Application of sodium aescinate or isomer thereof as penetration enhancer

Abstract

The invention discloses application of sodium aescinate or an isomer thereof as a novel penetration enhancer, and belongs to the technical field of pharmaceutical preparations. Sodium aescinate or an isomer thereof is used as a penetration enhancer, and by opening a close connection structure between cells, the penetration of a drug through a cell bypass is promoted, and the absorption efficiency of the drug is improved, so that the bioavailability of a mucous membrane administration preparation is further improved, especially the compatibility of low-permeability drugs. Compared with a traditional penetration enhancer, the sodium aescinate penetration enhancer is non-toxic and non-irritant, can quickly recover to the tight connectivity between mucous membrane tissue cells before administration, does not cause damage to mucous membrane tissues of an absorption part, shows excellent safety, and has a very wide application prospect.

Description

Technical Field

[0001] This invention relates to the field of pharmaceutical formulation technology, specifically to the application of sodium aescinate or its isomers as a penetration enhancer. Background Technology

[0002] Mucosal administration is a major non-invasive route of drug delivery, including routes via the oral mucosa, nasal mucosa, ocular mucosa, pulmonary mucosa, and rectal mucosa. However, due to the tight junctions between epithelial cells in biological mucosa, transmembrane delivery of drugs, especially BCS Class III and IV drugs and large molecule drugs such as peptides, is hindered, resulting in generally low bioavailability of mucosal drugs (Maher S, Mrsny RJ, Brayden DJ. Intestinal permeation enhancers for oral peptide delivery [J]. Adv Drug Deliv Rev, 2016,106: 277-319.). Tight junctions, also known as locked bands, are dense regions formed by the connection of adjacent cells in a biological membrane via cations and proteins. They are a type of tightly packed and impermeable junction that plays a crucial role in regulating drug transport via the cellular bypass pathway. Most large molecules cannot pass through them; only water and some small molecules can permeate through the pores at the junctions (Fasano A, Nataro JP. Intestinal epithelial tight junctions as targets for enteric bacteria-derived toxins [J]. Adv Drug Deliv Rev, 2004, 56: 795-807.). Furthermore, physiological barriers in brain tissue, such as the blood-brain barrier, are also protective structures formed by tight junctions between endothelial cells. These tight junctions protect brain tissue from damage by foreign substances while inhibiting drug accumulation in the brain. In recent years, researchers have made numerous attempts to address the problem of drugs' inability to penetrate tight junctions, such as preparing drugs into nano-formulations, microneedles, and other high-end formulations; modifying drugs to introduce hydrophobic functional groups to increase their lipophilicity; encapsulating drugs with positively charged excipients such as chitosan; and adding penetration enhancers to act on the tight junctions of mucosa. Studies have shown that these nano-formulations have not been widely used in practical applications due to their low drug loading capacity, complex preparation processes, and high requirements for scale-up production technology. The simplest and most effective way to solve the problem of low bioavailability of mucosal drug delivery is to use penetration enhancers. Therefore, the addition of penetration enhancers remains the main way to promote penetration and increase bioavailability at present (Ghadiri M, Young PM, Traini D. Strategies to enhance drug absorption via nasal and pulmonary routes [J]. Pharmaceutics, 2019, 11: 113.).

[0003] Permeation enhancers, also known as transdermal penetration enhancers, are a class of functional excipients in pharmaceutical formulations that improve the transmembrane absorption of active drugs, thereby increasing bioavailability and maximizing the therapeutic effect. There are many types of permeation enhancers. Commonly used traditional permeation enhancers include cyclodextrins, bile salts, fatty acids, sulfoxides, metal ion chelating agents, surfactants, protease inhibitors, and chitosans. Newer permeation enhancers include maltodextrins and polyethylene glycol dodecyl hydroxystearate (Solutol® HS15).Traditional penetration enhancers generally have toxicity issues. For example, dimethyl sulfoxide (DMSO) can cause erythema, blisters, and irreversible damage to the skin (Pathan IB, Setty CM. Chemical Penetration Enhancers for Transdermal Drug Delivery Systems[J]. Trop J PharmRes, 2009, 8(2): 172-179.); polysorbate and polyoxyethylene can cause dissolution of cell membrane structures and components during mucosal permeation (Dimitrijevic D, Shaw AJ, Florence A T. Effects of some non-ionic surfactants on transepithelial permeability in Caco-2 cells[J]. J PharmPharmacol, 2010, 52(2):157-162.); sodium dodecyl sulfate can cause irreversible damage to the mucosa (Lin PY, Chuang EY, Chiu YH, et al. Safety and efficacy of self-assembling bubblecarriers stabilized with sodium dodecyl sulfate for oral delivery of therapeutic proteins[J]. J Control Release, 2017, 259: 168-175.); Sodium laurate can induce apoptosis (Aungst B J. Absorption enhancers: applications and advances[J].The AAPS journal, 2012, 14(1): 10-18.); Excessive bile salt concentration can cause local irritation and cytotoxicity (Warnken ZN, Smyth HDC, Watts AB, et al. Formulation and device design to increase nose to brain drug delivery[J]. J Drug Deliv Sci Tec,2016, 35: 213-222.). Therefore, safety concerns limit the application of traditional penetration enhancers in pharmaceutical formulations.

