CN110575447B

CN110575447B - Pharmaceutical composition for preventing and treating diabetes and application thereof

Info

Publication number: CN110575447B
Application number: CN201810578941.1A
Authority: CN
Inventors: 张可; 谭昕; 于朝晖; 李向东; 连萌
Original assignee: Hanyi Bio Technology Beijing Co ltd
Current assignee: Hanyi Bio Technology Beijing Co ltd
Priority date: 2018-06-07
Filing date: 2018-06-07
Publication date: 2022-11-29
Anticipated expiration: 2038-06-07
Also published as: CN110575447A

Abstract

The invention belongs to the field of medicines, and relates to a pharmaceutical composition for preventing and treating diabetes and application thereof. In particular, the present invention relates to a pharmaceutical composition comprising: cannabidiol and/or a pharmaceutically acceptable salt or ester thereof; and one or more hypoglycemic agents; optionally, the pharmaceutical composition further comprises one or more pharmaceutically acceptable excipients. The pharmaceutical composition can effectively reduce blood sugar, obviously reduce adverse reactions of metformin medicaments or sulfonylurea secretagogues, and has good application prospect.

Description

Pharmaceutical composition for preventing and treating diabetes and application thereof

Technical Field

The invention belongs to the field of medicines, and relates to a pharmaceutical composition for preventing and treating diabetes and application thereof.

Background

At present, the prevalence rate of diabetes is on the rapid rise, and becomes another important chronic non-infectious disease after cardiovascular diseases and tumors, which seriously endangers the health of residents. Disability and lethality caused by diabetes and various acute and chronic complications thereof have become the third world diseases threatening human health, and seriously affecting the quality of life of patients.

Statistically, about 4.15 million people among 20-79 years of age worldwide suffer from diabetes (prevalence 8.8%) and additionally 3.18 million people suffer from impaired glucose tolerance (previous prevalence 6.7%). China is the first major country of diabetes patients all over the world, the number of patients reaches 1.096 hundred million in 2015, and 130 million people die of diabetes and complications thereof. Meanwhile, according to IDF prediction, if no intervention is added, the number of the global diabetic patients in 2040 years reaches 6.42 hundred million, the number of the early-stage diabetic patients reaches 4.81 million, and the number of the national patients rises to 1.54 million.

Metformin (metformin, abbreviated as Met) or metformin hydrochloride is the first choice for the clinical treatment of type two diabetes. Since 1957, metformin was formally marketed as a hypoglycemic agent for the treatment of type two diabetes. Through half a century of clinical use, more and more studies have confirmed the safety and effectiveness of metformin, and thus, metformin is gradually becoming the first-line drug for the treatment of diabetes. Because of low price, it has become one of the most widely used hypoglycemic drugs in clinical practice at present. In 2017, the Chinese guide for the prevention and treatment of type II diabetes takes metformin as a first-choice drug, and the important position of metformin is highlighted. The guidelines indicate that metformin can be used in combination with insulin secretagogues, α -glucosidase inhibitors, insulin sensitizers, and the like, respectively, in a dual drug. The structural formula of metformin is shown in the following formula A.

Metformin has various mechanisms of action including delaying the uptake of glucose from the gastrointestinal tract, increasing peripheral glucose utilization by increasing insulin sensitivity, and inhibiting excessive gluconeogenesis in the liver and kidney without lowering blood glucose levels in non-diabetic patients, and clinical trials have demonstrated that it can be taken for a long period of time. The most common adverse reactions of metformin are gastrointestinal reactions, manifested as epigastric discomfort, abdominal pain, diarrhea, nausea, vomiting, bloating, weakness, dyspepsia, etc. In addition, lactic acidosis occurs in 10% of diabetic patients after taking metformin. So that a proportion of patients were abandoned due to intolerance to metformin.

Glibenclamide is another common drug for the treatment of diabetes. The glibenclamide is specifically combined with a sulfonylurea receptor on a beta cell membrane to stimulate the beta cell to release insulin. However, chronic administration of glibenclamide can lead to islet atrophy; especially for patients with liver and kidney insufficiency, insulin dependent diabetes mellitus, non-insulin dependent diabetes mellitus with ketoacidosis, coma, severe burn, infection, trauma, and leukopenia. In addition, glibenclamide also causes the above-mentioned gastrointestinal reactions similar to metformin.

More importantly, metformin and glyburide are also the most common dual drugs for the treatment of type ii diabetes. The above adverse effects, which lead to a fraction of patients who are intolerant of metformin and/or glyburide therapy.

Therefore, the search for new specific therapeutic drugs, which reduce the side effects of metformin and/or glyburide while reducing blood glucose, is particularly urgent.

Disclosure of Invention

The inventor of the invention finds that the cannabidiol can effectively inhibit type I and type II diabetes and reduce adverse reactions caused by metformin and the like. The following invention is thus provided:

one aspect of the present invention relates to a pharmaceutical composition comprising:

cannabidiol and/or a pharmaceutically acceptable salt or ester thereof; and

one or more hypoglycemic agents;

optionally, the pharmaceutical composition further comprises one or more pharmaceutically acceptable excipients.

