TW201546086A - A novel triterpenoid and uses thereof - Google Patents

Abstract

Disclosed herein is a novel triterpenoid and its use in the treatment or prophylaxis of a kidney disease. The novel triterpenoid is capable of reducing the levels of blood urine nitrogen (BUN) and serum creatinine, which are factors associated with inflammation of kidney; as well as increasing the level of glomerular filtration rate (GFR). Therefore, the novel triterpenoid is useful for the manufacture of a medicament for the treatment and/or prophylaxis of a kidney disease, including acute kidney injury and chronic kidney inflammation.

Description

Novel triterpenoids and their uses

The present disclosure is directed to a novel triterpenoid compound and its use for the preparation of a medicament for the treatment of kidney diseases such as acute kidney damage and chronic kidney disease.

Acute kidney injury (AKI), also known as acute kidney failure, is caused by the rapid destruction of the renal filtration function, resulting in the rapid accumulation of nitrogen-containing metabolites (such as urea or creatinine) in the body. In the blood, the individual suffers from fatigue, loss of appetite, headache, nausea or vomiting, accompanied by a decrease in urine volume. AKI is prevalent in hospitalized patients, especially those who are seriously ill. There are many causes of AKI, such as tissue reperfusion after ischemia, serious injury, kidney disease or kidney transplantation. However, there is currently no clinically specific drug for the treatment of AKI, so supportive treatment (for example, dialysis) is often used to reduce symptoms for the main purpose. If patients can recover from the above diseases, they will often develop into long-term, chronic kidney disease patients, who need to rely on long-term dialysis, patients with serious illness, and even need a kidney transplant.

The current indicator used to determine the severity of AKI patients and whether they are likely to progress to CKD is glomerular filtration rate (glomerular Filtration rate, GFR). Decreased GFR value is often accompanied by an increase in blood urea nitrogen (BUN) and serum creatinine (CRE). Therefore, if a compound or a combination of compounds can effectively increase GFR, and inhibit BUN and CRE. It will be a lead compound that can potentially be used to prepare drugs that can treat or prevent kidney disease, including acute kidney damage and chronic kidney disease.

The present disclosure is based, at least in part, on the discovery of a novel triterpenoid compound isolated or purified from a fruiting or mycelium of Ganoderma lucidum and a composition comprising the novel triterpenoid compound, which reduces nephritis-related Biochemical indicators (including blood urea nitrogen and serum creatinine acid content) and increased renal spheroid filtration rate. This finding suggests that the novel triterpenoids of the present disclosure and compositions comprising the novel triterpenoids are useful as a medicament for the treatment or prevention of kidney diseases including acute kidney damage and chronic kidney disease.

Accordingly, a first object of the present disclosure is to provide a triterpene compound having the structure of the following formula (I),

According to an embodiment of the present invention, the structure of the formula (I) The compound itself can reduce the biochemical indicators related to nephritis, including blood urea nitrogen (BUN) and serum creatinine (CRE) content; and increase the glomerular filtration rate (GFR) Therefore, it can be used to manufacture drugs that can treat or prevent kidney diseases.

A second object of the present disclosure is to provide a treatment Or a pharmaceutical composition for preventing kidney disease. The pharmaceutical composition comprises a pharmaceutically effective amount of a triterpenoid compound of the above formula (I), a pharmaceutically acceptable salt, ester or solvate thereof; and a pharmaceutically acceptable carrier therefor.

According to an embodiment of the present invention, the disclosed pharmaceutical composition may further comprise at least another triterpene compound selected from the group consisting of: Iucidenic acid C (LAC), and Ricardonic acid N (Iucidenic acid N) , LAN), Ganoderma lucidum acid _{E 2 (Iucidenic acid E 2,} LAE- 2), Ganoderma lucidum acid A (Iucidenic acid A, LAA) , Ganoderma lucidum acid B (Iucidenic acid B, LAB) , Ganoderma lucidum acid D ₂ (Iucidenic acid D ₂ , LAD ₂ ) and combinations thereof.

According to a preferred embodiment, the disclosed pharmaceutical composition comprises a combination of a triterpenoid compound of the above formula (I) with LAA, LAB, LAC, LAD ₂ , LAE ₂ and LAN, and a pharmaceutically acceptable carrier therefor. Further, in the pharmaceutical composition, the amount of the triterpenoid compound and the LAC of the structure of the formula (I) is about 5 to 15% by weight based on the total amount of the triterpene compound; the amounts of LAA and LAD ₂ are respectively about three. 15-30% by weight of the total amount of the ruthenium compound; the amount of LAE ₂ is about 8-15% by weight of the total amount of the ruthenium compound; the amount of LAN and LAB is about 5 of the total amount of the ruthenium compound, respectively. -12% (% by weight).

Kidney suitable for treatment or prevention with the pharmaceutical composition Dirty diseases include acute kidney injury (AKI) and chronic kidney disease (CKD). In one example, the kidney disease is AKI, such as acute kidney inflammation. In another example, the kidney disease is CKD, such as chronic kidney inflammation.

If based on the total weight of the pharmaceutical or pharmaceutical composition, The total amount of the triterpene compound combination of the invention is from about 0.1% to about 99% by weight based on the total weight of the pharmaceutical composition. In certain embodiments, the total amount of the triterpenoid compound of the present invention is at least about 1% by weight based on the total weight of the pharmaceutical composition. In a particular embodiment, the total amount of the triterpenoid compound of the present invention is at least about 5% by weight based on the total weight of the pharmaceutical composition. In other embodiments, the total amount of the triterpenoid compound of the present invention is at least about 10% by weight based on the total weight of the pharmaceutical composition. In other embodiments, the total amount of the triterpenoid compound of the invention is at least about 25% by weight of the total weight of the pharmaceutical composition.

