MXPA00000620A - Crystal modification of a n-phenyl-2-pyrimidineamine derivative, processes for its manufacture and its use - Google Patents

Abstract

The invention relates to a new crystalline form of the methanesulfonic acid addition salt of 4-(4-methylpiperazin-1-ylmethyl)- N-[4-methyl-3-(4-pyridin-3-yl)pyrimidin-2-ylamino) phenyl]-benzamide of formula (I), which may be used for example for tumour therapy.

Description

MODIFICATION OF THE GLASS OF A N-PHENYL-2- PYRIMIDINAMINE DERIVATIVE. PROCESSES FOR ITS MANUFACTURE, AND ITS USE The invention relates to a particular form of the methanesulfonic acid addition salt of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) irimidin-2 -ylamino) phenyl] benzamide, which comprises certain crystals, to processes for their preparation, to pharmaceutical compositions containing this crystal form, and to their use in diagnostic methods or preferably for the therapeutic treatment of warm-blooded animals, especially human beings, or their use for the preparation of pharmaceutical preparations for use in diagnostic methods or preferably for the therapeutic treatment of warm-blooded animals, especially humans.

Background of the Invention The preparation of 4- (4-methyl-piperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) -pyrimidin-2-ylamino) -phenyl] -benzamide and its use, especially as an antitumor agent, they are described in Example 21 of European Patent Number EP-A-0, 564, 409, which was published on October 6, 1993, and in equivalent applications in numerous other countries. This compound is exemplified in these publications only in free form (river as a salt). Surprisingly, it has now been discovered that, under certain conditions, a crystal form can be found in the methanesulfonate salt of this compound, which is described hereinafter as a β-crystal form, and which has very convenient properties.

