CZ20032705A3 - Use of n-phenyl-2-pyrimidineamine derivativea against mast cell-based diseases like allergic disorders - Google Patents

Abstract

Use of the N-phenyl-2-pyrimidine-amine derivatives of formula I, in which the symbols and substituents have the meaning as given herein in free form or in pharmaceutically acceptable salt form in the manufacture of a pharmaceutical composition for the treatment of allergic rhinitis, allergic dermatitis, drug allergy or food allergy, angioedema, urticaria, sudden infant death syndrome, bronchopulmonary aspergillosis, multiple sclerosis or mastocytosis.

Description

Pharmaceutical compositions containing N-phenyl-2-pyridinamine derivatives for the treatment of mast cell diseases

Technical field

The present invention relates to a novel use of the N-phenyl-2-pyridinamine derivatives of the formula I in which the symbols and substituents have the meanings given below, in free form or in the form of a pharmaceutically acceptable salt for the preparation of a pharmaceutical composition for the treatment of allergic rhinitis, allergic dermatitis, allergy or food allergy, angioedema, urticaria, sudden infant death syndrome, bronchopulmonary aspergillosis, multiple sclerosis or mastocytosis; and a method of treating a warm-blooded animal, preferably a human, comprising administering to a warm-blooded animal suffering from one of the aforementioned diseases a therapeutically effective dose of a compound of Formula I.

BACKGROUND OF THE INVENTION

The compounds of the invention are generically and specifically described in patent applications EP 0 564 409 A1 and WO 99/03854.

In EP 0 564 409 A1 the compounds of the invention are described as being useful for the treatment of cancer, thrombosis, psoriasis, fibrosis, dermatosclerosis and arteriosclerosis. In accordance with the present invention, it has now been found that the compound of the invention surprisingly exhibits beneficial effects on allergic rhinitis, allergic dermatitis, drug allergy or food allergy, angioedema, urticaria, sudden infant death syndrome, bronchopulmonary aspergillosis, multiple sclerosis and especially mastocytosis. mastocytic neoplasms.

• · · · · · ·

SUMMARY OF THE INVENTION

The present invention relates to the use of N-phenyl-2-pyridinamine derivatives of the general formula I

in which

R 1 represents a 4-pyrazinyl group; 1-methyl-1H-pyrrolyl; an amino- or amino-lower alkyl-substituted phenyl group wherein the amino group is in each case free, alkylated or acylated; 1H-indolyl or nebo-imidazolyl attached to a carbon atom of a five-membered ring; or unsubstituted or lower alkyl substituted pyridyl group attached to a ring carbon atom and unsubstituted or substituted on nitrogen by an oxygen atom;

and R3 is each independently hydrogen or lower alkyl;

one or two of R 4, R 5, R 8, R 7 and R 8 are each nitro, fluoro-substituted lower alkoxy or a group of formula II

-N (R ₉ ) -C (= X) - (Y) _n -R 10 (II), wherein

R ₉ represents a hydrogen atom or a lower alkyl group

X is oxo, thio, imino, N-lower alkyl-imino, hydroxyimino or O-lower alkyl-hydroxyimino,

Y is oxygen or NH, n is 0 or 1 and

R ₁₀ represents an aliphatic group containing at least 5 carbon atoms or an aromatic, aromatic-aliphatic, cycloaliphatic, cycloaliphatic-aliphatic, heterocyclic or heterocyclicaliphatic group, and the remaining substituents R ₄ , R ₅ , R ₆ , R 7 and R ₈ each independently of the other hydrogen, lower alkyl which is unsubstituted or substituted by free or alkylated amino, piperazinyl, piperidinyl, pyrrolidinyl or morpholinyl, or lower alkanoyl, trifluoromethyl, free, etherified or esterified hydroxy, free, alkylated or acylated amino, or free or esterified carbox or a salt of such a compound comprising at least one salt-forming group, for the preparation of a medicament for the treatment of allergic rhinitis, allergic dermatitis, drug allergy or food allergy, angioedema, urticaria, sudden infant death syndrome, bronchopulmonary aspergillosis, mastocytosis or multiple sclerosis.

1-Methyl-1H-pyrrolyl is preferably 1-methyl-1H-pyrrol-2-yl or 1-methyl-1H-pyrrol-3-yl.

• ·

The amino or amino-lower alkyl-substituted phenyl group R 1, wherein the amino group is in each case free, alkylated or acylated, is a phenyl group substituted in any desired position (ortho, meta or para), the alkylated amino group preferably being mono- or di the lower alkylamino group, for example dimethylamino, and the lower alkyl portion of the amino-lower alkyl group is preferably a linear alkyl group having 1 to 3 carbon atoms, such as, in particular, methyl or ethyl.

The 1H-indolyl group attached to the carbon atom of the five-membered ring is 1H-indol-2-yl or 1H-indol-3-yl.

The unsubstituted or lower alkyl substituted by a pyridyl group attached to a ring carbon atom is a lower alkyl substituted or preferably unsubstituted 2-, 4- or preferably 3-pyridyl group, for example 3-pyridyl, 2-methyl-3-pyridyl or 4- methyl-3-pyridyl. The pyridyl group substituted on the nitrogen atom by an oxygen atom is a group derived from pyridine-N-oxide, i.e. the N-oxidopyridyl group.

A fluoro-substituted lower alkoxy group is a lower alkoxy group bearing at least one, but preferably several, fluorine atom substituents, in particular trifluoromethoxy or 1,1,2,2-tetrafluoroethoxy.

When X is oxo, thio, imino, N-lower alkyl-imino, hydroxyimino or O-lower alkyl-hydroxyimino, C = X, respectively, is C = O, C = S, C = NH, C = N-lower alkyl, C = N-OH and C = NO-lower alkyl, respectively. X is preferably oxo.

• ·

N preferably represents 0, i.e. the group Y is absent.

The group Y, if present, is preferably NH.

The term lower denotes within the scope of the present invention groups containing up to 7 carbon atoms inclusive, preferably up to 4 carbon atoms inclusive.

The lower alkyl groups R 1, R ₂ , R 3 and R ₉ are preferably methyl or ethyl.