A third object of the present disclosure is to provide a method of treating kidney disease. The method comprises administering to a subject in need of treatment a therapeutically effective amount of a triterpenoid compound of the formula (I) of the present invention; or a pharmaceutical composition as described above. According to a preferred embodiment, the pharmaceutical composition comprises a triterpenoid compound of the formula (I), a pharmaceutically acceptable salt, ester or solvate thereof, and LAA, LAB, LAC, LAD ₂ , LAE ₂ and LAN; and a pharmaceutically acceptable carrier thereof. Kidney diseases suitable for treatment by the methods of the invention include AKI and CKD. In one example, the kidney disease is AKI, such as acute kidney inflammation. In another example, the kidney disease is CKD, such as chronic kidney inflammation. An individual suitable for treatment by the methods of the invention may be a mammal, preferably a human.

Through the following detailed description and accompanying patent application scope These and other features of the present disclosure will be more fully appreciated. The above summary and the following detailed description are merely illustrative, and are not intended to limit the scope of the disclosure.

The above and other objects, features, advantages and embodiments of the present invention will become more apparent and understood. The description of the drawings is as follows: FIG. 1A is a diagram (I) of the present invention according to an embodiment of the present invention. The compound inhibits the apoptosis of renal cells induced by cisplatin; FIG. 1B is a view of the inhibition of cisplatin-induced renal cell apoptosis by the pharmaceutical composition YKII-10 of the present invention according to an embodiment of the present invention; 2A is a diagram showing inhibition of an increase in serum urea nitrogen content induced by cisplatin in vivo according to an embodiment of the present invention, YKII-10; FIG. 2B is a diagram of an embodiment of the present invention. The pharmaceutical composition YKII-10 of the present invention inhibits the increase of serum creatinine acid content induced by cisplatin in vivo; FIG. 3 is a diagram showing the compound of formula (I) of the present invention and pharmacy according to an embodiment of the present invention. The composition YKII-10 reduces the increase of urea nitrogen induced by lipopolysaccharide in vivo, respectively; FIG. 4A is a diagram of inhibiting aristolochic acid in vivo by the pharmaceutical composition YKII-10 of the present invention according to an embodiment of the present invention. Sour The increase in serum urea nitrogen content; 4B is a view showing the inhibition of the increase in serum creatinine acid content induced by aristolochic acid in vivo by the pharmaceutical composition YKII-10 of the present invention according to an embodiment of the present invention; FIG. 5 is an embodiment of the present invention. The pharmaceutical composition YKII-10 of the present invention is used to increase the glomerular filtration rate in vivo; and FIG. 6 is a view of the pharmaceutical composition YKII-10 of the present invention according to an embodiment of the present invention. A condition that improves kidney function in the body.

The description of the embodiments of the present invention is intended to be illustrative and not restrictive. The features of various specific embodiments, as well as the method steps and sequences thereof, are constructed and manipulated in the embodiments. However, other specific embodiments may be utilized to achieve the same or equivalent function and sequence of steps.

In the following, the term "treatment or treating" includes the prevention (ie, prophylactic), curative or palliative effects that may lead to a desired pharmaceutical and/or physiological effect. Sexual disposal. Preferably, the effect refers to medically partially or completely curing or preventing apoptosis of kidney cells. In addition, the term "treatment" is used herein to mean the occurrence of one or more symptoms based on partial or complete mitigation, delayed onset, inhibition of progression, reduction in severity, and/or reduction in a particular disease, disorder, and/or medical condition. For the purpose of being tested An individual (or patient), especially having a medical condition, a symptom of the medical condition, a disease or condition caused by the medical condition, or an individual who is likely to develop an advanced condition toward the medical condition, The compounds or pharmaceutical compositions of the present disclosure are applied. Individuals who have not developed a significant condition for a particular disease, abnormality, and/or medical condition, and/or individuals who have an early onset of the particular disease, abnormality, and/or medical condition may also be treated to reduce the occurrence of the particular disease, Risk of pathology associated with abnormalities and/or medical conditions. Herein, the condition, disease, abnormality, and/or medical condition may be acute kidney injury (AKI) or chronic kidney disease (CKD). Reducing one or more signs or clinical indicators means that the treatment is "effective." Like this article, " "

"Prophylaxis" here refers to a means of preventing a future event. The prevention of renal cell apoptosis due to the prevention of renal cells due to poisons or drugs herein, therefore, the prophylactic treatment described herein can be performed before, simultaneously or after the individual is exposed to the poison or drug. means.

The term "an effective amount" is intended to reduce the serum levels of urea nitrogen and creatinine, reduce renal cell apoptosis, or the like for the purpose of treating kidney disease after an appropriate period of administration. It is to improve the glomerular filtration rate and other purposes. According to certain embodiments, the compound of formula (I) or the composition comprising a compound of formula (I) is a toxic substance that is exposed to a kidney injury in a subject (eg, a chemotherapeutic drug, lipopolysaccharide (LPS) ) or before aristolochic acid I (AA-I), applied prophylactically to On the subject being tested.

The words "compound", "composition", "agent" or "medicine or medicament" are used interchangeably herein and refer to when applied to a body (human Or an animal, a compound or composition capable of inducing a desired pharmaceutical and/or physiological response through local and/or systemic action.

The term "administered, administered" or "administration" as used herein refers to the direct administration of the compound or composition, or the administration of a prodrug of the active compound (eg, an ester of the active compound), a derivative ( Derivatives, or analogs, and the like, can be formed in an individual to form a substantial amount of the active compound.

