptor. The results obtained are of undoubted interest, but it is not yet clear whether the modified hGH can be used in the clinic.

Cystic fibrosis

The most common lethal hereditary disease among Caucasians is cystic fibrosis. In the United States, 30,000 cases of this disease have been identified, in Canada and European countries - 23,000. Patients with cystic fibrosis often suffer from infectious diseases affecting the lungs. Treatment of recurrent infections with antibiotics eventually leads to the emergence of resistant strains of pathogenic bacteria. The bacteria and the products of their lysis cause the accumulation of viscous mucus in the lungs, which makes it difficult to breathe. One of the components of mucus is high molecular weight DNA, which is released from bacterial cells during lysis. Scientists from the biotechnology company Genentech (USA) have isolated and expressed the gene for DNase, an enzyme that breaks down high-molecular-weight DNA into shorter fragments. The purified enzyme is injected as part of an aerosol into the lungs of patients with cystic fibrosis, it breaks down DNA, the viscosity of mucus decreases, which makes breathing easier. Although these measures do not cure cystic fibrosis, they do provide relief. The enzyme was recently approved by the US Department of Food, Drug, and Cosmetic and generated approximately $ 100 million in 2000 sales.

Another biotechnological product that helps patients is alginate lyase. Alginate is a polysaccharide synthesized by a variety of algae as well as soil and marine bacteria. Its monomeric units are two saccharides - beta-D-mannuronate and alpha-1-guluronate, the relative content and distribution of which determine the properties of a particular alginate. Thus, the residues of a-L-guluronate form interchain and intrachain crosslinks by binding calcium ions; residues of beta-D-mannuronate bind ions of other metals. Alginate containing such crosslinks forms an elastic gel, the viscosity of which is directly proportional to the size of the polysaccharide molecules.

The release of alginate by mucous strains of Pseudomonas aeruginosa significantly increases the viscosity of mucus in patients with cystic fibrosis. To clear the airways and alleviate the condition of patients, in addition to treatment with DNase, depolymerization of alginate should be carried out using alginate lyase.

The alginate lyase gene was isolated from Flavobacterium sp., A gram-negative soil bacterium that actively produces this enzyme. On the basis of E. coli, a bank of Flavobacterium clones was created and those that synthesize alginate lyase were screened by plating all clones on a solid medium containing alginate with the addition of calcium ions. Under these conditions, all of the alginate in the medium, except for the one that surrounds the alginate-lyase-producing colonies, forms crosslinks and becomes cloudy. Hydrolyzed alginate loses its ability to form crosslinks, so the medium around the colonies synthesizing alginate lyase remains transparent. Analysis of the cloned DNA fragment present in one of the positive colonies showed the presence of an open reading frame encoding a polypeptide with a molecular weight of about 69,000. More detailed biochemical and genetic studies have shown that this polypeptide appears to be a precursor of three alginate lyases produced by Flavobacterium sp. First, some proteolytic enzyme cuts off an N-terminal peptide weighing about 6,000 from it. The remaining protein, with a molecular weight of 63,000, is capable of depolymerizing alginate produced by both bacteria and algae. When it is subsequently cut, a product with a molecular weight of 23,000 is formed, depolymerizing alginate of algae, and an enzyme with a molecular weight of 40,000 that destroys bacterial alginate. To obtain large amounts of the enzyme with a molecular weight of 40,000, the DNA encoding it was amplified by polymerase chain reaction (PCR) and then inserted into a plasmid vector isolated from B. subrjlis carrying the gene encoding the signal peptide of B. subrjlis a-amylase. Transcription was controlled using the penicillinase gene expression system. When B. subrjlis cells were transformed with the obtained plasmid and plated on a solid medium containing alginate with the addition of calcium ions, colonies with a large halo were formed. When such colonies were grown in liquid medium, the recombinant alginate lyase was released into the culture medium. Subsequent tests showed that this enzyme is able to effectively liquefy alginates synthesized by mucous strains of P. aeruginosa, which were isolated from the lungs of patients with cystic fibrosis. Further research is needed to determine whether clinical testing of recombinant alginate lyase is appropriate.

Prevention of organ transplant rejection

In the 1970s. views on passive immunization were revised: it began to be considered a prophylactic means of combating the rejection of transplanted organs. It was proposed to inject patients with specific antibodies that will bind to a certain type of lymphocytes, reducing the immune response against the transplanted organ.

Mouse monoclonal antibodies OCTZ were the first substances recommended by the US Department of Food, Drug, and Cosmetic for use as immunosuppressants in human organ transplantation. The so-called T cells, lymphocytes differentiating in the thymus, are responsible for organ rejection. OCTZ binds to a receptor on the surface of any T cell called CD3. This prevents the development of a complete immune response and rejection of the transplanted organ. This immunosuppression is very effective, although it has some side effects, such as fever and rash.

Techniques have been developed for the production of antibodies using E. coli. Hybridomas, like most other animal cell cultures, grow relatively slowly, do not reach high densities, and require complex and expensive media. The monoclonal antibodies obtained in this way are very expensive, which does not allow their widespread use in the clinic.

To solve this problem, attempts have been made to create a kind of "bioreactor" based on genetically modified bacteria, plants and animals. For this purpose, gene constructs capable of encoding individual regions of antibodies were introduced into the host genome. For the effective delivery and functioning of some immunotherapeutic agents, a single antigen-binding region of an antibody (Fab or Fv fragment) is often sufficient, i.e. the presence of the Fc portion of the antibody is optional.

GM plants - producers of pharmaceuticals

Today, the prospects for agricultural biotechnology to provide such plants that will be used as medicines or vaccines look more real. It is hard to imagine how important this could be for poor countries, where conventional pharmaceuticals are still a novelty and traditional WHO vaccination programs are too expensive and difficult to implement. This line of research must be supported in every possible way, including through cooperation between the public and private sectors of the economy.

Among the genes whose expression in plants is considered exotic, the genes encoding the synthesis of polypeptides of medical importance are the most important. Obviously, the Calgene patent on the expression of mouse interferon in plant cells should be considered the first study carried out in this area. Later, the synthesis of immunoglobulins in plant leaves was shown.

In addition, it is possible to introduce into the genome of a plant a gene encoding the envelope protein (s) of a virus. By consuming the plant for food, people will gradually acquire immunity to this virus. In essence, this is the creation of medicinal plants.

Transgenic plants have a number of advantages over the culture of cells of microorganisms, animals and humans for the production of recombinant proteins. Among the advantages of transgenic plants, we note the main ones: the possibility of large-scale production, cheapness, ease of cleaning, the absence of impurities that have allergenic, immunosuppressive, carcinogenic, teratogenic and other effects on humans. Plants can synthesize, glycosylate, and assemble mammalian proteins from subunits. When eating raw vegetables and fruits that carry genes that code for the synthesis of vaccine proteins, oral immunization occurs.