In one or more embodiments of the invention, the pharmaceutical composition, wherein the hypoglycemic agent is selected from the group consisting of:

biguanide hypoglycemic agents, sulfonylurea secretagogues, α -glucosidase inhibitors, thiazolidinedione derivative sensitizers, anthranilic acid derivative secretagogues, GLP-1 receptor agonists, DPP-4 enzyme inhibitors, insulin and insulin analogs;

preferably, the biguanide hypoglycemic agent is metformin or a pharmaceutically acceptable salt thereof, such as metformin hydrochloride;

preferably, the sulfonylurea secretagogue is selected from: glibenclamide, glibornuride, glimepiride, gliclazide, glipizide, and gliquidone.

metformin, metformin hydrochloride and glibenclamide.

In one or more embodiments of the present invention, the pharmaceutical composition comprises cannabidiol and/or pharmaceutically acceptable salts or esters thereof, one or more hypoglycemic agents, and one or more pharmaceutically acceptable excipients.

In one or more embodiments of the present invention, the pharmaceutical composition, wherein the weight ratio of the cannabidiol and/or pharmaceutically acceptable salt or ester thereof to the hypoglycemic agent is:

(10) to (1), (10). Here and in the following, if not specified otherwise, the weight ratio refers to the ratio of the total weight of cannabidiol and/or a pharmaceutically acceptable salt or ester thereof to the total weight of all hypoglycemic agents.

In one or more embodiments of the invention, the pharmaceutical composition wherein the cannabidiol and/or the pharmaceutically acceptable salt or ester thereof is present in an amount of 5-500mg, 10-300mg, 20-200mg, 50-150mg, 50-100mg, 100-300mg, 100mg, 200mg, 300mg, 400mg or 500mg by mass.

In one or more embodiments of the invention, the pharmaceutical composition is for use in the treatment and/or prevention of a condition or symptom selected from the group consisting of:

diabetes (e.g., type one or type two diabetes), hyperlipidemia, adverse reactions due to metformin or adverse reactions due to glyburide;

preferably, the diabetes is accompanied by at least one of the following conditions or symptoms: hepatic and renal insufficiency, ketoacidosis, coma, severe burn, infection, trauma, leukopenia, hyperlipidemia, retinopathy or chronic renal failure;

preferably, the metformin-induced adverse reaction is at least one of the following conditions or symptoms resulting from administration of metformin: epigastric discomfort, abdominal pain, diarrhea, nausea, vomiting, bloating, weakness, dyspepsia or lactic acidosis;

preferably, the adverse reaction caused by the glibenclamide is at least one of the following symptoms or symptoms caused by taking the glibenclamide: epigastric discomfort, abdominal pain, diarrhea, nausea, vomiting, bloating, weakness, dyspepsia, lactic acidosis or islet atrophy;

preferably, the cannabidiol and/or a pharmaceutically acceptable salt or ester thereof is administered in a dose of 0.1-50mg, 0.5-30mg, 0.5-20mg, 0.5-10mg, 0.5-5mg, 1-10mg, 2-8mg, 3-7mg, 4-6mg, 4mg, 4.5mg, 5mg, 5.5mg or 6mg per kg body weight per day.

The pharmaceutical composition may be formulated in any pharmaceutically acceptable dosage form including: tablets, sugar-coated tablets, film-coated tablets, enteric-coated tablets, capsules, hard capsules, soft capsules, oral liquids, buccal agents, granules, electuary, pills, powders, ointments, pellets, suspensions, powders, solutions, injections, suppositories, ointments, plasters, creams, sprays, drops, patches; oral dosage forms are preferred, such as: capsule, tablet, oral liquid, granule, pill, powder, pellet, and unguent. The oral dosage forms may contain conventional excipients such as binders, fillers, diluents, tabletting agents, lubricants, disintegrating agents, coloring agents, flavoring agents and wetting agents, and the tablets may be coated if necessary. Suitable fillers include cellulose, mannitol, lactose and other similar fillers; suitable disintegrants include starch, polyvinylpyrrolidone and starch derivatives such as sodium starch glycolate; suitable lubricants include, for example, magnesium stearate. Suitable pharmaceutically acceptable wetting agents include sodium lauryl sulphate.

Preferably, the pharmaceutical composition is an oral formulation.

Another aspect of the invention relates to a pharmaceutical product comprising an individually packaged first pharmaceutical formulation and an individually packaged second pharmaceutical formulation, wherein:

the first pharmaceutical formulation comprises cannabidiol and/or a pharmaceutically acceptable salt or ester thereof,

the second pharmaceutical formulation comprises one or more hypoglycemic agents;

optionally, the first pharmaceutical preparation and/or the second pharmaceutical preparation further comprise one or more pharmaceutically acceptable excipients.

In one or more embodiments of the invention, the pharmaceutical product, wherein the hypoglycemic agent is selected from the group consisting of:

metformin, metformin hydrochloride and glibenclamide.