The aliphatic group R ₁₀ containing at least 5 carbon atoms preferably contains no more than 22 carbon atoms, generally contains no more than 10 carbon atoms, and is such a substituted or preferably unsubstituted aliphatic hydrocarbon group, i. such a substituted or preferably unsubstituted alkynyl, alkenyl or preferably an alkyl group, such as an alkyl group containing 5 to 7 carbon atoms, for example an n-pentyl group. The aromatic group R ₀ having up to 20 carbon atoms and is unsubstituted or substituted, for example in each case unsubstituted or substituted naphthyl, such as especially 2-naphthyl, or preferably phenyl, the substituents are preferably selected from cyano, unsubstituted or hydroxy, amino or 4-methyl-piperazinyl-substituted lower alkyl groups, such as, in particular, methyl, trifluoromethyl, free, etherified or esterified hydroxy, free, alkylated or acylated amino, and free or esterified carboxy. In the aromatic-aliphatic group R ₁₀ , the aromatic moiety is as defined above, and the aliphatic moiety is preferably a lower alkyl group, such as, in particular, a C 1 -C 2 alkyl group which is substituted or preferably unsubstituted, for example benzyl. The cycloaliphatic group R ₁₀ preferably contains up to 30, more preferably up to 20 and most preferably up to 10 carbon atoms is

Is mono- or polycyclic and is substituted or preferably unsubstituted, such as a cycloalkyl group, in particular a five- or six-membered cycloalkyl group, such as preferably a cyclohexyl group. In a cycloaliphatic-group, R _xo is cycloaliphatic such as defined above and the aliphatic moiety is preferably lower alkyl, such as especially an alkyl group containing 1-2 carbon atoms which is substituted or preferably unsubstituted. The heterocyclic group R ₁₀ preferably contains up to 20 carbon atoms and is preferably a saturated or unsaturated monocyclic group containing 5 or 6 ring members and 1 to 3 heteroatoms, which is preferably selected from the group consisting of nitrogen, oxygen and sulfur atoms, in particular thienyl or 2-, 3- or 4-pyridyl, or a bi- or tricyclic group in which, for example, one or two benzene groups are fused to said monocyclic residue. In the heterocyclic-aliphatic group R 10, the heterocyclic moiety is as defined above and the aliphatic moiety is preferably a lower alkyl group, such as in particular an alkyl group having 1 to 2 carbon atoms, which is substituted or preferably unsubstituted.

The etherified hydroxy group is preferably a lower alkoxy group. The esterified hydroxy group is preferably a hydroxy group esterified with an organic carboxylic acid such as a lower alkanoic acid or a mineral acid such as a hydrohalic acid, for example a lower alkanoyloxy group, or especially a halogen atom such as iodine, bromine or especially fluorine or chlorine.

An alkylated amino group is, for example, a lower alkylamino group such as a methylamino group or a di-lower alkyl amino group such as a dimethylamino group. Acylated amino is, for example, lower alkanoylamino or benzoylamino.

An esterified carboxy group is, for example, a lower alkoxycarbonyl group such as a methoxycarbonyl group.

A substituted phenyl group can carry up to 5 substituents, such as a fluorine atom, but especially in the case of relatively large substituents, it is generally substituted with only 1 to 3 substituents. Examples of substituted phenyl which may be mentioned specifically are 4-chlorophenyl, pentafluorophenyl, 2-carboxyphenyl, 2-methoxyphenyl, 4-fluorophenyl, 4-cyanophenyl and 4-methylphenyl.

The salt-forming groups in the compound of formula I are groups or moieties with basic or acidic properties. Compounds containing at least one basic group or at least one basic radical, for example a free amino, pyrazinyl or pyridyl radical, may form acid addition salts, for example with inorganic acids such as hydrochloric acid, sulfuric acid or phosphoric acid, or with suitable organic carboxylic acids or sulfonic acids, for example aliphatic mono- or dicarboxylic acids such as trifluoroacetic acid, acetic acid, propionic acid, glycolic acid, succinic acid, maleic acid, fumaric acid, hydroxymaleic acid, malic acid, tartaric acid, citric acid or oxalic acid, or amino acids such as arginine or lysine, aromatic carboxylic acids such as benzoic acid, 2-phenoxybenzoic acid, 2-acetoxybenzoic acid, salicylic acid, 4-aminosalicylic acid, aromatic aliphatic organic carboxylic acids such as mandelic acid or cinnamic acid, heteroaromatic carboxylic acids such as nicotinic acid or isonicotinic acid, aliphatic sulfonic acids such as methane-, ethane- or 2-hydroxyethanesulfonic acid, or aromatic sulfonic acids, e.g. benzene-, p-toluene- or ························

-8- naphthalene-2-sulfonic acid. If several basic groups are present, mono- or poly-acid addition salts may be formed.

Compounds of formula I containing acidic groups, for example the free carboxy group in R ₁₀ , may form metal salts or ammonium salts, such as alkali metal or alkaline earth metal salts, for example sodium, potassium, magnesium or calcium salts, or ammonium salts with ammonia or with suitable organic amines such as tertiary monoamines, for example triethylamine or tri- (2-hydroxyethyl) amine, or with heterocyclic bases, for example N-ethylpiperidine or N, N'-dimethylpiperazine.

Compounds of formula I containing both acidic and basic groups can form internal salts.

For the purposes of isolation or purification, as well as compounds which are further used as intermediates, it is also possible to use pharmaceutically unacceptable salts. However, only pharmaceutically acceptable, non-toxic salts are used for therapeutic purposes, and these salts are therefore preferred.

The compounds of formula I can be used to treat or to prepare medicaments for treating warm-blooded animals, preferably humans.

The term allergic rhinitis as used herein means any allergic reaction of nasal mucosa. Such an allergic reaction may occur, eg, permanently, eg, spring conjunctivitis, or seasonally, eg, hay fever.

The term allergic dermatitis as used herein means, in particular, atopic dermatitis, allergic contact dermatitis and eczematous dermatitis, but also includes, for example, seborrheic dermatitis, Lichen planus, urticaria and acne. Atopic dermatitis, as defined herein, is chronic.

inflammatory skin disease observed in subjects with a hereditary predisposition to a decreased skin threshold to itching. It is characterized primarily by extreme itching, leading to scratching and friction, which in turn result in typical eczematic lesions. Allergic contact dermatitis as defined herein is a form of dermatitis that is caused by allergic sensitization to a variety of substances that cause inflammatory reactions in the skin of those who have acquired acquired allergen hypersensitivity as a result of prior exposure to the allergen.

The term drug allergy or food allergy, as used herein, refers to an allergic reaction caused by a drug or ingested antigens such as strawberries, milk or eggs.

The term bronchopulmonary aspergillosis refers to Aspergillus lung infection.

The term mastocytosis as used herein refers to systemic mastocytosis, for example mastocytoma, and especially canine mast cell neoplasms.

The compounds of formula I are hereinafter collectively referred to as COMPOUNDS OF THE INVENTION.

Mast cells play an important role as primary effector cells in the allergic disorders mentioned herein. Antigen-specific IgE-mediated mast cell degranulation results in subsequent release of chemical mediators and many cytokines and leukotriene synthesis. In addition, mast cells are involved in the pathogenesis of multiple sclerosis.