One of the ways to reduce the risk of gene leakage into the environment, used, in particular, in the creation of edible vaccines, is to introduce foreign genes into chloroplasts, and not into nuclear chromosomes, as usual. It is believed that this method will expand the field of application of GM plants. Despite the fact that it is much more difficult to introduce the desired genes into chloroplasts, this method has several advantages. One of them is that foreign DNA from chloroplasts cannot get into pollen. This completely eliminates the possibility of uncontrolled transfer of GM material.

Using DNA technology to develop vaccines

A promising direction is the creation of transgenic plants carrying genes for proteins characteristic of bacteria and viruses that cause infectious diseases. When consuming raw fruits and vegetables carrying such genes, or their freeze-dried juices, the body is vaccinated. For example, when the gene for the non-toxic subunit of cholera enterotoxin was introduced into potato plants and when raw tubers were fed to experimental mice, antibodies to cholera pathogens were formed in their bodies. It is clear that such edible vaccines can be an effective, simple and inexpensive way to protect people and ensure food safety in general.

The development of DNA technology in recent decades has revolutionized the development and production of new vaccines. Using the methods of molecular biology and genetic engineering, antigenic determinants of many infectious agents were identified, genes encoding the corresponding proteins were cloned, and, in some cases, the production of vaccines based on the protein subunits of these antigens was launched. Diarrhea caused by infection with Vibrio cholerae or enterotoxigenic E. coli (Escherichia coli) is one of the most dangerous diseases with a high percentage of deaths, especially in children. The total number of cholera diseases in the world exceeds 5 million cases annually, as a result of which about 200 thousand people die. Therefore, the World Health Organization (WHO) pays attention to the prevention of diarrheal infections, in every possible way stimulating the creation of a variety of vaccines against these diseases. Cholera outbreaks are also found in our country, especially in the southern regions.

Diarrheal bacterial diseases are also widespread in farm animals and poultry, primarily in young animals, which is the cause of large losses on farms as a result of weight loss and mortality.

A classic example of a recombinant vaccine obtained using microorganisms is the production of hepatitis B surface antigen. The viral HBsAg gene was inserted into a yeast plasmid, as a result of which a viral protein was synthesized in yeast in large quantities, which, after purification, is used for injection as an effective vaccine against hepatitis (Pelre et al., 1992).

Many southern countries with a high incidence of hepatitis are vaccinating the population, including children, against the disease. Unfortunately, the cost of such a vaccine is relatively high, which prevents the widespread adoption of universal vaccination programs for the population in countries with low living standards. In response to this situation, in the early 1990s, WHO launched an initiative to create new technologies for the production of inexpensive vaccines against infectious diseases available to all countries of the world.

Ten years ago, the concept of using transgenic plants for the production of so-called "edible" vaccines was put forward. Indeed, if any edible organ of a plant synthesizes a protein-antigen with strong oral immunogenic properties, then when these plants are eaten in food, the protein-antigen will be digested in parallel with the production of corresponding antibodies.

The obtained tobacco plants carry the gene encoding the hepatitis B virus envelope antigen under the plant promoter. The presence of the antigen in the leaves of transgenic plants was confirmed by enzyme immunoassay. The similarity of the physicochemical structure and immunological properties of the resulting recombinant antigen and the antigen of human serum has been shown.

The identification of antibodies produced in plants has shown the possibility of assembling two recombinant gene products into one protein molecule, which is impossible in prokaryotic cells. Antibody assembly took place when both chains were synthesized with a signal sequence. Moreover, along with the possibility of introducing two genes into one plant, it is also possible to combine individual polypeptide chains synthesized in different transgenic plants into a complete protein during hybridization of these two plants. It is possible to introduce several genes on one plasmid.

Transgenic autoantigens producing plants can also be used for other autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, insulin-dependent diabetes, and even organ transplant rejection. Insulin-dependent diabetes is an autoimmune disease in which the insulin-producing cells of the pancreas are destroyed by their own cytotoxic T lymphocytes. Oral prophylactic consumption of significant amounts of immunogenic proteins can lead to prevention and significant delay in the onset of symptoms of autoimmune diseases. However, it is possible only in the presence of a significant amount of autoantigens. The proteins insulin and pancreatic glutamic acid decarboxylase (GAD65) are considered as oral vaccines for the prevention of insulin-dependent diabetes. Recently, Canadian biotechnologists have obtained transgenic potato plants that synthesize pancreatic decarboxylase of glutamic acid. When feeding mice predisposed to diabetes, both a decrease in the incidence of diabetes and the magnitude of the autoimmune response were noted.

The above results of genetic engineering development convincingly indicate the possibility of creating "edible" vaccines based on transgenic plants. Given that the development of vaccines for humans will take much longer and more rigorous health screening, it is to be expected that the first edible vaccines will be developed for animals. Animal studies will help to reveal the mechanisms of action of "edible" vaccines and only then, after a long study and comprehensive assessment, such vaccines can be used in clinical practice. Nevertheless, work in this direction is actively continuing, and the idea of using plants for the production of vaccines has already been patented in the United States, which indicates the commercial interest in these developments.

Despite these encouraging results, the development of commercial “edible” diarrhea vaccines requires further research. In the pathogenesis of the enterotoxic form of bacterial and cholera diarrhea, the primary is to enable bacteria to multiply in the small intestine. This process depends on the ability of Escherichia coli to adhere, which is due to the presence on the surface of bacterial cells of special filamentous formations of a protein nature - fimbria. On the walls of the small intestine of patients with diarrhea, significantly more bacteria are found than in the lumen of the same part of the intestine, which is associated with the presence of fimbrial adhesins in Escherichia coli - proteins that provide binding to receptors on the surface of the intestinal epithelium.

Even non-pathogenic strains of Escherichia coll, which contained a plasmid encoding adhesin synthesis, were able to colonize the intestine and cause diarrhea without producing enterotoxins. In this regard, it is likely that immunity against toxins alone will not be sufficient to prevent the pathogenic effects caused by Vibrio cholerae or Escherichia coli. It is possible that in order to overcome these effects, in addition to the antigens of enterotoxins, it will be necessary to express neutralizing epitopes of structural antigens, such as lipopolysaccharides, proteins of the outer membrane of bacteria, or adhesins associated with the fimbriae of these bacteria, which are responsible for binding to the intestinal mucosa. Recently, one such adhesin, FimH, has been successfully used to immunize mice against bacterial diarrhea.

Another important problem associated with the development of "edible" vaccines is the level of expression of heterologous antigen in plants. Since oral administration of the vaccine requires larger amounts of antigen than with parenteral administration, the amount of antigen synthesized in plants, which is now no more than 0.3% of the total soluble protein, should be increased. At the same time, the level of expression must be high enough to induce an immune response, but less than the level that induces tolerance to the antigen, as is the case with substances consumed with normal food. And since the immune response (immunogenicity against tolerance) can be antigen-specific, the expression levels for each potential antigen will have to be selected individually.