In one or more embodiments of the invention, the pharmaceutical product,

wherein the pharmaceutical product consists of a first pharmaceutical formulation and a second pharmaceutical formulation,

wherein the first pharmaceutical preparation consists of cannabidiol and/or pharmaceutically acceptable salts or esters thereof, and one or more pharmaceutically acceptable auxiliary materials;

wherein the second medicinal preparation consists of one or more hypoglycemic agents and one or more pharmaceutically acceptable auxiliary materials.

In one or more embodiments of the invention, the pharmaceutical product is wherein the weight ratio of cannabidiol and/or a pharmaceutically acceptable salt or ester thereof to the hypoglycemic agent is:

(10) to (1), (10).

In one or more embodiments of the invention, the pharmaceutical product is one wherein,

the cannabidiol and/or pharmaceutically acceptable salt or ester thereof has a mass of 5-500mg, 10-300mg, 20-200mg, 50-150mg, 50-100mg, 100-300mg, 100mg, 200mg, 300mg, 400mg or 500mg.

In one or more embodiments of the invention, the pharmaceutical product is for use in the treatment and/or prevention of a condition or symptom selected from the group consisting of:

preferably, the metformin-induced adverse reaction is at least one of the following conditions or symptoms resulting from administration of metformin: upper abdominal discomfort, abdominal pain, diarrhea, nausea, vomiting, bloating, weakness, dyspepsia or lactic acidosis;

A further aspect of the invention relates to the use of a pharmaceutical composition according to any of the invention or a pharmaceutical product according to any of the invention in the manufacture of a medicament for the treatment and/or prevention of a condition or symptom selected from:

preferably, the adverse reaction caused by the glibenclamide is at least one of the following symptoms or symptoms caused by taking the glibenclamide: epigastric discomfort, abdominal pain, diarrhea, nausea, vomiting, bloating, weakness, dyspepsia, lactic acidosis or islet atrophy.

A further aspect of the invention relates to the use of cannabidiol and/or a pharmaceutically acceptable salt or ester thereof in the manufacture of a medicament for the treatment and/or prevention of a condition or symptom selected from:

diabetes (e.g., type one or type two diabetes), hyperlipidemia, diarrhea, elevated lactic acid, adverse reactions due to metformin or adverse reactions due to glyburide;

A further aspect of the present invention relates to cannabidiol and/or a pharmaceutically acceptable salt or ester thereof for use in the treatment and/or prevention of a condition or symptom selected from:

preferably, cannabidiol and/or a pharmaceutically acceptable salt or ester thereof is administered in an amount of 0.1-50mg, 0.5-30mg, 0.5-20mg, 0.5-10mg, 0.5-5mg, 1-10mg, 2-8mg, 3-7mg, 4-6mg, 4mg, 4.5mg, 5mg, 5.5mg or 6mg per kg body weight per day.

Yet another aspect of the invention relates to a method for the treatment and/or prevention of a disorder or condition selected from the group consisting of:

diabetes (e.g., type one or type two diabetes), hyperlipidemia, adverse reactions caused by metformin or adverse reactions caused by glyburide;

preferably, the adverse effect caused by metformin is at least one of the following conditions or symptoms caused by metformin administration: epigastric discomfort, abdominal pain, diarrhea, nausea, vomiting, bloating, weakness, dyspepsia or lactic acidosis;

Yet another aspect of the present invention relates to a method for the treatment and/or prevention of a condition or symptom selected from the group consisting of:

Yet another aspect of the present invention relates to a method of treatment and/or prevention of a condition or symptom selected from the group consisting of:

preferably, the cannabidiol and/or a pharmaceutically acceptable salt or ester thereof and the hypoglycemic agent are administered simultaneously or non-simultaneously;

It is noted that the dosage and method of administration of the active ingredient cannabidiol depends on a number of factors including the age, body weight, sex, physical condition, nutritional status, the strength of the activity of the compound, the time of administration, the metabolic rate, the severity of the condition, and the subjective judgment of the treating physician.

Some terms related to the present invention are explained below.

In the present invention, the term "Cannabidiol" (Cannabodiol, abbreviated CBD) is one of the cannabinoids, and has the following formula I:

the cannabidiol, i.e., the compound of formula I, may be purchased commercially (e.g., from Sigma, etc.) or synthesized by known techniques using commercially available starting materials. After synthesis, the product can be further purified by means of column chromatography, liquid-liquid extraction, molecular distillation or crystallization. Cannabidiol may also be extracted from industrial cannabis at any time.

In some embodiments of the present invention, the pharmaceutically acceptable salt of cannabidiol may be a salt of Cannabidiol (CBD) with sodium hydroxide, potassium hydroxide, calcium hydroxide, magnesium hydroxide, aluminum hydroxide, lithium hydroxide, zinc hydroxide, barium hydroxide, ammonia, methylamine, dimethylamine, diethylamine, picoline, ethanolamine, diethanolamine, triethanolamine, ethylenediamine, lysine, arginine, ornithine, choline, N' -benzhydrylethylenediamine, chloroprocaine, diethanolamine, procaine, N-benzylphenethylamine, N-methylglucamine piperazine, tris (hydroxymethyl) -aminomethane, or the like.

