CN112057606B

CN112057606B - Use of FGF19 in a medicament for the treatment and/or prevention of sepsis-induced organ damage

Info

Publication number: CN112057606B
Application number: CN202010959806.9A
Authority: CN
Inventors: 王春霞; 张育才; 唐晓梦
Original assignee: SHANGHAI CHILDREN'S HOSPITAL
Current assignee: SHANGHAI CHILDREN'S HOSPITAL
Priority date: 2020-09-14
Filing date: 2020-09-14
Publication date: 2023-09-19
Anticipated expiration: 2040-09-14
Also published as: CN112057606A

Abstract

The application relates to the field of biological medicine, in particular to application of FGF19 in medicines for treating and/or preventing sepsis-induced organ damage. FGF19 significantly reduces sepsis-induced organ damage, reduces inflammatory responses, and increases cell viability.

Description

Use of FGF19 in a medicament for the treatment and/or prevention of sepsis-induced organ damage

Technical Field

The application relates to the field of biological medicine, in particular to application of FGF19 in medicines for treating and/or preventing sepsis-induced organ damage.

Background

Sepsis (sepsis) refers to a loss of function of a life-threatening organ caused by uncontrolled inflammatory response of the body to an infection, and is mostly caused by infection of the body, and pathogens include viruses, bacteria, fungi, parasites, etc. Patients with severe basic diseases (such as diabetes, chronic obstructive bronchus, leukemia, aplastic anemia and the like), severe wounds, burns, sepsis after major surgery, worsening to septic shock, multiple organ failure and even death are the main causes of death for patients in intensive care units.

Sepsis-related organ dysfunction is a main cause of sepsis prognosis, and has significance in remedying sepsis by improving organ functions in occurrence and development of sepsis and reducing damage degree of the sepsis.

Disclosure of Invention

The present application relates to the use of FGF19 for the manufacture of a medicament for the treatment and/or prevention of sepsis-induced organ damage.

In some embodiments, the organ injury comprises at least one of liver injury, lung injury, kidney injury.

In some embodiments, the organ injury is inflammatory with the organ.

In some embodiments, the sepsis is caused by a microbial infection.

In some embodiments, the sepsis is caused by a gram-negative bacterial infection.

In some embodiments, the sepsis includes early sepsis, severe sepsis, and septic shock.

In some embodiments, the medicament comprises a pharmaceutically acceptable carrier.

In some embodiments, the drug is a lyophilized powder.

In some embodiments, the medicament is an injection.

In some embodiments, the injection is an intravenous injection or an intraperitoneal injection.

The beneficial effects of the application are as follows:

the application adopts intraperitoneal injection LPS to induce and prepare a mouse sepsis model, an experimental group uses FGF19 recombinant protein to carry out tail vein injection, and the influence of FGF19 on ALT, AST, IL-6 and TBIL in the serum of the sepsis model mouse is detected. The results show that after pretreatment, the levels of serum ALT, AST, IL-6 and TBIL are obviously reduced compared with those of LPS treatment groups, and the immunohistochemical staining results also show that the liver of the group pretreated by FGF19 can obviously improve pathological changes such as inflammatory infiltration, and the cell viability of the FGF19 pretreatment group is obviously better than that of the LPS treatment group. The present application thus provides the use of FGF19 for the manufacture of a medicament for the treatment and/or prevention of sepsis-induced organ damage.

Drawings

In order to more clearly illustrate the embodiments of the present application or the technical solutions in the prior art, the drawings that are needed in the description of the embodiments or the prior art will be briefly described, and it is obvious that the drawings in the description below are some embodiments of the present application, and other drawings can be obtained according to the drawings without inventive effort for a person skilled in the art.

FIG. 1 is a comparison of the levels of mouse serum ALT, AST, IL-6 and TBIL treated in different groups in one embodiment of the application;

FIG. 2 is a comparison of H & E staining sections of livers of mice treated in different groups in one embodiment of the present application;

FIG. 3 is a comparison of the results of the CCK-8 detection of living cells from different groups of treated Hepa1-6 cells in one embodiment of the application.

Detailed Description

Reference now will be made in detail to embodiments of the application, one or more examples of which are described below. Each example is provided by way of explanation, not limitation, of the application. Indeed, it will be apparent to those skilled in the art that various modifications and variations can be made to the present application without departing from the scope or spirit of the application. For example, features illustrated or described as part of one embodiment can be used on another embodiment to yield still a further embodiment.