Detailed Description of the Invention The invention is described in greater detail in the following, with the help of the drawings and other auxiliaries: Description of the Drawings Figure 1/3 shows the X-ray diffraction diagram of the crystal form a of the methanesulfonic acid addition salt of a compound of the formula I. Figure 2/3 shows the diffraction diagram of X-rays of the ß-crystal form of the methanesulfonic acid addition salt of a compound of the formula I. Figure 3/3 shows the crystals above the crystal form a, and below the β-crystal form, of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide methanesulfonate (= of the acid addition salt methanesulfonic acid of a compound of the formula I). In both X-ray diagrams, the refractive angle 2 teta is plotted on the horizontal axis (x axis), and the relative line intensity (peak intensity with corrected background) on the vertical (y axis). The diagrams are obtained as follows: first, the X-ray diffraction pattern is recorded on film using a Guinier camera (model Enraf-Nonius FR 552) with a Guinier film 258 -94c and copper radiation (Kal, wavelength? = 1.54060 Angstroms). The optical density of the lines in the film is proportional to the intensity of the light. The film is then scanned using a line scanner (LS 18, Johansson, T by, Sweden) with SCANPI software. According to Figure 2/3, there are lines that have a relative line strength of 20 or more at the following 2 tera refraction angles (the relative line strengths are given in parentheses): 9.7 ° (40), 13.9 ° (26), 14.7 ° (23), 17.5 ° (57), 18.2 ° (90), 20.0 ° (65), 20.6 ° (76), 21.1 ° (100), 22.1 ° (89), 22.7 ° (38 ), 23.8 ° (44), 29.8 ° (23), and 30.8 ° (20). The fact that in Figure 2/3 the line's relative line strength at 30.8 ° appears to be higher than that of the line at 29.8 °, is due to a closure by the additional line at 31.0 ° which has an intensity of relative line of 13. The melting points are determined by means of a differential scanning calorimetry thermogram, using a Mettler-Toledo TA8000. The DSC ("differential scanning calorimetry") is the technique of dynamic differential calorimetry. Using this technique, the melting temperature of both the crystal form a and the crystal form β can be measured by heating the samples until a thermal reaction is detected, that is to say an endothermic or exothermic reaction, by means of ultrasensitive sensors. The melting points indicated in this text are determined using a Mettler-Toledo TA8000 apparatus, measuring from about 5.5 to 6.5 milligrams of each sample in an aluminum crucible with a perforated lid under a calm atmosphere of air at a heating rate of 10. ° C / minute (starting at 20 ° C). The crystal form of the 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide methanesulfonate is characterized by crystals in the form of a needle, and it is hygroscopic. In this form, the crystals are not particularly suitable for a pharmaceutical formulation as solid dosage forms, due to their physical properties, for example, their flow characteristics, which are unfavorable. However, under certain conditions, it is possible to obtain 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] methanesulfonate] benzamide in a crystal shape that does not have the shape of needles. This form is described in the present text as the crystal form ß. The β-crystal form of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide methanesulfonate has the advantage that their flow properties are substantially more favorable than those of the crystal form. This crystal form has the additional advantage of being thermodynamically more stable at temperatures below 140 ° C. Finally, the crystal form β is less hygroscopic than the crystal form α, and therefore, it is also better stored and easier to process. The invention relates to an acid addition salt of a compound of the formula I comprising non-needle-shaped crystals, especially the β-crystal form of the methanesulfonic acid addition salt of the compound of the formula I. The invention relates especially to a particularly pure form of particular crystal, preferably that which is hereinafter referred to as the β-crystal form, of the methanesulfonic acid addition salt of 4- (4-methylpiperazine-1-methanesulfonate) -ylmethyl) -N- [4-methyl-3- (4-pyrid-3-yl) pyrimidin-2-ylamino) phenyl] benzamide of the formula I: Where the term methanesulfonic acid salt of a compound of the formula I or of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2 -ylamino) phenyl] benzamide is used hereinbefore and hereinafter, to mean especially the methanesulfonic acid salt of formula II: The term "essentially pure" is understood, in the context of the present invention, to mean especially that at least 90, preferably at least 95, and more preferably, at least 99 percent by weight of the crystals of a Acid addition salt of the formula I are present in the crystal form according to the invention, especially the β-crystal form. In the context which mentions that the acid addition salt of formula II exhibits an X-ray diffraction pattern essentially as in Figure 2/3, the term "essentially" means that at least the major lines of the diagram illustrated in Figure 2/3, that is, those that have a relative line intensity of more than 10 percent, especially more than 20 percent, compared to the most intense line in the diagram. The invention also expressly refers to those forms of the methanesulfonic acid addition salt of a compound of the formula I, wherein the crystals of the crystal form d according to the invention, especially the β-crystal form, are present in a essentially pure form, together with other crystal forms, and / or the amorphous form of the methanesulphonate of 4- (4-methyl-piperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridine 3- il) pyrimidin-2-ylamino) phenyl] benzamide. However, it prefers the acid addition salt of formula II, which is present in an essentially pure form, in the form of ß-crystal. The new crystal form, especially the d-crystal form, has the following properties: The melting point in the differential scanning calorimetry thermogram of the crystal form 8 is 217 ° C, that of the crystal form or; it is 226 ° C (beginning of the merger) The X-ray diffraction pattern of the crystal form ß does not show the peak of the crystal form a. marked co (1), and only to a very minor degree, shows the (3) mark (see Figures 1/3 and 2/3). In contrast, Figure 2/3 shows a new additional peak marked with (4). The new peak marked with (5) also appears in Figure 2/3. The X-ray diffraction diagrams also show other notable differences. In the preferred embodiment, the essentially pure methanesulfonic acid addition salt of a compound of formula I in the β-crystal form shows the X-ray diffraction pattern indicated in Figure 2/3. (i) A crystal form of the methanesulfonic acid addition salt of a compound of the formula I which does not show the peak marked with (1) in Figure 1/3 of the X-ray diffraction pattern is preferred. crystal form preferably present in an essentially pure form. (ii) A crystal form of the methanesulfonic acid addition salt of a compound of the formula II, which remains dry at 93 percent relative humidity, and at a temperature of 25 ° C, is also preferred. crystal preferably present in an essentially pure form. (iii) The invention relates preferably to the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, which is characterized by the presence of crystals exhibiting the form shown in Figure 3/3. later; especially the crystal form ß, in an essentially pure form. (iv) A greater preference is given for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, having a melting point of less than 225 ° C, especially between 217 ° C and 225 ° C. (v) A greater preference is also given for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, which has a melting point of less than 217 ° C, defined as the start of the fusion in the differential scanning calorimetry thermogram. (vi) A greater preference is also given for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I which, in the X-ray diffraction, shows the peak marked with (4) in the Figure 2/3. (vii) A greater preference is also given for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I which, in the X-ray diffraction, shows the peak marked with (5) in the Figure 2/3. (viii) There is still a greater preference for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, which shows an X-ray