CZ2010511A3 - Immunobiological formulation for treating autoimmune diseases of skin, especially psoriasis - Google Patents

Abstract

The present invention relates to an immunobiological composition comprising at least one active ingredient selected from the group consisting of inactivated Bacillus megaterium culture cells and Bacillus megaterium culture filtrate. This immunobiological agent can be used to treat skin autoimmune diseases, particularly in the treatment of psoriasis.

Description

The invention relates to a new immunobiological preparation suitable for the treatment of autoimmune skin diseases, in particular psoriasis, and to this preparation for use in therapy.

State of the art

Psonase (psoriasis) is a non-communicable chronic inflammatory skin disease, which is manifested by the sowing of red or purple plastic sharply demarcated deposits with a deposit of silvery scales. We are also talking about the so-called autoimmune disease, when an individual's immune system attacks his own cells. The most common sites are the elbows, knees, hips and foreheads.

The first mentions of it can be found in ancient civilizations, in Egyptian papyri. The word psoriasis comes from the Greek psora, ie itching, supina. In Central Europe, about 2 + 3 percent of the population suffers from psoriasis.

Although psoriasis has been recorded since biblical times, the treatment options for this skin disease, which excludes patients from life and causes disfigurement, did not improve until the second half of the 20th century. However, no cure has yet been found to cure it, although a number of topical, systemic or ultraviolet-based drugs are now available to patients. These can keep the disease under control and temporarily eliminate some of its symptoms. In the 21st century, a new type of psoriasis treatment was introduced, biological drugs that brought new hope to psoriasis sufferers around the world.

The treatment of psoriasis can be divided into the following basic categories: topical medications (skin medicines), phototherapy or a combination of phototherapy and other medicines, systemic treatment (medicines given as injections or tablets), biological treatment. A number of factors determine whether a particular treatment for a person with psoriasis will be appropriate, such as the type of psoriasis, the location of the affected area on the body, the severity of the symptoms, the patient's age and medical history, and the patient's response to previous treatment.

Topical therapy is usually used first in the treatment of psoriasis. Drugs work relatively quickly to remove immediate lesions after application and are also usually well tolerated. However, in order for topical preparations to be effective, they must be used repeatedly, as

it is not enough to maintain the temporary disappearance of psoriasis. They can also get dirty and can take a long time to apply if psoriasis affects most of the tchia. Dithranol (anthralin), for example, is used as a drug for local therapy, or in combination with UVB rays. However, this drug often causes skin irritation and burning, and anything that comes in contact with the drug is contaminated.

Topically applied corticosteroids are drugs similar to the naturally occurring hormones of the human body. They are available in many different strengths and dosage forms such as creams, solutions, emollients, sprays, ointments and medicated patches. Corticosteroids work by slowing down the growth of skin cells and reducing inflammation of the lesions in patients with psoriasis. Although corticosteroids can remove lesions quickly, it is typical for them that remission does not last long, so that psoriasis-associated lesions reappear after a short time. The side effects of long-term use of corticosteroids are numerous and include skin atrophy, the formation of telangiectasia (protruding dark red spots on the skin) and stretch marks (scratches on the skin), after which scars remain on the skin.

Topically applied coal tar can be used alone or in combination with UVB. The use of tar is severely limited by its ability to pollute everything and especially by the fact that coal is a proven carcinogen.

Retinoids (vitamin A derivatives) are more recently developed drugs for the local (topical) treatment of psoriasis and are used to treat mild to moderate focal psoriasis.

The vitamin D analog calcipotriol in the early 1990s provided some alternative to topically applied steroids, adithranol tar. Later, calcitriol and atacalcitol were introduced. Calcipotriol has been shown to be as effective or better than other products and is more cosmetically acceptable and generally well tolerated.

Salicylic acid is a substance that helps remove scales from lesions. This will allow the application of topical preparations and their better penetration into the skin. It can be used in the form of a paste, cream, ointment or solution.

For the treatment of psoriasis, some individuals can try a variety of over-the-counter products, including moisturizers, bath oils, magnesium sulfate products, and oatmeal with oil. They are not effective in everyone, but they are usually without side effects.

Patients with psoriasis who do not respond to topical treatment or for whom psoriasis is too prevalent are suitable for phototherapy. Its principle is the exposure of the skin to ultraviolet radiation, which has a therapeutic effect on psoriasis. This ultraviolet radiation can be both natural, as part of solar radiation, and artificial, from special light bulbs emitting radiation in a narrow band around the optimum wavelength of 311 nm.

