MXPA97006424A - Use of moxonidine for the treatment of the aterosclero - Google Patents

Abstract

The present invention relates to: The use of moxonidine for the treatment of atherosclerosis is described

Description

USE OF MOXONIDINE FOR THE TREATMENT OF ATHEROSCLEROSIS Description of the invention This invention describes the use of moxonidine in the treatment of atherosclerosis. Moxonidine is a well-known compound, which is described in, for example, U.S. Patent No. 4,323,570, for its properties as an agent for reducing blood pressure. The compound has the chemical formula 4-chloro-6-methoxy-2-methyl-5- (2-imidazolin-2-yl) aminopyrimidine. Atherosclerosis is a leading cause of ischemic death (ischemic heart disease) in industrialized countries. It is now accepted that atherosclerosis begins with local damage to the arterial endothelium which results in the proliferation of arterial smooth muscle cells with lipid deposition and accumulation of macrophages. As the atherosclerotic plaque develops, it progressively obstructs more and more the blood vessel and can thus lead to ischemia or infarction. It has now been found that moxonidine is useful in the treatment of atherosclerosis. Thus, the invention comprises the use of the moxonidine or a pharmaceutically acceptable acid addition salt thereof, in the treatment of atherosclerosis. More particularly, the invention comprises the use of the moxonidine or a pharmaceutically acceptable acid addition salt thereof, in the preparation of a medicament for the treatment of atherosclerosis. REF: 25389 It has been found in the test models that moxonidine significantly decreases cholesterol accumulation induced by atherogenic serum and also that it inhibits proliferation in cultured cells of an atherosclerotic plaque. As mentioned above, moxonidine is the compound 4-chloro-6-methoxy-2-methyl-5- (2-imidazolin-2-yl) aminopyrimidine and can also be used in the form of a pharmaceutically acceptable acid addition salt. Suitable acid addition salts are pharmaceutically acceptable non-toxic addition salts with appropriate acids, such as those with inorganic acids, for example hydrochloric, bromic, nitric, sulfuric or phosphoric acids or with organic acids, such as carboxylic acids organic, for example glycolic, maleic, hydroxymalonic, fumaric, malic, tartaric, citric, salicylic, o-acetoxybenzoic or organic ulphonic acid, 2-hydroxyethane sulphonic, toluene-p-sulphonic or naphthalene-2-sulfonic acid. The identification of those patients who are in need of treatment for atherosclerosis is in the skill and knowledge of the one experienced in the art. For example, individuals suffering from clinically significant atherosclerosis or who are at risk of developing clinically significant atherosclerosis are patients in need of treatment for atherosclerosis. A clinician skilled in the art can easily determine, by the use of clinical tests, physical examination and medical / family history, whether an individual is a patient in need of treatment for atherosclerosis. An effective antiatherosclerotic amount of a compound of formula (1) is an amount which is effective to inhibit the development or growth of atherosclerosis in a patient in need thereof. As such, it is understood that successful treatment of a patient for atherosclerosis includes effectively decreasing, stopping or stopping atherosclerotic lesion or growth or plaque growth and does not necessarily indicate a total elimination of atherosclerosis. It will be further understood and appreciated by those skilled in the art that successful treatment for atherosclerosis may include prophylaxis in the prevention of atherosclerotic lesion or plaque formation and inhibition of atherogenesis. For the purpose of the invention, moxonidine can be administered by several routes, for example by the oral or rectal route, topically or parenterally, for example by injection or infusion, it is usually employed in the form of a pharmaceutical composition. Such compositions are prepared in a manner well known in the pharmaceutical art. In making the composition, the active ingredient will usually be mixed with a carrier or diluted by a carrier and / or encased in a carrier which may be, for example, in the form of a capsule, sachet, paper or other container. When the carrier serves as a diluent, it can be a solid, semi-solid or liquid material which acts as a vehicle, excipient or medium for the active ingredient. Thus, the composition can be in the form of tablets, tablets, sachets, wafers, elixirs, suspensions, aerosols (as a solid or in a liquid medium), ointments containing for example up to 10% by weight of the active compound, gelatin capsules soft and hard, suppositories, injection solutions, suspensions, sterile packaged powders and as a topical patch. The preferred formulations are for oral dosing and are especially in the form of a tablet or capsule.