diffraction pattern of the type shown in Figure 2/3, especially one in which the relative peak intensities of each peak do not deviate by more than 10 percent from the relative peak intensities of the diagram shown in Figure 2/3, especially an X-ray diffraction pattern identical to that shown in the Figure 2/3. (ix) A greater preference is given for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, which has two of the properties mentioned in paragraphs (i) to (viii), the greater preference for three of the properties of these paragraphs, especially all the properties, and more especially those properties defined as preferred. In the same way, a crystal form as defined in one of paragraphs (i) to (ix) is more preferably in an essentially pure form. A particularly special preference is given for the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, which can be obtained as described in the examples. In all cases, a form of the methanesulfonic acid addition salt of a compound of the formula I, which comprises the aforementioned corresponding crystal form, is also to be understood in a broader aspect of the invention. The crystal form β (preferably essentially pure) can be obtained by the following: (a) digesting another form of crystal, especially the crystal form, or an amorphous starting material of the methanesulfonic acid addition salt of a compound of the formula I, with a suitable polar solvent, especially an alcohol, more especially methanol, or also a ketone (especially in a mixture with water, for example water / acetone), usually acetone, an N, N-lower dialkyl-alkane lower carboxamide, usually N, N-dimethyl formamide or acetamide, or a hydrophilic ether, usually dioxane, preferably in the presence of some water, or mixtures thereof, in suspension at a suitable temperature, preferably at a temperature between 20 ° C and 50 ° C, for example at about 25 ° C, or (b) dissolving another form of crystal, especially the crystal form, or an amorphous starting material of the methanesulfonic acid addition salt of a compound of the formula I, with a suitable polar solvent, such as especially an alcohol, usually methanol or ethanol, a ketone (especially in a mixture with water, for example water / acetone), usually acetone, an N, N- lower dialkyl-lower alkane-carboxamide, usually formamide or N, N-dimethyl acetamide, or a hydrophilic ether, usually dioxane, or mixtures thereof, preferably in the presence of some water, at a suitable temperature, especially after heat the solvent, or while heating during the dissolution process, in both cases preferably at 25 ° C and up to the reflux temperature of the reaction mixture, and then crystallization is initiated by the addition of a small amount of the ß crystal form as seed crystal at a suitable temperature, for example, between 0 ° C and 70 ° C, preferably between 20 ° C and 70 ° C. The educto, the crystal form of the methanesulfonic acid addition salt of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide, can be obtained, for example, by precipitation of the salt from a solution in a solvent other than an alcohol, such as methanol, and without adding a seed crystal of the β-crystal form. The above conditions on the selective preparation of the individual crystal forms are not conclusive. In general, for example, it is possible to vary the parameters, such as the weight ratio of the methanesulfonic acid addition salt of a compound of the formula I to the solvent. It is also possible to vary the time necessary for the preparation of the crystal form β, especially when the temperatures are adjusted at the same time. One of the advantages of the ß-crystal form is especially its more compact crystal form, which results in substantially more beneficial flow properties, and consequently, better processability of the methanesulfonic acid addition salt of a compound of the Formula I in the form of crystal ß against the crystal form OI, for example in the manufacture of pharmaceutical preparations. It is true to say that the crystal form a of the methanesulfonic acid addition salt of a compound of the formula I is metastable at room temperature. However, the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I is the thermodynamically stable form at room temperature. Therefore, greater stability should be expected. Finally, the crystal form ß is less hygroscopic than the crystal form o, of the addition salt of methanesulphonic acid of a compound of the formula I, as can be shown by the following table: When measuring the crystal forms to the point where the equilibrium is reached (no more adsorption) in a climatic chamber of glass at 25 ° C, and with the humidities shown below, the following water content values are found (the percentage values for the final water content refers to dry weight): Relative humidity Final water content on adsorption (%) crystal shape to crystal form ß (%) (molar) (%) (molar) 12 0.14 0.05 0.08 0.02 33 0.18 0.06 0.10 0.03 46 0.14 0.05 - - 54 0.13 0.04 0.14 0.05 66 0.07 0.02 0.09 0.03 75 0.49 0.16 - - 85 0.18 0.06 0.16 0.05 93 40 13.1 0.15 0.05 97 63 20.8 23 7.5 100 - - 37 12 It is shown that, at 25 ° C, the crystal form: is hygroscopic, and rapidly recovers water, such that, with a relative humidity of 93 percent, the sample is present to some degree in an amorphous form, while that the ß-crystal form remains dry under these conditions. Both crystal forms liquefy at 37 percent relative humidity, but this happens much more quickly with the crystal form than with the ß crystal form. The lower hygroscopicity is an additional advantage for the processing and storage of the acid addition salt in the ß crystal form. The methanesulfonic acid addition salt of a compound of the formula I, which is preferably used in the form of β-crystal (hereinafter, the addition salt of methanesulfonic acid always means the β-crystal form), as well as 4- (4-methyl-piperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) -pyrimidin-2-ylamino) -phenyl] -benzamide in free form, possesses valuable pharmacological properties, and , for example, it can be used as an antitumor agent, as an agent to treat atherosclerosis, as an agent to treat restenosis, for the prevention of disorders induced by transplants, such as bronchiolitis obliterative, and / or to prevent the invasion of cells from a warm-blooded animal by certain bacteria, such as Porphyromonas gingivalis. The phosphorylation of proteins has been known for a long time as an essential step in the differentiation and division of cells. Phosphorylation is catalyzed by protein kinases subdivided into serine, threonine and tyrosine kinases. Tyrosine kinases include the tyrosine kinase of the PDGF (Platelet Derived Growth Factor) receptor. PDGF (Platelet Derived Growth Factor) is a growth factor that occurs very commonly, which has an important role in both normal growth and pathological cell proliferation, as seen in carcinogenesis, and in diseases of the smooth muscle cells of the blood vessels, for example in atherosclerosis and thrombosis. Inhibition of receptor tyrosine kinase activity stimulated by platelet-derived growth factor in vitro is measured in immune complexes of the platelet-derived growth factor receptor of BALB / c 3T3 cells, as described by E. Andrejauskas-Buchdunger and U. Regenass in Cancer Research 52 .. 5353-5358 (1992). A compound of the formula I, described in greater detail hereinabove, such as especially its β-crystal form, inhibits the phosphorylation of the acellular receptor dependent on the Platelet-Derived Growth Factor. The inhibition of the tyrosine kinase of the platelet-derived growth factor receptor is measured in an ELISA microtiter assay (see Trinks et al., J. Med. Chem. 221, 1015-27 (1994)). 