Combining bitter taste receptor agonists and gut signaling compounds

Abstract

A combination oral dosage form pharmaceutical composition is disclosed that includes a bitter taste receptor agonist and a gut signaling compound, i.e., a gut signaling peptide analog and / or a gut signaling hormone enhancer.And a method for treating obesity, diabetes, metabolic syndrome, glycemic control hyperlipidemia and achieving weight loss is disclosed, which comprises administering an effective amount of a pharmaceutical composition that includes a bitter taste receptor agonist and a gut signaling compound, i.e., a gut signaling peptide analog and / or a gut signaling hormone enhancer as described above and herein.Furthermore, a method for preventing the progression and / or treating fatty liver disease is disclosed, which comprises administering an effective amount of a combination that includes a bitter taste receptor agonist, which includes a denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate; and a GLP-1 receptor agonist.

Description

[Technical field]

[0001] (CROSS REFERENCE TO RELATED APPLICATIONS) This application claims the benefit of priority to U.S. Provisional Application No. 63 / 180,224, filed April 27, 2021; U.S. Provisional Application No. 63 / 229,499, filed August 4, 2021; U.S. Provisional Application No. 63 / 245,925, filed September 19, 2021; and U.S. Provisional Application No. 63 / 305,037, filed January 31, 2022, each of which is incorporated by reference herein in its entirety for all purposes.

[0002] FIELD OF THEINVENTION The present disclosure provides a combination of bitter taste receptor agonist (alternatively referred to as TAS2R or T2R agonist) and at least one gut signaling compound selected from gut signaling peptide analogs and / or gut signaling hormone enhancers.Also provided is the therapeutic use of such combination for treating, for example, glucagon-related diseases, disorders and conditions as defined herein, including, for example, diabetes, pre-diabetes syndrome, obesity, weight and / or appetite control, hyperlipidemia and hyperglycemia.The present disclosure further provides a method for preventing the progression of, or treating, fatty liver disease, in particular, comprising administering a combination comprising bitter taste receptor agonist and GLP-1 receptor agonist. [Background technology]

[0003] (Introduction) Global obesity has more than doubled in the past 40 years. Obesity predisposes to metabolic syndrome and is associated with coronary heart disease, stroke, type 2 diabetes, and certain forms of cancer. Obesity is associated with increased risk of serious illness and increased risk of death from coronavirus, one of the most significant current global health challenges. Parallel to the emergence of this problem is an increasing understanding of the pathological mechanisms that link the obese state to disease development. Central to these mechanisms is the state of increased systemic inflammation resulting from obesity, which leads to numerous pathologies. Thus, there is a great need for treatments and preventative measures that address appetite and inflammatory signals and optimize metabolism. The present disclosure addresses this need and provides other benefits.

[0004] According to the HIS Division of Diabetes Treatment and Prevention (last updated April 2021), if a patient has metabolic syndrome and is overweight, the individual is at higher risk of developing type 2 diabetes. In general, the initial treatment algorithm for treating such patients (measured by fasting glucose and HbA1c for glycemic control) is to treat with oral agents before moving to injectable agents, which can be implemented when signs and symptoms indicate inadequate glycemic control.

[0005] For example, overweight patients presenting as diabetes or prediabetes can typically be treated by (1) starting with generic metformin; (2) if not adequately treated, then adding an oral DPP-4 inhibitor or using a combination metformin / DPP-4 inhibitor oral formulation; (3) if still not adequately treated, increasing the dose of metformin and DPP-4 inhibitor; and finally (4) if not adequately treated, switching to injectable insulin. There are also branded GLP-1 and GIP analogs, primarily for injection, labeled to lower HbA1c and currently undergoing clinical trials for weight loss. Both GLP-1 and GIP analogs, alone and in combination, are dual incretin peptidomimetic compounds that stimulate receptors for both human GIP and GLP-1. However, with the exception of Novo Nordisk's Ryvelsus® (semaglutide), an oral GLP-1 analog, most GLP-1 and GIP analogs are for injection. However, as published by Wadden et al. (JAMA 2021:325(14):1403-1413, published online February 24, 2021) and Aroda et al. ("PIONEER 1: Randomized Clinical Trial of the Efficacy and Safety of Oral Semaglutide Monotherapy in Comparison with Placebo in Patients with Type 2 Diabetes" Diabetes Care 43:1724-1732, 2019), the efficacy of oral semaglutide appears to be significantly reduced versus an injectable version of the same GLP-1 analogue.

[0006] Type 2 diabetes and weight loss DPP-4 inhibitors (DPP-4i) inhibit dipeptidyl peptidase-4 (DPP-4), thus resulting in an increase in endogenous incretin levels (including GLP-1 and GIP). DPP-4i are a class of effective oral therapeutics for treating diabetes, and sitagliptin (Januvia®) is a representative drug of this class. However, despite their usefulness, the usefulness of DPP-4i has been found to be limited by their ability to combat obesity. Although relatively little weight loss has been observed in obese patients taking DPP-4i drugs, their effects are often limited and temporary, possibly due to increased resistance to DPP-4i drugs through long-term and repeated exposure.

[0007] An 18-week trial of 800 patients with poorly controlled type 2 diabetes mellitus (T2DM) taking metformin, saxagliptin 5 mg daily versus sitagliptin 100 mg daily showed similar reductions in hemoglobin A1c (HbA1c) (-0.52 vs. -0.26%) (Scheen et al., "Efficacy and safety of saxagliptin in combination with metformin compared with sitagliptin in combination with metformin in adult patients with type 2 diabetes mellitus." Diabetes Metab. Res. Rev. (2010) 26:540-9. doi: 10.1002 / dmrr.1114). The risk of hypoglycemia with DPP-4 inhibitors is low given their GLP-1-mediated glucose-dependent mechanism of action.

[0008] GLP-1 receptor agonists (GLP-1 RAs) are peptide derivatives of either exendin-4 or human GLP-1 designed to resist the activity of DPP-4, and therefore have an extended half-life. In clinical trials, GLP-1 RAs have shown efficacy, improved weight loss and a low risk of hypoglycemia. However, GI adverse events, particularly nausea, vomiting and diarrhea, have been observed, as well as a strict black box warning for thyroid cancer.

[0009] Several clinical trials have directly compared the efficacy and safety of DPP-4 inhibitors and GLP-1 RAs. These trials have generally shown that GLP-1 RAs provide superior glycemic control and weight loss compared to DPP-4 inhibitors. Both treatments were associated with low and comparable incidences of hypoglycemia, while treatment with GLP-1 RAs was associated with a higher incidence of adverse events. According to current clinical guidelines, both GLP-1 RAs and DPP-4 inhibitors are indicated for glycemic control in T2DM patients across the disease spectrum. GLP-1 RAs may be preferable to DPP-4 inhibitors for many patients due to the greater reduction in hemoglobin A1c and weight loss observed in clinical trials. Thus, there is a need for better combinations with DPP-4 inhibitors for weight loss, without the severe side effects of GLP-1 agonists, given the better side effect profile.

[0010] Thus, there is a need in the art to delay the progression of type 2 diabetes to the need for insulin treatment or to delay or prevent the progression to a state requiring insulin treatment. This need can be addressed with the invention described herein, which can provide the surprising beneficial effects of (1) increasing the maximum effect of GLP-1 RA and / or GIP analogs with combined orally active agents (e.g., in some embodiments, weight loss to 10-15% of body weight) to enhance the treatment effectiveness of glycemic control or to delay the progression to insulin, and / or (2) improving the efficacy of DPP-4 inhibitors to be at least equal to or better than GLP-1 analogs (or combinations of GLP-1 analogs and GIP analogs), providing an oral administration alternative to injections.

[0011] Obesity and weight loss Obesity, defined in general terms as excess body fat relative to lean body mass, is now a worldwide epidemic and one of the most serious contributors to rising morbidity and mortality. Obesity is epidemic in the United States, affecting more than 61% of the total population (Flegal et al., Int. J. Obes. 22:39-47,1998). Obesity is more specifically defined by the Centers for Disease Control and Prevention (CDC) as an excessive amount of body fat or adipose tissue relative to lean body mass, and overweight is defined as an increase in weight for height compared to some standard of acceptable or desirable weight. Alternatively, the CDC defines overweight as a person with a body mass index (BMI) between 25.0 and 29.9, and obesity as a BMI of 30.0 or greater. Obesity is often associated with psychological and medical morbidity, the latter including increased joint problems, vascular diseases such as coronary artery disease, hypertension, stroke, and peripheral vascular disease. Obesity also leads to metabolic disorders such as insulin resistance and type II diabetes (non-insulin-dependent diabetes mellitus (NIDDM)), hyperlipidemia and endothelial dysfunction. These disorders predispose the vasculature to damage, cell proliferation and lipid oxidation, resulting in atherosclerosis leading to heart attacks, strokes and peripheral vascular disease. In 1998, consumers spent $33 billion in the United States on unsuccessful weight loss products and services (Serdula et al., JAMA 282:1353-1358,1999). Thus, obesity and its associated complications remain a major problem for health care systems worldwide.

[0012] Obesity is a significant clinical problem with widespread impact. Approaches are limited to diet and exercise (therapeutic lifestyle changes), surgery such as gastric bypass, and pharmacological agents including GLP-1 receptor agonists. Drug treatments of obesity have been disappointing because almost all drug treatments are associated with undesirable side effects that have contributed to their cessation and / or result in discontinuation of treatment and / or exhibit an unacceptable risk / benefit profile with very limited chances of success. Several monoamines and neuropeptides reduce food intake (Bray et al., Am. J. Clin. Nutr. 55:151S-319S, 1992). Available drug therapies include sibutramine (an appetite suppressant), orlistat (a lipase inhibitor), and the sympathomimetics fenfluramine and dexfenfluramine. Although weight loss is effective, sympathomimetics cause side effects including pulmonary hypertension, neuroanatomical changes, and atypical valvular heart disease. For example, fenfluramine and dexfenfluramine were withdrawn from the market in 1997 due to associated valvular heart disease. Therefore, nutritional and dietary restriction is the most desirable approach for weight loss. However, dietary restrictions have a low long-term success rate due to non-compliance.

[0013] Thus, there is no ideal treatment based on the biology of the primary metabolic abnormalities found in obesity and its related conditions, such as metabolic syndrome or atherosclerosis. Thus, there remains a need for new compositions and methods that address the treatment of individuals suffering from obesity and obesity-related disorders.

[0014] hyperlipidemia Hypercholesterolemia is a well-known risk factor for atherosclerotic cardiovascular disease (ASCVD), the leading cause of mortality in Western countries. Epidemiological studies have shown that pharmacological lowering of total cholesterol (TC) and low-density lipoprotein (LDL) cholesterol (LDL-C) is associated with a reduction in clinical cardiovascular events.

[0015] Triglycerides (TG) are a common type of fat (lipid) that, when present in normal amounts, is essential for good health. Higher than normal triglyceride levels are often associated with known risk factors for heart disease, such as obesity, low levels of high-density lipoprotein (HDL) ("good") cholesterol, and high levels of low-density lipoprotein (LDL) ("bad") cholesterol. Triglycerides may also contribute to the thickening of artery walls, a physical change that is considered a predictor of atherosclerosis. Thus, high triglyceride levels are at least a warning sign that a patient's heart health may be at risk.

[0016] Many treatments are currently available to lower serum cholesterol and triglycerides. However, each has its own drawbacks and limitations in terms of efficacy, side effects, and patient population qualification. Bile acid-binding resins, such as cholestyramine (Questran Light®, Bristol-Myers Squibb) and colestipol hydrochloride (Colestid®, The Upjohn Company), are a class of drugs that prevent the recycling of bile acids from the intestine to the liver. However, the use of such resins at best only lowers serum cholesterol levels by about 20% and is associated with gastrointestinal side effects, including constipation and certain vitamin deficiencies. Furthermore, because the resins bind to other drugs, other oral medications must be taken at least 1 hour before or 4-6 hours after ingestion of the resin, thus complicating the drug regimen of cardiac patients.

[0017] Statins are cholesterol-lowering drugs that block cholesterol synthesis by inhibiting HMGCoA reductase, a key enzyme involved in the cholesterol biosynthesis pathway.Statins, such as lovastatin (Mevacor®, Merck & Co., Inc.), simvastatin (Zocor®, Merck & Co., Inc.), atorvastatin (Lipitor®, Pfizer), rosuvastatin (Crestor®, Astra Zeneca) and pravastatin (Pravachol®, Bristol-Myers Squibb Co.), and combinations thereof, may be used in combination with bile acid-binding resins.Statins significantly reduce serum cholesterol and LDL serum levels, and slow the progression of coronary atherosclerosis.However, serum HDL cholesterol levels are only moderately increased. Side effects, including liver and kidney dysfunction, are associated with the use of these drugs (Physician's Desk Reference, Medical Economics Co., Inc., Montvale, NJ, 2004; hereafter "PDR"). The FDA has approved atorvastatin to treat rare but urgent cases of familial hypercholesterolemia.

[0018] Ezetimibe is a cholesterol absorption inhibitor that reduces the amount of cholesterol absorbed by the body. Ezetimibe is used to reduce the amount of total cholesterol, LDL cholesterol (about 18%), and apolipoprotein B. Ezetimibe is often used in conjunction with a low-cholesterol diet and, in some cases, other cholesterol-lowering drugs.

[0019] Niacin or nicotinic acid is a water-soluble vitamin B complex used as a dietary supplement and antihyperlipidemic agent. Niacin is effective in reducing the production of VLDL and lowering LDL. In some cases, niacin is used in combination with bile acid binding resins. NIASPAN® was approved to prevent recurrent heart attacks in patients with high cholesterol. Niacin can increase HDL when used in sufficient doses, but its usefulness is limited by serious side effects when used in such high doses.

[0020] Fibric acid derivatives ("fibrates") are a class of lipid-lowering drugs used to treat various forms of hyperlipidemia (i.e., elevated serum triglycerides) that may also be associated with hypercholesterolemia. Fibrates appear to reduce the VLDL fraction and slightly increase HDL. However, the effect of these drugs on serum cholesterol is variable. Fibrates are primarily used to lower high triglyceride levels. In the United States, fibrates are approved for use as antihyperlipidemic agents but not as hypercholesterolemic agents.

[0021] fatty liver disease Fatty liver disease is a term used to describe a group of liver diseases including nonalcoholic steatohepatitis (NASH), alcoholic steatohepatitis (ASH), nonalcoholic fatty liver disease (NAFLD) and HIV-associated steatohepatitis, with or without liver fibrosis. NASH is a common liver disease associated with increased morbidity and mortality. However, there are no FDA-approved treatment options, despite many compounds being tested in what are said to be NASH treatment models. NAFLD is a disorder that affects one in three to five adults and as many as one in ten children in the United States. These are conditions characterized by the accumulation of excess fat in the liver of people who drink little or no alcohol.