Neoplasms from mast cells occur in both humans and animals. In dogs, mast cell neoplasms are called mastocytomas and the disease is common, representing 7% to 21%

- 10 · canine tumors. A distinction must be made between human mastocytosis, which is usually transient or painless, and canine mast cell neoplasm, which behaves unpredictably and is often aggressive and metastatic. For example, human solitary mastocytomas are essentially never metastasized; in contrast, 50% of canine mastocytomas behave in a malignant manner, as determined by Hottendorf & Nielsen (1969) after viewing 46 published tumor reports in 938 dogs.

Cancer is a spontaneous disease in the domestic animal population. Pet owners, motivated by extending the quality of life of their animals, often seek specialized care and therapy for veterinary oncologists at private reference veterinary hospitals and veterinary teaching hospitals across the country. Therapeutic methods are similar to those in human veterinary cancer patients and include surgical treatment, chemotherapy, radiation therapy, and biotherapy. It has been found that there are 42 million dogs and approximately 20 million cats in the United States. Using rough estimates of cancer incidence, there are about 4 million newly diagnosed cancers per year in dogs and similar amounts in cats.

Cutaneous mast cell tumors are a common problem in dogs. Most mast cell tumors are benign and are cured by simple resection; however, if it is recurrent or metastasized to remote sites, therapeutic options are limited. Therapeutic options in recurrent lesions may include external radiation therapy. In distant metastases or disseminated disease, the use of chemotherapeutic protocols including Lomustine® and vinblastine shows some beneficial effects. The mast cell metastasis sites include skin, regional lymph nodes, spleen, liver and bone marrow.

·· ·· * ·

The observation that several mutations leading to substantial activation of KIT have been identified in a number of mast cell lines suggests the involvement of KIT receptors in the pathogenesis of mastocytosis. For example, a point mutation in human c-KIT, causing Val to Asp816 substitution in the phosphotransferase domain and receptor autoactivation, occurs in the long-term human mast cell leukemia (HMC-1) line and at the appropriate codon in two rodent mast cell lines. Furthermore, this activating mutation has been found in situ in some cases of human mastocytosis. Two additional activating mutations were found in the intracellular juxtamembrane region of KIT, i. Val560Gly substitution in the human mast cell line HMC-1, and deletion of seven amino acids (Thr573-His579) in the rodent mast cell line called FMA3.

Established test models, and in particular those described herein, can be shown that the COMPOUNDS OF THE INVENTION, or in any case a pharmaceutically acceptable salt thereof, results in effective prevention or, preferably, treatment of at least one of the diseases listed herein. One of ordinary skill in the art is fully able to select a relevant test model to verify the therapeutic indications and benefits listed above and below. For example, pharmacological activity can be demonstrated in a clinical study or test procedure as essentially described below.

DETAILED DESCRIPTION OF THE INVENTION

Part A

Example 1

This example shows the in vitro effects of the COMPOUNDS OF THE INVENTION on SCF-dependent development of growth of cultured human CD34 ⁺ umbilical cord blood cell cultured cells using the culture system described in Kinoshita T., Sawai N. et al., Blood 1999, 94, 496-49. 508. More than 90% of isolated cells are CD34-positive according to flow cytometric analysis.

Reagents and antibodies

The compounds of the invention are dissolved in PBS at a concentration

10 ² M (Sigma) v

The purified and stored acid stores -80 ° C.

at -80 ° C. All-trans retinoin is dissolved in ethanol at a concentration of 10 ^-2 M and ampoules protected from light at tryptase (MAB1222) can be obtained from Chemicon International Inc., CA. For flow cytometric analysis, mAbs for CD34 (8G12, FITC) and CD11b (Leul5, PE) from Becton Dickinson Immunocytometry Systems (Mountain View, CA) and mAb for CD41 (SZ22, FITC) from Immunotech SA (Marseilles, France) were obtained. A glycophorin A mAb (GPA, JC159, FITC) can be obtained from Dako (Glostrup, Denmark). For western blotting and immunoprecipitation, c-kit (NU-c-kit) and phosphotyrosine (4G10) mAbs can be obtained from Nichirei,

Biotechnology, Inc. (Lake Placid, NY).

respectively Upstate

Suspension culture

Sera devoid of liquid culture were run on twenty-four-well culture plates (# 3047; Becton Dickinson). Twenty thousand CD34 ⁺ are cultured in each well containing 2 ml α-medium supplemented with 1% BSA, 300 μg / ml fully iron-saturated human transferrin (approximately 98% pure, Sigma), 16 μg / ml soya lecithin (Sigma), 9.6 μg / ml cholesterol (Nakalai Chemicals Ltd., Japan) and 20 ng / ml SCF, 10 ng / ml GM-CSF, 2 U / ml EPO, 10 ng / ml TPO,

According to the invention, the evaluation of the effects of the COMPOUND alone in concentration. OF THE INVENTION

COMPOUNDS For the purpose of:

SCF-dependent mast cell development varies or

According to the target cells, cells of a ten-week culture cultured with 20 ng / ml SCF from CD34 ⁺ umbilical cord blood cells will be used. Five to ten x 10 ⁴ 10 weeks of cultured cells are incubated for 2 weeks

·· ·· «· • • · ·· • o · · • • • ·· · • • • • • • • · • • • • « • • · Φ · • ♦ • • · • · · * ·· • · • ·

on 24-well culture plates containing 20 ng / ml SCF with or without different concentrations of the COMPOUND OF THE INVENTION. The plates are incubated at 37 ° C in a humidified atmosphere, purged with 5% CO ₂ , 5% O ₂ and 90% N ₂ . If the culture is continued within 4 weeks, half of the culture medium is replaced every 2 weeks with fresh medium containing factor (s). The number of viable cells is determined by a tryptan blue exclusion assay using a hemocytometer.

Cultivation of clonal cells

Mast cell colony assays are performed in 35mm Lux suspension culture flasks (# 171099; Nunc, IL). The culture consists of 10-week cultured cells (4000 cells / ml) cultured with 10 ng / ml SCF, α-medium, 0.9% methylcellulose (Shinetsu Chemical, Japan), 1% BSA, 300 pg / ml fully iron-saturated human transferrin, 16 pg / ml soya lecithin, 9.6 µg / ml cholesterol, and 100 ng / ml SCF with or without 10 ^-8 COMPOUNDS OF THE INVENTION. The vessels were incubated at 37 ° C in a humidified atmosphere, purged with 5% CO ₂ , 5% O _2, and 90% N ₂ . After 4 weeks, aggregates containing 30 or more cells are recorded as mast cell colonies, and those consisting of 10-29 cells as mast cell clusters. Thirty individual colonies and clusters were picked and stained with antitryptase mAb or mouse IgG1 using the alkaline phosphatase-anti-alkaline phosphatase (APAAP) technique. Almost all the basic cells are tryptase positive.

