MXPA05002577A

MXPA05002577A - Use of histamine h4 receptor modulators for the treatment of allergy and asthma.

Info

Publication number: MXPA05002577A
Application number: MXPA05002577A
Authority: MX
Inventors: Robin L Thurmond
Original assignee: Johnson & Johnson
Priority date: 2002-09-06
Filing date: 2003-09-05
Publication date: 2005-09-20
Also published as: EP1545596A4; EP1545596A2; AU2003265961A1; CA2497788A1; JP2006510590A; WO2004021999A3; WO2004021999A2

Abstract

Methods are disclosed for identifying histamine receptor modulators that affect mast cell or basophil chemotaxis, and the use of such histamine H4 receptor modulators for the prevention, treatment, induction, or other desired modulation of asthma and/or allergic responses, or diseases and/or conditions that are modulated, affected or caused by asthma or allergic responses. Also disclosed is the use of histamine H4 receptor modulators for the prevention, treatment, induction, or other desired modulation of mast cell or basophil chemotactic responses, such as migration to a particular site, or diseases and/or conditions that are modulated, affected or caused by mast cell or basophil chemotaxis.

Description

USE OF HISTAMINE H4 RECEPTOR MODULATORS FOR THE TREATMENT OF ALLERGY AND ASTHMA RECIPROCAL REFERENCE TO RELATED REQUESTS This application claims the benefit of the provisional application of E.U.A. No. 60 / 408,736, filed September 6, 2002, which is hereby incorporated by reference in its entirety. Related applications include patent application of E.U.A. No. 10 / 094,357, filed March 8, 2002, provisional application of E.U.A. No. 60 / 408,569, filed on September 6, 2002, and provisional application of E.U.A. No. 60 / 408,579, also filed September 6, 2002, the descriptions of which are hereby incorporated by reference in their entirety.

FIELD OF THE INVENTION The present invention relates to the use of modulators of the histamine H4 receptor for the prevention, treatment, induction or other desired modulation of allergic responses, asthma or diseases and / or conditions that are modulated, affected or caused by asthma or allergic responses.

BACKGROUND OF THE INVENTION Histamine is a multifunctional chemical transmitter that signals through cell surface receptors that bind to intracellular pathways through guanine nucleotide binding proteins. This class of histamine-binding cell surface receptors is part of a broad family of receptors called G-protein coupled receptors or GPCRs. There are currently four subtypes of histamine receptors that have been defined pharmacologically and have been divided into the classifications H1, H2, H3 and H4 (Hill et al., Pharmacol. Rev. (1997) 49 (3): 253-278; , Mol, Pharmacol. (2001) 59: 415-419). The histamine H1 receptor has been cloned (Yamashita et al., Proc. Nati, Acad. Sci. USA (1991) 88 (24): 1 1515-11519), and is the target of drugs such as diphenhydramine, which blocks the effects of histamine on smooth muscle in allergic responses. The H2 receptor of histamine has been cloned (Gantz et al., Proc. Nati, Acad. Sel. USA (1991) 88 (2): 429-433), and is the target of drugs such as ranitidine that blocks the effects of histamine on the secretion of acid in the stomach. The histamine H3 receptor, which was hypothesized to exist in 1983 (Arrang et al., Nature (London) (1983) 302 (5911): 832-837), has been cloned (Lovenberg et al., Mol. (1999) 55: 1101- 107), and is currently a target for the development of drugs for the central nervous system. There are numerous additional functions of histamine in humans that can be mediated by histamine receptors of unknown class, for example, istamine is known to play a role in asthma; however, the current antihistamines which direct the H1 and H2 histamine receptors have little, if any, utility in the treatment of asthma (Larsen et al., Pharmacother. (2001) 21: 28S-33S).

BRIEF DESCRIPTION OF THE INVENTION The present invention relates to the use of modulators of the histamine H4 receptor for the treatment and / or prevention of asthma and / or allergic responses, and diseases and conditions mediated by asthma and / or allergic responses. Modulators of the histamine H4 receptor can be used to modulate allergic responses in mammals, including the induction, as well as the inhibition, of allergic responses, depending on whether the histamine H4 receptor modulator is an agonist, inverse agonist or activity antagonist. of the H4 receiver. Asthma and allergic responses mediated by leukocytes, basophils, eosinophils or mast cells are inhibited by treatment with histamine H4 receptor antagonists or inhibitors. The invention provides in one aspect methods for identifying compounds that modulate the activity of the mammalian histamine H4 receptor, which comprise: combining a putative modulator compound of mammalian histamine H4 receptor activity, with mammalian histamine H4 receptor and a ligand known for the histamine H4 receptor; and measuring an effect of the modulator on the function of the H4 receptor protein, or its ability to bind the ligand, wherein the effect is inhibition, activation, antagonist, agonist or inverse agonist activity, wherein said modulator compound is a modulator of chemotaxis of mast cells. Monospecific antibodies immunologically reactive with a mammalian histamine H4 receptor protein are also provided, wherein said antibody modulates chemotaxis of mast cells. In another aspect, the invention provides a method for identifying compounds that modulate the protein activity of the mammalian histamine H4 receptor, which comprises: combining a putative modulator compound of mammalian histamine H4 receptor protein activity, with receptor protein H4 of mammalian histamine and a known ligand of the histamine H4 receptor; and measuring an effect of the modulator on protein function or its ability to bind the ligand, wherein said effect is inhibition, activation, antagonist, agonist or inverse agonist activity, wherein said modulator compound is a chemotaxis modulator of basophils. Monospecific antibodies immunologically reactive with a mammalian histamine H4 receptor protein are also provided, wherein said antibody modulates basophil chemotaxis in vitro or in vivo. In another of its several aspects, the invention provides a method for identifying compounds that modulate the mast cell chemotaxis mediated by the mammalian histamine H4 receptor to histamine, which comprises: in the presence or absence of a modulator of the histamine H4 receptor , put mast cells in proximity to histamine under conditions that allow the mast cells to move towards histamine; and measuring an effect of the modulator of the histamine H4 receptor on the movement of the mast cells towards histamine, where an increase or decrease in the speed of movement of the mast cells towards histamine, or in the number of mast cells that are move towards histamine, is indicative that the test compound modulates chemotaxis mediated by the histamine H4 receptor of mast cells toward histamine. In another aspect, the invention provides methods for determining whether a modulator of the histamine H4 receptor modulates the subepithelial accumulation of mast cells in the trachea of a mammal in response to exposure to histamine or an allergen, the method comprising: in the presence or absence of pretreatment with a modulator of the histamine H4 receptor, exposing a mammal to an aerosol comprising histamine or an allergen under a regimen that would result, in the absence of the modulator, in a predetermined amount of subepithelial accumulation of mast cells in the trachea of the mammal; and comparing the amount of subepithelial accumulation of mast cells in the trachea of the mammal in the presence and absence of the histamine H4 receptor modulator, a change in the accumulation of mast cells in the presence of the modulator compared to the absence of the modulator, being indicative of that the modulator of the histamine H4 receptor modulates the subepithelial accumulation of mast cells in the trachea of the mammal in response to exposure to histamine or an allergen. Other features and advantages of the present invention will be understood by reference to the figures, detailed description and following examples: BRIEF DESCRIPTION OF THE DRAWINGS Figure 1: Chemotaxis of mast cells derived from mouse bone marrow in response to histamine. Figure 2: Chemotaxis of mast cells derived from mouse bone marrow in response to histamine can be blocked by a specific histamine H4 receptor antagonist, but not by H1, H2 or H3 receptor antagonists. Figure 3: Chemotaxis of mast cells derived from mouse bone marrow in response to histamine, can be blocked by a specific histamine H4 receptor antagonist, with an IC50 of 38 nM. Figure 4: the accumulation of mast cells induced by histamine in vivo in the trachea of the mouse, can be blocked by a specific antagonist of the histamine H4 receptor. Figures 5A and 5B: differential cell count in bronchoalveolar lavage fluid (BAL) from the model of ovalbumin-induced pulmonary inflammation in mice treated with the H 4 receptor antagonist (5-chloro-1 H-benzoimidazole-2) il) - (4-methyl-piperazin-1-yl) -methanone. The p-values of a t unpaired Student's test are given as follows: * represents p < 0.05, ** represents p < 0.01, and *** represents p < 0.001. Figure 5A: The symbols used for the treatments are the following: PBS control 1SI; OVA (ovalbumin) control, GZl; OVA + vehicle, ES; OVA + H4 antagonist at 50 mg / kg, [22; OVA + H4 antagonist at 20 mg / kg, | = j; OVA + H4 antagonist at 5 mg / kg, QH]. Figure 5B: The symbols used for the treatments are the following: PBS control, W, OVA (ovalbumin) control, GZ!; OVA + vehicle, IS3; OVA + H4 antagonist at 5 mg / kg, E22; OVA + H4 antagonist at 2 mg / kg, | =!; OVA + H4 antagonist at 0.5 mg / kg, OH]. Figure 6: differential cell count in bronchoalveolar lavage fluid from the model of ovalbumin-induced pulmonary inflammation in mice treated with the H4 receptor antagonist (5-cioro-1 H-indol-2-yl) - (4-methyl) -piperazin-1-yl) -metanone. The p-values of a t unpaired Student's test are given as follows: * represents p < 0.05, ** represents p < 0.0, and *** represents p < 0.001. The symbols used for the treatments are the following: PBS control, - S -; OVA (ovalbumin) control, -? -; OVA + vehicle,; OVA + H4 antagonist at 5 mg / kg, - tf-; OVA + H4 antagonist at 20 mg / kg, - ^ -; OVA + H4 antagonist at 50 mg / kg. Figure 7: Effects of the H4 receptor antagonist (5-chloro-1 H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone on airway hyperreactivity in the model of induced lung inflammation by ovalbumin in mice (n = 8). The p-values of a t unpaired Student's test are given as follows: * represents p < 0.05, ** represents p < 0.01, and *** represents p < 0.001. The symbols used for the treatments are the following: PBS control, 0; OVA control, ES; OVA + H4 antagonist at 20 mg / kg, EZl; OVA + H4 antagonist at 60 mg / kg, j = |; OVA + H4 antagonist at 100 mg / kg, ED.