4- (4-Methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide and the corresponding methanesulfonate salt, inhibit the tyrosine kinase activity of the Platelet-Derived Growth Factor receptor in an IC 50 (concentration where activity is inhibited by 50 percent, compared to the control) of approximately 120 nM and approximately 100 nM, respectively. Inhibition of Platelet Derived Growth Factor makes a compound of formula I also suitable for the treatment of tumoral diseases, such as gliomas, sarcomas, prostate tumors, and colon tumors, of the chest, and of the ovary. The methanesulfonic acid addition salt of a compound of the formula I also inhibits cell processes involving the so-called stem cell factor (SCF, also known as c-box ligand or steel factor), such as the autophosphorylation of the (cassette) stem cell factor receptor, and stem cell factor-stimulated activation of the MAPK kinase (mitogen-activated protein kinase). The methanesulfonic acid addition salt of a compound of the formula I, such as especially the β-crystal form thereof, consequently also inhibits the autophosphorylation of the stem cell factor receptor (and the c-kit, a proto- oncogene). The M07e cells are a human promegakaryotic leukemia cell line that depends on the stem cell factor for proliferation.These are obtained from Grover Bagby, Oregon Health Sciences University, USA Cells are grown in RPMI 1649 medium supplemented with serum fetal calf 10, and 2.5 nanograms / milliliter of GC-CMF GM-SCF and stem cell factor are commercially available.M07e cells deprived of serum are prepared and incubated for 90 minutes at 37 ° C with the test, before being stimulated with recombinant stem cell factor for 10 minutes at 37 ° C. Identical amounts of cell lysates are analyzed by Western Blot, using anti-phosphotyrosine antibodies (Buchdunger et al., Proc. Nati. Acad. Sci (EUA) 92, 2558-62 (1995).) Immunolabelled proteins are detected by means of the Western Blot ECL system from Amersham (Amersham, United Kingdom) A compound of the formula I, especially the The crystal structure of the methanesulfonate salt of the formula II inhibits the autophosphorylation of SCF-R on the micromolar scale. Based on the described properties, the methanesulfonic acid addition salt of a compound of the formula I, such as especially the β-crystal form thereof, can be used not only as a tumor inhibitory substance, for example in cancer of small cell lung, but also as an agent to treat non-malignant proliferative disorders, such as atherosclerosis, thrombosis, psoriasis, scleroderma, and fibrosis, as well as for the protection of stem cells, for example to combat the hemotoxic effect of chemotherapeutic agents, such as 5-fluorouracil, and in asthma. It can be used especially for the treatment of diseases that respond to an inhibition of the platelet-derived growth factor receptor kinase. In addition, the methanesulfonic acid addition salt of a compound of the formula I, such as especially its β-C crystal form, prevents the development of multidrug resistance in cancer therapy with other chemotherapeutic agents, or eliminates a resistance previously existing to other chemotherapeutic agents. Also, independently of the effect described hereinabove, the methanesulfonic acid addition salt of a compound of formula I, such as especially the β-crystal form thereof, can be used advantageously in combination with other antitumor agents. Also, the abl kinase, especially v-abl, is inhibited by 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) irimidin-2-ylamino ) phenyl] benzamide and its methanesulfonate salt. Inhibition of v-abl tyrosine kinase is determined by the methods of N. Lydon et al., Oncogene Research 5_, 161-173 (1990), and J.F. Geissler et al., Cancer Research 52., 4492-8 (1992). In these methods, [Val5] -angiotensin II and [? "32P] -ATP are used as substrates.Here, 4- (4-methyl-piperazin-1-ylmethyl) -N- [4-methyl-3-] (4-pyridin-3-yl) -pyrimidin-2-ylamino) phenyl] benzamide shows an IC50 of 38 nM By analogy, the salt of a compound of the formula I also inhibits the BCR-abl kinase (see Nature Medicine 2, 561-566 (1996)), and therefore, is suitable for the treatment of BCR-abl positive cancer, and tumor diseases, such as leukemia (especially chronic myeloid leukemia and acute lymphoblastic leukemia, wherein especially apoptotic action), and also shows effects on the subgroup of leukemic stem cells, as well as a potential for the purification of these cells in vi tro after removing said cells (for example, removal of the bone marrow), and of the reimplantation of the cells once they have been cleared of the cancer cells (for example, reimplantation of purified bone marrow cells). In addition, the methanesulfonic acid addition salt of a compound of the formula I, shows useful effects in the treatment of disorders that occur as a result of transplantation, for example allogeneic transplantation, especially tissue rejection, such as especially bronchiolitis oblitera -tiva (OB), that is, a chronic rejection of allogeneic lung transplants. In contrast to patients without obliterative bronchiolitis, those with obliterative bronchiolitis frequently show a high concentration of platelet-derived growth factor in the bronchoalveolar lavage fluids. If 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide methanesulfonate is administered, especially in the form of ß crystal, to rats with allogeneic tracheal transplants, for example in a dose of 50 milligrams / kilogram intraperitoneally, it can be shown, after removing 10 transplants per group after 10 and 30 days for morphometric analysis, possible epithelial lesions and occlusion of the airways, and the investigation to determine the immunohistochemical trajectories of action demonstrates that, although the methanesulfonic acid addition salt of a compound of formula I does not have a significant effect on epithelial necrosis or infiltration by inflammatory cells, it does reduce notably fibroproliferation and occlusion of the lumen compared to controls. It is possible to have synergistic effects with other immunomodulatory or anti-inflating substances, for example, when used in combination with cyclosporin, rapamycin, or ascomycin, or immunosuppressive analogs thereof, for example cyclosporine A (CsA), cyclosporin G, FK-506, rapamycin, or comparable compounds; corticosteroids; cyclophosphamide; azathioprine; methotrexate; brewing leflunomide; mizoribin; mycophenolic acid; mycophenolate mofetil; 15-deoxyspergualin; immunosuppressant antibodies, especially monoclonal antibodies to leukocyte receptors, for example MHC, CD2, CD3, CD4, CD7, CD25, CD28, B7, CD45, CD58 or their ligands; or other immunomodulatory compounds, such as CTLA41g. If CsA (1 milligram / kilogram subcutaneously) is combined, for example, with the acid addition salt of formula I (50 milligrams / kilogram), synergism can be observed. The methanesulfonic acid addition salt of a compound of the formula I is also effective in diseases associated with the migration and proliferation of vascular smooth muscle cells (where PDGF and PDGF-R often also play a role), such as restenosis and atherosclerosis. These effects and their consequences for the proliferation or migration of smooth muscle vascular cells in vi tro in vivo, can be demonstrated by administration of the methanesulfonic acid addition salt of a compound of the formula I, or also by research of its effect on the thickening of the vascular intima followed by mechanical injury in vivo. The methanesulfonic acid addition salt of a compound of the formula I is used in 0. IN HCl or in dimethyl sulfoxide, at a concentration of 10 M for the in vi tro studies. The delivery solution is further diluted with the cell culture medium, and used in concentrations of 10 to 0.1 μM for the experiments. For in vivo administration, the methanesulfonic acid addition salt of a compound of formula I is dissolved, for example, in dimethyl sulfoxide, at a concentration of 200 milligrams / milliliter, and then diluted 1:20 with Tween to 1 percent in a 0.9 percent saline solution. After sonication, a transparent solution is obtained. The supply solutions are prepared fresh every day before administration. (The compound of formula I can also be dissolved simply in deionized water for oral administration, or in a 0.9 percent saline solution for parenteral administration). The administration is done 24 hours before the operation. The methanesulfonic acid addition salt of a compound of formula I is administered to rats in a dose of 50 milligrams / intraperitoneally per day throughout the observation period. The control rats are given the same dose of substrate. Oral administration is also possible. The primary cultures of smooth muscle aortic cells are isolated from rat aorta DA (AG-B4, RTla) from 9 a 11 days of age, using a modification of the method described by Thyberg et al. (See Differentiation 25., 156-67). (1983)). The aorta is opened by means of a longitudinal incision, and the endothelium is carefully removed. The adventitia and the tunica media are separated, and the tunica media is digested with 0.1% collagenase and DNAse in physiological phosphate-regulated serum for 30 minutes at 37 ° C. The cells are centrifuged, suspended in the culture medium, and then allowed to grow in plastic bottles. The primary cells are used for the experiments after steps 2 to 6. The subcultures are maintained in DMEM (Eagle's Medium Modified by Dulbecco), supplemented with fetal calf serum at 10 percent, 2 mmol / ml glutamine, 100 millimoles / milliliter of streptomycin, and 100 international units / milliliter of penicillin. For identification purposes, cells are allowed to grow on glass slides, and stained on SMC-a actin (see below). The migration of smooth muscle cells is quantified in vi tro using a Transwell cell culture insert (Costar, Cambridge, MA) whose upper and lower compartments are separated by a polycarbonate membrane with a pore size of 8 microns. The cells (100 microliters at a concentration of 1 million cells / milliliter) are exposed in the upper compartment. After 2 hours, 60 nanograms / milliliter of PDGF-BB or PDGF-AA (Upstate Biotechnology Inc., Lake Placid, NY) are added to the lower compartment, supplemented with 0.5 percent fetal calf serum and bovine serum albumin. 