WO2021078246A1 - Composition pharmaceutique pour la prévention ou le traitement de la septicémie, kit, utilisation de celle-ci et procédé de traitement associé - Google Patents

Composition pharmaceutique pour la prévention ou le traitement de la septicémie, kit, utilisation de celle-ci et procédé de traitement associé Download PDF

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WO2021078246A1
WO2021078246A1 PCT/CN2020/123169 CN2020123169W WO2021078246A1 WO 2021078246 A1 WO2021078246 A1 WO 2021078246A1 CN 2020123169 W CN2020123169 W CN 2020123169W WO 2021078246 A1 WO2021078246 A1 WO 2021078246A1
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mitochondria
sepsis
mice
pharmaceutical composition
preventing
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Chinese (zh)
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唐凌峰
吴忆华
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深圳市脉唐生物科技有限公司
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Priority to CN202080002489.XA priority Critical patent/CN114599379A/zh
Publication of WO2021078246A1 publication Critical patent/WO2021078246A1/fr

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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/14—Blood; Artificial blood
    • A61K35/19—Platelets; Megacaryocytes
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K35/00—Medicinal preparations containing materials or reaction products thereof with undetermined constitution
    • A61K35/12—Materials from mammals; Compositions comprising non-specified tissues or cells; Compositions comprising non-embryonic stem cells; Genetically modified cells
    • A61K35/28—Bone marrow; Haematopoietic stem cells; Mesenchymal stem cells of any origin, e.g. adipose-derived stem cells
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K45/00—Medicinal preparations containing active ingredients not provided for in groups A61K31/00 - A61K41/00
    • A61K45/06—Mixtures of active ingredients without chemical characterisation, e.g. antiphlogistics and cardiaca
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P31/00—Antiinfectives, i.e. antibiotics, antiseptics, chemotherapeutics
    • Y—GENERAL TAGGING OF NEW TECHNOLOGICAL DEVELOPMENTS; GENERAL TAGGING OF CROSS-SECTIONAL TECHNOLOGIES SPANNING OVER SEVERAL SECTIONS OF THE IPC; TECHNICAL SUBJECTS COVERED BY FORMER USPC CROSS-REFERENCE ART COLLECTIONS [XRACs] AND DIGESTS
    • Y02—TECHNOLOGIES OR APPLICATIONS FOR MITIGATION OR ADAPTATION AGAINST CLIMATE CHANGE
    • Y02A—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE
    • Y02A50/00—TECHNOLOGIES FOR ADAPTATION TO CLIMATE CHANGE in human health protection, e.g. against extreme weather
    • Y02A50/30—Against vector-borne diseases, e.g. mosquito-borne, fly-borne, tick-borne or waterborne diseases whose impact is exacerbated by climate change

Definitions

  • the invention belongs to the field of biopharmaceuticals, and specifically relates to a pharmaceutical composition, a kit for preventing or treating sepsis, and an application and treatment method thereof.
  • Sepsis is an uncontrolled response of the host to infection and causes life-threatening organ dysfunction. There are approximately 48.9 million sepsis cases worldwide each year, of which 11 million deaths from sepsis, with a mortality rate of 22.5%. There is currently no specific medicine for sepsis, and conventional treatments have limited effects, including: fluid resuscitation, the use of broad-spectrum or narrow-spectrum antibiotics, surgery to remove the source of infection, and "symptomatic treatment.”
  • Fluid resuscitation is to give patients liquids such as crystalloid fluid through Early, goal-directed therapy (EGDT), so that the patient's central venous pressure and other physiological indicators meet the corresponding standards, so as to reduce the mortality of patients with sepsis.
  • EGDT Early, goal-directed therapy
  • the disadvantage of this method is that the dosage is difficult to determine, and the dosage window is narrow. Many clinical studies have shown that EGDT cannot reduce the mortality of patients with sepsis, and excessive fluid can easily cause damage to the kidneys, heart and other organs.
  • Antibiotics are used in the treatment of sepsis in two ways. One is the direct use of broad-spectrum antibiotics in the early stage of sepsis. The disadvantage of this method is that it is prone to produce drug-resistant bacteria and has greater side effects on the human body.
  • the second is to obtain biological evidence through bacterial culture, and then target antibiotic treatment.
