WO2022029029A1

WO2022029029A1 - Composition and method for modulating food intake

Info

Publication number: WO2022029029A1
Application number: PCT/EP2021/071394
Authority: WO
Inventors: Anna ARDÉVOL GRAU; Montserrat PINENT ARMENGOL; Carme GRAU BOVÉ; Mª Teresa BLAY OLIVÉ; Ximena Terra Barbadora; Raúl BELTRÁN DEBÓN; Esther RODRÍGUEZ GALLEGO; Carlos GONZÁLEZ QUÍLEN; Marta SIERRA CRUZ; Alba MIGUENS GÓMEZ
Original assignee: Universitat Rovira I Virgili
Priority date: 2020-08-07
Filing date: 2021-07-30
Publication date: 2022-02-10
Also published as: ES2894177A1

Abstract

The present invention relates to a composition based on mixed or specific agonists of the TAS2R5 receptors, mixed or specific agonists of the TAS2R39 receptors or in combinations thereof. The compositions of the present invention have the ability to modulate food intake either to decrease it or to increase it depending on whether there is greater activation or stimulation of TAS2R5 receptors or TAS2R39 receptors respectively. Likewise, the present invention relates to a method for modulating food intake based on the administration of the composition of the invention.

Description

COMPOSITION AND METHOD FOR MODULATING FOOD INTAKE

DESCRIPTION

FIELD OF THE INVENTION

The present invention belongs to the field of orexigenic/anorectic treatments for dietary and/or therapeutic purposes. More particularly, the present invention relates to a composition based on mixed or specific agonists of the TAS2R5 receptors, mixed or specific agonists of the TAS2R39 receptors or combinations thereof. The compositions of the present invention have the ability to modulate food intake either to decrease it or to increase it depending on whether there is greater activation or stimulation of TAS2R5 receptors or TAS2R39 receptors respectively. The activation of both receptors will modulate the intake in one sense or another depending on the potency in stimulating them and the profile of enterohormones that said activation produces in the gastrointestinal secretome. Likewise, the present invention relates to a method for modulating food intake based on the administration of the composition of the invention.

BACKGROUND OF THE INVENTION

Controlling food intake is a complex process that requires the integration of several signals from very different sources. The nervous and hormonal systems play a role in the case of animals, but in humans, feelings and sensations, among other environmental factors, are also involved [Blundell, J. E., et al. (2015). Physiology & Behavior. ; y Blundell, J.E et al. (2010). Obesidad Reviews : An Official Journal of the International Association for the Study of Obesity, 11(3), 251-270.]. To address the study and control of the regulation of food intake, different approaches have been adopted, including diet, physical activity, medical devices, pharmacotherapy and metabolic (bariatric) surgery.

One of the most effective treatments against obesity and its associated metabolic disorders is metabolic surgery [Mulla etal. (2018) Annals of the New York Academy of Sciences, 1411(1), 53-64], It produces an enormous change in metabolism and, among other effects, there is a modification in the gastrointestinal secretome of patients. Enterohormones reach various tissues in the body, including the brain and other peripheral tissues (fat, muscles, and the gastrointestinal tract itself) [Castagneto et al. (2018). Peptidos, 100(septiembre de 2017), 114-122], The most consolidated effect of bariatric surgery is the increase in GLP1 and PYY [Sweeney et al. 2014, Best Practice & Research Clinical Gastroenterology, 28(4), 727-740], The reproduction of this enterohormonal modulation without the use of surgery could be a mechanism to obtain some of the beneficial effects of surgery avoiding the use of this invasive method. Various nutritional approaches cause regulation of the enterohormonal profile [summarized in Serrano etal., 2017 Trends in Food Science & Technology, 68, 113-129] but their success in controlling food intake is not always effective, probably due to poor control on the composition of food components.

The bitter taste allows individuals to be protected against the unhealthy products of nature [Glendinning, 1994, Physiology & Behavior, 56(6), 1217-1227]. However, not all toxins are bitter and, vice versa, not all bitter compounds are toxic [Nissim et al, 2017, IUBMB Life, 69(12), 938-946], Many bitter compounds have health benefits, and healthier diets have been suggested to have a higher component of bitter-tasting ingredients, including bitter vegetables [Duffy et al., 2010, Chemosensory Perception, 3(3), 137-148], Bitter taste receptors (TAS2Rs) have recently been identified in places other than the mouth, where taste perception occurs, but a clear function for them has not yet been defined. [Foster et al , 2014, Pharmacology and Therapeutics, 142(1), 41-61], Meyerhof and collaborators did an arduous work of association of bitter ligands with the particular TAS2R through in vitro assays, through the visualization of changes in cytoplasmic calcium levels [Meyerhof et al., 2010, Chemical Senses , 35(2), 157-170], They, together with Di Pizio Bioorganic & Medicinal Chemistry, 23(14), 4082-4091] showed that in humans and mice, [Lossow et al., 2016, Journal of Biological Chemistry, 291(29), 15358-15377] where there is a relatively high number of bitter taste receptors (25 for humans and 35 for mice), the different types of receptors are differentiated in terms of their selectivity. There are selective receptors that only bind 1-3 ligands, such as hTAS2R5, less selective receptors, such as hTAS2R39, and highly promiscuous receptors that bind several ligands. A promiscuous TAS2R is one that can be activated by various ligands. In turn, the ligands can be specific or nonspecific for a given receptor [Pizio etal. 2015, Bioorganic & Medicinal Chemistry, 23(14), 4082- 4091], Species with a more limited number of bitter taste receptors contain only promiscuous receptors. The role of these bitter taste receptors away from the tongue papillae, where the taste is perceived, is being studied. They have been found in various places, for example in the lungs, [Grassin-Delyle et al., 2019, Frontiers in Physiology, 10(October), 1-13], Wu et al. showed an improvement in polycystic ovary syndrome in murine models [l/Vu et al. 2019, Endocrinologia, 160(1), 143- 155], The present inventors and others have previously shown that its stimulation induces ghrelin secretion in the murine grelinoma cell line [Serrano et al., 2016; Molecular Nutrition & Food Research, 60(12), 2554-2564] and in human fundic cells [Wang et al., 2019a, FASEB Journal : Publication oficial de la Federation de Sociedades Americanas de Biologla Experimental, 33(4), 4907-4920],

On the other hand, some have suggested that some bitter compounds with affinity for TAS2R receptors may have some utility in controlling appetite. LIS20190374489 describes a composition between a bitter compound, specifically a denatonium salt and a sweet antagonist useful in controlling appetite.