Experiments show that the level of expression of a heterologous antigen in plants can be increased by using tissue-specific promoters and enhancers, enhancers of transcription and translation, adding transport peptides, and also by changing the nucleotide sequence of the corresponding genes using codons preferred for plants. However, the question of which plants are best used and in which edible organ it is better to express the antigen requires further research, since various plants may contain substances that block or slow down the immune response or are simply toxic to humans and animals, such as alkaloids. in tobacco cells.

Health ABC - healthy foods

Achievements of scientific and technological progress have affected all spheres of human activity, from production to everyday life. For centuries, people have sought to free themselves from physical exertion, automating production, creating household appliances, etc. And, in general, they were released. As a result, the daily energy consumption of a person by the end of the XX century compared with its beginning decreased by 1.5-2 times.

Human health is determined mainly by hereditary predisposition (genetics) and nutrition. At all times, the creation of a food base has been the guarantee and basis for the prosperity of any state. Therefore, any state is interested in projects of prevention and health programs, improving the structure of nutrition, improving the quality of life, reducing morbidity and mortality. It is nutrition that closely connects us with the environment, and food is the material from which the human body is built. Therefore, knowledge of the laws of optimal nutrition allows you to ensure human health. This knowledge is simple and is as follows: consume as much energy as you spend. The energy value (caloric content) of the daily diet should correspond to the daily energy consumption. Another is the maximum variety of food, which will provide a variety of the chemical composition of nutrition to the physiological needs of humans for nutrients (about 600 items). The food consumed should contain proteins, fats, carbohydrates, vitamins, mineral salts, water, fiber, enzymes, flavoring and extractive substances, minor components - bioflavonoids, indoles, anthocyanides, isoflavones and many others. In case of insufficiency of at least one of these components, serious health problems are possible. And to prevent this from happening, a person's daily diet should include about 32 names of various foods.

The optimal ratio of nutrients entering the body contributes to the preservation of health and longevity. But, unfortunately, the majority of the world's population is characterized by a deficiency of the following nutrients: complete (animal) proteins; polyunsaturated fatty acids; vitamins C, B, B2, E, folic acid, retinol, beta-carotene and others; macro- and microelements: Ca, Fe, Zn, F, Se, I and others; dietary fiber. And excessive consumption of such animal fats and easily digestible carbohydrates.

The deficit of protein intake for the majority of the population is on average 20%, the content of most vitamins and microelements is 15-55% less than the calculated values of the need for them, and dietary fiber is 30% lower. Violation of the nutritional status inevitably leads to a deterioration in health and, as a consequence, to the development of diseases. If we take the entire population of the Russian Federation as 100%, then only 20% will be healthy, people in a state of maladaptation (with reduced adaptive resistance) - 40%, and in a state of pre-illness and disease - 20%, respectively.

Among the most common alimentary-dependent diseases are the following: atherosclerosis; hypertonic disease; hyperlipidemia; obesity; diabetes; osteoporosis; gout; some malignant neoplasms.

The dynamics of demographic indicators in the Russian Federation and Ukraine over the past 10 years is also characterized by extremely negative trends. Mortality is almost twice the birth rate, life expectancy is significantly inferior not only to developed countries ...

In the structure of the causes of death, the leading place is taken by pathologies of the cardiovascular system and oncological diseases - diseases, the risk of which, among other reasons, depends on nutritional disorders.

The shortage of food products in the world should also be taken into account. Over the 20th century, the world's population has increased from 1.5 to 6 billion people. It is assumed that by 2020 it will grow to 8 billion or more, depending on who counts and how. It is clear that the main issue is the nutrition of such a number of people. Despite the fact that agricultural production over the past 40 years, thanks to selection and improvement of agronomic methods, has grown on average 2.5 times, further growth seems unlikely. This means that the rate of production of agricultural food products in the future will increasingly lag behind the rate of population growth.

A modern man consumes about 800 g of food and 2 liters of water per day. Thus, in just a day, people eat more than 4 million tons of food. Already, the shortage of food products in the world exceeds 60 million tons, and the forecasts are disappointing ...

The solution to the problem of increasing food production using the old methods is no longer possible. In addition, traditional agricultural technologies are not renewable: over the past 20 years, mankind has lost over 15% of the fertile soil layer, and most of the suitable for cultivation has already been involved in agricultural production.

An analysis of the situation that has developed in recent years in the agro-industrial complex of Russia indicates a decrease in the living population and a fall in the production of all types of agricultural products by more than 1.5 times. With the remaining total volumes of natural and labor resources, the crisis caused a sharp deterioration in the use of arable land, a decrease in the productivity of agroecosystems, more than 30 million hectares of highly productive agrocenoses were withdrawn from circulation.

The measures taken so far to stabilize the situation on the agricultural market have proved ineffective and insufficient. And food imports have exceeded all reasonable limits and called food security into question.

Based on the importance of optimizing the structure of nutrition for the health of the nation, development and security of the country, a priority direction has been developed to improve the nutrition of the population of Russia: elimination of the deficit of complete protein; elimination of micronutrient deficiencies; creating conditions for optimal physical and mental development of children; ensuring the safety of domestic and imported food products; increasing the level of knowledge of the population in matters of healthy nutrition. The scientific basis of the modern strategy of food production is the search for new resources that provide the optimal ratio of the chemical components of food for the human body. The solution to this problem is primarily to find new sources of protein and vitamins.

For example, a plant containing a complete protein, which is not inferior to animal proteins in terms of a set of amino acids, is soy. The introduction of products from it into the diet allows you to compensate for the deficiency of protein, as well as various minor components, in particular, isoflavones.

One of the solutions to the food problem is the chemical synthesis of food products and their components, and some success has already been achieved in the production of vitamin preparations. A very promising and already used method for obtaining high-grade food products, as their enrichment with protein and vitamins in the process of technological processing, that is, the production of food with a given chemical composition.

Another way is to use microorganisms as individual components of food products, because the growth rate of microorganisms is a thousand times higher than the growth rate of farm animals and 500 times that of plants.

It is important that there is a possibility of directed genetic predetermination in microorganisms of their chemical composition, its improvement, which directly determines their nutritional value and prospects for use.

Thus, in the new century, food production cannot do without the use of high modern technologies and, in particular, without the use of biotechnology, the use of microorganisms to obtain food.

With the growing awareness of the importance of a healthy lifestyle, the demand for food products that do not contain harmful substances has increased. And here DNA technologists could not help but participate.

Above, we have already mentioned sugar beets, which produce fructan, a low-calorie substitute for sucrose. This result was obtained by inserting a gene from the Jerusalem artichoke into the beet genome, which encodes an enzyme that converts sucrose into fructan. Thus, 90% of the accumulated sucrose in transgenic beet plants is converted into fructan.