[0022] The most common form of NAFLD is a non-serious condition called hepatic steatosis (fatty liver), in which fat accumulates in liver cells. This is not normal, but in itself probably does not damage the liver. NAFLD most often appears in individuals with a set of risk factors called metabolic syndrome. Metabolic syndrome is characterized by elevated fasting plasma glucose (FPG) with or without intolerance to postprandial glucose, being overweight or obese, high blood lipids such as cholesterol and triglycerides (TG) and low high-density lipoprotein cholesterol (HDL-C) levels, and high blood pressure, although not all patients have all the signs of metabolic syndrome. Obesity is thought to be the most common cause of NAFLD, and some experts estimate that about two-thirds of obese adults and half of obese children may have fatty liver. The majority of individuals with NAFLD have no symptoms and their physical exam is normal (although the liver may be slightly enlarged). Children may present with symptoms such as abdominal pain and fatigue, and may have a patchy black discoloration of the skin (acanthosis nigricans). The diagnosis of NAFLD is usually first suspected in overweight or obese people who have mildly elevated liver blood tests during routine testing, but NAFLD may show normal liver blood tests or may be detected incidentally on imaging tests such as abdominal ultrasound or CT scan. NAFLD is confirmed by imaging tests, most commonly liver ultrasound or magnetic resonance imaging (MRI), and other causes are ruled out.

[0023] Some people with NAFLD may develop the serious condition NASH: approximately 2-5% of US adults and up to 20% of obese people may suffer from NASH. In NASH, accumulation of fat in the liver is associated with inflammation and varying degrees of scarring. NASH is a potentially serious condition with a substantial risk of progression to end-stage liver disease, cirrhosis, and hepatocellular carcinoma. Some patients who develop cirrhosis are at risk of liver failure and may ultimately require a liver transplant. Thus, weight loss is a recommended measure to prevent or slow the progression of NASH. However, weight loss has not been shown to treat NASH once liver fibrosis damage has occurred.

[0024] NAFLD can be differentiated from NASH by the sum of histopathology scores of liver biopsies for steatosis (0–3), lobular inflammation (0–2) and hepatocyte ballooning (0–2), the NAFLD activity score (NAS). An NAS of <3 corresponds to NAFLD, 3–4 to borderline NASH, and ≥5 to NASH. Biopsies are also scored for fibrosis (0–4).

[0025] NASH is the leading cause of end-stage liver disease.

[0026] Treatment of NAFLD and NASH No drugs are currently approved in the United States to prevent or treat NAFLD or NASH. Many pharmacological interventions have been attempted in NAFLD / NASH with limited overall benefit. Antioxidants halt lipid peroxidation and cytoprotectants stabilize phospholipid membranes, but agents that have been tried without success or with only modest benefit include ursodeoxycholic acid, vitamins E (α-tocopherol) and C, and pentoxifylline. Weight loss agents such as orlistat have not provided significant benefit compared with the use of diet and exercise alone to achieve weight loss ("weight loss alone").

[0027] Most weight loss trials in NAFLD / NASH have been pilot studies of short duration and limited success, reporting only modest improvements in necroinflammation or fibrosis. A randomized, double-blind, placebo-controlled 6-month trial of pioglitazone, a thiazolidinedione peroxisome proliferator-activated receptor-γ (PPARγ) agonist and insulin sensitizer, versus weight loss alone (Belfort, "A placebo-controlled trial of pioglitazone in subjects with nonalcoholic steatohepatitis", N. Engl. J. Med., 355, 2297-2307 (2006)) showed no improvement versus weight loss alone, but treatment with pioglitazone improved glycemic control, insulin sensitivity, indices of systemic inflammation (including hsCRP, tumor necrosis factor-α, and transforming growth factor-β), and liver histology in patients with NASH and IGT or T2DM. Treatment with pioglitazone also reversed adipose, liver and muscle IR and was associated with an approximately 50% reduction in necroinflammation (p<0.002) and a 37% reduction in fibrosis (p=0.08).

[0028] Improvements in hepatocellular injury and fibrosis have been reported in another controlled trial with pioglitazone over a 12-month period. In contrast, the first randomized clinical trial with rosiglitazone, another thiazolidinedione approved for diabetes treatment, in NASH showed reductions in IR, plasma alanine aminotransferase (ALT) levels, and steatosis, but rosiglitazone treatment had no significant effect on necrosis, inflammation, or fibrosis. It is important to note these results that reductions in ALT, insulin resistance, and other diabetes indices did not reduce liver fibrosis, an important indicator of NASH. Thus, diabetes control is not sufficient for the treatment or even prevention of NASH. Moreover, both pioglitazone and rosiglitazone have severe safety limitations. The preliminary report of the 2-year open-label, follow-up period of this trial was also disappointing, with no significant benefit from rosiglitazone treatment.

[0029] One pharmacological agent that has some efficacy in NASH is pioglitazone. Unfortunately, pioglitazone is also associated with significantly increased risk of weight gain, edema, congestive heart failure, and osteoporotic bone in both men and women.

[0030] A phase 2 clinical trial involving patients with NASH showed that daily subcutaneous semaglutide (a GLP-1 receptor agonist) treatment resulted in a higher proportion of patients with NASH resolution than placebo. However, the trial did not show a significant difference between groups in the proportion of patients with improvement in fibrosis stage (Newsome et al., N. Engl. J. Med. "A Placebo-Controlled Trial of Subcutaneous Semaglutide in Nonalcoholic Steatohepatitis" November 13, 2020). Unfortunately, "the proportion of patients in whom NASH resolution was achieved without worsening of fibrosis was 40% in the 0.1 mg group, 36% in the 0.2 mg group, 59% in the 0.4 mg group, and 17% in the placebo group (P=0.48). The mean percentage weight loss was 13% in the 0.4 mg group and 1% in the placebo group. The incidence of nausea, constipation, and vomiting was higher in the 0.4 mg group than in the placebo group (nausea, 42% vs 11%; constipation, 22% vs 12%; and vomiting, 15% vs 2%). Malignant neoplasms were reported in three patients (1%) receiving semaglutide and in none receiving placebo. Overall, neoplasms (benign, malignant, or unspecified) were reported in 15% of patients in the semaglutide group and 8% in the placebo group; no specific organ occurrence pattern was observed." Thus, even GLP-1 agonists such as semaglutide are not a safe treatment for preventing or treating NASH that justifies the risks of long-term administration necessary to treat, prevent or slow the progression of NASH.

[0031] Wilding et al., N. Engl. J. Med. Published 10 February 2021 conducted an obesity trial with a maintenance dose of 2.4 mg of semaglutide (or placebo) administered subcutaneously once weekly for 68 weeks. "In the semaglutide group, weight loss was observed from the first postrandomization assessment (week 4) onwards, reaching a nadir at week 60". However, many side effects were noted, including gastrointestinal disorders, "Gastrointestinal disorders (typically nausea, diarrhea, vomiting and constipation) were the most frequently reported events, occurring more frequently in participants receiving semaglutide than in those receiving placebo (74.2% vs 47.9%)". Of more concern was that "serious adverse events were reported in 9.8% and 6.4% of semaglutide and placebo participants, respectively". Many semaglutide participants discontinued due to the severity of side effects. Summary of the Invention

[0032] Furthermore, GLP-1 analogs exert GLP-1 activity rather than GLP-2 activity, and exert their effects primarily through hormone signaling pathways, sustained rather than the usual episodic action (coinciding with a temporary meal). Given sustained hormone pathway stimulation, the risk of side effects, thyroid c-cell tumors and pancreatitis, increases significantly. Furthermore, antibodies are often formed against synthetic GLP-1 analog derivatives (formed to prevent DPP-4 enzyme degradation). For example, 61% of patients developed antibodies against exenatide. Thus, there is a need in the art for better combinations with GLP-1 analogs to allow lower GLP-1 analog dosing and address the severe side effects observed with chronic dosing.

[0033] The summary of clinical data obtained shows that, even though weight loss may be an effective means for preventing NASH or may slow the progression of NASH, the treatment of NASH seems to be uncoupled from weight loss as a treatment method by any weight loss method.Therefore, there is a need for a more acceptable translational model for predicting the prevention, progression prevention and treatment of fatty liver disease, including NASH.Therefore, there is a need for effective and safer NASH treatment options, especially when the treatment can be delivered orally rather than by injection.There is also a need for safe agents that prevent the development of complete NASH liver disease and damage and slow the progression of NASH.

[0034] The present disclosure is made in part to address the aforementioned needs in the treatment and / or management of glucagon-related diseases, disorders or conditions, including (a) glycemic control / diabetes / metabolic syndrome (MetS), (b) weight loss and / or obesity, and (c) hyperlipidemia.Disclosed herein is the discovery of a combination of one or more bitter taste receptor agonists (otherwise referred to as TAS2R agonists) and at least one gut signaling compound, which provides significant benefits and advantages over currently available treatments of glucagon-related conditions, including the use of gut signaling compounds (i.e., gut signaling peptide analogs and gut signaling hormone enhancers).The present disclosure is also made in part to address the aforementioned needs in the treatment or prevention of fatty liver disease, such as NASH.

[0035] Preferably, the combination described herein (for example, with at least one gut signaling compound) that comprises bitter taste receptor agonist is formulated into pharmaceutical composition, more preferably into oral dosage form.The combination can provide great benefits in treating glucagon-related diseases, disorders and conditions, including, for example, diabetes, pre-diabetes syndrome, obesity, weight and / or appetite control, hyperlipidemia and hyperglycemia.One advantage of the combination of the present invention is that it provides equal or better efficacy with reduced dosage of gut signaling compound to achieve equal or better results with reduced side effects.

[0036] The present disclosure further provides methods for treating or preventing the progression of glucagon-related diseases, disorders and conditions, such as diabetes, pre-diabetic syndrome, obesity, weight and / or appetite control, hyperlipidemia and hyperglycemia, comprising administering a combination of one or more bitter taste receptor agonists and gut signaling compounds to a subject having such disease, disorder or condition.

[0037] The present disclosure further provides a method for treating or preventing the progression of fatty liver disease (e.g., selected from the group consisting of NASH, ASH, NAFLD or HIV-associated steatohepatitis with or without liver fibrosis) to a subject with fatty liver disease, comprising administering a combination comprising denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate; and a GLP-1 agonist drug.Preferably, the GLP-1 agonist drug is selected from the group consisting of semaglutide, glyburide, liraglutide, dulaglutide and / or albiglutide.

[0038] In some embodiments, the daily dose of the denatonium salt for a human adult is about 50 mg to about 3000 mg administered once a day (QD) or twice a day (BID). Preferably, the daily dose of the denatonium salt is about 100 mg to about 2000 mg administered QD or BID. Most preferably, the daily dose of the denatonium salt is about 200 mg to about 1000 mg administered QD or BID.

[0039] In some embodiments, the method further comprises administering acetic acid, for example, from about 0.5 g to about 5 g per dose. More preferably, the daily dose of acetic acid for an adult is from about 1.5 g to about 3 g. [Brief description of the drawings]

[0040] In the figure, the designation "ARD-101" means denatonium acetate (DA).

[0041] [Figure 1] FIG. 1 shows the average weight gain over the study period for all treatment groups in Example 2.

[0042] [Diagram 2] 2 shows serum triglyceride (TG) levels at the end of the study (day 31) for animals in each of the four treatment groups in Example 2. These data show that (1) treatment with DA, liraglutide, or a combination thereof significantly reduced serum TG levels in diet-induced obese (DIO) mice compared to vehicle; and (2) animals treated with the combination exhibited significantly lower serum TG levels compared to serum TG levels treated with DA or liraglutide alone, indicating a potential synergistic (or at least additive) effect between the two agents on serum TG levels.

[0043] [Diagram 3]3 shows serum glucose levels at the end of the study (day 31) for each animal in the four treatment groups in Example 2. Treatment with DA, liraglutide, or a combination thereof significantly reduced serum glucose levels in DIO mice when administered for 4 weeks.

[0044] [Figure 4] 4 shows serum HbA1c levels at the end of the study (day 31) for each animal in the four treatment groups in Example 2. The results suggest that treatment with DA, liraglutide, or their combination had no significant effect (p>0.05) on serum HbA1c levels in DIO mice after 4 weeks of treatment.

[0045] [Diagram 5] 5 depicts serum insulin levels at the end of the study (day 31) for each animal in the four treatment groups in Example 2. The data demonstrate that 4 weeks of treatment with DA, liraglutide, or their combination significantly reduced serum insulin levels in DIO mice.

[0046] [Figure 6] Figure 6 represents serum BA levels at the end of the study (day 31) for each animal in the four treatment groups in Example 2. When administered for 4 weeks, DA, liraglutide, or the combination thereof resulted in a significant increase in serum BA levels compared to the vehicle control.

[0047] [Figure 7] Figure 7 shows serum LDL levels at the end of the study (day 31) for animals in each of the four treatment groups in Example 2. There were no significant differences in serum LDL levels in animals treated with either vehicle or DA, liraglutide, or a combination thereof.

[0048] [Figure 8]Figure 8 shows serum HDL levels at the end of the study (day 31) for each animal in the four treatment groups in Example 2. The data demonstrate that, compared to vehicle, treatment with liraglutide, or the combination of DA + liraglutide, resulted in a significant decrease in serum HDL levels in DIO mice after 4 weeks of dosing.

[0049] [Figure 9] 9 shows the rate of change in GLP-1 from time 0 to 1 hour after oral administration in Example 3. With a small sample size, the difference was significant (p=0.0235).

[0050] [Figure 10] FIG. 10 shows the results for GLP-2 from Example 3, which showed a trend towards an increase in GLP-2 gut peptide hormone.

[0051] [Figure 11] FIG. 11 shows the results for PYY from Example 3, which showed a trend towards an increase in the PYY gut peptide hormone.

[0052] [Figure 12] Figures 12A and 12B show the relative body weight percentage (12A) and the relative body weight change (g) (12B) for the four groups of animals treated in Example 4. Treatment with sitagliptin alone had the least effect on body weight, consistent with previous studies and clinical experience. However, a much larger effect on body weight was seen with DA alone (ARD-101), while a significant or synergistic effect on body weight was seen with the combination of DA and sitagliptin.

[0053] [Figure 13] 13 shows that the combination of DA (ARD-101) and sitagliptin significantly reduced the weight gain of DIO mice on day 56 of the study in Example 4. The combination of DA and sitagliptin significantly reduced the weight gain of DIO mice compared to mice treated with the same sitagliptin at the same dose.

[0054] [Figure 14] 14A and 14B show that on day 56 of the study, treatment with DA, or its combination with sitagliptin, both showed significant effects on body weight in Example 4.

[0055] [Figure 15] Figures 15A and 15B show that DA alone and DA + sitagliptin significantly reduced fasting plasma glucose levels in DIO mice compared to vehicle controls on days 28 (Figure 15A) and 56 (Figure 15B).

[0056] [Figure 16] Figures 16A and 16B show that DA + sitagliptin significantly reduced HbA1c levels in DIO mice compared to vehicle controls at days 28 (Figure 16A) and 56 (Figure 16B). Baseline day 0 HbA1c levels were 4.7%.

[0057] [Figure 17] Figures 17A and 17B show that DA + sitagliptin significantly reduced insulin levels in DIO mice compared to vehicle controls at days 28 (Figure 17A) and 56 (Figure 17B). Baseline day 0 insulin levels were 1 ng / ml.

[0058] [Figure 18] Figures 18A and 18B show that DA + sitagliptin significantly reduced triglyceride (TG) levels in DIO mice compared to vehicle controls at days 28 (Figure 18A) and 56 (Figure 18B). Baseline day 0 triglyceride levels were 33.8 mmol / L.

[0059] [Figure 19]Figures 19A and 19B show that DA + sitagliptin significantly reduced bile acid (BA) levels in DIO mice compared to vehicle controls at days 28 (Figure 19A) and 56 (Figure 19B). Baseline day 0 bile acid levels were 27 μmol / L.

