Novel epithelial permeability enhancer

Abstract

The present invention relates to a composition comprising cold water-insoluble cross-linked dextrin and a fatty acid having 8 to 17 carbon atoms. The present invention also relates to a method for preparing such a composition. The present invention also relates to the use of a combination of cold water-insoluble cross-linked dextrin and a fatty acid having 8 to 17 carbon atoms for increasing epithelial permeability of the active ingredient or for epithelial delivery of the active ingredient.

Description

Technical Field

[0001] The present invention relates to the epithelial delivery of low-permeability drugs, particularly to the oral or rectal delivery of low-permeability drugs such as biopharmaceuticals, for example, therapeutic proteins.

Background Art

[0002] There is both great interest and medical need to improve the oral bioavailability of various drugs with low bioavailability. Maximizing oral bioavailability is therapeutically important because the degree of bioavailability directly affects plasma concentration, as well as the therapeutic and toxic effects that occur after oral drug administration. Drugs with low bioavailability are inefficient because most of the dose never reaches the plasma or does not exert its pharmacological effect. Furthermore, compilation of published results for structurally diverse drugs has shown that variability between subjects in bioavailability is inversely correlated with the degree of bioavailability. Thus, low oral bioavailability results in high variability and poor control of plasma concentration and effects.

[0003] Incomplete oral bioavailability has various causes. These include low solubility or low water solubility, degradation of the drug in gastric or intestinal fluid, low intestinal membrane permeability, and pre-systemic intestinal or hepatic metabolism.

[0004] Some physicochemical properties associated with low membrane permeability are low octanol / water partition, the presence of strongly charged functional groups, high molecular weight, a significant number of hydrogen bonding functional groups, and high polar surface area. Compounds that can most benefit from intestinal absorption-promoting formulations usually have one or more of these characteristics. Typically, drugs involved in absorption-promotion studies have been proteins, peptides, peptide analogs, or other polar high molecular weight drugs such as heparin.

[0005] Among the strategies developed to improve the intestinal permeability of these drugs, the use of the following permeation enhancers can be mentioned. - Surfactants (e.g., polyoxyethylene (POE) ether, POE ester, POE sorbitan ester, dodecyl maltoside, nonylphenoxypolyoxyethylene surfactant, sucrose laurate) - Fatty acids (e.g., sodium decanoate, caprylic acid, capric acid, oleic acid, linoleic acid, linolenic acid), -Medium-chain glycerides (e.g., monoglycerides and diglycerides of caprylic and capric acids, monohexanoin) that are ultimately combined with emulsifiers or solubilizers. - Steroidal cleansing agents (e.g., bile salts (sodium taurocholate), chenodeoxycholate, ursodeoxycholate, saponins, glycyrrhizinate, glycyrrhetinic acid, glycosylated bile acid analogs), - Acylcarnitines and alkanoylcholines (e.g., medium-chain and long-chain fatty acid esters of carnitine and choline, e.g., palmitoyl-DL-carnitine chloride, lauroylcholine), -N-acetylated α-amino acids and N-acetylated non-α-amino acids (e.g., N-cyclohexanoylleucine, N-(phenylsulfonyl)leucine, 4-[4-[(2-hydroxybenzoyl)amino]phenyl]butyric acid, N-[8-(2-hydroxybenzoyl)amino]caprylate (SNAC)), - Chitosan and other mucosal adhesive polymers (e.g., anionic polyacrylic acid derivatives, polycarbophils and carbomers (i.e., Carbopol 934P), cationic chitosan, trimethylchitosan), - Secretory transport inhibitors (e.g., P-glycoprotein (Pgp) inhibitors, e.g., SDZ PSC 833, cyclosporine, polysorbate 80, POE 35 castor oil (Cremophor EL®), poloxamer (Pluronic P85®), - Cyclodextrin.

[0006] However, the need for additional or more efficient intestinal permeability enhancers still exists.

[0007] The development of permeability enhancers useful for intestinal permeability will not only increase the bioavailability of orally administered low-permeability drugs, but also increase the bioavailability of those administered rectally. Furthermore, applications can be found in other epithelial routes of administration that require the active ingredient to cross the epithelium, such as the skin route, mucosal route (e.g., vaginal route, sublingual route, oral route), transdermal route, ocular route, nasal route, transnasal route, or bronchopulmonary route.

[0008] technical issues The object of the present invention was to provide an improved intestinal permeability enhancer, more broadly, an improved epithelial permeability enhancer, for low-permeability active ingredients such as biopharmaceuticals.

[0009] The object of the present invention was to provide a material that can improve the bioavailability of active ingredients such as biopharmaceuticals when ingested orally or rectally, or more broadly, when ingested via the epithelial pathway.

[0010] The object of the present invention was to solve this technical problem by providing a solution that has other properties required for its intended purpose, for example, from the viewpoint of purity and safety.

[0011] Presentation of the invention The inventors of this invention, -Cold water insoluble cross-linked dextrin, The above-mentioned problem was solved by providing an epithelial (e.g., intestinal) permeability enhancer that is a combination of a fatty acid having 8 to 17 carbon atoms.

[0012] As will be apparent from the following sections of examples in this specification, these substances act synergistically, thus enabling a high degree of intestinal permeability.

[0013] The inventors demonstrated that this synergistic effect requires this very specific combination. In fact, no synergistic effect was observed when permeabilis enhancers other than fatty acids having 8 to 17 carbon atoms were used (see Examples Sections 1.2 and 2.2, Experiment 1). Similarly, no synergistic effect was observed when non-crosslinked dextrin was used (see Examples Section 2.2, Experiment 2).

[0014] When such combinations are used with an active ingredient that needs to be improved for intestinal permeability, the active ingredient must be associated with a cold water-insoluble cross-linked dextrin (see Section 2.2 of the Examples).

[0015] Results obtained in the Caco-2 cell model suggest that it is a promising permeability enhancer for intestinal routes, such as oral and rectal routes. It is also a promising permeability enhancer for other epithelial routes, such as cutaneous routes, mucosal routes (e.g., vaginal, sublingual, oral routes), transdermal routes, ocular routes, nasal routes, transnasal routes, or bronchopulmonary routes. [Overview of the Initiative]

[0016] This disclosure is, firstly, -Cold water insoluble cross-linked dextrin, The present invention relates to a composition comprising a fatty acid having 8 to 17 carbon atoms.

[0017] Preferably, the dextrin is selected from pyrodextrin, maltodextrin, cyclodextrin, or a mixture thereof. Preferably, the cold water-insoluble crosslinked dextrin can be obtained by reacting dextrin with a crosslinking agent, the crosslinking agent being selected from trimetaphosphate, dicarboxylic acid, dianhydride, carbonyldiimidazole, diphenyl carbonate, triphosgene, acyl dichloride, diisocyanate, diepoxide, or any mixture thereof. Preferably, the cold water-insoluble crosslinked dextrin can be obtained by reacting dextrin with a crosslinking agent, the molar ratio of the crosslinking agent to the anhydrous glucose units of the dextrin is 0.1:1 or greater. Preferably, the fatty acid has at least 9 carbon atoms. Preferably, the fatty acid has up to 16 carbon atoms, more preferably up to 15 carbon atoms, even more preferably up to 14 carbon atoms, even more preferably up to 13 carbon atoms, even more preferably up to 12 carbon atoms, and even more preferably up to 11 carbon atoms. More preferably, the fatty acid has 10 carbon atoms. Preferably, the fatty acid is in the form of a salt, and more preferably in the form of a sodium salt. This is even more preferably sodium decanoate. Preferably, the cold water-insoluble cross-linked dextrin is in the form of particles.

[0018] This disclosure also states, -Cold water insoluble cross-linked dextrin, - Fatty acids having 8 to 17 carbon atoms, - Relating to a composition containing an active ingredient.

[0019] Preferably, the active ingredient is selected from BCS class III drugs, BCS class IV drugs, biopharmaceuticals, or any mixture thereof. Preferably, the active ingredient is selected from proteins. Preferably, the active ingredient is filled into the cold water-insoluble cross-linked dextrin.

[0020] The present disclosure also relates to such compositions for use as medicaments, and to the use of such compositions in food compositions such as food supplements, or dietary supplements, or cosmetic compositions.

[0021] Preferably, the compositions according to the present disclosure are intended to be administered by an epithelial route, preferably an intestinal route (including oral or rectal route), a skin route, a mucosal route (e.g., vaginal route, sublingual route, oral route), a transdermal route, an ocular route, a nasal route, a transmucosal nasal route, or a bronchopulmonary route. Preferably, the compositions according to the present disclosure are intended to be administered by an intestinal route, preferably an oral or rectal route, more preferably an oral route. In other words, these are preferably oral or rectal compositions, more preferably oral compositions.

