WO2020198832A1 - PEPTIDES SYNTHÉTIQUES À AFFINITÉ POUR LE RÉCEPTEUR DE TGF-β, COMPOSITION PHARMACEUTIQUE ET UTILISATION DE CEUX-CI COMME IMMUNOMODULATEURS DANS LE TRAITEMENT DE MALADIES AUTO-IMMUNES, INFLAMMATOIRES OU ALLERGIQUES - Google Patents

PEPTIDES SYNTHÉTIQUES À AFFINITÉ POUR LE RÉCEPTEUR DE TGF-β, COMPOSITION PHARMACEUTIQUE ET UTILISATION DE CEUX-CI COMME IMMUNOMODULATEURS DANS LE TRAITEMENT DE MALADIES AUTO-IMMUNES, INFLAMMATOIRES OU ALLERGIQUES Download PDF

Info

Publication number
WO2020198832A1
WO2020198832A1 PCT/BR2020/050117 BR2020050117W WO2020198832A1 WO 2020198832 A1 WO2020198832 A1 WO 2020198832A1 BR 2020050117 W BR2020050117 W BR 2020050117W WO 2020198832 A1 WO2020198832 A1 WO 2020198832A1
Authority
WO
WIPO (PCT)
Prior art keywords
seq
peptides
pep
cells
diseases
Prior art date
Application number
PCT/BR2020/050117
Other languages
English (en)
Portuguese (pt)
Inventor
Luiz Ricardo Goulart Filho
Fatima D. FERREIRA-BRIZA
Emília Rezende VAZ
Galber Rodrigues ARAUJO
Carlos Ueira VIEIRA
Lorenz AGLAS
Original Assignee
Universidade Federal de Uberlândia
Cirino Alberto Goulart Eireli Epp (Biogenetics)
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by Universidade Federal de Uberlândia, Cirino Alberto Goulart Eireli Epp (Biogenetics) filed Critical Universidade Federal de Uberlândia
Publication of WO2020198832A1 publication Critical patent/WO2020198832A1/fr

Links

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P29/00—Non-central analgesic, antipyretic or antiinflammatory agents, e.g. antirheumatic agents; Non-steroidal antiinflammatory drugs [NSAID]
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/02—Immunomodulators
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P37/00—Drugs for immunological or allergic disorders
    • A61P37/08—Antiallergic agents
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K7/00—Peptides having 5 to 20 amino acids in a fully defined sequence; Derivatives thereof
    • C07K7/04—Linear peptides containing only normal peptide links
    • C07K7/08—Linear peptides containing only normal peptide links having 12 to 20 amino acids