The words "subject" or "patient" are used interchangeably herein to mean an animal (including a human) that is treatable by the compounds and/or methods. "Individual" or "patient" is used herein to encompass both male and female genders unless otherwise specified. Therefore, "individual" or "patient" includes any mammal, including, but not limited to, human, non-human primates, such as mammals, dogs, cats, horses, sheep, pigs, cattle, etc. The compounds benefit from treatment. Animals suitable for treatment with the compounds and/or methods of the invention are preferably human. In general, the terms "patient" and "individual" are used interchangeably herein.

"An effective amount" or "a therapeutically effective amount" can be used interchangeably, meaning For example, a dose of a therapeutic compound or composition is administered. The measurement of efficacy can be objective (measured by some test or marker) or subjective (the subject gives a narrative of its effects and feelings). The effective amount of the above compound or composition may range from about 0.1 mg/kg/day to about 100 mg/kg/day, preferably from 1 mg/kg/day to about 50 mg/kg/day, more preferably From 5 mg/kg/day to about 10 mg/kg/day. The effective dose will also vary with the route of administration and the other agents that may be used in combination.

In the present specification, the term "triterpene compound" encompasses triterpenoid compounds and pharmaceutically acceptable salts, esters or solvates thereof, unless otherwise stated to the contrary. For example, the compound of formula (I) of the present disclosure is a triterpene compound, and thus, when the compounds of formula (I) of the present disclosure are described in the specification, the salts of the compounds of formula (I) of the present disclosure are also encompassed. , esters or solvates. The term "salt" as used herein refers to a salt formed by the reaction of a base with an acid (for example, a novel triterpenoid compound of the present disclosure). The salt obtained from a suitable base contains an alkali metal such as sodium. ), an alkaline earth metal (such as magnesium), an ammonium ion and a salt of N (alkyl) ₄ ⁺ . The term "ester" refers to an ester of a triterpene compound of the present disclosure, exemplified by a compound of formula (I) which is a carboxyl group of a compound of formula (I) and an alcohol (eg, a straight chain or a branch). a chain of C _1-6 alcohols formed after the reaction; or a hydroxyl group of the compound of formula (I) and another organic acid (R-COOH, wherein R is a linear or branched C _1-6 alkane Base) formed after the reaction. The term "solvate" as used herein, refers to the formation of a compound (eg, a compound of formula (I) in the present disclosure) that reacts with surrounding solvent molecules (eg, water, ethanol, etc.). The wrong compound. In one example, the bismuth compound is an ethanolate of the structure of formula (I).

Although numerical ranges and parameters are used to define a broad range of values for the present invention, the relevant values in the specific embodiments are presented as precisely as possible. However, any numerical value inherently inevitably contains standard deviations due to individual test methods. As used herein, "about" generally means that the actual value is within plus or minus 10%, 5%, 1%, or 0.5% of a particular value or range. Alternatively, the term "about" means that the actual value falls within the acceptable standard error of the average, depending on the considerations of those of ordinary skill in the art to which the invention pertains. Except for the experimental examples, or unless otherwise explicitly stated, all ranges, quantities, values, and percentages used herein are understood (eg, to describe the amount of material used, the length of time, the temperature, the operating conditions, the quantity ratio, and the like. Are all modified by "about". Therefore, unless otherwise indicated to the contrary, the numerical parameters disclosed in the specification and the appended claims are intended to be At a minimum, these numerical parameters should be understood as the number of significant digits indicated and the values obtained by applying the general carry method.

The scientific and technical terms used herein have the same meaning as commonly understood by those of ordinary skill in the art to which the invention pertains, unless otherwise defined herein. In addition, the singular noun used in this specification covers the plural of the noun in the case of no conflict with the context; the plural noun of the noun is also included in the plural noun used.

At least a portion of the present disclosure has been developed based on a novel triterpenoid compound that is isolated from the body or mycelium of the ganoderma lucidum, and the novel triterpenoid compound and the pharmaceutical composition comprising the novel triterpenoid compound have a reduced Serum urea nitrogen and creatinine acid levels and improve kidney The characteristics of the spheroidal filtration rate. Thus, the novel triterpenoids and pharmaceutical compositions comprising the novel triterpenoids are potentially useful agents for the manufacture of a medicament for the treatment or prevention of kidney disease.

Accordingly, a first aspect of the present disclosure is to provide a triterpene compound having the structure of formula (I),

According to a particular embodiment of the invention, the triterpene compound having the structure of formula (I) is itself capable of reducing the elevation of serum urea nitrogen or creatinine induced by a drug such as cisplatin or aristolochic acid. Or restore renal cell apoptosis caused by drugs such as cisplatin or aristolochic acid.

The inventors have also discovered a novel combination of triterpenoids in which each triterpene compound is isolated from the body or mycelium of the ganoderma lucidum, and the novel triterpene compound combination is the same as the compound of the above formula (I). Reduces the ability of serum urea nitrogen or creatinine to rise by drugs (eg, cisplatin or aristolochic acid), or restores kidney cells from drugs (eg, cisplatin or aristolochic acid) Death phenomenon.

Based on this, a second object of the present disclosure is to provide a pharmaceutical composition for treating or preventing kidney disease. This pharmaceutical composition A triterpenoid compound comprising at least one pharmaceutically effective amount of the above structure of formula (I); and a pharmaceutically acceptable carrier therefor.

According to an embodiment of the present invention, the disclosed pharmaceutical composition may further comprise at least another triterpene compound selected from the group consisting of: Iucidenic acid C (LAC), and Ricardonic acid N (Iucidenic acid N) , LAN), Ganoderma lucidum acid _{E 2 (Iucidenic acid E 2,} LAE- 2), Ganoderma lucidum acid A (Iucidenic acid A, LAA) , Ganoderma lucidum acid B (Iucidenic acid B, LAB) , Ganoderma lucidum acid D ₂ (Iucidenic acid D ₂ , LAD ₂ ) and combinations thereof.