DETAILED DESCRIPTION OF THE INVENTION AND MODALITIES PREFERRED DNA molecules that encode a mammalian histamine H4 receptor have been cloned and characterized, and represent members of the class of receptors that are coupled to G proteins (Liu et al., (2001) Mol.Pharmacol. (2001) 59 : 420-426; Liu et al., J. Pharmacol. Exp. Therapeut. (2001) 299 (1): 121-130). Through the use of a recombinant expression system, functional DNA molecules encoding these H4 histamine receptors have been isolated from mouse, rat, guinea pig and human. The recombinant protein is useful for a variety of purposes including, but not limited to, identification of human histamine H4 receptor modulators. The H4 histamine receptors of mouse, rat and guinea pig have a variety of uses including, but not limited to, resolving pharmacological differences observed between different mammalian species, particularly since guinea pig, rat and murine species are commonly used in the preclinical evaluation of new chemical entities that function as modulators. Said modulators may include, for example, agonists, antagonists and inverse agonists. The modulators identified in the tests described herein are useful, for example, as therapeutic agents, prophylactic agents and diagnostic agents. Indications for such therapeutic agents include, but are not limited to, asthma, allergy, inflammation, cardiovascular and cerebrovascular disorders, non-insulin dependent diabetes mellitus, hyperglycemia, constipation, arrhythmia, neuroendocrine system disorders, stress and spasticity, as well as acid secretion. , ulcers, airway constriction, and prostate dysfunction. In particular embodiments, modulators that down-regulate the expression, activity or accessibility of H4 receptors are used as therapeutic agents for the treatment of allergic rhinitis and / or asthma. The term "human histamine H4 receptor", as used herein, refers to protein of subclass H4 that can function as a specific receptor for histamine. Allergy and asthma are two of the most common respiratory problems. Allergy is typically characterized by sneezing, runny nose, watery or irritated eyes, and nasal congestion. The most severe allergy may be accompanied by other symptoms of increasing significance. In most cases, the allergy is triggered by exposure to an environmental allergen, for example, inhalation of airborne allergens such as pollen, dust mites, mold spores or animal dander. Other allergens are ingested with food or drinks. Like allergy, asthma can be caused by inhaled allergens, but it can be caused by non-specific irritants, and other factors such as exercise. It differs from allergy because it is characterized by airway inflammation, bronchial hyperresponsiveness and airway obstruction. Asthma symptoms typically include panting, coughing, chest tension and respiratory failure. IgE and mast cells play key roles in allergy and asthma. Of course, increases in the number of mast cells are found in chronic allergic rhinitis and allergy, as well as after exposure to antigens (Crimi et al., Am. Rev. Resp. Dis. (1991) 144: 1282; Kirby et al., Am. Rev. Resp. Dis. (1987) 136: 379; Slater et al., J. Laryn. Otol. (1996) 10: 929; Gauvreau et al., Am. J. Resp. Crit. Care Med. (2000) 161: 1473; Amin et al., Am. J. Resp. Crit. Care Med. (2000) 162: 2295; and Kassel et al., Clin. Exp. Allegy (2001) 31: 1432). A wide variety of stimuli can cause the activation of mast cells, and then cause them to migrate to a particular site (recruitment) and / or suffer degranulation. . These stimuli may be of an immunological nature (such as antigens or allergens) or non-immunological (such as chemical agents). Activated mast cells release many inflammatory mediators, such as histamine, which are involved in acute phase responses, including increased vascular permeability, bronchoconstriction, vasodilation, and inflammatory cell recruitment. The flow of inflammatory cells in turn leads to the release of other mediators that facilitate and prolong the response. It is this chronic inflammatory process, which leads to tissue remodeling associated with both conditions. Recently, the histamine H4 receptor has been cloned, and has been shown to be expressed in a variety of cells including, but not limited to, mast cells. Another important cell type is the basophils that, in contrast to the mast cells that normally reside in tissues that communicate directly with the environment, are circulating in the blood. In response to inflammatory stimuli, basophils migrate to the site of inflammation. Like mast cells, they respond to IgE or other agents, and release inflammatory mediators such as histamine. In allergic rhinitis and allergy, there is an increase in the number of basophils in the respiratory tract, where it is thought to play a role in late phase responses (Kirby et al., (1987), cited above, Gauvreau et al. (2000). ), cited above, and Braunstahl et al., Am. J. Resp. Crit. Care Med. (2001) 164: 858). In general, allergy can be controlled, for example, by antihistamines (all H1 receptor antagonists) and decongestants; however, there is still a need for agents that modify the disease. The same is applicable for asthma, where inhaled bronchodilators and steroids are the main therapies. Of note is the fact that, although it is thought that histamine and mast cells are important for both conditions, the currently available H1 and H2 receptor antagonists are only useful in the treatment of allergy and have little benefit, if any. , in asthma. This suggests that, for asthma, other histamine receptors, such as the H4 receptor, play a role. Numerous medical texts are published, and are available to experts in the relevant technical fields. In addition, numerous medical and scientific research publications have been published in the fields of allergy and asthma. Examples of available published texts on the subject of inflammation include Barnes et al., Asthma and COPD: Basic Mechanisms and Clinical Management, (Academic Press, London, 2002); Eric et al., Bronchial Asthma: Principles of Diagnosis and Treatment, (Humana Press, 2001); Mygind, Allergic and Non-Allergic Rhinitis: Clinical Aspects, (W. B. Saunders Company, 1993); Grammer and Greenberger, Patíerson's Allergic Diseases, (Lippincott Williams &Wilkins Publishers, 2002); Cecil et al., Textbook Of Medicine, 18th ed. (W. B. Saunders Company, 1988); and Steadman's Medical Dictionary. The present invention demonstrates that the histamine H4 receptor is involved in asthma and allergic responses, and is particularly involved in the recruitment of mast cells or basophils to the stimulation site, and that antagonists for this receptor are effective anti-asthmatic and / or antiallergic agents. The present invention provides, in certain currently preferred embodiments, methods for modulating asthma or allergic responses, which are mediated directly or indirectly by the histamine H 4 receptor. In one of its aspects, the present invention also provides methods for inhibiting, preventing, ameliorating, inducing or otherwise affecting asthma or allergic responses that are mediated by the histamine H4 receptor, by treating a mammal with modulators of the H4 receptor. of histamine. Modulators of the histamine H4 receptor that are useful in the method of the present invention include, but are not limited to, antibodies and antibody fragments that bind to the histamine H4 receptor, RNAi, antisense agents or other agents that modulate the expression of genes encoding the histamine H4 receptor, and inhibitors, activators, antagonists, agonists and inverse agonists of the histamine H4 receptor including, but not limited to, proteins, nucleic acids or other organic molecules. These modulators are useful for administration to humans in need thereof, and are also useful for veterinary purposes for administration to non-human animals including, but not limited to, non-human mammals. Monospecific antibodies to the mammalian histamine H4 receptor are purified from mammalian antisera containing antibodies reactive against the mammalian histamine H4 receptor, or are prepared as monoclonal antibodies reactive with the mammalian histamine H4 receptor, using the G technique. Kohler and C. Milstein (Nature (1975) 256: 495-497). A monospecific antibody, as used herein, is defined as a single antibody species or multiple antibody species with homogeneous binding characteristics for the mammalian histamine H4 receptor. Homogeneous binding, as used herein, refers to the ability of the antibody species to bind to a specific antigen or epitope, such as those associated with the mammalian histamine H4 receptor, as described above. Methods for preparing monoclonal and polyclonal monospecific antibodies are generally known in the art. It is also readily apparent to those skilled in the art, that well known methods can be used to produce monospecific antibodies that produce antibodies specific for polypeptide fragments of the mammalian histamine H4 receptor, or nascent full length polypeptides of the mammalian histamine H4 receptor, or the individual epitopes of the histamine H4 receptor mammal. Specifically, it is readily apparent to those skilled in the art, that monospecific antibodies can be generated that are specific for a portion of only one species of the mammalian histamine H4 receptor or the fully functional histamine H4 receptor. It is also readily apparent to those skilled in the art, that antibodies that are specific for the histamine H4 receptor can cause a change in the functional activity of the receptor including, but not limited to, causing the receptor to be activated or inactivated, blocked from binding to its ligand, blocked from releasing its bound ligand, or preventing it from functioning in the normal manner associated with a histamine H4 receptor. Nucleotide sequences that are complementary to the DNA sequence encoding the histamine H4 receptor of human or other mammal can be synthesized for antisense therapy. These antisense molecules may be DNA, stable derivatives of DNA, such as phosphorothioates or methylphosphonates, RNA, stable derivatives of RNA, such as 2'-0-alkylNAR, or other antisense oligonucleotide mimetics of the human histamine H4 receptor. The antisense molecules of the human histamine H4 receptor can be introduced into the cells by microinjection, encapsulation in liposomes, or by expression from vectors harboring the antisense sequence. Human histamine H4 receptor antisense therapy may be particularly useful for the treatment of diseases where it is beneficial to reduce the activity of the human histamine H4 receptor. Histamine H4 receptor gene therapy from human or other mammal can be used to introduce the histamine H4 receptor into the cells of target organisms. The histamine H4 receptor gene can be ligated into viral vectors that mediate the transfer of human histamine H4 receptor DNA, by infection of receptor host cells. Suitable viral vectors include retroviruses, adenoviruses, adeno-associated viruses, herpes viruses, vaccinia viruses, polioviruses, and the like. Alternatively, human histamine H4 receptor DNA can be transferred into cells for gene therapy by non-viral techniques, including receptor-directed DNA transfer using the DNA-ligand conjugates or adenovirus-ligand-DNA conjugates, fusion of membranes by lipofection, or direct microinjection. These methods and variations thereof are suitable for human histamine H4 receptor gene therapy both ex vivo and live. Human histamine H4 receptor gene therapy may be particularly useful for the treatment of diseases where it is beneficial to elevate the activity of the human histamine H4 receptor. Histamine is a biogenic amine transmitter that works with a certain capacity in almost all physiological and pathophysiological situations. Histamine acts as a neurotransmitter and neuromodulator in the central nervous system, mediates asthma and allergic responses, regulates the function of the respiratory tract, controls the secretion of acid in the stomach, regulates cardiovascular function, as well as arterial and venous responses, and It is likely to intervene in processes yet to be determined. The histamine receptors that mediate these effects are not fully characterized. One way to understand which histamine receptors are involved in these processes is to develop chemical modulators (such as agonists, antagonists and inverse agonists) of the receptors as research tools and therapeutic entities. Recombinant host cells expressing the mammalian histamine H4 receptor can be used to provide materials for a screening method that identifies such agonists and antagonists. As such, this invention provides a way to identify new histamine H4 receptor agonists and antagonists that may prove useful as research tools, or that may be used as a therapeutic to treat disorders that directly or indirectly include histamine receptors, such as allergic responses and asthma. Tests to detect compound modulation or interaction of the histamine H4 receptor include, but are not limited to, tests for direct binding to the ligand, competitive binding (or displacement) tests to the ligand, or functional tests that measure the response of the receptor to the ligand. ligand, for example, by cAMP production or, as in a preferred embodiment, chemotaxis of mast cells or basophils. Although tests of this general type are well known to those skilled in the art, they were not previously possible before obtaining the recombinant molecules described herein, nor were they appreciated to have practical utility prior to the inventors' discovery of the physiological effect of blocking the H4 receptor, for example, inhibition of mast cell chemotaxis, among other effects. An example of a competitive binding test includes the following steps: 1. Cultured mammalian cells are transiently transfected with a nucleic acid molecule that codes for a histamine H4 receptor and then grown in culture. 2. Cell membranes are prepared from the transfected cells by homogenization of the cells and separation of the membrane fraction, for example, by centrifugation. 3. For controls, cell membranes are incubated with detectably labeled histamine (e.g., tritiated histamine) in the presence or absence of more histamine. 4. For test samples, cell membranes are incubated with detectably labeled histamine as indicated above, in the presence of various concentrations of the compounds to be tested. 5. The values of K, are calculated according to known methods. Certain embodiments of the invention provide in vitro tests to measure the effect of a H4 modulator of histamine on the chemotaxis of mast cells or basophils to histamine. For these tests, as applied to mast cells, bone marrow is obtained from an animal source (eg, mice), and cultured for a suitable time in a medium that promotes the differentiation of mast cells. Segmented culture cavities of adequate pore size are lined with fibronectin. After the removal of fibronectin, culture medium containing histamine is added to the lower chambers of the segmented cavities. To the upper cavities are added several concentrations of the compounds to be tested, together with an aliquot of mast cells. The cavities are incubated under conditions that allow the migration of mast cells from the upper chambers to the lower chambers. After incubation, the number of cells in the lower chambers is counted, for example, by flow cytometry. In addition, in another of its several aspects, the invention provides animal models in vivo to evaluate the effect of a histamine H4 receptor modulator on the subepidermal accumulation of mast cells induced by histamine or allergen in the trachea. Test animals (e.g., mice) are exposed to an aerosol comprising saline (control) or histamine for a short period (e.g., 20 minutes) on a few (e.g., two) consecutive days. For test animals, mice previously dosed with histamine in a gas before each spray are dosed with saline or a compound to be tested. After the treatments, the animals are sacrificed and a section of the trachea is removed and sectioned. The mast cells are detected quantitatively by selective staining, for example, immunohistochemically or with toluidine blue. The mast cells can also be quantified as submucosa or subepithelial, depending on their location within the section of the trachea. The migration of mast cells in the subepithelial space is indicative of an allergic response. Preferred methods of the present invention are used to identify chemical compounds that act, for example, as agonists, antagonists or inverse agonists of the histamine H4 receptor. As described in greater detail in example 1, a screening test that includes binding of the H4 receptor to candidate compounds, has made it possible to identify several classes of compounds that bind to the receptor. The binding affinity of these compounds has been positively correlated with the ability of the compounds to inhibit the chemotaxis of mast cells to histamine. An example of a compound in an in vivo model has also been tested, where it has been shown that the compound reduces or inhibits the migration of mast cells mediated by histamine in the subepithelial space of the trachea. This movement is similar to what is thought to occur after exposure to an allergen. Accordingly, in addition to methods for identifying modulators of the histamine H4 receptor, the presently preferred embodiments of the present invention provide modulators of the H4 receptor identified by the various selection tests described herein. These modulators bind to the recombinant H4 receptor in vitro. In a preferred embodiment, they possess a Ki for the receiver, under conditions defined herein, less than 1 μ ?, more preferably less than 900 nM, 800 nM, 700 nM or 600 nM, respectively, in ascending preference order. In more preferred embodiments, they possess a Ki for the receiver less than 500 nM, even more preferably less than 400 nM, still more preferably less than 300 nM, and even more preferably less than 200 nM. In particularly preferred embodiments, the K i is less than 100 nM, 50 nM, 40 nM, 30 nM, 20 nM and 10 nM, respectively, in ascending preference order. In view of the fact that the K i of said compounds has been shown to be positively correlated with the ability of the compound to inhibit histamine-mediated chemotaxis of mast cells in vitro, those skilled in the art will appreciate that compounds with higher binding affinity can be used. for particular advantage as therapeutic agents in the treatment of pathological conditions associated with H4-mediated signal transduction including, but not limited to, allergy and asthma. In accordance with the present invention, it has been demonstrated that various classes of compounds possess the necessary chemotactic and binding modulatory characteristics mentioned above, as described in more detail in the examples below. In one of its several aspects, the present invention is also directed to methods for selecting compounds that modulate the expression of DNA or RNA encoding the mammalian histamine H4 receptor, as well as the protein function of the mammalian histamine H4 receptor in vitro or in vivo. Compounds that modulate these activities can be DNA, RNA, peptides, proteins, or non-proteinaceous organic molecules. The compounds can be modulated by enhancing or attenuating the expression of DNA or RNA encoding the mammalian histamine H4 receptor, or the protein function of the mammalian histamine H4 receptor. Compounds that modulate the expression of DNA or RNA encoding the mammalian histamine H4 receptor or the protein function of the mammalian histamine H4 receptor can be detected by various tests. The tests can be a simple "yes / no" test that determines whether there is a change in the expression of the nucleic acid encoding the receptor, or a change in the function or activity of the receptor protein. The test can be made quantitative, comparing the expression or function of a test sample with the levels of receptor expression or the function of the receptor protein in a standard sample. The modulators identified in this procedure are useful as therapeutic agents, research tools and diagnostic agents. Pharmaceutically useful compositions comprising modulators of mammalian histamine H4 receptor activity, by receptor binding or other mechanisms as described herein, can be formulated according to known methods, such as by mixing a pharmaceutically acceptable carrier. Examples of such vehicles and formulation methods can be found in Remington's Pharmaceutical Sciences. To form a pharmaceutically acceptable composition suitable for effective administration, said compositions will contain an effective amount of the modulator or other biologically active agent. Therapeutic or diagnostic compositions of the invention are administered to an individual in sufficient amounts to treat or diagnose disorders in which the modulation of activity related to the mammalian histamine H4 receptor is indicated. The effective amount may vary according to a variety of factors, such as the condition, weight, sex and age of the individual. Other factors include the mode of administration. The pharmaceutical compositions can be provided to the individual by a variety of routes, such as the subcutaneous, topical, oral, intranasal and intramuscular routes. The term "chemical derivative" describes a molecule that contains other chemical substituents or portions that are not normally a part of the base molecule. Said portions can improve the solubility, half-life, absorption, etc., of the base molecule. Alternatively, the portions may attenuate undesirable side effects of the base molecule, or decrease the toxicity of the base molecule. Examples of such portions are described in a variety of texts, such as Remington's Pharmaceutical Sciences. The compounds identified according to the methods described herein can be used alone at suitable dosages defined by the routine test, to obtain optimal inhibition of the mammalian histamine H4 receptor or its activity, while minimizing any potential toxicity. In addition, co-administration or sequential administration of other agents may be desirable. The present invention, in other of its various aspects, also provides topical, oral, systemic and parenteral pharmaceutical formulations suitable for use in the methods of treatment of the present invention. Compositions containing compounds or modulators identified in accordance with this invention as the active ingredient for use in the modulation of the mammalian histamine H4 receptor can be administered in a wide variety of therapeutic dosage forms in conventional vehicles for administration. For example, the compounds or modulators can be administered in oral dosage forms such as tablets, capsules (each including sustained release and delayed release formulations), pills, powders, granules, elixirs, dyes, solutions, suspensions, syrups and emulsions. , or by injection. Likewise, they can also be administered in intravenous (bolus or infusion), intraperitoneal, subcutaneous, topical with or without occlusion, or intramuscular form, all using forms well known to those skilled in the pharmaceutical arts. An effective but non-toxic amount of the desired compound can be used as a mammalian histamine H4 receptor modulating agent. The daily dosage of the products can be varied over a wide range from 0.01 to 1,000 mg per patient, per day. For oral administration, the compositions are preferably provided in the form of scored or non-scored tablets containing 0.01., 0.05, 0.1, 0.5, 1.0, 2.5, 5.0, 10.0, 15.0, 25.0 and 50.0 milligrams of the active ingredient for the symptomatic adjustment of the dosage to the patient to be treated. An effective amount of the drug is often provided at a dosage level of about 0.0001 mg / kg to about 100 mg / kg of body weight per day. The scale is more particularly around 0.001 mg / kg to 10 mg / kg of body weight per day. The dosages of the modulators of the mammalian histamine H4 receptor are adjusted when combined to achieve the desired effects. On the other hand, the dosages of these various agents can be optimized and combined independently to achieve a synergistic result, where the pathology is reduced more than it would be if any agent were used alone. For delivery to the airway of a patient in need of treatment, the compositions are preferably formulated as an aerosol, which can be delivered nasally or by an inhaler, as is well known in the art. The amounts of active ingredient are adjusted for this form of supply, according to standard methods. Advantageously, the compounds or modulators of the present invention can be administered in a single daily dose, or the total daily dosage can be administered in divided doses of two, three or four times a day. In addition to oral and intranasal / inhalation delivery, the compounds or modulators of the present invention can be administered by transdermal routes, using the forms of transdermal skin patches well known to those skilled in the art. To be administered in the form of a transdermal delivery system, the administration of the dosage will, in fact, continue more than intermittent throughout the dosing regimen. For combination treatment with more than one active agent wherein the active agents are in separate dosage formulations, the active agents may be administered concurrently, or each may be administered at separately staggered times. The dosage regimen using the compounds or modulators of the present invention is selected according to a variety of factors including type, species, age, weight, sex and medical condition of the patient.; the severity of the condition to be treated; the route of administration; the renal and hepatic function of the patient; and the particular compound thereof used. A physician or veterinarian of usual experience, can easily determine and prescribe the effective amount of the drug that is required to prevent, counteract or stop the progress of the condition. The optimal precision to achieve drug concentrations within the scale that gives efficacy without toxicity, requires a regimen based on the kinetics of drug availability towards target sites. This includes a consideration of the distribution, balance and elimination of a drug. In preferred methods of the present invention, an active drug component or ingredient may comprise one or more compounds or modulators described herein, and may be preferably administered in admixture with suitable diluents, excipients or pharmaceutical carriers (collectively referred to herein) as "vehicle" materials) selected suitably with respect to the desired form of administration, ie, oral tablets, capsules, elixirs, syrups, and the like, and consistent with conventional pharmaceutical practices.