The present invention is based on the inventors' discovery that stimulation of TAS2R5 receptors at the intestinal level produces an enterohormone secretion profile that limits food intake (anorectic effect) and, on the contrary, stimulation of TAS2R39 receptors produce a profile that tends to increase food intake (orexigenic effect). In this sense, the present invention is directed to compositions based on both TAS2R5 and TAS2R39 agonists (mixed or specific). The relative presence and concentration of these types of agonists in the compositions of the invention, the relative affinity for each of the receptors or the specificity for one of them will determine the nature, type and potency of TAS2R5 or TAS2R39 stimulation and will determine the enterohormone profile of the gastrointestinal secretome. In this way, the variation of these parameters makes it possible to design compositions that inhibit appetite and favor the reduction of food intake or, on the contrary, compositions that stimulate appetite and favor an increase in said intake. The compositions of the present invention have, therefore, application both in dietary treatments for weight gain or reduction, as well as in therapeutic treatments aimed at alleviating disorders related to food intake such as those related to obesity, but also others like bulimia or anorexia nervosa.

BRIEF DESCRIPTION OF THE FIGURES

Figure 1 : Stimulation of 1,10-phenanthroline-induced enterohormone secretions. Rat duodenum (for CCK) and ileum (for tGLPI and PYY) segments were treated with 150 pM 1 ,10-Phenanthroline for 30 minutes. The medium was then collected and the respective enterohormones were quantified by ELISA. The results are calculated against the respective basal secretion in each hormone. * denotes p <0.05 vs control.

Figure 2: Stimulation of thiamine-induced enterohormone secretions. Rat duodenum (for CCK) and ileum (for tGLP1 and PYY) segments were treated with 1 mM thiamine for 30 minutes. The medium was then collected and the respective enterohormones were quantified by ELISA. The results are calculated against the respective basal secretion in each hormone. * denotes p <0.05 vs control, # denotes p <0.1 vs control.

Figure 3: A. Total GLP1 secretion in rat ileum induced by simultaneous stimulation of the agonist hTA2R5 and hTAS2R39. The rat ileum segments were treated with 1 mM Epicatechin or 20 pM B2galate for 30 minutes. The medium was then collected and the total GLP1 in the medium was quantified by ELISA. * indicates p <0.05 vs control, # indicates p <0.1 vs control. B. CCK secretion in the rat duodenum induced by the agonist hTA2R5 and hTAS2R39. Rat duodenum segments were treated with 1mM Epicatechin or 20 pM B2galate for 30 minutes. The medium was then collected and the CCK in the medium was quantified by ELISA. * indicates p <0.05 vs control, # indicates p <0.1 vs control. C. Stimulation of enterohormonal secretions induced by Procyanidin B2. Rat ileum (for tGLPI , red columns) and duodenum (for CCK, blue columns) segments were treated with 67 or 300 pM B2 for 30 minutes, or 300 pM B2 + 1 mM epicatechin for 45 minutes. The medium was then collected and the respective enterohormones were quantified by ELISA. The results are calculated against the respective basal secretion in each hormone. * denotes p <0.05 vs control. Figure 4: Reduction of food intake (Fl) induced by an acute dose of 1,10- Phenanthroline in female rats. Animals were treated one hour before the dark period with an acute dose of 1 ,10-Phenanthroline 200 mg/kg ~ 290 pM (black columns) or water as a vehicle (white columns). Food intake was measured at the indicated times from the beginning of the dark period. * denotes p <0.05 vs. control; # denotes p <0.1 vs. control.

Figure 5: Reduction of food intake (Fl) induced by an acute dose of Epicatechin in female rats. The animals were treated one hour before the dark period with an acute dose of Epicatechin. The gray columns indicate the treatment with 244 mg/kg ~ 0.84 mM of Epicatechin, the black columns for a dose of 300 mg/kg ~ 1 mM of Epicatechin and the white columns for water as vehicle. Food intake was measured at the indicated times from the beginning of the dark period. * denotes p <0.05 vs control; # denotes p <0.1 vs control.

Figure 6: Food intake (Fl) after an acute dose of Epicatechin + Procyanidin B2 + Epicatechin Gallate in female rats. The animals were treated one hour before the dark period with an acute dose of Epicatechin + B2 + Epicatechin gallate (200 + 62 + 18 mg/kg) (black columns). The white columns indicate the control group treated with water as vehicle. Food intake was measured at the indicated times from the beginning of the dark period. # denotes p <0.1 vs control; * denotes p <0.05 vs control.

Figure 7: Increase in food intake (Fl) induced by an acute dose of Epicatechin + Epicatechin Gallate in female rats. The animals were treated one hour before the dark period with an acute dose of Epicatechin (234 mg/kg) + epicatechin gallate (14 mg/kg), a total dose ~ 0.84 mM (black columns). The white columns indicate the control group treated with water as the vehicle. Food intake was measured at the indicated times from the beginning of the dark period. * denotes p <0.05 vs control.

Figure 8: Increase in food intake (Fl) induced by an acute dose of Epicatechin + B2 in female rats. The animals were treated one hour before the dark period with an acute dose of Epicatechin (213 mg/kg) + B2 (62 mg/kg), a total dose of ~ 0.84 mM. The white columns indicate the control group treated with water as vehicle. Food intake was measured at the indicated times from the beginning of the dark period. # denotes p <0.1 vs control.

