KR100332138B1 - Compositions for the treatment and prevention of viruses, parasites, bacterial infections or fungal infections, including interleukin-4 receptors - Google Patents

Abstract

The present invention relates to the use of the interleukin-4 receptor or derivatives thereof for the treatment, prevention and diagnosis of allergies, viruses, parasites and bacterial diseases and fungal infections.

Description

Pharmaceutical compositions for the treatment and prevention of viruses, parasites, bacterial infections or fungal diseases comprising interleukin-4 receptor

There are serious problems with the treatment and prevention of numerous allergic, viral, parasitic and bacterial diseases. The present invention relates to the use of IL-4 receptors or derivatives thereof for the treatment, prevention and diagnosis of such diseases.

It is known that changes occur in subpopulations of lymphocytes and monocytes during the course of parasite, viral and bacterial diseases. This is associated with an increased incidence of, for example, so-called type 2 T-helper cells (hereinafter referred to as TH2 cells). T cells can generally be divided into subpopulations based on surface markers and their function. Thus, for example, T-helper lymphocytes carry CD40 surface molecules and secrete cytokines after their activation.

Cytokine pattern analysis of T-helper cells cloned from healthy mice or mice stimulated with allergen cells resulted in interleukin-2, interleukin-4, gamma-interferon, interleukin-5, interleukin-6, interleukin-10. And lymphotoxic toxins (so-called THO helper cells of type THO).

After stimulating mice with, for example, the bacterial antigen Brucella abortus or Mycobacterium tuberculosis, clones found in particular after cloning of T-helper cells are lymphotoxic toxins, Gamma-interferon and interleukin-2 are secreted, but little or no interleukin-4, interleukin-5, interleukin-6 and interleukin-10 (the so-called T1-helper cells of type TH1).

For example, after infecting susceptible mice with parasitic pathogens such as Leishmania major, the clones that occur, particularly during cloning of T-helper cells, have increased amounts of interleukin-4, interleukin-5 and interleukin- Produces 10, but produces reduced or undetectable amounts of interleukin-2 and gamma-interferon (T-helper cells of type TH2). Mosmaun et al., Immunological Reviews 1991, No. 123, 209-229; S. Romagnani, Immunology Today, 256-257, Vol. 12, No. 8 1991].

This increased incidence of TH2 lymphocytes has already been detected in some infectious diseases in animals and humans [see; Else and Grencis, Parasitology Today, Vol. 7, No. 11, 1991, pp, 313-316; Parasitology Today, Vol. 7, No. 10, 1991, p. 261)], which may also be reflected as secondary variables. Thus, mice infected with Leishmania majors typically have reduced gamma-interferon production resulting in a significant increase in serum IgE and eosinophils.

For example, in people infected with leprosy, leishmaniasis and schistosomiasis and infected with Mycobacterium tuberculosis, the serum IgE concentrations of these patients are found to be much higher than the serum levels of normal individuals. In parasitic infections, eosinophils are often observed during disease progression.

Atopic dermatitis and asthma as well as immediate types of IgE-mediated allergic reactions are also characterized by this type of dysregulation. For example, antigen-specific T-cell clones from skin biopsies of atopic dermatitis patients are particularly members of type TH2 [see; Kapsenberg et al., Immunology Today, Vol 12, No. 11, 1991,392-395].

It has now been found that diseases characterized by increased development of T-helper cells of type TH2 can be diagnosed and treated therapeutically and / or prophylactically with the help of IL-4R or derivatives thereof.

Accordingly, the present invention is directed to the use of an IL-4 receptor or a derivative thereof for the manufacture of a medicament for the treatment and / or prophylaxis of a disease with an increased incidence of T-helper cells of type TH2 or for the preparation of diagnostic aids for the identification of said disease. It is about.