  • the shortcomings of this method are obvious. Sepsis needs to be administered as soon as possible, but the microorganisms are cultured for a long time, and the culture results are not necessarily positive, and there are many multi-drug resistant bacteria, and the antibiotic treatment effect is very poor.
  • the common shortcoming of the two methods is that antibiotics can easily further aggravate the disorder of the immune system and cannot prevent sepsis. If sepsis is caused by surgical infection, the lesion can be removed or drained by surgery, but the disadvantage of this method is that it is difficult to determine the source of the infection.
  • Symptomatic treatment of sepsis refers to, for example, the use of glucocorticoids to control blood pressure, insulin to control blood sugar, and oxygen delivery to control blood oxygen content.
  • the main basis of this method is that the patient's physiological indicators are lower or higher than a certain alert value and given corresponding drugs for regulation.
  • the treatment effect of sepsis is not good, and it cannot prevent sepsis. It can only be used as an auxiliary treatment method. .
  • the mitochondria of patients with sepsis are damaged and the utilization of oxygen is low. The excess oxygen easily combines with electrons to form reactive oxygen species (ROS), which increases oxygen stress and exacerbates the condition and prognosis of patients with sepsis.
  • ROS reactive oxygen species
  • the present invention provides a pharmaceutical composition, kit, and application and treatment thereof for preventing or treating sepsis. method.
  • the present invention provides a pharmaceutical composition for preventing or treating sepsis, which includes physiologically active mitochondria as an active ingredient.
  • the present invention also provides a kit for preventing or treating sepsis, which includes the above-mentioned pharmaceutical composition.
  • the present invention also provides the application of mitochondria in the preparation of medicines, pharmaceutical compositions or kits for preventing or treating sepsis.
  • the source of the mitochondria is autologous, allogeneic or heterologous, and combinations thereof.
  • the source of the mitochondria is allogeneic.
  • the mitochondria are separated and extracted from cells or tissues, and the cells include somatic cells, germ cells, stem cells, and combinations thereof, and the tissues include heart, liver, spleen, kidney, brain, and combinations thereof.
  • the mitochondria are extracted and isolated from the tissues of the heart, liver, spleen, kidney and combinations thereof.
  • the pharmaceutical composition further includes a solvent.
  • the concentration of the mitochondria is 0.1 ⁇ g/ml to 900 mg/ml.
  • the solvent in the pharmaceutical composition includes physiological saline, phosphate buffer, culture fluid, tissue fluid, phospholipid or amino acid solution with drug properties, and combinations thereof.
  • the pharmaceutical composition further includes one or more of insulin, antibiotics, antiviral drugs, antifungal drugs, glucocorticoids, and cardiotonic drugs.
  • the present invention also provides a method for preventing or treating sepsis, including the following steps:
  • a pharmaceutical composition including mitochondria having physiological activity as an active ingredient is administered to the patient.
  • the administration modes of the pharmaceutical composition include intravenous injection, arterial injection, intraperitoneal injection, intramuscular injection, subcutaneous injection, intradermal injection, oral administration, sublingual administration, external application, inhalation, and oral, eye, and urinary Reproductive system mucosal administration and combinations thereof.
  • the method for preventing or treating sepsis further includes symptomatic and supportive therapy.
  • the method for preventing or treating sepsis further includes other symptomatic and supportive therapies, and the other symptomatic and supportive therapies include rehydration, cardiotonia, boosting, oxygen inhalation, assisted ventilation, enteral nutrition support, and parenteral nutrition. Support, ECG monitoring and their combination.
  • Mitochondria originated from archaea 1.5 billion years ago, so they have an "infection ability" similar to that of intracellular bacteria.
  • mitochondria can enter cells efficiently.
  • the inventor of the present invention realized that mitochondrial administration can be used as an effective means to treat sepsis, which can directly restore the energy supply level of the patient's body, thereby prolonging the patient's energy supply level. Survival, to achieve the purpose of treatment.
  • the inventors used E.
  • mice sepsis model isolated and extracted mitochondria from other mice, and performed a variety of administration methods on the mouse sepsis model. Mitochondria were applied, and the results showed that the mortality of mice was effectively reduced.
  • Mitochondria are natural medicines with low side effects and can be extracted from allogeneic cells, which will not cause or have only a weak immune rejection reaction when entering the patient's body;
  • Fig. 1 is a fluorescence image of active mitochondria provided in Example 1 of the present invention and mitochondria inactivated by two methods after being stained with MitoTracker Red under a confocal microscope.