DETAILED DESCRIPTION OF THE INVENTION

In order to facilitate understanding and clarify the meaning of certain terms in the context of the present invention, the following definitions are provided:

"Composition": refers to a set of components or substances that are formulated in a homogeneous or heterogeneous manner. The compositions in the context of the invention incorporate at least one mixed or specific agonist of the TAS2R5 receptor and/or one mixed or specific agonist of the TAS2R39 receptor. Compositions in the context of the present invention can be food compositions, food supplements or pharmaceutical compositions.

"Intake modulation": refers to the effect produced by the composition of the invention on food intake in the subject to whom it is administered. Depending on the components that make up the composition, the modulation of intake can be negative, reducing food intake (anorexigenic effect) or positive, increasing food intake (orexigenic effect). The positive or negative sense of modulation will depend on the presence in the composition of TAS2R5 agonists that produce a negative modulation or reduction of intake and of TAS2R39 agonists that produce a positive modulation or increase of intake. In the case of compositions that comprise both TAS2R5 and TAS2R39 agonists or those that comprise mixed agonists of both receptors, the positive or negative sense and the modulation potency will depend on the amount/concentration and the relative affinity (EC50) for each receptor of each TA2R5 and TAS2R39 agonist in the composition.

“Specific agonist”: refers to any compound or substance that is capable of binding to a receptor in a specific way by stimulating its activation. In the context of the present invention, specific agonists can be either of the TAS2R5 receptor or of the TAS2R39 receptor. Some specific agonists of the TAS2R5 receptor are, but not limited to: 1 ,10- phenanthroline, Procyanidin C2, Procyanidin B4 and Procyanindin B7. Some specific agonists of the TAS2R39 receptor are, although not limited thereto: thiamine, epicatechin gallate and acetaminophen.

“Mixed agonist”: It refers to any compound or substance that is capable of binding to one or more receptors, stimulating their activation. Typically, mixed agonists have different affinity for those receptors to which they can bind and will therefore bind more readily to those for which they have a lower EC50. In the context of the present invention, mixed agonists are those that have affinity for both the TAS2R5 receptor and the TAS2R39 receptor. Some TAS2R5/TAS2R39 mixed agonists in the context of the invention are: epicatechin, B2-gallate, epigallocatechin gallate (EGCG), Pentagaloylglucose (PGG), Denatonium Saccharide and Sucralose.

“TAS2R5 receptor”: refers to isoform number 5 of the family of G proteins that act as receptors located in the membrane of some cell populations, whose stimulation induces the sensation of bitter taste, encoded by the TAS2R5 gene.

“TAS2R39 receptor”: Isoform number 39 of the family of G proteins that act as receptors located in the membrane of some cell populations, whose stimulation induces the sensation of bitter taste, encoded by the TAS2R39 gene.

“Food composition”: in the context of the invention it refers to a food or drink processed with nutritional properties and comprising in its formulation at least one mixed or specific agonist of the TAS2R5 receptor and/or at least one mixed or specific agonist of the TAS2R39 receptor. Food compositions in the context of the invention can be, but are not limited to, dairy products, bakery products, natural juices or extracts of vegetables, cereals or similar products.

“Food supplement”: in the context of the invention it refers to a composition that serves to supplement certain nutritional elements in the diet of an individual and that comprises in its formulation at least one mixed or specific agonist of the TAS2R5 receptor and/or at least one mixed or specific agonist of the TAS2R39 receptor. Food supplements in the context of the invention may be, but are not limited to, vitamin supplements, protein supplements, vegetable concentrates or similar products. “Pharmaceutical composition”: in the context of the invention it refers to a formulation that serves for the preparation of a drug in any pharmaceutical form for preferably oral administration and that comprises in its formulation at least one mixed or specific agonist of the TAS2R5 receptor and/or at least one mixed or specific agonist of the TAS2R39 receptor, in addition to pharmaceutical excipients. The pharmaceutical compositions can preferably be formulated in the form of tablets, sugar-coated tablets, capsules, pills, chewing gums, powders, drops, gels, juices, syrups, solutions and suspensions.

The composition

The first aspect of the invention refers to a composition for the modulation of food intake comprising or consisting of: a) a combination of at least one mixed or specific agonist of the TAS2R5 receptor and at least one mixed or specific agonist of the TAS2R39 receptor; b) at least one Specific TAS2R5 receptor agonist; c) at least one Specific TAS2R39 receptor agonist; or d) at least one mixed agonist of the TAS2R5 and TAS2R39 receptors, with a relative affinity (EC50) differentiated between both receptors.

Throughout the document, a composition with these characteristics will be referred to as the composition of the invention or the composition according to the invention.

In the context of the present invention, the mixed agonist(s) of the TAS2R5 and TAS2R39 receptors are preferably selected from epicatechin, B2-gallate, epigallocatechin gallate (EGCG), Procyanidin B2, Pentagaloylglucose (PGG), Denatonium saccharide, Sucralose and a mixture thereof. Some of these agonists bind with a lower EC50 to TAS2R5 receptors such as epicatechin or B2-gallate, but others, such as EGCG have a lower EC50 to TAS2R39 receptors and, therefore, bind with greater affinity for this receptor.

On the other hand, the Specific TAS2R5 receptor agonist(s) are preferably selected from 1 ,10-phenanthroline, Procyanidin C2, Procyanidin B4 and Procyanindin B7 and a mixture thereof. For its part, the specific TAS2R39 receptor agonist(s) are preferably selected from thiamine, epicatechin gallate, acetaminophen and a mixture thereof.

Among the possible embodiments of the present invention, there is a specific one (option a) that refers to compositions that can have a positive (increase) or negative (decrease) modulating effect on intake depending on the agonists used, the concentration of them and the relative affinity for each of the TAS2R5 and TAS2R39 receptors or the specificity for one of them. Thus, in this specific embodiment, the composition of the invention comprises or consists of a combination of at least one mixed or specific agonist of the TAS2R5 receptor and at least one mixed or specific agonist of the TAS2R39 receptor.