[0060] [Figure 20] Figures 20A and 20B show that DA + sitagliptin significantly reduced total cholesterol (TC) levels in DIO mice compared to vehicle controls at days 28 (Figure 20A) and 56 (Figure 20B). Baseline day 0 total cholesterol levels were 110 μg / μL.

[0061] [Figure 21] Figures 21A and 21B show that DA + sitagliptin significantly reduced low-density lipoprotein (LDL) levels in DIO mice compared to vehicle controls at days 28 (Figure 21A) and 56 (Figure 21B). Baseline day 0 low-density lipoprotein (LDL) levels were 125 mg / dL.

[0062] [Figure 22] FIG. 22A shows that sitagliptin alone significantly reduced high density lipoprotein (HDL) levels compared to vehicle controls.

[0063] However, Figure 22B shows that sitagliptin alone, DA alone and DA + sitagliptin significantly reduced high density lipoprotein (HDL) levels in DIO mice compared to vehicle controls at day 56. Baseline day 0 high density lipoprotein (HDL) levels were 60 mg / dL.

[0064] [Diagram 23] FIG. 23 shows that treatment with DA (ARD-101), semaglutide, or their combination significantly improved NAFLD activity scores based on blinded histopathological review.

[0065] [Figure 24] Figures 24A and 24B show that treatment with DA (ARD-101), semaglutide, or their combination showed a significant effect on body weight (24A) and body weight change (24B) in trans fat-containing amylin liver NASH (AMLN) diet-induced mice containing the synergistic combination. Data are presented as means. Statistical analysis was performed using one-tailed t-test. ***P<0.001 (compared to vehicle); $$P<0.01 and $$$P<0.001 (compared to the combination)

[0066] [Diagram 25] Figures 25A and 25B show liver weights (Figure 25A) and liver / body weight ratios (Figure 25B) showing that (1) both treatments significantly reduced liver weights and liver / body weight ratios compared to vehicle; and (2) the effect of the combination of DA (ARD-101) and semaglutide was significantly greater than single-agent DA or semaglutide, indicating synergy between the two agents.

[0067] [Figure 26] FIG. 26A shows alanine aminotransferase (ALT) levels.

[0068] Figure 26B shows aspartate aminotransferase (AST) levels.At the end of the study, the two treatments each significantly reduced ALT and AST levels compared with vehicle control.In addition, the combination of DA (ARD-101) and semaglutide resulted in significantly lower ALT levels compared with either DA alone or semaglutide alone.This indicates a synergistic effect.

[0069] [Figure 27] Figures 27A, 27B and 27C show that at the end of the study of Example 6, DA (ARD-101) and semaglutide each significantly reduced TG (27A), LDL (27B) and HDL (27C), respectively.

[0070] [Figure 28] Figure 28 shows that at the end of the study of Example 6, the combination of DA (ARD-101) and semaglutide significantly attenuated the increase in fasting blood glucose levels induced by the AMLN diet compared to vehicle control.

[0071] [Figure 29] Figure 29 shows that at the end of the study, the combination of DA (ARD-101) and semaglutide significantly increased HbA1c compared to the vehicle control. Baseline HbA1c levels were 5.0%.

[0072] [Diagram 30] Figure 30 shows that at the end of the study, the combination of DA (ARD-101) and semaglutide significantly reduced insulin levels compared to the vehicle control. Baseline insulin levels were 1.5 ng / ml.

[0073] [Diagram 31] Figure 31 shows that the two treatments did not significantly affect bile acid levels compared to the vehicle control. Baseline bile acid levels were 30 μmol / L.

[0074] [Diagram 32] Figures 32A (CK-18) and 32B (TGF-β) show that the two treatments each significantly reduced CK-18 levels compared to the vehicle control (Figure 32A), and that only the combination of semaglutide and DA significantly reduced TGF-β1 levels compared to the vehicle control. These data provide further evidence of the synergistic effect of these two agents.

[0075] [Diagram 33] 33A and 33B show that at the end of the study of Example 6, the two treatments did not significantly affect IL-6 and TNF-α levels compared to vehicle. DETAILED DESCRIPTION OF THE PREFERRED EMBODIMENTS

[0076] The present disclosure is based in part on in vivo studies and clinical trials (as shown in the Examples herein) that have found surprising, beneficial and / or synergistic results in using a combination of a bitter taste receptor agonist, specifically an orally administered denatonium salt, wherein the denatonium salt is selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide, and denatonium tartrate; and at least one intestinal signaling compound, to treat glucagon-related diseases, disorders or conditions, including weight control and fatty liver disease, and to prevent the progression of fatty liver disease.

[0077] The section headings are provided solely for the convenience of the reader and are not intended to limit the disclosure.

[0078] To the extent that any matter incorporated by reference contradicts the explicit content of this disclosure, the explicit content controls.

[0079] (definition) "About" as used herein includes the exact amount modified by the term about, and an amount that is expected within experimental error, such as within 15%, 10% or 5%. For example, "about 5 mg" means "5 mg" and also means a range of mg that is within experimental error, such as plus or minus 15%, 10% or 5% of 5 mg. As used herein, the term "about" can be used to modify ranges and specific values ​​as well.

[0080] "Administering a combination" refers to any administration of more than one agent, whether the agents are administered simultaneously or sequentially; in the same composition or different compositions; and by the same route or different routes.

[0081] "API" means active pharmaceutical ingredient.

[0082] "Fatty liver disease" means any of a group of diseases characterized by the unwanted accumulation of fat in the liver, including nonalcoholic steatohepatitis (NASH), alcoholic steatohepatitis (ASH), nonalcoholic fatty liver disease (NAFLD), and HIV-associated steatohepatitis, with or without liver fibrosis.

[0083] A "glucagon-related" disease, disorder or condition, as used herein, means any unwanted condition in a subject, including any condition that can be mediated by intestinal signaling compounds that are mediated by the production, maintenance or metabolism of glucagon in a subject, or by the glucagon regulatory cycle.

[0084] "Gut signaling compound" refers to gut signaling peptide analogs and / or gut signaling hormone enhancers, such as, for example, compounds selected from GLP-1 receptor agonists (sometimes referred to as GLP-1 agonists or GLP-1 analogs), GLP-2 analogs, PYY analogs, DPP-4 inhibitors, GIP analogs and CCK analogs, as further described herein.

[0085] "Or" is used in its inclusive sense (equivalent to "and / or") unless the context otherwise requires.

[0086] As used herein, "synergy" or "synergistic" is used to convey the beneficial effect of API combinations that produce the effectiveness of multiple gut peptide hormone receptor signaling agonists rather than the simple increase of a single gut peptide hormone. In some embodiments, synergy is shown in which the combination of APIs administered during in vivo testing produces benefits that exceed additive benefits. Without being bound by theory, one hypothesis is that the beneficial effect of API combinations is due to (a) the transient increase of gut signaling hormones relative to long-acting GLP-1 receptor agonists, which have a high incidence of severe side effects that limit their use; and / or (b) the efficacy of multiple gut peptide hormone receptor signaling agonists rather than the simple increase of a single gut peptide hormone, and / or (c) the combined effect of temporarily increasing gut signaling peptides of long-acting baseline characteristics.

[0087] The terms and symbols "wt. %" and "%" refer to the weight percentage of excipients and API, and when used in reference to multi-layer tablets, refer to the "wt. %" of an individual layer, e.g., an "individual layer" being the first or second layer of a bilayer tablet.

[0088] A "therapeutically effective amount" of an API means an amount that, when administered to a human for treating a disease (e.g., fatty liver disease such as NAFLD or NASH), is sufficient to effect treatment of the disease state being treated. When applied to NAFLD or NASH in humans, "treating" or "treatment" refers to (1) preventing or reducing the risk of developing NAFLD or NASH, i.e., preventing the development of clinical symptoms of NAFLD or NASH in subjects who may be predisposed to NAFLD or NASH but who have not yet experienced or exhibited symptoms of NAFLD or NASH (i.e., prophylaxis); (2) inhibiting NAFLD or NASH, i.e., arresting or reducing the onset of NAFLD or NASH or clinical symptoms thereof; and (3) alleviating NAFLD or NASH, i.e., causing regression, reversal, or improvement of NAFLD or NASH, or reducing the number, frequency, duration, or severity of its clinical symptoms; This includes one or more of the following:

[0089] Similarly, "treating" or "treatment" as applied to T2DM includes treatment of diabetes and preventing the onset of diabetes by treating a pre-diabetic state or preventing the progression of T2DM requiring insulin treatment.

[0090] The therapeutically effective amount for a particular subject varies according to the health and condition of the subject being treated, the degree of disease progression (e.g., NAFLD or NASH), evaluation of medical condition and other related factors.It is expected that the therapeutically effective amount falls within a relatively broad range and can be determined by routine testing.

[0091] (Implementation) The present disclosure is based on the surprising discovery of synergistic combinations in (a) an in vivo weight loss study of two groups of compounds with different mechanisms of action measuring gut peptide hormone levels, (b) a Phase 1 clinical trial with oral administration measuring gut peptide hormones pre- and 1 hour after administration, and (c) a 56-day in vivo weight loss study in DIO mice showing the synergistic effect of the combination of two orally administered drugs compared to each drug administered alone. (a) The in vivo study (see Example 2 for results) was a long-term weight control study in DIO mice investigating the synergistic effect between denatonium salts (denatonium acetate or DA) and the GLP-1 receptor agonist riraglutide. These surprising findings are reflected in gut peptide hormones, GLP-1, CCK, PYY and GLP-2, as well as standard blood tests such as HbA1c and lipids.

[0092] A phase 1 clinical trial in which denatonium acetate was orally administered (see Example 3) resulted in increased gut hormone signaling of GLP-1 and two additional gut hormone peptides. Gut peptide hormone data from clinical trials in which DA was administered indicated that the possible mechanism of action of denatonium acetate for weight loss is based on signaling through multiple gut hormone peptides. This is because PK (pharmacokinetic) analysis showed that DA is substantially restricted to the gut, and pharmacokinetic data showed that DA is primarily restricted to the gut and does not affect weight loss through DA systemic concentration. Thus, the combination of denatonium salts with other gut peptide agonists, such as GLP-1RA, GIP analogs, PYY analogs and DPP-4 inhibitors, which act to increase the plasma half-life of gut signaling peptides GLP-1, PYY and CCK, can significantly increase their activity and allow for a lower dose of GLP-1RA to reduce side effects. Thus, gut peptide hormone data, both in vivo testing in the examples herein and Phase 1 clinical trial data in Example 3, show synergistic effects in the treatment and / or management of glucagon-related diseases, disorders or conditions, various indications. Clinical data showed that there are multiple gut peptide hormones (not just GLP-1) that DA affected. Clinical data also substantiated the DIO mouse data (Example 2). Nevertheless, marketers of GLP-1 agonists (such as the GLP-1 agonists available from Novo Nordisk, Lilly) claim that only GLP-1 is important for both diabetes and weight loss. Also, DPP-4 is an enzyme that breaks down GLP-1 and PYY, giving both of those hormones a short half-life. Thus, in some embodiments herein, DPP-4 inhibitors are used as part of API combinations.

[0093] In one embodiment herein, bitter taste receptor agonists (or TAS2R agonists) are substantially restricted to the gut and exert their activity through gut peptide hormones. DPP-4 inhibitors do not provide meaningful weight loss benefits. A 56-day in vivo weight loss study in DIO mice showed synergistic effects of the combination of two orally administered drugs (DA, a bitter taste receptor agonist substantially restricted to the gut, and the DPP-4 inhibitor sitagliptin phosphate) compared to each drug administered alone. Sitagliptin phosphate (Januvia®) caused slight weight loss over the first 30 days of administration, but as seen in patients, weight regain occurred and no weight loss effect was observed with longer periods of administration. Thus, sitagliptin phosphate showed its well-known lack of weight loss effect. DA caused significant weight loss. However, adding sitagliptin, which has no significant weight loss effect by itself, significantly increased the weight loss benefit of DA. This synergistic effect was seen with other measured metabolic parameters as well, including HbA1c, insulin, triglycerides, blood glucose, bile acids, cholesterol and low density lipoprotein (LDL), data presented in Example 4.

[0094] The data disclosed herein demonstrate that a combination of a bitter taste receptor agonist with either or both of a GLP-1RA (such as liraglutide or semaglutide) and a DPP-4 inhibitor, and optionally a GIP agonist, can (1) increase gut peptide hormone efficacy and (2) allow potentially lower dosing of difficult (with severe side effects) gut peptide hormone agents (such as semaglutide or other GLP-1 agonists) to reduce side effects while providing greater efficacy than higher doses of the individual therapeutic components alone.

[0095] More specifically, the findings show that the combination of bitter taste receptor agonists with gut peptide hormone agents selected from gut peptide analogs GLP-1, GLP-2, PYY, CCK and DPP-4 inhibitors (which increase the half-life of the natural gut peptide hormones GLP-1 and PYY) is synergistic or adds a "benefit". "Benefit" may refer to the ability to reduce the dosage of GLP-1 agonists, which can significantly reduce many of the severe side effects of GLP-1 agonist administration as indicated on the product label. From a mechanism of action perspective, denatonium salts are bitter taste receptor agonists, stimulating the episodic and endogenous secretion of multiple gut peptide hormones (such as GLP-1, GLP-2, PYY and CCK), providing multiple gut axis signals (i.e., a symphony) instead of just one gut peptide hormone such as GLP-1 (i.e., a violin), which is only one of the signals.

[0096] The data disclosed herein further show that the disclosed combination of bitter taste receptor agonist and gut signaling compound produces surprising beneficial results in treating fatty liver disease such as ASH, NASH and NAFLD. For example, after the in vivo test described in Example 6, the combination of DA and semaglutide significantly improved NAFLD activity score (Figure 23); showed a significant synergistic effect on body weight and body weight change in trans fat-containing amylin liver NASH (AMLN) diet-induced mice (Figures 24A, 24B); and significantly reduced liver weight and liver / body weight ratio compared to vehicle. The combination of DA and semaglutide also produced a significantly greater effect compared to single agent DA or semaglutide, showing a synergistic effect of the combination of the two agents (Figure 25). In particular, the data further show that the combination of the present invention had a surprising improvement in the effect of reducing ALT and AST levels (Figure 26) and reducing TG, LDL and HDL (Figures 27A-C) in AMLN diet-induced mice.

[0097] In view of the above-mentioned findings and discoveries, which will be further described below, the present disclosure provides in one embodiment a combination pharmaceutical composition comprising a bitter taste receptor agonist and a formulation of gut signaling compound, such as gut signaling peptide analog and / or gut signaling hormone enhancer.Preferably, the pharmaceutical combination further comprises a DPP-4 inhibitor, which acts to inhibit DPP-4 enzyme activity and destroy endogenous GLP-1 and PYY gut peptide hormones.

[0098] In another embodiment, the present disclosure provides a combination oral dosage pharmaceutical composition comprising a bitter taste receptor agonist and a DPP-4 inhibitor.

[0099] In another embodiment, the present disclosure provides a synergistic method for treating glucagon-related diseases, disorders or conditions, such as obesity, diabetes, glycemic control, metabolic syndrome, hyperlipidemia, and achieving weight loss, comprising administering an effective amount of a pharmaceutical composition comprising bitter taste receptor agonist and one or more intestinal signaling compounds.Preferably, the method further comprises administering an enhancer of endogenous GLP-1 and PYY activity-DPP-4 inhibitor.

