Methods for treating metabolic syndrome

Abstract

Disclosed herein are methods of treating metabolic syndrome in a subject using a vitamin D receptor agonist. The preferred vitamin D receptor agonist of the present invention is ellocalciferol.

Description

Technical Field

[0001] The invention disclosed herein generally relates to methods of treating metabolic syndrome in a subject using vitamin D receptor agonists. Background Art

[0002] Metabolic syndrome is a group of conditions that occur together and increase the risk of heart disease, stroke and type 2 diabetes. These conditions include excess body fat (visceral fat) around the waist, abnormal cholesterol or triglyceride levels (dyslipidemia), elevated blood pressure (hypertension), insulin resistance or glucose intolerance, high blood sugar (hyperglycemia), obesity, atherosclerosis, diabetes and hepatic steatosis. It has been found that the activation of NF-κB signaling is important in the development of metabolic syndrome; therefore, NF-κB inhibition provides a potential therapeutic approach to address metabolic syndrome. Vitamin D acts as an agonist of nuclear receptors (called vitamin D receptors), which control gene expression related to the regulation of calcium homeostasis and other cellular functions (such as inhibition of upregulated NF-κB signaling). Specifically, elocalciferol is a vitamin D analog and a high-affinity vitamin D receptor agonist that inhibits NF-κB and is therefore a good candidate for treating metabolic syndrome.

[0003] The prevalence of metabolic syndrome is estimated to be at least 34% in the United States and at least 25% worldwide. The combined health care costs and economic losses from cardiovascular and musculoskeletal complications, cancer, and neurodegenerative diseases stemming from metabolic syndrome are estimated to be in the trillions of dollars (Van Saklayen, 2018).

[0004] The exact underlying causes of metabolic syndrome vary, with genetic and lifestyle factors contributing significantly. Metabolic syndrome develops when chronic excessive energy intake triggers an imbalance in energy metabolism, which leads to chronic inflammation that further exacerbates the underlying condition and the emergence of pathological conditions (Chawla et al., 2011).

[0005] Since activation of NF-κB signaling is central to the development of metabolic syndrome by driving the expression of proinflammatory cytokines (such as TNFα, IL-1β, IL-18, IL6), priming of the NRLP3 inflammasome, and the emergence of insulin resistance (Shi et al. 2006; Hotamisligil 2006; He et al. 2016; Hotamisligil 2017), NF-κB inhibition offers a potential therapeutic avenue to address metabolic syndrome (Baker et al. 2011).

[0006] Vitamin D is a steroid produced in the body or obtained from food. It is converted into an active agent that regulates intestinal calcium absorption and plays a role in the proper homeostasis of calcium and phosphate in the serum, which are essential for bone mineralization and bone growth and remodeling. Vitamin D has been found to play a role in the development and function of multiple organs, including the immune and nervous systems. Vitamin D acts as an agonist for a nuclear receptor (called the vitamin D receptor), which controls the expression of genes related to the regulation of calcium homeostasis and other cellular functions, such as inhibition of upregulated NF-κB signaling. Although the natural ligand for the vitamin D receptor is 1,25(OH)2D3, its role as a vitamin D receptor can be mimicked by a large number of synthetic vitamin D analogs. As with other receptors, agonists need to mimic the effects of the natural ligand on its cognate receptor but do not necessarily need to be structural analogs of the same chemical class as the natural agonist. The functional class of vitamin D agonists includes vitamin D analogs, but also includes (by way of example, not exclusion) compounds not classified as steroids, peptides, and nucleic acids.

[0007] Metabolic syndrome is becoming increasingly common, affecting up to one-third of US adults. Therefore, there is a need for effective treatments for metabolic syndrome. BRIEF DESCRIPTION OF THE DRAWINGS

[0008] Figure 1 Shown are the mean body weights (±SD) of the experimental animals in the elocalciferol group, the vitamin D3 group, and the vehicle group.

[0009] Figure 2 The mean relative weight loss or gain of experimental animals in the elocalcitol group, the vitamin D3 group, and the vehicle group are shown.

[0010] Figures 3A to 3B Shown are the mean body weight and weight gain data of the animals: comparison of the mean body weights of four groups of mice (LFD, HFD, HFD+vitD, and HFD+Eloc) after 16 weeks of administration of a low-fat or high-fat diet and concomitant 1,25(OH)2D3 or elocalciferol to HFD-fed mice, where the data are expressed as mean ± SEM (compared with the LFD group; ** p<0.01, **** p<0.001; compared with the HFD group, # p<0.05, ### p<0.001; one-way ANOVA, n=15 / group)( Figure 3A ); and the relative average weight gain kinetics of animals in the different experimental groups, wherein the data are expressed as mean ± SEM (HFD + vitD group compared with HFD group, * p<0.05, **p<0.01; HFD+Eloc group compared with HFD group, ## p<0.01; two-way RM ANOVA followed by Tukey's multiple comparison post hoc test, n=15 / group) Figure 3B ).

[0011] Figures 4A to 4C Fat distribution and changes in visceral fat volume are shown. Figure 4D Lean body mass calculated from the volume of back muscles is shown. Figure 4A The distribution of visceral and subcutaneous fat in MRI axial slices at the level of the kidneys is shown; Figure 4B shows changes in visceral fat volume after 16 weeks of LFD or HFD diet and 1,25(OH)2D3 or elocalciferol treatment; and Figure 4C Shown are changes in subcutaneous fat volume after 16 weeks of LFD or HFD diet and 1,25(OH)2D3 or elocalciferol treatment. Figure 4B and Figure 4C The values ​​in the table are expressed as mean ± SEM ( * p < 0.05; ** p < 0.01; *** p < 0.001; **** p<0.0001, two-way ANOVA followed by Tukey's multiple comparison post hoc test; n=5 animals / group).