[0022] The term "epithelial route" classically refers to an administration route that requires crossing the epithelium. This may be a systemic route or a local administration route (e.g., as in the case of skin administration of an active ingredient that needs to cross the epidermis to reach the dermis). This is more preferably a systemic administration route.

[0023] The present disclosure also relates to a method for preparing such compositions, which comprises contacting the cold water-insoluble crosslinked dextrin with the fatty acid having 8 to 17 carbon atoms.

[0024] The present disclosure also relates to - a combination of a cold water-insoluble crosslinked dextrin and - a fatty acid having 8 to 17 carbon atoms, for use in increasing the epithelial permeation of an active ingredient, preferably increasing the intestinal permeation of an active ingredient.

[0025] The present disclosure also relates to - a combination of a cold water-insoluble crosslinked dextrin and - a fatty acid having 8 to 17 carbon atoms, for use in This invention relates to the use of an active ingredient for epithelial delivery, preferably for oral or rectal delivery, and more preferably for oral delivery.

[0026] Preferably, the active ingredient is selected from BCS class III drugs, BCS class IV drugs, biopharmaceuticals, or any mixture thereof. Preferably, the active ingredient is selected from proteins. [Brief explanation of the drawing]

[0027] Other features, details, and advantages are shown in the detailed description and drawings below. [Figure 1] Schematic diagram of a 24-well Caco-2 model (only the path from the apex to the side base). [Figure 2] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and sodium decanoate. [Figure 3] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and salcaprozate sodium. [Figure 4] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and lauryl-L-carnitine. [Figure 5] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and nonaethylene glycol monododecyl ether. [Figure 6] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and polyethylene glycol. [Figure 7] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and monodisperse 20 μm silica. [Figure 8]This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and monodisperse 150 nm silica. [Figure 9] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and kaolinite. [Figure 10] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and montmorillonite. [Figure 11] This bar graph shows the insulin permeability when using a combination of cold water-insoluble cross-linked dextrin (nanosponge) and various permeability enhancers (sodium decanoate, linoleic acid, sodium salcaprozate), when using nanosponge alone, or when using a permeability enhancer alone. [Figure 12] This bar graph shows the insulin permeability when cold water-insoluble cross-linked dextrin (nanosponge) is used in combination with sodium decanoate at different concentrations, when nanosponge is used alone, when sodium decanoate is used alone at different concentrations, or when uncross-linked dextrin is used. [Figure 13] Figure 12 is a bar graph showing the results of a barrier integrity test using Lucifer Yellow, conducted after the experiment whose results were shown in Figure 12. [Modes for carrying out the invention]

[0028] The drawings and the following detailed description contain several precise elements. These can be used to enhance the understanding of the invention and, where necessary, to define the invention.

[0029] Composition containing cold water-insoluble cross-linked dextrin and fatty acids The present invention, firstly, -Cold water insoluble cross-linked dextrin, The present invention relates to a composition comprising a fatty acid having 8 to 17 carbon atoms.

[0030] The term "dextrin" classically refers to products obtained from starch hydrolysis, including maltodextrin, glucose syrup having 20-30 dextrose equivalents (DE), pyrodextrin, and cyclodextrin. Preferably, the dextrin according to this disclosure is selected from maltodextrin, pyrodextrin, cyclodextrin, or any mixture thereof. This is more preferably selected from maltodextrin, pyrodextrin, or any mixture thereof. This is more preferably selected from maltodextrin or any mixture thereof, i.e., this is preferably cold water insoluble crosslinked maltodextrin.

[0031] The term "maltodextrin" classically refers to dextrin obtained by acid and / or enzymatic hydrolysis of starch. Maltodextrin is typically characterized by a DE of less than 20. Preferably, the maltodextrin according to this disclosure has a DE of 1 or more, preferably 5 or more, preferably 10 or more, and preferably 15 or more. This is preferably 19 or less. This is, for example, equal to about 17.

[0032] The term "pyrodextrin" classically refers to dextrin obtained by dry heating starch under acidic conditions, which generally results in the hydrolysis of starch and subsequent recombination via α-1,6 bonds. Generally, these pyrodextrins are classified as "white dextrin," "yellow dextrin," or "British gum" depending on the temperature, acidity, and humidity conditions under which they are used.

[0033] The term "cyclodextrin" generally refers to natural or substituted cyclic dextrins having 6 to 12 glucose units linked to each other via carbon atoms C1 and C4. Cyclodextrins include α-, β-, and γ-cyclodextrins, each having 6, 7, and 8 glucose units, respectively. Preferably, the cyclodextrins according to this disclosure are selected from α-cyclodextrin, β-cyclodextrin, γ-cyclodextrin, or any mixture thereof, more preferably from β-cyclodextrin. The cyclodextrins according to this disclosure may preferably be substituted with alkyl and / or hydroxyalkyl and / or sulfoalkyl groups, and may particularly be etherified. Preferably, the alkyl, hydroxyalkyl, and sulfoalkyl groups according to this disclosure have 1 to 5, preferably 1 to 4, preferably 1, 3, or 4 carbon atoms. Preferably, the substituted cyclodextrins according to the Disclosure are selected from methyl-cyclodextrin, hydroxypropyl-cyclodextrin, sulfobutyl ether-cyclodextrin, or any mixture thereof. In contrast to chemical substances with clearly defined structures, substituted cyclodextrins are generally mixtures of substituted cyclodextrins having different substitution patterns and are therefore structurally different. More preferably, the cyclodextrins according to the Disclosure are unsubstituted cyclodextrins (i.e., natural cyclodextrins), preferably unsubstituted β-cyclodextrins (i.e., natural β-cyclodextrins).

[0034] Dextrin is classically obtained from starch. The term “starch” classically refers to starch isolated from any suitable plant source by any technique well known to those skilled in the art. Isolated starch typically contains impurities of 3% by weight or less. This percentage is expressed as the dry weight of impurities relative to the total dry weight of isolated starch. These impurities typically include proteins, colloidal substances, and fibrous residues. Suitable plant sources include, for example, legumes, cereals, and tubers.

[0035] Therefore, the dextrins according to this disclosure can be selected from leguminous plant dextrins (e.g., pea dextrin, broad bean dextrin), cereal dextrins (e.g., maize dextrin, rice dextrin, wheat dextrin, oat dextrin), and tuber dextrins (e.g., potato dextrin, tapioca dextrin). Preferably, the dextrins according to this disclosure are selected from leguminous plant dextrins, cereal dextrins, or any mixture thereof.

[0036] Preferably, the leguminous plant dextrin is selected from pea dextrin, broad bean dextrin, or any mixture thereof, more preferably from pea dextrin. The term “pea” classically includes all wild and variant varieties of “smooth pea” and “wrinkled pea,” regardless of the generally intended use of the variety (human food, animal feed, and / or other uses). The term “pea” includes peas of Pisum genius, more specifically, peas of the sativum and aestivum species. The mutant varieties may be referred to as "r mutants," "rb mutants," "rug3 mutants," "rug4 mutants," "rug5 mutants," and "lam mutants," as described in the paper by CL HEYDLEY et al., titled "Developing novel pea starches," Proceedings of the Symposium of the Industrial Biochemistry and Biotechnology Group of the Biochemical Society, 1996, pages 77-87. Preferably, the dextrin according to this disclosure is selected from smooth pea dextrin, and more preferably from wild smooth pea dextrin.

[0037] Preferably, the cereal dextrin is corn dextrin.

[0038] Preferably, if the dextrin according to this disclosure is maltodextrin, it is leguminous maltodextrin, preferably pea maltodextrin, preferably smooth pea maltodextrin, preferably wild smooth pea maltodextrin.

[0039] Preferably, if the dextrin according to this disclosure is a pyrodextrin, it is a cereal pyrodextrin, preferably a maize pyrodextrin.

[0040] In preferred embodiments, the dextrin according to the present disclosure is obtained from starch having an amylose content of 20% or more, preferably 25% or more, preferably 30% or more, and preferably 35% or more, where this content is expressed as dry weight relative to the total dry weight of the starch. The amylose content is preferably 90% or less, preferably 60% or less, preferably 50% or less, preferably 45% or less, preferably 40% or less, and preferably 38% or less. Preferably, the starch is leguminous plant starch, preferably pea or broad bean starch, more preferably smooth pea starch, and even more preferably wild smooth pea starch. The amylose content can be determined by those skilled in the art by potentiometric analysis of iodine absorbed by amylose to form a complex.

[0041] Preferably, the dextrin according to the present disclosure, and in particular the maltodextrin according to the present disclosure, has a weight-average molecular weight (Mw) of 1000 Da or more, preferably 5000 Da or more, preferably 8000 Da or more, and preferably 10000 Da or more, as determined by liquid chromatography with detection by differential refractometer. This Mw is preferably 200000 Da, preferably 100000 Da or less, preferably 50000 Da or less, preferably 40000 Da or less, preferably 30000 Da or less, preferably 20000 Da or less, and preferably 15000 Da or less. This is, for example, equal to about 12000 Da.