[0100] In another embodiment, a synergistic method is described for treating multiple aspects of metabolic syndrome, including obesity, diabetes / MetS and hyperlipidemia, comprising administering DPP-4i and DA, which are administered simultaneously in a single dosage form or in separate dosage forms.No additive toxicity is noted.

[0101] In another embodiment, the present disclosure provides a method for treating hyperlipidemia comprising administering an effective amount of an orally administered pharmaceutical composition comprising a combination of a bitter taste receptor agonist and a gut signaling compound.

[0102] In another embodiment, the present disclosure provides a method for treating glycemic control, metabolic syndrome (MetS) and diabetes comprising administering an effective amount of an orally administered pharmaceutical composition comprising a combination of a bitter taste receptor agonist and an intestinal signaling compound.

[0103] In another embodiment, the disclosure provides a method for treating obesity and achieving weight loss comprising administering an effective amount of an orally administered pharmaceutical composition.

[0104] The present disclosure further provides a method for treating MetS and diabetes comprising administering an effective amount of an orally administered pharmaceutical composition comprising a combination of a bitter taste receptor agonist and a gut signaling compound.

[0105] In another embodiment, the present disclosure further provides a method for treating fatty liver disease, including ASH, NASH and NAFLD, (more preferably, for treating NASH), comprising administering an effective amount of an oral administration pharmaceutical composition comprising a combination of bitter taste receptor agonist and intestinal signaling compound.In one preferred embodiment, the present disclosure provides a method for treating NASH, comprising administering a combination of DA and intestinal signaling compound, more preferably, the intestinal signaling compound is selected from GLP-1 agonist, and even more preferably, the intestinal signaling compound is semaglutide.

[0106] Preferably, in each embodiment herein, the bitter taste receptor agonist is selected from the group consisting of denatonium salts (including DA, denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate), chlorpheniramine, diphenidol, famotidine, haloperidol, quinine, parthenolide and aristolochic acid. More preferably, the bitter taste receptor is a denatonium salt selected from DA, denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate, and even more preferably DA. It should be understood that these preferred choices for the bitter taste receptor agonist apply to each of the other embodiments and methods of use described herein, including the combination pharmaceutical composition of the present invention and the methods of use and treatment or prevention of glucagon-related diseases, disorders or conditions, and / or fatty liver diseases, including NASH, ASH and NAFLD.

[0107] In each embodiment disclosed herein, the DPP-4 inhibitor is selected from the group consisting of salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin salts (including phosphate salts), saxagliptin, linagliptin, alogliptin, and combinations thereof. Preferably, in one embodiment, the DPP-4 inhibitor is selected from sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin, and alogliptin.

[0108] In one embodiment, the DPP-4 inhibitor is sitagliptin phosphate combined in a single oral dosage form or taken together in two oral dosage forms.

[0109] Preferably, the dosages of commercially approved DPP-4 inhibitors used in the methods and combinations described herein are daily doses that are the approved daily doses for the particular DPP-4 inhibitor administered once daily (QD) or twice daily (BID) about 8 hours apart, and the bitter taste receptor agonist (preferably, a denatonium salt) is administered once or twice daily at a total daily dose (by weight of denatonium) of about 200 mg to about 480 mg.

[0110] For example, in one embodiment, when the DPP-4 inhibitor is sitagliptin, the total daily dose for a human adult is about 200mg to about 1000mg per day, administered either QD or BID simultaneously with or after the DPP-4 inhibitor. Regardless of whether the DPP-4 inhibitor is administered QD or BID, DA is preferably administered BID. Preferably, the single dosage form comprises a proportion of oral (PO) dosage selected from the group consisting of sitagliptin 50mg / DA 200mg PO BID, sitagliptin 50mg / DA 240mg PO BID, sitagliptin 100mg / DA 200mg PO QD, sitagliptin 100mg / DA 240mg PO QD, and sitagliptin 100mg / DA 480mg PO QD.

[0111] In a further embodiment, the combination pharmaceutical composition further comprises an oral dosage form of the GLP-1RA semaglutide.

[0112] In the inventive combinations and methods described herein, the bitter taste receptor agonist may be administered in a single dosage form or in two dosage forms.

[0113] In the inventive combinations, methods and uses described herein, the intestinal signalling compound is preferably selected from a GLP-IRA analogue, a GLP-1 receptor agonist, a GLP-2 analogue, a PYY analogue, a DPP-4 inhibitor, a GIP analogue and a CCK analogue.

[0114] More preferably, 1) the GLP-1RA is selected from the group consisting of semaglutide, glyburide, liraglutide, dulaglutide and albiglutide; 2) the PYY 1875 analog is selected from the group consisting of NN-9775 (Novo Nordisk) and JNJ-9321 (Johnson & Johnson); 3) the CCK analog is selected from the group consisting of C-2819 (Astra Zeneca), NN-9056 (Novo-Nordisk) and A-71378 (AbbVie); 4) the DPP-4 inhibitor is selected from the group consisting of sitagliptin phosphate, vildagliptin, linagliptin, alogliptin, saxagliptin (BMS47718), P93 / 01 (Prosidion), SYR322 (Takeda), GSK 823093, Roche 0730699, TS021 (Taisho), E3024 (Eisai) and PHX-1149 (Phenomix); and 5) The GLP-2 analogue is selected from the group consisting of teduglutide, grepaglutide, apraglutide, elsiglutide, HM-15912 (Hamni Pharmaceuticals), ZP-7570 (Zealand Pharma AS), GLP-2-ELP (PhaseBio Pharmaceuticals), MOD-1501 (OPKO Health) and HL-06 (Huons Global Co. Ltd.).

[0115] In one embodiment, the present disclosure provides a method for lowering the dosage of GLP-1RA, comprising co-administering a bitter taste receptor agonist as described herein.Preferably, the GLP-1RA is selected from the group consisting of semaglutide, glyburide, liraglutide, dulaglutide and albiglutide.

[0116] For indications other than obesity and weight loss, improvements in parameters represented by translationally relevant biomarkers (blood glucose, HbA1C, insulin, triglycerides, LDL cholesterol and total cholesterol) indicated that DA, or its combination with sitagliptin, showed superior benefit compared with sitagliptin alone.

[0117] Sitagliptin is excreted by kidney with minimal hepatic metabolism (through CYP3A4 and CYP2C8), whereas DA (Example 3) is approximately 99% restricted to the intestine, with limited systemic exposure.Therefore, both the pharmacokinetic data of clinical trials for DA (Example 3) and the published pharmacokinetic data for marketed sitagliptin support non-cumulative toxicity risk.Therefore, the DPP-4i drugs and denatonium salts listed herein are effective oral combinations for the treatment of obesity.

[0118] These data support the finding that DA alone or in combination with DPP-4i offers great benefits in the treatment of diabetes and metabolic syndrome in general, besides obesity. Despite hyperlipidemia being an important frequent comorbidity of diabetes or obesity, sitagliptin is not known to be effective in addressing hyperlipidemia (cholesterol and triglycerides) beyond its main intended effect on blood glucose. However, as Figures 20-23 show, sitagliptin in combination with DA showed efficacy against hyperlipidemia.

[0119] Table 1 compares the data on the DA + sitagliptin combination therapy presented in Example 4 below with four commercially available GLP-1 RAs (exenatide, dulaglutide, liraglutide and semaglutide) from similar and comparable published in vivo studies. [Table 1]

[0120] Table 1 shows a comparison of the combination of DA and sitagliptin versus various GLP-1 agonists on various parameters. # Based on a weight of 60 kg; ## Absolute weight change rate; * , ** and *** represents p<0.05, <0.01 and <0.001 vs. control, respectively. Abbreviations: HFD, high-fat diet; HED, human equivalent dose; BID, twice daily; QD, once daily; HbA1c, hemoglobin A1C; TG, triglycerides; TC, total cholesterol; LDL, low-density lipoprotein; PO, oral; SC, subcutaneous; IP, intraperitoneal. References: Sci Rep. 2019;9(1):15601; Int. J Obes. (Lond). 2020;44(4):937; Eur. J. Med. Chem. 2020;198:112389; Sci. Transl. Med. 2018; 10(472):eaat3392.

[0121] GLP-1RA drugs such as semaglutide also function along the GLP-1 axis, but they utilize a GLP-1-like structure that is not subject to rapid endogenous degradation. However, a drawback of GLP-1RAs is that they generally must be injected (with the exception of the approved oral semaglutide, Ryversus®) and are subject to a "black box" warning due to the associated increased risk of cancer and pancreatitis. In contrast to DPP-4i drugs, one of the more notable advantages of GLP-1RAs is their effect on weight loss, which has not been consistently replicated by oral DPP-4i drugs. The extent of weight loss measured relative to controls was 10-15%, but over a much longer period than the duration of the DA / DPP-4i combo study referenced in this application. Table 1 shows a comparison of the data in predictive in vivo models as DA+DPP-4i compared to GLP-1RA agents. Thus, taking into account the degree of improvement measured compared to control, DA plus sitagliptin (or another DPP-4i drug) appears to show similar efficacy and superior safety to GLP-1R agonists for the treatment of obesity, diabetes, or metabolic syndrome in general.

[0122] Preferably, administration in humans uses the current optimal doses of both the DPP-4i agent and the DA, at the recommended doses as single agents co-administered once or twice daily. For sitagliptin as a single agent, current guidelines indicate a daily dose of 100 mg PO QD (nevertheless, for those with renal impairment, the recommended dose may be as low as 25 mg to 50 mg PO QD). DA is in clinical trials and has been shown to be safely administered up to 240 mg PO BID in a Phase 1 clinical trial provided in Example 3 herein. Given the pharmacokinetics (substantially gut-confined) and relatively non-toxic nature of DA, the optimal dose range can be safely adjusted higher. An ongoing Phase 2 clinical trial is using 200 mg DA PO BID. DA can be taken once or twice daily.

[0123] Thus, some doses for a combination tablet / capsule formulation using both sitagliptin phosphate (total dose of 100 mg per day) and DA (total dose of about 200 mg to about 1000 mg per day based on the weight of denatonium) are as follows: 1. Sitagliptin 50mg / DA 200mg PO BID 2. Sitagliptin 50mg / DA 250mg PO BID 3. Sitagliptin 100mg / DA 200mg PO QD 4. Sitagliptin 100mg / DA 400mg PO QD 5. Sitagliptin 100mg / DA 500mg PO QD.

[0124] DA is preferably administered BID, since the appetite suppression effect observed in animals lasts for about 8 hours. Thus, twice-daily administration is preferred for suppressing appetite throughout the day. However, additional data has shown that DA confers metabolic benefits independent of weight loss, even when administered once-daily (and at levels lower than the BID doses in previous NASH studies). Therefore, the preferred administration for MetS treatment is QD.

[0125] Both QD and BID administration are effective for metabolic syndrome applications. However, if obesity is the primary indication for treatment, BID administration is the preferred embodiment. And if other aspects of metabolic syndrome (diabetes and hyperlipidemia) are the primary indication for treatment, either QD or BID administration may be preferred (QD for convenience and patient compliance).

[0126] GLP-1 receptor agonists The class of GLP-1 receptor agonists (sometimes simply referred to as GLP-1 agonists or GLP-1 analogues) includes dulaglutide (Trulicity®), which can be taken by weekly injection; liraglutide (Victoza®), which can be injected once daily; exenatide extended release semaglutide (Bydureon®), which can be taken by weekly injection; exenatide ER (Astra Zeneca), which can be taken by weekly injection; semaglutide (Ozempic®), which can be taken by weekly injection; semaglutide (Revelsus®), which can be taken orally once daily; lixisenatide (Adlyxin®), which can be taken by daily injection; and albiglutide (Tanzeum®), which can be injected once weekly. The Novo-Nordisk GLP-1 analogues semaglutide and liraglutide are fatty acid-modified GLP-1 protein receptor agonists. Dulaglutide and albiglutide, from Lilly and GSK, respectively, are fusion protein GLP-1 receptor agonists.

[0127] GLP-1 analogs have been approved for the treatment of type 2 diabetes as measured by glycemic control (HbA1c). GLP-1 analogs are also currently being evaluated in clinical trials for weight loss and obesity. GLP-1 induces numerous biological effects, such as stimulating insulin secretion, inhibiting glucagon secretion, inhibiting gastric emptying, inhibiting gastric or intestinal motility, and inducing weight loss. A hallmark of GLP-1 is its ability to stimulate insulin secretion without the associated risk of hypoglycemia seen when using insulin therapy or some types of oral therapy that act by increasing insulin expression.

[0128] GLP-1 / glucagon receptor coagonists are disclosed in WO2008 / 086086, WO2008 / 101017, WO2007 / 056362, WO2008 / 152403 and WO96 / 29342. Other glucagon analogs disclosed are pegylated (WO2007 / 056362) or acylated at specific positions of native human glucagon (WO96 / 29342). Glucagon peptides are disclosed in U.S. Patent No. 7,314,859. The disclosures of each of the aforementioned GLP-1 analogs are incorporated herein by reference.

[0129] Liraglutide is an analog of human GLP-1 and acts as a GLP-1 receptor agonist. Liraglutide is indicated for the treatment of type 2 diabetes patients to improve glycemic control. U.S. Patent No. 6,268,343 discloses liraglutide and its formulations. U.S. Patent No. 8,114,833 discloses a pharmaceutical formulation comprising a GLP-1 receptor agonist, disodium phosphate dihydrate buffer and propylene glycol, the propylene glycol being present in the formulation at a final concentration of 1 mg / mL to 100 mg / mL, the formulation having a pH of 7.0 to 10.0. U.S. Publication No. 2010 / 0234299 discloses a pharmaceutical formulation of a GLP-1 compound, an isotonicity agent, a buffer and a preservative, the formulation having a pH of 7.0 to 10.0, and specifies that mannitol or NaCl are not isotonicity agents if the isotonicity agent is present and the pH of the formulation is 7.4.

[0130] GLP-1 analogs are either short-acting or long-acting, requiring different dosing schedules. However, normal physiology experiences a transient GLP-1 bolus induced by a meal, and not a long-term or steady-state GLP-1 gut hormone stimulation. Table 2 provides a list of long- and short-acting GLP-1 analogs.

[0131] Injectable GLP-1 agonists such as semaglutide (up to 2 mg injectable) can cause a series of serious side effects including severe allergic reactions including medullary thyroid cancer, nephritis, pancreatitis, vision changes, gallbladder and angioedema. The serious side effects are dose-related. Thus, the disclosed combination with oral denatonium salts allows the use of lower and safer doses of GLP-1 agonists, providing a safer treatment option for the existing accepted indications of GLP-1 agonists (HbA1C reduction, weight loss, glycemic control). [Table 2]

[0132] DPP-4 inhibitors DPP-4 inhibitors are used with diet and exercise to lower blood sugar in adults with type 2 diabetes. If left untreated or inadequately treated, or even if adequately treated, type 2 diabetes can lead to serious problems, including glaucoma, blindness, nerve and kidney damage, and heart disease. DPP-4 inhibitors are available as single-component products and in combination with metformin. Available DPP-4 inhibitors are sitagliptin, saxagliptin, vildagliptin, linagliptin, and alogliptin. However, it is known that when used alone, DPP-4 inhibitors can cause joint pain that can be severe and disabling. For example, oral administration of vildagliptin or sitagliptin to human type 2 diabetes has been found to reduce fasting blood sugar and postprandial blood sugar excursions in association with a significant reduction in HbA1c levels.