  • Figure 2 is a graph showing the survival rate of septic mice constructed by intraperitoneal injection of E. coli by IP administration of mitochondria provided in Example 2 of the present invention.
  • Fig. 3 is a graph showing the survival rate of septic mice constructed by subcutaneous injection of Escherichia coli by IP administration of mitochondria provided in Example 3 of the present invention.
  • Fig. 4 shows the survival rate of the septic mice (male) provided in Example 4 of the present invention after inactivated or activated mitochondria were injected through the tail vein.
  • Fig. 5 is the result of the survival rate of the septic mice (female) provided in Example 4 of the present invention after inactivated or activated mitochondria were injected through the tail vein.
  • Fig. 6 is the result of the survival rate of the septic mice provided in Example 5 of the present invention after inactivated or activated mitochondria after intraperitoneal injection.
  • Fig. 7 is the result of the survival rate of septic mice after intramuscular injection of inactivated or activated mitochondria provided in Example 6 of the present invention.
  • Figure 8 is the result of the survival rate of the septic mice provided in Example 7 of the present invention after inactivated or activated mitochondria after intraperitoneal injection.
  • Fig. 9 is the result of the survival rate of the septic mice provided in Example 8 of the present invention after inactivated or activated mitochondria after intraperitoneal injection.
  • Fig. 10 is the result of the survival rate of the septic mice provided in Example 9 of the present invention after inactivated or activated mitochondria derived from 293T cells were injected through the tail vein.
  • Fig. 11 shows the survival rate of the septic mice provided in Example 10 of the present invention after a single injection of inactivated or activated mitochondria through the tail vein.
  • Figure 12 is the result of the survival rate of septic mice constructed by intraperitoneal injection of mouse fecal solution provided in Example 11 of the present invention after a single intraperitoneal injection of inactivated or activated mitochondria.
  • Figure 13 is the result of the survival rate of the septic mice provided in Example 12 of the present invention after inactivated or activated mitochondria after intraperitoneal injection.
  • Fig. 14 is the result of the survival rate of mice in the case of prior tail vein injection of inactivated or activated mitochondria and then induced sepsis in Example 13 of the present invention.
  • isolated and extracted refers to mitochondria or a composition containing mitochondria (for example, cytoplasm), which has been physically separated or removed from its natural biological environment.
  • the isolated mitochondria or the composition containing mitochondria may be purified during the isolation process, or need not be purified.
  • autologous refers to a biological composition obtained from the same organism.
  • allo refers to organisms of different genotypes in the same genus.
  • heterologous refers to organisms of different genotypes in different species.
  • necrosis refers to the life-threatening organ function damage caused by the imbalance of the host's response to infection, including infections (bacteria, fungi, mycoplasma, parasites, viruses and other infections), host reactions (inflammatory reactions and involving multiple systems Non-immune reactions) and organ dysfunction.
  • mitochondria with physiological activity refers to mitochondria that can perform basic physiological activities, such as energy conversion, and also includes mitochondria that dormant under certain conditions but whose physiological activity is not destroyed.
  • the physiological activity of mitochondria can be defined by the in vitro respiration rate of mitochondria.
  • various in vitro methods for assessing the physiological functions of mitochondria can also be used, including spectrophotometric enzyme determination, bioluminescence measurement of ATP production, MitoTracker staining intensity, JC- 1 Dyeing and so on.
  • various specific antibodies against mitochondrial proteins can be used in immunocytochemistry and Western blot analysis.
  • These antibodies include ATP synthase subunits, cytochrome c and cytochrome c oxidase, PGC-1 and mtTFA.
  • Two-dimensional polyacrylamide gel electrophoresis can be used, followed by Western blotting to analyze the content of mitochondrial protein.
  • the present invention provides a pharmaceutical composition for treating sepsis, which includes physiologically active mitochondria as an active ingredient.
  • the pharmaceutical composition can be administered in advance to achieve the effect of preventing sepsis. Therefore, the present invention also provides a pharmaceutical composition for preventing sepsis, which includes physiologically active mitochondria as an active ingredient.
  • the source of the mitochondria is autologous, allogeneic or xenogeneic, and combinations thereof.
  • the source of the mitochondria is allogeneic.
  • the source of the mitochondria is autologous or heterologous, and combinations thereof.