In some embodiments of the invention, the pharmaceutically acceptable ester of cannabidiol may be cannabidiol and one C ₀ -C ₆ Monoesters of alkylcarboxylic acids, also cannabidiol, with two identical or different C _0- C ₆ Diesters of alkyl carboxylic acids, said C ₀ -C ₆ The alkyl carboxylic acid may be a straight chain alkyl carboxylic acid, a branched alkyl carboxylic acid or a cycloalkyl carboxylic acid, e.g. HCOOH, CH ₃ COOH、CH ₃ CH ₂ COOH、CH ₃ (CH ₂ ) ₂ COOH、CH ₃ (CH ₂ ) ₃ COOH、CH ₃ (CH ₂ ) ₄ COOH、(CH ₃ ) ₂ CHCOOH、(CH ₃ ) ₃ CCOOH、(CH ₃ ) ₂ CHCH ₂ COOH、(CH ₃ ) ₂ CH(CH ₂ ) ₂ COOH、(CH ₃ ) ₂ CH(CH ₃ )CHCOOH、(CH ₃ ) ₃ CCH ₂ COOH、CH ₃ CH ₂ (CH ₃ ) ₂ CCOOH, cyclopropanecarboxylic acid, cyclobutanecarboxylic acid, cyclopentanecarboxylic acid.

The term "type one diabetes" has the meaning well known to those skilled in the art, its original name being insulin dependent diabetes mellitus, which occurs mostly in children and adolescents, but also at various ages, mainly autoimmune diseases, accounting for approximately 5% of all diabetes. The onset of the disease is rapid, the insulin in the body is absolutely insufficient, ketoacidosis is easy to occur, the satisfactory curative effect can be obtained only by using the insulin treatment, otherwise, the life is threatened.

The term "type II diabetes" has a meaning well known to those skilled in the art, and is primarily adult-onset diabetes, which occurs after 35 to 50 years of age and accounts for more than 90% of diabetic patients. The insulin producing ability of the patients with type II diabetes is not completely lost, and some patients even produce too much insulin, but the utilization effect of the insulin is poor, so that the insulin in the patients is relatively deficient, and the utilization efficiency of the insulin can be improved by oral medicines. Some patients still require insulin therapy to the end.

The term "effective amount" refers to a dose that achieves treatment, prevention, alleviation and/or amelioration of a disease or disorder described herein in a subject.

The term "subject" can refer to a patient or other animal, particularly a mammal, e.g., a human, dog, monkey, cow, horse, etc., that receives a composition of the invention to treat, prevent, ameliorate, and/or alleviate a disease or disorder described herein.

The term "disease and/or disorder" refers to a physical condition of the subject that is associated with the disease and/or disorder of the present invention.

In the present invention, the first and second pharmaceutical preparations are only for clarity and have no sequential meaning, if not otherwise specified.

Advantageous effects of the invention

The invention achieves one or more of the following technical effects:

(1) CBD can significantly inhibit type one and type two diabetes in mammals (e.g., mice or humans), significantly increase insulin sensitivity; the administration effect is better than that of the metformin alone;

(2) The CBD and metformin combined drug can remarkably reduce the hyperlipidemia and hypercholesterolemia induced by STZ and high-fat diet, and can increase the secretion of insulin;

(2) CBD can remarkably inhibit diarrhea and lactic acidemia caused by metformin;

(3) CBD was also able to significantly reduce total urine protein in doxorubicin-induced renal failure mice.

Drawings

FIG. 1A: fasting plasma glucose results were obtained for each group of mice.

FIG. 1B: results of mouse Glucose Tolerance Test (GTT).

FIG. 1C: results of mouse insulin resistance test (ITT).

FIG. 2A: mice fasting plasma glucose.

FIG. 2B: results of mouse Glucose Tolerance Test (GTT).

FIG. 2C: results of mouse insulin resistance test (ITT).

FIG. 3: relative blood glucose values after administration treatment of DB/DB mice.

FIG. 4A: results of day 1 diarrhea index induction in mice.

FIG. 4B: results of day 2 mice diarrhea index induction.

FIG. 4C: results of induction of day 3 mouse diarrhea index.

FIG. 4D: results of day 4 mice diarrhea index induction.

FIG. 4E: results of induction of day 5 mouse diarrhea index.

FIG. 4F: results of day 6 mice diarrhea index induction.

FIG. 4G: results of induction of day 7 mouse diarrhea index.

FIG. 5: the blood of the mice was tested for lactate concentration.

FIG. 6A: serum cholesterol insulin levels in mice.

FIG. 6B: triglyceride levels in the serum of mice.

FIG. 6C: high density cholesterol levels in the serum of mice.

FIG. 6D: low density cholesterol levels in the serum of mice.

FIG. 6E: insulin levels in the serum of mice.

FIG. 7 is a schematic view of: CBD reduces total protein levels in urine.

FIG. 8A: fasting plasma glucose values in 3 diabetic volunteers after CBD treatment.

FIG. 8B: after CBD treatment, 3 diabetic volunteers had glycated hemoglobin levels.

Detailed Description

Embodiments of the present invention will be described in detail below with reference to examples, but those skilled in the art will appreciate that the following examples are only illustrative of the present invention and should not be construed as limiting the scope of the present invention. The examples, in which specific conditions are not specified, were carried out according to conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are conventional products which are commercially available, and are not indicated by manufacturers.

