JP2002515046A - Methods for treating postmenopausal diseases including osteoporosis - Google Patents

Abstract

  (57) [Summary] Novel methods of treating osteoporosis are described. Idoxifen is a preferred compound.

Description

Description: FIELD OF THE INVENTION The present invention relates to therapeutic agents that bind to estrogen receptors and have been found to be useful in the treatment of osteoporosis. BACKGROUND OF THE INVENTION Estrogen depletion that occurs during menopause is an important etiology of the high incidence of osteoporotic fractures and cardiovascular disease in postmenopausal women. Although postmenopausal bone loss can be prevented by estrogen replacement therapy (ERT), unmatched ERT increases the risk of endometrial cancer. An ideal treatment would retain the desired skeletal and cardiovascular effects of estrogen without having undesirable effects on reproductive tissues. Tamoxifen is an antiestrogen that has been found to lower cholesterol levels and protect against bone loss in postmenopausal women. Tamoxifen is also effective in an ovariectomized rat model of osteoporosis. However, tamoxifen has been shown to have undesirable side effects, especially by causing endometrial hyperplasia and endometrial cancer. 1. Love RR, Wiebe DA, Newcomb PA, CameronL, Leventhal H, Jordan VC, Feyzi J, DeMets DL. (1991) .Effects of tamoxifen on cardiovascular risk factors in postmenopausal wom en. Annals of Internal Medicine, 115, 860. -864.2.Love RR, Mazess RB, Barden HS, Epstein S, Newcomb PA, Jordan VC, Carbone PP, DeMets DL. (1992) .Effects of tamoxifen on bone mineral density in postmenopausal women with breast cancer.New England Turner RT, Wakely GK, Hannon KS, Bell NH. (188) .Tamoxifen inhibits osteoclast-mediated resorpti on of trabecular bone in ovarian hormone-deficient rats.Endocrinology, 12, 2, 1146-1150. checking ... SUMMARY OF THE INVENTION The present invention provides methods for preventing and treating postmenopausal disease without apparent uterine nutritional effects. The method comprises administering to a human in need of such prevention and treatment an effective amount of Formula I: Wherein X is 3- or 4-iodo or bromo, and the symbols R ¹ and R ² may be the same or different and represent a C _1-3 alkyl group, especially a methyl or ethyl group. Or R ¹ is a hydrogen atom, R ² is C _1-3 alkyl, or R ¹ and R ² together with the nitrogen atom to which they are attached typically have 5 Or a saturated heterocyclic group having 6 ring atoms, especially a pyrrolidino, piperidino, 4-methylpiperidino or morpholino group), and pharmaceutically acceptable acid addition salts thereof. DETAILED DESCRIPTION OF THE INVENTION The present invention is a therapeutic method for treating post-menopausal disease using a group of compounds that have been manufactured and evaluated as being useful in treating estrogen receptor-positive breast cancer. These compounds are described in Formula I above and in US Pat. No. 4,839,155. Preferred compounds for the described methods of treatment are (E) -1- [2- [4- [1- (4-Iodophenyl) -2-phenyl-1-butenyl] phenoxy] pyrrolidine. The compounds are known to bind to the estrogen receptor and cause either an estrogen agonist or antagonism, depending on the tissue being studied. The term "post-menopausal disease" refers to osteoporosis and atherosclerotic cardiovascular diseases such as myocardial infarction and stroke and increases in plasma cholesterol. The methods of the present invention are useful for preventing bone loss and generating a plasma lipid profile associated with a reduced risk of atherosclerosis. The ability to prevent bone loss will be evaluated by studies in an ovariectomized rat model of osteoporosis and in postmenopausal women. Histomorphometric studies indicate that progressive bone loss occurs at the proximal tibial metaphysis when ovaries are removed from adult female rats. Three months after ovariectomy (OVX), increased bone turnover, where excessive bone resorption is predominant, removes 60-70% of the cancellous bone. At a slower rate, bone loss also occurs in the lumbar spine. OVX-induced bone loss in rats, which can be completely prevented by alternative administration of estrogens, forms the basis of the most widely used and best analyzed animal models of osteoporosis. Sprague-Dawley rats were used at 7-8 months of age. Baseline