[0004] Alkyl glycosides are a novel type of penetration enhancer, possessing advantages such as high efficiency, low toxicity, antibacterial properties, and biodegradability. They are already used in marketed drugs such as sumatriptan nasal spray (TOSYMRA) and diazepam nasal spray (VALTOCO). However, studies have shown that alkyl glycosides can cause mucosal damage with increasing carbon chain length and concentration (Maher S, Geoghhegan C, Brayden DJ. Intestinal permeation enhancers to improve oral bioavailability of macromolecules: reasons for low efficacy in humans [J]. Exp Opin Drug Deliv, 2021, 18: 273-300.). Furthermore, the limited variety of novel penetration enhancers cannot meet the diverse drug selection needs. Therefore, exploring a safe and efficient novel mucosal penetration enhancer remains extremely urgent. Summary of the Invention

[0005] Sodium aescinate (SA) is a triterpenoid saponin sodium salt with ester bonds, isolated and extracted from the mature seeds of *Castanopsis fargesii*, a plant in the Aesculaceae family. Its CAS number is 20977-05-3, and its structural formula is as follows:

[0006]

[0007] SA structure

[0008] SA has been clinically proven to have anti-exudative, anti-inflammatory, blood-stasis-reducing, and swelling-reducing effects. Its injections are commonly used in clinical practice to treat cerebral hemorrhage, cerebral infarction, and traumatic brain injury. To date, SA liniments, lyophilized powder injections, tablets, and compound SA gels have been developed, and their safety has been extensively verified.

[0009] According to the theory of biopharmaceutics, drugs can be classified into highly permeable and low-permeability drugs based on their permeability. The permeation efficiency of a drug into mucosal tissues determines its bioavailability. To improve the permeation effect of low-permeability drugs on mucosa, the inventors experimented with applying different types of permeation enhancers to these drugs. Through continuous screening and exploration, the inventors surprisingly discovered that sodium aescinate (SA) not only improves the permeation efficiency of low-permeability drugs on mucosa but is also non-toxic, non-irritating, and can quickly restore the tight junctions between mucosal tissue cells to their pre-drug-administered state, without causing damage to the mucosa at the absorption site, exhibiting excellent safety. Notably, the inventors clarified the mechanism of action of sodium aescinate as a permeabilizer: it promotes drug absorption via a cellular bypass pathway by opening the tight junctions between mucosal cells, particularly for low-permeability drugs or drugs requiring improved bioavailability.

[0010] Therefore, the purpose of this invention is to provide a novel, safe, and efficient SA penetration enhancer that can improve the bioavailability of transmucosal drug delivery formulations, especially for low-permeability drugs.

[0011] To achieve the above objectives, the technical solution adopted by the present invention is as follows:

[0012] In one aspect, the present invention provides the use of sodium aescinate or its isomers as a penetration enhancer.

[0013] In this invention, the penetration enhancer is used as a penetration enhancer for low-permeability drugs.

[0014] In this invention, the hypotonic drugs include BCS III and / or BCSIV classes in the biopharmaceutics classification system.