For example, for oral administration in the form of tablets or capsules, the active drug components can be combined with an oral, non-toxic and pharmaceutically acceptable inert carrier such as ethanol, glycerol, water, and the like. In addition, when desired or necessary, suitable binders, lubricants, disintegrating agents and coloring agents can also be incorporated into the mixture. Suitable binders include, without limitation, starch, gelatin, natural sugars such as glucose or beta-lactose, corn sweeteners, natural and synthetic gums such as acacia, tragacanth or sodium alginate, carboxymethylcellulose, polyethylene glycol, waxes, and the like. Lubricants used in these dosage forms include, without limitation, sodium oleate, sodium stearate, magnesium stearate, sodium benzoate, sodium acetate, sodium chloride, and the like. Disintegrators include, without limitation, starch, methylcellulose, agar, bentonite, xanthan gum, and the like. For liquid forms, the active drug components can be combined in suitably flavored dispersing or suspending agents, such as natural and synthetic gums, for example, tragacanth, acacia, methylcellulose, and the like. Other dispersing agents that can be used include glycerin, and the like. For parenteral administration, sterile solutions and suspensions are preferred. Isotonic preparations containing generally suitable preservatives are preferred, when intravenous administration is desired. Topical preparations containing an active drug component can be mixed with a variety of carrier materials well known in the art such as, for example, alcohols, Aloe vera gel, allantoin, glycerin, vitamin A and E oils, mineral oil , PPG2 myristyl propionate, and the like, to form, for example, alcoholic solutions, topical cleansers, cleansing creams, skin gels, skin lotions, and shampoos in cream or gel formulations. The compounds or modulators of the present invention can also be administered in the form of liposome delivery systems, such as small unilamellar vesicles, large unilamellar vesicles and multilamellar vesicles. Liposomes can be formed from a variety of phospholipids, such as cholesterol, stearylamine or phosphatidylcholines. The compounds of the present invention can also be delivered by the use of monoclonal antibodies as individual carriers to which the molecules of the compound are coupled. The compounds or modulators of the present invention can also be coupled with soluble polymers such as steerable drug vehicles. Such polymers may include polyvinylpyrrolidone, pyran copolymer, polyhydroxypropylmethacryl-amidophenol, polyhydroxy-ethylaspartamidophenol or polyethylen-eneoxidepolylysine substituted with palmitoyl residue. In addition, the compounds or modulators of the present invention can be coupled to a class of biodegradable polymers useful for achieving controlled release of a drug, for example, polyacetic acid, caprolactone, poly-epsilon, polyhydroxybutyric acid, polyorthoesters, polyacetals, polydihydropyrans, polycyanoacrylates, and flake! interspersed or amphipathic hydrogel block jets. For oral administration, the compounds or modulators may be administered in the form of a capsule, tablet or bolus, or alternatively may be mixed in the animal's food. The capsules, tablets and boluses comprise the active ingredient in combination with a suitable vehicle such as starch, talcum, magnesium stearate or dicalcium phosphate. These unit dosage forms are prepared by intimately mixing the active ingredient with suitable finely powdered inert ingredients including diluents, fillers, disintegrating agents and / or binders, so as to obtain a uniform mixture. An inert ingredient is one that will not react with compounds or modulators, and that is not toxic to the animal that is being treated. Suitable inert ingredients include starch, lactose, talc, magnesium stearate, gums and vegetable oils, and the like. These formulations may contain a widely variable amount of the active and inactive ingredients, depending on numerous factors such as the size and type of the animal species to be treated and the type and severity of the symptoms. The active ingredient can also be administered as an additive to the food, simply by mixing the compound with the forage, or by applying the compound to the surface of the food. Alternatively, the active ingredient may be mixed with an inert carrier, and the resulting composition may then be mixed with the feed, or provided directly to the animal. Suitable inert carriers include corn flour, citrus flour, fermentation residues, soy grains, dry grains, and the like. The active ingredients can be intimately mixed with these inert carriers by grinding, stirring, milling or tumbling, so that the final composition contains from 0.001 to 5% by weight of the active ingredient. The compounds or modulators can alternatively be administered parenterally by injection of a formulation consisting of the active ingredient dissolved in an inert liquid carrier. The injection can be intramuscular, intraruminal, intratracheal or subcutaneous. The injectable formulation consists of the active ingredient mixed with a suitable inert liquid vehicle. Acceptable liquid carriers include vegetable oils, such as peanut oil, cottonseed oil, sesame oil, and the like, as well as organic solvents such as solketal, glycerol formaldehyde, and the like. As an alternative, aqueous parenteral formulations can also be used. Vegetable oils are the preferred liquid vehicles. The formulations are prepared by dissolving or suspending the active ingredient in the liquid carrier, so that the final formulation contains from 0.005 to 10% by weight of the active ingredient. Topical application of the compounds or modulators is possible by the use of a liquid wetting or a shampoo containing the present compounds or modulators as a suspension or aqueous solution. These formulations generally contain a suspension agent ta! as bentonite, and will normally also contain an antifoaming agent. Formulations containing 0.005 to 10% by weight of the active ingredient are acceptable. Preferred formulations are those containing from 0.01 to 5% by weight of the present compounds or modulators. The following examples are provided for the purpose of illustrating the present invention without, however, limiting the same to them.