[0012] 5A to 5D Comparison of time points is shown: At study week 16, each time point ( Figure 5A and Figure 5C ) and AUC 葡萄糖 ( Figure 5B and Figure 5D ) comparison. Figure 5A and Figure 5C The values ​​in the table are given as mean ± SEM (HFD vs. LFD groups, * p<0.05, ** p<0.01, *** p<0.001; HFD+vitD compared with LFD group, ¤¤¤ p<0.001, ¤¤ p<0.01; HFD+Eloc compared with HFD group, # p<0.05, ## p<0.01, ### p<0.001; two-way ANOVA for RM followed by Tukey's multiple comparison post hoc test; n=9-10 / group). Figure 5B and Figure 5D The values ​​in are given as mean ± SEM ( * p<0.05,** p<0.01, *** p<0.001, one-way ANOVA followed by Tukey's multiple comparison post hoc test; n=9-10 / group). DETAILED DESCRIPTION

[0013] The present invention relates to methods of treating metabolic syndrome using vitamin D receptor agonists.

[0014] metabolic syndrome

[0015] Metabolic syndrome is a group of conditions that occur together and increase the risk of heart disease, stroke, and type 2 diabetes. These conditions include excess body fat around the waist (visceral fat), abnormal cholesterol or triglyceride levels (dyslipidemia), high blood pressure (hypertension), insulin resistance or glucose intolerance, high blood sugar (hyperglycemia), obesity, atherosclerosis, diabetes, and hepatic steatosis. Metabolic syndrome is the result of chronic nutritional overload. According to the National Cholesterol Education Program (NCEP) Adult Treatment Panel III (ATP III; 2011) definition, metabolic syndrome is present if at least three or more of the following criteria are met:

[0016] (i) Visceral fat, which is reflected by a disproportionate amount of adipose tissue in and around the abdomen;

[0017] (ii) atherogenic dyslipidemia (high triglycerides (≥150 mg / dL), high LDL cholesterol, and low HDL cholesterol (≤50 mg / dL));

[0018] (iv) hypertension (≥130 / 85 mmHg); and insulin resistance or glucose intolerance (inability to use insulin or blood sugar correctly, respectively).

[0019] These metabolic syndrome conditions are accompanied by a chronic proinflammatory state, which is reflected in higher than normal levels of high-sensitivity C-reactive protein (CRP) in the blood (Ridker et al., 2003). People with metabolic syndrome are at increased risk for type 2 diabetes, coronary heart disease, heart failure, other diseases related to plaque buildup in arterial walls (e.g., stroke, myocardial infarction, and peripheral vascular disease), and nonalcoholic steatohepatitis (NASH), a leading cause of cirrhosis and hepatocellular carcinoma.

[0020] Vitamin D receptor agonists

[0021] Elocalcitol is a non-hypercalcemic vitamin D analog and high-affinity vitamin D receptor agonist that increases bone metabolism (Peleg et al., 2002) and has antiproliferative and anti-inflammatory effects. It inhibits NF-κB by blocking the nuclear translocation of the p65 subunit (Penna et al., 2009). Consequently, elocalcitol has been studied in humans as an experimental drug for a variety of indications, including overactive bladder, male infertility, chronic nonbacterial prostatitis, benign prostatic hyperplasia, and adult osteoporosis. Elocalcitol is among the vitamin D receptor agonists with the lowest risk of causing hypercalcemia (Nagopal et al., 2005) and is generally considered safe for long-term treatment in adults at oral daily doses of up to 300 μg (Montorsi et al., 2008). Disclosed herein are methods for treating metabolic syndrome using vitamin D receptor agonists. In some embodiments, one or more symptoms of metabolic syndrome can be improved by administering a vitamin D receptor agonist or a pharmaceutical composition comprising a vitamin D receptor agonist. Also provided is the use of a vitamin D receptor agonist in the preparation of a medicament for treating metabolic syndrome. In some embodiments, the vitamin D receptor agonist of the present invention is elocalciferol.

[0022] In some embodiments, the vitamin D receptor agonist useful in the present invention is a vitamin D analog, elocalcitol, or a pharmaceutically acceptable salt thereof. elocalcitol is a synthetic, biologically active vitamin D analog with modified side chains and ring A.

[0023]

[0024] Examples of some vitamin D receptor agonists suitable for use in the present invention include those of formulas 1-01 to 1-57; with 1-57 being preferred.

[0025]

[0026]

[0027]

[0028]

[0029] Another preferred vitamin D receptor agonist suitable for use in the present invention includes inecalcitol (which is the international nonproprietary name of 19-nor-9,10-secocyl-14pH-cholesta-5(Z),7(E)-diene-23-ino-1α,3β,25-triol-23-yne (Formula II)) or a pharmaceutically acceptable salt thereof:

[0030]

[0031] Metal salts can be generated by adding an inorganic base to the compounds described herein. Examples of suitable metals include lithium, sodium, potassium, cesium, cerium, magnesium, manganese, iron, calcium, strontium, cobalt, titanium, aluminum, copper, cadmium, and zinc. Examples of suitable metal salts include lithium, sodium, potassium, cesium, cerium, magnesium, manganese, iron, calcium, strontium, cobalt, titanium, aluminum, copper, cadmium, and zinc salts.

[0032] Examples of suitable ammonium salts include triethylamine salts, diisopropylamine salts, ethanolamine salts, diethanolamine salts, triethanolamine salts, morpholine salts, N-methylmorpholine salts, piperidine salts, N-methylpiperidine salts, N-ethylpiperidine salts, dibenzylamine salts, piperazine salts, pyridine salts, pyrazole salts, piperyrazole salts, imidazole salts, pyrazine salts, piperyrazine salts, ethylenediamine salts, N,N′-dibenzylethylenediamine salts, procaine salts, chloroprocaine salts, choline salts, dicyclohexylamine salts, and N-methylglucamine salts.

