WO2024091037A1 - Formulation d'administration de médicament à nanoparticules lipidiques biodégradables ciblant les poumons - Google Patents

Formulation d'administration de médicament à nanoparticules lipidiques biodégradables ciblant les poumons Download PDF

Info

Publication number
WO2024091037A1
WO2024091037A1 PCT/KR2023/016792 KR2023016792W WO2024091037A1 WO 2024091037 A1 WO2024091037 A1 WO 2024091037A1 KR 2023016792 W KR2023016792 W KR 2023016792W WO 2024091037 A1 WO2024091037 A1 WO 2024091037A1
Authority
WO
WIPO (PCT)
Prior art keywords
lipid
peg
lipid nanoparticles
drug
composition
Prior art date
Application number
PCT/KR2023/016792
Other languages
English (en)
Korean (ko)
Inventor
김경진
양주성
이혁진
김민정
정예희
이경석
이예지
박정은
정혜인
임성은
Original Assignee
에스티팜 주식회사
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by 에스티팜 주식회사 filed Critical 에스티팜 주식회사
Publication of WO2024091037A1 publication Critical patent/WO2024091037A1/fr

Links

Images

Classifications

    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A61K9/5107—Excipients; Inactive ingredients
    • A61K9/5123—Organic compounds, e.g. fats, sugars
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K31/00—Medicinal preparations containing organic active ingredients
    • A61K31/70—Carbohydrates; Sugars; Derivatives thereof
    • A61K31/7088—Compounds having three or more nucleosides or nucleotides
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K48/00—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
    • A61K48/0008—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition
    • A61K48/0025—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition wherein the non-active part clearly interacts with the delivered nucleic acid
    • A61K48/0033—Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy characterised by an aspect of the 'non-active' part of the composition delivered, e.g. wherein such 'non-active' part is not delivered simultaneously with the 'active' part of the composition wherein the non-active part clearly interacts with the delivered nucleic acid the non-active part being non-polymeric
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K9/00—Medicinal preparations characterised by special physical form
    • A61K9/48—Preparations in capsules, e.g. of gelatin, of chocolate
    • A61K9/50—Microcapsules having a gas, liquid or semi-solid filling; Solid microparticles or pellets surrounded by a distinct coating layer, e.g. coated microspheres, coated drug crystals
    • A61K9/51—Nanocapsules; Nanoparticles
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
    • A61P11/00—Drugs for disorders of the respiratory system