The term "derivative" of IL-4R means, for the purposes of the present invention, amino acid 1 in functionally equivalent parts, mutants or variants of IL-4R, in particular the extracellular portion of IL-4R, specifically the mature protein of the human IL-4 receptor. To about 209 sites and glycosylation mutants thereof In addition, fusion containing IL-4R or derivatives thereof, and other proteins or portions thereof, preferably the Fc region of an antibody, in the use of a receptor according to the invention. Protein (IL-4R / Fc fusion protein) may also be used, and for the diagnosis, treatment and / or prophylactic treatment of the disease, IL-4R, derivatives thereof, preferably in a mixed product mixed with gamma-interferon Or fusion proteins can be used.

Furthermore, in the case of treatments in combination with purified allergens or portions thereof, in particular in the case of specific immunotherapy (desensitization), desensitization incorporating purified allergens, for example in patients with allergic rhinitis, is advantageous.

Furthermore, gamma-interferon and / or the cell surface molecule CD40 and its ligands, cell surface molecule CD40 ligands, preferably soluble variants of the CD40 surface molecule itself (e.g. CD40 / Ig fusion proteins or derivatives thereof corresponding to the following description). Combination treatment with substances that block the interaction is also advantageous.

By "derivative" of a soluble variant of a CD40 surface molecule is meant for the purposes of the present invention a functionally equivalent moiety or variant that blocks the interaction of the cell CD40 with the cell surface molecule CD40 ligand.

Diseases include allergic and infectious diseases, especially fungal infections, as well as viral, bacterial and parasitic infections; Preferably human immunodeficiency virus (HIV), mycobacteria, in particular mycobacterium reprae, Listeria, protozoa, in particular Leishmania and Plasmodium genus, worms, in particular Schistozoma, niposton gillus , Heligmosomoides infection and Trichuda, Trichinella, Taenia (Cisty cercus), Candida and Aspergillus infections. However, it is possible to diagnose, treat or prophylactically treat immediate types of allergic reactions, in particular IgE-mediated responses. These especially include atopic dermatitis and asthma.

Dosage forms generally depend on different diseases. Thus, for example, topical administration may be advantageous for some diseases. Advantageous examples include inhalation administration for asthma, eye drop administration for conjunctivitis, and dermal or intradermal administration for atopic dermatitis, since pathogenic TH2 cells can be detected particularly locally.

It has been reported that the human IL-4 receptor consists of a total of 825 amino acids. Idzerda, R. J. et al, (1990) J. Exp. Med. 171, 861-873. According to Izderda et al., 25 N-terminal amino acids act as signal peptides and the mature human IL-4 receptor consists of 800 amino acids, 1. Extracellular domain (207 amino acids), 2. Dural region (about 24 Dog amino acids), 3. A three-region structure comprising the cytoplasmic region (569 amino acids). The production of IL-4 receptors by genetic engineering is particularly advantageous because it is possible to produce the amount of material necessary for treatment directly.

IL-4R can be prepared, for example, as described in International Patent Application WO90 / 05183. According to this, cDNA gene banks are produced, for example, from T cell line T22 and screened with probes for IL-4R-specific DNA. Hybrid-excluded cDNAs of the mouse cell lines described in WO90 / 05183 can be used as probes. However, one or more probes of about 20 nucleotides in length at 193-210 can also be used to screen for IL-4R-specific hybridization probes, such as cDNA banks. Positive clones can be checked for IL-4R-specific cDNA, for example by sequencing, and then the found IL-4R DNA can be cloned into a suitable expression vector such as pCAV / NOT (WO90 / 05183). And can be expressed fully or partially in a suitable host cell, eg, COS-7, BHK-21 or CHO cells. When only cDNA sites encoding the extracellular region of the IL-4 receptor are used for expression, the extracellular region of the interleukin-4 receptor is generally secreted into the medium by the transfected cells. To this end, the cDNA is modified by a genetic engineering method corresponding to the state of the art, which introduces a termination codon after the coding sequence for the extracellular region of the receptor, or a desired site thereof, which is modified by the method. Is cloned into a suitable expression vector. Maliszewski et al. (1990), J. Immunol. 144. 3028-3033; Dower et al. (1989), J. Immunol. 142, 4314]. In many cases, for example, for the murine IL-4 receptor, the isolated cDNA encodes for the naturally occurring secreted form of the receptor (Mosley et al (1989), Cell 59, 335). These cDNAs can then be used directly for the production of expression vectors. Secreted proteins can be purified from the media, for example, by immunoaffinity chromatography using ligand affinity chromatography or specific monoclonal antibodies. Maliszewski et al (1990), J. Immunol 144, 3028-3033. In addition, as described in European patent application EP-Al-0 464533, IL-4R or derivatives thereof and other proteins, such as immunoglobulin sites, such as Fc regions of antibody molecules (hereinafter IL) by genetic engineering methods Fusion proteins (labeled -4R / Fc) can be prepared and expressed. The advantage of such fusion proteins is that the half-life is extended and the quality is enhanced and purified through protein A-Sepharose affinity chromatography.