[0033] Methods for treating metabolic syndrome

[0034] Disclosed herein are methods for treating metabolic syndrome using vitamin D receptor agonists such as elocalcitol, inecalcitol, and Formulas I-01 to I-57. The methods comprise administering an effective amount of a vitamin D receptor agonist or a pharmaceutically acceptable salt thereof to a subject in need thereof. As used herein, an "effective amount" is an amount effective to treat metabolic syndrome by ameliorating the pathological condition or reducing the symptoms of metabolic syndrome. In some embodiments, the therapeutic effect can be determined by reducing excess body fat around the waist, lowering high cholesterol or triglyceride levels, lowering high blood pressure, lowering insulin resistance or glucose intolerance, lowering high blood sugar, and reducing weight in obese individuals.

[0035] Also disclosed herein are methods for providing prophylactic treatment of metabolic syndrome in a patient in need thereof, the methods comprising administering to the patient an effective amount of a vitamin D analog. In some embodiments, the effectiveness of the prophylactic treatment can be determined by fasting blood glucose levels, AUC insulin levels, fasting insulin levels, cholesterol levels, AUC glucose levels, triglyceride levels, and / or body weight.

[0036] Also disclosed herein is a method for treating pre-existing abnormal levels of fasting blood glucose, AUC insulin, fasting insulin, cholesterol, AUC glucose and / or body weight in a patient in need thereof, the method comprising administering an effective amount of a vitamin D receptor agonist to the patient. In some embodiments, abnormal levels can be determined by levels higher than those detected in healthy individuals. In certain other embodiments, abnormal levels of fasting blood glucose, AUC insulin, fasting insulin, cholesterol, AUC glucose and / or body weight are associated with metabolic syndrome. Methods for measuring the level or extent of fasting blood glucose, AUC insulin, fasting insulin, cholesterol, AUC glucose and / or body weight are well known in the art.

[0037] In some embodiments, the level or extent of fasting glucose, AUC insulin, fasting insulin, cholesterol, AUC glucose, and / or body weight is reduced by about 5% to about 10%, about 10% to about 20%, about 20% to about 30%, about 30% to about 40%, about 40% to about 50%, about 50% to about 60%, about 60% to about 70%, about 70% to about 80%, about 80% to about 90%, or about 90% to about 100%.

[0038] In some embodiments, metabolic disorders can be relevant to type 2 diabetes.In certain embodiments, metabolic syndrome can be by reducing visceral fat, fasting blood sugar, HbA1c, non-fasting blood sugar, improve insulin resistance and / or reduce weight gain / weight loss and regulate.The regulation of some blood sugar parameters (that is, fasting blood sugar level) can be determined using any suitable method (such as oral glucose tolerance test).

[0039] In some embodiments, the methods of the present invention can be administered, guided, and modified based on a personalized medicine approach. In some embodiments, personalized medicine provides a health care approach adapted to the needs of a specific subject relative to the methods of establishing medical cohorts and epidemiological studies and subsequently applying to individuals. In some embodiments, personalized medicine allows healthcare providers to optimize the therapy for a specific subject based on a variety of factors (e.g., genetics, metabolism, family history, personal history, environment, behavior, diet, lifestyle, social tendencies, and personal goals). In some embodiments, at any time before or during treatment, a healthcare provider can investigate any relevant factors and use the information obtained to design or improve a treatment plan.

[0040] Investigations can include assays as described herein or personal consultations conducted between a healthcare provider and the subject. In some embodiments, personalized medicine allows for the combination of therapy with companion diagnostic tests, such as genotyping of genes involved in energy metabolism (e.g., leptin, leptin receptors, genes involved in immune metabolism, or any other genes that contribute to the clinical phenotype of metabolic syndrome) to select patient populations that will benefit from treatment as described herein.

[0041] The subject of any of the therapeutic methods disclosed herein can be a subject in need of treatment for one or more of the conditions disclosed herein. The subject can be a human. Other non-limiting examples of subjects include non-human mammals, such as companion animals, pets, livestock, service animals, guard animals, working animals, and zoo animals.

[0042] In some embodiments, metabolic syndrome includes at least one condition selected from the group consisting of excess body fat around the waist (visceral fat), abnormal cholesterol or triglyceride levels (dyslipidemia), elevated blood pressure (hypertension), insulin resistance or glucose intolerance, high blood sugar (hyperglycemia), obesity, atherosclerosis, diabetes, and hepatic steatosis. In some embodiments, the condition is excess body fat around the waist (visceral fat). In some embodiments, the condition is abnormal cholesterol or triglyceride levels (dyslipidemia). In some embodiments, the condition is elevated blood pressure (hypertension). In some embodiments, the condition is high blood sugar (hyperglycemia). In some embodiments, the condition is diabetes. In some embodiments, the condition is obesity.

[0043] In some embodiments, the vitamin D receptor agonist is administered subcutaneously, orally, topically, transdermally, intradermally, parenterally, intravenously, intraarterially, intramuscularly, intrarectally, intraventricularly, intraspinally, intraperitoneally, intranasally, intramuscularly, sublingually, buccally, mucosally, by aerosol, or by suppository. In preferred embodiments, the vitamin D receptor agonist is administered subcutaneously or orally.

[0044] In some embodiments, the dosage of the vitamin D receptor agonist is administered at a dose of about 0.1 mg / day to about 10 mg / day. In some embodiments, the dosage is between about 0.1 mg / day and about 1 mg / day, administered orally. In some embodiments, the dosage is between about 0.1 mg / week and about 10 mg / week. In some embodiments, the dosage is a weekly sustained-release dosage, for example, by sustained-release subcutaneous injection. In some embodiments, the dosage is between about 0.1 mg / month and about 300 mg / month. In some embodiments, the dosage is a monthly sustained-release dosage, for example, by sustained-release subcutaneous injection. In some embodiments, the dosage is between about 0.1 mg / 6 months and about 2 g / 6 months.

[0045] In some embodiments, vitamin D receptor agonists such as ilocalcitol can be administered chronically at therapeutic doses without affecting calcium levels. In some embodiments, ilocalcitol has been administered to adults for extended periods of up to 150 μg oral daily doses and has been shown to be safe (Montorsi et al., 2008). At oral doses above 300 μg / day, hypercalcemia may occur, a common side effect of long-term use of vitamin D analogs.