Definitions

  • the present invention relates to the selection and characterization of synthetic peptides, which have an affinity for receptors present in cells of the immune system, more particularly with affinity for the receptor of the Growth Transformation Factor - beta (of the acronym in English: TGF -B). These receptors are known to play an important role in modulating the immune, inflammatory and allergic responses.
  • the present invention also describes pharmaceutical compositions comprising at least one of these synthesized synthetic peptides and the use of these peptides to prepare medicaments or immunogenic compositions for the treatment and / or prophylaxis of diseases where there is an immunological disorder, more particularly in which the related diseases immunological disorders are: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • TGF transforming growth factor
  • pi The transforming growth factor (TGF) -pi is a pleiotropic cytokine that acts on several biological processes, including embryonic development and cell proliferation, differentiation, adhesion, migration and apoptosis. Its signaling has been proposed for the control of self-reactive peripheral T cells, present in patients with autoimmune diseases. This cytokine also plays an important role in the induction of auxiliary T cells capable of regulating cytokine production and activation of the pathway in order to repair the immune response.
  • TGF-bI is produced by several cell types, such as platelets, neutrophils, malignant cells, dendritic cells and macrophages. Consequently, this cytokine has an important effect on inflammatory, allergic and autoimmune diseases. Due to the diversity of TGF-bI, many factors are responsible for activating canonical or non-canonical pathways to induce multiple biological effects and interfere with different pro-inflammatory pathways. This cytokine can modulate different pathways directly related to TGF-b receptors and also in response to crosstalk signaling.
  • TGF-bI is a cytokine that plays an important role in allergic inflammation. It can suppress effector T cells and is involved in the development of conventional, innate and regulatory (Treg) T cells. T cells play a key role in maintaining peripheral tolerance against commensal, autologous and environmental antigens. In mice with the interrupted TGF-bI gene, and in mice without the II receptor (TORbRII), severe inflammatory responses, tissue necrosis and early death were observed, confirming the regulatory role of T ⁇ Rb1 in peripheral tolerance.
  • TORbRII II receptor
  • Allergic reactions are initiated when an allergen together with IgE binds to the high affinity Fc RI receptor on mast cells, which generally serves to alert the immune system to local infection. Once activated, mast cells induce inflammatory reactions by secreting chemical mediators; synthesizing leukotrienes, prostaglandins and proinflammatory cytokines, such as IL-4, IL-5 and IL-13; and recruiting other cells effectors, notably TH2 lymphocytes, basophils and eosinophils, which contribute to the exacerbation of the allergic response.
  • Treg cells are a heterogeneous population characterized by the constitutive expression of the transcription factor Foxp3. They are necessary for the maintenance of self-tolerance and immune homeostasis, and comprise several subsets. Among these subsets, Tregs TH3 cells (CD4 + CD25 + Foxp3 +), which have regulatory action through the production of TGF-bI, have been the focus of many studies. TH3 cells derived from the thymus are essential for immune tolerance against autoimmune and inflammatory diseases, allergies and several other events related to the breakdown of immune homeostasis. In addition, Treg cells consistently represent the specific dominant subgroup for common environmental allergens in healthy individuals.
  • Grass pollen is one of the most frequent allergenic sources responsible for IgE-mediated respiratory allergies. Approximately 50% of all patients with type I allergy are sensitized to grass pollen proteins. Phl p 5 is one of the main allergens found in grass pollen (Phleum pratense) and has great potential to induce TH2 response in mice. Allergen immunotherapy is a common treatment for patients with various allergic disorders. Allergen-specific immunotherapy (ASIT) involves administering different doses of allergenic extracts over 3-5 years to achieve clinical tolerance. In clinical practice, ASIT is most often performed through subcutaneous injections, which can lead to low rates of adhesion due to allergic local and systemic side effects.
  • Phage Display (PD) methodology may be a good option, as it allows us to select small molecules that can mimic specific amino acids of large proteins and induce the same or better response as expected with the natural molecule or in comparison with other recombinant molecules already described.
  • Peptides selected by PD have low toxicity and high receptor / protein affinity.
  • Therapeutic mimetic peptides are potential drug candidates for many different diseases, as they have the potential to antagonize, stimulate, increase or modulate the activity of natural ligands, binding to specific receptors on the cell surface and triggering intracellular pathways. Considering that numerous membrane-bound proteins can interact with members of the TGFp superfamily and control the availability and specificity of the ligand, short peptides that mimic the TGFpi binding domain could potentially bind to TGFpRIl with greater affinity and positively impact its target cells. .