The PUVA method, also called photochemotherapy, was developed in the 1970s and is based on a combination of a light-sensing agent (psoralen) followed by UVA ultraviolet radiation (similar to UVB, UVA is found in natural light). Psoralen causes the skin to be more sensitive and stronger to this type of ultraviolet radiation. Long-term treatment with PUVA can lead to premature skin aging and also significantly increases the risk of skin cancers (basal cell carcinoma and skin surface cells).

Laser therapy is a new method for the phototherapy of psoriasis. The excimer laser emits a beam of high-intensity ultraviolet light aimed at selected areas of the skin. The side effects of laser treatment include a small risk of scarring and bruising.

Patients who do not respond to phototherapy or cannot undergo the frequent irradiation necessary to remove the lesions can be treated systemically. The term systemic treatment means that it uses drugs that work throughout the body. These include methotrexate, cyclosporirl and orally administered retinoids. One of the biggest disadvantages of methotrexate is the fact that its long-term use is associated with liver damage, and therefore routine liver biopsies are required in many countries for patients taking long-term methotrexate. It works by binding to an enzyme that is involved in cell growth and therefore slows down pathologically altered skin cell growth in psoriasis. However, methotrexate also affects normal cells, including fetal cells, bone marrow cells and sperm cells. Cyclosporine slows the growth of skin cells by inhibiting the immune system. However, its long-term use is associated with nephrotoxicity. Retinoids affect cell behavior, including growth rate and their separation from the skin surface. Orally administered retinoids are of little use as monotherapy and treatment is associated with numerous side effects, such as hair loss and thinning of the nails, and can also harm the fetus. However, retinoids are very effective in small doses in combination with UVB phototherapy or PUVA, which allows them to substantially remove lesions even with a low number of phototherapy visits and few side effects.

Biological treatment of psoriasis represents a significant advance in the treatment of this disease.

It is an effective, long-term safe, comfortable and well-tolerated treatment for patients. It is intended for a defined group of psoriatic patients who cannot be effectively treated with current systemic therapy. It enables these disabled people to have a full personal life and employment in a society comparable to a healthy population. Biological treatment offers high effectiveness in the treatment of psoriasis

4 'and higher safety in that they are targeted drugs based on natural proteins that disrupt certain steps in the pathogenesis of psoriasis. Biological agents target the immune system, where they block the activity of certain cells of the immune system that are important in the pathogenesis of psoriasis. While other treatments for psoriasis, such as PUVA, mcthotrexate and cyclosporine, also affect the body's immune system, the effect of biologics is more specific and may be safer.

Two types of biological drugs are used to treat psoriasis - one acts on the T-cells of the immune system and thus directly affects psoriatic inflammation, others are non-specific, they act on cylocins (TNFα) and affect inflammation at many levels. The first group of drugs developed specifically for prosiasis includes efalizumab (Raptiva), which specifically affects T-cells and thus has a specific effect on the skin. Second group drugs /e.g. etanercept (Enbrel), infliximab (Remicade), adalimumab (Humiral)) act on cytokines and are therefore suitable for the treatment of joints and skin.

These drugs, because they act against the misguided immune response, have significantly fewer side effects. As with all advanced drugs, safety concerns have arisen.

up to 25 / o patients with moderate and severe psoriasis. These are candidates for lifelong overall treatment. Therefore, it is very important to make sure that the drug does not have serious side effects for them. Clinical observations showed significant and long-term improvement in 73% of patients treated with moderate and severe psoriasis who received efalizumab, and 40% of patients had complete resolution of symptoms. The side effects were comparable to those seen with placebo.

It is therefore necessary to further search for other drugs and methods suitable for the biological treatment of psoriasis, which have sufficient efficacy and the least possible side effects.

The essence of the invention

The invention relates to the use of an active ingredient selected from the group consisting of inactivated Bacillus megaterium culture cells and Bacillus megaterium culture filtrate for the manufacture of an immunobiological preparation for the treatment of autoimmune skin diseases, preferably for psoriasis.

The term "inactivated Bacillus megaterium culture cells" refers to cells that are prepared by any of the known inactivation methods, such as reactive sterilization.