Problem-free purification and possibly improved pharmacodynamic properties by affinity chromatography mean that the synthesis of the soluble form of the IL-4 receptor as an immunoglobulin fusion protein is particularly advantageous.

CD40 is a type I membrane protein. That is, it consists of an (amino-terminal) extracellular domain, a transmembrane region and a (carboxy-terminal) cytoplasmic region. CDNAs encoding CD40 are described, for example, in the panning method [Stamenkovic, I, et al. (1989) EMBO J., Vol. 8, pp. 1403-1410] can be used to isolate from cDNA banks of Raji cells. Various views, well known to the expert, are directed to causing expression of membrane-bound proteins, typically in soluble form. For example, in a polymerase chain reaction using a mutagenic primer, stop codons can be introduced at the ends of cDNA sequences encoding extracellular regions. The cDNA modified in this way then encodes a CD40 protein secreted by the cell similarly to other secreted proteins because it lacks a membrane anchor. Fanslow, W. C et al. (1992) J. Immunol., Pp. 655-660]. It may also be advantageous to use soluble recombinant immunoglobulin fusion proteins prepared by the preparations described in the examples in EP-A-0 464 533. In particular, CD40 / Ig fusion proteins are also described in Fanslow et al. (1992) J. Immunol., Vol. 149, pp. 655-660 and Noelle et al. (1992) Proc. Natl. Acad. Sci USA, Vol 89, pp. 6550-6554). Two publications describe a CD40 / Ig fusion protein consisting of the extracellular region of CD40 fused to a hinge, the CH2 and CH3 regions of the heavy chain of a human immunoglobulin G1 molecule. To prepare the corresponding DNA constructs, suitable restriction cleavage sites can be introduced into the CD40 cDNA using polymerase chain reaction using mutagenesis primers [Fanslow et al., (1992) J. Immunol., Vol. . 149, pp. 655-660] Use of naturally occurring restriction cleavage sites of CD40 cDNA (Noelle et al., (1992) Proc. Natl. Acad. Sci USA, Vol, 89, pp. 6550-6554].

Soluble forms of CD40 can be expressed in prokaryotic or eukaryotic systems known as recombinant proteins, but can preferably be expressed in mammalian cell culture and purified from culture supernatants or cell lysates by conventional methods. Regardless of the possible direct therapeutic use of soluble CD40 molecules, it can be used to identify other substances with therapeutic potential by blocking the interaction of membrane-bound CD40 and CD40 ligands. This can occur, for example, in cell-free receptor binding assays in which soluble CD40 molecules are present in the solid phase and binding of such soluble CD40 ligands takes place using suitable labeling or antibodies. Because of their automation potential, this type of analysis provides a method (receptor screening) for investigating a number of substances for the interaction of CD40 / CD40 ligands with them.