[0046] In some embodiments, the method further comprises administering to the subject at least one additional therapeutic agent. In some embodiments, the at least one therapeutic agent is selected from the group consisting of a glucagon-like peptide-1 (GLP-1) receptor agonist, a glucose-dependent insulinotropic polypeptide (GIP) agonist, a glucagon agonist, an amylin agonist, a farnesoid X receptor (FXR) agonist, a liver X receptor (LXR) agonist, a melanocortin 4 receptor (MC4R) agonist, a peroxisome proliferator-activated receptor (PPAR) agonist, a thyroid hormone receptor-β (TRβ) agonist, a fibroblast growth factor 21 (FGF21) analog, an activin type II receptor (ActRII) blocker, a statin, a sodium glucose cotransporter 2 (SGLT2) inhibitor, and a dipeptidyl peptidase 4 (DPP4) inhibitor.

[0047] In some embodiments, the at least one therapeutic agent is selected from the group consisting of semaglutide, liraglutide, tilportide, canagliflozin, lixisenatide, exenatide, albiglutide, dulaglutide, rosiglitazone, empagliflozin, dapagliflozin, canagliflozin, orlistat, bimacilumab, resmetirol, pezofungin, semaglutide, and metformin. In embodiments, the therapeutic agent is semaglutide.

[0048] In combination therapy for metabolic syndrome using a vitamin D receptor analog of the present invention and another therapeutic agent, a standard dose of the other therapeutic agent is administered, for example, semaglutide is administered by weekly injection of 1 mg to 2.4 mg, and telpotide is administered by weekly injection of 2.5 mg to 15 mg.

[0049] In some embodiments, the subject has been diagnosed with metabolic syndrome. In embodiments, the subject has three or more conditions selected from the group consisting of: (i) visceral fat, which is reflected as a disproportionate amount of adipose tissue in and around the abdomen; (ii) atherogenic dyslipidemia (high triglycerides (≥150 mg / dL), high LDL cholesterol, and low HDL cholesterol (≤50 mg / dL)); (iv) hypertension (≥130 / 85 mmHg); and

[0050] (v) Insulin resistance or glucose intolerance (inability to use insulin or blood sugar correctly, respectively).

[0051] Disclosed herein is the use of a vitamin D receptor agonist in the preparation of a medicament for treating metabolic syndrome in a subject in need thereof.

[0052] Also disclosed herein is a vitamin D receptor agonist or a pharmaceutically acceptable salt thereof for use as a pharmaceutical compound in a method for preventing and / or treating metabolic syndrome in a subject. Also disclosed herein is a compound selected from the group consisting of compounds of Formula I (elocalcitol), Formula II (inecalcitol), and Formulas I-01 to I-57 for use in a method for treating, preventing, and / or reducing metabolic syndrome in a subject.

[0053] Pharmaceutical composition

[0054] Pharmaceutical compositions of vitamin D receptor agonists can be administered to a subject together with a pharmaceutical excipient or diluent. These compositions can take the form of drops, solutions, suspensions, tablets, pills, capsules, powders, sustained-release, controlled-release, or immediate-release formulations, as well as other formulations known in the art. The pharmaceutical compositions of the present invention can be adjusted using suitable excipients and diluents.

[0055] The pharmaceutical composition of the present invention can be formulated into a unit dosage form, with each dose containing, for example, about 0.01 mg to 10 g of the vitamin D receptor agonist.

[0056] In some embodiments, the unit dosage form is administered to humans, domestic pets, livestock, or other animals together with a pharmaceutically acceptable diluent or excipient. In some embodiments, administration is topical, parenteral, intravenous, intraarterial, intraventricular, intraperitoneal, intranasal, intramuscular, subcutaneous, aerosol, oral, or by suppository.

[0057] In some embodiments, the dosage of the pharmaceutical composition of the present invention varies according to the symptoms, age and weight of the experimenter, the nature and severity of the disease to be treated, the route of administration and the form of the composition. In some embodiments, the pharmaceutical composition of the present invention is administered in a single dose or divided doses.

[0058] The combined use of multiple compounds in the pharmaceutical compositions of the present invention can reduce the required dosage of any individual compound. In such combination therapy, the compounds can be delivered together or separately, simultaneously or at different times.

[0059] The pharmaceutical composition of the present invention can be administered in various ways known in the art. For oral administration, the pharmaceutical composition of the present invention can be formulated as tablets, capsules, granules, powders or syrups. The pharmaceutical composition of the present invention can be administered parenterally as an injection (intravenous, intramuscular or subcutaneous), an infusion formulation or a suppository.

[0060] The present invention is further illustrated by the following examples, which should not be construed as limiting the invention in any way.

[0061] Example

[0062] Example 1: Weight gain or loss in animals after administration of elocalcitol

[0063] Table 1 lists the abbreviations used in this article.

[0064] Table 1. Abbreviations

[0065]

[0066]

[0067] Two-month-old male C57BL / 6J mice (4-5 mice per cage) were housed at 24-26°C with a 12-hour light / dark cycle and free access to food and water. Three to four days before the start of the study, mice were randomly divided into three experimental groups (N = 10 / group) using an online randomization tool (Graph Pad, USA). The body weight of each animal was measured daily throughout the study, and their health status was assessed. Elocalcitol (100 μg / kg), vitamin D3 (cholecalciferol, 100 μg / kg), or vehicle control was administered intraperitoneally once daily for 6 days.

[0068] By day 4 of treatment, the average body weight of the elocalcitol group had decreased by 10.7%. The average weight loss in the elocalcitol group reached 18.6% by day 6 of treatment. Elocalcitol administration was stopped on day 6, and the average body weight of the mice returned to normal within two days. No weight loss was observed in either the vitamin D3 group or the vehicle control group. No adverse changes in the health of the mice were observed in any group.