[0042] This Mw is preferably determined by liquid chromatography with detection by a differential refractometer, using a pullulan standard. This can be determined by those skilled in the art according to the following protocol. Use the following column set (for example, Shodex OH pak SB-800 QH) in the following order. A column with a particle size of -8 μm, a pore size of 100 Å, an inner diameter of 8.0 mm, and a length of 300 mm (e.g., OH pak SB-802 HQ-Waters ref. JWE 034256), A column having a particle size of -6 μm, a pore size of 800 Å, an inner diameter of 8.0 mm, and a length of 300 mm (e.g., OH pak SB-803 HQ-Waters ref. JWE 034257), A column with a particle size of -13 μm, a pore size of 7000 Å, an inner diameter of 8.0 mm, and a length of 300 mm (e.g., OH pak SB-805 HQ-Waters ref. JWE 034259).

[0043] Use the following Pull-Run standards with Mw values ​​(e.g., Kit Waters-Ref.JWE034207): P800, P400, P200, P100, P50, P20, P10, P5.

[0044] Conditions: Elution solvent: 0.1 M sodium nitrate aqueous solution containing 0.02% sodium azide filtered through a 0.02 μm filter, mobile phase flow rate: 0.5 mL / min, column temperature: 35 °C, injection volume: 100 μL, detector: RI decay: 16, internal temperature: 35 °C, analysis time: 80 minutes.

[0045] Calibration was performed using glycerol Mw=92, glucose Mw=180, maltose Mw=342, maltotriose Mw=504, maltotetraose Mw=667, maltopentaose Mw=829, maltohexaose Mw=991, and maltheptaose Mw=1153.

[0046] Suitable examples of dextrins are commercially available. For example, the following products commercially available from ROQUETTE can be cited: KLEPTOSE® β-cyclodextrin, KLEPTOSE® Linecaps (pea maltodextrin), GLUICIDEX® 2 (waxy maize pyrodextrin), STABILYS® A025 (maize pyrodextrin), STABILYS® A053 (maize pyrodextrin), and the range of TACKIDEX®.

[0047] The cold water-insoluble dextrins according to this disclosure are crosslinked, meaning that they can (or are obtained) by reacting dextrin with a crosslinking agent. The crosslinking agent is typically polyfunctional, i.e., it comprises at least two reactive groups. Preferably, the crosslinking agent is bifunctional. Typically, in this disclosure, the reactive groups are subject to nucleophilic attack, i.e., have atoms with a partial positive charge.

[0048] Preferably, the crosslinking agent according to the present disclosure is selected from trimetaphosphate, dicarboxylic acid, dianhydride, carbonyldiimidazole, diphenyl carbonate, triphosgene, acyl dichloride (acylic), diisocyanate, diepoxide, or any mixture thereof. Preferably, the dicarboxylic acid is selected from polyacrylic acid, butanetetracarboxylic acid, succinic acid, tartaric acid, citric acid, or a mixture thereof. More preferably, it is selected from citric acid and / or tartaric acid. Preferably, the dianhydride is selected from diethylenetriaminepentaacetic acid dianhydride, ethylenediaminetetraacetic acid dianhydride, benzophenone-3,3',4,4'-tetracarboxylic acid dianhydride, pyromellitic acid dianhydride, or any mixture thereof. More preferably, it is pyromellitic acid dianhydride. Preferably, the acyl chloride is selected from terephthaloyl chloride, sebacoyl chloride, succinyl chloride, or a mixture thereof. More preferably, it is terephthaloyl chloride. Preferably, the diisocyanate is selected from toluene diisocyanate, isophorone diisocyanate, 1,4-phenylenediisocyanate, poly(hexamethylene diisocyanate), hexamethylene diisocyanate, or a mixture thereof. More preferably, this is hexamethylene diisocyanate.

[0049] Preferably, the crosslinking agent according to this disclosure is selected from trimetaphosphate, pyromellitic dianhydride, 1,1'-carbonyldiimidazole, hexamethylene diisocyanate, citric acid, tartaric acid, or a mixture thereof.

[0050] In preferred embodiments, particularly when dextrin is selected from maltodextrin and / or pyrodextrin, the crosslinking agent is selected from trimetaphosphates. More preferably, this is sodium trimetaphosphate.

[0051] In preferred embodiments, particularly when the dextrin is selected from cyclodextrins, the crosslinking agent is pyromellitic dianhydride.

[0052] Preferably, the cold water-insoluble dextrin according to this disclosure can be obtained (or is obtained) by reacting dextrin with a crosslinking agent without undergoing a prior step of ethylamination (grafting with an ethylamine group).

[0053] Preferably, the molar ratio of the crosslinking agent dextrin to anhydrous glucose units is 0.1:1 or higher, preferably 0.2:1 or higher, and preferably 0.20:1 or higher. This is preferably 10:1 or lower, preferably 5:1 or lower, preferably 4:1 or lower, preferably 3:1 or lower, preferably 2:1 or lower, preferably 1:1 or lower, preferably 1.0:1 or lower, preferably 0.9:1 or lower, preferably 0.8:1 or lower, preferably 0.7:1 or lower, preferably 0.6:1 or lower, preferably 0.5:1 or lower, preferably 0.4:1 or lower, and preferably 0.40:1 or lower. This is, for example, equal to about 0.3:1.

[0054] In this disclosure, this is considered to be the amount or ratio of the components constituting the cold water-insoluble crosslinked dextrin. It should be understood that these amounts refer to the amounts of starting material used to produce the cold water-insoluble crosslinked dextrin (dextrin, crosslinking agent, and other final components). These amounts may differ slightly from the amounts actually present in the final matrix (i.e., in the cold water-insoluble crosslinked dextrin prepared therefrom), which, to the best of our knowledge, cannot be measured at present.

[0055] The dextrins or crosslinked dextrins according to this disclosure may undergo a physical modification process commonly referred to as “digestion” or “gelatinization.” This digestion process is well known to those skilled in the art and is typically carried out when the dextrin is at least partially in a granular state. This digestion process allows the polymer molecules constituting the dextrin to be completely and uniformly dispersed in a reaction solvent (e.g., water). This digestion process is particularly useful when the dextrin is a pyrodextrin. Preferably, digestion is carried out before the crosslinking process.

[0056] The cross-linked dextrins according to this disclosure are insoluble in cold water. The term "cold water" classically refers to water left at room temperature (generally 18-25°C). Preferably, the cross-linked dextrins according to this disclosure are insoluble in cold water at pH 7. Preferably, the cross-linked dextrins according to this disclosure are insoluble in cold water at pH 5. Preferably, the cross-linked dextrins according to this disclosure are insoluble in cold water at pH 9.

[0057] The cross-linked dextrins according to this disclosure are insoluble in cold water. That is, the cross-linked dextrins typically have a solubility of 20% or less in cold water, and this percentage is expressed as the dry weight of the soluble substance relative to the total dry weight of the cross-linked dextrin. This solubility is preferably 15% or less, preferably 10% or less, preferably 9% or less, preferably 8% or less, preferably 7% or less, preferably 6% or less, preferably 5% or less, preferably 4% or less, preferably 3% or less, preferably 2% or less, and preferably 1% or less. This solubility can be determined by those skilled in the art according to the following protocol: 2.5 g of the product to be tested (e.g., cross-linked dextrin particles obtained after, for example, grinding and sieving through a 315 μm sieve to remove coarse particles) is placed in 150 g of desalted water and stirred for 16 hours, then centrifuged at 4700 rpm for 15 minutes (e.g., using a VWR Mega Star 1.6 centrifuge). Next, the supernatant is placed in a crystallization apparatus that has been pre-tare-measured and left in a 55°C vacuum oven until there is no further weight loss (all water evaporates). The remainder (soluble substance) is then weighed. This soluble substance fraction may contain the soluble fraction of cross-linked dextrin, but may also contain some residual impurities.

[0058] Preferably, the cold water-insoluble crosslinked dextrin according to this disclosure is in the form of particles. These particles may have an average diameter selected from 1 nm to 1000 μm, for example, from 10 nm to 500 μm, and the diameter is measured in an aqueous suspension of the cold water-insoluble crosslinked dextrin. These particles may have an average diameter of 1000 nm or less (also referred to as "nanosponges").

[0059] This can be used in the form of a nano-suspension (nano-sponge suspension) of the particles of cold water-insoluble cross-linked dextrin. The suspension can be prepared according to the following protocol. Prepare a suspension starting from crude powder in distilled water with a concentration of 1.10 mg / mL by stirring at room temperature. 2. Disperse the suspension at 24,000 rpm for 10 minutes using a high-shear homogenizer (e.g., Ultraturrax®, IKA, Konigswinter, Germany). Further size reduction is achieved by high-pressure homogenization at a back pressure of 3,500 bar for 90 minutes (e.g., using an EmulsiFlex C5 instrument (Avastin, USA)). 4. The homogenized nanosuspension is purified by dialysis using a cellulose membrane (Spectrapor) with a cutoff of, for example, 12,000 Da, to remove any potentially present synthetic residues. 5. Store the nano-suspension at 4°C if necessary.