[0015] In this invention, the hypotonic drug includes small molecule drugs and / or large molecule drugs;

[0016] The small molecule drugs mentioned include glycopyrronium bromide, terbutaline, famotidine, bromocriptine, diazepam, budesonide, mometasone furoate, mannitol, fluticasone propionate, beclomethasone propionate, cisasonide, formoterol fumarate, dihydroergotamine mesylate, tiotropium bromide, nadolol, sodium valproate, ketamine, gabapentin, ketorolac tromethamine, metoprimidine, lisinopril, butorphanol tartrate, olopatadine hydrochloride, azelastine hydrochloride, oxcarbazepine, chlorothiazide, palonosetron, and zolmitrol. The following are listed as one or more of the following: naloxone hydrochloride, lorazepam, quetiapine fumarate, betahistine, carbamazepine, nalmefen hydrochloride, salmon calcitonin, levetiracetam, enalapril, alprazolam, xylometazoline hydrochloride, phosphatidylcholine, phenytoin sodium, indacaterol, umemetamine, polyethylene glycol 400, phenobarbital, salbutamol sulfate, ipratropium bromide, sulpiride, buspirone hydrochloride, ambroxol hydrochloride, olanzapine, acyclovir, lamotrigine, bromhexine hydrochloride, oxazepam, sumatriptan, and remimazolam.

[0017] The macromolecular drugs include one or more of the following: proteins, peptides, antibody-drug conjugates, peptide-drug conjugates, nucleic acid-drug conjugates, protein-drug conjugates, deoxyribonucleic acid, and ribonucleic acid.

[0018] In this invention, at least one of the sodium aescinate or its isomers is prepared with a pharmaceutically acceptable carrier into a clinically acceptable pharmaceutical formulation.

[0019] In some specific embodiments of the present invention, the pharmaceutical preparation is a mucosal delivery preparation;

[0020] In this invention, the mucosal drug delivery formulations include, but are not limited to, nasal mucosal drug delivery formulations, rectal mucosal drug delivery formulations, oral mucosal drug delivery formulations, pulmonary mucosal drug delivery formulations, and ocular mucosal drug delivery formulations.

[0021] In some specific embodiments of the present invention, the mucosal delivery preparation is a pulmonary mucosal delivery preparation.

[0022] In this invention, the pulmonary mucosal drug delivery formulation includes, but is not limited to, suspensions, solutions, emulsions, microspheres, liposomes, or powder formulations.

[0023] In some specific embodiments of the present invention, the mucosal delivery preparation is a nasal mucosal preparation.

[0024] In this invention, the nasal mucosal drug delivery formulation includes, but is not limited to, suspensions, solutions, emulsions, microspheres, liposomes, or powder formulations.

[0025] In this invention, the aescin or its isomers are used as permeation enhancers. Their mechanism of action is to promote drug absorption via a cellular bypass pathway by opening the tight junctions between mucosal cells, thereby further improving the bioavailability of mucosal drug delivery formulations, especially for formulations targeting low-permeability drugs.

[0026] In this invention, the penetration enhancer can be formulated into a pharmaceutical preparation with low-permeability drug components and other excipients, or the penetration enhancer can be directly formulated into a single-ingredient preparation for use in combination with other drugs.

[0027] In one aspect, the present invention provides a method for promoting drug penetration by simultaneously using sodium aescinate and / or its isomers when using a hypopermeable drug. For example, the hypopermeable drug can be used by simply mixing it with sodium aescinate and / or its isomers, or they can be formulated together for use.

[0028] In one aspect, the present invention provides a pharmaceutical composition for mucosal administration, comprising a low-permeability drug, sodium aescinate and / or its isomers.

[0029] The beneficial effects of this invention are as follows:

[0030] 1. This invention provides a novel, safe, and efficient penetration enhancer, sodium aescinate. This penetration enhancer can not only improve the penetration efficiency of low-permeability drugs on mucous membranes, but is also non-toxic, non-irritating, and can quickly restore the tight connection between mucosal tissue cells to the state before drug administration, without causing damage to the mucous membrane at the absorption site, thus exhibiting excellent safety.

[0031] 2. The mechanism of action of sodium aescinate permeation enhancer is to promote drug absorption through the bypass pathway by opening the tight junction structure between mucosal cells, thereby improving the bioavailability of the drug, and it has broad application prospects.

[0032] 3. The drug composition for mucosal administration provided by the present invention can improve the mucosal permeability of the low-permeability drug contained therein. Attached Figure Description

[0033] Figure 1 This refers to the regulatory effect of SA on the transmembrane resistance of different mucosal cell tissues in Embodiment 1 of the present invention;

[0034] Figure 2 This is the immunofluorescence of the tightly linked marker proteins Occludin and Claudin in Example 2 of this invention;

[0035] Figure 3 This describes the regulatory effect of SA, dodecyl maltodextrin, and tetradecyl maltodextrin on transmembrane resistance in the MDCK cell mucosal model in Example 3 of this invention.