  • patent WO2012167143 A1 reports the use of the phage display technology in obtaining a specific antibody binding to the TGF-b molecule that can be used in patients who have a disorder related to TGF-b expression, such as cancer. These results show the efficiency of the technique in the development of recombinant antibodies in the treatment of diseases
  • Patent application BR 102015003903-4 describes 27 synthetic peptides and their inverse sequences, selected by Phage Display, which have an affinity for the TGF-b receptor. Said patent application then describes the use of these peptide sequences synthesized in pharmaceutical compositions in the regulation of the immune system, in order to obtain control of inflammation or to allow the treatment of inflammatory and autoimmune diseases. Said patent application also describes the possibility of using such synthesized peptides in diagnostic or prognostic methods of related diseases.
  • the present patent application aimed to select new peptides similar to TGF-b, by PD technology, which had improved ability to modulate the immune response, more particularly, to control inflammatory and allergic responses.
  • the present patent application describes synthetic peptides with affinity for the TGF-b receptor. These peptides were synthesized and their three-dimensional structures were predicted. All of them were synthesized to present Histidine and Alanine as the first and second amino acid residues and a sequence of three Glycines and one Serine as the last four amino acid residues. Such peptides showed important activity on immunological modulatory and regulatory pathways. Therefore, they can be used in the treatment and / or prophylaxis of diseases related to immune disorders, such as: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • diseases related to immune disorders such as: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • Another embodiment of the present application is the methods of treatment and / or prophylactic of diseases related to immunological disorder, in which the diseases related to immunological disorder are: chronic or acute inflammatory diseases, allergic and / or autoimmune, comprising administering to a subject a pharmaceutically effective dose of at least one of the described synthetic peptides.
  • compositions that comprise at least one of the synthetic peptides described herein, plus at least one pharmaceutically acceptable carrier, carrier and / or compound.
  • the Pharmaceutical Composition is for use in the treatment and / or prophylaxis of diseases related to immunological disorders, in which the diseases related to immunological disorders are: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • Another embodiment of the present application is the use of the synthetic peptides or pharmaceutical compositions described herein, for the preparation of a drug or immunogenic composition for use in the treatment and / or prophylaxis of diseases related to immunological disorder, in which the diseases related immunological disorders are: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • FIGURE 1A shows the predicted three-dimensional structure of Pep
  • FIGURE 1 B shows Pep's predicted three-dimensional structure
  • FIGURE 1C shows Pep's predicted three-dimensional structure
  • FIGURE 1 D shows the predicted three-dimensional structure of Pep 4.
  • FIGURE 1 E shows the predicted three-dimensional structure of Pep
  • FIGURE 1 F shows Pep's predicted three-dimensional structure
  • FIGURE 1 G shows Pep's predicted three-dimensional structure
  • FIGURE 1 H shows Pep's predicted three-dimensional structure
  • FIGURE 2 shows an analysis of aggregation behavior of peptides synthesized in solution analyzed by DLS.
  • Figure 3 shows an analysis of recognition by the anti-TGF-bI antibody of the synthesized peptides, on the cell surface of the A549 strain.
  • Figure 4A shows activation of the dependent TFGPRII / SMAD pathway, by analyzing the production of SEAP (secreted alkaline phosphatase) after treatment with the peptides synthesized here for 12 hours.
  • Figure 4B shows the activation of the dependent TFGPRII / SMAD pathway, by analyzing the production of SEAP (secreted alkaline phosphatase) after treatment with the synthesized peptides described here for 24 hours.
  • SEAP secreted alkaline phosphatase
  • Figure 4C shows the activation of the dependent TFGPRII / SMAD pathway, by analyzing the production of SEAP (secreted alkaline phosphatase) after treatment with the synthesized peptides described here for 48 hours.
  • SEAP secreted alkaline phosphatase
  • Figure 5A shows an analysis of the effect of peptides synthesized in the NFkB pathway, after pre-treatment with the peptides for 1 hour, followed by TNF-a stimulation and analyzed after 12 hours.
  • Figure 5B shows an analysis of the effect of peptides synthesized in the NFkB pathway, after pre-treatment with the peptides for 1 hour, followed by TNF-a stimulation and analyzed after 24 hours.
  • Figure 5C shows an analysis of the effect of peptides synthesized in the NFkB pathway, after pre-treatment with the peptides for 1 hour, followed by TNF-a stimulation and analyzed after 48 hours.
  • Figure 5D shows an analysis of the effect of peptides synthesized in the NFkB pathway, after pre-stimulation with TNF-a for 1 hour, followed by the addition of the peptides and analyzed after 12 hours.
  • Figure 5E shows an analysis of the effect of peptides synthesized in the NFkB pathway, after pre-stimulation with TNF-a for 1 hour, followed by the addition of the peptides and analyzed after 24 hours.
  • Figure 5F shows an analysis of the effect of peptides synthesized in the NFkB pathway, after pre-stimulation with TNF-a for 1 hour, followed by the addition of the peptides and analyzed after 48 hours.