Accordingly, it is intended that the present application cover such modifications and variations as fall within the scope of the appended claims and their equivalents. Other objects, features and aspects of the present application will be disclosed in or be apparent from the following detailed description. It is to be understood by one of ordinary skill in the art that the present discussion is a description of exemplary embodiments only, and is not intended as limiting the broader aspects of the present application.

Fibroblast growth factor 19 (Fibroblast Growth Factor, FGF 19) is a newly discovered metabolic regulator of intestinal secretion, and bile acids stimulate intestinal secretion and expression after entering the intestine. The application proves that the novel intestinal secretion factor FGF19 can be used for treating liver injury under sepsis disease model for the first time.

In some embodiments, the organ injury is inflammatory with the organ.

In some embodiments, the sepsis is caused by a microbial infection.

Infectious microorganisms known to cause sepsis include, but are not limited to: gram-negative bacteria such as E.coli (E.coli), klebsiella, pseudomonas aeruginosa and Enterobacter; gram positive bacteria such as staphylococcus epidermidis and streptococcus faecalis.

The bacterial endotoxin produced by the bacteria produced by gram-negative bacteria is bacterial membrane Lipopolysaccharide (LPS). Lipopolysaccharide endotoxins trigger a range of events that lead to sepsis and ultimately may lead to septic shock and death chain phenomena, specifically in response to the presence of these lipopolysaccharides, the release of inflammatory response mediators including, but not limited to, tumor necrosis factor, interleukins, platelet activators, leukotrienes, prostaglandins, interferons, platelets, bradykinin. Releasing inflammatory mediators causes cell damage, which ultimately leads to cell destruction and ultimately to organ death.

Examples of pharmaceutically acceptable carrier ingredients include binders (syrup, gum arabic, gelatin, sorbitol, tragacanth (tragacanth), polyvinylpyrrolidone and the like), fillers (lactose, sucrose, starch, calcium phosphate, sorbitol, glycine and the like), lubricants (magnesium stearate, talc, polyethylene glycol and the like), disintegrants (starch, microcrystalline cellulose (microcrystalline cellulose) and the like), humectants (sodium dodecyl sulfate (sodium lauryl sulphate) and the like), suspending agents (sorbitol, syrup, methylcellulose, glucose syrup (glucose syrup), gelatin, hydrogenated edible fats and the like), emulsifying agents (lecithin, sorbitol monooleate, gum arabic and the like), nonaqueous carriers (almond oil, fractionated coconut oil or hydrophobic esters of glycerol, propylene glycol, ethanol and the like), preservatives (methylparaben or propylparaben, sorbic acid and the like), fragrances (synthetic flavors, natural flavors and the like), sweeteners (sucrose, sweet stevia, xylitol and the like), pH adjusting agents (sodium bicarbonate, potassium carbonate and the like), powders (thickening agents, dyes, resins and the like), colorants (gum arabic, methyl cellulose (E) and the like), antioxidants (vitamin E and the like).

The agent of the present application can be formulated into tablets, pills, powders, granules, capsules, liquids and solutions, suspensions, emulsions, injections, drops, and the like, as long as the agent can be administered.

In some embodiments, the drug is a lyophilized powder.

In some embodiments, the medicament is an injection.

The preferred mode of administration of the present application is intravenous or intraperitoneal; FGF19 may also be administered by any known administration method, for example, suitably selected from oral, transdermal mucosal administration (e.g., topical, sublingual, intranasal, and rectal), parenteral administration (e.g., via subcutaneous, intramuscular, intra-articular, intravenous, arterial, and inhalation administration, etc.). Thus, specific modes of administration include, but are not limited to, for example, oral, transdermal, mucosal, sublingual, intramuscular, intravenous, intraperitoneal, subcutaneous, and topical.

FGF19 may also be co-administered with drugs such as antibiotics or other cytokines that treat sepsis. The term "co-administration", as used herein, means that the subject is dosed with all types of adjuvants (i.e., compounding): simultaneously (simultaneously) and subsequently [ one drug is taken immediately after the other (administration) or one drug is taken after the other for a moment (administration) ]. FGF19 may be co-administered, for example, with certain antibiotic agents including, but not limited to, penicillins, cephalosporins, ciprofloxacin, quinolone lactam antibiotics, erythromycin, and amino sugars or antibiotics.