Preferably, the Bacillus megaterium is selected from the group consisting of Bacillus megaterium strains: 1/32. 2/37, 3/39, 4/58 and 5/74 according to CNCTC (Czech National Collection of Type Cultures), Catalog of Tribes - 5th edition, p.5.

The composition of the present invention is preferably administered by subcutaneous injection. It is given 2 to 3 weeks apart if the desired clinical effect does not occur within one week of the first dose.

As shown below, administration of the composition of the present invention has been shown to be very effective in the treatment of psoriasis.

Overview of figures in the drawings

Giant. 1 shows a table for calculating the PASSION value.

Giant. 2 is a graph showing changes in the average PACH value over time in the subjects of Example 3.

Examples of embodiments of the invention

Example 1

Cultivation of Bacillus megaterium bacteria, preparation of inactivated cells ·

In the presented examples, cultures of bacteria of the species Bacillus megaterium from the collection of the State Institute of Public Health, CNC1C - Czech National Collection of Type Cultures - Catalog of Strains - 5th Edition, p.5

Bacillus megaterium de Bary (hereinafter referred to as Bacillus megaterium), strains - 1/32. 2/37, 3/39, 4/58. 5/74.

6200 Brn 1/32 2/37 3/39 6756 4/58 6213 5/74

SZÚ, Mega I. (Medium 1 37C)

SZÚ, Africa. (Medium 1 37C)

SZÚ, Bacillus petasites. (Medium 1 37C)

I .Alf ~ ldi, Regional Hygiene Station, Szeged, Hungary, 25.

Producer of megacine. (Medium 1 37C)

ATCC, 13632. Non-spore-forming variant. (Medium 1 37C)

The lyophilisate culture is revived according to the following instructions:

1. Break off the incised upper end of the ampoule.

2. Cut the ampoule with a file.

3. The ampoule surface is disinfected.

4. Break the cut ampoule and remove the inner ampoule with dry matter.

5. Dissolve the dry matter by adding a few drops of No. 2 masopeptone broth (preparation described below)

6. This solution is sterile poured into 1 liter of masopepton broth No. 2, which is a universal nutrient medium for culturing microorganisms, followed by incubation.

Preparation of masopepton broth No. 2:

Ingredients: meat extract 10 g, peptone for bacto 10 g, sodium chloride 5 g.

Preparation: Weigh the above ingredients into 1000.0 ml of distilled water (sterile Erlenmeyer flask) and heat to complete dissolution. The flask was then sterilized in an autoclave at 121 ° C for 20 minutes.

pH: 7.2 ± 0.2 at 25 ° C.

Incubation:

• 7 ·

The flask is stored at 37 ° C for 1 week - aerobically. The growth of the culture is reflected in the turbidity of the contents of the flask.

Intermittent, fractional sterilization:

The flask was heated to 100 ° C for 30 minutes and then left at 37 ° C for 24 hours. In these conditions, any disputes present will arise. Grown vegetative forms are destroyed by repeating this process three times.

Example 2

Preparation and administration of immunobiological preparation

The inactivated cell suspension prepared according to Example 1, after cooling to 25 DEG C., is filtered using a sterile S-PAK Millipore 0.22 .mu.m filter and diluted 1:10 with water for injection (Aqua pro injectione, Czech Pharmacopoeia 2003). It is possible to use filters from other manufacturers with similar parameters.

An allergy test is then performed: the filtrate is injected intradermally into the patient in an amount of 0.1 mL with a syringe. If the allergy test is positive, the filtrate must be further diluted (1:20, 1:30).

After a negative allergy test, 1 ml of the product is injected subcutaneously into the patient's dorsolateral side of the arm.

Induration appears at the application site within 3 days. The antipsoriatic therapeutic effect usually begins 1 week after application. If the therapeutic effect does not occur, the application is repeated 2 to 3 weeks after the previous dose. This procedure can be repeated a total of 3 times with the application of the dose to the other arm than the one to which the previous dose was administered.

Care must be taken due to the possibility of allergies.

Example 3

Monitoring the efficacy and safety of an immunobiological product

The severity of psoriasis is assessed by the PASI (Psoroasis Area and Severity Index) - a scoring system that measures redness, induration and desquamation of the skin. where PASI 0 indicates the absence of psoriasis and PASJ 72 is the maximum possible skin condition.

Table for calculating PAS values! is shown in Fig.!.