Cytochemical and immunological staining

The cultured cells are plated on a glass plate using Cytospin II. Cytochemical reaction with peroxidase (POX) is carried out in a conventional manner. Reaction with tryptase mAbs is detected by the APAAP method (Dako APAAP Kit System, Dako Corp., CA), described by F. Ma, K. Koike et al., Br. J. Haematol. 1998, 100, 427-35.

• 4 4 4 4

4 4 44 4 4 4 4 4 · 444 444 444 444

44 44 44 44 44

Immunoprecipitation and Western blotting

Immunoprecipitation and Western blotting are performed as described by T. Kamijo, K. Koike et al., Leuk. Res. 1997, 21, 1097-106.

Flow cytometric analysis

To analyze surface markers on cultured cells, 1 to 2 × 10 ⁵ cells are collected in plastic tubes and cultured with appropriately diluted FITC- or PE-mAbs as described by Kinoshita T., Sawai N. et al., Blood 1999, 94, 496 to 508.

Cells are washed twice, after which their surface markers are analyzed with a FACScan flow cytometer. The viable cells go through their front light scattering characteristics and side scattering characteristics. The proportion of positive cells is determined by comparison to cells stained with FITC- or PE-conjugated mouse isotype matched Ig.

Detection of cell apoptosis

Cell apoptosis analysis is performed by flow cytometric analysis using propidium iodide (PI, Sigma) according to the procedure described by N. Sawai, K. Koike et al., Stem Cells. 1999, 17, 45-53. Percol gradient centrifugation can be used to reduce cells undergoing apoptosis, necrosis, or already dead. Ten weeks of cultured cells are plated on 27% Percoll (Sigma) in α-medium and 54% Percoll in PBS. After centrifugation, cells are collected from the interface of two different concentrations of Percoll solution, washed with PBS and treated with 1 ml Ortho PermeaFix ™ for 40 minutes at room temperature. The cells are then incubated with DNase-free RNase (Sigma) for 15 minutes at 37 ° C and stained with PI for 15 minutes. The DNA content of 2 × 10 ⁴ cells is monitored by

9 * 9 · · * • • • • · · 9 • • 9 • • • • · • • 9 • 9 ·· • • · • 9 ·· · * <·· 9 9 « • • · · ♦

flow cytometer. Ten weeks of cultured cells (2 x 10 ⁸ ) exposed to SCF or SCF and the COMPOUND OF THE INVENTION are lysed for 10 minutes on ice in 100 μΐ of hypotonic lysis buffer [10 mmol Tris (pH 7.5), 10 mmol EDTA, pH 8, 0.5%

Triton X-100]. After centrifugation for 10 minutes at 14000 g, the supernatant is transferred to a new tube and treated with 0.2 mg / ml RNase A (Sigma) and 0.2 mg / ml Proteinase K (Sigma). DNA is precipitated with 120 μΐ isopropanol and 20 μΐ 5M NaCl overnight at -20 ° C. After centrifugation at 14000 g, the pellets are dried, dissolved in 20 μΐ of Tris-EDTA and then analyzed by gel electrophoresis in 2% agarose and ethidium bromide staining.

Analysis of histamine, tryptase and cytokine levels

Histamine concentrations in cell lysates obtained by treating cultured cells with 0.5% Nonidet P-40 and supernatant are determined using the Histamine Radioimmunoassay (RIA) Kit (Immunotech) as described by Kinoshita T., Sawai N. et al., Blood 1999, 94 496-508.

Statistical analysis

All experiments should be performed at least three times. An unpaired t-test or Mann-Whitney-U test can be used to determine a significant difference between two independent groups if data are not normally distributed.

Results obtained for N- {5- [4- (4-methyl-piperazino-methyl) -benzoylamido] -2-methyl-phenyl} -4- (3-pyridyl) -2-pyrimidine-amine monomesylate (STI571B)

The addition of STI571B at a concentration of 10 ^-6 mol or higher to a culture with SCF almost completely inhibits offspring generation.

· ♦ 9 9 9 9 9 ♦ 9 9 9 9

In 9 9 9 4 9 9 9 10 11 12 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25

For SCF plus STI571B, the number of viable cells is unchanged on day 2 and then decreases with time. On day 14, the total cell count reaches a negligible level. Consistent with the results obtained with 10 weeks of cultured STI571B mast cells at a concentration of 10 ^-6 mol, apparently reduced cell formation in CD34 ⁺ cells under SCF stimulation. Further, STI571B induces programmatic death of cultured mast cells cultured under SCF stimulation. The percentage of sub-diploid nuclei increases with the culture period. This observation can be confirmed by DNA stratification of cells exposed to SCF plus STI571B by gel electrophoresis. STI571B exhibits its inhibitory effects in the early phase of mast cell development as well as the growth of later emerging mast cells.

The results obtained show the inhibition of SCF-dependent human mast cell growth by the COMPOUNDS OF THE INVENTION, eg STI571B. Therefore, they show that the COMPOUNDS OF THE INVENTION can be used to treat the diseases described herein.

Part B: Mastocytosis

Example 2: Methods

Reagents: Novartis Pharma; Basel, Switzerland provides SALT I for use in these experiments. Prior to each experiment, fresh 10 mM inhibitor stock solutions were prepared by dissolving the compound in 1 mL phosphate buffered saline (PBS; Gibco-BRL).

Antibodies

Ab-1) se

Cambridge, used in

Lexington, antibody antibody

Polyclonal rabbit anti-KIT antibody (c-kit used at 1: 500 dilution (c-kit Ab-1; Oncogene, MA). Anti-phosphotyrosine antibody (PY20) at 1: 1000 dilution (PY20 Transduction Laboratories;

KY). Peroxidase-conjugated goat anti-mouse was used at a 1: 5000 dilution and goat anti-rabbit at a 1: 10,000 dilution (Pierce; Rockford, IL).

• 9 9

Cell lines: BR and C2 lines of canine mastocytoma cells are obtained from Dr. George Caughey (University of California, San Francisco, CA). Both cell lines are stored in DMEM supplemented with 2% bovine calf, 1 mmol L-glutamine,

12.5 mmol HEPES (pH 7.5), 0.25 mg / ml histidine, 1% Penicillin-Streptomycin and 1% fungizone. BR and C2 cells originate from canine mast cell tumors and were initially introduced into long-term culture after initial passage in immunodeficient mice (DeVinney R. et al., Am J Respir Cell Mol Biol 1990; 3 (5): 413-420; Lazarus SC and et al., Am J Physiol 1986; 251 (6 Pt 1): C935 to C944). The BR cell line shows a point mutation (T1752C) resulting in the substitution of Leucine for Proline at amino acid 575 (juxtamembrane domain). The C2 cell line exhibits an internal tandem duplication (ITD) of the KIT of the juxtamembrane region. Translation of this ITD leads to reduplication of amino acid residues at the 3 'end of exon 11 (London CA et al., Exp Hematol 1999; 27 (4): 689-697; Ma Y. et al., J Invest Dermatol 1999; 112 (2) : 165-170).