bone mineral density (BMD) was measured by dual energy x-ray absorptiometry at 3-6 regions of the lumbar spine and at the proximal tibial metaphysis. Rats were then divided into groups of 8-10 with almost the same mean and standard deviation values for lumbar BMD. Groups of rats were ovariectomized bilaterally (OVX) and one group was mock-treated in each experiment. Idoxifen was prepared for oral administration as a suspension in a 1% aqueous solution of carboxymethylcellulose. Rats were dosed once daily by oral gavage. In each experiment, one OVX group and the sham group received vehicle by oral gavage once daily. Dosing started the day after surgery. Plasma cholesterol levels were determined two weeks after treatment. Lumbar and tibia BMD were measured at monthly intervals. The animals were killed, the uteri were removed and the wet weight was determined. Tibia were harvested post-mortem, fixed and sectioned for histomorphometry. The spongy bone area and perimeter were measured as a secondary spongy, 1.2 mm from the growth plate. Secondary structural parameters, column width, column number and column separation were calculated from the primary area and marginal measurements using equations developed by Parfitt et al. Initial dose-range study of the effects of idoxifen on bone loss, plasma cholesterol and uterine weight in an ovariectomized rat model of osteoporosis The purpose of this study was to determine the optimal dose of idoxifen for prevention of bone loss in an OVX rat model of osteoporosis Was to do. BMD was measured at 1, 2 and 3 months after treatment. In addition to removing the tibia for histomorphometry, the femur and vertebra were removed for ex vivo measurements of BMD (femur only) and mechanical testing. Idoxifen was administered at 2, 8, 40 and 200 micrograms / kg / d. The idoxifen dose ineffective for all measured parameters was 2 μg / kg. Only the dose of 200 μg / kg idoxifen caused significant prevention of OVX-induced reduction of BMD in the lumbar spine. This dose was effective and caused 100% inhibition of bone loss in January. After 3 months of treatment in the lumbar spine, 200 μg / kg idoxifen caused about a 50% inhibition of bone loss. In January at the proximal tibia, a dose of 8-200 μg / kg idoxifen caused about a 50% inhibition of bone loss. The degree of this protection fell to about 25%, which was not significant in March. This suggested that 200 μg / kg was not the optimal dose of idoxifen at this skeletal site. Bone mineral density measured ex vivo indicated that 200 μg / kg idoxifen maintained proximal femoral BMD at sham control levels. This dose also maintained mid-shaft medullary cross-sectional area at the sham control level, suggesting that idoxifen prevents cortical as well as cancellous bone loss. Idoxifen did not adversely affect the mechanical strength of the femoral shaft in the 3-point bending test or the L2 vertebral body in the axial compression test. Histomorphometry showed a nonsignificant but small effect of idoxifen on the proximal tibial cancellous bone area 3 months after treatment, consistent with BMD measurements. There were no differences between the groups with regard to pillar width. In the long term, the activity of idoxifen at doses as low as 40 μg / kg was evident with regard to column number and segregation, despite lack of effect on the tibial cancellous bone area. Idoxifen (200 μg / kg) significantly reduced plasma cholesterol (FIG. 5). After 3 months of treatment, all doses of idoxifen caused a very slight, but statistically significant increase in uterine weight. Detailed study of the dose of idoxifen on bone loss, plasma cholesterol and uterine weight in an ovariectomized rat model of osteoporosis The purpose of this study was to determine the optimal dose of idoxifen for prevention of bone loss in the OVX rat model of osteoporosis Met. BMD was measured at one month and treatment continued for another two weeks before the uterus and tibia were harvested. Idoxifen at 200 and 500 μg / kg completely prevented bone loss in the lumbar spine. No significant effect of 1000 μg / kg idoxifen was seen in the lumbar spine. 