[0060] Preferably, the cold water-insoluble crosslinked dextrin particles according to this disclosure have an average diameter of 1 to 1000 nm. This is preferably 10 nm or more, preferably 50 nm or more, and preferably 100 nm or more. This average diameter is the hydrodynamic diameter. This can be determined by those skilled in the art, for example, by laser light scattering on a nanosponge suspension obtained according to the protocol described above, preferably using a 90Plus apparatus (Brookhaven, NY, USA), which contains 10 mg / mL of nanosponge diluted with filtered (0.22 μm) distilled water using a dilution factor of 1 / 30 by volume.

[0061] Preferably, the polyvariance index with respect to the average diameter is less than 1.00. This polyvariance index is generally higher than 0.01, and more preferably 0.05 or higher.

[0062] Preferably, the cold water-insoluble dextrin according to this disclosure has a zeta potential of less than 0 mV, preferably -10 mV or less, and preferably -20 mV or less. This is generally -50 mV or more, and even more preferably -40 mV or more. This zeta potential can be determined by those skilled in the art using, for example, a 90Plus apparatus (Brookhaven, NY, USA) to the electrophoretic mobility of a crosslinked dextrin suspension by dynamic light scattering at a scattering angle of 90° at a temperature of 25°C. The sample is placed in an electrophoresis cell and an electric field of 15 V / cm is applied. Preferably, this zeta potential is determined, for example, in a nanosponge suspension obtained according to the protocol described above, where the suspension containing 10 mg / mL of nanosponge is diluted with filtered (0.22 μm) distilled water using a dilution factor of 1 / 30 by volume.

[0063] Preferably, the cold water-insoluble crosslinked dextrin according to this disclosure has a swelling index (SI%) of 200% or more, preferably 500% or more, preferably 600% or more, preferably 700% or more, preferably 800% or more, preferably 900% or more, preferably 1000% or more, preferably 1100% or more, preferably 1200% or more, preferably 1300% or more, preferably 1400% or more, preferably 1500% or more, and preferably 1600% or more. This is preferably 5000% or less, preferably 4000% or less, preferably 3000% or less, and preferably 2000% or less.

[0064] The swelling index of the matrix (e.g., cold water insoluble dextrin according to this disclosure) is defined by the following formula:

[0065]

number

[0066] To determine this SI%, 1 g (dry weight) of matrix (after being crushed and sieved through a 315 μm sieve to remove coarse particles) is dispersed in 100 mL of desalted water in a graduated cylinder and left to swell for 24 hours. After 24 hours of contact, the mixture of matrix dispersed in water is centrifuged to separate the supernatant (water) from the lower layer (swollen matrix or gel). The swollen matrix is ​​then weighed.

[0067] The dextrins according to this disclosure may undergo other chemical and / or physical modifications other than those described above (i.e., crosslinking and ultimately pulverization), as long as they do not impede the desired properties, particularly from the standpoint of safety and efficiency. However, since there is no need to solve the technical problems disclosed herein, the dextrins according to this disclosure are preferably not further modified.

[0068] The cold water-insoluble crosslinked dextrins according to this disclosure may contain other components in their structure besides the crosslinking pattern derived from the dextrin and the crosslinking agent, provided that such other components do not interfere with the desired properties of the cold water-insoluble crosslinked dextrin, particularly from the standpoint of efficiency and safety. The term “other components” is understood not to refer to small amounts of impurities ultimately resulting from the dextrin and the crosslinking agent. Examples of such other components are other polymers, such as proteins, which are typically crosslinked when used.

[0069] However, since there is no need to solve the technical problems disclosed herein, the cold water-insoluble crosslinked dextrin according to this disclosure preferably contains an amount of the other component equal to 30% or less, preferably 20% or less, preferably 10% or less, preferably 5% or less, preferably 1% or less, and preferably 0%, where the percentage is expressed as dry weight relative to the total dry weight of the cold water-insoluble dextrin. More preferably, the cold water-insoluble crosslinked dextrin according to this disclosure does not contain the other component.

[0070] Therefore, in preferred embodiments, the cold water-insoluble crosslinked dextrin according to this disclosure is composed of a crosslinked dextrin. Preferably, the dextrin is selected from maltodextrin, pyrodextrin, cyclodextrin, or any mixture thereof. More preferably, the cold water-insoluble dextrin is composed of a crosslinked maltodextrin, or a crosslinked pyrodextrin, or a crosslinked cyclodextrin.

[0071] Examples of suitable cold water-insoluble crosslinked dextrins and processes for producing them are described in International Publication No. 2016 / 004974(A1)(ROQUETTE) and International Publication No. 2021 / 254662(A1)(ROQUETTE).

[0072] The compositions according to this disclosure also include fatty acids having 8 to 17 carbon atoms.

[0073] The fatty acid may be in the form of a salt or a protonated form. Preferably, it is a salt of the fatty acid.

[0074] Preferably, the fatty acid according to the disclosure has at least 9 carbon atoms. Preferably, the fatty acid according to the disclosure has up to 16 carbon atoms, preferably up to 15 carbon atoms, preferably up to 14 carbon atoms, preferably up to 13 carbon atoms, preferably up to 12 carbon atoms, preferably up to 11 carbon atoms. More preferably, the fatty acid according to the disclosure has 10 carbon atoms.

[0075] Preferably, the fatty acids according to this disclosure are saturated fatty acids.

[0076] Preferably, the fatty acid according to this disclosure is a decanoate, decanoic acid, or any mixture thereof, and more preferably a decanoate. Preferably, the decanoate is sodium decanoate.

[0077] The fatty acids according to this disclosure may be slightly chemically modified, in particular from the viewpoint of efficiency and safety, as long as this does not interfere with the desired properties of the fatty acids. Preferably, the fatty acids according to this disclosure are unchemically modified, as there is no need to solve the technical problems disclosed herein.

[0078] Preferably, the dry weight ratio of fatty acids to cold water-insoluble cross-linked dextrin is 0.05:1 to 5:1. This is preferably 0.10:1 or higher, preferably 0.15:1 or higher, preferably 0.2:1 or higher, preferably 0.3:1 or higher, and preferably 0.30:1 or higher. This is preferably 4:1 or lower, preferably 3:1 or lower, preferably 2:1 or lower, preferably 1.5:1 or lower, preferably 1:1 or lower, preferably 1.0:1 or lower, preferably 0.9:1 or lower, preferably 0.8:1 or lower, preferably 0.7:1 or lower, preferably 0.6:1 or lower, preferably 0.5:1 or lower, preferably 0.50:1 or lower, and preferably 0.40:1 or lower. This is, for example, selected from about 0.2 to about 0.8, preferably about 0.3:1 to about 0.4:1.

[0079] In preferred embodiments, the compositions according to this disclosure do not contain cold-water-insoluble cross-linked dextrins grafted with fatty acids having 8 to 17 carbon atoms. "Grafting" is understood to mean that fatty acids are covalently bonded to the cross-linked dextrin.

[0080] The present invention also relates to the use of the compositions according to this disclosure in pharmaceuticals, or in food compositions such as dietary supplements, or in nutritional supplements or cosmetic compositions.

[0081] The compositions according to this disclosure may be products intended for administration to an individual, or compositions useful for preparing compositions intended for administration to an individual. These may be powdered compositions, tablets, or suspensions. The powdered composition may be administered as is, or ultimately with the addition of water. It may also be encapsulated, for example, in a hard capsule. The suspension may be administered as is, or encapsulated, for example, in a soft capsule.

[0082] A composition comprising cold water-insoluble cross-linked dextrin, fatty acids, and an active ingredient. The combination of cold water-insoluble cross-linked dextrin and a fatty acid having 8 to 17 carbon atoms according to this disclosure enables increased intestinal permeability (and more broadly, epithelial permeability) of the active ingredient.

[0083] Therefore, this disclosure also, -Cold water insoluble cross-linked dextrin, - Fatty acids having 8 to 17 carbon atoms, - Relating to a composition containing an active ingredient.

[0084] Preferably, the composition is as described above. Preferably, the cold water-insoluble crosslinked dextrin is as described above. Preferably, the fatty acid having 8 to 17 carbon atoms is as described above.