Within this embodiment, and more particularly, the composition comprises at least one mixed TAS2R5 and TAS2R39 agonist selected from epicatechin, B2-gallate, epigallocatechin gallate (EGCG), Procyanidin B2, Pentagaloylglucose (PGG), Denatonium saccharide, Sucralose, preferably epicatechin, B2-gallate, epigallocatechin gallate (EGCG), procyanidin B2; and at least one specific TAS2R5 agonist selected from 1 ,10-phenanthroline, Procyanidin C2, Procyanidin B4 and Procyanidin B7, preferably 1 ,10-phenanthroline.

Still within this embodiment, and also more particularly, the composition comprises at least one mixed TAS2R5 and TAS2R39 agonist selected from epicatechin, B2- gallate, epigallocatechin gallate (EGCG), Procyanidin B2, Pentagaloylglucose (PGG), Denatonium saccharide, Sucralose, preferably epicatechin, B2-gallate, epigallocatechin gallate (EGCG), Procyanidin B2; and at least one specific TAS2R39 agonist selected from thiamine, epicatechin gallate and acetaminophen, preferably thiamine or epicatechin gallate. For example, as demonstrated in the examples (see figure 6) a composition containing epicatechin, procyanidin B2 and epicatechin gallate where the TAS2R5 stimulation pathway prevails produces a reduction in food intake. However, a composition with epicatechin and epicatechin gallate makes the TAS2R39 receptor stimulation pathway prevail, which increases food intake (see figure 7). Still within this embodiment, and also more particularly, the composition comprises at least one specific TAS2R5 agonist selected from 1 ,10-phenanthroline, Procyanidin C2, Procyanidin B4 and Procyanidin B7, preferably 1 ,10-phenanthroline and at least one specific TAS2R39 agonist selected from thiamine, epicatechin gallate and acetaminophen, preferably thiamine. A composition according to this embodiment comprises or consists of a combination of 1 ,10-phenanthroline and thiamine.

Another specific embodiment of the invention (option b) contemplates those compositions that only have a negative modulating effect, that is, that favor the reduction of food intake. In this specific embodiment of the invention, the composition comprises or consists of at least one Specific TAS2R5 receptor agonist.

Within this embodiment, the specific TAS2R5 agonist(s) are selected from 1 ,10- phenanthroline, Procyanidin C2, Procyanidin B4 and Procyanidin B7 or a combination thereof. The preferred option of this embodiment contemplates a composition comprising or consisting of 1 ,10-phenanthroline. As shown in Figure 1 , a composition based solely on the specific TAS2R5 agonist 1 ,10-phenanthroline produces a significant increase in the enterohormones GLP1 in the ileum and CCK in the duodenum. Furthermore, the composition based only on 1 ,10-phenanthroline produces a significant reduction in food intake in vivo in rats (see figure 4).

Another additional specific embodiment of the invention (option c) refers to those compositions that only have a positive modulating effect, that is, that favor increased food intake. In this specific embodiment of the invention, the composition comprises or consists of at least one Specific TAS2R39 receptor agonist.

Within this embodiment, the specific TAS2R39 agonist(s) are selected from thiamine, epicatechin gallate, acetaminophen, and a mixture thereof. The preferred option of this embodiment contemplates a composition comprising or consisting of thiamine or epicatechin gallate, more preferably a composition comprising or consisting of thiamine. As seen in Figure 2, a composition based on thiamine produces an enterohormone release profile where GLP1 is not affected, but CCK is significantly decreased. For its part, PYY shows an upward trend due to the TAS2R39-specific stimulation of thiamine. A last specific embodiment of the invention (option d), refers to those compositions based on mixed agonists of the TAS2R5 and TAS2R39 receptors that can have a positive (increased intake) or negative (decreased intake) modulating effect depending on of the relative affinity (EC₅o) of the agonist for one or another receptor. In a case like this the agonist will preferentially bind and stimulate the receptor for which it has a higher affinity and, therefore, a lower EC50. If the receptor for which the agonist(s) show(s) a higher affinity is a TAS2R5 receptor, the composition will show a positive modulatory effect of ingestion, whereas if the agonist's higher relative affinity is for the TAS2R39 receptor, the composition will have the effect of reducing the food intake. Therefore, in this last specific embodiment of the invention, the composition comprises or consists of at least one mixed agonist of the TAS2R5 and TAS2R39 receptors, with a relative affinity (EC50) differentiated between both receptors.

Within this embodiment, the mixed agonist(s) of the TAS2R5 and TAS2R39 receptors are selected from epicatechin, B2-gallate, epigallocatechin gallate (EGCG), Procyanidin B2, Pentagaloylglucose (PGG), Denatonium saccharide and Sucralose.

A preferred option within this specific embodiment of the invention is a composition comprising or consisting of epicatechin which is a mixed TAS2R5/TAS2R39 agonist although it has a lower EC50 for the TAS2R5 receptor. In this sense, as shown in Figures 3A and 3B, it has an enterohormone release profile that is more similar to that of a TAS2R5 agonist but is not the same, since although CCK levels in the duodenum are significantly increased, levels of GLP1 are not altered. This does not prevent that, as observed in figure 5, a composition based on epicatechin has a significant effect in reducing food intake in rats, although it should be noted that this effect is dosedependent so that at a low dose the effect lasts only three hours, while at a higher dose the effect of reducing the intake is maintained over time.

Another preferred option within this specific embodiment is a composition comprising or consisting of procyanidin B2 and epicatechin, both mixed TAS2R5/TAS2R39 agonists. As seen in Figure 3C, procyanidin B2 alone appears to have an enterohormone stimulation profile similar to that of a specific TAS2R5 agonist, producing a significant increase in GLP1 and CCK release. However, when administered together with epicatechin this effect appears to be neutralized. These results suggest that the presence of procyanidin competes with epicatechin (which in principle has a greater affinity for TAS2R5 than for TAS2R39) to bind to the TAS2R5 receptor, which shifts the balance and causes epicatechin to bind in a greater proportion to TAS2R39 and makes a response more reminiscent of TAS2R39 stimulation to prevail. In fact, as shown in figure 8, this composition (epicatechin + procyanidin 2) produces a significant increase in food intake, although of lower potency than that of epicatechin with a selective TAS2R39 agonist such as epicatechin gallate (see figure 7).