[0069] Table 2. Mean body weight (±SD) and body weight of experimental animals in the elocalciferol group, vitamin D3 group, and vehicle group Relative weight gain / loss.

[0070]

[0071] Example 2. Prevention of Total Weight Gain

[0072] Six-week-old male C57BL / 6J mice were divided into four experimental groups (N = 15 / group) and fed with a low-fat diet (LFD) or a high-fat diet (HFD) that provided 10% (3, 61 kcal / kg) or 45% (4, 65 kcal / kg) of total energy as fat (the main fat source was lard; ssniff GmbH): (Group 1) mice fed with LFD, (Group 2) mice fed with HFD, (Group 3) mice fed with HFD and treated with 1,25(OH)2D3 (HFD+VitD), and (Group 4) mice fed with HFD and treated with elocalciferol (HFD+Eloc). Mice were group housed (4-5 mice per cage) at an appropriate temperature of 24°C-26°C with a 12-hour light / dark cycle, with free access to food and water. 1,25(OH)2D3 (1,25-dihydroxycholecalciferol, 15 μg / kg) or elocalciferol (15 μg / kg) was administered intraperitoneally to the HFD+VitD and HFD+Eloc groups twice a week for 16 weeks, respectively, while the LFD and HFD groups were administered with vehicle solution. Body weight was measured once a week, and the amount of food and water consumed by the animals was measured once a month.

[0073] The average weight gain of mice in the HFD group was significantly higher than that of mice in the LFD group at week 16 of the study ( Figure 3A Compared with vehicle-treated HFD mice, elocalcitol and 1,25(OH)2D3 treatment resulted in 20% and 10% lower relative body weight gain, respectively ( Figure 3A and Figure 3B Compared to mice treated with 1,25(OH)2D3, inhibition of HFD-induced weight gain by elocalcitol reached statistical significance much earlier (study week 12 vs. study week 7, respectively) ( Figure 3B ).

[0074] Example 3. Normalization of visceral and subcutaneous fat volume

[0075] Six-week-old male C57BL / 6J mice were divided into four experimental groups (N = 15 / group) and received either a low-fat diet (LFD) or a high-fat diet (HFD) (the main fat source was lard; ssniff) that provided 10% (3, 61 kcal / kg) or 45% (4, 65 kcal / kg) of total energy as fat. GmbH): (Group 1) mice fed an LFD, (Group 2) mice fed an HFD, (Group 3) mice fed an HFD and treated with 1,25(OH)2D3 (HFD+VitD), and (Group 4) mice fed an HFD and treated with elocalciferol (HFD+Eloc). Mice were group-housed (4-5 mice per cage) at a controlled temperature of 24°C-26°C with a 12-hour light / dark cycle, with free access to food and water. 1,25(OH)2D3 (1,25-dihydroxycholecalciferol, 15 μg / kg) or elocalciferol (15 μg / kg) was administered intraperitoneally twice weekly to the HFD+VitD and HFD+Eloc groups, respectively, for 16 weeks, while the LFD and HFD groups were administered vehicle solution.

[0076] Before the experiment and once a month thereafter during the experiment, the abdominal area of ​​the animals was scanned to assess visceral and subcutaneous fat load using a magnetic resonance imaging system (MRI, Bruker BioSpin Group, Bruker Corporations, Germany). The study procedure was performed as follows: Mice (n = 5 / group) were anesthetized with isoflurane (1.5%-2.5% in 1.5 1 / min medical oxygen) and placed on a heated animal bed throughout the MRI procedure. The MRI was performed using a 1H circularly polarized transceiver coil connected to a ParaVision Scans were performed on a 9.4T Bruker BioSpec 94 / 21USR system using the software (Bruker BioSpin Group, Bruker Corporation, Germany). Respiration was monitored using a breathing pillow (SA Instruments Inc., Stony Brook, USA), and the respiratory rate was maintained between 35 and 70 breaths per minute. Two directional guidance scans were performed to determine the position of the animal and identify anatomical landmarks relevant to the planned subsequent scans. The final T1-weighted Bruker: RARE sequence was performed using the following parameters: repetition time (TR) 1164 ms, echo time (TE) 6 ms, flip angle 90 degrees, average number 2, imaging matrix 320×320×40, spatial resolution 0.125×0.125×0.5 mm. Volumes were manually segmented using ITK-SNAP (V3.8.0) by an observer who was blinded to the experiment. Visceral and subcutaneous fat volumes were measured using 40x zoom, and the kidneys were the starting point for fat volume analysis.

[0077] Representative images of visceral and subcutaneous fat volume analysis by MRI in Figure 4 show that at study week 16, HFD resulted in significantly higher visceral fat levels compared with LFD ( ****p < 0.0001, two-way ANOVA followed by Tukey's multiple comparison post hoc test, n = 5 / group). Treatment with elocalciferol resulted in a significant decrease in visceral and subcutaneous fat levels in the HFD+Eloc group compared with the HFD group (Figure 4; A, B, and C). In fact, elocalciferol treatment reversed the visceral and subcutaneous fat levels in HFD mice to the levels observed in LFD mice ( Figure 4A 、 Figure 4B and Figure 4C 1,25(OH)2D3 treatment had no statistically significant effect on either visceral fat volume or subcutaneous fat distribution in HFD-fed mice ( Figure 4A and Figure 4B Lean body mass, calculated from back muscle volume, was maintained ( Figure 4D Analysis of epididymal fat deposit weight at the end of the study confirmed the MRI results, showing that the weight of fat deposits in the LFD and HFD+Eloc groups were significantly lower than those in the HFD and HFD+vitD groups (Table 3).

[0078] Table 3. Effects of diet, 1,25(OH)2D3, and elocalciferol on organ weights. Absolute organ weight (mg) and Organ weights were normalized (organ weight / body weight, mg / g).