[0085] The term “active ingredient” classically refers to any substance that is the subject of a pharmaceutical, veterinary, food, dietary supplement, or cosmetic product. Preferably, the active ingredients according to this disclosure are pharmaceutical, dietary supplement, cosmetic, or veterinary active ingredients, and more preferably, pharmacoactive ingredients. The active ingredients according to this disclosure, in particular pharmacoactive ingredients, may be selected from so-called small molecules or so-called large molecules (also referred to as “biopharmaceuticals”), for example, proteins, nucleic acids, viruses, and cells. Biopharmaceuticals are typically not bioavailable when taken orally, for example, because this route of administration causes their degradation and / or because they cannot cross biological membranes (including epithelium). In other words, these are typically active ingredients that need to be administered parenterally and for which systemic effects are desired. Non-limiting examples of biopharmaceuticals include vaccines, blood components (e.g., coagulation factors, blood fractions), antibodies (e.g., monoclonal antibodies, humanized antibodies, chimeric antibodies, monoclonal antibody fragments, e.g., variable fragments of antibodies), allergens, hormones such as insulin, gene therapies, tissues, cell therapies, and recombinant therapeutic proteins. Preferably, the biopharmaceuticals according to this disclosure are selected from proteins. The term “protein” should be understood in a broad sense. It includes proteins in particular, regardless of their manufacturing process or the number of their subunits. It also includes protein fragments, peptides, and oligopeptides. It may be selected from natural proteins, recombinant proteins, fusion proteins, or any mixture thereof. If the protein is derived from a natural product, e.g., from a plant, it is preferably understood to be an isolated protein.

[0086] Preferably, the protein according to the disclosure has at least 5 amino acids, preferably at least 10, preferably at least 20, preferably at least 30, preferably at least 40, and preferably at least 50 amino acids. Preferably, the protein according to the disclosure has up to 5000 amino acids, preferably up to 1000, preferably up to 500, preferably up to 400, preferably up to 300, preferably up to 200, preferably up to 100, preferably up to 90, preferably up to 80, preferably up to 70, and preferably up to 60 amino acids.

[0087] Preferably, the protein according to this disclosure (e.g., a pharmaceutical protein) is selected from enzymes, cytokines, hormones, growth factors, plasma factors, vaccines, and antibodies. This is preferably insulin. The term "insulin" encompasses insulin or any pharmaceutically active derivative thereof, preferably insulin.

[0088] Preferably, the active ingredient according to this disclosure is an active ingredient that needs to increase epithelial (preferably intestinal) permeability. This is preferably selected from BCS class III and / or IV drugs (the Biopharmaceutical Classification System of the U.S. Food and Drug Administration, effective November 1, 2022).

[0089] Non-limiting examples of active ingredients that need to be increased in epithelial (especially intestinal) permeability include erythromycin, colistin, cephamandol, cefotaxime, moxalactam, mezlocillin, penicillin G, ampicillin, cefoxitin, carmonamgentamicin, vancomycin; octreotide; calcitonin; cromolyn; insulin; glucagon; recombinant human growth hormone; and low-absorption antibodies such as doxorubicin, paclitaxel, etoposide, azidodeoxythymidine, and argvasopressin. Preferably, the active ingredient according to this disclosure is a biopharmaceutical, more preferably a protein, and even more preferably insulin.

[0090] Preferably, the active ingredient according to this disclosure is an active ingredient whose absorption can be increased by transport through tight bonds.

[0091] Preferably, the active ingredient according to this disclosure is intended to be administered by a route selected from the following: intestinal route (including oral or intrarectal route), cutaneous route, mucosal route (e.g., vaginal route, sublingual route, oral route), transdermal route, ocular route, nasal route, transnasal route, or bronchopulmonary route. Preferably, the active ingredient according to this disclosure is intended to be administered by an intestinal route, preferably intrarectally or orally. More preferably, it is intended to be administered orally.

[0092] Preferably, the active ingredient is filled into the cold water-insoluble crosslinked dextrin according to this disclosure. To the extent that the nature of the association between the crosslinked dextrin and the active ingredient is unknown, the term “filled” is intended to mean “associated,” whether within or on the surface of the matrix formed by the crosslinked dextrin.

[0093] The filling can be carried out, for example, by adding the active ingredient to a pre-formed nanosponge suspension prepared as described above. The mixture is then stirred, for example, at room temperature for 30 minutes to incorporate the active ingredient. The medium is then centrifuged and the precipitate is collected. This can be freeze-dried for future use.

[0094] Preferably, the packing volume of the cold water-insoluble cross-linked dextrin according to this disclosure is 1% or more, and this percentage is expressed as the dry weight of the active ingredient relative to the total dry weight of the packed cold water-insoluble cross-linked dextrin. This packing volume is preferably 5% or more, preferably 10% or more, and preferably 15% or more. Generally, it is 50% or less, more preferably 40% or less, more preferably 30% or less, and more preferably 20% or less. This packing volume can be determined by those skilled in the art according to the following protocol, and the packing volume is preferably determined from a lyophilized packing sample prepared as described above. Briefly, a weighed amount of 2-3 mg of the lyophilized delivery system packed with the active ingredient is dispersed in 5 mL of distilled water. Sonication (15 minutes, 100 W) and centrifugation are performed to release the active ingredient from the cross-linked dextrin. The supernatant is then analyzed to quantify the active ingredient. The packing volume of the delivery system is calculated as follows. [Dry weight of active ingredient / Dry weight of freeze-dried packing sample] × 100.

[0095] The compositions comprising the active ingredient according to this disclosure may be dosage forms, i.e., products intended to be administered to an individual, or compositions useful for preparing dosage forms. These may be powdered compositions, tablets, or suspensions. The powdered composition may be administered as is, or ultimately with the addition of water. It may also be encapsulated, for example, in a hard capsule. The suspension may be administered as is, or encapsulated, for example, in a soft capsule.

[0096] In a preferred embodiment, the composition according to the present disclosure is a suspension.

[0097] Use of the composition as disclosed herein The present invention also relates to compositions according to the present disclosure, comprising an active ingredient, for use as pharmaceuticals, and to the use of compositions according to the present disclosure in food compositions such as dietary supplements, or nutritional supplements, or cosmetic compositions. In the context of the present invention, use as a pharmaceutical is intended to be for human or veterinary use, preferably human.

[0098] The present invention also relates to a method for treating or preventing a disease in an organism that requires treatment or prevention of the disease, the method comprising administering to the organism a composition according to the present disclosure comprising an active ingredient.

[0099] The present invention also relates to a method for feeding an organism in need of feeding, comprising administering to the organism a composition according to the present disclosure containing an active ingredient.

[0100] Preferably, the chemical and composition are as described above. In particular, the active ingredient is preferably filled in cold water-insoluble cross-linked dextrin.

[0101] Preferably, the drug and composition are intended to be administered by a route selected from the following: intestinal route (including oral or rectal route), cutaneous route, mucosal route (e.g., vaginal route, sublingual route, oral route), transdermal route, ocular route, nasal route, transnasal route, or bronchopulmonary route. Preferably, the drug and composition are intended to be administered by an intestinal route, preferably rectally or orally. More preferably, they are intended to be administered orally.

[0102] Therefore, preferably, the pharmaceuticals and compositions are oral or rectal compositions, respectively, for the oral or rectal administration of the active ingredient. More preferably, the pharmaceuticals and compositions are oral compositions, particularly for the oral administration of the active ingredient.

[0103] Preferably, the disease treated in this disclosure is diabetes mellitus, preferably insulin-dependent diabetes mellitus, and more preferably type 1 diabetes mellitus and / or gestational diabetes mellitus.

[0104] Preferably, the compositions according to this disclosure are for organisms suffering from diabetes, preferably insulin-dependent diabetes, more preferably type 1 diabetes and / or gestational diabetes. Preferably, the organism is a human or animal, preferably a mammal, more preferably a human.

[0105] Preferably, the compositions according to this disclosure are compositions intended to be administered by an epithelial route. These are preferably oral or rectal compositions, more preferably oral compositions, i.e., compositions intended to be taken orally. These may be compositions intended to be administered as is, for example, in a dosage form containing an active ingredient, or compositions useful for preparing compositions intended to be administered as is, for example, in a dosage form. These are preferably intended to be administered to humans or animals, preferably mammals, and more preferably humans. Preferably, the compositions containing an active ingredient according to this disclosure are pharmaceuticals, or food compositions, or nutritional supplement compositions, or cosmetic compositions.

[0106] Other ingredients The compositions according to this disclosure may contain other components, insofar as they do not interfere with the desired properties of the composition, particularly from the viewpoint of efficiency and safety. If the composition is a dosage form, these further components typically depend on the final Galenic form. Non-limiting examples of such other components include binders and fillers (e.g., lactose, microcrystalline cellulose, mannitol), (super)disintegrants (e.g., sodium starch glycolate, crospovidone, croscarmellose), minerals, granulators (e.g., polyvinylpyrrolidone, cellulose derivatives, acacia gum, dextrose, gelatin, maltodextrin, starch, starch derivatives, tragacanth gum), flavoring agents, colorants, flow accelerators (e.g., silicon dioxide), anti-sticking agents (talc), lubricants (e.g., magnesium stearate), solvents (preferably water), buffers, and other permeation accelerators. However, since it is not necessary to obtain the desired effects in this disclosure, the compositions according to this disclosure preferably do not contain further permeation accelerators.