0.1 percent, and the test compound is added in concentrations of 3, 1, 0.3, 0.1, 0.03, 0.01, and 0.003 μM. To measure fibronectin-dependent migration, Transwell chambers are covered with fibronectin at a concentration of 10 micrograms / milliliter for 24 hours at 4 ° C (human cell fibronectin, Upstate Biotechnology Inc.). After a 24-hour migration, the filters are removed, fixed in methanol, and stained with Mayer's hematoxylin and eosin. The cells migrated on the underside of the filter membrane are determined by counting the sectional fields specified on the filters, with the help of a light microscope, with a magnification of 400x. The inhibition of emigration is. quantifies in terms of the percentage of cells against control. To exclude the possibility of a toxic effect, the viability of the cells is tested by the incorporation of 3H-thymidine in DMEM, supplemented with 10 percent fetal calf serum. An inhibition of migration induced by PDGF-AA and especially by PDGF-BB is observed. Experimental animals: the aorta and carotid artery of male Wistar rats is separated (acquired at the Laboratory Animal Center of the University of Helsinki, Finland). The rats are anesthetized with 240 milligrams / kilogram of chloral hydrate intraperitoneally. Buprenorphine is given (Temgesic, Reckitt &Coleman, Hull, United Kingdom) for preoperative and postoperative pain relief. All animals are given human care, keeping with the "Principles of Laboratory Animal Care "and NIH's" Guide for the Care and Use of Laboratory Animals "(NIH, Publication 86-23, revised 1985). Rats weighing 200 to 300 grams were used for the separation procedure The left common carotid artery is separated from the endothelium through the intralu-minal passage of a 2F embolectomy catheter (Baxter Healthcare Corporation, Santa Ana, CA, 27) .To remove the endothelium, the catheter is passed through the lumen three. Sometimes, inflated with 0.2 milliliters of air, the external carotid is ligated after removing the catheter, and the wound is closed.The histological changes are evaluated by referring to the sections of the carotid media 4 days after separation. Thoracic aorta of the endothelium using a Fogarty 2F arterial embolectomy catheter The catheter is inserted into the thoracic aorta through the left iliac artery, it is inflated with 0.2 milliliters of air, and it is passed through the lumen five times to remove the endothelium. Then the iliac artery is ligated. Three times (3, 7, and 14 days) are selected for the evaluation of histological changes. To quantify the proliferating cells, three different methods are used to label the cells with bromodeoxyuridine (BrdU) after separating the carotid from the rat. In this model, the average cell proliferation begins 24 hours after separation; intimal cells appear first after 72 to 96 hours. To quantify the proliferation of smooth muscle cells prior to the appearance of cells in the intima, a labeling reagent of 0.1 milliliter BrdU (ZYMΔD, San Francisco, CA) is administered intravenously during the postoperative period from 0 to 72 hours after separation (in total, 0.1 milliliters 6 times). To quantify the proliferation during the initial migration wave, the rats were given 3 x 0.1 milliliters of BrdU labeling reagent at 8 hour intervals for a period of 72 to 96 hours after the operation. To quantify proliferation at the end of the initial wave of emigration, a third group of rats was given a pulse dose of 0.3 milliliters of BrdU 3 hours before slaughter. Histological samples are fixed in a 3 percent paraformaldehyde solution for 4 hours to be embedded in paraffin. The morphological changes are evaluated from paraffin sections stained with Mayer's hematoxylin-eosin. The cell counts of different vessel sections are calculated at a magnification of 400x. To identify the cells of the culture and the cells that appear in the neo-intima within 4 days of the separation lesion, an immunohistochemical staining of samples fixed in acetone is performed, using an anti-a-actin antibody, obtained from cells of smooth muscle (Bio-Makor, Rehovot, Israel). The primary smooth muscle cells are identified on glass slides fixed with acetone, using the same dyeing method. The sections are incubated with the primary antibody (dilution of 1: 2000), washed, and incubated consecutively with rabbit Ig against mouse conjugated with peroxidase and goat Ig against rabbit, followed by a treatment with a substrate solution with the 3-amino-9-ethylcarbazole chromogen and hydrogen peroxide. BrdU stains are prepared from paraffin sections using a primary mouse antibody (Bu20a, Dako, A / S, Denmark) and the Vectastain Elite ABC-Kit (Vector Laboratories, Burliname, CA). The sections are deparaffinized and treated with microwaves at 500 W (2 x 5 minutes in a citrate regulator 0.1M, pH of 6), followed by a treatment with 95 percent formamide in 0.15M trisodium citrate for 45 minutes at 70 ° C. Antibody dilutions are prepared according to the manufacturer's specifications. The sections are counter-stained with Mayer's hematoxylin and eosin, and the positive cells are counted separately for the intima, media, and adventitia. In the carotid of treated animals, a significant decrease in cell count is found for smooth muscle cells. The adventitia and the average showed a significant reduction in the cell count. As a result of the methanesulfonic acid addition salt of a compound of the formula I, there is a slight decrease in the absolute number of BrdU-labeled cells in the intima, media, and adventitia, during the first two marking periods (0 to 72 hours and 72 to 96 hours), and after 93 to 96 hours, there is a decrease in the number of marked cells in all compartments. In the same way, decreases in the number of smooth muscle cells are found in the animals with the aorta separated. According to these findings, the methanesulfonic acid addition salt of a compound of formula I, therefore, can inhibit proliferation, and especially the migration of vascular smooth muscle cells. The methanesulfonic acid addition salt of a compound of the formula I, especially the β-crystal form, can also inhibit angiogenesis. This can be demonstrated as follows: a chamber containing agar (0.8%) and heparin (2 units / milliliter) with or without growth factor (VEGF 3 micrograms / milliliter, PDGF 1 microgram / milliliter, or bFGF) is implanted subcutaneously. 0.3 micrograms / milliliter) in normal mice (C57 BL / 6). The metansulphonic acid addition salt of a compound of the formula I is administered orally, in a dose showing good antitumor activity in a nude mouse xenotransplant model. The dosage is started one day before the implantation of the cameras. The cameras are removed after 5 days. Angiogenic efficacy is quantified by measuring both the vascularized tissue that has grown around the implant and the blood content of this tissue (external blood). Blood is determined by measuring hemoglobin. Although the vessels do not grow into the agar, the agar becomes intensely red if an anti-angiogenic effect is present. If a compound inhibits the increase in blood that is induced by the growth factor, this is seen as an indication that the compound in question is blocking the angiogenic effect of the growth factor concerned. The inhibition of weight but not of blood volume suggests an effect on the proliferation of fibroblasts.