Ultimately, these examples demonstrate that the potency of the orexigenic or anorectic effect of the compositions of the invention can be modulated depending on whether the signaling pathway mediated by the TAS2R5 receptor or the pathway mediated by the TAS2R39 receptor is activated more or less. The prevalence of one or the other pathway and the potency with which it is activated allows the compositions of the invention to be adapted according to the needs, in such a way that the orexigenic or anorexigenic treatment can be modulated in a targeted manner.

In relation to the concentration of the different agonists in the compositions of the invention, this must be at a level that is equal to or greater than the effective concentration, that is, the minimum concentration that produces a response. Said effective concentration for the different mentioned compounds is summarized in the following table 1 :

Table 1

Preferably, the different agonists are found in the compositions of the invention within the concentration ranges and are administered in the doses specified in the following table 2:

Table 2

The composition of the present invention can be formulated as a food composition, a food supplement or a pharmaceutical composition.

A food composition generally refers to a processed food or drink that incorporates in its composition at least one mixed or specific agonist of the TAS2R5 receptor and/or at least one mixed or specific agonist of the TAS2R39 receptor according to the invention described above.

In the context of the invention the food composition can be, but is not limited to, a dairy product, a bakery product, natural juices or extracts of vegetables, cereals or similar products. The composition of the invention can also be formulated as a food supplement, that is, a composition that serves to supplement certain nutritional elements in the diet of an individual and that comprises in its formulation at least one mixed or specific agonist of the TAS2R5 receptor and/or at least a mixed or specific agonist of the TAS2R39 receptor according to the invention described above.

In the context of the invention the food composition may be, but is not limited to, vitamin supplements, protein supplements, plant extracts or concentrates or similar products.

Both food compositions and food supplements described in the context of the invention are aimed at dietary and/or aesthetic treatment, but do not have a therapeutic purpose.

Finally, the composition of the invention can also be formulated as a pharmaceutical composition or medicine. Preferably, the pharmaceutical composition will be formulated for oral administration and comprises in its formulation at least one mixed or specific agonist of the TAS2R5 receptor and/or at least one mixed or specific agonist of the TAS2R39 receptor according to the invention described above. Furthermore, in its formulation the pharmaceutical composition includes one or more pharmaceutical excipients commonly used in the galenic formulation.

The preferred administration forms of the pharmaceutical compositions according to the invention are tablets, sugar-coated tablets, capsules, pills, chewing gums, powders, drops, gels, juices, syrups, solutions and suspensions.

The pharmaceutical compositions according to the invention are directed to medical treatments and therefore have a therapeutic purpose.

Use of the composition

A second aspect of the invention relates to the use of the composition of the invention. In general, the composition of the present invention is used for the modulation of food intake in animals, preferably mammals, or in humans.

Modulation of intake can be understood in a negative sense. This occurs when the composition of the invention is used to reduce appetite and food intake. In this case, the modulation of intake would be mainly directed towards an anorectic treatment for weight reduction, either for dietary and/or aesthetic purposes in people who want to lose kilos, particularly in the form of body fat, or for therapeutic purposes for the treatment of obesity and obesity-associated disorders such as diabetes, hyperlipidemia or metabolic syndrome.

On the other hand, modulation of intake can be understood in a positive sense, that is, when the composition is used to increase appetite and food intake. In this case, the modulation of the intake would be mainly directed towards an orexigenic treatment for weight gain either for dietary and/or aesthetic purposes in people who want to gain weight in the form of muscle mass or body fat, or for therapeutic purposes for the treatment of diseases such as anorexia nervosa or anorexia derived from any pathological process or pathologies such as bulimia nervosa.

As has already been explained, the fact that the composition is used in one sense or another depends on the specific composition used, on the concentration of the components and on the extent to which they further favor a given enterohormonal profile because they favor a greater activation of the signaling pathway mediated by the TAS2R5 or TAS2R39 receptor.

Intake modulation method

A third aspect of the invention relates to a method for modulating food intake in a subject which comprises administering the composition of the invention to the subject.

In a particular embodiment of the method of the invention, modulation involves the decrease in food intake that involves administering a composition that comprises: a) at least one Specific TAS2R5 receptor agonist; b) a mixed TAS2R5 and TAS2R39 receptor agonist having a lower EC50 for the TAS2R5 receptor than for the TAS2R39 receptor; or c) a combination of at least one Specific TAS2R5 receptor agonist and at least one Specific TAS2R39 receptor agonist, where the stimulating effect of the TAS2R5 receptor agonist prevails.

In another particular embodiment of the method of the invention, modulation involves increasing food intake that involves administering a composition that comprises: a) at least one Specific TAS2R39 receptor agonist b) a mixed TAS2R5 and TAS2R39 receptor agonist having a lower EC50 for the TAS2R39 receptor than for the TAS2R5 receptor; or c) a combination of at least one Specific TAS2R5 receptor agonist and at least one Specific TAS2R39 receptor agonist, wherein the stimulating effect of the TAS2R39 receptor agonist prevails.

For each of these embodiments, both specific and mixed agonists should be administered at a concentration range and according to a dosage range such as those in Table 2 above, without being limiting.

The following examples are intended to illustrate the invention, but are not intended to be limiting thereof.