  • the mitochondria are isolated and extracted from cells or tissues.
  • Mitochondria can be mitochondria obtained from non-human mammals (such as mice, rabbits, pigs, sheep, goats, cattle, and higher primates), or mitochondria obtained from humans. Specifically, mitochondria may be mitochondria isolated from cells or tissues. In a specific embodiment, the cells or tissues are all cells or tissues in vitro.
  • the cell may be any one selected from the group consisting of somatic cells, germ cells, stem cells, and combinations thereof.
  • mitochondria may be mitochondria obtained from somatic cells, germ cells, or stem cells.
  • the mitochondria may be normal mitochondria obtained from cells with normal mitochondrial biological activity.
  • the mitochondria may be mitochondria cultured in vitro.
  • the tissues include heart, liver, spleen, kidney, brain, and combinations thereof.
  • the mitochondria are extracted and isolated from tissues of heart, liver, spleen, kidney, and combinations thereof.
  • the pharmaceutical composition further includes a vehicle.
  • the concentration of the mitochondria is 0.1 ⁇ g/ml to 900 mg/ml.
  • the concentration of the mitochondria may be 0.1 ⁇ g/ml, 0.15 ⁇ g/ml, 0.6 ⁇ g/ml, 1 ⁇ g/ml, 2 ⁇ g/ml, 5 ⁇ g/ml, 10 ⁇ g/ml, 20 ⁇ g/ml, 50 ⁇ g/ml, 100 ⁇ g/ml, 200 ⁇ g/ml, 500 ⁇ g/ml, 1mg/ml, 5mg/ml, 10mg/ml, 20mg/ml, 50mg/ml, 100mg/ml, 300mg/ml, 500mg/ml, 700mg/ ml or 900mg/ml.
  • the solvent in the pharmaceutical composition includes physiological saline, phosphate buffer, culture fluid, tissue fluid, phospholipid or amino acid solution with drug properties, and combinations thereof.
  • the pharmaceutical composition further includes a stabilizer.
  • the stabilizer includes glucose, sucrose, fetal bovine serum, ADP, ATP, amino acids, glycerol, propylene glycol, sodium glycocholate, cholesterol, mannitol, albumin or sodium citrate and combinations thereof.
  • the pharmaceutical composition further includes other active agents with disease prevention or treatment effects.
  • the active agent may be an agent for synergistic prevention or treatment of sepsis, or an agent for prevention or treatment of other diseases.
  • the pharmaceutical composition further includes one or more of antibiotics, antiviral drugs, antifungal drugs, glucocorticoids, insulin, and cardiotonic drugs.
  • Another embodiment of the present invention provides a kit for preventing or treating sepsis, including the pharmaceutical composition as described above.
  • Another embodiment of the present invention provides the use of mitochondria in the preparation of drugs, pharmaceutical compositions or kits for preventing or treating sepsis.
  • the characteristic requirements of the mitochondria as a medicine, pharmaceutical composition or kit for the prevention or treatment of sepsis are consistent with the description in the above-mentioned pharmaceutical composition and kit, and will not be repeated.
  • Another embodiment of the present invention provides a method for preventing or treating sepsis, including the following steps:
  • a pharmaceutical composition including mitochondria having physiological activity as an active ingredient is administered to the patient.
  • the administration mode of the pharmaceutical composition includes intravenous injection, arterial injection, intraperitoneal injection, intramuscular injection, subcutaneous injection, intradermal injection, oral administration, sublingual administration, topical application, inhalation, and oral and eye , Urogenital system mucosal administration and combinations thereof.
  • the method for preventing or treating sepsis further includes other symptomatic and supportive therapies.
  • the other symptomatic and supportive therapies include fluid rehydration, cardiotonic, boosting, oxygen inhalation, assisted ventilation, enteral nutrition support, and intestinal support. External nutritional support, ECG monitoring and their combination.
  • Escherichia coli (DH 5 ⁇ ) was cultured in TB medium and harvested by centrifugation. Prepare a 20% E. coli solution (20% E. coli, 30% glycerol, 50% TB medium, calculated by volume). The same batch of E. coli is frozen and stored at -80°C in the refrigerator. Kunming mice (KM mice) were used to test the toxicity of different batches of E. coli, including male and female mice.