See Idzerda et al. (1990, J. Exp. Med. 171, 861-873) and Maliszewski et al. (1990, J. Immunol. 144, 3028-3033), in the examples detailed below, methotrexate and G418 (EP-A-0 330 977) and human and rat IL-4 receptors secreted into culture medium by stable expression of BHK cells as soluble proteins after double selection and purified by immunoaffinity chromatography (hereafter huIL-4R and naturally occurring soluble forms of the muIL-4S and extracellular domains are used. In addition, extracellular regions of human or murine IL-4 receptors and hinges, CH2 and CH3 regions of human IgG1 or murine IgG2b molecules (see Zettlmeiβl et al. (1190) DNA and Cell Biol. 9, 347-353] (hereinafter referred to as huIL-4R / Fc and muIL-4R / Fc, respectively) and likewise expressed in BHK cells and purified by protein A-Sepharose affinity chromatography Globulin fusion protein (EP-Al-0 464 533) is used.

IL-4R and IL-4R / Fc are equally effective in neutralizing the same species of interleukin-4 in each case in the bioassay (Example 1). In addition, inhibition of human T-cell clone of type TH2 can be detected by huIL-4R / Fc in vitro. IL-4 formed by these clones in vitro is neutralized by huIL-4R / Fc. This reduces proliferation. Proliferation of control clones of type TH1 is not affected by huIL-4R / Fc (Example 4).

In addition, according to the present invention huIL-4R / Fc is synthesized in vitro by human peripheral blood mononuclear cells induced by IL-4 of IgE (Example 2), and surprisingly also antibody-specific synthesis (Example 3) can be suppressed. Surprisingly, an animal model of rat listeriosis, an animal model of Cysticercus crassiceps infection (mouse), an animal model of systemic lupus erythematosus (MRL / 1 mouse), chronic graft-versus-host response As can be demonstrated using the example (Example 5) for soluble interleukin-4 receptors in rats in animal models (mouse) and animal models for allergen-induced asthma, the infection and disease of the immune system It has a therapeutic or prophylactic effect.

Example 1

Biological Activity of Variants of Human and Rat IL-4R and IL-4R / Fc

Biological activity is measured by bioassay. IL-4 binds strictly to the IL-4 receptor. For this reason, cell lines of murine origin and murine IL-4 receptors are associated with the membrane are used. This cell line is transfected with a complete gene for the human interleukin-4 receptor. Mice and human membrane-bound receptors are functionally expressed simultaneously by these cell lines and the proliferation of these cell lines depends on both IL-4 of mice and humans. See Mosley et al., Cell, Vol. 59, 335-348 October 20, 1989].

Murine IL-4 is used to detect muIL-4R and muIL-4R / Fc (Table 1). The proliferation of this cell line depends on rat IL-4 (Table 1). Constant use of interleukin-4 and simultaneous addition of muIL-4R or muIL-4R / Fc result in concentration-dependent neutralization of interleukin-4. Interleukin-4 decreases proliferation because it is less effective against cell lines. Control proteins with the same Fc site do not show this effect. The difference between muIL-4R and muIL-4R / Fc in concentration-dependent inhibition in Table 1 may be due to different molecular weights.

Table 2 shows the bioactivity of human interleukin-4 and the specific neutralization action of huIL-4R and huIL-4R / Fc corresponding to Table 1. The difference between huIL-4R and huIL-4R / Fc in concentration-dependent inhibition may be due to different molecular weights.