Another example of work on the creation of "functional nutrition" products is the attempt to create caffeine-free coffee. A team of scientists in Hawaii has isolated a gene for the enzyme xanthosine-N7-methyltransferase, which catalyzes the critical first step in the synthesis of caffeine in coffee leaves and beans. With the help of Agrobacterium, an antisense version of this gene was inserted into the tissue culture cells of Arabica coffee. Studies of transformed cells have shown that the level of caffeine in them is only 2% of normal. If the work on the regeneration and reproduction of transformed plants is successful, then their use will allow avoiding the process of chemical decaffeination of coffee, which will not only save $ 2.00 per kilogram of coffee (the cost of the process), but also preserve the taste of the drink spoiled in this way, which is partially lost during decaffeination. ...

Developing countries, where hundreds of millions of people are starving, are in particular need of improved food quality. For example, legumes grown all over the world lack some sulfur-containing amino acids, including methionine. Active attempts are now being made to increase the concentration of methionine in legumes. In GM plants, it is possible to increase the storage protein content by 25% (this has been done so far for some varieties of beans). Another example already mentioned is the beta-carotene-enriched “golden rice” obtained by Professor Potrikus of the Technical University in Zurich. Obtaining an industrial grade would be a remarkable achievement. Attempts are also being made to enrich rice with vitamin B, the lack of which leads to anemia and other diseases.

The work on improving the quality characteristics of crop products illustrates well the capabilities of modern DNA technologies in solving a wide variety of problems.

Food as medicine

The term "biotechnology" refers to a set of industrial methods that use living organisms and biological processes for production. Biotechnological methods are as old as the world - winemaking, baking, brewing, cheese making are based on the use of microorganisms and also belong to biotechnology.

Modern biotechnology is based on cellular and genetic engineering, which makes it possible to obtain valuable biologically active substances - antibiotics, hormones, enzymes, immunomodulators, synthetic vaccines, amino acids, as well as food proteins, to create new varieties of plants and animal breeds. The main advantage of using new approaches is a decrease in the dependence of production on natural resources, the use of ecologically and economically the most profitable methods of farming.

The creation of genetically modified plants makes it possible to speed up the process of breeding cultivated varieties many times over, as well as to obtain crops with properties that cannot be bred using traditional methods. Genetic modification of crops makes them resistant to pesticides, pests, diseases, reducing losses during cultivation, storage and improving product quality.

What is typical for the second generation of transgenic crops that are already being produced on an industrial scale? They have higher agrotechnical characteristics, that is, greater resistance to pests and weeds, and, consequently, higher yields.

From the point of view of medicine, important advantages of transgenic products are that, firstly, it was possible to significantly reduce the residual amount of pesticides, which made it possible to reduce the chemical load on the human body in an unfavorable environmental situation. Secondly, to give insecticidal properties to plants, which leads to a decrease in their infestation by insects, and this greatly reduces the infestation of grain crops by molds. It is known that they produce mycotoxins (in particular, fumonisins - natural contaminants of cereals) that are toxic to humans.

Thus, both the first generation and the second generation GM products have a positive effect on human health not only indirectly - through improving the environment, but also directly - through reducing the residual amount of pesticides and the content of mycotoxins. It is not surprising that the areas occupied by transgenic crops are increasing from year to year.

But now the greatest attention will be paid to the creation of third-generation products, with improved or modified nutritional value, resistant to climatic factors, soil salinity, as well as having a prolonged shelf life and improved taste properties, characterized by the absence of allergens.

For crops of the fourth generation, in addition to the above qualities, a change in plant architecture (for example, short stature), a change in the time of flowering and fruiting, will be characteristic, which will make it possible to grow tropical fruits in the middle lane, a change in the size, shape and number of fruits, an increase in the efficiency of photosynthesis, the production of nutrients with an increased level of assimilation, that is, better absorbed by the body.

Improving the methods of genetic modification, as well as deepening knowledge about the functions of food and about the metabolism in the human body, will make it possible to produce products intended not only to ensure adequate nutrition, but also to further improve health and prevent diseases.

Plants bioreactors

One of the promising areas of plant DNA technology is the creation of bioreactor plants capable of producing proteins required in medicine, pharmacology, etc. The advantages of bioreactor plants include the absence of the need for feeding and maintenance, the relative ease of creation and reproduction, and high productivity. In addition, foreign proteins do not induce immune responses in plants, which is difficult to achieve in animals.

There is a need to obtain a whole set of biologically active proteins, which, due to a very low level of synthesis in specific tissues or products, are not available for study by the mechanism of action, widespread use or determination of areas of additional use. These proteins include, for example, lactoferrin, which is found in small amounts in mammalian milk, blood leukocytes.

Human lactoferrin (hLF) is promising to be used as a dietary supplement and medicinal preparation for the prevention and treatment of infectious diseases of the gastrointestinal tract of young children, increasing the body's immune response in malignant and a number of viral (AIDS) diseases. The production of lactoferrin from cattle milk, due to its low content, leads to a high cost of the drug. With the introduction of cDNA of the lactoferrin gene into tobacco cells, a number of callus tissues were obtained, synthesizing truncated lactoferrin, the antibacterial properties of which were much stronger than the antibacterial properties of native lactoferrin. The concentration of this truncated lactoferrin in tobacco cells was 0.6-2.5%.

The genes are inserted into the plant genome, the products of which induce an immune response in humans and animals, for example, against the envelope proteins of causative agents of various diseases, in particular, cholera, hepatitis, diarrhea, as well as against antigens of the plasma membranes of some tumors.

Transgenic plants are created that carry genes that produce some hormones necessary for hormone therapy in humans, and so on.

An example of the use of plants to create vaccines is the work done at Stanford University. In this work, antibodies to one of the forms of cancer were obtained using a modernized tobacco mosaic virus, into which a hypervariable region of lymphoma immunoglobulin was inserted. Plants infected with the modernized virus produced antibodies of the correct conformation in an amount sufficient for clinical use. 80% of the mice that received the antibodies survived the lymphoma, while all the mice that did not receive the vaccine died. The proposed method makes it possible to quickly obtain patient-specific antibodies in an amount sufficient for clinical use.

The prospects for using plants for the production of antibodies are great. Kevin Uzil and co-workers showed that the antibodies produced by soy were effective in protecting mice from infection with the herpes virus. Compared to antibodies produced in mammalian cell cultures, antibodies produced by plants had similar physical properties, remained stable in human cells, and did not differ in their ability to bind and neutralize virus. Clinical trials have shown that the use of antibodies produced by tobacco effectively inhibited the multiplication of mutant streptococci that cause tooth decay.

The creation of a vaccine produced by potatoes against insulin-dependent diabetes was carried out. In potato tubers, a chimeric protein was accumulated, consisting of the B subunit of cholera toxin and proinsulin. The presence of the B subunit makes it easier for the cells to consume this product, which makes the vaccine 100 times more effective. Feeding tubers with microgram amounts of insulin to diabetic mice slowed down the progression of the disease.