Proliferation assays: Cells are plated on ninety-six well plates at a density of 40,000 cells / well in normal culture medium and various concentrations of SOLI I. Proliferation is assessed at 48-72 hours using XTT-based analysis (Roche Molecular Biochemicals; Indianapolis, IN) ( Heinrich MC et al., Blood 2000; 96 (3): 925-32).

Protein Lysates: BR and C2 cells are washed twice in PBS and then allowed to stand in Optimem (Gibco-BRL) at 37 ° C for approximately 18 hours. The cells are then incubated for 90 minutes in the presence of different concentrations of SOLI I. After this incubation, the cells are pelleted and lysed with 100 to 250 μΐ protein lysis buffer (50 mmol Tris, 150 mmol NaCl, 1% NP-40, 0.25% deoxycholate, with the addition of aprotinin, leupeptin, pepstatin, PMSF and sodium orthovanadate inhibitors [Sigma]). Western immunoblot analysis was performed as previously described

(Hoatlin, E. E. et al., Blood 1998; 91 (4): 1418-1425;

Heinrich M. C. et al., Blood 2000; 96 (3): 925-932).

Example 3: COMPOUND I inhibits constitutively activated KIT kinase associated with canine mast cell tumors.

Two cell lines (BR and C2) that express two different constitutively activated isoforms of KIT are used to test the efficacy of COMPOUND I to inhibit the kinase activity of mutant forms of canine KIT. KIT mutations in these cell lines are both located in the juxtamembrane domain and are homologous to those observed in human gastrointestinal stromal tumors (GISTs) (Lux ML et al., Am J Pathol 2000; 156 (3): 791-795; Ruby Β. P). et al., Cancer Res 2001; 61 (22): 8118-8121). Lysates prepared from BR or C2 cells are screened with anti-P-Tyr antibody and the activation of the KIT receptor is assessed by measuring autophosphorylation. As previously described, KIT autophosphorylation is observed in these cells in the absence of SLF (Ma Y. et al., J Invest Dermatol 1999; 112 (2): 165-170; Ma Y. et al., Journal of Investigative Dermatology 2000; 114 (2): 392-1

394). Inhibition of KIT autophosphorylation by COMPOUND I is dose-dependent with complete inhibition observed using 10 and Ι, ΟμΜ doses. Nearly complete inhibition was observed using the Ο, ΙμΜ dose. Limited c-kit autophosphorylation is observed using 0.001 to Ο ΟμΜ doses of COMPOUND I. Thus, Compound I not only inhibits the autophosphorylation of the mutated c-kit receptor in these cells, but is also a more potent inhibitor of this mutated receptor than the untreated c-kit receptor (IC ₅₀ 100-200 nmol) (Heinrich MC et al., Blood 2000;

96 (3): 925-932). To determine whether COMPOUND I modulates KIT protein expression, the membrane is peeled and re-screened with anti-c-kit antibody. No change in c-kit protein expression is observed in cells treated with COMPOUND I. Therefore, COMPOUND I reduces autophosphorylation of the mutant canine KIT polypeptide by inhibiting KIT kinase activity rather than

• fc * fcfc by regulation to reduce the expression of KIT protein.

Example 4: COMPOUND I inhibits the proliferation of canine mast cell tumor cell lines.

To test the biological effect of inhibiting the kinase activity of mutant c-kit receptors, BR or C2 cells are cultured for 48 to 72 hours in the presence of different concentrations of COMPOUND I. At inhibitor concentrations of 0.1 to 10 pmol, proliferation decreases by 90 to 95% , compared to cells treated with media only. Partial inhibition of proliferation was observed at doses of 0.001 to 0.01 μπιοί of COMPOUND I. The reduction in proliferation observed at doses of 0.01 to 10 μπιοί of inhibitor was statistically significant (p <0.001). Therefore, COMPOUND I inhibits proliferation of BR and C2 cells in the same dose response range as observed for inhibition of receptor autophosphorylation. Morphological observations of inhibitor-treated cells reveal changes consistent with apoptosis (data not shown).

Table 1: BR cells

Diameters O. O SD % SD Cells 0,929 100% 0,030447 3% 5 umol 0,083 9% 0.001732 0% 1 umol 0.105 11% 0,002 0% , 1 µmol 0.105 11% 0.002082 0% .01 umol 0.479 52% 0,043016 5% , 001 umol 0,781 84% 0.033081 4%

Table 2: C2 cells

Diameters O. 0 SD % SD Cells 1,236 100% 0.04417 4% 5 umol 0,032 3% 0.005686 0% 1 umol 0, 037 3% 0.013868 1% , 1 µmol 0,028 2% 0.003512 0% .01 umol 0,754 61% 0.185236 15% , 001 umol 1,065 86% 0,055245 4%

Tables 1 and 2: BR or C2 cells are plated in ninety-six well plates at a concentration of 40,000 cells / well and cultured in normal culture medium and various concentrations of COMPOUND I. Cell proliferation is measured at 72h using an XTT-based assay system. Each concentration of COMPOUND I is analyzed in triplicate. Results are expressed as a percentage of maximum proliferation (cells only, no COMPOUND I) ± 1 standard deviation (SD). Representative results of one of six experiments are shown.

Compositions for pets and people

Example 5: 4 - [(4-methyl-1-piperazin-1-ylmethyl) -N- [4-methyl-3 - [[4- (3-pyridinyl) -2-pyrimidinyl] amino] phenyl] benzamide capsule -methanesulfonate, crystal form β

Capsules containing 11.95 mg of the title compound (= SALT I), corresponding to 10.0 mg of COMPOUND I (free base), are prepared as active ingredient as follows:

♦ ·

Ingredients :

SALT I 11.95 mg

Cellulose MK GR 9.2 mg

Crospovidone XL 1.5 mg

Aerosil 200 0.2 mg

Magnesium stearate 0.15 mg

23.0 mg

Capsules are prepared by mixing the ingredients and filling the mixture into hard gelatin capsules of size 1.

Example 6: 4 - [(4-Methyl-1-piperazin-1-ylmethyl) -N- [4-methyl-3 - [[4- (3-pyridinyl) -2-pyrimidinyl] amino] phenyl] benzamide capsule -methanesulfonate, crystal form β

Capsules containing 10.0 mg of the title compound (= SALT I) as active substance are prepared as follows:

Ingredients:

Active substance 10.0 mg

Avicel 20,0mg

PVPPXL 1.5 mg

Aerosil 0.2 mg

Magnesium stearate 0.15 mg

31.85 mg

Capsules are prepared by mixing the ingredients and filling the mixture into hard gelatin capsules of size 1.