EXAMPLE 1 Cloning of human histamine H4 receptor cDNA into a mammalian expression vector Human histamine H4 receptor (cited collectively as pH4R) cDNA molecules were cloned into the mammalian expression vector pCIneo. The human histamine H4 receptor cDNA clone was isolated from the human thalamus cDNA library. The full-length cDNA was used as a template for PCR using specific primers with EcoR1 and Not1 sites for cloning. The PCR product was purified on a column (DNA purification equipment by PCR Wizard from Promega), and digested with Not1 and EcoR1 (NEB) to create cohesive ends. The product was purified by low melting point agarose gel electrophoresis. The pCIneo vector was digested with EcoR1 and Not1 enzymes, and subsequently purified on a low melting point agarose gel. The linear vector was used to ligate to cDNA inserts of human histamine H4 receptor. The recombinants were isolated, designated as human histamine H4 receptor, and used to transfect mammalian cells (SK-N-MC cells) by precipitation of DNA with CaP04. Stable cell clones were selected by growth in the presence of G418. Individual clones resistant to G418 were isolated, and were shown to contain the intact human histamine H4 receptor gene. Clones containing human histamine H4 receptor cDNA molecules were analyzed for the expression of pH4R by measuring the inhibition of adenylate cyclase in response to histamine, according to the method of Konig et al. . { Mol. Cell. Neurosci. (1991) 2 (4): 331-337), or by directly measuring cAMP accumulation by radioimmunoassay using Flashplates (NEN). The expression was also analyzed using [3H] -histamine binding assays (Clark et al., Eur. J. Pharmacol. (1992) 210 (1): 31-35). Recombinant plasmids containing DNA encoding human histamine H4 receptor were used to transform mammalian COS7 or CHO cells or HEK293 or L cells or SK-N-MC cells. Cells expressing the human histamine H4 receptor, stably or transiently, were used to test the expression of the human histamine H4 receptor, and for [3H] -histamine binding activity. These cells were used to identify and examine other compounds for their ability to modulate, inhibit or activate the human histamine H4 receptor, and to compete for binding to radioactive histamine. Cassettes containing the human histamine H4 receptor cDNA in the positive orientation with respect to the promoter were ligated into suitable 3 'restriction sites of the promoter, and identified by sequencing and / or restriction site mapping. These cDNA expression vectors were introduced into fibroblast host cells, for example, COS-7 (ATCC, # CRL1651) and CV-1 tat (Sieckevitz et al., Science (1987) 238: 1575-1578], 293, L (ATCC, # CRL6362), SK-N-C (ATCC # HTB-10), by standard methods including, but not limited to, electroporation or chemical procedures (cationic liposomes, DEAE dextran, calcium phosphate). of cultured cells and transfected cells were harvested and analyzed for expression of the human histamine H4 receptor as described herein.All vectors used for transient expression in mammals can be used to establish stable cell lines expressing the receptor Human histamine H4 It is expected that unchanged human histamine H4 receptor cDNA constructs cloned into expression vectors, program host cells to obtain human histamine H4 receptor proteins. host cells for transfection include, but are not limited to, CV-1-P [Sieckevitz et al., cited above], tk-L [Wigler et al., Cell (1977) 11 (1): 223-232] , NS / 0 and dHFr-CHO [Randall J. Kaufman and Phillip A. Sharp, J. Mol. B / o /. (1982) 159: 601-621]. The cotransfection of any vector containing human histamine H4 receptor cDNA with a drug selection plasmid including, but not limited to, G418, aminoglycoside phosphotransferase; hygromycin, hygromycin-B phosphotransferase; APRT, xanthine-guanine phosphoribosyl-transferase, will allow the selection of stably transfected clones. Human histamine H 4 receptor levels are quantified by the tests described herein. Human histamine H4 receptor cDNA constructs were also ligated into vectors containing amplifiable drug resistance markers, for the production of mammalian cell clones that synthesize the highest possible levels of the human histamine H4 receptor. After the introduction of these constructs into cells, clones containing the plasmid were selected with the appropriate agent, and isolation of a clone overexpressing with a high number of plasmid copies was achieved, by selection in increasing doses of the agent. Expression of the recombinant human histamine H4 receptor was achieved by transfecting full-length human histamine H4 receptor cDNA into a mammalian host cell.