Gene technologies in the fight against environmental pollution. Phytoremediation

By his actions, man intervened in the course of the evolutionary development of life on Earth and destroyed the existence of the biosphere independent of man. But he failed to abolish the fundamental laws governing the biosphere and free himself from their influence.

Reborn after the next cataclysm from the remaining foci, adapting and evolving, life, nevertheless, at all times had the main direction of development. It was determined by the law of historical development of Rulier, according to which, within the framework of the progress of life and the irreversibility of evolution, everything strives for independence from environmental conditions. In the historical process, such a striving is realized by increasing the complexity of the organization, which is expressed in the growing differentiation of structure and functions. Thus, at each successive turn of the spiral of evolution, organisms appear with an increasingly complex nervous system and its center - the brain. 19th century evolutionary scientists they called this direction of evolution "cephalization" (from the Greek "cephalon" - the brain) However, the cephalization of primates and the complication of their body ultimately put humanity as a biological species on the brink of extinction according to the biological rule of accelerating evolution, according to which the complication of the biological system means a reduction in the average duration of existence species and an increase in the rate of its evolution. For example, the average life span of a bird species is 2 million years, mammals - 800 thousand years, ancestral forms of man - 200-500 thousand years. The modern subspecies of man has existed, according to some ideas, only from 50 to 100 thousand years, but many scientists believe that his genetic potential and reserves are exhausted (Dlekseenko, Keisevich, 1997).

The ancestors of modern man set foot on the path that intensifies confrontation with the biosphere and leads to catastrophe about 1.5-3 million years ago, when they first began to use fire. From that moment on, the paths of man and the biosphere diverged, their opposition began, the result of which may be the collapse of the biosphere or the disappearance of man as a species.

Humanity cannot abandon any of the achievements of civilization, even if they are disastrous: unlike animals that use only renewable energy sources, and in quantities adequate to the biosphere's ability to self-reproduce biomass, humanity can exist using not so much renewable as non-renewable energy carriers and energy sources. New inventions in the field only reinforce this opposition.

One of the latest trends in the use of transgenic plants is their application for phytoremediation - cleaning of soils, water pounds, etc. - from pollutants: heavy metals, radionuclides and other harmful compounds.

Environmental pollution by natural substances (oil, heavy metals, etc.) and synthetic compounds (xenobiotics), often toxic to all living things, increases from year to year. How to prevent further contamination of the biosphere and eliminate its existing foci? One of the solutions is to use genetic technologies. For example, living organisms, primarily microorganisms. This approach is called "bioremediation" - biotechnology aimed at protecting the environment. Unlike industrial biotechnologies, the main goal of which is to obtain useful metabolites of microorganisms, the fight against pollution is inevitably associated with the "release" of microorganisms into the environment, which requires an in-depth understanding of their interaction with it. Microorganisms produce biodegradation - the destruction of hazardous compounds that are not a common substrate for most of them. Biochemical pathways for the degradation of complex organic compounds can be quite extensive (for example, naphthalene and its derivatives are destroyed by a dozen different enzymes).

The degradation of organic compounds in bacteria is most often controlled by plasmids. They are called degradation plasmids, or D-plasmids. They decompose compounds such as salicylate, naphthalene, camphor, octane, toluene, xylene, biphenyl, etc. Most D-plasmids were isolated in soil strains of bacteria of the genus Pseudomonas. But other bacteria also have them: Alcalkjenes, Flavobacterium, Artrobacter, etc. Plasmids that control resistance to heavy metals have been found in many pseudomonads. Almost all D-plasmids are said to be conjugative, i.e. are able to independently transfer into the cells of a potential recipient.

D-plasmids can control both the initial stages of degradation of an organic compound and its complete degradation. The first type is the OST plasmid, which controls the oxidation of aliphatic hydrocarbons to aldehydes. The genes contained in it control the expression of two enzymes: hydroxylase, which converts hydrocarbons into alcohol, and alcohol dehydrogenase, which oxidizes alcohol to aldehyde. Further oxidation is carried out by enzymes, for the synthesis of which the genes of the chromosomes are "responsible". However, most of the D-plasmids belong to the second type.

Mercury-resistant bacteria express the mer A gene, which encodes a protein for the transfer and detoxification of mercury. The modified mer A gene construct was used to transform tobacco, rapeseed, poplar, and Arabidopsis. In hydroponic culture, plants with this gene were extracted from the aquatic environment up to 80% of mercury ions. At the same time, the growth and metabolism of transgenic plants were not suppressed. Mercury resistance has been passed down through seed generations.

Upon the introduction of three modified constructs of the mer A gene into a tulip tree (Liriodendron tulipifera), plants of one of the lines obtained were characterized by a rapid growth rate in the presence of mercury chloride (HgCl 2) concentrations hazardous to control plants. Plants of this line absorbed and converted into a less toxic elemental form of mercury and evaporated up to 10 times more ionic mercury than control plants. Scientists believe that elemental mercury vaporized by transgenic trees of this species will immediately dissipate into the air.

Heavy metals are an integral part of land pollutants used in agricultural production. In the case of cadmium, it is known that most plants accumulate it in the roots, while some plants, such as lettuce and tobacco, accumulate it mainly in the leaves. Cadmium enters the soil mainly from industrial emissions and as an impurity in phosphorus fertilizers.

One of the approaches to reducing the intake of cadmium into the human and animal body can be the production of transgenic plants that accumulate a smaller amount of this metal in the leaves. This approach is valuable for those plant species whose leaves are used for food or for animal feed.

You can also use metallothioneins, small cysteine-rich proteins that can bind heavy metals. Mammalian metallothionein has been shown to be functional in plants. Transgenic plants expressing genes of metallothioneins were obtained, and it was shown that these plants were more resistant to cadmium than control plants.

Transgenic plants with the mammalian hMTII gene had 60-70% lower cadmium concentration in stems compared to control, and the transfer of cadmium from roots to stems was also reduced - only 20% of the absorbed cadmium was transported to the stems.

Plants are known to accumulate heavy metals by extracting them from soil or water. Phytoremediation is based on this property, subdivided into phytoextraction and rhizofiltration. Phytoextraction refers to the use of fast-growing plants to extract heavy metals from the soil. Rhizofiltration is the absorption and concentration of toxic metals from water by plant roots. Plants that have absorbed metals are composted or burned. Plants differ markedly in their storage capacity. Thus, Brussels sprouts can accumulate up to 3.5% lead (based on dry weight of plants), and its roots - up to 20%. This plant also successfully accumulates copper, nickel, chromium, zinc, etc. Phytoremediation is also promising for the purification of soil and water from radionuclides. But toxic organic compounds are not decomposed by plants; it is more promising to use microorganisms here. Although some authors insist on reducing the concentration of organic pollutants during phytoremediation, they are mainly destroyed not by plants, but by microorganisms living in their rhizosphere.

The symbiotic nitrogen fixer of alfalfa, Rhlzobium melitotj, has been integrated with a number of genes that decompose gasoline, toluine, and xylene contained in fuel. The deep root system of alfalfa allows cleaning the soil contaminated with oil products to a depth of 2-2.5 meters.