In certain embodiments, the amount of the triterpenoid compound of the formula (I) and the LAC in the pharmaceutical composition is about 5-15% by weight, for example about 5, 5.5, 6 of the total triterpenoid content, respectively. 6.5, 7, 7.5, 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5, 12, 12.5, 13, 13.5, 14, 14.5 or 15% (% by weight), which is contained in the composition The total amount of the triterpenoid compound is calculated as 100%. In a preferred embodiment, the amount of the triterpenoid compound and LAC of the structure of formula (I) is about 10% of the total amount of the triterpene compound, respectively.

In other embodiments, the amount of LAA and LAD ₂ in the pharmaceutical composition is about 15-30% by weight, respectively, of the total amount of the triterpene compound; for example, about 15, 15.5, 16, 16.5, 17, 17.5, 18, 18.5, 19, 19.5, 20, 20.5, 21, 21.5, 22, 22.5, 23, 23.5, 24, 24.525, 25.5, 26, 26.5, 27, 27.5, 28, 28.5, 29, 29.5 or 30% (weight %), which is calculated by taking the total amount of the triterpenoid compound contained in the composition as 100%. In one example, the amount of LAA and LAD ₂ is about 25% of the total amount of triterpenoids in the composition, respectively.

In other embodiments, the amount of LAE ₂ in the pharmaceutical composition is from about 8 to 15% by weight of the total amount of the triterpenoid compound; for example, about 8, 8.5, 9, 9.5, 10, 10.5, 11, 11.5 , 12, 12.5, 13, 13.5, 14, 14.5 or 15% (% by weight), which is calculated as the total amount of the triterpenoid compound contained in the composition as 100%. In one example, the amount of LAE ₂ is about 12% by weight of the total amount of the triterpenoid compound in the pharmaceutical composition.

In certain embodiments, the amount of the LAN and LAB in the pharmaceutical composition is from about 5 to 12% by weight of the total triterpenoid content, for example, about 5, 5.5, 6, 6.5, 7, 7.5, respectively. 8, 8, 9.5, 9, 9.5, 10, 10.5, 11, 11.5 or 12% (% by weight), which is calculated as the total amount of the triterpenoid compound contained in the composition as 100%. In a preferred embodiment, the amount of LAN and LAB is about 8.5% (% by weight) of the total amount of the triterpenoid compound in the pharmaceutical composition, respectively.

According to a preferred embodiment, the disclosed pharmaceutical composition comprises a triterpenoid compound of the above formula (I), LAC, LAN, LAE ₂ , LAA, LAB and LAD ₂ , and a pharmaceutically acceptable carrier thereof; The amount of the triterpene compound of the formula (I) and the amounts of LAC, LAN, LAE ₂ , LAA, LAB, and LAD ₂ are about 10%, 10%, 8.5%, 15%, 25%, and the total triterpene compound content, respectively. 25% (% by weight).

The triterpenoid compound of the present invention, the triterpenoid compound including the structure of the above formula (I), and LAC, LAN, LAE ₂ , LAA, LAB and LAD ₂ can be separated from the Ganoderma genus according to the method disclosed in Example 1, For example, it is isolated from the fruit body or mycelium of Ganoderma lucidum . Regardless of whether the raw material is derived from the fruit body or the mycelium of the ganoderma lucidum, the separation and purification method is generally carried out by using a solvent (preferably an alcohol solution) at a temperature higher than room temperature, and then the extract is subjected to a column layer. The analysis includes, but is not limited to, high-efficiency liquid chromatography (HPLC) and reverse phase liquid chromatography (Reverse Phase Liquid Chromatograpy), and the like, and steps of concentration, drying, and the like until a dry powder of the active compound of the ganoderma lucidum is obtained.

Kidney diseases suitable for treatment or prevention with the pharmaceutical composition include acute kidney injury (AKI) and chronic kidney disease (CKD). In one example, the kidney disease is AKI, such as acute kidney inflammation. In another example, the kidney disease is CKD, such as chronic kidney inflammation.

Generally, the total amount of the triterpenoid compound of the present invention is from about 0.1% to about 99% by weight based on the total weight of the pharmaceutical or pharmaceutical composition. In certain embodiments, the total amount of the triterpenoid compound of the present invention is at least about 1% by weight based on the total weight of the pharmaceutical composition. In a particular embodiment, the total amount of the triterpenoid compound of the present invention is at least about 5% by weight based on the total weight of the pharmaceutical composition. In other embodiments, the total amount of the triterpenoid compound of the present invention is at least about 10% by weight based on the total weight of the pharmaceutical composition. In other embodiments, the total amount of the triterpenoid compound of the invention is at least about 25% by weight of the total weight of the pharmaceutical composition.

Kidney diseases suitable for treatment by the methods of the invention include AKI and CKD. In one example, the kidney disease is AKI, such as acute kidney inflammation. In another example, the kidney disease is CKD, such as chronic kidney inflammation. An individual suitable for treatment by the methods of the invention may be a mammal, preferably a human.

The above drugs can be prepared according to the accepted pharmaceutical process. Or a pharmaceutical composition such as that disclosed in Remington's Pharmaceutical Sciences (17th edition, ed. Alfonoso R. Gennaro, Mack Publishing Company, Easton, Pa (1985)). A pharmaceutically acceptable excipient is meant to be compatible with the other ingredients of the pharmaceutical formulation and compatible with the organism.