[0036] Figure 4 This illustrates the permeation regulation effect of SA on zero-permeability drugs in different mucosal tissues in Example 4 of this invention.

[0037] Figure 5 This illustrates the permeation regulation effect of SA on hypotonic drugs in different mucosal tissues in Example 5 of the present invention. Detailed Implementation

[0038] The following detailed embodiments further illustrate the above-described content of the present invention. However, this should not be construed as limiting the scope of the present invention to the following embodiments. All technologies implemented based on the above-described content of the present invention fall within the scope of the present invention. Where specific techniques or conditions are not specified in the embodiments, they shall be performed in accordance with the techniques or conditions described in the literature in the field or according to the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased commercially.

[0039] In this invention, the "penetration enhancer" is a class of excipients that can reversibly and temporarily alter the physicochemical properties of biological barriers (such as skin or mucous membranes), thereby significantly reducing their barrier capacity and improving drug penetration and absorption efficiency. They typically do not exert pharmacological effects themselves, but rather function through mechanisms such as dissolving lipid interstitial spaces, liquefying cellular lipids, or forming drug reservoirs. Acting as "molecular keys," they help active pharmaceutical ingredients (APIs), especially large molecules or low-permeability drugs that are difficult to absorb, to more effectively cross biological barriers, ultimately achieving enhanced efficacy, reduced dosage, or delivery to specific sites.

[0040] In this invention, the sodium aescinate is a mixture containing four isomers, namely sodium aescinate A (CAS No.: 123748-68-5), sodium aescinate B (CAS No.: 26339-90-2), sodium aescinate C (CAS No.: 219944-39-5), and sodium aescinate D (CAS No.: 219944-46-4).

[0041] In some specific embodiments of the present invention, the penetration enhancer is one or more of sodium aescinate or its isomers.

[0042] In this invention, "low-permeability drugs" refer to drugs with poor self-permeability and difficulty in crossing biological barriers (such as the intestinal mucosa, blood-brain barrier, stratum corneum of skin, or cell membrane). These drugs typically have characteristics such as large molecular weight, low lipid solubility, or excessive polarity, making them unable to be effectively absorbed through the lipid bilayer of cells (transcellular pathway) or tight junctions (bypass pathway), resulting in low oral bioavailability or poor local administration efficacy. In the Biopharmaceutics Classification System (BCS), they mostly belong to Class III (high solubility, low permeability) or Class IV (low solubility, low permeability) drugs. To solve their delivery challenges, drug delivery technologies such as permeation enhancers, liposomes, and nanoparticles are often used to assist their absorption.

[0043] In the Biopharmaceutics Classification System (BCS), drugs are classified into four categories based on two key factors: solubility and intestinal permeability: BCS Class I (hyperosmolar and highly soluble), BCS Class II (hyperosmolar and poorly soluble), BCS Class III (hypoosmolar and highly soluble), and BCS Class IV (hypoosmolar and poorly soluble). In this invention, the hypoosmolar drugs include those in BCS Class III and / or BCS Class IV, such as ranitidine, famotidine, atenolol, insulin, amphotericin B, ketoconazole (oral), and paclitaxel.

[0044] In this invention, the term "pharmaceutically acceptable carrier" refers to the collective term for all additives in a pharmaceutical preparation other than the active ingredient. These carriers must be safe, non-toxic, and compatible with the active pharmaceutical ingredient. Their core function is to ensure that the drug can be formulated into a stable, effective, and easy-to-use dosage form. Common carriers include: solvents such as water for injection and ethanol, used to dissolve drugs to form injections, tinctures, or other liquid preparations; excipients such as lactose and microcrystalline cellulose, used as fillers to provide volume in tablets, capsules, etc.; binders such as starch paste and hydroxypropyl methylcellulose, used to bind powders for tableting or granulation; disintegrants such as sodium croscarmellose, which promote rapid disintegration of tablets in vivo for absorption; lubricants such as magnesium stearate, which prevent material adhesion during production; preservatives such as sodium benzoate, which inhibit microbial growth in liquid preparations; and sustained-release materials such as ethyl cellulose, used to control the drug release rate.