• · · · · ···

·· · »

Example 7: 4 - [(4-Methyl-1-piperazin-1-ylmethyl) -N- [4-methyl-3 - [[4- (3-pyridinyl) -2-pyrimidinyl] amino] phenyl] benzamide methanesulfonate capsule , crystal form β

Capsules containing 119.5 mg of the title compound (= mesylate of COMPOUND I), corresponding to 100 mg of COMPOUND I (free base), are prepared as active ingredient as follows:

Ingredients:

Mesylate COMPOUNDS I 119.5 mg Cellulose MK GR 92 mg

Crospovidone XL 15 mg

Aerosil 200 2 mg

Magnesium stearate 1.5 mg

230 mg

Capsules are prepared by mixing the ingredients and filling the mixture into hard gelatin capsules of size 1.

Example 8: An example of a prospective series of pet cases - dogs with measurable mast cell skin tumors

The patients studied are pets - dogs with measurable and histologically confirmed mast cell tumors. Cases are limited to cases with measurable lesions accessible for biopsy.

The classification criteria are:

- histologically confirmed measurable mast cell skin tumors,

- cases will require serial biopsy using 2mm Keyes puncture before and during therapy,

- histological grade (II-medium or III-poorly differentiated),

- performance status 0 or 1 (Modified Karnofsky - Table 3),

- informed consent of the owner.

(a) The exclusion criteria are:

- concomitant cytotoxic chemotherapy,

- prednisone and non-steroidal anti-inflammatory drugs should not be started within 30 days of the study; if prednisone or non-steroidal anti-inflammatory drugs are administered for more than 30 days, they may be continued,

- abnormal serum bile acid test (liver function).

Table 3: Physical Activity Status (Modified Karnofsky)

Grade Description 0 Normal activity

Limited activity; decreased activity compared to the pre-disease state

Weakened; mobile only for life activities; it is emptied and urinated in acceptable places consistently; must be forced into food; unable to keep urination and emptying to acceptable locations

Dead

Pre-treatment evaluations include physical examination, complete blood count, buffy coat, serum biochemistry, urine analysis, serum bile acid (fasting and post-prandial), regional lymph node size documentation, abdominal radiographs and abdominal ultrasound. The therapeutic regimen is 25 mg / kg PO QD x 60 days SOLI I.

In all cases, therapy is continued for 60 days unless disease progression is noted. In cases • · • · · · • ·

recording a partial response or a complete response may be considered to continue therapy for an additional 60 days. Cases of successfully completing therapy are eligible for repeated entry into the study.

Table 4: Treatment and clinical trial plan

Activity Day 0 Day 7 Day 14 Day 28 ql4 days Clinical Raindance Connect ¹ X X X Physical exam X X X X X Tumor volume measurement ² X X X X X Start SOLI I 25 mg / kg QD X Pharmacokinetics ³ X Incision biopsy ⁴ X X Repeated fixation X

¹ Initial Raindance Connect includes a physical examination, CBC, buffycoat, serum biochemistry tests bile acids), urinalysis, abdominal ultrasound. Reassessing physical examination and measuring tumor volume or repeated clinical stability.

² The tumor volume was measured at day 0, 7, 14, 28 and every 14 days. The therapeutic response is defined against the measurable skin lesion (s) and other lesions found during fixation (CR, PR, SD, PD - defined below).

³ Plasma collection from the first five incoming cases is performed at 0, 0.5, 1, 2, 5, 8, 12, 16, 24 after the first dose of SOLI I.

⁴ Incision biopsies from defined measurable lesion (s) are taken on day 0 and 28 in all cases. Further biopsies are taken at partial response time (PR) and after complete objective response.

liver function (serum abdominal radiographs and evaluation may include

The efficacy of COMPOUND I against measurable cutaneous mast cell tumors was evaluated by clinical outcomes.

· · • · * * *

Biological outcomes can be obtained from serial biopsies taken from skin tumors and from blood samples available during treatment.

Clinical outcomes include measurable tumor response rates, objective response to measurable tumors, and time to measurable tumor progression. All adverse side effects shall be recorded.

Objective tumor responses (s) as defined below are observed in treatment with COMPOUND I and show the efficacy of the therapeutic regimen.

In particular, complete responses and partial responses to Compound I treatment can be observed. Furthermore, it can be observed that more animals receiving therapy show a stable disease, while less treated animals show progressive disease. It can also be seen that fewer animals receiving therapy show a relapse of the disease compared to untreated animals. Time to progression, duration of remission and survival may increase in animals receiving Compound I therapy.

A complete response (CR) is defined as the disappearance of all clinical signs of cancer and any signs of cancer.

The partial response (PR) is defined as a 50% or greater reduction in the sum of the measurement results of representative lesions, without increasing the size of any lesion or the appearance of any new lesions.

Stable disease (SD) is defined as no response or response less than the response defined for a partial response or progressive disease without the appearance of any new lesions or worsening clinical signs.

• · ····

Progressive disease (PD) is defined as an unquestionable increase, at least 50%, in the size of any measurable lesion or the appearance of new lesions.

Relapse (R) is defined as the appearance of new lesions or the reoccurrence of old lesions in dogs that showed a complete response; in dogs that showed only a partial response, relapse is defined as an at least 50% increase in the sum of the measurement results of representative lesions compared to those obtained at the time of maximal response.

Time to progression ( TTP) is recorded from on 0 protocol. TTP is defined as number of days from start therapy (from day 0) to relapse (R). Time duration of remission se defined as count days from

objective responses (PR or CR) to relapse.

Survival is defined as the number of days from the start of Compound I therapy to death. The cause of death is recorded, but may include disease progression, toxicity, and others.

The results obtained show the inhibition of mast cell neoplasms by the COMPOUNDS OF THE INVENTION, eg STI571B.

Preferred are COMPOUNDS OF THE INVENTION in which one or two of R ₄ , R ₅ , R ₆ , R ₇ and R ₈ are each nitro or a group of formula II wherein R ₉ is hydrogen or lower alkyl, X is oxo , thio, imino, N-lower alkyl-imino, hydroxyimino or O-lower alkyl-hydroxyimino, Y is O or NH, n is 0 or 1 and R 10 is an aliphatic group containing at least 5 carbon atoms or an aromatic, aromatic-aliphatic,

The cycloaliphatic, cycloaliphatic-aliphatic, heterocyclic or heterocyclicaliphatic group, and the remaining substituents R ₄ , R ₅ , R ₆ , R ₇ and R 8 are each independently of each other a hydrogen atom, a lower alkyl group which is unsubstituted or substituted by a free or alkylated amino, piperazinyl, piperidinyl, pyrrolidinyl or morpholinyl group, or a lower alkanoyl, trifluoromethyl, free, etherified or esterified hydroxy group, free, alkylated or an acylated amino group or a free or esterified carboxy group, and the remaining substituents have the meanings defined above.