In accordance with the disclosed embodiments, the medicament or pharmaceutical composition disclosed herein may be administered via any suitable route of administration, such as a capsule, suspension or tablet taken orally, or without gastrointestinal tract. Way to apply. The mode of administration without gastrointestinal route includes systemic administration such as intramuscular injection, intravenous injection, subcutaneous injection or intraperitoneal injection. Alternatively, it can be applied by means of a transdermal membrane, such as topical skin application or inhalation (eg, intrabronchial, intranasal, intraoral or nasal drops, etc.); or intrarectal administration. Administration may be carried out alone or in combination with conventional pharmaceutically acceptable adjuvants. In a preferred embodiment, the compounds of the invention can be administered to an individual via oral means (e.g., through food).

If administered orally, the triterpenoid compound of the present invention or a novel combination thereof can be formulated to contain various adjuvants (eg, microcrystalline cellulose, calcium carbonate, dicalcium phosphate, and glycine); various disintegrants ( Such as starch, alginic acid and specific citrate); and granule binders (eg, polyvinylpyrrolidone, sucrose, gelatin and acacia). In addition, lubricants such as magnesium stearate, sodium lauryl sulfate, and talc may also be included. Preferred materials associated therewith include sugars in lactose or milk as well as high molecular weight polyethylene glycols. The above solid dosage form may also optionally include a coating or shell, such as a casing coating, and to improve the release of active ingredients of any of the drugs. Rate of coating. Examples of such coatings are well known to those skilled in the art. In one example, the pharmaceutical composition is formulated as a tablet. In another example, the pharmaceutical composition is a granule formulated to be filled in a soft or hard gelatin capsule or encapsulated in a biodegradable kit. When used in the form of oral suspensions and / or special effects (elixirs), the active ingredients may be combined with various sweeteners or flavors, colorants or dyes, and emulsifiers and/or suspensions may be added if desired. And diluents such as water, alcohol, propylene glycol, glycerin. In certain embodiments, the pharmaceutical compositions of the invention are formulated as liquid dosage forms suitable for oral administration. Such liquid formulations may further include a buffer to maintain the pH. This liquid formulation can also be filled in a soft capsule. The liquid dosage form can be a solution, suspension, emulsion, microemulsion, precipitate or can carry a compound of the invention, a pharmaceutically acceptable derivative, isomer, metabolite, salt or solvate thereof Any liquid medium. The liquid can be designed to improve the solubility of the pharmaceutically acceptable salts of the compounds of the invention to form a drug-containing emulsion or dispersion. When such a dosage form is employed, it is formed by mixing the active compound with at least one pharmaceutically acceptable adjuvant, including, but not limited to, the above-described adjuvants.

If administered in a parenterally manner, the triterpenoid compound of the present invention or a novel combination thereof can be formulated into a liquid pharmaceutical composition which can be administered intravenously, intramuscularly, subcutaneously or intraperitoneally. Sterile solutions or suspensions for administration. Diluents which can be used in the manufacture of such sterile injectable solutions or suspensions include, but are not limited to, 1,3-butanediol, mannitol, water, Ringer's solution, isotonic sodium chloride solution. It can also be derived from fatty acids (eg, oleic acid) and its glycerides. Or a naturally pharmaceutically acceptable oil (eg, olive oil or rapeseed oil) to make a solution or suspension for injection. Alcohols or carboxymethylcellulose or similar dispersing agents for dilution may also be included in such oily solutions or suspensions. Other commonly used surfactants (e.g., Tweens or Spans series) or emulsifiers, or agents commonly used in the pharmaceutical arts to enhance bioavailability can be used.

The above pharmaceuticals or pharmaceutical compositions of the present invention may also be formulated into a variety of dosage forms suitable for mucosal applications, such as buccal and/or sublingual pharmaceutical dosage units, for delivery of drugs through Oral mucosa. A wide variety of biodegradable and pharmaceutically acceptable polymeric adjuvants can be used which provide the pharmaceutical compositions with acceptable adsorption and desired drug release patterns, as well as buccal and/or sublingual drugs. The active ingredient or other ingredients to be administered in the dosage unit are compatible. In general, the above polymeric adjuvants comprise a hydrophilic polymer that adheres to the wetted surface of the oral mucosa. Examples of polymeric adjuvants include, but are not limited to, acrylic acid polymers and copolymers; hydrolyzed polyvinyl alcohol; polyethylene oxides; polyacrylates; Vinyl polymers and copolymers; polyvinylpyrrolidine; dextran; guar gum; pectins; starch; and cellulosic polymers.

It will be understood that the dosage of the triterpenoids of the invention or their novel combinations will vary from individual to individual, not only because of the particular combination employed. The difference in factors such as the desired response to the substance or composition, the route of administration, the compound (alone or in combination with one or more drugs), may also be affected by other factors, such as the condition or severity of the condition to be treated. Degree; the age, sex or weight of the patient, the health of the patient; the severity of the pathological condition to be treated, other medical or special dietary content of the patient at the same time; and other factors conceivable by those of ordinary skill in the art; The medical personnel responsible for the care can ultimately determine the appropriate dose based on these factors. The dosage and form of administration can be adjusted to provide a preferred therapeutic response. A therapeutically effective amount also refers to a toxic or detrimental effect of a compound or composition that is less than the therapeutic benefit it brings. In a preferred embodiment, the combination of the triterpenoids of the present invention, when administered, should be administered in an appropriate dosage for a period of time to reduce the number and/or severity of symptoms.