[0133] DPP-4 inhibitors act by inhibiting the degradation of GLP-1, GLP-2 and PYY, which all have essentially short half-lives.DPP-4 inhibitors have no effect on gastric emptying, are weight neutral, and have little or no perceptible effect on appetite.Therefore, DPP-4 inhibitors are only indicated for diabetes / glycemic control, and not for weight loss, obesity or hyperlipidemia. Reviews on the application of DPP-4 inhibitors for the treatment of type 2 diabetes include: (1) Demuth, et al., "Type 2 diabetes-Therapy with dipeptidyl peptidase IV inhibitors, Biochim. Biophys. Acta, 1751: 33-44 (2005); and (2) Augustyns et al., "Inhibitors of proline-specific dipeptidyl peptidases: DPP-4 inhibitors as a novel approach for the treatment of Type 2 diabetes," Expert Opin. Ther. Patents, 15: 1387-1407 (2005).

[0134] Sitagliptin phosphate has formula I below and is the dihydrogen phosphate salt of (2R)-4-oxo-4-[3-(trifluoromethyl)-5,6-dihydro[1,2,4]triazolo[4,3-a]pyrazin-7(8H)-yl]-1-(2,4,5-trifluorophenyl)butan-2-amine. [ka]

[0135] DPP-4 inhibitors are used in combination with other glycemic agents, such as metformin hydrochloride (U.S. Pat. No. 8,414,921). However, there is a need to find synergistic or additive combinations with bitter agonists, such as denatonium salts, based on the increased serum half-life of GLP-1 and PYY(1-36).

[0136] Regarding combination therapy, the oral GLP-RA semaglutide was compared with the oral DPP-4 inhibitors in Table 3. Standard measures of glycemic control, weight change and serum lipids were compared based on published results at different doses of each marketed drug. [Table 3]

[0137] Given the similar results achieved by oral administration of denatonium salts with different mechanisms of action, combination treatment with either oral GLP-1RA analogs or DPP-4 inhibitors would be effective in combination to treat diabetes / MetS / glycemic control, weight loss / obesity and hyperlipidemia.

[0138] GIP analog Another combination is the GIP and GLP-1 co-analog combination. Incretins are a group of metabolic hormones released in the intestine that stimulate a glucose-dependent decrease in blood glucose levels. Incretins include the peptide hormones GLP-1 and GIP. Incretin hormones are released in enteroendocrine cells after a meal. Both are dual incretin peptidomimetic compounds that stimulate receptors for both human GIP and GLP-1.

[0139] GLP-2 analogue There is one approved GLP-2 analogue, teduglutide (Gattex®). Teduglutide is a 33 amino acid glucagon-like peptide-2 analogue made in Escherichia coli (E. coli) by recombinant processing (without glycosylation). Teduglutide is injected sc (0.05 mg / kg) and is indicated for short bowel syndrome. Teduglutide has a half-life of 0.7-1.3 hours and is associated with a number of adverse events, including fluid retention (1%-12%); gastrointestinal reactions (12%-30%); antibody development (3%-54%; incidence increasing with prolonged use); injection site reactions (13%); upper respiratory tract infections (21%); and intestinal stoma complications (42%).

[0140] In addition, 11 other GLP-2 analogs have been identified in various stages of clinical or preclinical development for short bowel syndrome or chemotherapy-induced diarrhea. These agents are listed in Table 4: [Table 4]

[0141] PYY analogue PYY is released during a meal from L-cells in the distal small intestine and colon. PYY is known to have peripheral effects in the gastrointestinal (GI) tract. PYY is naturally secreted as a 36 amino acid peptide with a C-terminal amide (PYY(1-36)), but is cleaved to PYY(3-36), which accounts for approximately 50% of circulating PYY. The enzyme responsible for the degradation is dipeptidyl peptidase IV (DPP-4). PYY(3-36) is rapidly excreted by proteases and other clearance mechanisms. The half-life of PYY(3-36) has been reported to be <30 minutes in pigs (Ito T et al, Journal of Endocrinology (2006), 191, ppl13-119). Thus, PYY exhibits suboptimal pharmacokinetic properties. This means that the peptide must be administered at least twice daily, and possibly once daily, together with a DPP-4 inhibitor.

[0142] Whereas PYY(1-36) activates the Y1, Y2 and Y5 receptors with little selectivity and the Y4 receptor to a lesser extent, DPP-4-processed PYY(3-36) retains some Y1 and Y5 affinity but shows enhanced selectivity for the Y2 receptor relative to the Y1, Y4 and Y5 receptors. Y1 and Y5 receptor activation leads to increased appetite and food intake, whereas Y2 receptor activation reduces appetite and food intake. Furthermore, Y1 and Y5 receptor activation can lead to increased blood pressure.

[0143] Based on the effects shown, for example, in Zucker rats and diet-induced obese (DIO) mice, Y2-selective PYY(3-36) analogs have shown positive effects on glucose (van den Hoek A. et al., Am. J. Physiol. Endocrinol. Meta. (2006), 292, ppE238-E245; and Ortiz A. et al, The Journal of Pharmacology and Experimental Therapeutics (2007), 323, pp 692-700). WO2009 / 138511, WO2011 / 033068 and WO2011 / 058165 disclose long-acting Y2 and / or Y4 receptor agonists, PYY analogs stabilized against C-terminal proteolysis, and Y2 receptor agonists with prolonged pharmacokinetic properties, respectively.

[0144] There are three PYY analogs found in development, including NN-9775 (Novo-Nordisk), a synthetic peptide PYY analog that activates the hypothalamic NPY-Y2 autoreceptor in Phase 1 clinical trials for obesity; JNJ-0321 (J&J) synthetic peptide as a long-acting PYY analog for obesity in preclinical development; and Zihipp, Ltd. PYY analogs in very early stages for obesity. Similar to GLP-2 or GLP-2 analogs, PYY analogs also have the challenges of retaining function at the target receptor, increasing immunogenicity (antibody formation), and raising the potential risk of adverse effects due to long-acting signaling that does not reflect normal physiological function. Thus, despite the development of several such gut hormone analogs, almost all of which require injection (with the exception of oral GLP-1 analog Ryversus®, which has a lipid additive for daily oral administration but requires a much higher dose), such biopeptide gut hormone analogs are not suitable for oral delivery.

[0145] CCK analog CCK is also a gut-secreted peptide hormone with appetite suppressant properties. However, unlike PYY, which has a half-life of 9-14 minutes, CCK half-life is 2-3 minutes. Developing CCK analogs include C-2816 (Astra-Zeneca), a fusion peptide for both receptors, GLP-1R agonist AC3174+CCKR1 agonist AC17022; a synthetic peptide CCK analog for obesity, NN-9056 (Novo-Nordisk); University of Nebraska CCKR8 analog synthetic peptide; and A-71378 (AbbVie) CCK-8 analog synthetic peptide for obesity, which appears to have been discontinued.

[0146] Gut Peptide Hormones (GPH) Symphony Orchestra Each of the described gut peptide hormones (GLP-1, GLP-2, GIP, PYY and CCK) has various analogs on the market or in development individually for glycemic control / diabetes and weight loss / obesity. Additionally, DPP-4 is the enzyme that breaks down GLP-1 and PYY, and several orally active enzyme inhibitors are available. The present disclosure provides a conductor to the symphony orchestra of multiple gut peptide hormones, bitter taste receptor agonists (which can also play a role in all sections of the gut peptide hormone orchestra). Because gut signaling is driven by multiple gut peptide hormones, not just one, there is a need to treat glycemic control / diabetes, weight loss / obesity and hyperlipidemia by addressing multiple gut peptide hormones, not just a single one. Thus, given the surprising data provided herein showing multiple relevant gut peptide hormone increases in all animal studies and Phase 1 human clinical trials, the combination disclosed herein enhances single gut peptide hormone treatment by providing agonist activity to multiple gut peptide hormones with the addition of the denatonium salt component of the combination.

[0147] Pharmaceutical Compositions and Pharmaceutically Acceptable Carriers The pharmaceutical composition described herein and / or for use in the method described herein may further comprise a pharma- ceutically acceptable carrier.In some embodiments, pharma-ceutically acceptable carrier means a pharma-ceutically acceptable substance, material, composition or vehicle, such as a diluent, solid excipient, additive or manufacturing aid (e.g., lubricant, talc, magnesium, calcium or zinc stearate or steric acid), that aids in the process of delivery of API to patient and / or stabilizes API during transportation for delivery to patient.The term "acceptable" when used in this sense means that the material is compatible with other components of the formulation and does not cause intolerable adverse effects that are harmful to the patient.

[0148] In the solid pharmaceutical dosage forms of the invention for oral administration disclosed herein (capsules, tablets, pills, powders, granules, etc.), the API may be mixed with one or more pharma- ceutically acceptable excipients, such as sodium citrate or dicalcium phosphate, and / or a pharma- ceutically acceptable carrier, including any of the following: (1) excipients or fillers, such as starch, lactose, sucrose, glucose, mannitol, and / or silicic acid; (2) binders, such as carboxymethylcellulose, alginates, gelatin, polyvinylpyrrolidone, sucrose, and / or acacia; (3) humectants, such as glycerol; (4) disintegrants, such as agar-agar, calcium carbonate, potato starch, or tapioca starch, Alginic acid, certain silicates and sodium carbonate; (5) solution disintegrants, such as paraffin; (6) absorption enhancers, such as quaternary ammonium compounds, and surfactants, such as poloxamers and sodium lauryl sulfate; (7) wetting agents, such as cetyl alcohol, glycerol monostearate and nonionic surfactants; (8) absorbents, such as kaolin and bentonite clay; (9) lubricants, such as talc, calcium, magnesium stearate, solid polyethylene glycols, sodium lauryl sulfate, zinc stearate, sodium stearate, stearic acid, and mixtures thereof; (10) coloring agents; and (11) controlled release agents, such as crospovidone or ethylcellulose. In the case of capsules, tablets and pills, the pharmaceutical compositions may also contain buffering agents. Solid compositions of a similar type may also be used as excipients for soft and hard shell gelatin capsules using additives such as lactose or milk sugar, and high molecular weight polyethylene glycols, etc.

[0149] Further Alternative Embodiments The present disclosure provides the following embodiment of a method for treating or preventing the progression of fatty liver disease ("Method 1"), comprising administering to a subject with fatty liver disease a combination of a denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate; and a GLP-1 receptor agonist (e.g., semaglutide, glyburide, liraglutide, dulaglutide, albiglutide, exenatide or lixisenatide). In some embodiments, the method further comprises administering about 0.5 g to about 5 g acetic acid. Preferably, the daily dosage of acetic acid for an adult is about 1.5 g to about 3 g. 1.1 Method 1, wherein the fatty liver disease is NASH. 1.2 Method 1 or 1.1, wherein the fatty liver disease is NASH or NAFLD or ASH. 1.3 Any one of the preceding processes, wherein the denatonium salt is denatonium acetate. 1.4 Any one of the preceding methods, wherein the denatonium salt is denatonium citrate. 1.5 Any one of the preceding processes, wherein the denatonium salt is denatonium maleate. 1.6 Any one of the preceding methods wherein the daily dose of the denatonium salt is 200 mg administered QD or BID. 1.7 Any one of the preceding methods wherein the daily dose of the denatonium salt is 400 mg administered BID. 1.8 Any one of the preceding methods, wherein the daily dose of the denatonium salt is 600 mg administered QD or BID. 1.9 Any one of the preceding methods wherein the daily dose of the denatonium salt is 1000 mg administered BID. 1.10 Any one of the preceding methods wherein the GLP-1 receptor agonist is semaglutide. 1.11 Any one of methods 1.1 to 1.10, wherein the GLP-1 receptor agonist is glyburide. 1.12 Any one of methods 1.1 to 1.10, wherein the GLP-1 receptor agonist is liraglutide. 1.13 Any one of methods 1.1 to 1.10, wherein the GLP-1 receptor agonist is dulaglutide. 1.14 Any one of methods 1.1 to 1.10, wherein the GLP-1 receptor agonist is albiglutide. 1.15 Any one of methods 1.1 to 1.10, wherein the GLP-1 receptor agonist is exenatide. 1.16 Any one of methods 1.1 to 1.10, wherein the GLP-1 receptor agonist is or lixisenatide.

[0150] In another embodiment, the present disclosure provides Method 2, which includes a method for treating obesity and / or achieving weight loss by administering an effective amount of an orally administered pharmaceutical composition in a single dosage form or in two dosage forms, wherein the pharmaceutical composition comprises a bitter taste receptor agonist and a gut signaling compound. Preferably, in method 2, the bitter taste receptor agonist is a denatonium salt, and the denatonium salt is selected from the group consisting of DA, denatonium citrate, denatonium maleate, denatonium saccharide, and denatonium tartrate; and the gut signaling compound is (a) a gut peptide hormone analog selected from the group consisting of GLP-1RA, GLP-2 analogs, PYY analogs, GIP analogs, CCK analogs, and combinations thereof; or (b) a DPP-4 inhibitor selected from the group consisting of salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin, and alogliptin.

[0151] Further, another embodiment of Method 2 includes: 2.1 Method 2, wherein the GLP-1 analogue is semaglutide, glyburide, liraglutide, dulaglutide or albiglutide. 2.2 Method 2, wherein the PYY 1875 analog is NN-9775 or JNJ-9321. 2.3 Method 2, wherein the CCK analog is C-2819, NN-9056 or A-71378. 2.4 Method 2, wherein the GLP-2 analogue is teduglutide, grepaglutide, apraglutide, elsigutide, HM-15912, ZP-7570 GLP-2-ELP MOD-1501 or HL-06. 2.5 Method 2, wherein the gut signaling compound is a DPP-4 inhibitor selected from the group consisting of sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin, and alogliptin, administered in a daily dose that is the approved daily dose for the particular DPP-4 inhibitor. 2.6 Method 2.5, wherein the DPP-4 inhibitor is administered once daily (QD) or twice daily (BID) about 8 hours apart, and the denatonium salt is administered once or twice daily at a total daily dose (by weight of denatonium) of about 50 mg to about 3000 mg, preferably about 100 mg to about 2000 mg, and most preferably about 200 mg to about 1000 mg.

[0152] In another embodiment, a bitter taste receptor agonist comprising a denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide, and denatonium tartrate; and (a) a gut peptide hormone analog selected from the group consisting of GLP-1RA, GLP-2 analogs, PYY analogs, GIP analogs, CCK analogs, and combinations thereof; or (b) a salt of a medium chain fatty acid, N-(8-(2-hydroxybenzoyl)acetate, N-(8 ... Method 3 includes a method for treating glycemic control, metabolic syndrome (MetS) and / or diabetes by administering an effective amount of an orally administered pharmaceutical composition comprising a combination of either a salt of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin and alogliptin, in a single dosage form or in two dosage forms.