Medicaments for intravenous administration are generally in the form of solid sterile compositions. These compositions may also contain additives, in particular mannitol, dextran, hydrolyzed gelatin, sodium citrate, glycine, and the like. Is dissolved in sterilized injectable water or other injectable sterilizing medium.

The human urinary kallidinogenase composition administered by intravenous injection may also be in the form of an aqueous solution. The composition may also contain additives, in particular mannitol, sodium chloride, glucose, etc.

The pharmaceutical formulation is preferably in unit dosage form. In this form, the formulation is subdivided into unit doses containing appropriate quantities of the active component. The unit dosage form may be a packaged formulation containing discrete amounts of formulation, such as packaged tablets, capsules, and powders in vials or ampoules. Furthermore, the unit dosage form may be a capsule, tablet, cachet, or lozenge itself, or it may be the packaging of any of a suitable number of these dosage forms. Tablets or capsules for oral administration and liquids for intravenous administration and continuous infusion are preferred compositions.

According to yet another aspect of the present application, there is also provided a method of treating, preventing sepsis comprising the step of administering to a subject a safe and effective amount of a medicament as described above.

The phrase "safe and effective amount". As used herein, means that the amount of a compound or composition within the scope of reasonable pharmaceutical modulation is large enough to significantly effectively alleviate the symptoms or conditions being treated, but small enough to avoid serious side effects (at a reasonable benefit/risk ratio). The safe and effective amount of the active ingredient in the pharmaceutical compositions used in the methods of the present application will vary with the particular condition being treated, the age and physical condition of the patient being treated, the severity of the disease, the time of treatment, the concurrent treatment conditions, the particular active ingredient being used, the particular pharmaceutically acceptable excipients used, and such factors including the knowledge and skill of the attending physician.

The term "subject" as used herein may refer to a patient or other animal receiving an agent or medicament as described herein for the treatment, prevention, alleviation and/or alleviation of a disease or a disorder or a condition of the present application, which subject includes a warm-blooded animal, such as a mammal, e.g., a primate, and preferably a human. Non-human primates are also individuals. The term individual includes domestic animals such as cats, dogs, etc., domestic animals (e.g., cattle, horses, pigs, sheep, goats, etc.), and laboratory animals (e.g., mice, rabbits, rats, gerbils, guinea pigs, etc.).

In one embodiment of the application, the drug is administered locally or systemically or by a combination of both routes.

The application is further illustrated in the following drawings and specific examples, which are not intended to limit the application in any way. It will be apparent to those skilled in the art that various changes, modifications, substitutions, combinations, and simplifications can be made without departing from the spirit and principles of the application and these are intended to be equivalent arrangements.

Unless specifically stated otherwise, the reagents, methods and apparatus employed in the present application are those conventional in the art. Reagents and materials used in the following examples are commercially available unless otherwise specified.

Unless otherwise indicated, the immunology, biochemistry, chemistry, molecular biology, microbiology, cell biology, genomics, recombinant DNA, etc., employed by this application are within the skill of the art. See Sambrook (Sambrook), friech (Fritsch) and manitis (Maniatis), molecular cloning: laboratory Manual (MOLECULAR CLONING: A LABORATORY MANUAL), edit 2 (1989); the handbook of contemporary molecular biology (CURRENT PROTOCOLS IN MOLECULAR BIOLOGY) (edited by f.m. ausubel (f.m. ausubel) et al, (1987)); series (academic publishing company) of methods in enzymology (METHODS IN ENZYMOLOGY): PCR2 practical methods (PCR 2:A PRACTICAL APPROACH) (M.J. MaxFrisson (M.J. MacPherson), B.D. Black (B.D. Hames) and G.R. Taylor (G.R. Taylor) editions (1995)), harlow and Lane editions (1988) antibodies: laboratory Manual (ANTIBODIES, A LABORATORY MANUAL), animal cell CULTURE (ANIMAL CELL CULTURE) (R.I. French Lei Xieni (R.I. Freshney) eds. (1987)).

Embodiments of the present application will be described in detail below with reference to examples.

Examples

Experimental animals and materials

The C57BL/6 mice used in the application were purchased from Shanghai's mode biological research center. The Hepa1-6 cell line was purchased from the Shanghai cell Bank of the Chinese sciences. FGF19 for use in the present application was purchased from offshore protein technologies Inc., and lipopolysaccharide was purchased from Sigma-Aldrich Inc. CCK-8 is purchased from Shanghai Kogyo Biotech Co.