  • Mitochondria were isolated and extracted from the heart, liver, kidney and spleen of Kunming mice. In short, these organs were homogenized in 1x PBS using an electronic homogenizer, and centrifuged at 1015 g for 10 min in a 50 mL centrifuge tube. Subsequently, the supernatant was transferred to a new 50ml centrifuge tube and centrifuged at 14610g for 10min. The supernatant was discarded, the pellets were combined and resuspended in DMEM.1xPS.10% FBS (DMEM is high glucose, containing 1x Pen-Strep and 10% FBS) cell culture medium.
  • DMEM is high glucose, containing 1x Pen-Strep and 10% FBS
  • inactive mitochondria In order to use inactive mitochondria as a negative control, each gram of mitochondrial pellet was resuspended in 5ml PBS and 2.5ml 75% ethanol. Let it stand at room temperature for 5 min, and then centrifuge at 14610 g for 10 min to obtain inactivated mitochondria (IAM). The inactivated mitochondria were then resuspended in DMEM.1xPS.10% FBS cell culture medium at the same concentration as the active mitochondria.
  • IAM inactivated mitochondria
  • E. coli was diluted with 1xPBS and injected into Kunming mice through the intraperitoneal cavity.
  • the dose of E. coli that killed about 50% of Kunming mice within 6 hours Construct a murine sepsis model.
  • mice using the same number of male and female mice weighing 20-26g, by intraperitoneal injection (IP), with 0.1ml/10g 50% mt.DMEM (50% active mitochondria in DMEM.1xPS.10% FBS culture Resuspension in the basal medium) or 50% IAM.DMEM (50% inactivated mitochondria are resuspended in DMEM.1xPS.10% FBS medium) dose administration, record the death of mice every 6 hours, and continue to observe 24 hours.
  • IP intraperitoneal injection
  • Figure 2 shows the construction of a Kunming mouse sepsis model induced by intraperitoneal injection (IP) of Escherichia coli, and it was administered in high glucose DMEM.1xPS.10% FBS by IP or IV methods. 50% active mitochondria diluted in solution (Treatment) or inactivated mitochondria (Control).
  • Figure 3 shows the construction of a Kunming mouse sepsis model induced by subcutaneous injection (SC) of Escherichia coli, and 50% active mitochondria diluted in high glucose DMEM.1xPS.10% FBS solution by intraperitoneal injection (IP) ( Treatment or inactivation of mitochondria (Control).
  • IP intraperitoneal injection
  • Newborn mice extract mitochondria take newborn mice within 1-3 days, sterilize with alcohol, cut into 4-5 cuts and place in a beaker, add 1 ⁇ PBS (phosphate buffered saline solution) at 10mL/piece, and then add according to the volume The final concentration is 5mM EDTA (ethylenediaminetetraacetic acid) and the final concentration is 0.2mM PMSF, homogenized in a homogenizer at 6000rpm for 2min. After homogenization, dispensed into 50mL centrifuge tubes, centrifuged at 1015g, 4°C for 10min. Take the supernatant and centrifuge at 14600g for 10 min at 4°C. The supernatant was discarded, the precipitate was filtered and dried and weighed, and the mitochondrial solution was prepared with the required solution directly or after inactivation.
  • PBS phosphate buffered saline solution
  • Ethanol inactivation In the above mitochondrial extraction step, 14,600g, centrifuged at 4°C for 10 minutes, discard the supernatant, filter the precipitate and weigh it, add 5 times 1 ⁇ PBS according to the weight, and add 75% at the volume of 50%. Mix it with ethanol, leave it at room temperature for 5 minutes, then centrifuge at 14600g for 10 minutes, discard the supernatant, filter the precipitate, and weigh it. According to the weight, use DMEM.1xPS.10% FBS cell culture medium to configure 500mg/mL inactivated mitochondria (Inactivated Mitochondria). , IAM) solution.
  • Freeze-thaw inactivation place the active mitochondrial solution extracted in the above mitochondrial extraction step in a refrigerator at -80°C or room temperature, and freeze and thaw three times repeatedly to obtain an inactivated mitochondrial solution.