[0153] Further embodiments of Method 3 include: 3.1 Method 3, wherein the method further comprises administering a DPP-4 inhibitor selected from salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin, saxagliptin, linagliptin, and alogliptin. 3.2 Method 3, wherein the GLP-1 analogue is semaglutide, glyburide, liraglutide, dulaglutide or albiglutide. 3.3 Method 3, wherein the PYY 1875 analog is NN-9775 or JNJ-9321. 3.4 Method 3, wherein the DPP-4 inhibitor is selected from sitagliptin phosphate, vildagliptin, linagliptin, alogliptin, saxagliptin, P93 / 01, SYR322, GSK 823093, Roche 0730699, TS021, E3024, and PHX-1149. 3.5 Method 3, wherein the GLP-2 analogue is teduglutide, grepaglutide, apraglutide, elsigutide, HM-15912, ZP-7570, GLP-2-ELP, MOD-1501 or HL-06. 3.6 Method 3, wherein the orally administered pharmaceutical composition is administered in a dosage of a commercially approved DPP-4 inhibitor selected from the group consisting of sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin, and alogliptin, with the daily dosage being the approved daily dose for the particular DPP-4 inhibitor administered once daily (QD) or twice daily (BID) about 8 hours apart; and the denatonium salt is administered once or twice daily in a total daily dose (per weight of denatonium) of about 50 mg to about 3000 mg, preferably about 100 mg to about 2000 mg, and most preferably about 200 mg to about 1000 mg.

[0154] In another embodiment, there is provided a method 4, which is a method for treating hyperlipidemia, comprising administering an effective amount of an orally administered pharmaceutical composition comprising a bitter taste receptor agonist comprising a denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide, and denatonium tartrate; and either (a) a gut peptide hormone analog selected from the group consisting of glucagon-like peptide (GLP-1) analogs, GLP-2 analogs, PYY analogs, GIP analogs, CCK analogs, and combinations thereof; or (b) a DPP-4 inhibitor selected from the group consisting of salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin, and alogliptin, in a single dosage form or in two dosage forms.

[0155] A further embodiment of this method 4 includes: 4.1 Method 4, wherein the method further comprises administering a DPP-4 inhibitor selected from salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin, saxagliptin, linagliptin, and alogliptin. 4.2 Method 4, wherein the GLP-1 analogue is semaglutide, glyburide, liraglutide, dulaglutide or albiglutide. 4.3 Method 4, wherein the PYY 1875 analog is NN-9775 or JNJ-9321. 4.4 Method 4, wherein the DPP-4 inhibitor is selected from sitagliptin phosphate, vildagliptin, linagliptin, alogliptin, saxagliptin, P93 / 01, SYR322, GSK 823093, Roche 0730699, TS021, E3024 and PHX-1149. 4.5 Method 4, wherein the GLP-2 analogue is teduglutide, grepaglutide, apraglutide, elsigutide, HM-15912, ZP-7570, GLP-2-ELP, MOD-1501 or HL-06. 4.6 Method 4, wherein the orally administered pharmaceutical composition is administered in a dosage of a commercially approved DPP-4 inhibitor selected from the group consisting of sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin, and alogliptin, in a daily dosage that is the approved daily dosage for the particular DPP-4 inhibitor administered once daily (QD) or twice daily (BID) about 8 hours apart; and the denatonium salt is administered once or twice daily in a total daily dosage of about 50 mg to about 3000 mg, preferably about 100 mg to about 2000 mg, and most preferably about 200 mg to about 1000 mg (by weight of denatonium).

[0156] In another embodiment, Method 5 is provided, which is a method for lowering the administered dose of a GLP-1RA drug, comprising co-administering a bitter taste receptor agonist comprising a denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide, and denatonium tartrate with the GLP-1RA drug.

[0157] A further embodiment of Method 5 includes: 5.1 Method 5, wherein the GLP-1RA is semaglutide, glyburide, liraglutide, dulaglutide or albiglutide. 5.2 Method 5, wherein the denatonium salt is DA.

[0158] Alternatively, the present disclosure further provides a combination product. For example, according to one embodiment (combination 1) of the present invention, an oral dosage form pharmaceutical composition is provided that includes a bitter taste receptor agonist and a dipeptidyl peptidase-4 (DPP-4) inhibitor.

[0159] Further embodiments of Combination 1 include: 1.1 Combination 1, wherein the bitter taste receptor agonist is selected from the group consisting of denatonium salts, including acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate, chlorpheniramine, diphenidol, famotidine, haloperidol, quinine, parthenolide and aristolochic acid. 1.2 Combination 1, wherein the DPP-4 inhibitor is selected from the group consisting of salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin, saxagliptin, linagliptin, alogliptin, and combinations thereof.

[0160] Alternatively, according to another embodiment (Combination 2), there is provided a synergistic combination pharmaceutical composition comprising a formulation (oral formulation) of a bitter taste receptor agonist and a gut signaling peptide analog and a gut signaling hormone enhancer selected from the group consisting of a glucagon-like peptide (GLP-1) analog, a peptide YY (PYY) analog, a dipeptidyl peptidase-4 (DPP-4) inhibitor and a glucose-dependent insulinotropic polypeptide (GIP) analog.

[0161] Further embodiments of Combination 2 include: 2.1 Combination 2, wherein the pharmaceutical combination further comprises a DPP-4 inhibitor. 2.2 Combination 2, wherein the bitter taste receptor agonist is selected from the group consisting of denatonium salts, including acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate, chlorpheniramine, diphenidol, famotidine, haloperidol, quinine, parthenolide and aristolochic acid. 2.3 Combination 2, wherein the DPP-4 inhibitor is selected from the group consisting of salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin, saxagliptin, linagliptin, alogliptin, and combinations thereof.

[0162] According to another embodiment of the present invention (Combination 3), there is provided a combination pharmaceutical composition for oral administration comprising a formulation of a bitter taste receptor agonist comprising a denatonium salt selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide, and denatonium tartrate; and a DPP-4 inhibitor selected from the group consisting of salts of medium chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin salts, saxagliptin, linagliptin, alogliptin, and combinations thereof.

[0163] Further embodiments of Combination 3 include: 3.1 Combination 3, wherein the denatonium salt in the single oral dosage form is denatonium acetate and the DPP-4 inhibitor is sitagliptin. 3.2 Combination 3, further comprising an oral dosage form of the GLP-1RA semaglutide. 3.3 Combination 3, in which the dosage of a commercially approved DPP-4 inhibitor selected from the group consisting of sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin phosphate, saxagliptin, linagliptin and alogliptin is a daily dose that is the approved daily dose for the particular DPP-4 inhibitor administered once daily (QD) or twice daily (BID) about 8 hours apart, and the denatonium salt is administered once or twice daily at a total daily dose (per weight of denatonium) of about 50 mg to about 3000 mg, preferably about 100 mg to about 2000 mg, and most preferably about 200 mg to about 1000 mg. In one embodiment, the dosage is 1000 mg.

[0164] For each of the methods and combinations enumerated herein (e.g., Methods 1-5 and Combinations 1-3 above and each subpart thereof), it is contemplated that different embodiments and dosages of denatonium salts may be used and administered to a human patient. For example, in some embodiments, the dosage of denatonium salts is about 0.5 mg / kg BID to about 30 mg / kg BID, or, optionally, about 1 mg / kg BID to about 20 mg / kg BID. In other embodiments, the dosage of denatonium salts administered to a human patient is about 1.0 mg / kg / day to about 60 mg / kg / day; in some embodiments, about 2 mg / kg / day to about 40 mg / kg / day; and, optionally, about 4 mg / kg / day to about 20 mg / kg / day. In another embodiment, the dosage range of the denatonium salts used in the methods and combinations of the invention is from about 0.1 mg / kg / day to about 32 mg / kg / day, preferably from about 0.25 mg / kg / day to about 16 mg / kg / day; and most preferably from about 0.5 mg / kg / day to about 8 mg / kg / day. EXAMPLES

[0165] Reference will now be made in detail to certain specific embodiments which are illustrated in the following examples and accompanying drawings. It will be understood that while the disclosure provides exemplary embodiments, the examples are not intended to limit the disclosure to those embodiments. Rather, the present invention is intended to encompass all alternatives, modifications and equivalents which may be appreciated by one skilled in the art from the present disclosure.

[0166] [Example 1a] Preparation of DA, bitter taste receptor agonists [ka] Denatonium acetate anhydride, or DA, is an anhydrous salt in which there are 83 mg of denatonium cation and 17 mg of acetate anion per 100 ml of DA.

[0167] This Scheme A describes the synthesis of denatonium acetate (DA).

[0168] Step 1: Synthesis of denatonium hydroxide from lidocaine

[0169] Add 25 g of lidocaine, 60 ml of water and 17.5 g of benzyl chloride to the reflux apparatus with stirring and heating at 70-90 ° C. The solution should be heated and stirred at the previously given values ​​for 24 hours, and the solution should be cooled to 30 ° C. Remove the unreacted reagents with 3 × 10 mL of toluene. Dissolve 65 g of sodium hydroxide in 65 mL of cold water with stirring and add this to the aqueous solution with stirring over 3 hours. Filter the mixture, wash with water and dry in open air. Recrystallize with hot chloroform or hot ethanol. [ka]

[0170] Step 2: Preparation of denatonium acetate anhydride from denatonium hydroxide

[0171] Add 10 g of denatonium hydroxide (MW: 342.475 g / mol, 0.029 mol), 20 mL of acetone, and 2 g of glacial acetic acid (0.033 mol) dissolved in 15 mL of acetone to the reflux apparatus, stir and heat the mixture for 3 hours at 35° C. Then evaporate to dryness and recrystallize with hot acetone. [ka]

[0172] [Example 1b] Preparation of pharmaceutical compositions containing DA A formulation of DA particles that can be formed into tablets or filled into capsules for oral delivery avoiding oral exposure.

[0173] This example provides an immediate release 50 mg granule formulation of denatonium acetate (DA) as the free base as an immediate gastric release oral pharmaceutical formulation. The detailed manufacturing process is described below.

[0174] 1. Drug layering process-drug layered pellets The drug layering process was carried out in a fluid bed granulator equipped with a rotor insert (rotor granulator). The drug solution was prepared by solubilizing povidone K30 (Kollidon 30) and denatonium acetate in ethyl alcohol. The drug solution was sprayed tangentially onto the bottom of white sugar spherical granules (35 / 45 mesh) moving in a circular motion in the rotor granulator. The final drug-loaded pellets were then dried in the rotor granulator for 10 minutes, discharged, and screened through a #20 mesh.

[0175] 2. Seal Coating Process-Seal Coating Pellets The seal coating dispersion was prepared by dissolving Hypromellose E5 separately in a mixture of ethyl alcohol and purified water (1:1) until a clear solution was obtained. The remaining amount of ethyl alcohol was then added to the above solution followed by the addition of talc. The dispersion was mixed for 20 minutes to allow for uniform dispersion of the talc. The seal coating dispersion was sprayed tangentially onto the drug loaded pellets to achieve a 5% weight gain. The seal coated pellets were then dried in a rotor granulator for 5 minutes, discharged and further dried in a tray drier / oven at 55°C for 2 hours. The seal coated pellets were then screened through a #20 mesh.

[0176] 3. Final Mix – Denatonium Immediate Release (IR) Pellets The seal coated pellets were mixed with talc screened through mesh #60 using a V-blender for 10 minutes and discharged. The mixed seal coated beads, denatonium IR pellets, were used for encapsulation.

[0177] 4. Encapsulation – Denatonium Capsules 50mg Denatonium IR pellets (50 mg) from step 3 were filled into size 1, white opaque hard gelatin capsules using an automated capsule filling machine. The capsules were then passed through an in-line capsule polisher and metal detector. In-process control of capsule weight and appearance was performed during the encapsulation process. Acceptable Quality Level (AQL) sampling and testing was performed by Quality Assurance (QA) on composite samples during the encapsulation process. Final product composite samples were collected and analyzed per release testing specifications.

[0178] 5. Packaging - Capsules 50mg - 30 pieces The 50 mg capsules were packaged in 30-packs into 50 / 60cc white HDPE round S-line bottles with 33mm white CRC caps. The bottles were rolled and sealed using an induction sealer.

[0179] Table 5 shows the qualitative and quantitative formulation composition of DA. [Table 5]

[0180] [Example 2] This example describes an in vivo study in a high-fat diet-induced obesity (DIO) mouse model after 4 weeks of treatment with denatonium acetate, liraglutide (GLP-1 agonist), or their combination. This study included 4 treatment groups, with 15 mice assigned to each group: (1) a vehicle treatment group, with distilled water, administered orally (PO) twice a day (BID), and sterile 0.9% saline solution, administered subcutaneously (SC) BID; (2) a denatonium acetate (DA) treatment group, with DA 75 mg / kg (denatonium salt weight), administered PO BID; (3) a liraglutide treatment group, with liraglutide 200 μg / kg, administered SC BID; and (4) a combination treatment group, with DA 75 mg / kg (denatonium salt weight), administered PO BID, plus liraglutide 200 μg / kg, administered SC BID. The dosing regimen used is shown in Table 6. [Table 6]

[0181] Upon arrival at the testing facility, all animals were acclimated to the vivarium for a period of at least 3 days, kept on a 60% fat rodent diet (D12492; research diet), a 12:12 dark / light cycle, and group-housed 2–3 animals in hepatofilter cages. All treatments lasted for 4 weeks. During the study period, gross observations (animal behavior and clinical signs) were performed, and body weight measurements were taken for each animal three times per week. At the end of the study, all animals were fasted overnight before measuring serum levels of glucose, insulin, hemoglobin A1c (HbA1c), low-density lipoprotein (LDL) cholesterol, high-density lipoprotein (HDL) cholesterol, triglycerides (TG), and bile acids (BA). Statistical analysis of the data was performed using a one-tailed Student's t-test in Excel.

[0182] Figure 1 shows the average weight gain over the study period for all treatment groups. Table 7 shows the weight gain at the end of the study (day 31) for each animal (anima) in all treatment groups. These data demonstrate that (1) weight gain was significantly reduced in DIO mice when treated with DA, liraglutide, or their combination; and (2) the combination of DA and liraglutide resulted in significantly lower weight gain compared to DA or liraglutide alone, indicating a potential synergistic (or at least additive) effect between the two agents on weight gain. [Table 7-1] [Table 7-2]

[0183] Table 8 and Figure 2 show the serum TG levels at the end of the study (day 31) for each animal in the four treatment groups. These data show that (1) treatment with DA, liraglutide, or their combination significantly reduced serum TG levels in DIO mice compared to vehicle; and (2) animals treated with the combination exhibited significantly lower serum TG levels compared to those treated with DA or liraglutide alone, indicating a potential synergistic (or at least additive) effect between the two agents on serum TG levels. [Table 8-1] [Table 8-2]

[0184] Figure 3 shows serum glucose levels at the end of the study (day 31) for each animal in the four treatment groups. Treatment with DA, liraglutide, or their combination significantly reduced serum glucose levels in DIO mice when administered for 4 weeks.

[0185] Figure 4 shows serum HbA1c levels at the end of the study (day 31) for each animal in the four treatment groups. The results suggest that treatment with DA, liraglutide, or their combination had no significant effect (p>0.05) on serum HbA1c levels in DIO mice after 4 weeks of treatment.

[0186] Figure 5 shows serum insulin levels at the end of the study (day 31) for each animal in the four treatment groups. The data demonstrate that 4 weeks of treatment with DA, liraglutide, or their combination significantly reduced serum insulin levels in DIO mice.