Testing took place under the following conditions:

Entry criteria for evaluation subjects:

A / PACH value> 22

B / Men and women over the age of 18

C / Without concomitant treatment of psoriasis (psoriasis), outside the location of the emollient (substances that soften, smooth and hydrate the skin)

Exclusion criteria for evaluation subjects:

A / Systemic treatment with acitretin, cyclosporine, methotrexate, light therapy or other general treatment for psoriasis during the last 3 months and throughout the follow-up from administration until day 180.

B / Topical (local) treatment of psoriasis after Day -14 (so-called washout period)

C / Pregnancy or breast-feeding. Women of childbearing potential were excluded from participation if they did not use a reliable method of contraception

D / Acute infectious diseases

E / Immunosuppressive therapy

F / Cancer

Time frame:

AI Elution period of 2 weeks, when subjects evaluated only local emollients to treat psoriasis.

B / Evaluation intervals:

Day -14, 0. 10, 14, 21, 28, 56, 90. 180.

Methodology of administration:

After a negative sc allergy test, 1 ml of filtrate is administered subcutaneously to the dorsolateral side of the arm.

The therapeutic antipsoriatic effect of sc usually begins to appear after 1 week of application. If the sc does not appear, the sc application is repeated 2 to 3 weeks after the previous dose, this procedure is possible

- 9 'repeat a total of 3 times with the application of the dose to the other arm than the one to which the previous dose was administered.

Effectiveness evaluation methodology:

The evaluation was based on the assessment of psoriasis skin involvement in the categories of redness, induration, desquamation in individual subjects of evaluation. PAŠI values were calculated according to the table shown in Fig. 1, on Days -14, 0, 10, 14. 21, 28, 56, 90 and 180.

The overall treatment effect for the subject was evaluated on Day 180 as follows;

Deterioration was defined as a PASS value higher than a PASS on Day 0.

No improvement was defined as a reduction in PASI of 0 to 25% compared to PASI on Day 0.

A slight improvement was defined as a 26 to 50% reduction in PASI compared to PASI on Day 0.

Improvement or positive clinical response as a 51 to 75% reduction in PASI compared to PASI on Day 0.

Very significant improvement as a 76 to 100% reduction in the value of the PASS compared to the PASS on Day 0.

The results are reported in Table 1 below.

Safety assessment methodology - monitoring of adverse reactions:

The incidence of adverse events was monitored and recorded from the time of administration (Day 0) to Day 180. The results are reported in Table 1 below.

Characteristics of subjects evaluated:

Number of subjects: 25, of which 7 women and 18 men

Average age: 47.4 years (19-68)

Average duration of psoriasis; 15.4 years (2 * 36)

The average value of PAŠI on Day 0: 45.3

Evaluation results:

Table 1

Deterioration (Ο (about%) No improvement 1 (4%) Slight improvement 2 (8%) Improvement 5 (20%) Very significant improvement 17 (68%) Side effects lx inflammation at the injection site with adjustment within one week 3x transient mild itching around the injection site The need to repeat the application The application was repeated once at 3 subjects on Day 14

The graph showing the change in the average PAŠI value for all monitored subjects at the time of evaluation is shown in Fig. 2.

Conclusion:

The use of the preparation prepared and applied according to the above procedure was very well tolerated and had a significant antipsoriatic therapeutic effect, with 68% of subjects having a very significant improvement in their skin psoriasis in the observed period. There was an improvement in 20% of subjects and a slight improvement in 8% of subjects in the observed period. No improvement was seen in 4% of subjects and no subject had a worsening of psoriasis during the study period. The mean PAH value for all subjects on Day 180 was 9.1, corresponding to a significant antipsoriatic treatment effect that was statistically significant at a significance level of 5%. The adverse reactions were mild and transient and did not exclude any subject from the evaluation. The product has proven to be very effective, well tolerated and suitable for the treatment of psoriasis with a long-term therapeutic effect.

- 1Γ

Claims (2)

- 1Γ PATENT CLAIMS Use of an active ingredient selected from the group consisting of inactivated Bacillus megaterium culture cells and Bacillus megaterium culture filtrate for the manufacture of an immunobiological preparation for the treatment of autoimmune skin diseases, preferably for the treatment of psoriasis. Use according to claim 1, characterized in that the Bacillus megaterium is selected from the group consisting of Bacillus megaterium strains 1/32, 2/37, 3/39, 4/58 and 5/74 (CNCTC).