  • Mitochondria were extracted from Kunming mouse heart, liver, spleen, kidney and other organs. Active mitochondria were prepared using DMEM.1xPS.10% FBS to prepare 50mg/mL mt.DMEM. Inactivated mitochondria were inactivated by ethanol and three freeze-thaw methods. For inactivation, use DMEM.1xPS.10% FBS to prepare 50mg/mL IAM.DMEM. Using MitoTracker Red staining (dilution ratio 1:5000), MitoTracker Red enters through the mitochondrial membrane potential and remains in the mitochondrial matrix. A strong signal indicates good mitochondrial activity, and no signal indicates loss of mitochondrial membrane potential, which is a manifestation of mitochondrial inactivation.
  • Example 2 is a detailed description of the example in the priority document.
  • the mouse sepsis model is induced by the dose of intraperitoneal injection (IP) of Escherichia coli in Kunming mice with a body weight of 20-26 g that kills about 50% within 6 hours.
  • E. coli was diluted with 1 ⁇ PBS and injected by IP into 24 male Kunming mice of equal weight (12 in the control group and 12 in the treatment group). Mitochondria are extracted from mouse internal organs.
  • IP intraperitoneal injection
  • mt.DMEM 50% active mitochondria resuspended in DMEM.1xPS.10% FBS
  • 500mg/mL IAM.DMEM 50% inactivated mitochondria resuspended in DMEM
  • Death records are recorded every 6 hours for 24 hours.
  • Example 3 is a detailed description of the example in the priority document.
  • the mouse sepsis model is induced by subcutaneous injection (SC) of E. coli in Kunming mice by killing about 50% of the weight of 20-26g in 5 days.
  • E. coli was diluted with 1 ⁇ PBS and injected by SC into 24 male Kunming mice of equal weight (12 in the control group and 12 in the treatment group). Mitochondria are extracted from mouse internal organs.
  • the Kunming mice were randomly divided into two groups (male: 24 in the control group, 24 in the treatment group, female: 8 in the treatment group, 8 in the control group), the weight range of 22-36g, the treatment group and the control group were paired in pairs. The difference is within 1g.
  • the E. coli solution with a concentration of 2mg/0.1mL was injected intraperitoneally with 0.1mL/10g into the mice, and the mice developed sepsis symptoms 2 hours later.
  • the inactivated or active mitochondria extracted from newborn mice were diluted with PBS into a 25mg/mL mitochondrial suspension, and the mice with sepsis symptoms were injected with 0.05mL/10g tail vein, and the control group was injected with IAM.PBS for treatment The group was injected with mt.PBS, and the survival of the mice was continuously observed until the 7th day.
  • the results are shown in Figures 4 and 5.
  • a single injection of 25 mg/mL mitochondria in the tail vein can increase the survival rate of mice by about 17%-25%.
  • mice 34 male Kunming mice were randomly divided into two groups (the treatment group and the control group each had 17), the weight range was 22-36g (the treatment group and the control group were paired, the weight difference was within 1g), and the concentration was 2mg/0.1mL
  • the Escherichia coli strain was injected intraperitoneally with 0.1mL/10g into the mice, and the mice developed sepsis symptoms 2 hours later.
  • the inactivated or active mitochondria extracted from newborn mice were diluted with DMEM into a 250mg/mL mitochondrial suspension, and injected intraperitoneally with 0.1mL/10g to mice with sepsis symptoms.
  • the control group was injected with IAM.DMEM, and the treatment group was injected with IAM.DMEM.
  • Mt.DMEM was injected and repeated intraperitoneal injections with the same dose on the second and third days, and the survival of the mice was continuously observed until the 7th day.
  • the results are shown in Figure 6. Intraperitoneal injection of mitochondria for 3 consecutive days can increase the survival rate of mice by about 30%.
  • mice 34 male Kunming mice were randomly divided into two groups (the treatment group and the control group each had 17), the weight range was 22-36g (the treatment group and the control group were paired, the weight difference was within 1g), and the concentration was 2mg/0.1mL
  • the Escherichia coli bacteria solution was injected into the left abdominal cavity of 0.1mL/10g into the mice, and the mice developed sepsis symptoms 2 hours later.
  • Inactivated or active mitochondria extracted from newborn mice were diluted with DMEM into a 250mg/mL suspension, and then injected into the right abdominal cavity of the mice at 0.1mL/10g.
  • the control group was injected with IAM.DMEM, and the treatment group was injected with IAM.DMEM.
  • mice Twenty-six male Kunming mice were randomly divided into two groups (13 in the treatment group and 13 in the control group), with a weight range of 20-26g (the treatment group and the control group were paired with a weight difference of less than 1g), and the concentration was 2mg/0.1mL
  • the Escherichia coli bacteria solution was injected intraperitoneally with 0.1 mL/10 g into the mice, and the mice developed sepsis symptoms 2 hours later.