[0079]

[0080]

[0081] *Data are expressed as mean ± SEM (compared with LFD, * p<0.05, **** p<0.0001; HFD+vitD compared with HFD+Eloc, ¤¤ p<0.01; HFD vs. HFD+Eloc, ## p<0.01, ### p<0.001; n=10 / group, one-way ANOVA followed by Tukey's multiple comparison post hoc test).

[0082] Example 4. Reversal of glucose intolerance and insulin resistance

[0083] Six-week-old male C57BL / 6J mice were divided into four experimental groups (N = 15 / group) and received either a low-fat diet (LFD) or a high-fat diet (HFD) (the main fat source was lard; ssniff) that provided 10% (3, 61 kcal / kg) or 45% (4, 65 kcal / kg) of total energy as fat. GmbH): (Group 1) mice fed an LFD, (Group 2) mice fed an HFD, (Group 3) mice fed an HFD and treated with 1,25(OH)2D3 (HFD+VitD), and (Group 4) mice fed an HFD and treated with elocalciferol (HFD+Eloc). Mice were group-housed (4-5 mice per cage) at a controlled temperature of 24°C-26°C with a 12-hour light / dark cycle, with free access to food and water. The HFD+VitD and HFD+Eloc groups were administered 1,25(OH)2D3 (1,25-dihydroxycholecalciferol, 15 μg / kg) or elocalciferol (15 μg / kg) intraperitoneally (ip), while the LFD and HFD groups were administered vehicle solution twice a week for 16 weeks.

[0084] To determine the effects of elocalciferol and 1,25(OH)2D3 on hyperglycemia and insulin sensitivity in mice, a glucose tolerance test (GTT) was performed at baseline and once a month during the study. For the GTT, mice were fasted overnight for 10-12 hours, and blood glucose was measured in tail blood using an Accu-Chek Performa system glucose meter (Roche, Germany), and the baseline glucose value (0 minute) was recorded. The animals were then injected intraperitoneally with glucose (2 g / kg), and after the glucose injection, blood glucose in tail blood was measured according to this schedule: 15 minutes, 30 minutes, 60 minutes, 90 minutes, and 120 minutes.

[0085] At the end of the study (research the 16th week), insulin tolerance test (ITT) was carried out. For ITT, mice were fasted for 6 hours, and Accu-Chek Performa system glucose meter (Roche, Germany) was used to measure the blood glucose in the tail blood, and baseline glucose value was recorded (0 minute). Animals were then injected with insulin intraperitoneally (0,5 U / kg; diluted with saline 0,9% from 100 U / ml (Novorapid, Novo Nordisk A / S)), and after the glucose injection, the blood glucose in the tail blood was measured according to the schedule: 15 minutes, 30 minutes, 60 minutes, 90 minutes and 120 minutes.

[0086] At study week 16, following glucose administration (2 g / kg, ip), HFD-fed mice exhibited significantly impaired glucose tolerance compared with LFD controls, as determined by comparison of individual time points and glucose area under the curve analysis ( Figure 5A and Figure 5BGlucose area under the curve (glucose AUC) analysis, an indicator of the entire glucose excursion after a glucose load, showed that treatment with elocalciferol, but not 1,25(OH)2D3, resulted in a statistically significant improvement in glucose tolerance in HFD-fed mice ( Figure 5B Results of the insulin tolerance test showed that treatment with elocalcitol, but not 1,25(OH)2D3, reversed the increased insulin resistance observed in the HFD in a statistically significant manner ( Figure 5C and Figure 5D ).

[0087] Example 5. Normalization of triglyceride and cholesterol levels

[0088] Six-week-old male C57BL / 6J mice were divided into four experimental groups (N = 15 / group) and received either a low-fat diet (LFD) or a high-fat diet (HFD) (the main fat source was lard; ssniff) that provided 10% (3, 61 kcal / kg) or 45% (4, 65 kcal / kg) of total energy as fat. GmbH): (Group 1) mice fed with LFD, (Group 2) mice fed with HFD, (Group 3) mice fed with HFD and treated with 1,25(OH)2D3 (HFD+VitD), and (Group 4) mice fed with HFD and treated with elocalciferol (HFD+Eloc). Mice were group-housed (4-5 mice per cage) at an appropriate temperature of 24°C-26°C with a 12-hour light / dark cycle, with free access to food and water. 1,25(OH)2D3 (1,25-dihydroxycholecalciferol, 15 μg / kg) or elocalciferol (15 μg / kg) was administered intraperitoneally to the HFD+VitD and HFD+Eloc groups, while the LFD and HFD groups were administered vehicle solution twice a week for 16 weeks.

[0089] 24 hours after the last study treatment administration, mice were deeply anesthetized with a final dose of phenobarbital (200 mg / kg-300 mg / kg), and blood samples were collected from mice of all groups via cardiac puncture using a standardized protocol. The collected blood was allowed to clot and then centrifuged at 2000 × g for 20 minutes, after which the serum was stored at -80°C until analysis.

[0090] Triglyceride levels were significantly increased in the HFD group compared to the LFD group (p < 0.05, one-way ANOVA followed by Tukey's multiple comparison post hoc test, Table 3). Treatment with both elocalcitol and 1,25(OH)2D3 reduced triglyceride levels similar to those in the LFD group, while treatment with elocalcitol had a statistically significant cholesterol (combined HDL and LDL) lowering effect (Table 4).

[0091] Table 4. Blood metabolism in mice in the LFD, HFD, HFD+VitD, and HFD+Eloc groups at week 16 of the study landmark.