[0045] In this invention, the nasal mucosal preparation is used to enhance the absorption of drugs across the blood-brain barrier into brain tissue.

[0046] In this invention, the term "suspension" refers to a liquid dosage form, specifically a heterogeneous system in which an insoluble solid drug is dispersed and suspended in a liquid (such as water or oil) as fine particles (typically with a particle size of 0.5-10 micrometers). Because the solid particles will settle, the mixture needs to be shaken well before use to ensure accurate dosage each time. Examples include ibuprofen suspension (Motrin) and certain antibiotic dry suspensions.

[0047] In this invention, the "solution" is a homogeneous, clear liquid dosage form formed by completely dissolving a drug in a liquid solvent (such as water, ethanol, glycerol, or mixtures thereof) in a molecular or ionic state. Examples include oral acetaminophen solutions, eye drops, etc.

[0048] In this invention, the "emulsion" is a heterogeneous system formed by dispersing one phase as tiny droplets within the other of two immiscible liquids (usually oil and water) under the action of an emulsifier. It is classified into oil-in-water and water-in-oil types. It is typically milky white in appearance and can be used orally, topically, or by injection, simultaneously improving the bioavailability of both oil-soluble and water-soluble drugs. Examples include cod liver oil emulsions and certain fat emulsion injections.

[0049] In this invention, "microspheres" refer to tiny spherical entities formed when a drug is dissolved or dispersed in a polymer material, typically with a particle size between 1 and 300 micrometers. Microspheres can serve as a "drug reservoir," allowing for the slow release of the drug into the body over weeks or even months by controlling the degradation rate of the polymer material, thus achieving a long-lasting effect. They are commonly used for macromolecular drugs such as proteins and peptides.

[0050] In this invention, a "liposome" is a tiny, spherical vesicle whose structure consists of a phospholipid bilayer membrane surrounding an aqueous core, similar to a human cell membrane. Drugs can be encapsulated in either the aqueous core or the lipid bilayer, depending on their solubility. This structure protects drugs (especially easily degradable nucleic acid and protein drugs), reduces toxicity, and promotes drug absorption and targeted delivery by fusing with cell membranes.

[0051] In this invention, the term "powder formulation" refers to a dry powder form for inhalation administration. It consists of micronized drug particles encapsulated in capsules or blister packs. Patients inhale the powder through a dedicated inhalation device, where a deep inhalation disperses the drug powder deep into their lungs.

[0052] In this invention, RMPI2650 nasal epithelial cells, MDCK Coxsbane canine kidney cells, and Calu-3 human lung adenocarcinoma cells are commonly used to evaluate mucosal absorption models of drugs, especially absorption through the mucous membranes of the respiratory system such as the nasal cavity and lungs.

[0053] In this invention, "DAPI (4′,6-diamidinyl-2-phenylindole)" is a blue fluorescent DNA dye that exhibits approximately 20-fold fluorescence enhancement when bound to the AT region of dsDNA. DAPI is excited by a 405 nm laser and is commonly used as a nuclear counterstain in fluorescence microscopy, flow cytometry, and chromosome staining.

[0054] In this invention, the "TEER (transmembrane resistance)" provides information on particle flow resistance across a single cell layer. It is related to the integrity of the tight connections between cells and is an important indicator for evaluating the permeability of a cell mucosa model. Changes in TEER reflect the degree of tightness of cell connections. A higher TEER value indicates higher cell mucosa integrity and lower permeability, suggesting tighter connections between mucosal tissue cells. Conversely, a lower TEER value indicates lower model integrity and higher permeability, suggesting looser connections between mucosal tissue cells.

[0055] In some embodiments of the present invention, when sodium aescinate or its isomer is used as a permeation enhancer, its concentration is above 1 μg / mL.

[0056] In some embodiments of the present invention, when sodium aescinate or its isomer is used as a permeation enhancer, its concentration is above 5 μg / mL.

[0057] In some embodiments of the present invention, when sodium aescinate or its isomer is used as a permeation enhancer, its concentration is above 10 μg / mL.

[0058] In some embodiments of the present invention, when sodium aescinate or its isomer is used as a permeation enhancer, its concentration is above 20 μg / mL.

[0059] In some embodiments of the present invention, when sodium aescinate or its isomers are used as permeation enhancers, their concentration is above 100 μg / mL.