When the triterpenoid compound of the formula (I) or the pharmaceutical composition of the present invention is applied to a body, the effective amount to be administered is about 0.1 to 100 mg/kg/day of the individual body weight. The pharmaceutical composition to be applied to the individual per day is used in an amount of about 0.1, 0.5, 1, 2, 5, 7, 9, 10, 12, 15, 17, 19, 20, 22, 25, 27, 29, 30 , 32, 35, 37, 39, 40, 42, 45, 47, 49, 50, 52, 55, 57, 59, 60, 62, 65, 67, 69, 70, 72, 75, 77, 79, 80 , 82, 85, 87, 89, 90, 92, 95, 97, 99, or 100 mg/kg body weight; preferably about 1-50 mg/kg body weight, for example about 1, 2, 5, 7, 8, 9, 10, 12, 15, 17, 19, 20, 22, 25, 27, 29, 30, 32, 35, 37, 39, 40, 42, 45, 47, 49, 50 mg / kg body weight; better Is about 5-10 mg / kg body weight, for example about 5, 7, 8, 9 or 10 mg / kg body weight. These doses can be administered in a single administration or divided into multiple administrations within one day.

A third object of the present disclosure is to provide a method of treating kidney disease. The method comprises administering to a subject in need of treatment a drug or composition that treats a kidney disease. When the drug or composition is administered to a mammalian subject, preferably a human, in need of treatment, an effective amount of a triterpenoid compound of the formula (I) of the present invention or a pharmaceutical combination of the present invention described above may be delivered. Things. The drug can be administered by a suitable route, such as oral, nasal, transpulmonary, transdermal, or the like, or intravenously, intramuscularly, subcutaneously, intrarectally, intraocularly, or the like.

The following examples are presented to illustrate certain aspects of the present disclosure and to assist those skilled in the art to understand and practice the present disclosure. However, the scope of the disclosure is not limited to these embodiments.

Example

Materials and Methods

Cell culture

This study used MDCK large kidney cell line, inoculated in a 96-well culture dish at a density of ^1.5 ×10 ⁵ cells/plate, and cultured in Dulbecco's modified Eagle's medium (DMEM). And added 10% fetal calf serum (FCS), 100 units/ml of penicillin, 100 ng/ml of streptomycin (Invitrogen, Carlsbad, CA), 2 mM L-glutamic acid and non-in the medium. Necessary amino acids and sodium pyruvate were maintained and maintained at 37 ° C in a humid environment (5% CO ₂ and 95% air).

Experimental animal

In this experiment, BLb/c mice were used, kept in the environment free of pathogenic bacteria, maintained at 22±3°C at room temperature, maintained at 50±20% relative humidity, maintained 12/12 hours in day and night, and free access to drinking water. food. Animal feed was used with experimental rodent diet 5001 (purchased from PMI ^® Nutrition International, Inc., MO, USA). For drinking water, boiled tap water is used. All animal experiments were carried out in accordance with the "Guidelines for the Management and Use of Laboratory Animals" published by the Chinese Society of Laboratory Animals.

Example 1 Compounds of the Invention and Pharmaceutical Compositions

1.1 Isolation and purification of compounds of formula (I)

After the dry weight of 50 g of Ganoderma lucidum fruit body was broken by a powder machine, the Ganoderma lucidum powder was placed in an oscillating extraction device, and ethanol was added in a ratio of dry powder of Ganoderma lucidum fruit body: alcohol volume = 1:24, and then 37 Concentrate after one day of cooking at °C. The foregoing extraction step was repeated several times in succession, and the obtained extract was filtered, combined, and concentrated under reduced pressure to give a crude extract of 5 g of a ginseng fruit solid in a yield of about 10%. The crude extract of the Ganoderma lucidum fruit body obtained above is filtered by a vacuum drying system at a constant temperature of 40 ° C overnight to remove residual solution. Agent.

The above-obtained dry weight 5.0 g of the crude extract of the ginseng fruit body alcohol was extracted by adding ethyl acetate containing an equal volume of water to obtain another aqueous layer solution and an ethyl acetate extraction layer solution. After the ethyl ester extraction layer solution is concentrated and dried, the residual solvent is removed by vacuum filtration at a constant temperature of 40 ° C overnight, and the yield is about 40% (2 g) of the total dry weight of the crude extract of the G. lucidum fruit body. Solid ethyl acetate extract.

The dried ethyl acetate extract obtained above is dissolved in alcohol and then separated by semi-preparative HPLC. The mobile phase is cyanomethane/2% acetic acid (1/2 ^to 1/4), and the formula (I) can be obtained. Compound (8 mg) in a yield of 0.4%.

The ¹³ C, ¹ H, and Heteronuclear Multiple Bond Correlation (HMBC) data for the compound of formula (I) are as follows:

MS (EI, 20eV), m/z (rel.int.): 460 [M+] (53.56), 442 (22.56), 339 (16.46), 322 (100), 304 (46.41); HRMS m/z = 460.2834 (calculated molecular weight of C ₂₇ H ₄₀ O ₆ : 460.2819)

IR ν _max 3445, 1736, 1707, 1661 cm ^-1

UV(MeOH)λ _max 252nm

Melting point: 124-126 ° C

1.2 Preparation of a pharmaceutical composition containing a compound of formula (I) YKII-10

The pharmaceutical composition YKII-10 which specifically treats kidney diseases in the present disclosure is a mixture of the novel compound of the formula (I) isolated in the embodiment 1.1 and other conventional triterpenoid compounds according to the formulation shown in Table 2 below. Thereafter, the triterpene compounds can be isolated or purified from the body or mycelium of the ganoderma lucidum according to any of the disclosed separation or purification methods.

Example 2 The compound of the formula (I) and the pharmaceutical composition of YKII-10 can effectively alleviate or ameliorate the inflammation of the kidney induced by cisplatin

In this embodiment, cisplatin is used to induce apoptosis of kidney cells in vitro or in vivo, and then the compound of the formula (I) or the YKII-10 composition of the present invention is administered to evaluate the treatment of kidney diseases. efficacy.