Example 2

Inhibition of IL-4-induced IgE Synthesis by huIL-4R / Fc

Human peripheral heel liquid mononuclear cells are isolated and incubated for 48 hours in Iscove's Cultule medium containing 5% FCS with 1 × 10 6 cells in culture volume Iml per culture well in 24-well plates. 300 ng / ml human phosphorus-4 is added to the medium. Membrane-bound IgE already present is released into the culture medium during this prestimulation. After 48 hours, cells are washed to remove released IgE. The cells are subsequently cultured in isocove culture medium containing 5% FCS at 1 × 10 5 cells in a 200 μl culture volume in a flat 96-well culture plate. To this medium add 0 g / ml, 3 ng / ml and 30 ng / ml Interleukin-4. The batch is performed with or without 3 μg / ml huIL-4R / Fc or huCD4 / Fc in each case. huCD4 / Fc acts as a control for the Fc region of IL-4R / Fc and is prepared as described in EP-A2-0325 262. The Fc region of huCD4 / Fc and huIL-4R / Fc are identical (IgG1). After incubation for 10 days, the culture medium is removed and assayed in ELISA for human IgE. Table 3 shows the results of this test. IL-4-induced IgE synthesis is specifically inhibited by the addition of HuIL-4R / Fc.

Example 3

Isolate human peripheral blood mononuclear cells. 5000 cells per well in a 96-well culture plate are incubated for 14 days with 200 μl of isocove culture medium containing 10% FCS. At the beginning of the cultivation, nothing further is added or purified mite antigen [Derm. Pt., Order no. Sq 503, manufactured by Scherax, Hamburg] 100 SQ units (manufacturer description) alone or with a tick antigen and 3 μg / mL of huIL-4R / Fc are added. A mixture of tick antigen and 3 μg / ml of TW1 acts as an Fc control. TW1 is a human monoclonal antibody that has specificity for rabbit antigens and has the same isotype as huIL-4R / Fc. TW1 is prepared as described in European Patent Application No. 0 445 625 Al. The supernatant is then removed and assayed for human IgE by ELISA. It was found that antigen-induced production of IgE was specifically inhibited by huIL-4R / Fc (Table 4).

Example 4

Inhibition of antigen-induced proliferation of T-cell clones of TH2 subtype

T-cell clones are isolated from the skin of patients with atopic dermatitis. After stimulating cytokines of type TH2 in vitro, clones are secreted at high rates and type TH1 is found only at consumption. Proliferation assays are performed in 96-well microtiter plates with 1 × 10 4 clone T cells and 1 × 10 5 irradiated autologous PBLs as stimulatory cells in 4% human AB serum. This incubation is carried out for 18 hours in a 37 ° C. incubator while passing CO 2 with 0.5 μCi / culture well of 3H-thymidine.

After 18 hours, cell proliferation is measured using incorporated thymidine. For cytokine production in supernatants, 1 × 10 6 / ml cloned T cells are stimulated with isocobe, 10% FCS and 10 μg / m concanavalin A in 24-well culture plates. The supernatant incubated for 24 hours is removed while passing CO 2 in a 37 ° C. incubator. This culture supernatant is assayed for the content of human interleukin-2 and human interleukin-4 by bioassay.

This bioassay is based on cell lines whose proliferation depends on human interleukin-2 and human interleukin-4, respectively. Mosley et al. Cell, Vol. 59, 335-348, 20 October 1989]. IL-4 is detected in large quantities in one supernatant of TH2 clones, but not IL-2. By adding 3 μg / ml huIL-4R / Fc, only 10 ng / ml IL-4 can be detected instead of 49 ng / ml IL-4. Supernatants of TH1 clones contain relatively large amounts of IL-2 and only very small amounts of IL-4. Interleukin-2 concentration in the supernatant of TH1 clone was not significantly affected by the addition of huIL-4R / Fc during incubation. Proliferation experiments demonstrate that the proliferation of TH2 clones but not TH1 clones is affected by the addition of 3 μg / mL of huIL-4R / Fc (Table 5). It is also possible to demonstrate that antigen-specific proliferation of tick-specific TH2 clones other than TH1 clones is inhibited by the addition of huIL-4R / Fc. Further experimental studies may demonstrate that IgE synthesis induced by tick antigens in a mixture with B lymphocytes of autologous tissue from the same donor as the TH2 clone can be inhibited by the addition of huIL-4R / Fc.