200-1000 μg / kg idoxifen completely prevented bone loss at the proximal tibial metaphysis. Histomorphometry showed that idoxifen optimally prevented OVX-induced loss of spongy bone at 500 μg / kg. Prevention of the OVX-induced decrease in column width occurred significantly at 200 and 500 μg / kg idoxifen. Pillar numbers were significantly protected with 500 and 1000 μg / kg idoxifen. Idoxifen at 200-1000 [mu] g / kg significantly prevented OVX-induced increase in column separation. All doses of idoxifen significantly reduced plasma cholesterol levels (FIG. 11). There was no effect of idoxifen on uterine wet weight at any of the doses tested. 500 μg / kg idoxifen was identified in OVX rats after 6 weeks of treatment, consistent with being optimal for all measured parameters. Over a treatment period of 6 weeks, the optimal dose of idoxifen was shown to be 500 μg / kg. The minimally effective dose for preventing bone loss in the spine was 200 μg / kg and the cholesterol lowering effect was 100 μg / kg. In summary, the osteo-protective and cholesterol-lowering effects of idoxifen are useful in preventing post-menopausal disease without having an apparent uterotrophic effect. Three different doses of idoxifene (2.5, 5, and 10 mg / day) were compared to placebo in a 3-month study in postmenopausal women. These women had proven to have low bone mineral density at the start of the study. The presence of an effect on bone, consistent with a reduction in the rate of bone loss, was measured for bone resorption (measured as urinary collagen cross-link elimination, C-telopeptide elimination, free and total cross-links) and bone formation (serum osteocalcin) Were detected by measuring changes in biochemical markers. The dose-related decrease observed at the level of these biochemical markers (see below) is indicative of a decrease in bone turnover, consistent with estrogen's effect on bone turnover in postmenopausal women. All changes are reported as percentage changes from baseline. Statistically significant differences from placebo are indicated by the following notations: ^* = p <0.01, ^** = p <0.001. Changes in the levels of different lipid parameters, fibrinogen and other coagulation / fibrinolysis parameters were also measured before and after treatment as indicators of the likely effect of idoxifen on the risk of cardiovascular disease. The results of this study are described below. The compounds of the present invention and their pharmaceutically acceptable salts that are active when administered orally can be formulated as solutions, for example, syrups, suspensions or emulsions, tablets, capsules and lozenges. Liquid formulations generally comprise a suspending agent, preservative, flavoring or coloring agent and a suitable liquid carrier (s) such as, for example, ethanol, glycerin, a non-aqueous solvent such as polyethylene glycol, oil or water. Or a suspension or solution of the compound or a pharmaceutically acceptable salt thereof. A composition in the form of a tablet can be prepared using any suitable pharmaceutical carrier (s) routinely used for preparing solid formulations. Examples of such carriers include magnesium stearate, starch, lactose, sucrose and cellulose. A composition in the form of a capsule can be prepared using conventional encapsulation methods. For example, a pellet containing the active ingredient can be prepared using a standard carrier, and then filled into a hard gelatin capsule; alternatively, the dispersion or suspension can be prepared using a suitable liquid, such as an aqueous gum, cellulose, silicate or oil. It can be prepared by using a suitable pharmaceutical carrier (s), and then filling the dispersion or suspension in a soft gelatin capsule. The compounds of the present invention and their pharmaceutically acceptable salts that are active when administered parenterally (by injection or infusion) can be formulated as solutions or suspensions. Compositions for parenteral administration are generally prepared from a solution or suspension of the active ingredient in a sterile aqueous carrier or parenterally acceptable oil, such as polyethylene glycol, polyvinylpyrrolidone, lectin, peanut oil or sesame oil. Become. Alternatively, the solution can be lyophilized and then reconstituted with a suitable solvent just prior to administration. A typical suppository composition may contain a compound of the present invention or a pharmaceutically acceptable salt thereof, which is active when so administered, may contain a binding and / or lubricating agent such as a polymeric glycol, gelatin or