It should be remembered that most of the xenobiotics have appeared in the environment in the last 50 years. But in nature there are already microorganisms capable of utilizing them. This suggests that in populations of microorganisms, genetic events occur rather quickly, which determine their evolution, more precisely, microevolution. Since the number of xenobiotics in connection with our technogenic civilization is becoming more and more, it is important to have a general understanding of the metabolism of microorganisms and their metabolic capabilities. All this required the development of a new science - metabolomics. It is based on the fact that bacteria can acquire the ability to process new compounds as a result of mutations. As a rule, this requires several successive mutations or the insertion of new gene systems from those already existing in other types of microorganisms. For example, for the decomposition of a stable organohalogen compound, genetic information is needed, which is in the cells of various microorganisms. In nature, this exchange of information occurs due to horizontal gene transfer, and in laboratories, DNA technology methods taken from nature are used.

Further development of phyto- and bioremediation is a complex problem associated, in particular, with the use of plants and rhizosphere microorganisms. Plants will successfully extract heavy metals from the soil, and rhizosphere bacteria will decompose organic compounds, increasing the efficiency of phytoremediation, promoting plant growth, and plants - the development of microorganisms living on their roots.

Environmental pollution can be considered a disease of ecosystems, and bioremediation can be considered a treatment. It should be considered as the prevention of numerous human diseases caused by environmental pollution. Compared to other cleaning methods, this one is much cheaper. With diffuse pollution (pesticides, oil and oil products, trinitrotoluene, which contaminated numerous lands), there is no alternative to it. In cleaning the environment from pollution, it is important to correctly prioritize, minimizing the risks associated with this or that pollution, and taking into account the properties of a particular compound and its effect, first of all, on human health. There is a need for legislative acts and rules governing the introduction of GM microorganisms into the environment, with which there are special hopes for cleaning from any pollutants. Unlike industrial biotechnology, where it is possible to strictly control all parameters of the technological process, bioremediation is carried out in an open system, where such control is difficult. To a certain extent, this is always "know-how", a kind of art.

The advantage of microorganisms in the purification of oil products was fully demonstrated when, after the tanker disaster, 5000 m 3 of oil spilled into the sea off the coast of Alaska. About 1.5 thousand km of the coastline was contaminated with oil. The mechanical cleaning involved 11,000 workers and a variety of equipment (it cost $ 1 million per day). But there was another way: at the same time, nitrogen fertilization was introduced into the soil to clean the coast, which accelerated the development of natural microbial communities. This accelerated oil decomposition 3-5 times. As a result, the pollution, the consequences of which, according to calculations, could affect even after 10 years, was completely eliminated in 2 years, having spent less than 1 million dollars on bioremediation.

The development of bioremediation, technologies and methods of its application require an interdisciplinary approach and cooperation of specialists in the field of genetics and molecular biology, ecology, and other disciplines. Thus, the areas of use of genetic engineering are very diverse and extensive, and some of them are fantastic and at the same time very promising in terms of attainable results.

The study of the response of living organisms to environmental changes is extremely important for assessing the impact of these changes, especially those of anthropogenic origin, on biodiversity, the preservation of which is the most important task of human civilization.

According to the Organization for Economic Cooperation and Development (OECD), the potential market for bioremediation is more than $ 75 billion. The accelerated introduction of biotechnologies for environmental protection is due, in particular, to the fact that they are much cheaper than other cleaning technologies. According to the OECD, bioremediation is of local, regional and global importance, and both natural organisms and GMOs will increasingly be used for purification.

Biofuels

Given the limited reserves of fossil energy, special attention should now be paid to the possibility of using new types of fuel - methane, hydrogen, etc., as well as renewable energy sources. However, in the general energy balance, such environmentally friendly energy sources as the energy of the Sun, sea currents, water, wind, etc., can make up no more than 20% of their total production. In this situation, biomass is becoming one of the most promising renewable energy sources, the methods of using which are constantly being improved. At the same time, along with direct combustion, bioconversion processes are widely used, for example, alcohol and anaerobic fermentation, thermal conversions, gasification, pyrolysis, etc. used as a fuel additive instead of imported oil. For the same purpose, the exploitation of natural thickets of black vine, which occupies about 6 million hectares in the northeastern regions of the country, has begun.

If in India, China and some other countries, agricultural waste is utilized for the purpose of obtaining biogas, then in Sweden, Germany, Brazil, the USA, Canada, agricultural crops are specially grown for the production of fuel ethanol. An effective substitute for fossil fuels is rapeseed and rapeseed oil, the spring forms of which can be cultivated in Russia up to the Arctic Circle. Soybean, sunflower and other crops can also be a source of vegetable oils for biofuel production. Sugar cane is increasingly used to produce fuel ethanol in Brazil, and corn is used in the United States.

The energy output coefficient (the ratio of the total energy equivalent of useful products to all energy costs for its production) is 1.3 for sugar beet; forage grasses - 2.1; rapeseed - 2.6; wheat straw - 2.9. At the same time, due to the use of 60 centners of wheat straw as a raw material from each hectare, it is possible to obtain 10 thousand m 3 of generator gas, or 57.1 GJ.

Due to the rapid depletion of natural resources of oil, gas and coal in many countries, special attention is paid to the so-called oil-bearing plants - Euphorbia lathyris (euphorbia) and E.tirucallii from the euphorbia family (Kupharbiacea), containing latex, the composition of terpenes of which is close in their characteristics to high quality oil. At the same time, the yield of dry matter of these plants is about 20 t / ha, and the yield of an oil-like product under the conditions of Northern California (i.e., in a zone of 200-400 mm of precipitation per year) can reach 65 barrels of raw materials per 1 ha. Therefore, it is more profitable to grow plant substitutes for fossil fuels, since more than 3,600 petrodollars can be obtained from each hectare, which in grain equivalent will amount to 460 c / ha, i.e. 20 times the average wheat yield in the United States and Canada. If we recall the well-known US slogan "for every barrel of oil there is a bushel of grain", then at today's prices for oil, gas and grain, this means an exchange - 1 grain dollar for about 25 petrodollars. Of course, a barrel of oil will not replace a bushel of grain in the literal sense, and far from every zone will be able to cultivate these types of plants. But obtaining alternative fuels through targeted plant breeding also turns the technogenic and energy component of highly productive agrophytocenoses into a reproducible and environmentally friendly factor for intensifying crop production, and, of course, this is one of the most painless solutions for such states as Ukraine - to use plants on a large scale in as renewable resources, including energy (biodiesel, lubricants, etc.). For example, the production of winter rapeseed already provides a ratio of 1: 5 energy consumption to energy output.

GMOs and biodiversity

The fundamental moment of the modern stage of breeding is a clear understanding that the basis for its development, including the use of genetic engineering techniques, is biodiversity.