Characterization of Human Histamine H4 Receptor Human SK-N-MC cells were transfected with pH4R, and selected in the presence of neomycin for ten days. Individual colonies were collected, and developed in six-well plates. The cells were then placed in 96-well plates, and developed to confluence. The cells were incubated for 20 minutes with isobutylmethylxanthine (1 mM). The cells were stimulated with histamine (100 pM - 100 μ?) For 5 minutes. The cells were then stimulated with forskolin (3 μ?), And were left incubating at 37 ° C for 20 minutes. The cells were treated with HCI at 0.1 N. The cells were frozen and thawed. Aliquots of the supernatant were analyzed for their cyclic AMP content, using standard cAMP radioimmunoassay equipment (Flashplates, NEN). Treatment with forskolin raises the intracellular concentration of cAMP. It was considered that any cell that responds to histamine by decreasing the cAMP content in response to forskolin, expresses the active functional histamine H4 receptor of human. The recombinant human histamine H4 receptor expressed from the DNA molecule encoding the human histamine H4 receptor described herein was shown to be activated specifically by histamine.

EXAMPLE 2 Binding test at the histamine H4 receptor of recombinant human SK-N-MC cells or COS7 cells were transiently transfected with pH4R, and grown in 150 cm2 tissue culture plates. The cells were washed with saline, scraped with a cell scraper, and harvested by centrifugation (1000 rpm, 5 minutes). The cell membranes were prepared by homogenizing the cell pellet in Tris-HCl at 20 mM with a Polytron tissue homogenizer for 10 seconds at high speed. The homogenate was centrifuged at 1000 rpm for 5 minutes at 4 ° C. The supernatant was then collected and centrifuged at 20,000 x g for 45 minutes at 4 ° C. The final pellet was resuspended in Tris-HCl at 50 mM. The cell membranes were incubated with 3H-histamine (5-70 nM) in the presence or absence of more histamine (10000 nM). Incubation occurred at room temperature for 45 minutes. Membranes were harvested by rapid filtration on GF / C Whatman filters, and washed 4 times with ice-cold 50 mM Tris-HCl. The filters were then dried, mixed with scintillant, and the radioactivity counted. SK-N-MC or COS7 cells expressing the human histamine H4 receptor were used to measure the binding affinity of other compounds, and their ability to displace binding to the 3H-ligand by incubating the mixture described above in the presence of several concentrations of inhibitor or compound to be tested. For studies of competitive binding using 3 H-histamine, the K 2 values were calculated, based on an experimentally determined KD value of 5 nM, and a ligand concentration of 5 nM, according to Cheng and Prusoff. { Biochem. Pharmacol. (1973) 22: 3099-3108): K i = (IC 50) / (1 + [L] / (K D)). The results of the competitive binding studies are set forth in Table 1 for several compounds within three classes of compounds, which can be prepared as described in the patent application of E.U.A. No. 10 / 094,357 (see also international publication No. WO 02/072548), provisional application of E.U.A. No. 60 / 408,569, and provisional application No. 60 / 408,723, the disclosure of which is incorporated herein by reference. A class comprises the main group f1 / p.21 1 H-indole, and is referred to herein as the class of "indoi". Another class comprises one of the following major groups, and is referred to herein as the "bicyclic pyrrole" class: f.2 / p.21 6H-thieno [2,3] pyrrole f.3 / p.21 4H- thieno [3,2-b] pyrrole f.4 / p.21 4tf-furo [3,2-b] pyrrole The third class comprises the major group f.1 / p.22 1H-benzimidazole, and is referred to in present as the "benzoimidazole" class.