  • the inactivated or active mitochondria extracted from newborn mice were diluted with DMEM into a 500mg/mL mitochondrial suspension, and then injected into the sepsis-symptomatic mice with 0.05mL/10g right thigh muscle, and the control group was injected with IAM.
  • DMEM the treatment group was injected with mt.DMEM, and the survival of the mice was continuously observed until day 7. The results are shown in Figure 8.
  • a single intramuscular injection of mitochondria can increase the survival rate of mice by about 30%.
  • 76 female Kunming mice were randomly divided into two groups (38 in the treatment group and 38 in the control group), with a weight range of 20-27g (the treatment group and the control group were paired with a weight difference of less than 1g), and the concentration was 10mg/0.1mL
  • the Escherichia coli bacteria solution was injected subcutaneously into the neck of 0.1mL/10g mice, and the mice developed sepsis symptoms 4 hours later.
  • the inactivated or active mitochondria extracted from newborn mice were diluted with DMEM into a 250mg/mL mitochondrial suspension, and then injected intraperitoneally with 0.1mL/10g to mice with sepsis symptoms.
  • the control group was injected with IAM.DMEM for treatment
  • the group was injected with mt.DMEM, and repeated intraperitoneal injections on the second and third days.
  • the survival of the mice was continuously observed until the 7th day.
  • the results are shown in Figure 9. Intraperitoneal injection of mitochondria for 3 consecutive days can increase the survival rate of mice by about 31.5%.
  • the inactivated or activated mitochondria extracted from 293T cells were diluted with DMEM into a 50mg/mL mitochondrial suspension, and then injected into the sepsis-symptomatic mice with 0.05mL/10g tail vein, and the control group was injected with IAM.DMEM for treatment The group was injected with mt.DMEM, and the survival of the mice was continuously observed until the 7th day.
  • the results are shown in Figure 10.
  • a single tail vein injection of mitochondria can increase the survival rate of mice by about 17.5%.
  • mice Forty male Kunming mice were randomly divided into two groups (20 in the treatment group and 20 in the control group), with a weight range of 22-30g (the treatment group and the control group were paired with a weight difference of less than 1g), and the concentration was 2mg/0.1mL
  • the Escherichia coli bacteria solution of 0.1mL/10g was injected subcutaneously into the neck of the mice, and the mice developed sepsis symptoms 2 hours later.
  • the mitochondria extracted from newborn mice, after inactivation or alcohol inactivation, are diluted with DMEM.10% FBS solution containing 20% sucrose at a concentration of 200 mg/mL, and placed in the program cooling box for one week at -80 degrees. the above.
  • mice with sepsis symptoms were injected with 0.05mL/10g tail vein.
  • the control group was injected with IAM.DMEM, and the treatment group was injected with mt.DMEM.
  • the survival of the mice was continuously observed until the 7th day. The results are shown in Figure 11.
  • a single tail vein injection of mitochondria can increase the survival rate of mice by about 19%.
  • Fifty male Kunming mice were randomly divided into two groups (25 in the treatment group and 25 in the control group), with a weight range of 22-30g (the treatment group and the control group were paired with a weight difference of less than 1g), and the concentration was 5% mice
  • the stool solution was injected intraperitoneally with 0.12mL/10g into the mice, and the mice developed sepsis symptoms 4 hours later.
  • the inactivated or active mitochondria extracted from newborn mice were diluted with DMEM into a 250mg/mL mitochondrial suspension, and then injected intraperitoneally with 0.1mL/10g to mice with sepsis symptoms.
  • the control group was injected with IAM.DMEM for treatment
  • the group was injected with mt.DMEM, and the survival of the mice was continuously observed until the 7th day.
  • the results are shown in Figure 12.
  • a single intraperitoneal injection of mitochondria can increase the survival rate of mice by about 22%.
  • 29 male Kunming mice were randomly divided into two groups (15 in the treatment group and 14 in the control group), with a weight range of 22-30g (the treatment group and the control group were paired with a weight difference of less than 1g), and the concentration was 10mg/0.1mL
  • the Escherichia coli bacteria solution was injected subcutaneously in the neck of 0.1mL/10g into the mice, and the mice developed sepsis symptoms 4 hours later.
