Recombinant interferon: types, classification and mechanism of action

Recombinant interferons are a group of antiviral drugs used for therapeutic and prophylactic purposes. These biologically active proteins are naturally synthesized in human cells in response to the penetration of foreign agents. In modern medicine, these drugs are recognized as the most effective and safe for the treatment of viral diseases.

Classification

In microbiology, more than 20 types of interferons (IFN) are distinguished, which differ in properties and biological structure. Medicines based on them are classified as follows:

• By type of active component: o alpha interferon (or leukocyte); o beta interferon (fibroblast); o gamma interferon (immune); o lambda interferon.
• According to the production method: o natural, obtained from human blood leukocytes; o recombinant human interferon, produced synthetically (by genetic engineering).

Alpha and beta interferons are combined into a type I family due to the similarity of their functions in the body and the same amino acid sequences. Gamma and lambda interferons are isolated into a separate type II and III, respectively. The first generation of natural proteins had a big drawback - they required the use of scarce raw materials (donated blood) and a high degree of purification from foreign proteins. This led to their high cost and low efficiency. Recombinant alpha interferons currently occupy a leading position among drugs of this type in terms of degree of knowledge and breadth of use in medical practice.

Features

In addition to the above classification, these proteins differ in subtypes. So, the category of recombinant alpha 2 interferons includes no less than 24 subtypes, which differ in 24 genes. They are not completely identical in primary structure.

Unlike alpha interferons, the beta modification is encoded by only one known gene. Both types of proteins are activated by viruses and use the same receptors in their mechanism of action on other cells.

The subtype of human recombinant interferon alpha-2b differs from alpha-2a in two amino acid residues in the structure. The rest of them (and there are more than a hundred of them in total) are the same. Therefore, the diseases in which they are used, as well as the side effects, are the same, but the reaction of the body (antibody production) is different.

Natural leukocyte interferons are also classified according to the degree of purification:

• Native, characterized by shallow cleaning and as close as possible to the feedstock. They have the greatest potential for immunobiological effects.
• Concentrated, highly purified. They are used most often in cases where it is necessary to introduce a large single dose. The uniformity of the composition of these drugs reaches 90%.
• Combined. They are obtained with gentle cleaning methods. The presence of additional cytokines makes standardization of these substances difficult. At the same time, thanks to this factor, they have a higher immunomodulating effect, which contributes to the expansion of their scope.

Recombinant human interferon incorporates a monospecific protein. It belongs to one of the subtypes. Type b1a in the preparations of recombinant alpha interferon is characterized by a glycolized form (non-enzymatic addition of sugar residues to organic protein molecules), and b1b is non-glycolized. Such interferons are 98% homogeneous in composition.

These features of natural and artificially synthesized proteins cause a difference in the field of their application. Recombinant interferons are dominated by antiviral and antitumor effects. In natural ones, it is immunomodulatory, and there is also a higher activity against bacterial and purulent-septic pathologies.

Recombinant interferon preparations

The most widely used of this group of drugs are:

• alpha 2a interferon: "Reaferon", "Viferon", "Roferon", "Interal";
• alpha 2b interferon: Intron-A, Lifeferon, Peginterferon, Infagel, Inrek;
• alpha 2c interferon: Berofor;
• beta-interferon: "Interferon-beta-1a", "Fron", "Rebif", "Avonex", "Betaseron", "Betaferon";
• gamma interferon: "Aktimmun", "Gammaferon", "Ingaron", "Imukin".

Diseases

Recombinant interferons are active in the treatment of diseases such as:

• dermatological pathologies: genital herpes, warts, condylomas, papillomatosis, shingles;
• ophthalmic diseases: inflammation of the cornea of ​​the eye due to herpetic or adenovirus infection (reduction in the duration of the disease, increase in inter-relapse periods);
• acute infectious diseases of the upper respiratory tract: influenza, acute respiratory viral infections (emergency prevention for people at risk, as well as for medicinal purposes);
• pathology of the hepatobiliary system: viral hepatitis B, C in acute and chronic form (pronounced clinical effect, mortality reduction up to 60%);
• AIDS: normalization of immunity, reduction of disease severity in more than half of patients; reducing the risk of developing Kaposi’s sarcoma related to AIDS;
• other pathologies: CMVI (cytomegalovirus infection), which occurs against the background of immunodeficiency states (interferons are used to prevent it), as well as after transplant operations; sclerosing panencephalitis (inflammation of the brain).