[0092]

[0093] Example 6. Biphasic Treatment for Weight Loss Using Variable Strength Sustained-Release Elocalcitol Formulations

[0094] Subjects experiencing symptoms of metabolic disease and diagnosed with obesity will be prescribed a monotherapy of a vitamin D receptor agonist, wherein the vitamin D receptor agonist is elocalciferol. Initially, elocalciferol is administered in the form of a pre-filled syringe via weekly subcutaneous injection. The pre-filled syringe contains a sterile liquid sustained-release formulation of elocalciferol in a polar lipid phase consisting of a mixture of phospholipids, diacylglycerols, ethanol, and a buffer, thereby producing a daily effective dose of approximately 0.6 mg to 1.2 mg of elocalciferol. The initial treatment with elocalciferol will last for up to six months. Thereafter, elocalciferol is administered in the form of a pre-filled syringe via quarterly subcutaneous injection. The pre-filled syringe contains a sterile liquid sustained-release formulation of elocalciferol in a polar lipid phase consisting of a mixture of phospholipids, diacylglycerols, ethanol, and a citrate buffer, or an in situ polymer gel extended delivery system, thereby producing a daily effective dose of approximately 0.3 mg of elocalciferol. As long as the therapeutic efficacy is detectable, the therapy will be continuous. Subjects are monitored throughout therapy to determine efficacy of treatment, minimization of side effects (eg, hypercalcemia), and any need for changes in dosage or prescribed treatment regimen.

[0095] Example 7. Weight Loss Using a Variable Strength Sustained-Release Elocalcitol Formulation in Combination with a GLP-1 Agonist Phase Treatment

[0096] Subjects experiencing symptoms of metabolic disease and diagnosed with obesity will be prescribed a monotherapy with a vitamin D receptor agonist, wherein the vitamin D receptor agonist is elocalcitol. Elocalcitol will initially be administered weekly subcutaneously in a prefilled syringe containing a sterile liquid sustained-release formulation of elocalcitol in a polar lipid phase consisting of a mixture of phospholipids, diacylglycerols, ethanol, and buffer, resulting in the release of an effective daily dose of approximately 0.6 mg to 1.2 mg of elocalcitol. Simultaneously, subjects will be treated with weekly subcutaneous injections of 2.4 mg of semaglutide. Initial treatment with this combination of elocalcitol and semaglutide will continue for up to six months. Thereafter, ilocalcitol is administered by subcutaneous injection quarterly in the form of a pre-filled syringe containing a sterile liquid sustained-release formulation of ilocalcitol in a polar lipid phase consisting of a mixture of phospholipids, diacylglycerols, ethanol, and citrate buffer or an in situ polymer gel extended delivery system, resulting in the release of a daily effective dose of approximately 0.3 mg of ilocalcitol. Therapy will be continued as long as therapeutic efficacy is detectable. Subjects are monitored throughout therapy to determine therapeutic efficacy, minimized side effects (e.g., hypercalcemia), and any need to change the dose or prescribed treatment regimen.

[0097] Example 8. Biphasic Treatment of Glucose Intolerance Using a Variable Strength Sustained-Release Elocalcitol Formulation

[0098] Subjects experiencing symptoms of metabolic disease and diagnosed with type 2 diabetes will be prescribed a monotherapy of a vitamin D receptor agonist, wherein the vitamin D receptor agonist is elocalciferol. Initially, elocalciferol is administered weekly in the form of a pre-filled syringe containing a sterile liquid sustained-release formulation of elocalciferol in a polar lipid phase consisting of a mixture of phospholipids, diacylglycerols, ethanol, and a buffer, thereby producing a daily effective dose of approximately 0.6 mg to 1.2 mg of elocalciferol. The initial treatment with elocalciferol will last for up to six months. Thereafter, elocalciferol is administered quarterly in the form of a pre-filled syringe containing a sterile liquid sustained-release formulation of elocalciferol in a polar lipid phase consisting of a mixture of phospholipids, diacylglycerols, ethanol, and a citrate buffer, or an in situ polymer gel extended delivery system, thereby producing a daily effective dose of approximately 0.3 mg of elocalciferol. As long as the therapeutic efficacy is detectable, the therapy will be continuous. Subjects are monitored throughout therapy to determine efficacy of treatment, minimization of side effects (eg, hypercalcemia), and any need for changes in dosage or prescribed treatment regimen.

[0099] Exemplary embodiments

[0100] In one embodiment, disclosed herein is a method of treating metabolic syndrome in a subject or reducing the effects of metabolic syndrome in a subject, the method comprising: administering to a subject in need thereof an effective amount of a vitamin D receptor agonist or a pharmaceutically acceptable salt thereof, wherein the vitamin D receptor agonist is selected from the group consisting of elocalciferol, inacalcitol, I-01, I-02, I-03, I-04, I-05, I-06, I-07, I-08, I-09, I-10, I-11, I-12, I-13, I-14, I-15, I-16, I-17, I-18, I-19, I-20, I-21, I-22, I-23, I-24, I-25, I-26, I-27, I-28, I-29, I-30, I-31, I-32, I-33, I-34, I-35, I-36, I-37, I-38, I-39, I-40, I-41, I-42 , I-43, I-44, I-45, I-46, I-47, I-48, I-49, I-50, I-51, I-52, I-53, I-54, I-55, I-56 and I-57 compounds, and wherein the effective amount of the vitamin D receptor agonist does not exceed the threshold for inducing hypercalcemia.

[0101] In one embodiment, the vitamin D receptor agonist is elocalcitol, inecalcitol, or formula 1-57.

[0102] In one embodiment, metabolic syndrome includes at least one condition selected from the group consisting of excess body fat around the waist (visceral fat), abnormal cholesterol or triglyceride levels (dyslipidemia), elevated blood pressure (hypertension), insulin resistance or glucose intolerance, high blood sugar (hyperglycemia), obesity, atherosclerosis, diabetes, and hepatic steatosis.

[0103] In one embodiment, the condition is excess body fat around the waist (visceral fat).

[0104] In one embodiment, the disorder is abnormal cholesterol or triglyceride levels (dyslipidemia).

[0105] In one embodiment, the disorder is elevated blood pressure (hypertension).

[0106] In one embodiment, the disorder is high blood sugar (hyperglycemia).

[0107] In one embodiment, the disorder is diabetes.

[0108] In one embodiment, the disorder is obesity.

[0109] In one embodiment, the threshold value is less than or equal to 1,200 μg / day.