  • the inactivated or active mitochondria extracted from newborn mice were diluted with DMEM into a 250mg/mL mitochondrial suspension, and then injected intraperitoneally with 0.1mL/10g to mice with sepsis symptoms.
  • the control group was injected with IAM.DMEM for treatment
  • the group was injected with mt.DMEM and repeated intraperitoneal injections on the 2nd to 5th days.
  • the survival of the mice was continuously observed until the 7th day.
  • the results are shown in Figure 13. Intraperitoneal injection of mitochondria for 5 consecutive days can increase the survival rate of mice by about 53%.

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Abstract

L'invention vise à surmonter les problèmes d'effets thérapeutiques médiocres et d'effets secondaires importants des procédés de traitement et des médicaments thérapeutiques existants pour la septicémie, et produit à cet effet une composition pharmaceutique pour la prévention ou le traitement de la septicémie, comprenant des mitochondries ayant une activité physiologique en tant que principe actif. L'invention concerne également un kit qui contient la composition pharmaceutique décrite. De plus, l'invention concerne l'utilisation de mitochondries dans la préparation d'un médicament, d'une composition pharmaceutique ou d'un kit pour la prévention ou le traitement de la septicémie, et un procédé de prévention ou de traitement de la septicémie. Différents modes d'administration (tels que l'injection intrapéritonéale IP, l'injection intraveineuse IV et l'injection sous-cutanée SC) ainsi que les dosages d'administration sont définis, les mitochondries actives et les mitochondries inactivées sont transplantées pour traiter une souris atteinte de septicémie. Il a été découvert que la transplantation de mitochondries actives autologues ou allogéniques peut améliorer le taux de survie des souris de 30 à 50 % pendant le processus de construction d'un modèle de septicémie de souris induite par Escherichia coli, ce qui a des effets thérapeutiques significatifs.
PCT/CN2020/123169 2019-10-24 2020-10-23 Composition pharmaceutique pour la prévention ou le traitement de la septicémie, kit, utilisation de celle-ci et procédé de traitement associé WO2021078246A1 (fr)

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Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115005162A (zh) * 2022-07-01 2022-09-06 天津市第一中心医院 贴合临床现状的老年脓毒症血小板减少小鼠模型构建方法

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090181367A1 (en) * 2002-07-05 2009-07-16 Helene Cote Diagnosis of sepsis using mitochondrial nucleic acid assays
CN104777109A (zh) * 2015-03-16 2015-07-15 首都儿科研究所附属儿童医院 一种脓毒症诊断方法及试剂
WO2017044940A1 (fr) * 2015-09-10 2017-03-16 Washington State University Nanovésicules à membrane cellulaire et leurs procédés d'utilisation
CN108796060A (zh) * 2017-07-26 2018-11-13 朱海燕 线粒体mt-co1在筛查脓毒症中的用途

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20090181367A1 (en) * 2002-07-05 2009-07-16 Helene Cote Diagnosis of sepsis using mitochondrial nucleic acid assays
CN104777109A (zh) * 2015-03-16 2015-07-15 首都儿科研究所附属儿童医院 一种脓毒症诊断方法及试剂
WO2017044940A1 (fr) * 2015-09-10 2017-03-16 Washington State University Nanovésicules à membrane cellulaire et leurs procédés d'utilisation
CN108796060A (zh) * 2017-07-26 2018-11-13 朱海燕 线粒体mt-co1在筛查脓毒症中的用途

Non-Patent Citations (2)

* Cited by examiner, † Cited by third party
Title
QIAN, J. ET AL.: "Research progress of transfering mitochondria application in nanotubes in treatment of acute lung injury sepsis", JOURNAL OF ACUTE DISEASE, vol. 6, no. 6, 1 November 2017 (2017-11-01), XP055756376, DOI: 10.4103/2221-6189.221286 *
XIAO, YAWEN ET AL.: "Research Progress on Mitochondrial Dysfunction in Sepsis", XINJIANG MEDICAL JOURNAL, vol. 49, no. 3, 31 March 2019 (2019-03-31), pages 225 - 229, XP055803552 *

Cited By (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN115005162A (zh) * 2022-07-01 2022-09-06 天津市第一中心医院 贴合临床现状的老年脓毒症血小板减少小鼠模型构建方法

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