  • Figure 6A shows analysis to investigate the action of peptides in the TLR4 pathway, after pretreatment with the synthesized peptides, followed by LPS stimulation and analyzed after 12 hours.
  • Figure 6B shows analysis to investigate the action of peptides in the TLR4 pathway, after pre-stimulation with LPS, followed by the addition of the peptides and analyzed after 12 hours.
  • Figure 7A shows an ELISA assay to assess the binding of rTGF1 1 and Pep 2 to the Jurkat cell surface.
  • Figure 7B shows a luciferase reporter assay showing IL-8 gene expression in A549 cells stimulated with TNF-a and treated with Pep 2.
  • Figure 7C shows an IL-8 production / secretion assay in A549 cells stimulated by TNF-a and treated with Pep 2.
  • Figure 7D shows a test of the action of Pep 2 on the secretion of
  • Figure 7E shows a test of the action of Pep 2 on the secretion of
  • Figure 7F shows a mediator release assay based on rat basophilic leukemia (RBL) cells that have a humanized Fc RI receptor to investigate the effect of Pep 2 on IgE-mediated basophil degranulation.
  • RBL rat basophilic leukemia
  • Figure 8A shows an immunization schedule in animals.
  • Figure 8B shows an immunization assay with recombinant Phl p5, showing the action of Pep 2 on the degranulation of basophils.
  • Figure 8C shows a recombinant Phl p5 immunization assay, showing the action of Pep 2 on IgE levels.
  • Figure 8D shows an immunization assay with recombinant Phl p5, showing the action of Pep 2 on lgG1 levels.
  • Figure 8E shows a recombinant Phl p5 immunization assay, showing the action of Pep 2 on levels of lgG2a.
  • Figure 8F shows an immunization assay with recombinant Phl p5, showing the action of Pep 2 on IgA levels.
  • Figure 9A shows an immunization assay with recombinant Phl p5 in splenocytes, showing the action of Pep 2 on IFN-g levels.
  • Figure 9B shows an immunization assay with recombinant Phl p5 in splenocytes, showing the action of Pep 2 on IL-4.e levels
  • Figure 9C shows an immunization assay with recombinant Phl p5 in splenocytes, showing the action of Pep 2 in inducing IL-10.
  • Figure 9D shows a re-immunization assay with recombinant Phl p5 in splenocyte supernatant, showing the action of Pep 2 on IFN-g levels.
  • Figure 9E shows a re-immunization assay with recombinant Phl p5 in splenocyte supernatant, showing the action of Pep 2 on IL-2 levels.
  • Figure 9F shows a re-immunization assay with recombinant Phl p5 in splenocyte supernatant, showing the action of Pep 2 on IL-4 levels.
  • Figure 9G shows a re-immunization assay with recombinant Phl p5 in splenocyte supernatant, showing the action of Pep 2 on IL-5 levels.
  • Figure 9H shows a re-immunization assay with recombinant Phl p5 in splenocyte supernatant, showing the action of Pep 2 on IL-13 levels.
  • Figure 9I shows a re-immunization assay with recombinant Phl p5 in splenocyte supernatant, showing the action of Pep 2 in the induction of IL-10.
  • Figure 10A shows an assay to investigate CD4 cells
  • Figure 10B shows an assay to investigate CD4 cells
  • Figure 10C shows an assay to investigate CD4 cells
  • Figure 10D shows an assay to investigate CD4 cells
  • Figure 10E shows an assay to investigate CD4 cells
  • Figure 10F shows an assay to investigate CD4 cells
  • Figure 1 1A shows the analysis of GFP + CD4 + lymphocytes isolated from 4get mice 5 days after immunization Mice.
  • Figure 1 1 B shows that the synthesized peptide administered over the grass pollen extract suppresses the polarization of TH2.
  • Figure 11 C shows that the synthesized peptide administered throughout the grass pollen extract suppresses the polarization of TH2.
  • the present patent application describes 8 synthetic peptides with TGF-b receptor affinity, which comprise or consist of the amino acid sequences: peptide 1 (Pep 1): SEQ ID NO 2 01; peptide 2 (Pep 2): SEQ ID NO 2 02; peptide 03; peptide 4 (Pep 4) SEQ ID NO 2 04; peptide 5 (Pep 5) SEQ ID NO 2 05; peptide 6 (Pep 6) SEQ ID 06; peptide 7 (Pep 7) SEQ ID NO 2 07; and peptide 8 (Pep 8) SEQ ID NO 2 08.
  • Such peptides showed important activity on immune modulatory and regulatory pathways. Therefore, they can be used in the treatment and / or prophylaxis of diseases related to immune disorders, such as: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases
  • the peptides described in the present patent application were selected using the Phage Display (PD) methodology. The selection was carried out by evaluating the ability of said peptides to recognize the TGF-bI receptor in A549 cells, using an ELISA assay. 96-well MaxiSorp microtiter plates were coated with 1x10 6 cells diluted in PBS and incubated overnight at 4 ° C. The cells were blocked with 3% PBS-BSA for 1 h at 37 ° C, washed once with PBS and incubated with pg / ml of each TGF-bI-like peptide or 5 ng / ml of recombinant TGF-bI for 1 ha 37 ° C.
  • PD Phage Display
  • the plate was incubated for 1 h at 37 ° C with anti-TGF-bI antibody.
  • the plate was washed and incubated with HRP-labeled anti-human IgG antibody for 1 h at 37 ° C.
  • the TMB substrate solution was added to the plate; the reaction was stopped by heating the 2 N H2SO4 solution and read at 492 nm in a microplate reader.
  • Peptide 1 SEQ ID N-01, was synthesized inducing the formation of disulfide bridges between cystine residues and has its three-dimensional structure predicted in Figure 1A.
  • Peptide 2 (Pep 2), SEQ ID N-02, was synthesized inducing the formation of disulfide bridges between cystine residues and has its three-dimensional structure predicted in Figure 1 B.
  • Peptide 3 (Pep 3), SEQ ID NO 2 03, is free of disulfide bridges and has its three-dimensional structure predicted in Figure 1C.