CBD, available from the Hanyi Biotechnology (Beijing) Ltd;

metformin, available from aladine (formula a as above);

insulin, purchased from school hospitals of chinese agriculture university;

streptozotocin (STZ), purchased from alatin.

C57 male mouse, purchased from vindolizhihua;

ICR female mouse, purchased from vindolite;

kunming white male mouse, purchased from Wintonlihua.

Example 1: establishment of mouse type I diabetes model and drug treatment

1) Conventional type I diabetes model induction method (multiple small dose induction method)

Using 4-6 week old C57 male mice as experimental animals, after the mice are fasted for 10-12h, injecting 50mg/kg Streptozotocin (STZ) into the abdominal cavity, continuously injecting for 5 days, and detecting whether the detection of the fasting blood sugar value of the mice on the 6 th day and the 10 th day is successful; successfully inducing with fasting blood glucose of more than 11.1 mmol/L; applied to the second to sixth groups in the following step 2).

2) Experimental grouping and administration

The mice were randomly divided into 6 groups of 10 mice each:

a first group: feeding common feed to the blank control group, and performing intraperitoneal injection of buffer solution at a dosage of 100 μ l/group for 5 days, wherein the administration is performed for 1 time per day;

wherein the buffer solution is 0.1mol/L and the citric acid buffer solution with pH of 4.2, and is sterilized by suction filtration;

second group: a diabetes model group, which is fed with common feed and injected with streptozotocin in the abdominal cavity;

third group: feeding common feed to a CBD prevention group, carrying out intraperitoneal injection of streptozotocin, and inducing that the CBD administration is carried out by intragastric administration for 1 time every day at 50mg/kg on the first day;

and a fourth group: feeding common feed to a CBD treatment group, carrying out intraperitoneal injection of streptozotocin, and after successful induction, carrying out CBD administration by intragastric administration for 1 time every day at a dose of 50mg/kg;

and a fifth group: feeding a positive drug control group with common feed, injecting streptozotocin into abdominal cavity, and starting injecting insulin 0.2U/kg after induction is successful, wherein administration is carried out for 1 time every day;

a sixth group: the metformin group was fed with ordinary feed, streptozotocin was intraperitoneally injected, and after induction was successful, metformin administration by intragastric gavage was started, 50mg/kg, 1 administration per day.

3) Sugar tolerance test (GTT) and insulin resistance test (ITT)

Testing fasting blood glucose of each group of mice every three to four weeks; one month after administration (model stabilization), a Glucose Tolerance Test (GTT) was performed, the mice were fasted for 10h, and based on the body weight (10. Mu.l/g), the blood glucose levels of the mice before (0 min) injection and at the time points 15min, 30min, 60min and 120min after injection were measured, respectively.

An insulin resistance test (ITT) was performed one week after GTT, mice were fasted for 10h, and were intraperitoneally injected with 0.75U/kg insulin, and blood glucose levels were measured at time points of 15min, 30min, 60min, and 120min before (0 min) and after injection, respectively.

4) Results of the experiment

As shown in fig. 1A, 1B and 1C, respectively.

As can be seen in fig. 1A, STZ was able to induce a mouse model of diabetes; both the CBD prevention group and the CBD treatment group were able to significantly reduce the STZ-induced blood glucose rise from 10 days later. In addition, the CBD prevention group was more effective than the CBD treatment group.

As can be seen from the GTT results of fig. 1B, CBD was able to significantly enhance the regulation of glucose in mice. In addition, the CBD prevention group was more effective than the CBD treatment group.

As can be seen from the ITT results in fig. 1C, CBD significantly enhanced insulin sensitivity in mice, modulated hyperglycemia, and reduced symptoms of type one diabetes. In addition, the CBD prevention group was more effective than the CBD treatment group.

In addition, metformin should have no effect on type one diabetes mellitus, and metformin is not clinically used as a clinical drug for treating type one diabetes mellitus.

Example 2: establishment of mouse type II diabetes model and drug treatment

1) Type II diabetes model induction method (multiple small dose induction method and high fat diet induction method)

The experimental animal uses C57 male mice with the age of 4-6 weeks, the mice are fed with high-fat feed, after the mice are fasted for 10-12h, 50mg/kg Streptozotocin (STZ) is injected into the abdominal cavity, the continuous injection is carried out for 5 days, and whether the detection of the fasting blood sugar value of the mice on the 6 th day and the 10 th day is successful or not is detected (the success of the induction is detected by taking fasting blood sugar larger than 11.1mmol/L, and the success of the induction is found on the 32 th day in the experiment); day 6, the dose of streptozotocin injected into the abdominal cavity of mice with substandard blood sugar (without induction success) on day 8 is 80mg/kg; applied to the second group to the eighth group in the following step 2).

The high-fat feed is formed by adding 60% of fat on the basis of common feed.