EXAMPLES

Materials and methods

Chemicals and reagents

1 ,10-Phenanthroline, (-)-Epicatechin, Thiamine, were purchased from Sigma (Barcelona, Spain). Procyanidin gallate B2, epigallocatechin gallate (EGCG) and epicatechin gallate were purchased from Extrasynthese (Genay, France) and procyanidin B2 was purchased from Adooq-Bioscience (Irvine, CA, USA). Krebs-Ringer (KRB)'s bicarbonate buffer solution was used (11.5 mM Hepes, 2.6 mM CaCI₂, 1.2 mM MgCI₂, 5.5 mM KCI, 138 mM NaCI, 4.2 mM NaHCO₃, 1.2 mM NaH₂PC>4) pH 7.4, supplemented with 10 mM D-Glucose (KRB-D-Glucose buffer) or 10 mM D-Mannitol (KRB-D-Mannitol buffer). For the enterohormone secretion studies, KRB-D-Glucose was supplemented with protease inhibitors: 10 pM of amastatin (Enzo Life Sciences, Madrid, Spain), 100 Kill of aprotinin (Sigma, Barcelona, Spain) and 0, 1 % fatty acid-free bovine serum albumin.

Animals

26 male Fischer-344 rats (Charles River Laboratories, Barcelona, Spain), and 20 female Wistar rats (Envigo, Barcelona, Spain) were used. Most of the animals were housed in the Rovira i Virgili University facility. Ten female Wistar rats were raised and housed in the Faculty of Biology of the University of Barcelona. In all cases, the rats were housed under standard conditions: caged in pairs at an ambient temperature of 23 °C with a standard 12-hour light and dark cycle (lights on at 7 in the morning), ventilation and ad libitum access to tap water and a standard feed diet (Fischer-344 was fed a standard feed diet by SAFE -Cat No: Rats A04, SAFE, Augy, France and Wistar were fed the standard Teklad diet -Cat No : Teklad 2014, Envigo, Barcelona, Spain-). All procedures were approved by the respective ethics committees of the Rovira I Virgili University and the University of Barcelona.

Ex vivo treatments of intestinal segments

26 male Fischer-344 rats weighing 350-400g were used. After a short period of fasting (1-3 hours), they were euthanized by decapitation and their guts were removed. Samples were collected from the proximal duodenum and distal ileum. The tissue was rinsed with KRB-D-Mannitol ice buffer and dissected into segments (0.5 cm diameter). After a 15 minute wash period, the tissue segments were placed in a prewarmed (37 ° C) KRB-D-Glucose 0.1 % DMSO buffer containing the compounds to be tested. The duodenal and ileal segments were treated with different compounds or mixture of compounds (Table 3) in a humidified incubator at 37 ° C, 95% O₂ andy 5% CO₂. After 30 minutes of treatment, the entire volume was frozen and stored at -80 ° C for the quantification of enterohormones.

Table 3 Compound Concentration

Tested on intestinal segments

1 ,10-Phenanthroline 150pM

Thiamine 1 mM

1 ,10-Phenanthroline + Thiamine 150 M+1 mM

Epicatechin 1 mM

Procyanidin B2 (B2) 67 o 300 pM

Epicatechin + B2 1 mM + 300 pM

Procyanidin gallate B2 20 pM

Food intake studies

Ten female Wistar rats were housed in pairs for a week of adaptation. After this adaptation period, the animals were housed in individual cages and a daily food deprivation was introduced for 4 h before the light was turned off (from 3:00 p.m. to 7:00 p.m.) in order to habituate the subjects to the experimental schedule and they were trained for intragastric oral intubation with water 1 h before the onset of darkness (6:00 p.m.). One experiment per week was performed in a crossover design for all food intake studies. In each experiment, the trained animals were treated with different compounds or mixtures of compounds at defined concentrations (see table 4) intragastrically by oral intubation 1 h before the onset of darkness (18:00 h), using water as a vehicle. Parallel controls were performed by administering the vehicle intragastrically. The feed diet was administered at the beginning of the dark (7:00 p.m.) and the feed intake was measured 3, 12 and 20 hours later.

Table 4 Compound Concentration

Food consumption was checked

Epicatechin 244 mg/kg o 300 mg/kg

Procyanidin B2 62 mg/kg

Epicatechin + B2 213+62 mg/kg

Epicatechin + B2 + Epicatechin

200+62+18 mg/kg

Gallate Determination of the effects of an acute dose on the secretion of enterohormones in the portal vein in intraqastric treatments

Intragastric treatments were performed in two different groups of animals. The first set consisted of 10 female Wistar rats that received a specific intragastric dose of 1 ,10-phenanthroline. The second set consisted of the 10 female Wistar rats that received an intragastric dose of (-)-Epicatechin. The same procedure was applied to both sets of rats, which were indicated differently. The animals were randomly divided into two groups: a control group and a treated group. The rats were fasted from 10:00 p.m. to 7:00 a.m. before treatment and were anesthetized 5 minutes later with inhaled isoflurane (5% for induction, followed by 3% for maintenance), in the case of the 1.10 phenanthroline test, or pentobarbital (70mg/kg) in the case of the (-) - Epicatechin test. An incision was made in the abdominal cavity through the midline and the portal vein was catheterized with a PE tube (ID 0.28 mm, OD 0.61 mm; Becton Dickinson, Sparks, MD, USA) using a standard procedure. The catheter was fixed with cyanoacrylate and the abdominal cavity was closed with surgical forceps. Body temperature was kept constant at 37 ° C by means of a heated surgical table and ceiling lamps. At time zero, 200 pl of blood was obtained, and then filled with saline. Then the specific treatment or the tap water as a vehicle was pierced in the anterior stomach. After treatment, two portal blood samples (200 pl) were taken (detailed in the results); and each time, the catheter was filled with 0.9% heparinized NaCI. The blood was transferred to heparinized tubes and a 1 : 100 volume of a 1 : 1 mixture of the commercial inhibitor of Dipeptidyl peptidase-4 (DPPIV, Millipore, Madrid, Spain) and a serine protease inhibitor (complete™ ULTRA Tablets, Roche, Barcelona, Spain). Plasma was collected by centrifugation at 1500g for 15 min at 4 °C and immediately frozen at -80 °C for the quantification of enterohormones. Rats were sacrificed by bilateral thoracotomy.