  • Peptide 4 (Pep 4), SEQ ID NO 2 04, was synthesized inducing the formation of disulfide bridges between cystine residues and has its three-dimensional structure predicted in Figure 1 D.
  • Peptide 5 (Pep 5), SEQ ID NO 2 05, is free of disulfide bridges and has its three-dimensional structure predicted in Figure 1 E.
  • Peptide 6 (Pep 6), SEQ ID NO 2 06, is free of disulfide bridges and has its three-dimensional structure predicted in Figure 1 F.
  • Peptide 7 (Pep 7), SEQ ID NO 2 07, is free of disulfide bridges and has its three-dimensional structure predicted in Figure 1 G.
  • Peptide 8 (Pep 8), SEQ ID NO 2 08, is free of disulfide bridges and has its three-dimensional structure predicted in Figure 1 H.
  • the peptides synthesized comprising or consisting of a of SEQ IDs No. 01, 02 or 04 and may present a three-dimensional shape with disulfide bridges.
  • the peptides Pep 1 and Pep 5; Pep 2 and Pep 3; and Pep 4 and Pep 6; were synthesized with an identity or similarity of at least 85% between them, respectively.
  • the amino acid residues in positions three (3) and eleven (11) were replaced from Serina in the peptides Pep 3, Pep 5 and Pep 6, to Cysteine in the papteides Pep 1, Pep 2 and Pep 4.
  • Pep 1 (0.1 mM and 1 pM), Pep 2 (0.1 pM and 10 pM), Pep 3 (0.1 pM, 10 pM and 100 pM), Pep 4 (0, 1 pM:
  • the assay was performed so that approximately 2.5x10 4 cells / well (100 pl) were treated with 100 pl of all synthetic peptides in different concentrations (100 pM, 10 pM, 1 pM and 0.1 pM) , while rTGF-bI (recombinant antibody) at 100 ng / mL was used as a positive control.
  • rTGF-bI recombinant antibody
  • 20 ⁇ l of supernatant was transferred to a Greiner plate to which 80 ⁇ l of Quanti-blue solution was added. The plate was incubated for 1 hour at 37 ° C in 5% CO2 and the production of SEAP (secreted alkaline phosphatase) was detected at 655 nm. All tests were performed in triplicate.
  • the assay was performed so that a total of 3x10 5 cells / well (100 pl) was pretreated with 100 mI of a composition comprising one of the synthetic peptides at concentrations of 100 mM, 10 mM, 1 mM and 0 , 1 mM for 1 hour at 37 ° C in 5% CO 2 , followed by stimulation with TNF-a (200ng / mL) for 12, 24 and 48 hours.
  • TNF-a 200 ng / mL
  • rll_-10 0.4ng / mL
  • the assay was performed using HEK-Blue hTLR4 reporter cells (InvivoGen) to investigate the action of peptides on the TLR4 pathway.
  • a total of 2.3 x 10 4 cells / well (100 ⁇ l) was pretreated with 100 ⁇ l of composition comprising at least one of the synthetic peptides at concentrations of 100 ⁇ M, 10 ⁇ M, 1 ⁇ M and 0.1 ⁇ M for 1 hour at 37 ° C in 5% CO2 followed by LPS stimulation at 100 ng / mL for 12 hours.
  • the same stimuli were applied to cells pretreated with LPS at 100 ng / mL for 1 hour at 37 ° C in 5% CO2, followed by stimuli with peptides at 100 pM, 10 pM, 1 pM and 0.1 pM during 12 hours.
  • the stimuli were added in Quanti-blue medium and the SEAP production was measured at 655nm.
  • peptides at the same concentrations were tested alone to investigate whether they could activate the TLR4 pathway and LPS was used as a positive control.
  • Example 5 The synthesized peptides have anti-inflammatory properties and suppress IgE-mediated basophil degranulation in vitro
  • both Pep 2 and tT ⁇ RbI decreased the gene expression of IL-8 (Fig 7B), while only Pep 2 decreased IL-8 production / secretion (Fig. 10).
  • both the Pep 2 mimetic peptide and rTGFpi significantly decreased TNF-a secretion (Fig. 7D) and increased IL-10 secretion (Fig. 7E).
  • a mediator release assay based on rat basophilic leukemia (RBL) cells that have a humanized Fc RI receptor was performed to investigate the effect of synthesized peptides on IgE-mediated basophil degranulation in an already sensitized system.
  • This assay based on sensitive cells can help to understand the action of the mimetic peptide in an already established allergic microenvironment.
  • Phl p 5 grass antigen
  • Example 6 The synthesized peptides modulate the antibody response in a murine sensitization model
  • Example 7 The peptides synthesized regulate the production of cytokines in splenocytes re-stimulated with Phl p5
  • Example 8 The peptides synthesized induce the production of Treg cells
  • TH2 lymphocytes produce cytokines such as IL-4, IL-5 and IL-13 and lead to a class change in the production of B-cell immunoglobulins to IgE, which triggers the allergic reaction.
  • cytokines such as IL-4, IL-5 and IL-13
  • IgE B-cell immunoglobulins to IgE
  • IL-4 reporter mouse 4get mice express GFP as part of an IL-4 IRES-GFP bicistronic mRNA, allowing the identification of cells that express IL-4 in situ during an allergen-induced TH2 response.
  • the present patent application demonstrates the ability of the 8 synthesized peptides to bind to the TGF-b receptor.
  • the commercial anti-TGF-bI antibody was also able to recognize the synthesized peptides of the present invention.
  • the ability of the synthesized peptides, when in different concentrations, to interfere in the activation of the NFKB and TLR4 signaling pathways was also demonstrated in this application.
  • the peptides synthesized and described in the present patent application can induce the activation of different pathways and, consequently, can be used for the development of new drugs with immunomodulatory action.
  • the TGF-bI molecule binds to the receptor and activates the SMAD-dependent pathway or interferes with SMAD-independent pathways, a response that is dependent on the microenvironment.
  • the cytokine binds to TGFpRIl and TGFpi, respectively, both of which have serine / threonine activity.