Example 5

Inhibition of skin test response and other variables by treatment with muIL-4R after lung allergy in mice

Balb / c mice are sensitized by inhaling an ovalbumin solution (500 μg / 7 mL aerosol in PBS. This procedure is described in Renz et al. 1992, J. Allergy Clin. Immunol. 89: 112). This aerosol is prepared in an ultrasonic nebulizer (Pulmosonic Model 25, Ths De Vilbiss Co, Somerset, PA.) More than 90% of the particles produced are 1-5 μm in size. Exposure to ovalbumin-containing aerosol for 20 minutes on days 7, 7, 14 and 21. To test the effects of soluble muIL-4R in the model, FBS and 11B11 (rat monoclonal antibody against rat IL-4, Ohara , J. and WI Poul, 1985, Nature 315: 333) are administered on days -2, -1, 1, 2, 7, 8, 14, 15, 21 and 22, respectively.

On day 23 of the experiment, skin tests are performed using ozalbumin (OVA) for each group. This process is described in Saloga et al. 1993, J. Clin. Invest. 91,133-140.

The results are summarized in Table 6 and show a marked decrease from 83.3% to 22.2% in the number of animals with a positive skin test response.

The effect of muIL-4R on OVA-specific IgE and tracheal contraction of isolated organs of these animals are investigated in the same experimental system. This research method is described in Renz, H. et al. 1992, J. Allergy Clin, Immunology, 89: 112 and Larsen et al. 1992, J. Clin. Invest, 89747. This experiment is performed using the same albumin and muIL-4R administration mode and the same dosage as described above in this example.

The results are summarized in Table 7. Tracheal contraction is performed by electric field stimulation ex vivo (ES50 unit data). IgE concentrations are determined from serum by ELISA (relative units of data based on rat control serum with high anti-VA titers). This serum is taken on day 23 of the experiment. This result shows a pronounced normalization of organ function and a decrease in IgE titers of 2.47 to 3.4 ES50. OVA-specific IgG1 was measured in parallel and similarly reduced (not described above). An increase in IgG2a is also observed at the same time.

Lung sensitization with ovalbumin (using the same mode of administration, dose, and method as described above in the same example), and skin test response after lung treatment with muIL-4R in parallel in the same experimental system as described above Try it. For this experiment, muil-4R 500 / μg in 7 ml of PBS was similarly used with the obalbumin method using an ultrasonic nebulizer on days -2, -1, 1, 2, 7, 8, 14, 15, 21 and 22, respectively. Spray with aerosol. Animals are exposed to aerosols for 20 minutes in closed containers (4 animals per nebulizer). This is ensured by a preliminary test that maintains the biological activity of the muIL-4R protein in the ultrasonic spray phase. In addition, the animals are set to inhale about 0.5-5% of the 500 μg muIL-4L total amount. The skin test is performed on day 23 of the experiment described above. The results are shown in Table 8. Although small doses of muIL-4R per animal were administered (the final dose inhaled was 0.5-5% / animal of 125 μl muIL-4R), the topical applied skin test effect was similar to that of intraperitoneal treatment (150 μg / animal). It is observed to be nearly identical.

Example 6

Effect of Interleukin-4 Receptor on Induced Chronic Graft-versus-Host Disease (cGvHD)