TABLE 1 Compound K (nM) (5-Chloro-7-methyl-1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -metanone 1 (2-Chloro-3-methyl-4H-thieno [3,2-b] pyrrol-5-yl) - (4-methyl-1-piperazin-1-yl) -3 methanone (5-Chloro-1 H) -indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 5 (2,3-Dimethyl-4H-thieno [3,2-b] pyrrol-5-yl) - (4-methyl) -piperazin-1-yl) -5 methanone (2,3-Dichloro-4H-thieno [3,2-b] pyrrol-5-yl) - (4-methyl-piperazin-1-yl) - 5.5 methanone (7-Methyl-1 H -indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 6.6 (7-Amino-1 H -indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 7 (5-Bromo-1 H -indole-2-yl) - (4-methyl-piperazin-1-yl) -methanone 8 (5,7-Dichloro-1 H -indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 10 (5-Chloro-1 H -indole-2-yl) -piperazin-1-yl-methanone 10 (2,3-Dichloro-6H-thieno [2,3-b] pyrrol-5-yl) - (4-methyl-piperazin-1-yl) -10 methanone (5-Methyl-1 H-indoI-2- il) - (4-methyl-piperazin-1-yl) -methanone (4,5-Dichloro-1 H -indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 1 (4-Methyl-piperazin-1-yl) - (5-trifluoromethyl-1 H -benzoimidazol-2-yl) -11 methanone (5-Fluoro-1 H -indol-2-yl) - (4-methyl) -piperazin-1-yl) -metanone 18 (5,7-Difluoro-1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 19 (5-Amino-1 H -indol-2-yl) - (4-methyl-p-piperazin-1-yl) -methanone 19 (5-Hydroxy-1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 19 (4-Met.l-piperazin-1-yl) - (3-methyl-4H-thieno [3,2-b] pyrrol-5-yl) -methanone 21 (7-Chloro-1 H-indole-2- il) - (4-methyl-p¡perazin-1-yl) -methanone 23 (5-Chloro-1 H-indol-2-yl) - [4- (2-hydroxy-ethyl) -piperazin-1-yl] -methanone (2-Chloro-6H-thieno [2,3-b] pyrrol-5-yl) - (4-methyl-p¡perazin-1-yl) -methanone 25 (5-Chloro-1 H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 25 (5-Chloro-1 H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 26 (5-Chloro-H-indol-2-yl) - (3,4-dimethyl-piperazin-1-yl) -methanone 27 (5,6-Difluoro-H-benzoamidazol-2-yl) - (4-methyl-piperazin-1-yl) -28 methanone TABLE 1 (CONTINUATION) (5,7-DimetiI-1H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 30 (2-Chloro-3-methyl-4H-t-ene [3,2-b] pyrrol-5-yl) -piperazin-1-yl-methanone; (4-Methyl-1H-benzoimidazol-2-yl) - (4-metii-piperazin-1-yl) -methanone 31 (1 H-Benzoxydazol-2-yl) - (4-methyl-p-piperazin-1-N) -methanone 32 (6-Hydroxy-1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 32.5 (5-Cioro1 H -indole-2-yl) - ((R) -3-methyl-p-piperazin-1-yl) -methanone 34 (5-Chloro-1 H-indol-2-yl) - ((S) -3-methyl-piperazin-1-yl) -methanone 36 (4-Bromo-1 H -indole-2-yl) - (4-methyl-piperazin-1-yl) -methanone 40 (2-Chloro-4H-t-ene [3,2-b] pyrrol-5-yl) - (4-methyl-p-piperazin-1-yl) -methanone 40 (5-Chloro-1 H -indole-2-yl) - (3-methyl-piperazin-1-yl) -metanone 41 (5-Fluoro-1 H-benzoimidazol-2-yl) -piperazin-1-yl-methanone 42 (7-Amino-1 H -indol-2-yl) -piperazin-1-yl-methanone 43 (4-Met.l-piperazin-1-yl) - (5-nitro-1 H -indole-2-yl) -methanone 46 (7-Hydroxy-1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 47 (6-Chloro-5-fIuoro-1H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -53 methanone (7-Bromo-1 H-indol-2-yl) - ( 4-methyl-p-piperazin-1-yl) -metanone 55 (2-Chloro-6H-thieno [2,3-b] pyrrol-5-yl) -piperazin-1-yl-methanone 56 (3-Bromo-4H-thieno [3,2-b] pyrrol-5-yl) - (4-methyl-piperazin-1-yl) -methanone 56 (3-Methyl-4H-t-ene [3,2-b] pyrrol-5-yl) -piperazin-1-yl-methanone 80 (4-Methyl-pperazin-1-yl) - (6H-thieno [2,3-b] pyrrol-5-yl) -methanone 85 (5-Chloro-1 H-benzoimidazol-2-yl) -piperazin-1-yl-methanone 87 (8-Methyl-8-aza-bicyclo [3.2.1] oct-3-yl) -amide of 1H-89-benzoimidazole-2-carboxylic acid (5-Bromo-benzofuran-2-yl) - (4-methyl) -piperazin-1-yl) -metanone 95 (1 H-indol-2-yl) - (3-methyl-piperazin-1-yl) -methanone 100 (1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 117 (6-Chloro-1 H -indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 124 (4-Met.l-piperazin-1-yl) - (4H-thieno [3,2-b] pyrrol-5-yl) -methanone 125 (1 H -indole-2-yl) - (4-meth yl-piperazin-1-yl) -methanethione 132 (4-Methyl-1 H-benzoimidazol-2-yl) -piperazin-1-l-methanone 135 (2,3-Dimethyl-4H-furo [3,2-b] pyrrol-5-yl) - (4-methyl-p¡perazin-1-yl) -140 methanone [5- (3-Methoxy- phenyl) -1 H-indol-2-yl] - (4-methyl-p¡perazin-1-yl) -methanone 145 (4-Methyl-1H-benzolmidazol-2-yl) - (3-methyl-piperazin-1-yl) -methanone 156 (3-Methyl-p-piperazin-1-yl) - (3-methyl-4H-thieno [3,2-b] pyrrol-5-yl) -methanone 161 (2-Chloro-6H-thieno [2,3-b] pyrrol-5-yl) - (hexahydro-pyrrolo [1,2-a] pyrazin-2-yl-yl) -methanone (6-Bromo-1 H- indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 188 (8-Methyl-8-aza-bicyclo [3.2.1] oct-3-yl) -amide of 5-methyl-H-613-benzoimidazole-2-carboxylic acid (3-Bromo-4H-t-ene [3,2 -b] pyrrol-5-yl) - (3-methyl-piperazin-1-yl) -methanone 980 EXAMPLE 3 Ligand binding to mammalian histamine H4 receptors Affinity of 3H-histamine was determined by rat, mouse, guinea pig and human histamine H4 receptors, using standard techniques as described herein. Saturation binding was performed on membranes of stably transfected SK-N-MC cells with the appropriate histamine H4 receptor. The values of Kp were derived from a -1 relation / slope of the linear regression of a Scatchard plot (united / free versus united). The results are shown in table 2: TABLE 2 The relative affinity of several known histamine receptor ligands was determined by competitive binding of 3 H-histamine to 30 nM. The values of Kj were calculated for each ligand, according to the method of Cheng and Pruscoff (K, = IC5o / (1 + [3H-histamine] / Kd) .The KD values for 3H-histamine were the ones exposed in table 2. The results are shown in table 3: TABLE 3 EXAMPLE 4 Inhibition of mast cell chemotaxis induced by histamine in vitro, by histamine H 4 receptor antagonists This example demonstrates for the first time the discovery that histamine H 4 receptor antagonists can block the chemotactic response of mast cells in response to a stimulus.

Methods Culture of marrow cells from bone marrow Marrow cells marrow derived from BALB / c or C57b1 / 6j mice were differentiated. Mice were sacrificed by asphyxia under C02 at 95%, and the femurs were removed. The bone marrow of the femurs was aseptically isolated. Cells (5x105 / mL) were cultured at 37 ° C with 5% C02 in culture medium consisting of R.PMI medium with 10% FCS, 0.1 nM non-essential amino acids, 50 μg / mL penicillin / streptomycin and WEHI-3 medium conditioned at 20%. Conditioned WEHI-3 medium was prepared from WEHI-3 cells (ATCC), which were cultured in Iscoves modified Dulbecco's medium with 10% FCS, 4 mM L-glutamine, 1.5 g / L carbonate sodium, beta-mercaptoethanol at 0.05 μ? and 50 μg / mL of penicillin / streptomycin. The filtered supernatant was used as the conditioned WEHI-3 medium. After 16 hours in culture, the bone marrow cells were transferred to a new flask. The medium was renewed once a week. After four weeks, the cells were tested by flow cytometry for IgE receptor and expression of CD117 (c-kit). The mast cells were incubated with anti-DNP IgE (ICN Pharmaceuticals, Costa Mesa, CA) or vehicle for 30 minutes, followed by FITC-labeled anti-IgE (Pharmingen) or FITC-labeled CD117 (Pharmingen) for 30 minutes on ice. Cultured mast cells consisted of a homogeneous population that were more than 99% positive for the IgE receptor, and more than 99% positive for CD117. Baited cells from four to eight weeks of culture were used for the experiments.

Chemotaxis test Transwells (Costar, Cambridge, MA) of a pore size of 8 μ were coated? with 100 μ? of 100 ng / mL of human fibronectin (Sigma) for 2 hours at room temperature. After removal of fibronectin, 600 μm of RPM medium was added! with 5% BSA in the presence of histamine (varying from 1.25-20 μ) to the lower chamber. To test the various histamine receptor antagonists, 10 μl solutions of the compounds were added to the upper and lower chambers. Baited cells (2x105 / cavity) were added to the upper chamber. The plates were incubated for 3 hours at 37 ° C. The transwells were removed, and the number of cells in the lower chamber was counted for 60 seconds using a flow cytometer.

Results Histamine mediates chemotaxis through the H4 receptor The chemotactic capacity of mast cells towards histamine was investigated using a transwell system. Baited cells were added to the upper chamber, while histamine was added to the lower chamber.

Histamine induced a dose-dependent increase in the mast cells that migrated in the lower chamber (Figure 1). Histamine receptor-specific antagonists were used to select which histamine receptor is responsible for the chemotaxis towards histamine. The specific antagonists for the histamine H1, H2 or H3 receptors did not alter the chemotaxis induced by histamine (figure 2). However, a specific histamine H 4 receptor antagonist inhibited mast cell chemotaxis (Figures 2 and 3) in a dose-dependent manner. The results shown in Table 2 below show a positive correlation between the K i of the compound and its ability to inhibit the chemotaxis of mast cells; that is, in general, the more powerful the compound, the better is the inhibition.