[0110] In one embodiment, the threshold is greater than or equal to about 150 μg / day and less than or equal to about 1,200 μg / day.

[0111] In one embodiment, the vitamin D receptor agonist is topical, transdermal, intradermal, parenteral, intravenous, intraarterial, subcutaneous, intramuscular, intracranial, intrarectal, intraorbital, ocular, intraventricular, intracapsular, intraspinal, intracranial, intraperitoneal, intranasal, intramuscular, subcutaneous,

[0112] Administered sublingually, buccally, mucosally, by aerosol, orally, or by suppository.

[0113] In one embodiment, the vitamin D receptor agonist is administered subcutaneously.

[0114] In one embodiment, the method further comprises administering to the subject one or more pharmaceutically acceptable excipients.

[0115] In one embodiment, the method further comprises administering to the subject at least one therapeutic agent or a pharmaceutically acceptable salt thereof.

[0116] In one embodiment, the at least one therapeutic agent is selected from the group consisting of a glucagon-like peptide-1 (GLP-1) receptor agonist, a glucose-dependent insulinotropic polypeptide (GIP) agonist, a glucagon agonist, an amylin agonist, a farnesoid X receptor (FXR) agonist, a liver X receptor (LXR) agonist, a melanocortin 4 receptor (MC4R) agonist, a peroxisome proliferator-activated receptor (PPAR) agonist, a thyroid hormone receptor-β (TRβ) agonist, a fibroblast growth factor 21 (FGF21) analog, an activin type II receptor (ActRII) blocker, a statin, a sodium glucose cotransporter 2 (SGLT2) inhibitor, and a dipeptidyl peptidase 4 (DPP4) inhibitor.

[0117] In one embodiment, the at least one therapeutic agent is selected from the group consisting of semaglutide, liraglutide, tilportide, canagliflozin, lixisenatide, exenatide, albiglutide, dulaglutide, rosiglitazone, empagliflozin, dapagliflozin, canagliflozin, orlistat, bimacilumab, resmetirol, pezofungin, semaglutide, and metformin.

[0118] In one embodiment, at least one therapeutic agent is administered to a subject at a dose of less than or equal to about 1000 milligrams (mg).

[0119] In one embodiment, the vitamin D receptor agonist is administered to a subject at a dose of less than or equal to about 1000 milligrams (mg).

[0120] In one embodiment, the subject has metabolic syndrome.

[0121] References

[0122] Baker RGet al.,CellMetab.2011;13(1):11-22.

[0123] Chawla A. et al., Nat. Rev. Immunol. 2011; 11(11): 738-749. He Y. et al., Trends Biochem. Sci. 2016; 41(12): 1012-1021. Hotamisligil GS, Immunity 2017; 47(3): 406-420.

[0124] Hotamisligil GS,Nature 2006;444(7121):860-867.

[0125] Montorsi F.et al.,J.Urol.2008;179(suppl):700;Abstract 2035.NagpalS.etal.,Endocrine Rev.2005;26(5):662-687.

[0126] National Cholesterol Education Program (NCEP) Adult Treatment Panel III (ATP III), JAMA 2001; 285(19):2486-2497.

[0127] Peleg S. et al., Endocrinology 2002;143(5):1625-1636.

[0128] Penna G.et al.,Prostate 2009;69(5):480-493.

[0129] Ridker PM et al.,Circulation 2003;107(3):391-397.

[0130] Shi H.et al.,J.Clin.Invest.2006;116(11):3015-3025.

[0131] Van Saklayen MG, Curr. Hypertens. Rep. 2018;20(2):12.

Claims

1. A method of treating metabolic syndrome in a subject, the method comprising: administering an effective amount of a vitamin D receptor agonist or a pharmaceutically acceptable salt thereof to a subject in need thereof, Wherein the vitamin D receptor agonist is selected from the group consisting of: allocalcidol, inecalcidol, formula I-01, I-02, I-03, I-04, I-05, I-06, I-07, I-08, I-09, I-10, I-11, I-12, I-13, I-14, I-15, I-16, I-17, I-18, I-19, I-20, I-21, I-22, I-23, I-24, I-25, I -26, I-27, I-28, I-29, I-30, I-31, I-32, I-33, I-34, I-35, I-36, I-37, I-38, I-39, I-40, I-41, I- 42. Compounds of I-43, I-44, I-45, I-46, I-47, I-48, I-49, I-50, I-51, I-52, I-53, I-54, I-55, I-56 and I-57.

2. The method of claim 1, wherein the vitamin D receptor agonist is elocalciferol.

3. The method of claim 1, wherein the vitamin D receptor agonist is inecalcitol.

4. The method of claim 1, wherein the vitamin D receptor agonist is Formula 1-57.

5. The method according to any one of claims 1 to 4, wherein the metabolic syndrome comprises at least one condition selected from the group consisting of excess body fat around the waist, abnormal cholesterol or triglyceride levels, elevated blood pressure, insulin resistance or glucose intolerance, hyperglycemia, obesity, atherosclerosis, diabetes, and hepatic steatosis.

6. The method of claim 5, wherein the condition is excess body fat around the waist.

7. The method of claim 5, wherein the condition is abnormal cholesterol or triglyceride levels.

8. The method of claim 5, wherein the condition is elevated blood pressure.

9. The method of claim 5, wherein the condition is hyperglycemia.

10. The method of claim 5, wherein the condition is diabetes.

11. The method of claim 5, wherein the condition is obesity.

12. The method of any one of claims 1 to 11, wherein the vitamin D receptor agonist is administered orally or subcutaneously.

13. The method according to any one of claims 1 to 12, further comprising administering at least one therapeutic agent selected from the group consisting of semaglutide, liraglutide, tilportide, canagliflozin, lixisenatide, exenatide, albiglutide, dulaglutide, rosiglitazone, empagliflozin, dapagliflozin, canagliflozin, orlistat, bimacilumab, resmetirol, pezofungin, semaglutide, and metformin.

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