Preferred are COMPOUNDS OF THE INVENTION in which

R 1 is a pyridyl group or an N-oxidopyridyl group, each of which is attached to a carbon atom,

R2 and _R3 are each hydrogen,

R ₄ is hydrogen or lower alkyl,

R ₅ represents a hydrogen atom, a lower alkyl group or a trifluoromethyl group,

Rg represents a hydrogen atom,

R ₇ is nitro, fluoro-substituted lower alkoxy or a group of formula II wherein

R 8 represents a hydrogen atom, represents an oxo group,

n stands for Ο and

R 10 represents a pyridyl group attached to a carbon atom, a phenyl group which is unsubstituted or substituted by a halogen atom, a cyano group, a lower alkoxy group, a carboxy group, a lower alkyl group or a 4-methylpiperazinylmethyl group or an alkyl group containing 5 to 7 carbon atoms, thienyl, 2-naphthyl or cyclohexyl, and

R 8 represents a hydrogen atom.

Especially preferred are COMPOUNDS OF THE INVENTION wherein at least one of R ₄ and R 8 is lower alkyl and the remaining substituents have the meanings defined above.

Particularly preferred are the COMPOUNDS OF THE INVENTION in which

R _x represents a pyridyl group attached to a carbon atom,

_R2, R3, R5, R6 and Rs are each hydrogen,

R ₄ represents a lower alkyl group,

R ₇ represents a group of formula II wherein

Rg represents a hydrogen atom,

X is oxo, n is 0 and

R 10 represents a 4-methyl-piperazinyl-methyl group.

Most preferred is a compound of the formula I, which is CGP 57148B {N- {5- [4- (4-methyl-piperazinomethyl)].

9

* Benzoylamido] -2-methylphenyl} -4- (3-pyridyl) -2-pyrimidineamine monomesylate}.

Very preferably, the compound of the invention is used in the form of its monomesylate salt.

According to one preferred embodiment of the invention, the disease to be treated is selected from the group consisting of allergic rhinitis, allergic dermatitis, drug allergy and food allergy. In another embodiment of the invention, the disease to be treated is multiple sclerosis. According to another embodiment of the invention, the disease to be treated is selected from the group consisting of angioedema, urticaria, sudden infant death syndrome and bronchopulmonary aspergillosis. Further, the COMPOUNDS OF THE INVENTION can be used to treat systemic mastocytosis, particularly mastocytoma. The latter is a malignant disease with extensive metastasis, especially in dogs. Thus, the COMPOUNDS OF THE INVENTION are particularly useful for the preparation of a medicament for the treatment of canine mastocytic neoplasms.

COMPOUNDS OF THE INVENTION are generically and specifically described in patent applications EP 0 564 409 A1 and WO 99/03854, in particular in the claims relating to the compounds and end products of the Examples, the nature of the end products, pharmaceutical compositions and claims is hereby incorporated by reference into these applications. publications. Also included are the respective stereoisomers as well as the corresponding polymorphs, e.g., crystal modifications described herein.

In EP 0 564 409 A1, COMPOUNDS OF THE INVENTION are described as being useful for the treatment of cancer, thrombosis, psoriasis, fibrosis, dermatosclerosis and arteriosclerosis. In accordance with the present invention, it has now been found that the COMPOUNDS OF THE INVENTION surprisingly exhibit beneficial effects on allergic rhinitis, allergic dermatitis, drug allergy or food allergy, angioedema, urticaria, sudden death syndrome · «♦♦ ··

of the child, bronchopulmonary aspergillosis, multiple sclerosis, and mastocytosis, especially canine mastocytic neoplasms.

In accordance with particular findings of the invention, the present invention also provides a method of treating a warm-blooded animal, including a human, comprising administering to said warm-blooded animal, preferably a human or dog, most preferably a human suffering from one of the diseases described herein.

The invention also relates to a method of administering to a dog with canine mastocytic neoplasms a compound of the invention or a pharmaceutically acceptable salt thereof, which comprises administering to the dog once a day for a period preferably exceeding 1 month, 2 months or even 3 months. In particular, the invention relates to such a method in which a daily dose of approximately 20 to 200 mg, preferably 80 to 160 mg, in particular 125 mg of SOLI I is administered.

Preferably, the invention relates to a use or process wherein the daily dose of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino monomethanesulfonate salt is present. The compound of formula I is administered to a dog and said daily dose comprises an amount of said monomethanesulfonate salt sufficient to maintain plasma levels of at least 0.2 µmol.

The term method of treatment as used herein also refers in particular to a method of preventing the diseases mentioned herein, i. prophylactically administering to a healthy patient a pharmaceutical composition comprising the COMPOUND OF THE INVENTION to prevent outbreaks of the diseases mentioned herein.

The present invention also relates to a pharmaceutical composition for the treatment of at least one of the diseases mentioned herein comprising

glycol, contain

A COMPOUND OF THE INVENTION. Pharmaceutical compositions for treating at least one of the diseases described herein comprise an effective amount of the COMPOUNDS OF THE INVENTION together with pharmaceutically acceptable carriers that are suitable for topical, enteral, e.g. oral or rectal, or parenteral administration, and may be inorganic or organic, solid or liquid. For oral administration, in particular tablets or gelatin capsules containing the COMPOUNDS OF THE INVENTION are used together with diluents and / or lubricants, for example silicic acid, talc, stearic acid or salts thereof, and / or polyethylene, but also lotions, gels or creams. The tablets may be binders, starches, gelatin, methylcellulose, sodium carboxymethylcellulose and / or polyvinylpyrrolidone, disintegrants and / or effervescent mixtures, or adsorbents, colorants, flavors and sweeteners. The compounds of the invention may also be used in the form of parenterally administrable compositions or in the form of infusion solutions. Topical administration is, for example, administration to the skin. Another form of topical administration is to the eye, e.g., for the treatment of spring conjunctivitis. The pharmaceutical compositions may be sterilized and / or may contain adjuvants such as preservatives, stabilizing emulsifying agents, solubilizing osmotic pressure and / or buffers.

The present pharmaceutical compositions are prepared in a manner known per se, for example by means of conventional mixing, granulating, coating, dissolving or lyophilizing, and comprise from about 1% to 100%, in particular from about 1% to about 20%, of the active substance (s). ).

agents, agents, humectants and / or salts for control

For example, the compounds of the invention may be formulated as described in Examples 4 and 6 of WO 99/03854.