[0187] Figure 6 shows serum BA levels at the end of the study (day 31) for each animal in the four treatment groups. When administered for 4 weeks, DA, liraglutide, or the combination resulted in a significant increase in serum BA levels compared to the vehicle control.

[0188] Figure 7 shows serum LDL levels at the end of the study (day 31) for animals in each of the four treatment groups. There were no significant differences in serum LDL levels in animals treated with either vehicle or DA, liraglutide, or their combinations.

[0189] Figure 8 shows serum HDL levels at the end of the study (day 31) for each animal in the four treatment groups. The data reveal that compared to vehicle, treatment with liraglutide, or the combination of DA + liraglutide, resulted in a significant decrease in serum HDL levels in DIO mice after 4 weeks of treatment.

[0190] [Example 3] This example provides data for three gut peptide hormones from a Phase 1 clinical trial of denatonium acetate after a single dose of DA in a 240 mg tablet of the formulation disclosed herein. Subjects received either placebo or 240 mg DA, and blood samples were taken immediately prior to dosing and 1 hour after oral administration. Figure 9 shows the percent change in GLP-1 from 0 hours to 1 hour after oral administration. Even with a small sample size, the difference was significant (p=0.0235). Figure 10 shows the results for GLP-2, which showed a trend toward increased gut peptide hormones. Figure 11 shows the results for PYY, which showed a trend toward increased gut peptide hormones. Thus, data from this human clinical trial showed that DA exerts its weight loss effects in the GI tract, but stimulates the release of gut hormones GLP-1, GLP-2, and PYY.

[0191] [Example 4] This example provides results from an in vivo study of denatonium acetate and sitagliptin in high-fat diet-induced obese (DIO) mice. C57BL / 6Ntac mice (at least 12 weeks old) were fed a high-fat diet. All mice were orally administered: (a) vehicle group (N=15) receiving distilled water, BID; (b) DA group (N=15) receiving 75 mg / kg (denatonium weight), BID; (c) sitagliptin group (N=15) receiving 10 mg / kg, QD by gavage; and (d) DA+sitagliptin group (N=15) treated with DA (denatonium weight) 75 mg / kg, BID, and sitagliptin 10 mg / kg by gavage, QD. The groups were administered for 8 weeks followed by a 5-7 day sitagliptin period. Body weight and body weight change were measured 3 times a week. Blood glucose (after 6–8 h of fasting), blood insulin, percentage of blood HvA1c, serum biomarker levels of HDL, LDL, total triglycerides (TG), total cholesterol (TC) and bile acids were measured twice during the study, on day 28 and at the end of the study (day 56). Cumulative food intake and water consumption of each animal were measured.

[0192] All 60 animals tolerated the given treatment well, and no significant toxic side effects were observed during the study. Figures 12A and 12B show the relative body weight percentage (12A) and relative body weight change (g) (12B) for the four groups of animals. Treatment with sitagliptin alone had the least effect on body weight, consistent with previous studies and clinical experience. However, a much larger effect on body weight was seen with DA alone (ARD-101), while a significant or synergistic effect on body weight was seen with the combination of DA and sitagliptin.

[0193] Thus, in Example 3 and the accompanying figures, DA (denatonium salt with denatonium as cation and acetate as anion) showed superiority to sitagliptin (representative of all other DPP-4i class drugs) in a one-to-one fashion. A statistically significant as well as significant weight loss was observed in the DA group versus the sitagliptin group. Furthermore, unlike the "rebound" effect observed over time with sitagliptin (consistent with Example 4 and clinical observations of the effect of DPP-4i drugs on obesity), DA continued to show sustained weight loss and other parameter effects throughout the 56-day study period (Example 3). It should be noted that the combination of sitagliptin and DA together induced even more significant and statistically significant weight loss benefits compared to either drug alone. These data showed significant synergy (the total effect measured was greater than the sum of each of the component drugs in the combination). Furthermore, no toxic effects were observed in animals with the combination of both sitagliptin (and by extension other DPP-4i drugs) and DA, indicating that there are significant cumulative toxic effects of concern that would limit either co-administration or a single combination dosage form (Example 3 data).

[0194] Figures 13 to 22B show data from this Example 4, which is further described above under the heading "Brief Description of the Figures". Data from this study showed that the combination of bitter taste receptor agonist (denatonium acetate or ARD-101) and DPP-4 inhibitor (sitagliptin) showed a synergistic and significant effect on the body weight of DIO mice. Given that DPP-4 inhibitors have not achieved the efficacy of weight loss as marketed drugs (but have shown to be effective treatments for type 2 diabetes and HbA1c reduction), combining DPP-4 inhibitors with bitter taste receptor agonists such as denatonium acetate provides synergistic benefits for weight loss (treatment of obesity), diabetes / glycemic control and hyperlipidemia.

[0195] [Example 5] Combined pharmaceutical composition for oral administration Combining two or more oral antidiabetic agents into a single tablet offers a potential means of delivering combination therapy without adding to the complexity of a patient's daily regimen. Such formulations have been well accepted in other disease indications, such as for hypertension (HYZAAR™, a combination of losartan potassium and hydrochlorothiazide) and cholesterol lowering (VYTORIN™, a combination of simvastatin and ezetimibe). Examples of commercially available combination tablets containing two oral antidiabetic agents include metformin and the DPP-4 inhibitor sitagliptin (Janumet®), saxagliptin (Kombiglyze®), linagliptin (Jentadueto®), and alogliptin (Kazano®).

[0196] This example provides a formulation of a DA tablet mixed with a DPP-4 inhibitor, provided that the DPP-4 inhibitor is released immediately before the denatonium salt, such that the circulating DPP-4 inhibitor can increase the half-life of the GLP-1 and PYY gut peptide hormones that are stimulated for release by the denatonium salt in the small intestine.

[0197] This example provides an immediate release 100mg particle formulation of denatonium acetate (DA) as free base and a non-granular water-soluble DPP-4 inhibitor as an immediate gastric release oral pharmaceutical formulation.In one embodiment, immediate release includes release in the stomach or intestine to avoid or minimize oral cavity exposure.This embodiment provides the advantage of avoiding subjective taste aversion to API.

[0198] The detailed manufacturing process is described below.

[0199] 1.~3. Drug layer / seal coating / final blending The drug layering, seal coating and final blending process described above in Example 1B was carried out to produce the blended seal coated beads, denatonium IR pellets, used for encapsulation.

[0200] 4. Encapsulation – Denatonium Capsules 100mg Denatonium IR pellets (100 mg) were filled into size 1, white opaque hard gelatin capsules using an automated capsule filling machine. The capsules were then passed through an in-line capsule polisher and metal detector. In-process control of capsule weight and appearance was performed during the encapsulation process. Acceptable Quality Level (AQL) sampling and testing was performed by Quality Assurance (QA) on composite samples during the encapsulation process. Final product composite samples were collected and analyzed per release testing specifications.

[0201] 5. Packaging - Capsules 100mg - 30pcs The 100 mg capsules were packaged in 30-packs in 50 / 60cc white HDPE round S-line bottles with 33mm white CRC caps. The bottles were rolled and sealed using an induction sealer.

[0202] Table 9 shows the qualitative and quantitative formulation composition of the DA / DDP4i combination capsule according to this example. [Table 9]

[0203] The bilayer tablet will contain a first layer of a bitter taste receptor agonist, preferably a denatonium salt, and a second layer of a DPP-4 inhibitor. The DPP-4 inhibitor is selected from the group consisting of sitagliptin, vildagliptin, saxagliptin, P93 / 01, SYR322, GSK 823093, Roche 0730699, TS021, E3024 and PHX-1149. Preferably, the DPP-4 inhibitor is alogliptin, carmegliptin, melogliptin, dutogliptin, denagliptin, linagliptin, sitagliptin, vildagliptin or saxagliptin. In a subclass of this class, the DPP-4 inhibitor is sitagliptin.

[0204] A preferred pharma- ceutically acceptable salt of sitagliptin is the dihydrogen phosphate salt (sitagliptin phosphate). A preferred form of sitagliptin dihydrogen phosphate salt is the crystalline monohydrate (sitagliptin phosphate monohydrate) disclosed in WO 2005 / 0031335, the disclosure of which is incorporated herein by reference.

[0205] The preparation of sitagliptin phosphate monohydrate is disclosed in International Patent Publication WO 2005 / 0031335, published Jan. 13, 2005, the contents of which are incorporated by reference.

[0206] Dosage strengths of the DPP-4 inhibitor for incorporation into pharmaceutical compositions are from about 1 mg to about 250 mg of active moiety. Preferred dosage strengths of the DPP-4 inhibitor are from about 25 mg to about 200 mg of active moiety. Individual dosage strengths are 25, 50, 75, 100, 150 and 200 mg equivalents of the DPP-4 inhibitor active moiety. By "active moiety" is meant the free base form of the DPP-4 inhibitor as the anhydrate.

[0207] The unit dosage strengths of sitagliptin free base anhydrate (active moiety) for inclusion in the fixed dose combination pharmaceutical composition are 25, 50, 75, 100, 150 or 200 mg. The preferred dosage strengths of sitagliptin are 50 (for BID) or 100 mg daily. Equivalent amounts of sitagliptin phosphate monohydrate to sitagliptin free base anhydrate, i.e., 32.13, 64.25, 96.38, 128.5, 192.75 and 257 mg, respectively, are used in the pharmaceutical composition.

[0208] Dosage strengths of denatonium salts are from about 50 mg to about 3000 mg, preferably from about 100 mg to about 2000 mg, and most preferably from about 200 mg to about 1000 mg total daily (by weight of denatonium) administered once or twice daily.

[0209] The pharmaceutical composition comprises: (a) a second layer comprising about 20-45% by weight of a dipeptidyl peptidase-4 inhibitor or a pharma- ceutical acceptable salt thereof; and (b) a first layer comprising about 7-24% by weight of a bitter taste receptor agonist; Includes.

[0210] The second layer further comprises one or more additives selected from the group consisting of: (i) a diluent; (ii) a disintegrant; and (iii) a lubricant. In a subclass of this class, the first layer further comprises one or more additives selected from the group consisting of: (i) two diluents; (ii) a disintegrant; and (iii) two lubricants.

[0211] The second layer further comprises one or more additives selected from the group consisting of: (i) about 40-80% by weight of a diluent; (ii) about 0.5-6% by weight of a disintegrant; and (iii) about 0.75-10% by weight of a lubricant. In a subclass of this class, the second layer further comprises one or more additives selected from the group consisting of: (i) about 40-80% by weight of two diluents; (ii) about 0.5-6% by weight of a disintegrant; and (iii) about 0.75-10% by weight of two lubricants.

[0212] Alternatively, the second layer further comprises one or more additives selected from the group consisting of: (i) about 20-40% by weight of a first diluent; (ii) about 20-40% by weight of a second layer diluent; (iii) about 0.5-6% by weight of a disintegrant; (iv) about 0.25-4% by weight of a first lubricant; and (v) about 0.5-6% by weight of a second lubricant. In a subclass of this class, the first diluent is microcrystalline cellulose; the second diluent is anhydrous calcium hydrogen phosphate; the disintegrant is croscarmellose sodium; the first lubricant is magnesium stearate; and the second lubricant is sodium stearyl fumarate.

[0213] The dipeptidyl peptidase-4 inhibitor is selected from the group consisting of alogliptin, carmegliptin, denagliptin, dutogliptin, linagliptin, melogliptin, saxagliptin, sitagliptin and vildagliptin, or a pharmaceutically acceptable salt thereof. In another class of this embodiment, the dipeptidyl peptidase-4 inhibitor is selected from the group consisting of sitagliptin, vildagliptin and saxagliptin, or a pharmaceutically acceptable salt thereof. In a subclass of this class, the dipeptidyl peptidase-4 inhibitor is sitagliptin or its dihydrogen phosphate salt.

[0214] Alternatively, the pharmaceutical composition comprises: (a) a second layer comprising: (i) about 20-45% by weight of a dipeptidyl peptidase-4 inhibitor or a pharma- ceutically acceptable salt thereof; (ii) about 40-80% by weight of a diluent; (iii) about 0.5-6% by weight of a disintegrant; and (iv) about 0.75-10% by weight of a lubricant; and (b) a first layer comprising: (i) about 7-24% by weight of a denatonium salt; (ii) about 60-80% by weight of a diluent; (iii) about 2-12% by weight of a disintegrant; (iv) about 1-7% by weight of a binder, and (v) about 0.25-4% by weight of a lubricant. Includes.

[0215] The dipeptidyl peptidase-4 inhibitor is selected from the group consisting of alogliptin, carmegliptin, denagliptin, dutogliptin, linagliptin, melogliptin, saxagliptin, sitagliptin and vildagliptin, or their respective pharmaceutically acceptable salts. In another class, the dipeptidyl peptidase-4 inhibitor is selected from the group consisting of sitagliptin, vildagliptin and saxagliptin, or their respective pharmaceutically acceptable salts. In a subclass of this class, the dipeptidyl peptidase-4 inhibitor is sitagliptin or its dihydrogen phosphate salt.

[0216] Glycemic control / diabetes combination therapy Table 10 shows a comparison of denatonium acetate (DA) and DPP-4 inhibitors in a similar ob / ob mouse model. Data for DPP-4 inhibitors were taken from J Clin. Biochem. Nutr. 2015; 57(3):244-53. Acta Pharmacol. Sin. 2012; 33(8):1013-22. J Pharmacol. Exp. Ther. 2012; 342(1):71-80; and Eur. J. Pharmacol. 2008;588(2-3):325-32. [Table 10]

[0217] Similarly, Table 11 shows a comparison of denatonium acetate (DA) and DPP-4 inhibitors in a similar ob / ob mouse model. Data for DPP-4 inhibitors were taken from Am. J Physiol. Endocrinol. Metab. 2011; 300(2); E4l0-E42l.Biochim. Biophys. Acta Gen. Subj. 2018; 1862(3): 403-413. PLoS One. 2012; 7(6): e38744; and Aging Cell. 2019; 18(2): el2883. [Table 11]

[0218] [Example 5A] This embodiment provides an oral formulation combination bilayer tablet containing fixed doses of DPP4 inhibitor and bitter taste receptor agonist.Preferably, the bilayer components are sitagliptin and denatonium acetate (200mg per bilayer tablet), and sitagliptin (50mg per bilayer tablet) is designed to be either one bilayer tablet or two single layer tablets administered daily or BID (at least 6 hours apart).The in vivo data provided in Example 4 herein shows the synergistic effect of this combination, either combination tablet can be administered or separate tablets can be administered together.

[0219] Sitagliptin dihydrogen phosphate monohydrate is an orally active inhibitor of the DPP-4 enzyme chemically designed as 7-[(3R)-3-amino-1-oxo-4-(2,4,5-trifluorophenyl)butyl]-5,6,7,8-tetrahydro-3-(trifluoromethyl)-1,2,4-triazolo[4,3-a]pyrazine phosphate (1:1) monohydrate. Sitagliptin dihydrogen phosphate monohydrate is indicated as an adjunct to diet and exercise to improve glycemic control in adults with type 2 diabetes. However, sitagliptin does not achieve weight loss.

[0220] U.S. Patent No. 7,326,708 (hereby incorporated by reference) discloses, particularly in Example 7, a process for the manufacture of sitagliptin phosphate. Film-coated tablets Januvia® are sold in the United States by Merck. Januvia® tablets contain 32.13, 64.25 or 128.5 mg of sitagliptin phosphate monohydrate, corresponding to 25, 50 or 100 mg of the free base, respectively.