  • the binding to the receptors causes the phosphorylation of SMAD2 and SMAD3 initiating the signaling.
  • SMAD2 and SMAD3 bind to Co-SMAD4, creating a protein complex capable of penetrating the nucleus and promoting the transcription of different genes.
  • SMAD6 and SMAD7 are proteins that inhibit the phosphorylation of SMAD2 and SMAD3, directly related to pro-inflammatory signaling.
  • TGF-bI can decrease SMAD7 induction and, consequently, the production of pro-inflammatory cytokines.
  • the independent SMAD pathway plays a role in signaling p38MAPK, RHO, PI3K-AKT, ERK, JNK and NF-kB12.
  • NFkB signaling is expressed in response to a pathological and inflammatory environment, thus acting on cell adhesion molecules, cytokines and other transcription genes. Therefore, controlling this pathway is an important approach to regulate a severe inflammatory response.
  • the synthesized peptides After the Jurkat cells were pretreated with the synthesized peptides, followed by TNF-a stimuli, the synthesized peptides, at a specific time and concentration, had an effect in decreasing NFKB expression.
  • Pep 8 (0.1 mM) was the one that decreased activation in 24 hours more significantly.
  • TNF-a stimuli negatively regulate TGFpRI, which is necessary to initiate the SMAD-dependent pathway, this can be explained by the fact that the cells were treated with TNF-a before or after rTGF-bI stimuli ; only at a specific time and concentration, the molecule reduced the expression of NFkB (figure not shown).
  • TNF-a stimuli can also induce SMAD7 protein expression by activating RelA / NFKB19 and, controversially, SMAD7 can disturb the formation of TRAF-2-TAK1-TAB2 / 320 complex inhibitor NFKB expression. This may be a response to how TGF-bI-like peptides interfere in the SMAD-independent pathway, blocking the expression of the SMAD7 and NFKB protein.
  • TLR4 Another important pro-inflammatory pathway analyzed in this study is TLR4, responsible for the recognition of lipopolysaccharides (LPS), leading to the expression of NFKB.
  • LPS lipopolysaccharides
  • each of the eight peptides causes a specific action according to the analyzed path and this effect is dependent on conformation, concentration and time. According to the diversity of TGF-bI in the immune system and which response the molecule can induce, the effect is dependent on several factors, such as ligands, receptors, proteins, crosstalk and other signaling pathways and the environment. Like the molecule, each peptide has a specific action according to the target and the pathways analyzed, so that this specificity can be used to modulate a specific path of interest or in combination with another peptide to modulate different routes of interest.
  • the stimuli applied by pretreating the cells with the peptides and analyzing the interference of the peptides in the already activated cell pathway give us some indication of how the peptides can be used to develop new strategies to control the inflammatory response, in chronic or acute inflammatory, allergic and / or autoimmune diseases.
  • the peptide may represent a valuable tool for the treatment of other inflammatory conditions, including autoimmune diseases.
  • all peptides synthesized and described in the present patent application have demonstrated the ability to modulate an allergic or autoimmune inflammatory response and these can be used in the development of pharmaceutical compositions, alone or in combination with other peptides or pharmaceutically acceptable compounds, such as for example allergens, to develop new treatments or to prevent chronic or acute inflammatory, allergic and / or autoimmune diseases.
  • the present patent application describes synthetic peptides with TGF-b receptor affinity comprising or consisting of an amino acid sequence selected from the group comprising: SEQ ID N 2 01, SEQ ID N 2 02, SEQ ID N 2 03, SEQ ID NO 2 04, SEQ ID NO 2 05, SEQ ID NO 2 06,
  • SEQ ID NO 2 07 and SEQ ID NO 2 08 or a sequence having at least 85% identity or similarity with one of the amino acid sequences selected from the group consisting of: SEQ ID NO 2 01, SEQ ID NO 2 02, SEQ ID NO 2 03, SEQ ID NO 2 04, SEQ ID NO 2 05, SEQ ID NO 2 06, SEQ ID NO 2 07 and SEQ ID NO 2 08.
  • Peptides comprising the sequences SEQ ID NO: 01, SEQ ID NO: 02 and SEQ ID N ° 04, which may have three-dimensional deformation with disulfide bonds, and the peptides SEQ ID N ° 03, SEQ ID N ° 05, SEQ ID N ° 06, SEQ ID N ° 07 and SEQ ID N ° 08 presenting three-dimensional structure without disulfide bridges.
  • Synthetic peptides are to be used in pharmaceutical compositions for use in the treatment and / or prophylaxis of diseases related to immunological disorders, in which the diseases related to immunological disorders are: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • compositions that comprise at least one of the synthetic peptides described herein, plus at least one pharmaceutically acceptable carrier or compound. More particularly where the pharmaceutically acceptable compound is selected from an allergen of interest.
  • the Pharmaceutical Composition is for use in the treatment and / or prophylaxis of diseases related to immunological disorders, in which the diseases related to immunological disorders are: chronic or acute inflammatory diseases, allergic and / or autoimmune diseases.
  • the present application also describes the use of the synthetic peptides or pharmaceutical compositions described herein, for the preparation of a medicament or immunogenic composition for use in the treatment and / or prophylaxis of diseases related to immune disorder.