Quantification of enterohormones

Enterohormone secretions from intestinal segments and plasma were measured with commercial kits. Total and active GLP-1 were measured with ELISA kits from Millipore (Cat #: EZGLPT1-36k and EGLP-35K, respectively, Burlington, MA, USA). PYY was measured with a fluorescent immunoassay kit (Catalog No. FEK-059-03, Phoenix Pharmaceuticals, Burlingame, CA, USA). Total CCK was measured with an ELISA kit (Catalog No. EKE-069-04, Phoenix Pharmaceuticals, Burlingame, CA, USA). Statistic analysis

Results are presented as mean ± SEM. The data were analyzed with the statistical software XLSTAT 2020.1 (Addinsoft, Spain). Statistical differences were evaluated by Student's t tests and P <0.05 was considered statistically significant.

Example 1 : Stimulation of the secretion of enterohormones of the TAS2R5 specific agonist 1,10-phenanthroline

The results on the secretion of enterohormones with 150 pM of 1 ,10-phenanthroline one of the few TAS2R5 specific agonists are shown in Figure 1.

In this, it is observed that the stimulation of the TAS2R5 receptor by 1 ,10- phenanthroline produces a significant increase in the secretion of GLP1 and CCK while the PYY does not vary significantly with respect to the control.

This enterohormonal secretion profile can be considered the type profile of stimulation by the TAS2R5 signaling pathway since, as previously mentioned, 1 ,10- phenanthroline is a pure and specific agonist of this receptor.

Example 2: Stimulation of the secretion of enterohormones of the TAS2R39- specific agonist Thiamine

The results on the secretion of enterohormones with 1 mM thiamine a TAS2R39 specific agonist are shown in Figure 2.

In this study, it is observed that stimulation by thiamine of the TAS2R39 receptor produces a significant increase in PYY secretion and a significant reduction in CCK. The GLP1 related to the feeling of satiety does not vary significantly with respect to the control.

This enterohormonal secretion profile can be understood as the type of stimulation profile by the TAS2R39 signaling pathway since, as mentioned above, thiamine is a pure and specific agonist of this receptor.

Example 3: Stimulation of enterohormone secretion of mixed TAS2R5/TAS2R39 agonists Epicatechin, Gallate B2 and Procyanidin B2

Epicatechin, B2 gallate and procyanidin B2 are all ligands that show a mixed binding profile to both TAS2R5 and TAS2R39 receptors. Each one shows a different affinity for each receptor, so it is expected that each receptor will show its own stimulation profile for these receptors based on their relative affinity for each receptor. Figures 3A and 3B show the secretion profile of GLP1 and CCK respectively induced by epicatechin and B2 gallate. As can be seen, epicatechin does not significantly affect GLP1 secretion, but it does affect CCK secretion, which is significantly increased. For its part, B2 gallate seems to have a significant impact on the increase in the secretion of both enterohormones compared to the control. If the secretion profile of B2 gallate is compared with the secretion profile of epicatechin, although the latter does not produce significant changes in GLP1 secretion, the stimulation of CCK secretion is much more powerful with epicatechin than with B2-gallate. In any case, both epicatechin and B2 gallate show a stimulation profile more similar to that of 1 ,10- phenanthroline (TAS2R5 agonist) than to thiamine (TAS2R39 agonist) not surprisingly, both compounds have a higher affinity for TAS2R5 receptors than for TAS2R39 receptors.

Figure 3C shows the secretion profile of GLP1 and CCK with procyanidin B2, as well as the combination of this with epicatechin. Procyanidin B2 demonstrates an effective and significant stimulation of the secretion of both hormones when compared to the control. However, interestingly, this stimulating effect on GLP1 and CCK secretion appears to be significantly neutralized by combined treatment with epicatechin. Without wishing to be bound by theory, it appears that epicatechin and procyanidin B2 compete to bind to the TAS25R receptor, shifting the balance towards greater activation of the TAS2R39 pathway resulting in a completely different enterohormone secretion profile and more similar to the activation of the TAS2R39 pathway.

These results demonstrate that the potency and direction of enterohormone release can be modulated by treatment with different TAS2R5 and TAS2R39 agonists.

Example 4: Analysis of the intake induced by an acute dose of 1,10- phenanthroline

The results of ingestion induced by 1 ,10-phenanthroline are shown in Figure 4. Rats treated with an acute dose of 200 mg/kg 290 pM of 1 ,10-phenanthroline showed a feeling of satiety and a significant reduction of the food intake from 12 hours and up to at least 20 hours after treatment.

From these results in conjunction with those of example 1 it can be inferred that the stimulation of TAS2R5 receptors produces a reduction in intake mediated by an enterohormone profile where both high levels of GLP1 and high CCK appear to be relevant. Example 5: Analysis of ingestion induced by an acute dose of epicatechin

The results of epicatechin-induced ingestion are shown in Figure 5. Rats treated with an acute dose of 244 mg/kg 0.84 mM epicatechin showed a feeling of satiety and a significant reduction in food intake at 3 hours, but not at 12 and 20 hours.

In contrast, rats treated with a higher acute dose (300 mg/kg 1 mM) of epicatechin showed a feeling of satiety and a significant reduction in food intake both at 3 hours and at 12 and 20 hours from treatment.

These results indicate that the stimulating effect of the compounds is dosedependent, which makes higher and more concentrated dosages produce more potent and long-lasting responses.

Example 6: Analysis of the intake induced by an acute dose of epicatechin, procyanidin B2 and epicatechin gallate.

The results of the ingestion induced by epicatechin + procyanidin B2 + epicatechin gallate are shown in figure 6. As can be seen, the rats treated with an acute dose of epicatechin + procyanidin B2 + epicatechin gallate (200 + 62 + 18 mg / kg) showed a feeling of satiety and a significant reduction in food intake both at 3 hours and at 12 and 20 hours after treatment.