[0060] In some embodiments of the present invention, when sodium aescinate or its isomers are used as a permeation enhancer, the concentration is 100 μg / mL to 10000 μg / mL.

[0061] Information on reagents, consumables, equipment, and cells is shown in Tables 1-3:

[0062] Table 1. Information on experimental reagents

[0063]

[0064] Table 2. Information on experimental consumables and equipment

[0065]

[0066] Table 3 Cell-related information

[0067]

[0068] Example 1: Study on the effects of sodium aescinate on mucosal tissue structure

[0069] In vitro mucosal models were constructed using RPMI2650 cells, Calu-3 cells, and MDCK cells, respectively, to assess the impact of sarcopenia (SA) on mucosal tissue structure. The specific procedures are as follows:

[0070] RPMI2650, MDCK, and Calu-3 cells were respectively loaded at approximately 1.8 × 10⁻⁶. 5 / well density seeded into the Transwell chambers of a 12-well plate (1.12 cm²) 2 (3μm, polycarbonate membrane). RPMI2650 cells were continuously incubated for approximately 15 days at 37℃ and 5% CO2, Calu-3 cells for approximately 10 days, and MDCK cells for approximately 3-5 days. The TEER of the RPMI2650, Calu-3, and MDCK cell mucosal models was measured to be 250 Ω*cm. 2 1000 Ω*cm 2 and 3500 Ω*cm 2 It served as a control group (see Figure 1 (Control group). The TEER values ​​of the three cell mucosal models showed that the MDCK cell model was the most compact, followed by Calu-3 cells, while the RPMI2650 cell model had a relatively loose structure. After successfully establishing the three mucosal models, SA at concentrations of 5 μg / mL, 10 μg / mL, and 20 μg / mL were added for further incubation, and the transmembrane resistance of the mucosal tissues in each group was measured at different time points.

[0071] The test results are as follows Figure 1 As shown, after the addition of SA, the TEER values ​​of the mucosal tissues in all groups decreased with the extension of co-incubation time, indicating that SA can alter the intercellular tightness of RPMI2650, MDCK, and Calu-3 mucosal tissues and increase the intercellular space. SA significantly reduced the TEER values ​​in the MDCK and Calu-3 groups, while the reduction in the TEER value of the RPMI2650 group was relatively smaller. This is because the intercellular tightness of the RPMI2650 cell mucosal model is lower, and changes in the tight junction structure have no significant effect on the permeability of the tissue. Furthermore, the TEER values ​​of each group decreased with increasing SA dosage, indicating that the regulatory effect of SA on mucosal tissue structure is dose-dependent.

[0072] The above results indicate that SA can increase intercellular spaces and enhance the permeability of mucosal tissues by reducing the tightness of intercellular connections.

[0073] Example 2: Study on the mechanism by which sodium aescinate alters mucosal tissue permeability

[0074] Based on the findings of Example 1, the expression of tight junction proteins between cells was investigated using Calu-3 and MDCK cells as models. The specific operational steps are as follows:

[0075] MDCK and Calu-3 cells in logarithmic growth phase were harvested and injected with 1.8 × 10⁻⁶ cells. 5 Cells were seeded per well in 12-well plates and cultured in an incubator. MDCK cells were incubated for 48 hours, and Calu-3 cells for 10 days. Then, 10 μg / mL SA solution was added to each well as the experimental group, and the same volume of culture medium was added as the control group. Incubation was continued for 2 hours. After incubation, 300 μL of primary antibody (Occludin and Claudin antibodies, respectively) was added to each well and incubated overnight in a humidified chamber at 4°C. After washing away excess primary antibody, approximately 1 μg / mL of fluorescent secondary antibody was added and incubated in the dark for 1 hour. After removing unbound secondary antibody, DAPI was added, and the cell nuclei were stained and observed under a 200x fluorescence microscope, and photographs were taken.

[0076] The test results are as follows Figure 2 As shown, Occludin and Claudin, marker proteins of tight junction structures, were observed in the control groups of both cell types, indicating that relatively dense tight junction structures formed between cells during incubation. After the addition of SA, the immunofluorescence intensity in the SA group was significantly reduced compared to the control group, indicating downregulation of Occludin and Claudin expression levels, suggesting that SA inhibited the formation of tight junction structures. Therefore, SA can disrupt tight junction structures and increase intercellular gaps by inhibiting the expression of tight junction proteins.