These drugs are characterized by a universal spectrum of antiviral activity. Unlike chemotherapeutic agents, they do not lead to the emergence of resistant forms of pathogens, but act on factors of innate, natural immunity.

Discovery story

Interferons were discovered almost 50 years ago. The first drugs were obtained from the blood of donors. To do this, blood cells were treated with viruses, after which they began to produce proteins with protective properties. The interferon thus obtained was highly effective, but its production on a large scale was difficult due to a shortage of raw materials. For example, to obtain the volume of the drug needed to treat 1 cancer patient, it was necessary to collect blood from 200 donors.

In the mid 80-ies of the XX century, the first prerequisites for the production of synthetic recombinant interferons arose. The rapid development of genetic engineering in these years has led to the creation of a new technology - the introduction of the corresponding gene in the colony of bacterial cells of Pseudomonas putida, which can multiply rapidly. This allowed us to synthesize human recombinant alpha 2b interferon on an industrial scale. The first drug created in the USSR was called Reaferon.

In subsequent years, thorough studies of this drug in animals were carried out for teratogenic and toxic properties. Tests have confirmed its safety for the fetus and the absence of differences between the side effects of artificially synthesized and natural interferon.

Later, E. coli bacteria began to be used to produce recombinant interferon, since they produce this substance faster. The first drug obtained on their basis was called Reaferon-EC (from the abbreviation Latin name of this microorganism Escherichia coli). These bacteria are also used in most modern production of recombinant interferons.

Operating principle

Interferons are a kind of biological mediators that activate the human immune system. They contribute to the recognition and suppression of foreign genetic information. With the introduction of viruses into the cell, after a few minutes, the number of pathogens increases many times. They spread further, affecting healthy cells and multiplying again. Especially quickly this process occurs at the initial stage of the disease, since during this period the human body is not able to produce the necessary amount of interferon.

Thanks to these proteins, the synthesis of a number of enzymes, antibodies and other components of the immune defense is triggered. As a result, cells become immune to viruses. The following mechanisms are also distinguished in which interferons are involved:

• stimulation of macrophages, their activation to the absorption of non-viable, affected cells;
• suppression of growth and destruction of atypical cells (antitumor effect);
• effects on immunocytes (lymphocytes produced in the bone marrow) - the main cells of the immune system: NK cells, T-lymphocytes, monocytes, macrophages and granulocytes; stimulation of their non-specific cytotoxicity;
• activation of the synthesis of proteins that increase the resistance of cells to foreign agents, the transfer of these proteins to neighboring cells;
• launching a cascade of reactions that stimulate the production of anti-inflammatory factors (anti-inflammatory effect);
• activation of the synthesis of their own IFN, which allows to reduce the recovery time.

A particularly vivid antiviral effect is characteristic of recombinant interferons alpha 2b, 2a and beta. They block the production of viral proteins and prevent the propagation of pathogens. One of the important advantages of drugs based on them is minimal toxicity and the possibility of prescribing in childhood.

Synthesis

The production of recombinant interferons occurs in stages:

• Isolation of messenger RNA after activation of interferon production in a bacterial culture;
• synthesis of complementary RNA-based DNA;
• embedding the DNA obtained in the previous step into plasmid vectors — extrachromosomal DNA molecules capable of independent copying inside bacterial cells and responsible for the production of proteins;
• obtaining recombinant DNA;
• synthesis of clones of microorganisms that produce interferon;
• propagation of a bacterial culture in a nutrient medium;
• isolation of bacterial cells by centrifugation;
• precipitation of interferon proteins from solution;
• purification of recombinant interferon by the method of affinity chromatography or other methods.

Propagation of the clone culture occurs under industrial conditions in reactors, and the previous steps in laboratories. Recombinant IFNs are produced outside the human body, a human interferon gene is built into their genetic material.