TABLE 4 Compound K%% of (n) 10 μ inhibition inhibition? 1 μ (5-Chloro-7-methyl-1 H -indole-2-yl) - (4-methyl-p-eperazin-1-yl) -1- 97 methanone (7-Amino-1 H -indole-2- L) - (4-methyl-p¡perazin-1-yl) -methanone 7 99 (5-Chloro-1 H -indol-2-yl) -piperazin-1-yl-methanone 10 99 (5,7-Difluoro-1 H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone 19 100 (4-Methyl-piperazin-1-yl) - (3-methyl-4H-thieno [3,2-b] pyrrol-5-yl-) 60 methanone (2-Chloro-6H-thieno [2] , 3-b] pyrrol-5-yl) - (4-methyl-piperazin-1-yl) -25- 106 103 methanone (5-Chloro-1 H -benzoimidazol-2-yl) - (4 -met.l-piperazin-1-yl) -25-97 84 methanone (5,6-difluoro-1 H -benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -28-97 72 methanone ( 2-Chloro-4H-t-ene [3,2-b] pyrrol-5-yl) - (4-methyl-piperazin-1-yl) - 40 92 methanone (5-Chloro-1 H-benzoimidazole- 2-yl) -piperazin-1-yl-methanone 87 101 8 (2-Chloro-6H-thieno [2,3-b] pyrrol-5-yl) -hexahydro-pyrrolo [1, 2- 176 0 0 a] pyrazin-2-yl) -methanone (8-methyl) -8-aza-bicyclo [3.2.1] oct-3-yl) -amide 5-methyI-613 27 66 1 H-benzoimidazole-2-carboxylic acid (3-Bromo-4H-thieno [3,2-b] ] pyrrol-5-yl) - (3-methyl-piperazin-1-yl) -980 0 0 methanone Indol, control > 10k 0 0 Bicyclic pyrrole, control > 10k 0 0 Benzoimidazole, control > 10k 0 0 EXAMPLE 5 Inhibition of mast cell chemotaxis induced by histamine. by histamine H4 receptor antagonists in vivo Groups of ten female Balb / c mice (8-12 weeks) were exposed to an aerosol of control saline or histamine at 0.1 M for 20 minutes on 2 consecutive days. Mice that were dispersed histamine in a gas, were previously dosed 15 minutes before each spray with saline or 20 mg / kg of the modulator of H4 (5-chloro-1 H-indol-2-yl) - ( 4-methyl-piperazin-1-yl) -methanone by the subcutaneous route (5 ml / kg). Four hours after the final administration of the aerosol, the mice were sacrificed using an overdose of pentobarbital, and a section of the trachea was removed and fixed in formalin. Paraffin inclusion and longitudinal sectioning of the tracheas was carried out, and the mast cells were stained with toluidine blue. The mast cells were quantified as submucosa or subepithelial, depending on their location within each section of the trachea. Statistics were performed using the unpaired Student t test. The results (figure 4) show that histamine induces migration of mast cells in the subepithelial space. This movement is similar to what is thought to occur after exposure to allergens. This migration can be blocked by a specific H4 receptor antagonist.

EXAMPLE 6 Inhibition of mast cell chemotaxis by the H 4 receptor antagonist in an animal model of asthma and allergic rhinitis The animal model set forth in this example is used to test the observation that mast cells accumulate in response to allergic inflammation, and that this accumulation can be blocked by the H 4 receptor antagonists. The compounds of the present invention are tested in this model to demonstrate their use as a treatment for allergic rhinitis or asthma. Mice are sensitized by intraperitoneal injection of ovalbumin / alum (10 μg in 0.2 mL AI (OH) 3; 2%) on day 0 and day 14. On days 21 to 23, mice are challenged by PBS or ovalbumin, and sacrificed thereto 24 hours after the last challenge on day 24. A section of the The trachea is removed and fixed in formalin. Inclusion in paraffin and longitudinal sectioning of the tracheas is carried out, followed by staining of the mast cells with toluidine blue. Alternatively, the tracheas are frozen in OCT for frozen sectioning, and the mast cells are identified by IgE staining. The mast cells are quantified as submucosal or subepithelial, depending on their location within each tracheal section. Exposure to the allergen increases the number of subepithelial mast cells, and the ability of the H 4 receptor antagonists to block this effect is measured.

EXAMPLE 7 Inhibition of Basophil Chemotaxis by Histamine H4 Receptor Antagonists Basophils are isolated from human blood, using standard methods (Tsang et al., Immunological Methods (2000) 233 (1-2): 13-20). Chemotaxis tests can be carried out using transwells (Costar, Cambridge, MA) of a pore size of 8 μ? T? coated with 100 of 100 ng / mL of human fibronectin (Sigma) for 2 hours at room temperature. After the removal of fibronectin, 600 μL · of RPMI medium with 5% BSA in the presence of histamine (varying from 1.25 to 20 μm) to the lower chamber is added. To test the various histamine receptor antagonists, 10 μ? Solutions can be added. of the compounds to the upper and lower chambers. Basophils can be added to the upper chamber. The plates are incubated for 3 hours at 37 ° C. The transwells are removed, and the number of cells in the lower chamber counted for 60 seconds using a flow cytometer, or they can be quantified by staining using Wright's stain.

EXAMPLE 8 Measurement of increases in mast cell and basophil populations in patients after antigen challenge Patients with allergies to particular allergens such as cat dander or grass pollen are challenged with suitable antigens by direct bronchial administration or other methods. The number of mast cells and basophils in the bronchial mucosa and nasal mucosa is quantified using standard immunohistochemical staining methods after tissue biopsy. The effects of a modulator of the histamine H4 receptor on the increase in mast cells and basophils after challenge of the antigen are studied, administering the modulator by many different routes before the challenge with the antigen.

EXAMPLE 9 Measurement of basophil and tissue mast cell populations in patients It is known that patients with allergic rhinitis and / or asthma have increases in mast cells and basophils in their airways compared to healthy subjects. The effects of a modulator of the histamine H4 receptor on the population of mast cells and basophils residing in the respiratory tract are examined, administering the modulator by many different routes for a determined period. A comparison can be made of the number of mast cells and basophils present in the airways before, during and after treatment. Cells are quantified using standard immunohistochemical staining methods after tissue biopsy.

EXAMPLE 10 Treatment of asthma or allergic responses in patients, using histamine H 4 receptor antagonists A placebo or histamine H4 receptor antagonist is given to asthmatic or allergic patients for a period of time. Throughout the course of asthma treatment, the severity rating of the patient's condition, forced expiratory volume in one second (FEV1), and bronchial hyperreactivity (BHR) for bronchoconstrictors are measured. Decreases in asthma severity rating, increases in FEV1 and decrements in BHR after treatment with a histamine H4 receptor antagonist, are indicative of a positive effect on the disease. In addition, decreases in the inflammatory response can be determined after treatment with histamine H4 receptor antagonists, by changes in serum concentrations of the soluble interleukin-2 receptor (slL-2R), IL-4 and soluble intercellular adhesion molecule 1 ( slCAM-1); eosinophil count in peripheral blood; and eosinophilic cationic protein (ECP). In addition, bronchial biopsies can be used to quantify the change in inflammatory cell populations, such as eosinophils, T cells, mast cells, basophils, and the like.

EXAMPLE 11 Treatment of allergic responses in patients using histamine H4 receptor antagonists A histamine H4 receptor antagonist or placebo is given to patients with allergic rhinitis for a certain period of time. Efficacy is measured by comparing the qualification of the nasal symptoms in the day (average of congestion, itching and sneezing), ocular symptoms, nocturnal symptoms, nasal symptoms of the individuals in the day, global evaluations (by the patient and by the doctor) and quality of life qualifications. In addition, the effects on allergic conjunctivitis can be quantified using measurements of ocular redness, itching and days without symptoms.

EXAMPLE 12 Inhibition of eosinophil shape change induced by histamine. by histamine H4 receptor antagonists This example demonstrates that histamine H 4 receptor antagonists can block the response of the change of shape from human eosinophils to histamine. The change of form in eosinoyls is an early event that leads to one or more of several different biological outcomes, including adhesion, chemotaxis, degranulation and phagocytosis. These downstream events are associated with allergic responses and other inflammatory / immune responses, including allergic rhinitis and asthma.

Methods Human granulocytes were isolated from human blood, using a Ficoll gradient. Red blood cells were lysed with 5-1 OX of Qiagen lysis pH buffer at room temperature for 5 to 7 minutes. The granulocytes were harvested and washed once with a pH regulator for FACS. The cells were resuspended at a density of 2 x 10 cells / mL in reaction pH buffer. To test the inhibition by specific histamine receptor antagonists, 90 μL · of the cell suspension (~ 2 x 10 5 cells) were incubated with 10 μ? of one of several solutions of the test compound. After 30 minutes, 11 μl of one of the various concentrations of histamine was added. Ten minutes later, the cells were transferred to ice, and fixed with 250 μ? of pH regulator for ice-cold fixative (formaldehyde at 2%) for 1 minute. The change in shape was quantified using a forward valve autofluorescence with gate valve (GAFS) test (Byran et al., Am. J. Crit. Care Med. 165: 1602-1609, 2002).

Results The data in the following table show that histamine induces a dose-dependent change in the form of eosinophils. Histamine receptor (HR) antagonists were used to select which histamine receptor is responsible for the change in shape. Specific antagonists for the H1 receptor (diphenhydramine) or the H2 receptor (ranitidine) of histamine, did not alter the change in shape induced by histamine. However, a double H3 / H4 antagonist (thioperamide), and a specific histamine H4 receptor antagonist, (5-chloro-1 H -indol-2-yl) - (4-methyl-piperazin-1-yl) -metanone (K, = 5 nM), inhibited the change of shape of the eosinophils induced by histamine, with IC 50 values of 1.5 and 0.27 μ, respectively. The data is shown in table 5: TABLE 5 Change in number of times Histamine (μ?): ~? 10 1 0.1 0.01 0 Without antagonist of 1.34 1.31 HR 1.21 1.01 1.00 Antagonist from H4 to 1.09 1.05 1.05 1.01 10 μ? 1.00 Thioperamide at 10 μ? 1.08 1.05 1.01 1.04 1.00 Diphenhydramine at 10 1.63 1.50 1.18 1.03 1.00 μ? Ranitidine at 10 μ? 1.64 1.49 1.21 1.04 1.00 EXAMPLE 13 H4 Receptor Antagonists Modulate the Response in a Human Allergic Inflammation Model This example demonstrates that H 4 receptor antagonists modulate the allergic response in animals to ovalbumin-induced lung inflammation, a common animal model for human allergic inflammation.