Abstract

La présente invention concerne la sélection et la caractérisation de peptides synthétiques possédant une affinité pour les récepteurs présents dans les cellules du système immunitaire, et notamment une affinité pour le récepteur de TGF-β, ainsi que des compositions pharmaceutiques et l'utilisation de ces peptides pour préparer des médicaments ou des compositions immunogènes. Ces peptides synthétiques peuvent se lier aux récepteurs cellulaires et promouvoir un profil de régulation important pour le traitement et/ou la prophylaxie de maladies caractérisées par un trouble immunologique. Plus particulièrement, les maladies associées au trouble immunologique sont : les maladies inflammatoires chroniques ou aiguës, allergiques et/ou auto-immunes.
PCT/BR2020/050117 2019-04-05 2020-04-06 PEPTIDES SYNTHÉTIQUES À AFFINITÉ POUR LE RÉCEPTEUR DE TGF-β, COMPOSITION PHARMACEUTIQUE ET UTILISATION DE CEUX-CI COMME IMMUNOMODULATEURS DANS LE TRAITEMENT DE MALADIES AUTO-IMMUNES, INFLAMMATOIRES OU ALLERGIQUES WO2020198832A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
BRBR1020190070455 2019-04-05
BR102019007045A BR102019007045A8 (pt) 2019-04-05 2019-04-05 Peptídeos sintéticos com afinidade ao receptor de tgf-beta, composição farmacêutica e uso dos mesmos como imunomuduladores no tratamento de doenças autoimunes, inflamatórias ou alérgicas

Publications (1)

Publication Number Publication Date
WO2020198832A1 true WO2020198832A1 (fr) 2020-10-08

Family

ID=72664368

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/BR2020/050117 WO2020198832A1 (fr) 2019-04-05 2020-04-06 PEPTIDES SYNTHÉTIQUES À AFFINITÉ POUR LE RÉCEPTEUR DE TGF-β, COMPOSITION PHARMACEUTIQUE ET UTILISATION DE CEUX-CI COMME IMMUNOMODULATEURS DANS LE TRAITEMENT DE MALADIES AUTO-IMMUNES, INFLAMMATOIRES OU ALLERGIQUES

Country Status (2)

Country Link
BR (1) BR102019007045A8 (fr)
WO (1) WO2020198832A1 (fr)

Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012167143A1 (fr) * 2011-06-03 2012-12-06 Xoma Technology Ltd. Anticorps spécifiques du tgf bêta
BR102015003903A2 (pt) * 2015-02-23 2016-11-22 Univ Fed De Uberlândia peptídeos anti-inflamatórios e imuno moduladores

Patent Citations (2)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO2012167143A1 (fr) * 2011-06-03 2012-12-06 Xoma Technology Ltd. Anticorps spécifiques du tgf bêta
BR102015003903A2 (pt) * 2015-02-23 2016-11-22 Univ Fed De Uberlândia peptídeos anti-inflamatórios e imuno moduladores

Non-Patent Citations (4)

* Cited by examiner, † Cited by third party
Title
EMÍLIA R VAZ; FUJIMURA PATRÍCIA T; ARAUJO GALBER R; DA SILVA CARLOS A T; SILVA RANGEL L; CUNHA THIAGO M; LOPES-FERREIRA MÔNICA; LI: "A short peptide that mimics the binding domain of TGF-betaI presents potent anti-inflammatory activity", PIOS ONE, vol. 10, no. 8, 27 August 2015 (2015-08-27), pages e0136116, XP055746234 *
LIESBETH CAROLIEN VAN NIEUWENHOVE, ROGÉ STIJN, BALHARBI FATIMA, DIELTJENS TESSA, LAURENT THIERRY, GUISEZ YVES, BÜSCHER PHILIPPE, L: "Identification of peptide mimotopes of Trypanosoma brucei gambiense variant surface glycoproteins", PLOS NEGLECTED TROPICAL DISEASES, vol. 5, no. 6, 14 June 2011 (2011-06-14), pages e1189, XP055747370 *
LINGYIN LI; BRENDAN P ORNER; TAO HUANG; ANDREW P HINCK; LAURA L KIESSLING: "Peptide ligands that use a novel binding site to target both TGF-beta receptors", MOL BIOSYST., vol. 6, no. 12, 2010, pages 2392 - 402, XP055102599, DOI: 10.1039/c0mb00115e *
ZONG, X. L. ET AL.: "Transforming growth factor-betaiota with tonos phage model peptides isolated from a phage display 7-mer peptide library can inhibit the activity of keloid fibroblasts", CHIN MED J., vol. 124, no. 3, February 2011 (2011-02-01), pages 429 - 435 *