The dosage range of the COMPOUNDS OF THE INVENTION used depends on factors known to those skilled in the art, including the species of warm-blooded animal, body weight and age, route of administration, the particular compound employed and the disease being treated. If

Unless otherwise stated herein, the COMPOUNDS OF THE INVENTION are administered preferably one to four times daily or as soon as an allergic reaction is observed. Further, the COMPOUNDS OF THE INVENTION, in particular N- {5- [4- (4-methylpiperazinomethyl) benzoylamido] -2-methylphenyl} -4- (3-pyridyl) -2-pyrimidineamine (STI571B), preferably administered to a warm-blooded animal, especially a human, at a dose ranging from about 10 to 750 mg / day, preferably 30 to 600 mg / day, more preferably 30 to 300 mg / day.

In dogs, a warm-blooded animal weighing about 5 kg of an effective dose, for example a daily dose of about 20 to 200 mg, preferably 80 to 160 mg, is administered, depending on the species, age, particular condition, route of administration and the clinical picture. 125 mg. In adult dogs weighing approximately 5 kg with unresectable and / or metastatic malignant canine mast cells, a starting dose of 125 mg per day may be recommended. In dogs with inadequate responses after evaluating the response to 125 mg therapy per day, dose escalation can be safely considered and dogs can be treated as long as they benefit from the therapy and in the absence of limiting toxicities. Doses can be titrated to achieve plasma levels of at least 0.2 µmol (micromole), preferably at least 0.5 µmol, more preferably at least 1 µmol. Achieving and / or maintaining plasma levels of about 1 µmol is particularly preferred.

Claims (10)

PATENT CLAIMS Use of an N-phenyl-2-pyridinamine derivative of the general formula I in which R 1 represents a 4-pyrazinyl group; 1-methyl-1H-pyrrolyl; an amino- or amino-lower alkyl-substituted phenyl group wherein the amino group is in each case free, alkylated or acylated; 1H-indolyl or nebo-imidazolyl attached to a carbon atom of a five-membered ring; or unsubstituted or lower alkyl substituted pyridyl group attached to a ring carbon atom and unsubstituted or substituted on nitrogen by an oxygen atom; R ₂ and R ₃ are each independently of the other hydrogen or lower alkyl; one or two of R ₄ , R ₅ , R ₆ , R ₆ , R ₆ and R ₈ are each nitro, fluoro-substituted lower alkoxy or a group of formula II -N (R ₉ ) -C (= X) - (Y) _n -R ₁₀ (II), wherein Rg represents a hydrogen atom or a lower alkyl group ·· · ♦♦ · X is oxo, thio, imino, N-lower alkyl-imino, hydroxyimino or O-lower alkyl-hydroxyimino, Y is oxygen or NH, n is 0 or 1 and R ₁₀ represents an aliphatic group containing at least 5 carbon atoms or an aromatic, aromatic-aliphatic, cycloaliphatic, cycloaliphatic-aliphatic, heterocyclic or heterocyclic-aliphatic group, and the remaining substituents R ₄ , R ₅ , R ₆ , R 7 and R ₈ each independently of one another hydrogen, lower alkyl which is unsubstituted or substituted by free or alkylated amino, piperazinyl, piperidinyl, pyrrolidinyl or morpholinyl, or lower alkanoyl, trifluoromethyl, free, etherified or esterified hydroxy, free, alkylated or acylated amino, or free or esterified carbox or a salt of such a compound containing at least one salt-forming group, for the preparation of a medicament for the treatment of allergic rhinitis, allergic dermatitis, drug allergy or food allergy, angioedema, urticaria, sudden infant death syndrome, bronchopulmonary aspergillosis, mastocytosis or multiple sclerosis. Use according to claim 1 of a compound of formula I, wherein one or two of R ₄ , R ₅ , R ₆ , R ₇ and R ₈ are each nitro or a group of formula II, wherein: R 8 represents a hydrogen atom or a lower alkyl group, X is oxo, thio, imino, N-lower alkyl-imino, hydroxyimino or O-lower alkyl-hydroxyimino, Y is oxygen or NH, n is 0 or 1 and R ₁₀ represents an aliphatic group containing at least 5 carbon atoms or an aromatic, aromatic-aliphatic, cycloaliphatic, cycloaliphatic-aliphatic, heterocyclic or heterocyclic-aliphatic group, and the remaining substituents R ₄ , R ₅ , R ₆ , R ₇ and R _g are each independently hydrogen, lower alkyl a group which is unsubstituted or substituted by a free or alkylated amino, piperazinyl, piperidinyl, pyrrolidinyl or morpholinyl group, or a lower alkanoyl, trifluoromethyl, free, etherified or esterified hydroxy group, a free, alkylated or acylated amino group or a free or esterified group , and the remaining substituents have the meanings defined in claim 1, or a pharmaceutically acceptable salt of such a compound that comprises at least one salt-forming group. Use according to claim 1 of a compound of the general formula wherein I, ve 99 9 * 9 99 99 9 9 9 99 ** 9 9 9 9 <♦ 9 9 «· 9 R 1 represents a pyridyl group attached to a carbon atom, R 2, R 3, R 5, R 5 and R 5 are each hydrogen, R ₄ represents a lower alkyl group, R ₇ represents a group of formula II wherein Rg represents a hydrogen atom, X is oxo, n is 0 and R 10 represents a 4-methyl-piperazinyl-methyl group, or a pharmaceutically acceptable salt thereof. Use of a compound of formula I according to claim 1, which compound is N- {5- [4- (4-methyl-piperazinomethyl) benzoylamido] -2-methylphenyl} -4- (3-pyridyl) -2-pyrimidine -amine or a pharmaceutically acceptable salt thereof. Use of a compound of formula I according to any one of claims 1 to 4, wherein the compound is used in the form of its monomesylate salt. Use of a compound of formula I according to any one of claims 1 to 5 for the preparation of a medicament for the treatment of canine neoplasms from mast cells. A method of treating a warm-blooded animal with allergic rhinitis, allergic dermatitis, drug allergy or food allergy, angioedema, urticaria, sudden infant death syndrome, bronchopulmonary aspergillosis, mastocytosis, or multiple sclerosis, comprising administering a compound The compound of formula (I) according to any one of claims 1 to 5 to an animal in an amount that is therapeutically effective against at least one of said diseases, wherein the compound of formula (I) may also be in the form of a pharmaceutically acceptable salt. A method of treating dogs suffering from canine mast cell neoplasms comprising administering to said dog a dose effective against canine mast cell neoplasms of a compound of formula (I) according to any one of claims 1 to 5. The use or method of claim 6 or 8, wherein the adult dog is administered a daily dose of 20 to 200 mg 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide or a pharmaceutically acceptable salt thereof. The use or method of claim 1 or 7, wherein the human is administered a daily dose of 30 to 400 mg 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide or a pharmaceutically acceptable salt thereof.