Methods Sensitization and challenge of the animals Mice were sensitized (n = 8 per group) by intraperitoneal injection of ovalbumin / alum on days 0 and 14. On each of the days 21 to 24, the mice were previously treated for 15 minutes with vehicle or with (5-chloro-H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone before challenge for 20 minutes with PBS or ovalbumin.

Administration of examples of H 4 receptor antagonists In one experiment, (5-chloro-1 H -benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone was orally administered at 5, 20 or 50 mg / kg. In a second experiment, (5-chloro-1 H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone was administered orally at 0.5, 2 or 5 mg / kg. In a third experiment, (5-chloro-1H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone was administered subcutaneously at 20, 60 or 100 mg / kg.

Cell counting / data collection Twenty-four hours after the last challenge, the mice were sacrificed, and the total number of cells was determined, and a differential cell count was made in the bronchoalveolar lavage fluid (BAL).

Monitoring of airway hypersensitivity Airway hypersensitivity in ovalbumin-sensitized animals in conscious mice was monitored by whole-body plethysmography (Buxco). The mice were treated (n = 8 per group) as described above, for cell counting.

Results Analysis of the BAL fluid After challenge with ovalbumin, there was a dramatic increase in the total number of cells in the BAL fluid that was mainly due to increases in the number of eosinophils. As shown in Figures 5A and 5B, the total number of cells and the number of eosinophils decreased significantly at all doses of 5 mg / kg and more than (5-chloro-H-benzoimidazol-2-yl) - (4 -methyl-pperazin-1-yl) -metanone. The maximum reduction in the number of eosinophils was 50%. The same doses that provided maximum reduction in eosinophils, also significantly reduced the number of lymphocytes. At the highest dose tested (50 mg / kg), there was a reduction in the number of macrophages and an increase in the number of neutrophils. A reduction in macrophages was also observed at a dose of 5 mg / kg in the second experiment. These data indicate that H 4 receptor antagonists such as (5-chloro-1 H -benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone, have anti-inflammatory properties. Another H4 receptor antagonist, (5-chloro-1H-indol-2-yl) - (4-methyl-piperazin-1-yl) -methanone, was also tested in this model. Figure 6 shows the results for the differential cell count in the BAL fluid. Both the total number of cells and the number of eosinophils, decreased significantly to 60 and 100 mg / kg of (5-chloro-1 H-indol-2-i!) - (4-methyl-piperazin-1-yl) - methanone The maximum reduction in the number of eosinophils was 50%. At the same doses that provided reduction in eosinophils, the number of lymphocytes decreased significantly.

Analysis of airway hypersensitivity The airway hypersensitivity response to methacholine is shown in figure 7. Twenty-four hours after the final challenge of animals sensitized with ovalbumin, there was a dramatic increase in airway response to methacholine, measured by Penh (intensified pause), an index of bronchoconstriction. This hypersensitivity was decreased by treatment with (5-chloro-1H-benzoimidazol-2-yl) - (4-methyl-piperazin-1-yl) -methanone at any of the first three doses. These data indicate that H 4 receptor antagonists not only block airway inflammation, but also affect allergic hyperactivity of the respiratory tract, and thus, will be useful in the treatment of allergies and / or asthma in humans. The features and advantages of the invention are apparent to those skilled in the art. Based on this description, including the summary aspects, detailed description, background of the invention, examples, drawings and claims, the person skilled in the art will be able to make modifications and adaptations to various conditions and uses. These other embodiments are also within the scope of the invention. The publications referred to herein are incorporated herein by reference in their entirety.

Claims

NOVELTY OF THE INVENTION CLAIMS 1. - A method for identifying compounds that modulate mammalian histamine H4 receptor activity, comprising: a) combining a putative modulator compound of mammalian histamine H4 receptor activity with mammalian histamine H4 receptor and a known ligand of the receptor Histamine H4; and b) measuring an effect of the modulator on protein function or its ability to bind the ligand, wherein said effect is a modulation selected from the group consisting of inhibition, activation, antagonist, agonist and inverse agonist activity, wherein said compound modulator is a modulator of mast cell chemotaxis or chemotaxis of basophils. 2. The method according to claim 1, further characterized in that the effect measured in step b) is inhibition by competition between the modulator of step a) and a ligand of the known histamine H4 receptor by binding to the receptor. 3. The method according to claim 1, further characterized in that the effect measured in step b) is modulation of a second intracellular messenger of the histamine H4 receptor. 4. - The method according to claim 3, further characterized in that the second intracellular messenger is selected from the group consisting of cAMP, calcium and a reporter gene product. 5. The method according to claim 1, further characterized in that said compound is a chemotaxis modulator of mast cells. 6. The method according to claim 1, further characterized in that said compound is a modulator of chemotaxis of basophils. 7. A compound identified using the method claimed in claim 1, wherein said compound is an inhibitor of a mammalian histamine H4 receptor function and an inhibitor of mast cell chemotaxis or basophil chemotaxis in vivo or in vitro 8. The compound identified using the method according to claim 1, further characterized in that said compound is an agonist, antagonist or inverse agonist of a mammalian histamine H4 receptor. 9. The compound identified using the method according to claim 1, further characterized in that said compound modulates the expression of a gene encoding the mammalian histamine H4 receptor. 10. A monospecific antibody immunologically reactive with a protein of the mammalian histamine H4 receptor, wherein said antibody modulates chemotaxis of mast cells and chemotaxis of basophils. 11. The antibody according to claim 10, further characterized in that the antibody blocks binding to histamine or activation of the mammalian histamine H4 receptor protein. 12. A pharmaceutical composition comprising an active compound in the method claimed in claim 1 and a pharmaceutically acceptable carrier, wherein said compound is an asthma modulator or allergic responses. 13: - The use of the pharmaceutical composition claimed in claim 12, for preparing a medicament for treating a patient to modulate asthma or allergic responses or a disease or condition that is mediated by asthma or allergic responses, and an H4 receptor of histamine. 14. A pharmaceutical composition comprising an active compound in the method claimed in claim 1 and a pharmaceutically acceptable carrier, wherein said compound is a chemoattractant modulator of mast cells. 15. The use of the pharmaceutical composition claimed in claim 14 for preparing a medicament for treating a patient to modulate asthma or allergic responses or a disease or condition that is mediated by mast cell chemotaxis, and an H4 receptor histamine. 16. - A pharmaceutical composition comprising an active compound in the method claimed in claim 1 and a pharmaceutically acceptable carrier, wherein said compound is a modulator of basophil chemotaxis. 17. - The use of the pharmaceutical composition claimed in claim 16 for preparing a medicament for treating a patient to modulate asthma or allergic responses or a disease or condition that is mediated by chemotaxis of basophils, and a histamine H4 receptor . 18. - A pharmaceutical composition comprising a compound that modulates the activity of the mammalian histamine H4 receptor and a pharmaceutically acceptable carrier, wherein said compound is a chemotaxis modulator of mast cells or basophils in vitro or in vivo. 19. - The pharmaceutical composition according to claim 18, further characterized in that the compound inhibits mast cell chemotaxis. 20. - The pharmaceutical composition according to claim 18, further characterized in that the compound inhibits basophil chemotaxis. 21. - The pharmaceutical composition according to claim 18, further characterized in that the compound is an agonist, antagonist or inverse agonist of a mammalian histamine H4 receptor. 22. - The pharmaceutical composition according to claim 18, further characterized in that the compound modulates the expression of a gene encoding the mammalian histamine H4 receptor. 23. - The pharmaceutical composition according to claim 18, further characterized in that the compound further modulates the eosinophil shape change in vitro or in vivo. 24. - The use of a compound that modulates the activity of the mammalian histamine H4 receptor and a pharmaceutically acceptable carrier, wherein said compound is a chemotaxis modulator of mast cells or basophils in vitro or in vivo to prepare a pharmaceutical composition for treating a patient for modulating asthma or allergic responses or a disease or condition that is mediated by mast cell or basophil chemotaxis, and a histamine H4 receptor. 25. - The use as claimed in claim 24, wherein the pharmaceutical composition comprises a compound that inhibits the chemotaxis of mast cells. 26. The use as claimed in claim 24, wherein the pharmaceutical composition comprises a compound that inhibits the chemotaxis of basophils. 27. The use as claimed in claim 24, wherein the pharmaceutical composition comprises a compound that is an agonist, antagonist or inverse agonist of a mammalian histamine H4 receptor. 28. The use as claimed in claim 24, wherein the pharmaceutical composition comprises a compound that modulates the expression of a gene encoding the mammalian histamine H4 receptor. 29. The use as claimed in claim 24, wherein the pharmaceutical composition comprises a compound that modulates the eosinophil shape change in vitro or in vivo. 30. A method for identifying compounds that modulate the mast cell chemotaxis mediated by the mammalian histamine H4 receptor to histamine, the method comprising: a) in the presence or absence of a test compound that is tested as a histamine H4 receptor modulator, put the mast cells in proximity to the histamine under conditions that allow the movement of mast cells towards histamine; and b) measuring an effect of the modulator of the histamine H4 receptor on the movement of the mast cells towards histamine, wherein an increase or decrease in the speed of movement of the mast cells towards the histamine or in the number of mast cells that are move towards histamine, is indicative that the test compound modulates the chemotaxis of mast cells mediated by histamine H4 receptor to histamine.