cocoa. Include with avatar or other low melting point edible or synthetic wax or fat. Typical transdermal formulations include a conventional aqueous or non-aqueous vehicle, such as a cream, ointment lotion or paste, or are in the form of a medicated plaster, patch or film. For topical administration, the applied pharmaceutical compositions include solutions, suspensions, ointments and solid inserts. Topically pharmaceutically acceptable carriers include, for example, water, water and water-miscible solvents, such as mixtures of lower alcohols or edible oils, and water-soluble, ophthalmically acceptable non-toxic polymers, such as methyl cellulose. Of cellulose derivatives. Pharmaceutical preparations include emulsifiers, preservatives, wetting agents and thickeners, non-toxic adjuvants such as polyethylene glycol; antimicrobial components such as quaternary ammonium compounds; buffer components such as alkali metal chlorides; Antioxidants; and other conventional ingredients such as sorbitan monolaurate. Preferably, the compositions are in unit dosage form. The dose of the compound of the present invention in a drug dosage unit is selected from an effective, non-toxic amount, in the range of 0.01-200 mg / kg of the active compound, preferably, in a range of 0.1-100 mg / kg. You. The selected dose may be adjusted for a human patient in need of treatment or prevention of osteoporosis, or in need of lowering plasma cholesterol, or in need of prevention of cardiovascular disease, 1 to 6 times a day, It is administered orally, rectally, topically, by injection or continuously by infusion. Oral dosage units for human administration preferably contain 10 to 500 mg of active compound. Generally, lower dosages are used for parenteral administration. Oral administration is used where safe, effective and convenient for the patient. No unacceptable toxic effects are expected when a compound of the present invention is administered in accordance with the present invention. Example 1 An oral dosage form for oral administration of an orally active compound of formula (I) is prepared by screening, mixing and filling the following proportions of the ingredients, for example, and filling into hard gelatin capsules. To manufacture. Component amount (E) -1- [2- [4- [1- (4-Iodophenyl) -2-phenyl-1-butenyl] phenoxy] pyrrolidine 100 mg Magnesium stearate 10 mg Lactose 100 mg Example 2 Sucrose Calcium phosphate dihydrate The hydrate and the orally active compound of formula (I) were mixed and granulated with a 10% gelling solution. The wet granules were screened, dried, mixed with starch, talc and stearic acid, screened and compressed into tablets. Component amount (E) -1- [2- [4- [1- (4-Iodophenyl) -2-phenyl-1-butenyl] phenoxy] pyrrolidine 75 mg Calcium sulfate dihydrate 100 mg Sucrose 15 mg Starch 8 mg Talc 4 mg Stearic acid 2 mg Example 3 (E) -1- [2- [4- [1- (4-Iodophenyl) -2-phenyl-1-butenyl] phenoxy] pyrrolidine, 50 mg, was dispersed in 25 ml of normal saline. An injectable preparation was prepared. Example 4 This experiment was performed to compare the mechanism of action of idoxifen and raloxifene on osteoblasts. Using a construct containing an estrogen response element (ERE) upstream of the luciferase reporter gene (described below), it has now been shown that idoxifen, like estrogen, is a pure agonist mediated by ERE in osteoblasts. Efficacy of agonist action was similar between the natural steroid hormones estrogen and idoxifen. Raloxifene at the same concentration as idoxifene (0.01-10 μM) showed a very weak signal via the ERE with a similar intensity as the vehicle control. Competition experiments were performed to confirm the mechanism of action through this response element. Raloxifene inhibited the agonist activity of both estrogen and idoxifen via the ERE. At a dose of 100 nM ligand (either estrogen or idoxifen), a maximal agonist response was obtained. Co-treatment with 500 nM raloxifene reduced reporter gene activity to vehicle control levels. In contrast, 500 nM idoxifen did not reduce the maximal agonism of 100 nM estrogen in osteoblasts. With sub-maximal concentrations of idoxifen and estrogen, an additive effect of the two agonists was seen via ER in osteoblasts. Experimental Methods Cells were seeded in 6-well plates at 1.5 × 10 ⁵ cells / well or in 24-well plates at 1.5 × 10 ⁴ cells / well in phenol red-free medium. A DNA construct containing the mouse mammary tumor virus promoter in which the glucocorticoid response element was replaced with 5 copies of a 33-base pair vitellogenin estrogen response element was used. This is upstream of the luciferase reporter gene (MMTV-ERE-Luc) (Wen, DX, Xu, MKGoldman and P. McDonnell. 