A suppression of the control response suggests an inhibition of wound healing. In an oral dose of 50 milligrams / kilogram once a day, the compound of formula I inhibits the angiogenic effect of the three growth factors (VEGF, PDFG, bFGF). It remains without saying that all the inhibitory and pharmacological effects indicated, are also found with the free base, 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide, or other salts of it. The present invention relates especially to the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, in the treatment of one of these diseases, or in the preparation of a pharmacological agent for its treatment. The antiproliferative, especially antitumor, activity of the methanesulfonic acid addition salt of a compound of formula I in vivo, for example, is described for the treatment of abl-dependent tumors, in Nature Med. 2, 561-6 (1996 ). The invention also relates to a process for the treatment of warm-blooded animals suffering from these diseases, especially a tumor disease, wherein an amount of the β-crystal form of the methanesulfonic acid addition salt of a compound of the Formula I, which is effective against the disease concerned, especially an amount with an antiproliferative and especially tumor inhibitory efficacy, to warm-blooded animals in need of this treatment. The invention further relates to the use of the β-crystal form of the methanesulfonic acid addition salt of a compound of the formula I, for the inhibition of the aforementioned tyrosine kinases, especially the derived growth factor receptor kinase. of platelets, the v-abl kinase, and / or the c-receptor kinase of the kit-c, or for the preparation of pharmaceutical compositions for use in the treatment of the human or animal body, especially for the treatment of tumors, such as gliomas, ovarian tumors, prostate tumors, colon tumors, and lung tumors, such as especially small cell lung carcinoma, and breast tumors or other gynecological tumors. Depending on the species, the age, the individual condition, the mode of administration, and the clinical picture in question, effective doses are administered, for example daily doses of approximately 1 to 2500 milligrams, preferably 1 to 1000 milligrams, especially from 5 to 500 milligrams to warm-blooded animals of a body weight of approximately 70 kilograms. The invention also relates to pharmaceutical preparations containing an effective amount, especially an effective amount for the prevention or treatment of one of these diseases, of the methanesulphonic acid addition salt of a compound of the formula I in the form ß crystal, together with pharmaceutically acceptable carriers that are suitable for topical, enteral, for example oral or rectal, or parenteral administration, and can be inorganic or organic, and solid or liquid. For oral administration, tablets or gelatin capsules containing the active substance together with diluents, for example lactose, dextrose, sucrose, mannitol, sorbitol, cellulose, and / or glycerin, and / or lubricants, for example silica, are especially used for oral administration. , talc, stearic acid, or salts thereof, usually magnesium or calcium stearate, and / or polyethylene glycol. In the same way, the tablets may contain binders, for example, magnesium aluminum silicate, starches, usually corn starch, wheat starch, or rice starch, gelatin, methyl cellulose, sodium carboxymethyl cellulose, and / or polyvinyl pyrrolidone, and disintegrants are desired, for example, starches, agar, alginic acid, or a salt thereof, typically sodium alginate, and / or effervescent mixtures, or adsorbents, coloring agents, flavorings, and sweetening agents. The pharmacologically active compounds of the present invention can be further used in the form of preparations for parenteral administration or solutions for infusion. These solutions are preferably isotonic aqueous solutions or suspensions, possibly prepared before use, for example in the case of freeze-dried preparations containing the active substance alone or together with a vehicle, for example mannitol. The pharmaceutical substances can be sterilized and / or contain excipients, for example preservatives, stabilizers, wetting agents, and / or emulsifiers, solubilizers, salts for regulation of the osmotic pressure, and / or pH regulators. The present pharmaceutical preparations which, if desired, may contain other pharmacologically active substances, such as antibiotics, are prepared in a manner known per se, for example by means of conventional mixing, granulating, coating, dissolving, or lyophilizing processes, and contain from about 1 percent to 100 percent especially from about 1 to about 20 percent of the substance or active substances. The following Examples illustrate the invention without limiting its scope. The Rf values are determined on thin layer chromatography plates coated with silica gel (Merck, Darmstadt, Germany). The ratio of the solvents to each other in the solvent systems used is indicated by volume (volume / volume), and temperatures are given in degrees Celsius (° C).