In Example 3 and Figure 3C it was observed that the combination of epicatechin and procyanidin B2 neutralized each other the "anorectic enterohormonal profile" that each of them exhibited separately. In fact, as will be seen in Example 8, the combination between epicatechin and procyanidin B2 actually produces an orexigenic effect by increasing food intake in rats.

Interestingly, when a Specific TAS2R39 receptor agonist is added to this combination, an orexigenic response could be expected, and yet what is observed is an anorectic response. Without wishing to be bound by any theory, it appears that the presence of a specific and pure agonist of the TAS2R39 receptor such as epicatechin gallate may displace the competition for binding to TAS2R5 of epicatechin and procyanidin 2 resulting in a much more potent activation of the TAS2R5 receptor and therefore, in an anorectic response.

Again this example shows that the effect on intake in a positive or negative sense can be effectively modulated by playing not only with the dose of the different compounds but also with their relative and I or absolute affinities for the TAS2R5 and TAS2R39 receptors. The management of these parameters seems to have a very important impact on the response obtained at the compositional level of the gastrointestinal secretome and therefore on the feeling of appetite I satiety.

Example 7: Analysis of the intake induced by an acute dose of epicatechin and epicatechin gallate.

The results of the intake induced by epicatechin + epicatechin gallate are shown in figure 7. Rats treated with an acute dose of epicatechin (234 mg I kg) and epicatechin gallate (14 mg / kg) showed a significant increase in the intake of food both at 3 hours and at 12 and 20 hours after treatment.

This combination of a mixed TAS2R5 I TAS2R39 agonist and a specific TAS2R39 agonist seems to tend towards greater activation of the TAS2R39 stimulation pathway, the result being an orexigenic effect translated into an increase in food intake.

Example 8: Analysis of the intake induced by an acute dose of epicatechin and procyanidin B2.

The results of the intake induced by epicatechin + procyanidin B2 are shown in figure 8. Rats treated with an acute dose of epicatechin (213 mg / kg) and procyanidin B2 (62 mg I kg) showed a significant increase in the intake of food at 3 and 12 hours, but not at 20 hours after treatment.

This combination of mixed TAS2R5 I TAS2R39 agonists produces a stimulation similar to that of the combination of Example 7 (epicatechin + epicatechin gallate) but less potent. Furthermore, at 20 hours the orexigenic effect of the epicatechin + procyanidin B2 combination disappears while that of the epicatechin + epicatechin gallate combination still remains.

Claims

25 CLAIMS

1. A composition for modulating food intake comprising: a) a combination of at least one mixed or specific agonist of the TAS2R5 receptor and at least one mixed or specific agonist of the TAS2R39 receptor; b) at least one Specific TAS2R5 receptor agonist; c) at least one Specific TAS2R39 receptor agonist; or d) at least one mixed TAS2R5 and TAS2R39 receptor agonist, with a relative affinity (EC50) differentiated between both receptors.

2. A composition according to claim 1 , wherein the mixed TAS2R5 and TAS2R39 receptor agonist is selected from epicatechin, B2-gallate, epigallocatechin gallate (EGCG), Procyanidin B2, Pentagaloylglucose (PGG), Denatonium saccharide, Sucralose and a mixture thereof.

3. A composition according to claim 1 , wherein the Specific TAS2R5 receptor agonist is selected from 1 ,10-phenanthroline, Procyanidin C2, Procyanidin B4 and Procyanindin B7 and a mixture thereof.

3. A composition according to any of the preceding claims, wherein the Specific TAS2R39 receptor agonist is selected from thiamine, epicatechin gallate, acetaminophen and a mixture thereof.

4. A composition according to claim 1 , comprising 1 ,10-phenanthroline.

5. A composition according to claim 1 , comprising thiamine or epicatechin gallate.

6. A composition according to claim 1 , comprising 1 ,10-phenanthroline as an agonist of the TAS2R5 receptor and thiamine as an agonist of the TAS2R39 receptor.

7. A composition according to claim 1 , comprising epicatechin.

8. A composition according to claim 1 , comprising epicatechin and epicatechin gallate.

9. A composition according to claim 1 , comprising epicatechin, procyanidin B2 and epicatechin gallate.

10. A composition according to claim 1 , comprising epicatechin, procyanidin B2.

11. A composition according to any of the preceding claims, which is a food composition, a food supplement or a pharmaceutical composition.

12. A composition according to any of the preceding claims, for use in modulating food intake.

13. A composition for use according to claim 12, wherein the modulation of food intake involves a decrease in food intake.

14. A composition for use according to claim 12, wherein the modulation of food intake involves an increase in food intake.

15. A composition for use according to any one of claims 12 to 14, wherein the modulation of food intake has orexigenic or anorectic purposes in dietary treatments; or a therapeutic purpose in the treatment of diabetes, hyperlipidemia, metabolic syndrome or other disorders associated with obesity, anorexia nervosa or anorexia derived from any pathological process or bulimia nervosa.

16. A method for modulating food intake in a subject comprising administering to the subject a composition according to any of claims 1 to 11.

17. A method according to claim 16, wherein the modulation involves a decrease in food intake comprising administering a composition comprising: a) at least one Specific TAS2R5 receptor agonist; b) a mixed TAS2R5 and TAS2R39 receptor agonist having a lower EC50 for the TAS2R5 receptor than for the TAS2R39 receptor; or c) a combination of at least one Specific TAS2R5 receptor agonist and at least one Specific TAS2R39 receptor agonist, wherein the stimulating effect of the TAS2R5 receptor agonist prevails.

18. A method according to claim 14, wherein the modulation involves increasing food intake comprising administering a composition comprising: a) at least one Specific TAS2R39 receptor agonist b) a mixed TAS2R5 and TAS2R39 receptor agonist having a lower EC50 for the TAS2R39 receptor than for the TAS2R5 receptor; or c) a combination of at least one Specific TAS2R5 receptor agonist and at least one Specific TAS2R39 receptor agonist, where the stimulating effect of the TAS2R39 receptor agonist prevails.