Also Published As

Publication number Publication date
BR102019007045A8 (pt) 2023-05-09
BR102019007045A2 (pt) 2021-12-28

Similar Documents

Publication Publication Date Title
Fornasa et al. Dichotomy of short and long thymic stromal lymphopoietin isoforms in inflammatory disorders of the bowel and skin
Mirabella et al. Prenatal interleukin 6 elevation increases glutamatergic synapse density and disrupts hippocampal connectivity in offspring
Xu et al. Transplanted L1 expressing radial glia and astrocytes enhance recovery after spinal cord injury
Nishida et al. Claudin-2 regulates colorectal inflammation via myosin light chain kinase-dependent signaling
Bergquist Schistosomiasis vaccine development: approaches and prospects
Brandner et al. Hereditary barrier-related diseases involving the tight junction: lessons from skin and intestine
Zhang et al. Protein tyrosine phosphatase 1B deficiency ameliorates murine experimental colitis via the expansion of myeloid-derived suppressor cells
Cohen et al. Newly formed endothelial cells regulate myeloid cell activity following spinal cord injury via expression of CD200 ligand
Nie et al. Phenotypic switch in lung interstitial macrophage polarization in an ovalbumin‑induced mouse model of asthma
Liu et al. Functional macrophages and gastrointestinal disorders
Anders et al. Proteomic profiling reveals crucial retinal protein alterations in the early phase of an experimental glaucoma model
Cui et al. From birth to death: The hardworking life of Paneth cell in the small intestine
EP2988775B1 (fr) Modulation immune pour le traitement d'une maladie auto-immune
WO2020198832A1 (fr) PEPTIDES SYNTHÉTIQUES À AFFINITÉ POUR LE RÉCEPTEUR DE TGF-β, COMPOSITION PHARMACEUTIQUE ET UTILISATION DE CEUX-CI COMME IMMUNOMODULATEURS DANS LE TRAITEMENT DE MALADIES AUTO-IMMUNES, INFLAMMATOIRES OU ALLERGIQUES
WO2020198833A1 (fr) COMBINAISON DE PEPTIDES SYNTHÉTIQUES À AFFINITÉ POUR LE RÉCEPTEUR DE TGF-β ET À AFFINITÉ POUR LE RÉCEPTEUR DE L'IL-10, COMPOSITION PHARMACEUTIQUE ET UTILISATION DE CEUX-CI COMME IMMUNOMODULATEURS DANS LE TRAITEMENT DE MALADIES AUTO-IMMUNES, INFLAMMATOIRES OU ALLERGIQUES
CN109468280B (zh) 过表达circScmh1基因的神经干细胞外泌体及其制备方法和应用
Mazloomi et al. Evaluation of the efficacy of nicotine in treatment of allergic asthma in BALB/c mice
Geng et al. Induced regulatory T cells superimpose their suppressive capacity with effector T cells in lymph nodes via antigen-specific S1p1-dependent egress blockage
Yang et al. Lentinan inhibited the activation of Th2 cells in allergic mice by reducing the amplitude of changes in biological rhythm
Assayag et al. The effect of omalizumab treatment on the low affinity immunoglobulin E receptor (CD23/fc epsilon RII) in patients with severe allergic asthma.
KR20180072579A (ko) 변이된 타우 단백질 단편 및 이의 용도
CN102319431B (zh) 用于检测和治疗老年性痴呆的单克隆抗体
JP2002507573A (ja) トロポミオシンアイソフォームとトロポミオシンアイソフォームに対するモノクローナル抗体による潰瘍性結腸炎の治療
Tworek et al. Human hemopoietic progenitor cell toll-like and thymic stromal lymphopoietin receptor expression and function in allergic asthmatic subjects
Sokol et al. CCR8 Mediated Cell Migration Controls Th2 Differentiation

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 20782600

Country of ref document: EP

Kind code of ref document: A1

NENP Non-entry into the national phase

Ref country code: DE

122 Ep: pct application non-entry in european phase

Ref document number: 20782600

Country of ref document: EP

Kind code of ref document: A1