Examples of suitable carriers are lactose, dextrose, sucrose, sorbitol, mannitol, starches, ethylcellulose, acacia gum, calcium phosphate, alginates, tragacanth, gelatin, syrup, methylcellulose, methyl- and propyl-hydroxybenzoate, talc, magnesium stearate and mineral oil. The compositions of the invention may, as is well known in the art, be formulated to provide a rapid, sustained or delayed release of the active ingredient after administration to the patient. When the composition is formulated in unit dosage form, it is preferred that each unit dosage form contains from 0.01 mg to 2.0 mg for example, from 0.05 mg to 1.0 mg. The term "unit dosage form" refers to physically discrete units, suitable as unitary dosages for human and animal subjects, each unit containing a predetermined amount of the active material calculated to produce the desired therapeutic effect, in association with the required pharmaceutical carrier. The active compound is effective over a broad dosage range and, for example, dosages per day will normally fall in the range of 0.01 to 2.0 mg, more usually in the range of 0.1 to 1.0 mg. Usually one dose a day, preferably in the morning, is sufficient. However, it will be understood that the amount administered will be determined by the physician in light of the relevant circumstances including the condition to be treated and the route of administration chosen and therefore the above dosage ranges are not intended to limit the scope of the invention in no way.

The following example illustrates the activity of moxonidine in human cell tissue models.

Example The antiatherosclerotic and antiatherogenic effects of moxonidine are demonstrated in the following manner.

Materials Cell culture Sub-endothelial cells were isolated from the thick normal intima (free of atherosclerosis) or atherosclerotic plaque of the human aorta derived from the recent autopsy within 1-3 hours after sudden death. Methods for the isolation and culture of human aortic cells are described in Orekhov et al. 1986. (Principal culture of human aortic intimal cells as a model for testing antiatherosclerotic drugs. "Effects of cyclic AMP, prostaglandins, calcium antagonists , antioxidants, and lipid-lowering agents. "Atherosclerosis 60: 101-110).

Methods Antiatherosclerotic effects Moxonidine dissolved in 10 μl of Medium 199 was added to 9 wells of a 7-day cell culture derived from the atherosclerotic plaque.

Simultaneously, 10 ml of Medium 199 containing 1 μCi / ml thymidine [3 H] (21 Ci / mmol, Amersham International, Amersham, U.K.) are added to three of the nine wells. To the other three cavities, 10 μl of Medium 199 containing leucine [1 C] 1 μCi / ml (135 Ci / mmol, Amersham) is added. The remaining three cavities were supplemented with 10 μl of Medium 199. After an incubation of 24 hours (on the eighth day of culture), the intracellular cholesterol content and the DNA synthesis were determined. The total intracellular cholesterol was estimated in three free cavities of radioactive precursors. The intracellular lipids were extracted with a mixture of hexane-isopropanol (3: 2, v / v) [Will A .. Radin N.S .. 1978. Lipid extraction of tissue with low-toxicity solvent, Anal. Biochem. 90: 420-426]. The content of total cholesterol in lipid extracts is determined as described by fOrekhov et al. 1986] when using the CHOD-PAP cholesterol method of Boehringer Mannheim Monotest (Boehringer Mannheim GmbH Mannheim, Germany). DNA synthesis is evaluated by incorporation of [3 H] thymidine into the acid-insoluble cell fraction as described in Orekhov et al. 1986. After incubation with thymidine [3 H] the cells were rinsed three times with PBS and three times with trichloroacetic acid (TCA) at 5%. The cells were then dissociated with 0.1 N NaOH and after neutralization with HCl, the reductivity was measured in a liquid scintillation counter using the Triton X-100-toluene-based scintillation liquid. The total cholesterol content and DNA synthesis are determined in cell cultures (eight cavities for each parameter) incubated without medication. The values obtained serve as the reference values. The results are expressed as reference percent for each atherosclerotic cell parameter.

For the confirmation of antiatherosclerotic activity, additional independent experiments on cell cultures obtained from two other cell isolates were carried out. Thus, the final conclusion was based on the results of at least three independent experiments.