Eluents (gradients); High performance liquid chromatphy gradient: 0 percent b) in a) for 20 minutes, then 0 percent? 30 percent of b) in a) for 10 minutes, then 30 percent of b) in a) for 5 minutes. Eluent a): ion and methanol match reagent (420 milliliters + 580 milliliters). Eluent b): ion and methanol match reagent (40 milliliters plus 960 milliliters). Ion pairing reagent: 7.5 grams of 1-octane sulfonic acid dissolved in approximately 800 milliliters of water, pH value adjusted to 2.5 with phosphoric acid, and diluted with water to 1000 milliliters. Column: 150 X 3.9 mm, packed with Symmetry C18 5μ (Waters), previously balanced with the eluent a). Flow rate: 1.2 milliliters / minute; ultraviolet detection at 267 nanometers.

Examples; Example 1. Preparation of the β-crystal form of metansul-fonate of 4- (4-methylpiperazin-1-ylmethyl) -N-r 4 -methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) feni11 benzamide - Variant 1. An 11 percent (w / w) suspension of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) methanesulfonate ) -pyrimidin-2-ylamino) phenyl] benzamide in the crystal form OI, is digested in methanol for two days at about 25 ° C. The crystals are isolated by filtration on a glass filter with a G4 frit, and dried overnight at room temperature on filter paper. Smp (by differential scanning calorimetry): 217 ° C (start of fusion). The starting material, 4- (4-methylpi-perazin-1-ylmethyl) -N- [4-methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide methsulfonate was Prepare as follows: 98.6 grams (0.2 moles) of 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl- "- (4-methyl-3- (4-pyridin-3-yl) pyrimidin are added. 2-ylamino) phenyl] benzamide free (for the preparation, see, for example, European Patent Number EP-A-0, 564, 409) to 1.4 liters of ethanol To this beige suspension, 19.2 grams are added ( 0.2 mole) of methanesulfonic acid per drop over a period of 20 minutes The solution is heated under reflux for 20 minutes, and then filtered to a transparent temperature at 65 ° C. The filtrate is evaporated to 50 percent, and the residue is filtered at 25 ° C (filter material A) .The mother liquor is evaporated to dryness.This residue and the filter material A are suspended in 2.2 liters of ethanol, and dissolved under reflux with the addition of 30 milliliters. of water, cooling during the overnight at 25 ° C, filtration, and drying at 65 ° C until a weight constancy is reached, result in 4- (4-methylpiperazin-1-ylmethyl) -N- [4-methyl-3 - (4-pyridin-3-yl) pyrimidin-2-ylamino) phenyl] benzamide as a light beige crystalline mesylate (crystal form). Example 2: Preparation of the β-crystal form of metansul-fonate of 4- (4-mefcylpiperazin-1-ylmefcyl) -N-r 4 -methyl-3- (4-pyridin-3-yl) pyrimidin-2-ylamino ) fenill benzamide - Variant 2. 50.0 grams (101 millimoles) of 4- [(4-methyl-1-piperazinyl) methyl] -N- [4-methyl-3 - [[4- (3-pyridinyl) - are suspended. 2-pyrimidinyl] amino] phenyl] benzamide in methanol (480 milliliters). 9.71 grams (101 millimoles) of methanesulfonic acid and methanol (20 milliliters) are added, heated to 50 ° C, activated charcoal (5.0 grams) is added, and the mixture is boiled under reflux for 30 minutes, filtered, and Concentrate by evaporation. The residue is dissolved in methanol (150 milliliters), and inoculated with 4- [(4-methyl-l-piperazinyl) methyl] -N- [4-methyl-3- [[4- (3-pyridinyl) methanesulfonate]. -2-pyrimidinyl] amino] phenyl] benzamide (modification ß, a few milligrams), leading to the crystallization of the product. Drying at 50 mbar and 60 ° C leads to the raetan-sulfonate of 4- [[4-methyl-l-piperazinyl) methyl] -N- [4-methyl-3- [[4- (3-pyridinyl) -2] -pyrimidinyl] amino] phenyl] benzamide, modification ß; Rf = 0.58 (methylene chloride: ethyl acetate: methanol: concentrated aqueous solution of ammonium hydroxide = 60: 10: 30: 2); High performance liquid chromatography: tret = 10.2 minutes. Example 3: Preparation of the β-crystal form of methansulphonate Le 4- (4-methylpiperazin-1-ylmethyl) -N-r 4 -methyl-3- (4-pyridin-3-yl) pyrimidin-3-ylamino ) feni11 benzamide - Variant 3. 670 grams (1136 millimoles) of 4- [(4-methyl-l-piperazin-1-yl) methyl] -N- [4-methyl-3- [[4- (3-pyridinyl) -2-pyrimidinyl] amino] phenyl] benzamide, modification OI, are heated in methanol (1680 milliliters) The solution is inoculated at 60 ° C with methanesulfonate of 4- [(4-methyl-1-piperazin-1-yl) methyl] -N- [4-methyl-3- [[4- (3-pyridin-nil-2-pyrimidinyl] amino] phenyl] benzamide (modification β, 55 milligrams), upon which the product begins to crystallize. Drying at 50 mbar and at 100 ° C leads to the methanesulfonate of 4- [(4-methyl-l-piperazinyl) methyl] -N- [4-methyl-3 - [[4- (3-pyridinyl) -2-pyrimidinyl] ] amino] phenyl] benzamide, modification ß; Rf = 0.58 (methylene chloride: ethyl acetate: methanol: concentrated aqueous solution of amino hydroxide = 60: 10: 30: 2); High performance liquid chromatography: tr 10.2 minutes. Example 4: Tablets with me ansulfonate Le-4- (4-methyl-1-piperazin-1-ylmethyl) -N-r4-methyl-3-? f4- (3-pyridinyl) -2-pyrimidi-nyl] amino) enyl] benzamide, crystal form ß Tablets containing 100 milligrams of the active substance mentioned in the title are normally prepared in the following composition: Compo ición nt Active ingredient 100 milligrams Crystalline lactose 240 milligrams Avicel 80 milligrams PVPPXL 20 milligrams Aerosil 2 milligrams Magnesium stearate 5 milligrams 447 milligrams Preparation: The active substance is mixed with the vehicle materials, and compressed in a tablet press (Korsch EKO, drilling diameter of 10 millimeters). Avicel is microcrystalline cellulose (FMC, Philadelphia, USA). PVPPXL is cross-linked polyvinylpolypyrrolidone (BASF, Germany). Aerosil is silicon dioxide (Degussa, Germany).