There are several bacterial cultures on the basis of which these proteins are obtained. Below is what recombinant alpha 2b interferon is made of:

• Escherichia coli E. coli (product accumulation occurs intracellularly);
• Bacillussubtilis hay bacillus (interferons are released into the environment);
• Pseudomonas aeruginosa Pseudomonas aeruginosa;
• yeast Saccharomycopsis fibuligera.

The latter type of producers has the following advantages compared to others:

• the possibility of using cheap culture media;
• easy separation during separation;
• high productivity of the process (more than 10 times in comparison with others);
• the process of joining carbohydrate groups, similar in mechanism to that in animal cells.

Release Forms

Recombinant interferons 2b, 2a and beta are available in the following dosage forms:

• solutions for injection;
• lyophilisates;
• drops and films for the eyes;
• solutions for oral administration;
• suppositories and microclysters for rectal and vaginal administration;
• ointments;
• gels;
• tablets;
• aerosols;
• spherical vesicles (liposomes).

Recombinant Alpha IFN

Synthetic alpha interferons are fully consistent with natural proteins. They play an important role in triggering the immune response in the human body, activate the production of important cytokines, mediate between innate and acquired immunity, and provide an immunological "memory".

The table below shows the main characteristics of some preparations of recombinant interferon of this type:

The new generation of drugs is pegylated (or conjugated) alpha-IFN, which are characterized by a prolonged effect. They show high efficiency in the treatment of viral hepatitis. These include Pegasys (IFN-α-2a) and Pegintron (human recombinant interferon 2b).

Recombinant Beta IFN

Among the beta-interferons, 2 subtypes are currently distinguished - b1a (glycolized) and b1b (non-glycolized). In addition to antiviral and immunomodulatory effects, they affect the nervous system and are used to treat multiple sclerosis. Drugs are administered subcutaneously or intramuscularly. It has been clinically proven that the decrease in the frequency of exacerbations of the disease occurs by almost a third, but clear criteria for assessing effectiveness do not yet exist.

The mechanism of action of such drugs is based on the following phenomena:

• The binding of interferons to specific receptors on the cell surface, resulting in the activation of the production of proteins with an antiviral, antitumor anti-inflammatory effect.
• Reducing the number of new foci of sclerotic brain damage and atrophic changes in its tissues (confirmed by MRI data).
• Inhibition of leukocyte division and their migration to the area of ​​inflammation by reducing the production of proteolytic enzymes.
• Enhanced decay of gamma interferon, which plays an important role in the development of multiple sclerosis.

Recombinant Gamma IFN

In Russia, the most widely produced recombinant gamma interferon in the composition of the drug "Ingaron". It is used in the treatment of diseases such as:

• flu (including "swine");
• otitis media (aerosol form of the drug);
• granulomatous disease;
• osteopetrosis (congenital familial osteosclerosis);
• chronic viral hepatitis B, C;
• AIDS;
• pulmonary tuberculosis;
• oncological pathologies;
• urogenital infections;
• genital herpes and herpes zoster;
• HPV;
• chronic prostatitis.

The aerosol form of human recombinant gamma interferon is also used to prevent influenza (irrigation of the nose and nasopharynx). This substance blocks the production of polypeptides responsible for the development of fibrotic changes in the liver and lung tissue.

Side effects

When treating with alpha and gamma interferons, the most common side effects are flu-like syndrome. It includes features such as:

• increase in body temperature;
• headaches and muscle aches;
• chills;
• weakness.

These symptoms usually appear during the first or second week of treatment. They can be eliminated by lowering the dosage.

The following side effects are less common:

• digestive upset;
• sleep impairment;
• thrombocytopenia;
• a decrease in the level of white blood cells in the blood;
• intoxication with thyroid hormones.

When taking beta-interferons, the following negative phenomena may also occur:

• hypertension;
• tachycardia;
• arrhythmia;
• heartache;
• heart failure;
• decreased intelligence;
• mental disorders - depression, suicidal ideation, depersonization, epileptic seizures.