1994. The A and B isoforms of the human progesterone receptor operate through distinct signaling pathways within targe. t cells.Molec.Cell.Biol.14: 8356-8364). Transfection efficiencies were corrected using the renilla-luciferase vector and the dual-luciferase detection method (Promega, Madison, WI). DNA was introduced into rat osteosarcoma (Ros 17 / 2.8) cells by the lipofectin method (Life Technologies, Gaithersburg, MD). Cells were co-transfected with 2 μg per well in a 6-well plate and 140 ng and 25 ng of a control retinal luciferase vector (pRL-CMV) in a 24-well plate of MMTV ERE-Luc. Transfection efficiency was corrected by co-transfection with a Renilla luciferase vector utilizing a different substrate, coelenterazine, for its bioluminescence readout (Promega, Madis on WI). Cells were cultured overnight. The transfection medium was then removed and the cells were cultured with or without hormone for 48 hours. The cells were washed in phosphate buffered saline and then lysed with 500 μl / well of 1 × passive lysis buffer (PLB) for 15 minutes while rocking the sample on a rocking table. The lysate was centrifuged at 12,000 g for 30 seconds and the clear lysate was transferred to a tube for reporter enzyme analysis. Samples (20 μl) were transferred to a 96-well luminescence detection plate and reacted with 100 μl of each assay reagent (Promega, Madison, WI). Each assay reagent was injected with a microlumat LB96P luminometer, Wallac, Gaithersburg, MD, which measures luciferase activity. Luciferase activity provides a surrogate for the transcriptional activity of estrogen responsive genes, including estrogen responsive elements (EREs). Therefore, up-regulation of luciferase activity indicates an agonistic effect via ERE, and down-regulation indicates an antagonistic effect. Discussion: Idoxifen is an agonist via the estrogen response element (ERE) in osteoblasts. In contrast to idoxifene, raloxifene is an ERE-mediated antagonist in osteoblasts at the doses tested, suggesting a different mechanism for the bone deficient effects seen with raloxifene. Therefore, raloxifene can exert its biological effects via non-ERE containing sequences present on the 5'-untranslated region of the human TGFβ3 promoter. In the same cell system, raloxifene inhibited ERE-containing vitellogenin promoter expression and therefore showed pure estrogen antagonism. Raloxifene response element (Yang, N, N., Venugopalan, M., Hardikar, S., and Glasebrook, A. (1996) Identification of an estrogen response element activated by metabolites of 17beta-estradiol and raloxifen Science 273: 1222-1225 ) Is not on the same gene as the ERE, suggesting that regulation through this response element acts on a different gene. This distinguishes the mechanism of action of idoxifene from that of the selective-estrogen receptor modulator (SERM) raloxifene, a more classical estrogen-like mechanism that exerts its biological agonism in osteoblasts via the ERE. Idoxifene is arranged. The effects of idoxifen and estrogen are specific for a reporter gene construct that carries the classic ERE. This system has been shown to be sensitive to cell-specific factors and is, therefore, an effective model of action on endogenous gene transcription.

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Claims (1)

[Claims] 1. A method for preventing and treating postmenopausal diseases, which requires the treatment and prevention. Administering to the subject an effective amount of a compound of formula I. 2. The method according to claim 1, wherein the postmenopausal disease is osteoporosis. 3. The compound of formula I is (E) -1- [2- [4- [1- (4-iodophenyl)- 2-phenyl-1-butenyl] phenoxy] ethyl] pyrrolidine. The described method. 4. Compounds of Formula I in the manufacture of a medicament for use in the treatment of postmenopausal disease Use of. 5. The use according to claim 4, wherein the postmenopausal disease is osteoporosis. 6. The compound of formula I is (E) -1- [2- [4- [1- (4-iodophenyl) -2]. -Phenyl-1-butenyl] phenoxy] ethyl] pyrrolidine. Use.