[0107] Therefore, the present invention also, -Cold water insoluble cross-linked dextrin, preferably cold water insoluble cross-linked maltodextrin, A fatty acid having 8 to 17 carbon atoms, preferably sodium decanoate, -Optionally, the active ingredient and, - Relating to compositions comprising other components of optional choice.

[0108] Preferably, the cold water-insoluble cross-linked dextrin is as described above. Preferably, the fatty acid having 8 to 17 carbon atoms is as described above. Preferably, the active ingredient is as described above. This is typically filled into the cold water-insoluble cross-linked dextrin. Preferably, the further components are as described above. Preferably, the amounts of the components are as described above.

[0109] Method for preparing the composition according to this disclosure The present invention also relates to a method for preparing compositions according to the present disclosure, comprising contacting a cold water-insoluble crosslinked dextrin with a fatty acid having 8 to 17 carbon atoms. The present invention also relates to compositions obtained or that can be obtained by such method.

[0110] Preferably, the composition is as described above. Preferably, the cold water-insoluble crosslinked dextrin is as described above. Preferably, the fat having 8 to 17 carbon atoms is as described above.

[0111] If the composition contains an active ingredient, the method may advantageously include the step of filling (or “associating”) the active ingredient into the cold water-insoluble crosslinked dextrin. To carry out the filling, the cold water-insoluble crosslinked dextrin according to the Disclosure may be used in a liquid, solid, or semi-solid state. For example, the cold water-insoluble crosslinked dextrin may be mixed with a small amount of water to obtain a gel. This gel may then be mixed with the active ingredient to be filled by kneading and / or mixing, the active ingredient being in powder form or dissolved in a suitable solvent. Alternatively, a select amount of cold water-insoluble crosslinked dextrin may be added together with an excess of a guest active ingredient dissolved in a suitable solvent, and then stirred overnight at room temperature to obtain filled cold water-insoluble crosslinked dextrin. The filled cold water-insoluble crosslinked dextrin can be recovered by filtration under vacuum.

[0112] Next, the filled, cold-water-insoluble cross-linked dextrin can be blended with fatty acids.

[0113] Use of a combination of cold water-insoluble cross-linked dextrin and fatty acids having 8 to 17 carbon atoms. The present invention also, -Cold water insoluble cross-linked dextrin, - The use of a combination with fatty acids having 8 to 17 carbon atoms, The present invention relates to uses for increasing epithelial permeability of the active ingredient, preferably for increasing intestinal permeability of the active ingredient, and / or for epithelial delivery of the active ingredient, preferably for oral or rectal delivery of the active ingredient, preferably for oral delivery of the active ingredient.

[0114] The present invention also relates to a method for increasing epithelial permeability of an active ingredient and / or a method for epithelial delivery of an active ingredient, comprising administering a combination of a cold water-insoluble cross-linked dextrin and a fatty acid having 8 to 17 carbon atoms.

[0115] Preferably, the cold water-insoluble cross-linked dextrin is as described above. Preferably, the fatty acid having 8 to 17 carbon atoms is as described above. Preferably, the active ingredient is as described above. This is preferably a biopharmaceutical, more preferably a protein, and even more preferably insulin. Preferably, the amount and / or ratio of the cold water-insoluble cross-linked dextrin and / or fatty acid having 8 to 17 carbon atoms in the combination is as described above for the composition according to the present disclosure.

[0116] The term "increases epithelial permeability" classically means that the combination can increase the passage of the active ingredient from the apical side of the epithelium to the basolateral side of the epithelium. In other words, this means that the combination can increase the cross-section of the active ingredient through the epithelium. The term "increases intestinal permeability" classically means that the combination can increase the passage of the active ingredient from the intestinal lumen to the blood compartment. This ability to increase intestinal permeability can be assessed by comparing the permeability of the active ingredient with the permeability of the active ingredient in the presence of the claimed permeability enhancer. This can be determined, for example, by a Caco-2 cell permeability assay, in which the absorption of the active ingredient is measured. This can be determined according to the detailed protocols shown in the Examples section below.

[0117] Preferably, the combination does not impair the integrity of the cell barrier. This cell barrier integrity can be assessed by those skilled in the art by a Caco-2 cell permeability assay in which Lucifer Yellow absorption is measured after exposure to the combination being tested. This can be assessed according to the protocol shown in the following Examples section of this specification ("Lucifer Yellow Test"). According to this test, the percentage of absorbed Lucifer Yellow is 2.0% dry weight or less, more preferably 1.5% or less, and even more preferably 0.7% or less. The integrity of this membrane can be assessed by those skilled in the art by a well-known TEER measurement, for example, according to the protocol shown in the following Examples section of this specification ("TEER Measurement").

[0118] The combination may be used by administering cold water-insoluble cross-linked dextrin, a fatty acid having 8 to 17 carbon atoms, and an active ingredient independently or as a composition. Preferably, the active ingredient is filled into the cold water-insoluble cross-linked dextrin. Therefore, the method / use advantageously includes the step of filling (or “associating”) the active ingredient into the cold water-insoluble cross-linked dextrin.

[0119] Preferably, the combination is used in the form of the composition according to this disclosure, preferably as described above. This is preferably a composition comprising the active ingredients described above.

[0120] Therefore, preferably, the present invention is -Cold water insoluble cross-linked dextrin, - Fatty acids having 8 to 17 carbon atoms, -A composition containing an active ingredient, The cold water-insoluble cross-linked dextrin is filled with the active ingredient. The present invention relates to the use of the active ingredient for increasing epithelial permeability (preferably for increasing intestinal permeability) and / or for epithelial delivery of the active ingredient (preferably for oral or rectal delivery, more preferably for oral delivery).

[0121] Therefore, preferably, the present invention also provides a method for increasing epithelial permeability of an active ingredient and / or a method for epithelial delivery of an active ingredient to an organism that requires it. -Cold water insoluble cross-linked dextrin, - Fatty acids having 8 to 17 carbon atoms, - This includes administering a composition containing the active ingredient, The present invention relates to a method in which the cold water-insoluble cross-linked dextrin is filled with the active ingredient.

[0122] In this disclosure, the amount of an ingredient may be expressed as a weight percentage. Unless otherwise specified, these weights are the raw amount of the ingredient in powder or oil form. Powdered ingredients generally contain small amounts of water (also referred to as moisture % or "loss on drying") and / or small amounts of impurities. Conversely, where dry weight is referred to in this disclosure, this well refers to the anhydrous weight.

[0123] Other features and advantages of the present invention will be clearly understood by reading the examples provided later in this specification, which illustrate but do not limit the present invention. [Examples]

[0124] 1. Evaluation of intestinal permeability of active ingredients using cold water-insoluble cross-linked dextrin (nanosponge) in combination with various permeability enhancers. First, the inventors screened various permeability enhancers in combination with cold water-insoluble cross-linked dextrin nanoparticles (hereinafter referred to as "nanosponges"). Insulin was selected as the active ingredient.

[0125] 1.1. Test Materials Insulin permeability was assayed for the different samples mentioned below (Table 1).

[0126] [Table 1]

[0127] The nanosponge was composed of cold water-insoluble pea maltodextrin (pea maltodextrin KLEPTOSE® Linecaps, ROQUETTE) crosslinked with sodium trimetaphosphate, having a DE of 17 and an Mw of 12000 Da, obtained according to International Publication No. 2021 / 254662 (A1), Example 2 (pages 13, line 8 to page 14, line 2) (the latter being incorporated herein by reference).

[0128] The permeation enhancers tested were sodium decanoate, sodium salcaprozate (SNAC), lauroyl-L-carnitine, nonaethylene glycol monododecyl ether, polyethylene glycol 3000 50% (w / v) solution, SiO2 silica 20 μm, SiO2 silica 150 nm, kaolinite, and montmorillonite.

[0129] Insulin used in all experiments was bovine insulin (Sigma / I5500).

[0130] The nanosponge suspension was prepared as follows. A suspension is prepared by stirring at room temperature, starting with crude powder of cold water-insoluble cross-linked dextrin in distilled water at a concentration of -10 mg / mL. - Disperse the suspension at 24,000 rpm for 10 minutes using a high-shear homogenizer (Ultraturrax®, IKA, Konigswinter, Germany). - Further size reduction is achieved by using high-pressure homogenization with a back pressure of 500 bar for 90 minutes using an EmulsiFlex C5 instrument (Avastin, USA). - The homogenized nano-suspension is purified by dialysis (Spectrapore, cellulose membrane, cutoff 12000 Da) to remove any potentially present synthetic residues. Store the nanosuspension at -4°C.