2.1 cisplatin-induced renal cell apoptosis

MDCK large kidney cells were seeded in a flat-bottomed 6-well culture dish at a density of 1 × 10 ⁵ cells/well, cultured overnight in an incubator, and then pre-treated with 50 M cisplatin (Cisplatin). The cells were treated with the compound of formula (I) (50 μg/mL or 100 μg/mL) or the YKII-10 pharmaceutical composition of Example 1 (25-100 μg/mL) for treatment for 4 hours, and the cells were treated directly after 24 hours. Add 50nM DiOC6 dye to the mitochondrial membrane potential staining, react at 37 ° C for 30 minutes, then wash the cells twice with PBs, observe the mitochondrial membrane potential staining using inverted fluorescence microscope and photograph it. Dead kidney cells cannot be stained with DioC6 to assess apoptosis. The results are shown in Figures 1A and 1B, respectively.

As shown in the bar graph of Figure 1A, the compound of formula (I) (50 μg/mL) alone can effectively restore the decrease of tubular membrane potential induced by cisplatin. When the concentration is increased to 100 μg/mL, the membrane can be reversed. The potential drops more than 80%, making it tend to control the membrane potential of the group. Similarly, if different doses of YKII-10 pharmaceutical composition (25-100mg/Kg) are administered, the tubular membrane potential drop can also be effectively restored, and the degree of recovery is The YKII-10 pharmaceutical composition is increased in dose (Fig. 1B). It can be seen that the compound of the formula (I) of the present invention or the pharmaceutical composition YKII-10 comprising the compound of the formula (I) can effectively reverse the apoptosis of renal tubular cells, and thus can be used for alleviating or treating nephritis.

2.2 cisplatin-induced inflammation of mouse kidney

In this example, 6-week-old C57BL/6 mice were selected, and the experimental group was orally administered the YKII-10 pharmaceutical composition of Example 1.2 (10 mL/Kg) for 5 consecutive days, and the control mice were followed. Distilled water was given, and all animals were given cisplatin (15 mg/kg, ip) once a day by intraperitoneal injection on day 2, and the blood of the mice was collected 3 days later. The blood was centrifuged to obtain serum and then biochemical analyzer (TOSHIBA, TBA) was used. -40FR) Blood urea nitrogen (BUN) and serum creatinine (CRE) were measured to evaluate renal function in mice. The results are shown in Figure 2.

As shown in Figures 2A and 2B, administration of the YKII-10 pharmaceutical composition (10 mL/Kg) of Example 1.2 of the present invention to an experimental animal orally can effectively reduce serum biochemical indicators associated with inflammation, including BUN and CRE. Proximate to the control group, it is indicated that the pharmaceutical compositions of the present invention comprising a compound of formula (I) are effective for the relief or treatment of nephritis.

Example 3 The compound of the formula (I) and the YKII-10 pharmaceutical composition of Example 1 can effectively alleviate or improve lipopolysaccharide-induced nephritis in vivo.

In the present embodiment, mouse kidney inflammation is induced by lipopolysaccharide (LPS), followed by administration of the compound of the formula (I) or YKII-10 of the present invention to evaluate the therapeutic kidney. Dirty and inflamed effect.

Six weeks old BLb/c mice were used in this study. Animals were grouped by body weight one day before the test, and there was no statistically significant difference in body weight between the groups after grouping. After the start of the experiment, LPS (2.5 mg/Kg) was intraperitoneally injected into the mice every day for 5 days to induce inflammation of the kidneys of the mice. At the same time, the experimental group of the compound of the formula (I) of Example 1.1 (100 mg / Kg) or the YKII-10 composition of Example 1.2 (dose of 250 mg / Kg) was administered once a day orally, and the control group was small. The rats were given distilled water. Animal urine was collected on the 5th and 10th day after the start of the test, and the urine protein content was measured by an ARKRAY urine analyzer to evaluate the kidney function of the mouse. The results are shown in Figure 3.

As shown in the bar graph of Figure 3, inflammation of the mouse kidney induced by LPS, whether using the compound of formula (I) alone (100 mg/Kg) or in combination with other known triterpenoids (e.g., Example 1.2) The YKII-10 pharmaceutical composition (250mg/Kg)) can effectively reduce the amount of protein in the urine of the experimental animal, and represents the compound of the formula (I) of the present invention or the pharmaceutical composition YKII-containing the compound of the formula (I). 10 can effectively relieve or treat kidney inflammation.

Example 4 The YKII-10 pharmaceutical composition of Example 1 can effectively alleviate or improve acute kidney damage in vivo.

In this example, aristolochic acid I (AA-I) was used to induce acute kidney injury in mice, followed by administration of the YKII-10 pharmaceutical composition of Example 1.2 of the present invention. To assess its efficacy in treating acute kidney damage.

4.1 AA-I induces renal function damage animal model

Pick 6-week-old Balb/c mice daily for oral administration The mice were given AA-I (2.5 mg/kg) for 5 consecutive days, and the experimental group mice were orally administered with the YKII-10 pharmaceutical composition of Example 1.2 (300 mg/kg) once daily for continuous administration. On the 12th, the control group was given distilled water. The blood of the mice was collected on the 12th day, and the serum was obtained by centrifugation. The values of urea nitrogen (BUN) and Creatinine (CRE) were measured by a biochemical analyzer (TOSHIBA, TBA-40FR) to evaluate the renal function of the mice. The results are shown in Figures 4A and 4B.

The result of Fig. 4 shows that the YKII-10 pharmaceutical composition of Example 1.2 of the present invention can effectively reduce BUN and CRE induced by AAI, and indicates that the pharmaceutical composition YKII-10 of Example 1.2 of the present invention can effectively alleviate or treat acute Kidney damage.