Antiartergenic effects Atherogenesis in primary smooth muscle cells derived from the thick uncoated human aortic intima was initiated by the addition of atherogenic blood serum from patients with coronary atherosclerosis determined angiographically. Previously it had been found that such serum causes atherosclerotic manifestations at the cellular level, ie: induction of lipid accumulation in cultured cells, stimulation of cell proliferation and synthesis of extracellular matrix fQrekhov et al .. 1990b, Triggerlike stimulation of accumulation of cholesterol and DNA and extracellular matrix synthesis induced by atherogenic serum or low density lipoprotein in cultured cells. Circ. Res. 66: 311-320]. The atherogenic serum capable of inducing the significant increase in the total cholesterol levels of the cells cultured in the 24 hour incubation was prepared from blood taken from normolipidemic, non-diabetic subjects with coronary atherosclerosis determined angiographically. The atherogenic blood serum was prepared from the accumulated blood of 10-15 patients and was tested for atherogenicity as described [Chazov et al. 1986, Atherogenicity of blood serum from patients with coronary / heart disease, Lancet 2 , 595-598, Qrekhov et al. 1988, Blood serum atherogenicity associated with coronary / atherosclerosis. Evidence for nonlipid factor providing atherogenicity of low-density lipoproteins and an approach to its elimination. Circ. Res. 62. 421-429]. The detailed characteristics of atherogenic serum are described elsewhere [Chazov et al .. 1986]. Only serum that causes a statistically significant elevation of total intracellular cholesterol in normal aortic cells cultured within 24 hours was used. Forty microliters of the arteriogenic serum are added to each well of 96-well tissue culture plates containing 60 μl of culture medium. Ten microliters of the dissolved moxonidine are added to 9 cavities of a seven-day cell culture simultaneously with the atherogenic serum. At the same time, 10 μl of Medium 199 containing thymidine [3 H] 1 μCi / ml is added to three of the six cavities. To the other three cavities are added 10 μl of Medium 199 containing leucine [1 C] 1 μCi / ml. The remaining three cavities are supplemented with 10 μl of Medium 199. After an incubation period of 24 hours (on the eighth day in the culture) the total intracellular cholesterol content also as the DNA synthesis are determined as described above. The cell parameters are determined in cell cultures (eight cavities for each parameter) incubated without medication and patient serum (standard conditions). The values obtained serve as the standard values. All parameters examined were determined in cells incubated for 24 hours with atherogenic serum but without moxonidine. The results are expressed as a percent increase over the standard value for each atherosclerotic cell parameter.

For a confirmation of the atherogenic activity, additional independent experiments on cell cultures obtained from two other cell isolates were carried out. Thus, the final conclusion was based on the results of at least three independent experiments.

Results Antiatherosclerotic Effects Antiathermaltogenic effects, that is, effects mimicking the regression of atherosclerosis in a cell culture, were examined on smooth muscle cells derived from an atherosclerotic plaque, as described above. These cells differed considerably from the normal cultured cells of the intima not involved in their cholesterol content. The average cholesterol content in the cultured cells of the plate was five times higher than that in the cultured cells of the intima not involved. The prolific activity was approximately twice as high as in a primary cell culture derived from the intima not involved. During a 24-hour incubation period with plaque cells, moxonidine at 10 ^ -IO "5 M significantly decreased the intracellular cholesterol content by 20-30% .The concentration dependence curve for the effect of moxonidine on the cholesterol content in plaque-derived cells had a Bell-like shape, similar to experiments carried out on a culture of normal cells.Moxonidine also inhibited the prolific activity of cultured cells of an arteriosclerotic plaque at 10 ~ 7 M.

Ffer.tn & antiaterogenic With the aim of revealing the antiatherogenic activity, that is, the activity that mimics the prevention of atherosclerosis at the cellular level, smooth cells of non-involved human aortic intima and atherogenic serum obtained from patients with coronary atherosclerosis were used, as described above. . This serum induces a statistically significant 1.3 to 1.6 fold increase in the cholesterol content of the cultured cells. The increase in cholesterol content above the standard level was assumed as a 100% atherogenic effect. At concentrations of 10"7-10" 5 M, moxonidine significantly reduced the accumulation of intracellular cholesterol induced by the atherogenic effect. At concentrations of lO ^ -IO "5 M, the effect of moxonidine was significant.Atherogenic serum significantly stimulated proliferous activity of intimal smooth muscle cells estimated by the incorporation of [3 H] thymidine into cellular DNA. Thymidine incorporation was increased by 30-90% in the presence of atherogenic serum.At 10'8-10'5 M, moxonidine inhibits the stimulation of proliferative activity induced by atherogenic serum. in relation to this date, the best method known by the applicant to carry out said invention is the conventional one for the manufacture of the objects to which it refers.The invention having been described as above, property is claimed as content in the following

Claims (2)

1. Claims 1. The use of the moxonidine or a pharmaceutically acceptable acid addition salt thereof, characterized in that it is applied in the preparation of a medicament for treating atherosclerosis.
2. 2. The use of moxonidine according to claim 1, characterized in that it is applied in the preparation of a medicament in unit dosage form comprising 0.01 to 2.0 mg of moxonidine.