2) Experimental grouping and administration

The mice were randomly divided into 8 groups of 10 mice each:

a first group: feeding high-fat feed and injecting buffer solution into abdominal cavity of blank control group at dosage of 100 μ l/body, administering 1 time per day for 5 days;

second group: in the diabetes model group, high-fat feed is fed, and streptozotocin is injected in the abdominal cavity;

third group: CBD prevention group, feeding high fat feed, injecting streptozotocin in abdominal cavity, inducing the first day to start CBD administration through intragastric gavage, 50mg/kg, 1 time per day;

and a fourth group: CBD treatment group, high-fat diet, intraperitoneal injection of streptozotocin, CBD administration (50 mg/kg) by gavage daily (day 32) after successful induction;

and a fifth group: metformin group, feed high-fat feed, intraperitoneal injection streptozotocin, on day 32, carry on metformin administration, 50mg/kg, administer 1 time per day;

a sixth group: metformin plus CBD group, high-fat diet, injected streptozotocin in abdominal cavity, and gavage administration (metformin 50mg/kg + CBD 50 mg/kg) every day beginning on day 32;

a seventh group: feeding a high-fat feed to a metformin and Glibenclamide (Glibenclamide, glibenclamide for short) group, carrying out intraperitoneal injection on streptozotocin, and carrying out intragastric administration (50 mg/kg of metformin and 0.758mg/kg of Glibenclamide) every day after induction is successful;

group eight: the glibenclamide and CBD group is fed with high-fat feed, streptozotocin is injected into the abdominal cavity, and the stomach filling administration is started every day after the induction is successful (Gli 0.758mg/kg + CBD 50 mg/kg).

3) Sugar tolerance test (GTT) and insulin resistance test (ITT)

One month after administration (after the type two diabetes model was stabilized), a Glucose Tolerance Test (GTT) was performed, and the mice were fasted for 10 hours, and blood glucose levels of the mice at the time points of 15min, 30min, 60min, and 120min before (0 min) and after injection were measured, respectively, based on the body weight (10 μ l/g) of intraperitoneally injected 20% (w/w) glucose.

One month after administration (after stabilization of the model), an insulin resistance test (ITT) was performed, the mice were fasted for 10h, and the mice were intraperitoneally injected with 0.75U/kg insulin, and blood glucose values at the time points of 15min, 30min, 60min, and 120min before (0 min) injection and after injection were respectively measured.

4) The experimental results are shown in fig. 2A, 2B and 2C, respectively.

From the results of fig. 2A, it can be seen that STZ and high fat diet can cause blood glucose elevation in mice, and CBD can significantly reduce blood glucose elevation induced by STZ and high fat diet.

As can be seen from the results of the glucose tolerance test in mice shown in FIG. 2B, CBD can improve the self-glucose regulation induced by STZ and high-fat diet, and better control blood sugar.

As shown in FIG. 2C, the results of the insulin resistance test in mice show that CBD can improve the sensitivity of mice to insulin and regulate their blood sugar. Alleviating the symptoms of type II diabetes.

As can be seen from fig. 2A to 2C, the blood glucose of the mice of the CBD and metformin combination (group six) was significantly lower than that of the other groups, and both the glucose tolerance and insulin sensitivity of the mice of this group were significantly improved.

In addition, fig. 2A to 2C all show that the CBD prevention group was more effective than the CBD treatment group.

Example 3: effect study of CBD and metformin combined use on type II diabetes

To investigate the effect of the combined use of CBD and metformin, the present inventors bought adult DB/DB mice (spontaneous type two diabetic mice, purchased from the model animals center of Nanjing university) randomly divided into 5 groups of 6 animals each:

the first group was a control group, which was not treated;

the second group was CBD single dose group (50 mg/kg);

the third group was a metformin low dose group (100 mg/kg, converted with reference to the lowest dose administered to humans, administered to mice =9.1X dose administered to humans);

the fourth group is a combination of metformin low dose (100 mg/kg) and CBD (50 mg/kg);

the fifth group was the metformin high dose group (250 mg/kg, converted with reference to the highest dose administered to humans, and the dose administered to mice =9.1X dose administered to humans).

The third and fifth groups were positive controls of the treatment group, the third group was a metformin low dose group, the fifth group was a metformin high dose group, and the patients initially received a low dose of metformin therapy in clinical practice, and then gradually increased the amount of metformin in accordance with the change in blood glucose, but not more than the maximum amount, and the experiment of this example was conducted in order to demonstrate that the effect of the combination of CBD and metformin is superior to that of the low dose of metformin and the high dose of metformin.

The above 5 groups of administration routes are intragastric administration once a day.

The blood glucose of each group of mice was measured 4 weeks after the administration of each group, and the results of the relative blood glucose of each group of mice are shown in FIG. 3.

The results showed that the blood glucose dropped most rapidly and most significantly in the CBD and metformin combination (fourth group) mice during the dosing period. The fasting blood glucose of mice in the CBD and metformin combination group was significantly lower than that in the metformin alone treatment group (third and fifth groups) and also lower than that in the CBD alone treatment group (second group) after 5 weeks of administration.

The results show that the CBD and the combination of the CBD and the metformin can reduce the blood sugar of DB/DB mice, and the combination of the CBD and the metformin has better effect.