The evolution of the plant kingdom followed the path of multiplying the number of species and their "ecological specialization". This fact indicates the danger of a decrease in biological (genetic) diversity in the biosphere in general and in agroecosystems in particular. A sharp reduction in species and genetic diversity has reduced not only the resistance of crop production to the vagaries of the weather and climate change, but also the ability to more efficiently utilize solar energy and other inexhaustible resources of the natural environment (carbon, oxygen, hydrogen, nitrogen and other biophilic elements), which, as it is known to account for 90-95% of dry matter of phytomass. In addition, this leads to the disappearance of genes and gene combinations that could be used in the breeding work of the future.

One and the same area, emphasized Charles Darwin (1859), can provide more life, the more diverse the forms inhabiting it. Each cultivated plant species, in connection with its evolutionary history and specific work of the breeder, is characterized by its own "agroecological passport", i.e. the confinement of the size and quality of the crop to a certain combination of temperature, humidity, lighting, the content of mineral nutrition elements, as well as their uneven distribution in time and space. Therefore, a decrease in biological diversity in agricultural landscapes reduces, among other things, the possibility of differentiated use of natural resources, and, consequently, the implementation of differential land rent of I and II types. At the same time, the ecological stability of agroecosystems is weakened, especially in unfavorable soil, climatic and weather conditions.

The scale of the disaster caused by the defeat of potatoes by late blight and nematode, the catastrophic loss of wheat due to the defeat of rust, corn due to the epiphytoty of helminthosporiosis, the destruction of reed plantations due to viruses, etc. are known.

A sharp decrease in the genetic diversity of plant species cultivated at the beginning of the 21st century is clearly evidenced by the fact that out of 250 thousand species of flowering plants over the past 10 thousand years, man introduced into culture 5-7 thousand species, of which only 20 crops (14 of them belong to grains and legumes) form the basis of the modern diet of the world's population. In general, to date, about 60% of food products are produced due to the cultivation of several grain crops, and over 90% of human needs for food is provided by 15 species of agricultural plants and 8 domesticated animal species. So, of 1940 million tons of grain production, almost 98% falls on wheat (589 million tons), rice (563 million tons), corn (604 million tons) and barley (138 million tons). Of the 22 known types of rice (genus Oryza), only two are widely cultivated (Oryza glaberrima and O.sativa). A similar situation has developed with legumes, the gross production of the 25 most important species of which is only about 200 million tons. And most of them are soybeans and peanuts, cultivated mainly as oilseeds. For this reason, the variety of organic compounds in the human diet has significantly decreased. It can be assumed that for Homo sapiens, as one of the biological species, the need for high biochemical variability of food is recorded in the evolutionary "memory". Therefore, the upward trend in its uniformity can have the most negative consequences for health. Due to the wide spread of oncological diseases, atherosclerosis, depression and other diseases, attention is drawn to the lack of vitamins, tonic substances, polyunsaturated fats and other biologically valuable substances.

It is obvious that an important factor in the spread of a particular valuable culture is the scale of its use. Thus, the rapid increase in the area of soybeans and corn in the United States and other countries is due to the production of hundreds of names of the corresponding products. The task of diversification is very relevant for other crops as well (for example, high-quality beer began to be obtained from sorghum, whiskey from rye, etc.).

Increasing the area under crops of such valuable crops as buckwheat (Fagopyrum), which has high adaptive capabilities in various, including unfavorable environmental conditions, amaranth (Amaranthus), deserves more attention in terms of solving interrelated problems of healthy food and increasing the species diversity of agroecosystems, quinoa (Chenopodium quinoa), rapeseed, mustard and even potatoes.

With the development of geographical discoveries and world trade, the introduction of new plant species has become widespread. Written monuments testify, for example, that as early as 1500 BC. Egyptian Pharaoh Hatshepsut sent ships to East Africa to collect plants used in religious rites. In Japan, there is a monument to Taji Mamori, who, on the orders of the emperor, traveled to China to collect citrus plants. The development of agriculture has played a special role in the mobilization of plant genetic resources. From the history of the United States it is known that already in 1897 Niels Hansen arrived in Siberia in search of alfalfa and other forage plants capable of growing successfully in the arid and cold conditions of the prairies of North America. It is believed that it was from Russia during that period that such important fodder crops as fire, pigs, fescue, hedgehog, white bent, alfalfa, clover and many others were introduced to the United States. At about the same time, Mark Carleton was collecting wheat varieties in Russia, of which the Kharkov variety for a long period occupied more than 21 million acres annually in the United States and became the basis for durum wheat production in the Northern Plains zone (Zhuchenko, 2004).

The introduction of new plant species into the culture continues at the present time. In the Peruvian Andes, a variety of lupine (tarvi) was found, which was eaten by the ancestors of modern Indians, which surpasses even soy in protein content. In addition, tarvi is resistant to low temperatures, undemanding to soil fertility. Breeders managed to obtain tarvi forms containing less than 0.025% alkaloids versus 3.3% in the starting material. Economic species also include the Australian herb (Echinochloa lurnerana), which can be an excellent millet-like crop for very arid areas. Among promising crops, the species Bauhinia esculenta deserves attention, which, like Psophocarpus tetragonolobus, forms tubers, and its seeds contain more than 30% protein and fat. In very arid conditions, the species Voandzeia subterranea can be used, which is not only rich in protein, but also more drought tolerant than peanuts, and also better resists diseases and pests. For arid and barren oilseed lands, Cucurbita foetidissima from the Cucurbitaceae family is considered promising, and for saline pasture lands, some species of the Atriplex genus of the Chenopodiaceae family, which secrete excess salt through the leaves.

Currently, in many countries of the world, active breeding work is underway with the amaranth (Amaranthus), a forgotten culture of the Incas, the seeds of which, in comparison with the used cereal spike plant species, contain twice as much protein, including 2-3 times more lysine and methionine, 2-4 times more fat, etc. Lines of maize have been found that, due to the presence of bacteria Spirillum lipoferum on their roots, fix atmospheric nitrogen in the same amount as soybean plants. It was found that nitrogen-fixing bacteria also function on the roots of a number of tropical grass species, assimilating nitrogen no less actively than bacteria of the genus Rhizobium in legumes. Thus, it was possible to find species of tropical grasses capable of fixing up to 1.7 kg of nitrogen per day per hectare, i.e. 620 kg / year.

In many countries, including European ones, potatoes are the main source of vitamin C, as they are consumed in large quantities. It is known that the production of potatoes in the world is about 300 million tons.

At the same time, out of 154 known potato species, only one, Solanum tuberosum, is ubiquitous. It is obvious that in connection with the increased possibilities of breeding to increase the potential productivity of plants, as well as the need to increase the ecological stability of agrocenoses and the development of areas of little use for plant growing, the scale of human activity to introduce new plant species into culture will significantly increase. Ultimately, "unconscious" (Darwin's term) and conscious selection led to the fact that the adaptive potential of cultivated plants significantly differs from that of their wild ancestors, not only due to differences in the criteria of adaptability themselves, but also in its main components: potential productivity, resistance to abiotic and biotic stresses, the content of economically valuable substances.