[0131] Insulin-filled nanosponges were prepared as follows: Using insulin powder, a 2 mg / mL solution was prepared in distilled water adjusted to pH 2.3 using phosphoric acid. The insulin solution was added to the nanosponge suspension prepared as described above in a weight ratio of insulin solution to nanosponge suspension of 1:5. The mixture was stirred at room temperature for 30 minutes and then centrifuged. The supernatant was separated from the precipitate, and the precipitate was collected and freeze-dried. The filling volume of the nanosponges was 14 ± 1%, and this percentage is expressed as the dry weight of insulin relative to the total dry weight of the nanosponges.

[0132] 1.2. Intestinal permeability assay (Caco-2 assay) Insulin permeability was evaluated by the Caco-2 assay using a 24-well Readycell Caco-2 plate (CacoReady 24 Transwell (Costar)-KRECECCR01). Briefly (Figure 1), samples were prepared at the desired concentration in Caco-2 buffer A (Hanks' Balanced Salt solution (HBSS) + 5 mM 2-(N-morpholino)ethanesulfonic acid (MES) pH 6.5), and 250 μL of each sample was placed in the apical chamber of the Caco-2 plate. 750 μL of buffer B HBSS-HEPES (4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid) (pH 7.4) was added to the bottom chamber. The Caco-2 plate was incubated at 37°C in 5% CO2, and insulin permeability was evaluated by measuring the amount of insulin that reached the bottom chamber. All materials were tested at non-cytotoxic concentrations, and all samples had an insulin concentration of 7.5 UI / mL.

[0133] The detailed protocol is as follows:

[0134] First, the following stock solution was prepared. - Free insulin stock solution: Insulin was solubilized in H2O / HCl pH2 at a rate of 2 mg dry weight / mL. - Insulin-filled nanosponge stock solution (immediate preparation): 15 mg of lyophilized insulin-filled nanosponge (equivalent to 2.1 mg of dry weight insulin) was added to 1 mL of autoclaved saline (water + 0.9% NaCl), and the suspension was gently dispersed until it was homogeneous from visual inspection. Thus, the stock solution contained 2.1 mg of dry weight insulin / mL, corresponding to 60 UI / mL of insulin. - Permeation enhancer stock solution: Each permeation enhancer was prepared in HBSS buffer (10-fold concentrated compared to the highest concentration tested).

[0135] Next, samples were prepared from these stock solutions in buffer A to obtain the following concentrations (weights are expressed as dry weight). - "Free insulin": 7.5 UI / mL of insulin, - "Insulin-filled nanosponge": 7.5 UI / mL insulin, - "Insulin-filled nanosponge + sodium decanoate": 7.5 UI / mL insulin + 6.5 mM, 3.25 mM, or 1.625 mM sodium decanoate (Sigma / C4151), - "Insulin-filled nanosponge + salcaprozate sodium": 7.5 IU / mL insulin + 1 mg / mL, 0.5 mg / mL, or 0.25 mg / mL salcaprozate sodium (SNAC) 203787-91-1 - "Insulin-filled nanosponge + lauroyl-L-carnitine": 7.5 UI / mL insulin + 0.25 mM, 0.125 mM, or 0.625 mM lauroyl-L-carnitine (Sigma / 39953) - "Insulin-filled nanosponge + nonaethylene glycol monododecyl ether": 7.5 UI / mL insulin + 0.01 mM, 0.05 mM, or 0.025 mM nonaethylene glycol monododecyl ether (Sigma / P9641), - "Insulin-filled nanosponge + polyethylene glycol": 7.5 UI / mL insulin + 0.2% or 0.1% or 0.05% (w / v) polyethylene glycol 3000 (Sigma / 81269), - "Insulin-filled nanosponge + SiO2 silica 20μm": 7.5 UI / mL insulin + 0.8% or 0.4% or 0.2% (w / v) SiO2 silica 20μm (Sigma / 904376) - "Insulin-filled nanosponge + SiO2 silica 150nm": 7.5 UI / mL insulin + 0.8% or 0.4% or 0.2% (w / v) SiO2 silica 150nm (Sigma / 904414), - "Insulin-filled nanosponge + kaolinite": 7.5 UI / mL insulin + 0.2%, 0.1%, or 0.05% (w / v) kaolinite (Sigma / 03584), - "Insulin-filled nanosponge + montmorillonite": 7.5 UI / mL of insulin + 0.4%, 0.2%, or 0.1% (w / v) of montmorillonite (Sigma / 69866).

[0136] For the "insulin-filled nanosponge + permeation enhancer" sample, the permeation enhancer stock solution was first diluted in buffer A, and then the insulin-filled nanosponge stock solution was added at the target concentration and gently dispersed.

[0137] Caco-2 growth medium was removed from the 24-well Readycell Caco-2 plate. Then, 250 μL of each sample, prepared in buffer A [HBSS-MES (pH 6.5)], was added to the apical chamber. 800 μL of buffer B [HBSS-HEPES (pH 7.4)] was added to the bottom chamber. 50 μL was collected and transferred to a Greiner 651201 plate for analysis (blank = 0 minutes, incubation time = t0). All assays were performed three times (n = 3).

[0138] The apical chamber was placed in the lateral-bottom chamber and incubated for 120 minutes (37°C, 5% CO2).

[0139] At t0+15, t0+30, t0+60, and t0+120 minutes, 50 μL of culture medium was collected in the bottom chamber and transferred to Greiner plate 651201 for analysis. Insulin detection was performed using ultra-high-performance liquid chromatography combined with triple quadrupole mass spectrometry (UHPLC-QqQ), which has detection limits optimized for testing with all permeabilizers. 50 μL of fresh buffer B was added to the bottom chamber. The apical chamber was returned to the bottom chamber.

[0140] The results are shown in Figures 2 to 10.

[0141] 1.3.Results By comparing both controls ("free insulin" vs. "insulin-filled nanosponge"), it was found that the nanosponge increased insulin permeability. When the nanosponge was combined with sodium decanoate, an unexpectedly high level of permeability enhancement was obtained (Figure 2). This enhancement was much greater than when the nanosponge was used alone. With other permeability enhancers, no permeability enhancement was observed when used in combination with the nanosponge (Figures 3-10). The permeability levels were similar to those obtained with free insulin and lower than those obtained with the nanosponge alone. In other words, these permeability enhancers inhibited the permeability effect of the nanosponge.

[0142] Following these favorable and surprising results, the combination of nanosponge and sodium decanoate was further evaluated to detect the ultimate synergistic effect. In fact, in this previous experimental setup, the use of the permeation enhancer alone (i.e., without nanosponge) was not tested, and therefore it was not possible to confirm whether nanosponge and sodium decanoate acted synergistically.

[0143] 2. Evaluation of synergistic effects. In this section, the combination of nanosponge and sodium decanoate was further evaluated to detect the final synergistic effect. Combinations with sodium salcaprozate (also tested in a previous experiment) or linoleic acid (a fatty acid with 18 carbon atoms) were also evaluated (Experiment 1, Table 1). Uncrosslinked dextrin was also evaluated, and different concentrations of sodium decanoate were tested (Experiment 2). In particular, the following were evaluated (Table 2).

[0144] [Table 2]

[0145] [Table 3]

[0146] 2.1. Intestinal permeability assay (Caco-2 assay) Insulin permeability was assayed as described in Section 1.2, except that the Elisa assay was used to quantify insulin (instead of UHPLC-QqQ) because it is more sensitive to the advantages of the assay.

[0147] Furthermore, since the experiments were conducted in different laboratories, the compositions of buffers A and B were slightly different (this should not have significantly affected the results obtained). -Buffer A:HBSS (Gibco 14025-050) + 5mM MES (Sigma Aldrich M2933, lot SLCH7805) pH 6.5 + 10 μg / mL trypsin inhibitor (T9003, Sigma Aldrich lot SLCG7982) + 1 μg / mL leupeptin inhibitor (Sigma L9783, lot 147407) - pH 6.5. - Buffer B: HBSS + 10mM Hepes (Gibco 15630-080) pH 7.4 + 10 μg / mL trypsin inhibitor (T9003, Sigma Aldrich lot SLCG7982) + 1 μg / mL leupeptin inhibitor (Sigma L9783, lot 147407) - pH 7.4.

[0148] For samples containing empty nanosponges, samples were prepared from a raw suspension of nanosponges containing 15 mg / mL of nanosponges prepared in Buffer A. To prepare samples containing dextrin and insulin, powdered dextrin and insulin were first blended to achieve final association or filling of insulin. The powdered blend was then prepared either directly in Buffer A (for the "insulin-dextrin blend" sample) or in a permeabilis enhancer prepared in Buffer A (for the "insulin-dextrin blend + sodium decanoate" sample), and then gently dispersed. In all sample preparations, insulin (whether filled or not) was always the last substance added and then gently dispersed. The final concentration of insulin was always 7.5 UI / mL.