[0221] A preferred combination oral dosage form contains two drug compartments stacked on top of each other: The denatonium acetate compartment has a compressed particle formulation.

[0222] It provides an immediate gastric release oral pharmaceutical formulation and an immediate release 200 mg particulate formulation of denatonium acetate (DA) as a non-particulate water soluble DPP-4 inhibitor.

[0223] Table 12 shows the qualitative and quantitative formulation composition of DA in this example. [Table 12]

[0224] The detailed manufacturing process is described below.

[0225] 1.~3. Drug layer / seal coating / final blending The drug layering, seal coating and final blending process described above in Example 1B was carried out to produce the blended seal coated beads, denatonium IR pellets, used for compression into tablets.

[0226] 4. Tablet compression – Denatonium / DPP4i tablets 100mg Denatonium IR pellets, 100 mg, were compressed into a tablet layer and layered on top of the DPP-4 inhibitor tablet layer described below.

[0227] The second layer comprises a dipeptidyl peptidase-4 inhibitor or a pharma- ceutically acceptable salt thereof. The second bilayer further comprises one or more additives selected from the group consisting of: (i) a diluent; (ii) a disintegrant; and (iii) a lubricant. In another embodiment of the present invention, the second bilayer further comprises one or more surfactants or wetting agents; and one or more antioxidants.

[0228] The pharmaceutical bilayer composition is prepared by dry and wet processing methods. The DA layer is prepared by wet processing methods, preferably wet granulation methods. Either high shear granulation or fluid bed granulation may be used together with wet granulation. Alternatively, the DA layer is prepared by fluid bed granulation. Fluid bed granulation processing has the advantage of providing the tablet with higher diametral strength. The wet processing method enhances the chemical stability of DA. Alternatively, the DPP-4 layer is prepared by dry processing methods. In this class of embodiments, the DPP-4 inhibitor layer is prepared by direct compression. In addition, a bilayer tablet with a separate DA layer containing a disintegrant such as crospovidone is used to further increase the stability of the tablet.

[0229] The pharmaceutical compositions obtained by dry and wet processing methods can be compressed into tablets, encapsulated or metered into sachets.

[0230] The pharmaceutical composition contains one or more lubricants or glidants.Examples of lubricants include magnesium stearate, calcium stearate, stearic acid, sodium stearyl fumarate, hydrogenated castor oil, and mixtures thereof.In one embodiment, the lubricant is magnesium stearate or sodium stearyl fumarate, or mixtures thereof.Or, the lubricant is magnesium stearate or sodium stearyl fumarate.Examples of lubricants include colloidal silicon dioxide, tricalcium phosphate, magnesium silicate, and talc.

[0231] The pharmaceutical bilayer tablet composition may also contain one or more binders. Embodiments of binders include hydroxypropyl cellulose (HPC), hydroxypropyl methylcellulose (HPMC), hydroxyethyl cellulose, starch 1500, polyvinylpyrrolidone (povidone), copovidone, and polyvinylpyrrolidone.

[0232] The pharmaceutical bilayer tablet composition may also contain one or more diluents. Examples of diluents include mannitol, sorbitol, anhydrous calcium hydrogen phosphate, lactose monohydrate, calcium hydrogen phosphate dihydrate, microcrystalline cellulose, powdered cellulose, and combinations thereof. An example of a combination is mannitol, anhydrous calcium hydrogen phosphate, lactose monohydrate and microcrystalline cellulose, or a mixture of any two, three or four of them. Another example of a diluent combination is selected from anhydrous calcium hydrogen phosphate, lactose monohydrate and microcrystalline cellulose, or a mixture of any two or three of them. Microcrystalline cellulose is available from several suppliers, including Avicel, Avicel PH 101, Avicel PH 102, Avicel PH 103, Avicel PH 105 and Avicel PH 200 manufactured by FMC Corporation. Another example of a diluent is a mixture of microcrystalline cellulose and mannitol, the diluent being a 2:1 to 1:2 mixture of microcrystalline cellulose to mannitol.

[0233] The pharmaceutical bilayer tablet composition may also contain a disintegrant, which may be one of several modified starches, modified cellulose polymers or polycarboxylic acids, such as croscarmellose sodium, sodium starch glycolate, polacrilin potassium, carboxymethylcellulose calcium (CMC calcium) and crospovidone.

[0234] The pharmaceutical bilayer tablet composition may also contain one or more surfactants or wetting agents. The surfactants may be anionic, cationic or neutral. Anionic surfactants include sodium lauryl sulfate, sodium dodecane sulfonate, sodium oleyl sulfate and sodium laurate mixed with stearates and talc. Cationic surfactants include benzalkonium chloride and alkyltrimethylammonium bromide. Neutral surfactants include glyceryl monooleate, polyoxyethylene sorbitan fatty acid esters, polyvinyl alcohol and sorbitan esters. Wetting agent embodiments include poloxamers, polyoxyethylene alkyl ethers, polyoxyethylene castor oil derivatives and polyoxyethylene stearates.

[0235] The pharmaceutical bilayer tablet composition may also contain an antioxidant, which can be added to the formulation to provide chemical stability.The antioxidant is selected from the group consisting of α-tocopherol, γ-tocopherol, δ-tocopherol, tocopherol-rich natural extracts, L-ascorbic acid and its sodium or calcium salts, ascorbyl palmitate, progesterone gallate, octyl gallate, dodecyl gallate, butylhydroxytoluene (BHT) and butylhydroxyanisole (BHA).In one embodiment, the antioxidant is BHT or BHA.

[0236] The preferred dosage form of the pharmaceutical composition is a tablet produced by compression method. Such tablets may be film-coated with a mixture of hydroxypropanol cellulose and hydroxypropanol methylcellulose containing titanium dioxide and / or other coloring agents, such as iron oxide, pigments and lakes; a mixture of polyvinyl alcohol (PVA) and polyethylene glycol (PEG) containing titanium dioxide and / or other coloring agents, such as iron oxide, pigments and lakes; or any other suitable immediate release film coating agent. The coating provides taste masking and additional stability to the final tablet. A commercially available film coating agent is Opadry®, a formulated powder blend provided by Colorcon. Embodiments of Opadry® useful in the present invention include, but are not limited to, Opadry® I (HPC / HPMC), Opadry® 20A18334, Opadry® II, Opadry® II HP (PVA-PEG) or another suitable Opacity® suspension (such as polyvinyl alcohol, polyethylene glycol, titanium dioxide and talc, with or without colorants).

[0237] Finally, if desired, sweetening and / or flavoring agents may be added.

[0238] [Example 6] This example provides results from an in vivo mouse model of fatty liver disease treatment to investigate the therapeutic effect of DA on the treatment of NASH, compared to the positive control semaglutide (a GLP-1 agonist marketed drug for lowering HbA1c). The study used the positive control semaglutide, a vehicle control, the test drug DA, and a combination of semaglutide and DA. A mouse strain (B6 mice) was used. The animals were already adults (23 weeks old) when the study began, since they had already been fed an AMLN diet for 17 weeks before the study began. The test dose started at 75 mg / kg BID. However, after 2 weeks of administration, this DA dose was found to be poorly tolerated, and was therefore reduced to 50 mg / kg BID for the remaining 10 weeks of administration (total of 12 weeks).

[0239] The study included three groups of 10 mice each: (A) vehicle control with distilled water by gavage BID, (B) DA by gavage BID, and (C) semaglutide 10 mmol / kg sc QD. Body weight and changes were measured 3x per week. Serum metabolic markers (blood glucose, blood insulin, HbA1c, HDL, LDL, triglycerides and bile acids) were measured at the start of treatment (baseline) and at the end of the study. At the end of the study, histopathology of liver samples and serum levels of inflammatory biomarkers (IL-6, TNFα, CK-18 and TGF-β) were evaluated. Histopathology was performed blinded with a grading scale according to NAFLD activity score and fibrosis score according to Table 13. Table 14 identifies the kits and supplies used to measure serum parameters. [Table 13] [Table 14]

[0240] Figures 23-33B show data from this Example 6, further described above under the heading Brief Description of Figures. A summary and comparison of NASH in vivo data from multiple studies is shown in Table 15 below. [Table 15-1] [Table 15-2] [Table 15-3] [Table 15-4] [Table 15-5]

[0241] Table 15 shows widely different results in widely different NASH in vivo models, making direct comparison of the data difficult. The study corresponding to column 1 (called Aardvark Therapeutics) is provided in PCT patent application PCT / US2022 / 014550, filed January 31, 2022.

[0242] Studies emphasizing weight loss as a model for NASH treatment appear to be more directed at treating existing NASH conditions. Thus, in some embodiments, the dosage range of denatonium salts for methods of treating NASH and related liver disease is about 1.0 mg / kg / day to about 60 mg / kg / day; in some embodiments, about 2 mg / kg / day to about 40 mg / kg / day; and optionally, about 4 mg / kg / day to about 20 mg / kg / day. In another embodiment, the dosage range of denatonium salts for methods of preventing NASH and related liver disease and methods of slowing the progression of NASH and related liver disease is about 0.1 mg / kg / day to about 32 mg / kg / day, preferably about 0.25 mg / kg / day to about 16 mg / kg / day; most preferably about 0.5 mg / kg / day to about 8 mg / kg / day.

[0243] FDA Guidance for Clinical Trials to Treat NASH On January 29, 2021, the Food and Drug Administration (FDA) held a short seminar on NASH with fibrosis, how treatment drug candidates can show efficacy in animal models and clinical trials. The FDA confirmed that NASH (with fibrosis, hereafter NASH) is a serious condition and that the clinical use of surrogate endpoints can predict clinical benefit. Although in animal studies (such as those provided in Example 1 herein), histopathological examination is a better evidence of treatment, prevention and progression of disease benefit (depending on the length of the animal study). Therefore, in clinical trials, the FDA will accept surrogate endpoints and liver biopsies as a means of showing clinical benefit (or lack thereof). The FDA acknowledged that the challenge of NASH drug development stems from the gradual and slow progression of chronic inflammatory changes in the liver, and that any NASH drug to prevent or treat or slow progression of complete NASH (advanced liver fibrosis) is a potentially lifelong treatment. Regarding surrogate endpoints, the FDA suggests histopathology diagnosis as having "substantial predictive potential for clinical benefit." The FDA has shown that advanced NASH liver "fibrosis stage (but without other histologic features of steatohepatitis) is independently associated with increased mortality, transplantation, and liver-related events" (citing Angulo et al. Gastroenterology, 149:389-397, 2015).

[0244] In conducting clinical trials, the FDA suggests that initial trials will assess liver stiffness using non-invasive disease-specific biomarkers (e.g., aminotransferases), total bilirubin and radiological modalities (elastography, MRI-PDFF, etc.). To receive approval, the FDA will allow improvement in liver histology. "Liver biopsy will be a surrogate based on studies showing that improvement in histology may predict improved clinical outcomes in patients with NASH." Liver fibrosis is staged as stage 0 (none), stage, stage 2, stage 3 and stage 4 (cirrhosis). The recommended endpoints for NASH are (1) resolution of steatohepatitis and no worsening of liver fibrosis; or (2) improvement of liver fibrosis and no worsening of steatohepatitis; or (3) both resolution of steatohepatitis and improvement of fibrosis.

Claims

1. A pharmaceutical composition comprising a combination of a bitter taste receptor agonist and a GLP-1 receptor agonist or a DPP-4 inhibitor, wherein the bitter taste receptor agonist is a denatonium salt selected from denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate, and the pharmaceutical composition is for the treatment or prevention of diabetes, pre-diabetic syndrome, obesity, weight and / or appetite control, hyperlipidemia, fatty liver disease and / or hyperglycemia.

2. The pharmaceutical composition of claim 1, wherein the pharmaceutical composition is for the treatment or prevention of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) or alcoholic steatohepatitis (ASH).

3. The pharmaceutical composition of claim 1, wherein the GLP-1 receptor agonist or DPP-4 inhibitor is selected from salts of medium-chain fatty acids, salts of N-(8-(2-hydroxybenzoyl)amino)caprylic acid, sodium N-(8-(2-hydroxybenzoyl)amino)caprylate (SNAC), sitagliptin or a salt thereof, saxagliptin, linagliptin, alogliptin, semaglutide, glyburide, liraglutide, dulaglutide, albiglutide, vildagliptin, linagliptin, alogliptin, P93 / 01, SYR322, GSK 823093, Roche 0730699, TS021, E3024 or PHX-1149.

4. The pharmaceutical composition of claim 3, wherein the GLP-1 receptor agonist or DPP-4 inhibitor is sitagliptin or a salt thereof.

5. The pharmaceutical composition of claim 3, wherein the GLP-1 receptor agonist or DPP-4 inhibitor is sitagliptin, semaglutide or liraglutide.

6. The pharmaceutical composition of claim 1, wherein the bitter taste receptor agonist is administered as a racemic mixture or as an enantiomer or diastereoisomer.

7. The pharmaceutical composition of claim 1, wherein the bitter taste receptor agonist is denatonium acetate (DA).

8. 1. A pharmaceutical composition comprising a bitter taste receptor agonist, including a denatonium salt, in combination with a GLP-1 receptor agonist for the treatment or prevention of the progression of non-alcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH) or alcoholic steatohepatitis (ASH), wherein the denatonium salt is selected from the group consisting of denatonium acetate (DA), denatonium citrate, denatonium maleate, denatonium saccharide and denatonium tartrate, and wherein the denatonium salt is administered as a racemic mixture or as an enantiomer or diastereoisomer.

9. 9. The pharmaceutical composition of claim 8, wherein the dosage range of the denatonium salt for the treatment of NASH and related liver diseases in adult humans is from about 50 mg / day to about 3000 mg / day or from about 100 mg / day to about 2000 mg / day.

10. 9. The pharmaceutical composition of claim 8, wherein the dosage range of the denatonium salt for the treatment of NASH and related liver disease in adult humans is from about 0.5 mg / kg BID to about 30 mg / kg BID or from about 1 mg / kg BID to about 20 mg / kg BID.

11. 9. The pharmaceutical composition of claim 1 or 8, wherein the daily dose of the denatonium salt is administered once a day, twice a day or three times a day.

12. 2. The pharmaceutical composition of claim 1, wherein the fatty liver disease is selected from non-alcoholic steatohepatitis (NASH), alcoholic steatohepatitis (ASH), non-alcoholic fatty liver disease (NAFLD), HIV-associated steatohepatitis and liver fibrosis.

13. The pharmaceutical composition of claim 1, wherein the fatty liver disease does not include liver fibrosis.

14. A pharmaceutical composition according to claim 1 or 8, wherein the GLP-1 receptor agonist or DPP-4 inhibitor is a GLP-1 receptor agonist selected from semaglutide, liraglutide, dulaglutide, albiglutide, glucagon, exenatide or lixisenatide.

15. The pharmaceutical composition of claim 14, wherein the GLP-1 receptor agonist is semaglutide.

16. The pharmaceutical composition of claim 6, wherein the GLP-1 receptor agonist or DPP-4 inhibitor is sitagliptin.

17. The pharmaceutical composition of claim 14, wherein the GLP-1 receptor agonist is liraglutide.

18. 9. The pharmaceutical composition of claim 8, wherein the bitter taste receptor agonist is denatonium acetate (DA).

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