Example 4: establishment of mouse diarrhea model and drug treatment

1) Mouse diarrhea model induction method (senna leaf induction mouse diarrhea method)

The experimental animals are 4-6 weeks old ICR female mice, the mice are fed with normal diet and water, 8% (w/w) senna leaf solution is adopted to perfuse the mice with stomach, 0.2 ml/mouse is used, and the diarrhea index of the mice is counted every other half hour for 6 hours. Senna (Folium Sennae) was purchased from the same Tang pharmacia, beijing, and 8 parts by weight of senna and 92 parts by weight of water were added to an erlenmeyer flask and boiled for 10min to obtain an 8% (w/w) senna solution.

2) Experimental grouping and administration

The mice were randomly divided into 7 groups of 10 mice each:

the first group irrigated mice with equal volumes of water per day;

the second group infused mice with 150mg/kg metformin per day;

in the third group, mice were filled with 50mg/kg CBD per day;

the fourth group perfused mice with 8% senna leaf solution every day;

in the fifth group, mice were filled with a mixture of 8% senna leaves and 150mg/kg metformin per day;

in the sixth group, mice were filled with a mixture of 8% senna leaves and 50mg/kg CBD per day;

in the seventh group, mice were filled with a mixture of 8% senna leaves 150mg/kg metformin and 50mg/kg CBD daily.

3) Detection and statistical method

The diarrhea index of each mouse was counted every half hour 4 times (1 time every 6 hours) every day, starting the time after gavage.

Diarrhea index = stool rarity rate + stool rarity grade

Wherein, the first and the second end of the pipe are connected with each other,

the diarrhea rate is as follows: number of stool dilution animals/total animals in the group 100%;

the excrement dilution rate is as follows: number of stool excreted per animal/total stool;

diluting the excrement: the hardness of the thin stool is expressed as a measure of how thin the stool forms a stain on the filter paper. As shown in table 1 below.

Table 1: toilet rarity grade

4) Results of the experiment

As shown in fig. 4A-4G, respectively.

Fig. 4A to 4G are the results of inducing the index of diarrhea in mice from the first day to the seventh day. From the statistical results of the diarrhea indexes of the groups of mice shown in fig. 4A to 4G, it can be seen that senna can cause diarrhea in the mice, metformin can aggravate the senna-induced diarrhea in the mice, and CBD can significantly slow down the senna-induced diarrhea in the mice as well as the metformin-induced and aggravated diarrhea in the mice.

Example 5: establishment of mouse lactic acidosis model and drug treatment

1) Mouse lactic acidosis model induction method

Experimental animals 4-6 weeks old ICR male mice were used for three times of induction, each time with metformin gavage (260 mg/kg), and the interval of 3 gavages was 12 hours.

2) Experimental groups and dosing

The experiment randomly divided the mice into seven groups of 5 mice each:

a first group: control group, no treatment;

second group: irrigating stomach and supplying water every day;

third group: feeding CBD 1 time per day at a dose of 50mg/kg;

and a fourth group: on the first day, 260mg/kg of metformin is administrated by intragastric administration in the morning and evening, on the next morning, 260mg/kg of metformin is administrated by intragastric administration, and on the next afternoon, samples are collected;

a fifth group: the first day, morning and evening with 260mg/kg metformin by gavage, and the second day, morning with 260mg/kg metformin by gavage, wherein 50mg/kg CBD is administered at the same time as each administration of metformin by gavage; collecting samples at four noon on the next day;

a sixth group: injecting 260mg/kg of metformin into a tail vein, injecting once every one hour for four times, and collecting a sample one hour after the fourth injection;

a seventh group: 50mg/kg CBD was administered by gavage half an hour before tail vein injection, and 260mg/kg metformin was injected half an hour after tail vein injection, once every other hour, four times for total injection, and one hour after the fourth injection for sample collection.

3) Experiment design: blood is collected before treatment to detect the concentration of lactic acid, and blood of each group of mice is collected after treatment to detect the concentration of lactic acid.

4) The results of the experiment are shown in FIG. 5.

The results indicate that metformin can induce lactate anemia in mice, while CBD can significantly inhibit metformin-induced elevation of lactate in blood.

Example 6: study of the combination of CBD and metformin on the reduction of hyperlipidemic induced blood lipid levels in mice

1) Diabetes model induction method (multiple small dose induction method)

Experimental animals are male Kunming mice with 4-6 weeks old, after the mice are fasted for 10-12h, 50mg/kg Streptozotocin (STZ) is injected into the abdominal cavity, the injection is continuously carried out for 5 days, and whether the detection of the fasting blood sugar value of the mice on the 6 th day and the 10 th day is successful or not is detected; the fasting blood glucose is more than 11.1mmol/L as successful induction; applied to the second to fifth groups in the following step 3).

2) High-fat induction method

Mice were fed a high fat diet containing 60% of fat on a regular diet basis during the test period, and mice in the second to fifth groups were all fed a high fat diet.