[0077] Example 3: Study on the efficacy and safety of sodium aescinate

[0078] Dodecyl maltoside and tetradecyl maltoside are novel permeation enhancers currently approved for clinical use, and their safety and permeation-enhancing efficiency are widely recognized. In this example, the two alkyl maltosides mentioned above are used as positive controls to evaluate the permeation-enhancing effect and safety of SA.

[0079] Following the method described in Example 1, an in vitro mucosal model based on MDCK cells was constructed. After approximately 3 days of incubation, 10 μg / mL of SA, dodecyl maltodextrin, and tetradecyl maltodextrin permeation enhancers were added, and incubation continued. After 3 hours, the medium was changed to remove each permeation enhancer. During incubation, changes in transmembrane resistance of the model were detected at set time points.

[0080] The test results are as follows Figure 3 As shown, similar to the positive control agents dodecyl maltodextrin and tetradecyl maltodextrin, SA rapidly reduced the TEER value in the cell model and increased mucosal permeability. After removal of the permeation enhancers, the transmembrane resistance of both the SA group and the dodecyl maltodextrin group quickly recovered to the pre-drug TEER value, and SA exhibited superior tissue structure recovery efficiency compared to tetradecyl maltodextrin. Therefore, given that both dodecyl maltodextrin and tetradecyl maltodextrin are approved for clinical use as permeation enhancers in pharmaceutical formulations, this indicates that SA also possesses high permeation-enhancing activity and safety.

[0081] Example 4: Permeation-enhancing effect of sodium aescinate on zero-permeability drugs

[0082] Sodium fluorescein, due to its high water solubility, low toxicity, and high fluorescence response value, is often used as a zero-permeability model drug in permeation studies (Montermini D, Winlove C. P, Michel C. C, Effects of perfusionrate on permeability of frog and rat mesenteric microvessels to sodium fluorescein [J]. Journal of Physiology, 2002, 543(3):959-975.). This example uses sodium fluorescein as a model drug to evaluate the permeation-enhancing effect of sodium fluorescein (SA) on zero-permeability drugs in RPMI2650, MDCK, and Calu-3 cell mucosal models. The specific operating steps are as follows:

[0083] The cell mucosal model was constructed using the same method as in Example 1. After construction, 10 μg / mL of sodium fluorescein and different doses of SA were added to the mucosal model. At different time points, 0.5 mL of the lower chamber solution of each group was taken, and the content of sodium fluorescein was measured using a microplate reader under Em498nm / Ex517nm conditions. The transmittance coefficient (Papp) at each time point was calculated using the following formula:

[0084] Papp=(dQ / dt) / (C0*A)

[0085] dQ is the total amount of fluorescein sodium contained in the following formula, dt is the permeation time, C0 is the drug concentration, and A is the mucosal surface area.

[0086] The test results are as follows Figure 4As shown, the permeation efficiency of sodium fluorescein varied among different mucosal model control groups. The RPMI2650 cell model group had the highest Papp value, while the Calu-3 and MDCK cell model groups had lower Papp values. This is because the RPMI2650 cell model has a relatively loose structure with larger intercellular spaces and tissue permeability, allowing sodium fluorescein to permeate more efficiently; whereas the Calu-3 and MDCK cell models have a relatively dense structure, resulting in the opposite effect (see...). Figure 4 (Control group). After the addition of SA, the Papp value of sodium fluorescein increased in all groups, but the increase in Papp value in the RPMI2650 group was relatively lower compared to the MDCK and Calu-3 groups; while the increase in Papp value was more significant in the MDCK and Calu-3 groups. This indicates that the permeation-enhancing effect of SA was more pronounced in the relatively tightly packed Calu-3 and MDCK cell models. Furthermore, the permeation-enhancing effect of sodium fluorescein became more significant with increasing SA dosage, demonstrating a clear dose-response relationship.

[0087] In summary, the results indicate that SA mainly improves the permeability of zero-permeability drugs by regulating the tight junction structure between cells and increasing the intercellular space.

[0088] Example 5: Permeation-enhancing effect of sodium aescinate on low-permeability drugs

[0089] This embodiment uses terbutaline, famotidine, and glycopyrronium bromide, all low-permeability drugs recommended in the "Guidelines for Exemption from Human Bioequivalence Trials," as model drugs to evaluate the permeation-enhancing effect of SA on low-permeability drugs in MDCK and Calu-3 cell mucosal models.