CGvHD in female B6D2F1 hybrid animals (parent species C57Bl / g and DBA / 2) weighing 15-18 g by intravenous injection of 1 × 108 spleen cells from DBA / 2 mice per injection for four consecutive weeks. Induce. Expressed proteinuria is monitored by the N-comber test (Boehringer Mannheim GmbH, Hannheim, Germany). 10 animals are used per group. One group received 2 mg / kg of mouse interleukin-4 receptor (muIL-4R / Fc) at 27, 28, 29, 30 and 31 days, respectively, and the other group, human interleukin-4 receptor ( huIL-4R / Fc) at the same dose and mode of administration, and one control is treated with PBS. Table 9 shows the surviving animals as fractions for the animals in the group, and Table 10 shows the change in the mean protein levels in the urine of the animals in the various groups. Significant effects of muIL-4R can be seen in both the number of surviving animals and proteinuria. Another experiment is performed by inducing cGvHD as described above. The group classification is the same, but treatment occurs after glomerulonephritis on days 63, 64, 65, 66 and 67, i.e. therapeutically (2 mg / kg huIL-4R / Fc or muIL-4RFc administered on the same day, respectively) ) Fractions of surviving animals and protein levels in animal urine are shown in Tables 11 and 12, respectively. In another experiment in which cGvHD was induced identically and grouped identically, monomeric muIL-4R or monomeric huIL-4R was administered intravenously five times every three days, beginning at day 24 at 2 mg / kg per week. . Again, similar to the previous experiments of this example, an increase in animal survival and significant inhibition of the cGvHD factor was observed in the group treated with muIL-4R. In this experiment, blood is taken from the animal during the experiment. The immunoglobulin E (IgE) of rats in these sera is measured by ELISA, where the IgE concentration increases during the disease and decreases again after measurement on day 59 in the above experiments. When treated with muIL-4R, after the measurement on day 45, this reduction begins early. Table 13 shows the change in mean IgE concentration in three animals randomly selected from the group treated with muIL-4R and the control group (treated with PBS).

Example 7

Treatment of Spontaneous Autoimmune Disease in MRL / 1 Mice That Resemble Human Systemic Lupus Erythematosus

Approximately 12-week-old MRL / Mp-Lpr-Lpr (MRL / 1) mice were treated for 5 consecutive days in buffer control (PBS), mouse interleukin-4-receptor-Fc fusion protein (muIL-4R / Fc) or human interleukin-4 Receptor-Fc fusion protein (huIL-4R / Fc) is treated by intravenous administration as a control protein. Remove the spleen, lymph nodes and blood. The spleen and lymph nodes are weighed and various variables from blood or serum are measured. Antibodies against double-stranded DNA (dsDNA) and rheumatoid disease factor (RF) are described in reference; Schorlemmer et al. , Int. J. Immunotherapy 7: 169, 1991], as measured by ELISA. Changes in viability are shown in Table 14 and other variables at week 25 are shown in Table 15. The marked action of muIL-4R / Fc on disease is evident from both changes in the number of surviving animals and other variables at week 25.

Example 8

Treatment of Cystycerus (Tania) Krasiceps Infection in Mice Using the Interleukin-4 Receptor in Mice

For experiments, on day 0, NMRI mice weighing 15 to 25 g were taken, each of 5 metacestodes of cystineserjus (Tania) Kracisces strain MR-1 in 1 ml of PBS per mouse. Infection by intraperitoneal injection. Ten animals are used per experimental group. One control group is not infected (injected with metacetodes free PBS). One group is infected but not treated. One group is infected and treated intraperitoneally with 100 μg muIL-4R on days -1, 0, 1, 7, 14, 20, 30, 35, 40, 45, 50, 55 and 63, respectively. Another group is infected and treated with huIL-4R as a control protein similar to treatment with muIL-4R. On day 76, animals are weighed and dissected. Animals that can find metacestodes in the abdominal cavity are measured. The wet weight of metacetodes found in each animal is measured for the positive animal. A significant decrease in the overall weight of metacetodes is found in the group treated with the murine interleukin-4 receptor (Table 16).

Example 9

Effect of muIL-4R on Rat Candida albicans Infection

Hybrid mice (BALB / cCrxDBA / 2Cr) F1 (CD2F1) females are infected with highly pathogenic Candida albicans strain CA-6 (5 × 10 4 cells, intravenously). Animals were administered intraperitoneally with 100 μg of muIL-4R in 250 μl of PBS 24 times before infection once on day -1, twice on day 0 and day 1 and once on day 2 and day 3 respectively. Meanwhile, control animals receive mouse serum albumin (MSA), huIL-4R or PBS purified in the same manner. All animals treated with mouse serum albumin (16), huIL-4R (16) and PBS (24) result in progressive Candida infection and exhibit an average survival time of 14 to 17 days after infection. 91.6% (23) of the animals treated with muIL-4R survive. These surviving animals are reinfected with Candida albicans (106 cells) 8 weeks after infection. All animals survived this reinfection. In normal untreated animals such infections result in lethal infections with a survival time of two to three days after infection.