Experimental method

1. Method of administration in mice

Male C57BL/6j mice were housed in SPF-class animal houses 40 (8 weeks old, 22-25g weight) and after 1 week of adaptive feeding were randomly divided into control (n=6), LPS (n=6), FGF19 (n=6) and LPS+FGF19 (n=6).

(1) FGF19:0.1 mg/(kg.d), tail vein injection, pretreatment for 7 consecutive days;

(2) LPS:5mg/kg, intraperitoneal injection, 24 hours after treatment, blood was taken and levels of alanine Aminotransferase (ALT), aspartic acid Aminotransferase (AST), interleukin-6 (IL-6) and Total Bilirubin (TBIL) were measured.

2. Detecting biochemical indexes of mouse blood

Mice ALT, AST, IL-6 and TBIL levels were measured using a biochemical detector and plotted using "graphpad" software.

3. Cell administration method

The Hepa1-6 cells were seeded in 96-well plates with a cell number of 1 x 10 x 6 cells per well, and the experimental group was divided into control group (n=10), LPS group (n=10), FGF19 group (n=10), lps+fgf19 group (n=10). While blank sets are set.

(1) FGF19:10ng/ml, pretreatment for 48 hours

(2) LPS:1ug/ml, for 24 hours

Detection of Hepa1-6 cell Activity by 4CCK-8 experiment

Four groups of Hepa1-6 cells were assayed for viability using the CCK-8 assay, the drug-containing medium was discarded, fresh 10% FBS-containing DMEM medium was added 100 ul/well, then 10ul of CCK-8 assay was added to each well, incubated for 30 minutes at 37℃in an incubator, and the corresponding absorbance was measured at 450nm using an ELISA. Cell viability = (experimental well absorbance-blank well absorbance)/(control well absorbance-blank well absorbance) ×100% and is plotted using "graphpad" software.

Experimental results

1. Animal serum detection

The levels of ALT, AST, IL-6 and TBIL in the four groups of mice are shown in FIG. 1: the levels of ALT, AST, IL-6 and TBIL were significantly increased after LPS treatment, and the FGF19 pretreatment group, serum ALT, AST, IL-6 and TBIL were significantly reduced compared to the LPS treatment group.

2. Animal H & E staining results

The H & E stained sections of the livers of the four groups of mice are shown in fig. 2: after LPS, inflammatory infiltration of liver tissue is obvious, liver Dou Xiazhai almost disappears, and liver plate is disordered; FGF19 pretreatment can obviously improve pathological changes such as inflammatory infiltration.

Hepa1-6 cell viability

The results of CCK-8 detection of the viable cell count of the four Hepa1-6 cells are shown in FIG. 3, and the viable cell count is significantly reduced after LPS action. The FGF19 pretreatment group had significantly better cell viability than the LPS treatment group.

The technical features of the above-described embodiments may be arbitrarily combined, and all possible combinations of the technical features in the above-described embodiments are not described for brevity of description, however, as long as there is no contradiction between the combinations of the technical features, they should be considered as the scope of the description.

The above examples illustrate only a few embodiments of the application, which are described in detail and are not to be construed as limiting the scope of the application. It should be noted that it will be apparent to those skilled in the art that several variations and modifications can be made without departing from the spirit of the application, which are all within the scope of the application. Accordingly, the scope of protection of the present application is to be determined by the appended claims.

Claims

1. Use of FGF19 for the manufacture of a medicament for the treatment and/or prevention of sepsis-induced liver injury.

2. The use of claim 1, wherein the liver injury is accompanied by liver inflammation.

3. The use of claim 1, wherein the sepsis is caused by a microbial infection.

4. The use of claim 3, wherein the sepsis is caused by a gram-negative bacterial infection.

5. The use of claim 1, wherein the sepsis is early sepsis, severe sepsis, and septic shock.

6. The use of claim 1, wherein the medicament comprises a pharmaceutically acceptable carrier.

7. The use according to claim 1, wherein the medicament is a lyophilized powder.

8. The use according to claim 1, wherein the medicament is an injection.

9. The use according to claim 8, wherein the injection is an intravenous injection or an intraperitoneal injection.