[0090] Calu-3 and MDCK cell mucosal models were constructed in Transwell chambers using the method described in Example 1. 100 μg / mL of the model drug and 10 μg / mL of SA were added to each experimental group model, while the control group received the same dose of the model drug but without SA. Furthermore, Papp values ​​were measured at 30 min, 60 min, and 90 min after drug administration, and the mean values ​​were calculated.

[0091] The test results are as follows Figure 5 As shown, compared with the control group, the Papp value in the experimental group after adding SA increased by more than 4 times, reaching the standard of hypertonic drugs.

[0092] The above results indicate that SA can improve the penetration efficiency of low-permeability drugs by regulating the tight junction structure between cells and increasing the intercellular space.

[0093] Finally, it should be noted that the above descriptions are merely embodiments of this application, used only to illustrate the technical solutions of the present invention, and are not intended to limit this application. Various modifications and variations can be made to this application by those skilled in the art. Any modifications, equivalent substitutions, improvements, etc., made within the spirit and principle of this application should be included within the scope of the claims of this application.

Claims

1. Application of sodium aescinate or its isomers as penetration enhancers.

2. The application according to claim 1, characterized in that, The penetration enhancer is used as a penetration enhancer for drugs with low permeability.

3. The application according to claim 2, characterized in that, The low-permeability drugs mentioned include BCS Class III and / or BCS Class IV in the biopharmaceutics classification system.

4. The application according to claim 2, characterized in that, The low-permeability drugs mentioned include small molecule drugs and / or large molecule drugs; The small molecule drugs include glycopyrronium bromide, terbutaline, famotidine, bromocriptine, diazepam, budesonide, mometasone furoate, mannitol, fluticasone propionate, beclomethasone propionate, cisaconazole, formoterol fumarate, dihydroergotamine mesylate, tiotropium bromide, naldolol, sodium valproate, ketamine, gabapentin, ketorolac tromethamine, metoprolol, lisinopril, butorphanol tartrate, olopatadine hydrochloride, azelastine hydrochloride, oxcarbazepine, chlorothiazide, palonosetron, and zolmitriptan. Naloxone hydrochloride, lorazepam, quetiapine fumarate, betahistine, carbamazepine, nalmefen hydrochloride, salmon calcitonin, levetiracetam, enalapril, alprazolam, xylometazoline hydrochloride, phosphatidylcholine, phenytoin sodium, indacaterol, umemetrine, polyethylene glycol 400, phenobarbital, salbutamol sulfate, ipratropium bromide, sulpiride, buspirone hydrochloride, ambroxol hydrochloride, olanzapine, acyclovir, lamotrigine, bromhexine hydrochloride, oxazepam, sumatriptan, remimazolam, or one or more of these drugs. The macromolecular drugs include one or more of the following: proteins, peptides, antibody-drug conjugates, peptide-drug conjugates, nucleic acid-drug conjugates, protein-drug conjugates, deoxyribonucleic acid, and ribonucleic acid.

5. The application according to any one of claims 1 to 4, characterized in that, At least one of sodium aescinate or its isomers as described in any one of claims 1 to 4 is prepared with a pharmaceutically acceptable carrier to form a clinically acceptable pharmaceutical preparation.

6. The application according to claim 5, characterized in that, The pharmaceutical preparation is a mucosal delivery preparation; preferably, the mucosal delivery preparation includes nasal mucosa, rectal mucosa, oral mucosa, lung mucosa, and ocular mucosa delivery preparations; more preferably, the mucosal delivery preparation is a lung mucosa and nasal mucosa delivery preparation.

7. The application according to claim 6, wherein the pulmonary mucosal delivery formulation comprises a suspension, solution, emulsion, microsphere, liposome, or powder formulation.

8. The application according to claim 6, characterized in that, The nasal mucosal delivery formulations include suspensions, solutions, emulsions, microspheres, liposomes, or powder formulations.

9. The application according to any one of claims 1 to 4, characterized in that, The sodium aescinate or its isomers promote drug absorption via the cellular bypass pathway by opening the tight junction structure between mucosal cells.

10. A method for promoting drug penetration, characterized in that, When using low-permeability drugs, sodium aescinate and / or its isomers may be used concurrently.

11. A pharmaceutical composition for mucosal administration, characterized in that, This includes low-permeability drugs, sodium aescinate, and / or its isomers.

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