3) Experimental grouping and administration

The mice were randomly divided into 5 groups of 10 mice each:

wherein the buffer solution is 0.1mol/L and the pH value is citric acid buffer solution of 4.2, and the citric acid buffer solution is sterilized by suction filtration;

third group: in the metformin low-dose group, high-fat feed is fed, streptozotocin is injected in the abdominal cavity, and low-dose metformin administration is induced to start by intragastric administration on the first day, wherein the administration is 100mg/kg and is performed for 1 time every day;

and a fourth group: the metformin low dose + CBD group is used for intragastric administration of the metformin with low dose, 100mg/kg is administered for 1 time per day; adding gavage CBD,50mg/kg, 1 time per day, feeding high-fat feed, injecting streptozotocin into abdominal cavity, inducing the first day to start low-dose metformin and CBD administration through gavage;

and a fifth group: in the metformin high dose group, 250mg/kg was administered 1 time per day, and high-dose metformin administration by intragastric gavage was started the first day after feeding high-fat diet and induction of streptozotocin injection into the abdominal cavity. The fifth group served as the positive control treatment group.

4) Detection method

After two months of drug treatment, mice were sacrificed, blood was collected from the heart, and serum was taken and tested for cholesterol, triglyceride, high density cholesterol, low density cholesterol and insulin levels, respectively.

5) Results of the experiment

As shown in fig. 6A to 6E, respectively.

As can be seen from fig. 6A to 6E, the combination of CBD and metformin can significantly reduce the hyperlipidemia and hypercholesterolemia induced by STZ and high fat diet, and can increase insulin secretion. In addition, figure 6E shows that metformin administered alone does not increase insulin secretion.

Example 7: CBD significantly reduced total urine protein levels in doxorubicin-induced chronic renal failure mice

According to a conventional method for inducing a kidney injury model, a doxorubicin-induced chronic renal failure mouse model is constructed. Total protein levels in urine were measured 14 days after CBD treatment. The specific method comprises the following steps:

1) Diabetic nephropathy model induction method (multiple small dose induction method)

The experimental animal uses Kunming white male mice with the age of 4-6 weeks, after the mice are fasted for 10-12h, 50mg/kg Streptozotocin (STZ) is injected into the abdominal cavity, the injection is continuously carried out for 5 days, and whether the detection of the fasting blood sugar value of the mice on the 6 th day and the 10 th day succeeds or not is detected (the fasting blood sugar value is more than 11.1mmol/L and is used for successfully inducing diabetes); on the 7 th day after the onset of diabetes induction, the mice in the renal disease group were intravenously injected with 10mg/kg of doxorubicin; the third to sixth groups in step 2) below.

2) Experimental groups and dosing

Experimental animals were randomly divided into 6 groups of 10 animals each:

a first group: the control group was given physiological saline;

second group: a diabetic group;

third group: diabetes mellitus and CBD group, wherein CBD is administrated by intragastric administration, 50mg/kg, intragastric administration is carried out, and 1 time per day;

and a fourth group: diabetes plus doxorubicin group;

and a fifth group: diabetes mellitus is added with adriamycin and CBD, CBD is administrated by intragastric administration, 50mg/kg, intragastric administration is carried out, and 1 time per day;

a sixth group: the group of diabetes mellitus plus adriamycin plus metformin is administrated by intragastric administration of metformin, 50mg/kg, once a day.

The second and third groups above are for the purpose of measuring the total protein content in urine when chronic renal failure is not induced, but only diabetes is induced.

3) Detection method

After the mouse diabetes model is successfully induced, doxorubicin is injected into the tail vein of the mouse to induce the kidney disease model, the urine of the mouse is collected on the 14 th day of the kidney disease model, and the total protein of the urine is detected.

4) Results of the experiment

As shown in fig. 7.

The results show that CBD can significantly reduce the total urinary protein levels in doxorubicin-induced chronic renal failure mice. Nephropathy is one of the complications of diabetes, and CBD is seen to treat chronic renal failure caused by diabetes.

Example 8: CBD significantly reduces fasting plasma glucose and glycated hemoglobin levels in type II diabetics

There are three volunteers with diabetes clinically: two men weighing about 65 and 70kg, respectively, and aged 47 and 52 years; one woman, weighing about 55kg, was age 74.

Each patient receives 300 mg/day oral CBD treatment, and fasting blood glucose values are detected at 8 weeks, 0, 4 and 8 weeks; glycated hemoglobin values were measured at

weeks

0 and 8. The results are shown in fig. 8A and 8B, respectively.

Fig. 8A shows that fasting glucose was significantly decreased (although not decreased to the normal range) in all three subjects 8 weeks after treatment with CBD;

figure 8B shows that three people had a significant improvement in glycated hemoglobin after 8 weeks of CBD treatment (although not falling into the normal range, i.e., 4% -6%) as before treatment.

Although specific embodiments of the invention have been described in detail, those skilled in the art will appreciate. Many modifications and variations of those details may be made in light of the overall teachings of the disclosure, and such variations are within the scope of the invention. The full scope of the invention is given by the appended claims and any equivalents thereof.

Claims

1. The use of cannabidiol or a pharmaceutically acceptable salt thereof in the manufacture of a medicament for the treatment or prevention of adverse reactions caused by metformin,

wherein the adverse reaction caused by the metformin is at least one of the following symptoms or symptoms caused by taking the metformin: diarrhea or lactic acidosis.