4.2 glomerular filtration rate (GFR) test

In this embodiment, the fluorescent labeling of inulin is used as the sole excretion pathway of the kidney spheroid, injected into the mouse, and the fluorescence emission rate is monitored by a living fluorescent image detector to reflect the kidney. Silk ball filtration efficiency.

Six-week-old Balb/c mice were selected, and the YKII-10+AAI experimental group was given a drug (dose of 300 mg/Kg) for 8 consecutive days. On the 4th day, AA-I (2.5 mg/kg) was administered intraperitoneally. On the 5th; YKII-10 experimental group mice were given drugs (300mg/kg) for 8 consecutive days; AA-I experimental group mice were given distilled water for 3 days, and on the 4th day, AA-I (2.5mg/kg) was continuously injected intraperitoneally. For 5 days, the control group was given distilled water. Eight days after the test, fluorescently labeled inulin (GFR-Vivo 680) (2 x 10 ^-9 mol) was injected into the mice and injected by Perkin Elmer In Vitro Fluorescence Image Detector (FMT 4000). Fluorescence images were detected at 1, 5, 15, 30, and 45 minutes, and the concentration of fluorescently labeled inulin in the mice was calculated, and the renal spheroid filtration curve was calculated to compare the kidneys of the four groups of mice. Ball filtration efficiency. The results are shown in Figure 5.

Figure 5 is a bar graph showing that when the mouse is exposed to AAI treatment, the renal spheroid filtration rate (GFR) drops 10 times from the original normal value (about 250 μL/min) to about 25 μL/min, if the pharmaceutical composition is Treatment with YKII-10, although unable to completely reverse the acute kidney damage caused by AAI, can effectively increase the GFR by about 4 times to about 100 μL/min.

Example 5 The YKII-10 pharmaceutical composition of Example 1 can effectively alleviate or improve living chronic kidney disease

In the present embodiment, by removing a part of the kidney of the experimental animal by surgery, the renal function is lowered to about 1/6 of that of the normal animal, and then the YKII-10 pharmaceutical composition of Example 1.2 of the present invention is applied to evaluate the improvement. The efficacy of kidney function, thereby reflecting the efficacy of the YKII-10 pharmaceutical composition of Example 1.2 of the present invention for treating chronic kidney disease.

Eight-week-old adult male rats were randomly divided into experimental group and control group (3 rats in each group). Each group of animals was surgically removed 2/3 of the left kidney, and the animals were rested for one week before the right kidney was removed. The renal function of the test animals was reduced to about 1/6 of that of normal animals to simulate the renal function of chronic kidney patients. The drug was started in the second week after the operation. The experimental group was given 300 mg/kg of the YKII-10 pharmaceutical composition of Example 1.2 per day, and the control group was given water. At the same time, blood was collected from the tail of the animal to determine the creatinine in the blood of the experimental animal. The content was converted to eGFR (estimated GFR) by the following equation to evaluate renal function, and the pre-experimental eGFR was used as a baseline for 100% renal function: eGFR (ml/min/1.73 m ^ 2 ) = 186 x (creatinine) Acid value) ^-1.154 x(30) ^-0.203 Next, the creatinine acid content was measured every two weeks and converted to eGFR by the same formula. The result is shown in Fig. 6.

As shown in the results of Figure 6, the kidney function of the test animals was about 44% of the normal rats at the beginning of the administration. At 6 weeks after the operation, the renal function of the experimental animals in the administration group was significantly increased to 59% of the normal rats. There was no change in the control group at 15%, and there was a statistically significant difference between the experimental group and the control group in the sixth week after T-test calculation.

In summary, the novel triterpenoids of the present disclosure (i.e., the compounds of formula (I)) and pharmaceutical compositions comprising the compounds of formula (I) are potential agents for the treatment of acute or chronic kidney disease.

It is to be understood that the above-described embodiments and examples are merely illustrative, and various modifications may be made by those skilled in the art. The description, examples, and materials set forth above are intended to be illustrative of the present invention and are illustrative of the invention. The present disclosure has been disclosed in the above embodiments, but it is not intended to limit the disclosure, and any person skilled in the art can make various changes and refinements without departing from the spirit and scope of the disclosure. The scope of protection of the disclosure is subject to the definition of the scope of the patent application.

Claims (8)

a triterpene compound having the structure of the following formula (I), And pharmaceutically acceptable salts, esters or solvates thereof. A pharmaceutical composition for slowing or treating kidney disease comprising an effective amount of a triterpenoid compound as claimed in claim 1; and a pharmaceutically acceptable carrier therefor. The pharmaceutical composition according to claim 2, further comprising another triterpenoid compound selected from the group consisting of: cinnamic acid C, cinnamic acid N, cinnamic acid E ₂ , cinnamic acid A, cinnamic acid B , 赤芝酸 D ₂ and combinations thereof. The pharmaceutical composition according to claim 3, wherein the amount of the compound of the formula (I) and the amount of cinnamic acid C is about 5-15% by weight based on the total amount of the triterpenoid compound in the pharmaceutical composition. The pharmaceutical composition according to claim 4, wherein the amount of gibberellic acid A and anthoic acid D ₂ is about 15-30% by weight, respectively, based on the total amount of the triterpenoid compound in the pharmaceutical composition. The pharmaceutical composition according to claim 4, wherein the amount of erythric acid E _{2 is} from about 8 to 15% by weight based on the total amount of the triterpenoid compound in the pharmaceutical composition. The pharmaceutical composition according to claim 4, wherein the amount of cinnamic acid B and gibberic acid N is from about 5 to 12% by weight based on the total amount of the triterpenoid compound in the pharmaceutical composition. The pharmaceutical composition according to claim 3, wherein the kidney disease is acute kidney injury or chronic kidney disease.