Example 6: Capsules with 4-r (4-methyl-1-piperazin-1-ylmethyl) -N-f 4 -methyl-3-f 4 (3-pyridinyl) -2-pyrimidinyl] amino] iuetansulonate] phenyl] benzamide, crystal form ß Capsules containing 100 milligrams of the compound mentioned in the title as an active substance are normally prepared in the following composition: Composition Active ingredient 100 milligrams Avicel 200 milligrams PVPPXL 15 milligrams Aerosil 2 milligrams Magnesium stearate 1.5 milligrams 318. 5 miles The capsules are prepared by mixing the components, and filling the mixture into hard gelatin capsules, size 1.

Claims (12)

1. A form of the monomethane sulphonic acid addition salt of a compound of the formula I: which comprises at least 90 weight percent of crystals of the β modification, these crystals being non-hygroscopic, and remaining essentially dry in a glass climatic chamber at 25 ° C and with relative humidity up to and including 93 percent .

2. A crystalline form according to claim 1, of the methanesulfonic acid addition salt of a > The compound of formula I, which comprises at least 95 weight percent of crystals of the β modification, and remains dry at 93 percent relative humidity and at 25 ° C.

3. A crystalline form according to claim 1, of the methanesulfonic acid addition salt of a The compound of formula I, which comprises at least 99 weight percent of crystals of the β modification, and remains dry at 93 percent relative humidity and at 25 ° C.

4. A crystalline form according to claim 1, of the methanesulfonic acid addition salt of a compound of the formula I., which comprises at least 99 weight percent of crystals of the β modification, and has a melting point less than 225 ° C.

5. A ß crystal form according to claim 1 of the methanesulfonic acid addition salt of a compound of the formula I, which comprises at least 99 weight percent of crystals of the β modification, and has a melting point lower than 217 ° C, defined as the start of fusion in the differential scanning calorimetry thermogram.

6. The ß crystal form according to claim 1 of the methanesulfonic acid addition salt of a compound of the formula I, which shows, in X-ray diffraction, a peak at a refractive angle 2T of 20 °, this peak having a relative line intensity of 65, compared to the most intense line in the diagram. The ß crystal form according to claim 3 of the methanesulfonic acid addition salt of a compound of the formula I, which shows, in an X-ray diffraction diagram, lines having a relative line intensity , compared to the most intense line in the diagram, of 20 or more at the following 2T refractive angles (the relative line strengths are given in parentheses): 9.7 ° (40), 13.9 ° (26), 14.7 ° (23) , 17.5 ° (57), 18.2 ° (90), 20.0 ° (65), 20.6 ° (76), 21.1 ° (100), 22.1 ° (89), 22.

7 ° (38), 23.8 ° (44) , 29.8 ° (23) and 30.8 ° (20).

8. The ß-crystal form according to claim 5 of the methanesulfonic acid addition salt of a compound of the formula I, which has a melting point of 217 ° C, defined as the onset of the fusion in the differential scanning calorimetry diagram, and that essentially shows the following X-ray diffraction diagram:

9. The ß-crystal form according to any of claims 1 to 8 of the methanesulfonic acid addition salt of a compound of the formula I, for use in a process for the diagnosis or therapeutic treatment of the human or animal body.

10. A pharmaceutical composition, which comprises a β-crystal form according to any of claims 1 to 8 of the methanesulfonic acid addition salt of a compound of the formula I, and a pharmaceutically acceptable carrier.

11. The use of a β-crystal form according to any of claims 1 to 8 of the methanesulfonic acid addition salt of a compound of the formula I, for the preparation of a pharmacological agent for the treatment of a disease tumor.

12. Processes for the preparation of the β-crystal form according to claim 1, of the methanesulfonic acid addition salt of a compound of the formula I, characterized by: a) digesting another form of crystal or a starting material amorphous of the methanesulfonic acid addition salt of a compound of the formula I, with a suitable polar solvent in suspension, at a temperature between 20 ° C and 50 ° C, or b) dissolving another form of crystal or an amorphous starting material of the methanesulfonic acid addition salt of a compound of the formula I, in a polar solvent, at a suitable temperature of 25 ° C up to the reflux temperature of the reaction mixture, and then start the crystallization by adding a small amount of the crystal form ß as the crystal sowing, at a temperature between 20 ° C and 70 ° C.