Along with the preservation of the plant gene pool in reserves, wildlife sanctuaries and national eco-parks, i.e. in situ, the creation of “gene banks” or “germplasm banks” to ensure the safe preservation of ex situ collections will play an increasingly important role in the coming period. The initiator of the organization of the latter was N.I. Vavilov, who collected in VIR the largest bank of plant resources in the world at that time, which served as an example and a basis for all subsequent banks, and most importantly, more than once saved a number of countries from devastation and hunger (for example, thanks to the presence of resistance genes in the VIR genebank).

Thanks to the continuation of the ideology of N.I. Vavilov, by the end of the 90s, national and international collections of plants numbered over 6 million samples, including more than 1.2 million cereals, 400 thousand food legumes, 215 thousand fodder, 140 thousand vegetables, over 70 thousand .root crops. At the same time, 32% of the samples are kept in Europe, 25% - in Asia, 12% - in North America, 10% each - in Latin America and International Centers, 6% - in Africa, 5% - in the Middle East.

Holders of the largest in terms of quantity and quality of samples of genetic collections are the United States (550 thousand), China (440 thousand), India (345 thousand) and Russia (320 thousand). Along with the conservation of plant resources in genebanks, the creation of natural reserves of flora and fauna is becoming more widespread. Due to the dramatically increased integration of the world food market, the exchange of plant genetic resources between countries has also increased significantly. At the heart of these processes is the understanding that no country or region is self-sufficient in terms of the provision of genetic resources. The creation of national botanical gardens in a number of countries has greatly contributed to the mobilization of genetic resources. Among them, for example, the botanical garden, created in London in 1760 and constantly importing exotic plant species from colonial countries.

Currently, the International Council for Plant Genetic Resources (IBPGR) is coordinating work on the conservation of the plant gene pool in the world. Since 1980, the European Genetic Resources Cooperation Program has been implemented. An important role in this is also played by the FAO Commission on Plant Genetic Resources, decisions of international conferences, adopted in 1992, the Convention on Biological Diversity. At the same time, gene banks of different types function. Some of them support only one culture and its wild relatives, others - several crops of a certain soil and climatic zone; while some contain basic collections for long-term storage, while others are focused on meeting the needs of selection centers and research institutions. So, in the gene bank in Kew Gardens (England) only wild plants (about 5000 species) are stored.

The adaptive strategy of agricultural intensification puts forward qualitatively new requirements for the mobilization of the world's plant resources in terms of the collection, storage and use of the gene pool, including the introduction of new plant species into culture. Currently, over 25 thousand species of higher plants are under threat of complete destruction in the world, including in Europe - every third of 11.5 thousand species. Many primitive forms of wheat, barley, rye, lentils and other crops have been lost forever. Local varieties and weedy species disappear especially quickly. So, if in China and India in the early 50s. XX century. thousands of varieties of wheat were used, then already in the 70s - only dozens. At the same time, each species, ecotype, local cultivar is a unique complex of coadapted gene blocks created during long-term natural or artificial selection, which ultimately ensure the most efficient utilization of natural and anthropogenic resources in a particular ecological niche.

Understanding of the retrospective nature of the evolutionary "memory" of higher plants clearly indicates the need to preserve the species diversity of flora not only in gene banks and centers of genetic resources, but also in natural conditions, i.e. in a state of constantly evolving dynamic system. At the same time, the creation of genetic collections of genetic systems for the transformation of genetic information, including res-systems, mei-mutants, gametocidal genes, polyploid structures, different types of recombination systems, reproductive isolation systems, etc., deserves much more attention. It is clear that they can be essential for development breeding of the future using genetic engineering technologies. It is also important to identify and preserve the genetic determinants of the formation of stable homeostatic systems, synergistic, cumulative, compensatory and other coenotic reactions that provide ecological "buffering" and dynamic equilibrium of the biocenotic environment. Such genetically determined plant traits as competitiveness, allelopathic and symbiotic interactions, and other environment-forming effects realized at the biocenotic level also deserve more attention. Particular attention should be paid to plant species with constitutive resistance to environmental stressors. It is known that in the second half of the XX century. in a number of countries, the area under this type of crops has increased significantly (sometimes by 60-80 times).

Currently, over 1460 national gene banks operate in the world, including about 300 large ones, in which, in ex situ conditions, guaranteed storage of samples of cultivated plants and their wild relatives is ensured. The ex situ collections are also kept by botanical gardens, of which there are about 2 thousand in the world (about 80 thousand plant species, 4 million samples and 600 seed banks). Their presence is a sign of national sovereignty, the level of culture, concern for the future of the country and the world. By 2002, more than 532 thousand plant specimens were preserved in international centers under the control of the FDO advisory group, of which 73% belong to traditional and landraces, as well as wild relatives of cultivated plants. As noted by Dleksanyan (2003), one should distinguish between the concepts of “genebank” and “ex silu collections”. If the first is the guaranteed storage of the gene pool in specially equipped premises, then "ex situ collections" include samples that are of interest to their holders.

In the early 50s. XX century, the first semi-dwarf rice cultivar was obtained by using the dwarfism gene of the Chinese cultivar Fee-geo-woo, and the wheat cultivar Gaines in the irrigated lands of the Pacific Northwest of the United States gave a record harvest of 141 kg / ha. In 1966, the IR 8 variety was created, which received the nickname "miracle rice". With high agricultural technology, these varieties gave 80 and even 130 c / ha. Similar results were obtained with millet. While the old varieties had a yield index of 30-40%, the new ones had 50-60% and more.

Further opportunities to increase yields by increasing the yield index are limited. Therefore, much more attention should be paid to increasing the amount of net photosynthesis. It is necessary to focus on the wide species and varietal heterogeneity of agroecosystems and agricultural landscapes in field crop production conditions, along with the selection of insurance crops, as well as crops and mutually insured varieties, it also includes a differentiated approach to the implementation of the adaptive potential of each of them. The high potential productivity of the variety and the agroecosystem, achieved by (and sometimes by) reducing their ecological resistance to environmental factors limiting the size and quality of the crop, as well as the functioning of excessively bioenergy-consuming ecological sustainability, cannot be considered as adaptive, since for cultivated plants, the main indicator of adaptability ultimately is to ensure a high value and quality of the crop. Gene pools accumulated in genebanks can be a source for scientifically grounded breeding to create the necessary varieties.

It should be emphasized that millions of accessions have been collected in the world genebanks of cultivated plants; however, so far only 1% of them have been studied in relation to their potential properties (Zhuchenko, 2004). At the same time, the control and improvement of their genetic component - the gene pools of agricultural species, which determines the characteristics of local agricultural systems - is of leading importance for the creation of sustainable agricultural systems.