Table 1

Bioactivity of the Rat Interleukin-4 Receptor

TABLE 2

Bioactivity of the Human Interleukin-4 Receptor

TABLE 3

Inhibition of Human IgE Synthesis

Table 4

Inhibition of synthesis of allergen-specific IgE

Table 5

Tick-specific T-cell clones from the skin of atopic dermatitis patients

Table 6

Inhibition of skin test response by muIL-4R

TABLE 7

Inhibition of antigen-specific IgE after organ contraction by muIL-4R

Table 8

Lung sensitization and reduced number of positive skin tested animals by pulmonary administration of muIL-4R

Table 9

Effect of Interleukin-4 Receptor on Induced Chronic Graft-versus-Host Disease (cGvHD)

Table 10

Effect of Interleukin-4 Receptor on Induced Chronic Graft-versus-Host Disease (cGvHD)

Table 11

Effect of Interleukin-4 Receptor on Induced Chronic Graft-versus-Host Disease (cGvHD)

Table 12

Effect of Interleukin-4 Receptor on Induced Chronic Graft-versus-Host Disease (cGvHD)

Table 13

Effect of Interleukin-4 Receptor on Induced Chronic Graft Versus Host Disease (cGvHB)

Table 14

Treatment of Spontaneous Autoimmune Disease in MRL / 1 Mice That Resemble Human Systemic Lupus Erythematosus

Table 15

Treatment of Spontaneous Autoimmune Disease in MRL / 1 Mice That Resemble Human Systemic Lupus Erythematosus

Table 16

Treatment of Mice Infected with Cystyrcus (Tania) Krasiceps Using the Rat Interleukin-4 Receptor

Claims (11)

From the group consisting of retroviruses, suprafungus, yeast-like fungi, mycobacteria, listeria, protozoa, tapeworms or worm infections, which harbor the IL-4 receptor (IL-4R) or extracellular domain of IL-4R. A pharmaceutical composition for the treatment and prevention of infectious diseases in which the occurrence of selected TH-type T-helper cells is increased. The pharmaceutical composition of claim 1, wherein the extracellular region of the IL-4 receptor or IL-4R is a member of a fusion protein. The pharmaceutical composition of claim 2, wherein the fusion protein contains an extracellular region of an IL-4 receptor or IL-4R and an Fc region of an antibody. The method of claim 1, wherein the infection is human immunodeficiency virus (HIV), Mycobacterium leprae, Leishmania, Plasmodium, Schistozoma, Nipostrongills, Triturida, Trichinella, A pharmaceutical composition that is Taenia (cystisercus), Candida, Aspergillus, Cyclopirida, or Heligmosomosides infection. The pharmaceutical composition according to any one of claims 1, 2 and 3, wherein the extracellular region of IL-4k or IL-4R or the IL-4R / Fc fusion protein is used as a mixed product. The pharmaceutical composition of claim 5, wherein the mixed product contains gamma-interferon. 6. The pharmaceutical composition of claim 5, wherein the mixed product contains allergens. The pharmaceutical composition of claim 5, wherein the mixed product contains one or more substances that inhibit the interaction of cell surface protein CD40 ligand with cell surface protein CD40. The pharmaceutical composition of claim 8, wherein at least one of the substances is a soluble determinant of the CD40 surface molecule. The pharmaceutical composition according to any one of claims 1, 2 and 3 for topical administration. The pharmaceutical composition of claim 10 for intradermal, subcutaneous, dermal, nasal or pulmonary administration.