[0149] Prior to the Caco-2 assay, cell barrier integrity was also evaluated by measuring TEER (Transpeithelial electrical resistance) to confirm the integrity of the cell barrier used in the assay. For Experiment 2, the barrier integrity after exposure to the tested sample was further assayed by measuring the passage of Lucifer Yellow after the assay to verify the effect of the tested sample on barrier integrity and identify the ideal concentration for in vitro use. According to this test, the percentage of absorbed Lucifer Yellow is preferably 2.0% dry weight or less, more preferably 1.5% or less, and even more preferably 0.7% or less.

[0150] TEER measurement Before the experiment, barrier integrity was measured using Millicell ERS-2 (Merck Millipore MERS00002) electrodes. Briefly, the electrodes were sterilized in a 70% ethanol solution. They were then equilibrated in pre-warmed culture medium at room temperature (RT). Since TEER measurements are performed at RT, cells were removed from the incubator 20 minutes prior to the experiment.

[0151] Lucifer Yellow Test After sampling the supernatant last, the apical and basal compartments were washed with HBSS medium. Lucifer Yellow was added to the apical compartment. After incubation at 37°C for 1 hour, the basal supernatant was collected, and fluorescence was measured using Spectramax (Molecular Devices) with a 485 nm wavelength laser, with readings taken at 527 nm. As a standard, a dilution series of Lucifer Yellow ranging from 100 μM to 0.1 μM was prepared.

[0152] ELISA dose Samples at t0, t0+15 min, t0+30 min, t0+60 min, and t0+120 min were analyzed using an insulin ELISA kit (Sigma Aldrich-RAB0568) according to the manufacturer's recommendations. Briefly, samples were diluted in buffer and immunoassay was performed. After steps with different washing and incubation times, absorbance was read at 450 nm using a Spectramax (Molecular Devices) spectrophotometer.

[0153] The results of the permeability tests in Experiment 1 and Experiment 2 are shown in Figures 11 and 12, respectively. The results of the barrier integrity test (Yellow Lucifer test) in Experiment 2 are shown in Figure 13.

[0154] 2.2.Results Results of Experiment 1 (Figure 11) By comparing the results obtained with the "insulin-filled nanosponge" sample with those obtained with the "free insulin + nanosponge" and "free insulin" samples, it can be concluded that the nanosponge can significantly improve insulin passage when insulin is filled into it. Conversely, free insulin cannot cross the cell barrier. When the nanosponge is used in combination with sodium decanoate, a synergistic effect is observed. Insulin passage is much higher than that obtained with sodium decanoate alone, and much higher than that obtained with the nanosponge alone (whether filled or not). More specifically, the insulin passage obtained with the "insulin-filled nanosponge" was 67 μU / mL, while the insulin passage obtained with "insulin + sodium decanoate 6.5 mM" was 34 μU / mL. Therefore, the expected insulin passage for "insulin-filled nanosponge + sodium decanoate 6.5 mM" should be 67 + 34 = 101 μU / mL. However, the actual passage obtained was 3907 μU / mL, which is almost 40 times higher than the value expected by a simple additive effect.

[0155] This synergistic effect is not observed when nanosponges are combined with linoleic acid or sodium salcaprozate. On the contrary, the addition of these permeability enhancers to insulin-filled nanosponges appears to be detrimental to insulin permeation.

[0156] For all groups tested, cell barrier integrity was measured according to the CaCo-2 plate manufacturer's reference (TEER: 2347, 73Ω.cm²). 2 They agreed on the matter (data not shown).

[0157] Results of Experiment 2 (Figures 12 and 13) In this case as well, a synergistic effect can be observed when insulin-filled nanosponges are used in combination with sodium decanoate (Figure 12). This effect can be seen at all concentrations tested and is dose-dependent. No synergistic effect occurs when uncrosslinked dextrin is used.

[0158] Note: The values ​​obtained in Experiment 2 are not of the same magnitude as those obtained in Experiment 1. This is because the obtained values ​​can vary significantly from one experimental set to the next. This means that values ​​obtained in one experimental set can be compared with each other, but values ​​obtained in different experimental sets cannot. This is why controls and comparisons ("culture medium", "free insulin", etc.) were repeated in Experiment 2.

[0159] Results obtained using the Lucifer Yellow test (Figure 13) suggest that concentrations of sodium decanoate lower than 6.5 mM should be preferable for in vitro assays. Indeed, when using 6.5 mM sodium decanoate with insulin-filled nanosponges, the pass-through rate of Lucifer Yellow was 1.49%, which is higher than the upper limit of the ideal standard value of 0.7% (the value indicated in the user manual by the manufacturer of the 24-well Readycell Caco-2 plate). However, it should be noted that this percentage of 1.45% is still very low, and the 6.5 mM concentration has been found to be non-cytotoxic in previous experiments (data not shown). Therefore, this change in membrane integrity is likely to be very limited and reversible. When using lower concentrations of sodium decanoate, barrier integrity was not compromised at all.

[0160] For all groups tested, cell barrier integrity was measured according to the CaCo-2 plate manufacturer's reference (TEER: 2347, 73Ω.cm²). 2 They agreed on (data not shown).

Claims

1. - Cold water insoluble cross-linked dextrin, A composition comprising a fatty acid having 8 to 17 carbon atoms.

2. The composition according to claim 1, wherein the dextrin is selected from pyrodextrin, maltodextrin, cyclodextrin, or a mixture thereof.

3. The composition according to claim 1 or 2, wherein the cold water-insoluble crosslinked dextrin can be obtained by reacting dextrin with a crosslinking agent, the crosslinking agent being selected from trimetaphosphate, dicarboxylic acid, dianhydride, carbonyldiimidazole, diphenyl carbonate, triphosgene, acyl dichloride, diisocyanate, diepoxide, or any mixture thereof.

4. The composition according to any one of claims 1 to 3, wherein the cold water-insoluble crosslinked dextrin can be obtained by reacting dextrin with a crosslinking agent, and the molar ratio of the crosslinking agent to the dextrin with respect to anhydrous glucose units is 0.1:1 or greater.

5. The composition according to any one of claims 1 to 4, wherein the fatty acid has at least nine carbon atoms.

6. The composition according to any one of claims 1 to 5, wherein the fatty acid has a maximum of 16 carbon atoms, preferably a maximum of 15 carbon atoms, more preferably a maximum of 14 carbon atoms, more preferably a maximum of 13 carbon atoms, more preferably a maximum of 12 carbon atoms, and more preferably a maximum of 11 carbon atoms.

7. The composition according to any one of claims 1 to 6, wherein the fatty acid has 10 carbon atoms.

8. The composition according to any one of claims 1 to 7, wherein the fatty acid is in the form of a salt.

9. The composition according to claim 8, wherein the fatty acid is in the form of a sodium salt.

10. The composition according to claim 9, wherein the fatty acid is sodium decanoate.

11. The composition according to any one of claims 1 to 10, wherein the cold water-insoluble crosslinked dextrin is in the form of particles.

12. The composition according to any one of claims 1 to 11, wherein the composition is intended to be administered via an epithelial pathway.

13. The composition according to any one of claims 1 to 12, wherein the composition is an oral or rectal composition.

14. The composition according to any one of claims 1 to 13, further comprising an active ingredient.

15. The composition according to claim 14, wherein the active ingredient is selected from a BCS class III drug, a BCS class IV drug, a biopharmaceutical, or any mixture thereof.

16. The composition according to claim 14 or 15, wherein the active ingredient is selected from proteins.

17. The composition according to any one of claims 14 to 16, wherein the active ingredient is filled in the cold water-insoluble crosslinked dextrin.

18. A composition according to any one of claims 14 to 17 for use as a pharmaceutical.

19. Non-medicinal use of a composition according to any one of claims 14 to 17 in a food, nutritional supplement, or cosmetic composition.

20. A method for preparing the composition according to any one of claims 1 to 18, comprising contacting the cold water-insoluble crosslinked dextrin with the fatty acid having 8 to 17 carbon atoms.

21. - Cold water insoluble cross-linked dextrin, - The use of a combination with fatty acids having 8 to 17 carbon atoms, Use to increase epithelial permeability of the active ingredient, preferably to increase intestinal permeability of the active ingredient.

22. - Cold water insoluble cross-linked dextrin, - The use of a combination with fatty acids having 8 to 17 carbon atoms, Use for epithelial delivery of the active ingredient, preferably for oral or rectal delivery of the active ingredient, more preferably for oral delivery of the active ingredient.

23. The use according to claim 21 or 22, wherein the active ingredient is selected from a BCS class III drug, a BCS class IV drug, a biopharmaceutical, or any mixture thereof.

24. The use according to any one of claims 21 to 23, wherein the active ingredient is selected from proteins.

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