Definitions

  • the present invention relates to a composition for transpulmonary delivery of a drug, comprising lipid nanoparticles containing ionizable lipids and cationic lipids of a specific structure, and its use in the prevention or treatment of lung diseases.
  • Nucleic acids such as antisense RNA and mRNA are substances that can inhibit the expression of specific proteins in vivo, and are attracting attention as important tools in the treatment of cancer, genetic diseases, infectious diseases, and autoimmune diseases (Novina and Sharp, Nature, 430, 161-164, 2004). However, since nucleic acids are difficult to transfer directly into cells and are easily decomposed by enzymes in the blood, many studies are being conducted to overcome this problem.
  • Drug Delivery System is a technology designed to efficiently deliver the required amount of drugs by reducing the side effects of drugs and maximizing their efficacy and effectiveness.
  • conventional viral carriers have proven to be effective as drug carriers in gene therapy, but the use of viruses as a gene delivery system has been discouraged due to several drawbacks such as immunogenicity, limitations in the size of the injected DNA, and difficulties in mass production. It is being restricted.
  • the method of transporting nucleic acids into cells is currently mixing them with positively charged lipids or polymers (named lipid-DNA conjugates (lipoplex) and polymer-DNA conjugates (polyplex), respectively). ) is mainly used (Hirko et al., Curr, Med, Chem., 10, 1185-1193, 2003; Merdan et al., Adv. Drug. Deliv.Rev., 54, 715-758, 2002; Spagnou et al. al., Biochemistry, 43, 13348-13386, 2004).
  • lipid-DNA conjugates are widely used at the cellular level because they bind to nucleic acids and deliver nucleic acids well into cells, but when injected locally in vivo, they often cause inflammation in the body (Filonand and Phillips, Biochim. Biophys/Acta , 1329, 345-356, 1997), it has the disadvantage of accumulating in tissues such as the lung, liver, and spleen, which are the first passage organs during intravascular injection (Ren et al., Gene Therapy. 7, 764-768, 2000 ).
  • the present inventors have made diligent efforts to develop a delivery vehicle that can effectively deliver anionic drugs such as nucleic acids specifically to lung tissue.
  • a lipid nanoparticle formulation containing cationic lipids in addition to ionizable lipids with a specific structure has been developed into an excellent lung tissue.
  • the present invention was completed by confirming that it had specific drug delivery efficacy.
  • One object of the present invention is to provide a composition for transpulmonary delivery of a drug, comprising lipid nanoparticles containing ionizable lipids and cationic lipids having a specific structure.
  • Another object of the present invention is to provide a pharmaceutical composition for preventing or treating lung diseases, comprising the lipid nanoparticles and anionic drugs.
  • Lipid nanoparticles containing ionizable lipids and cationic lipids of the present invention are specifically delivered to lung tissue, have excellent biocompatibility, and can deliver anionic drugs with high efficiency, thereby enabling related technologies such as lipid nanoparticle-mediated gene therapy. It can be useful in this field.
  • Figure 1 shows the results of hEPO levels analyzed by intravenous injection into mice and blood collection to confirm in vivo delivery of hEPO mRNA-encapsulated 244-cis lipid nanoparticles.
  • Figure 2 shows the results of confirming the EPO protein level and MCP-1 level in the blood after intravenous injection into mice to confirm the in vivo delivery efficacy of hEPO mRNA-encapsulated 244-cis lipid nanoparticles.
  • Figure 3 shows the results of confirming the level of MCP-1 secretion depending on whether hEPO mRNA-encapsulated 244-cis lipid nanoparticles contain DOTAP.
  • Figure 4 shows the results of confirming the bioluminescence of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing DOPE.
  • Figure 5 shows the results of bioluminescence after intravenous injection into mice to confirm the in vivo delivery efficacy of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing 10 mol% of DOTAP.
  • Figure 6 shows the results of bioluminescence after intravenous injection into mice to confirm the in vivo delivery efficacy of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing 20 mol% DOTAP.
  • Figure 7 shows the results of bioluminescence after intravenous injection into mice to confirm the in vivo delivery efficacy of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing 40 mol% of DOTAP.
  • Figure 8 shows the results of bioluminescence after intravenous injection into mice to confirm the in vivo delivery efficacy of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing 60 mol% of DOTAP.
  • Figure 9 shows the results of comparing the distribution of luminescence in each organ after intravenous injection into mice to confirm the in vivo delivery efficacy according to the DOTAP-containing ratio of lipid nanoparticles encapsulated with fLuc mRNA.
  • Figure 10 shows the results of bioluminescence after intravenous injection into mice to confirm the in vivo delivery efficacy of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing 20 mol% phosphatidylcholine.
  • Figure 11 shows the results of bioluminescence after intravenous injection into mice to confirm the in vivo delivery efficacy of 244-cis lipid nanoparticles encapsulated with fLuc mRNA and containing 40 mol% phosphatidylcholine.
  • Figure 12 shows the results of comparing fluorescence expression in each lung cell after intravenous injection of Cre mRNA-encapsulated 244-cis lipid nanoparticles into LSL-tdTomato mice.
  • Figure 13 shows the results of comparing the fluorescence expression of each lung cell after intravenous injection of Cre mRNA-encapsulated 244-cis lipid nanoparticles into LSL-tdTomato mice with lung fibrosis.
  • Figure 14 shows the results of confirming fluorescence expression in Sca-1 + cells of fibroblasts after intravenous injection of Cre mRNA-encapsulated 244-cis lipid nanoparticles into LSL-tdTomato mice with lung fibrosis.
  • LNPs lipid nanoparticles
  • ionizable lipids ionizable lipids
  • cationic lipids represented by the following formula (1): , a composition for transpulmonary delivery of drugs.
  • ionizable lipid refers to an amine-containing lipid that can be easily protonated, and is also called a lipid analog (lipidoid). Since the charge state of the ionizable lipid can change depending on the surrounding pH, it plays a role in ensuring that the drug is encapsulated within the lipid nanoparticle with high efficiency through electrostatic interaction with the anionic drug, and the structure of the lipid nanoparticle contributes to forming
  • One feature of the lipid nanoparticles of the present invention is that they contain an ionizable lipid of the above formula (1).
  • the ionizable lipid of Formula 1 is a substance having a head group having a plurality of tertiary amines, an ester functional group, and a chain tail containing a carbon double bond, and is named “244-cis” in the present invention.
  • the present inventors found that ionizable lipids with 244-cis or similar structures can be used as a component of lipid nanoparticles, and that the lipid nanoparticles are excellent for delivering anionic drugs such as mRNA. It has been reported that it is effective.
  • the present inventors found that the target tissue of the ionizable lipid surprisingly varies depending on the type of helper lipid with which it is combined, and in particular, when lipid nanoparticles are manufactured using cationic lipid as a helper lipid, it is targeted specifically at the lungs. Delivery was confirmed. In addition, it was confirmed that the lipid nanoparticles of the present invention stably delivered mRNA with little immunogenicity and were expressed at a high level at the target site.
  • the present invention can be applied not only to the ionizable lipid represented by Formula 1, that is, 244-cis, but also to the ionizable lipid having a similar structure.
  • ionizable lipid of Formula 1 that is, 244-cis
  • Those skilled in the art can fully expect that beyond the scope of the ionizable lipid of Formula 1, for example, other ionizable lipids with similar structures described in WO2023/136689 can show the same effects as those confirmed in the present invention. Therefore, all ionizable lipids with structures similar to 244-cis can also be included within the scope of the present invention as equivalents.
  • the ionizable lipid of the present invention may be a compound represented by the following formula (2).
  • Y is -O- or -NH-
  • n 1 to 2
  • n is an integer from 3 to 7
  • x is an integer from 1 to 3
  • l is an integer from 3 to 4, and
  • p is an integer from 0 to 2.
  • the compound represented by Formula 2 may be selected from the group consisting of compounds shown in Table 1 below.
  • alkyl refers to a straight-chain or branched-chain acyclic saturated hydrocarbon, unless otherwise specified.
  • C 1-6 alkyl may mean alkyl containing 1 to 6 carbon atoms.
  • lipid nanoparticles of the present invention contain cationic lipids.
  • the cationic lipid corresponds to a helper lipid in lipid nanoparticles and, together with the ionizable lipid of the present invention, plays a role in delivering anionic drugs specifically to the lungs.
  • helper lipid serves to surround and protect the core formed by the interaction of ionizable lipids and drugs within lipid nanoparticles, and binds to the phospholipid bilayer of the target cell to protect the drug. When delivered intracellularly, it facilitates cell membrane passage and endosomal escape.
  • the “cationic lipid” may be DOTAP (1,2-dioleoyl-3-trimethylammonium propane) or phosphatidylcholine, but is not limited thereto.
  • phosphatidylcholine refers to a type of phospholipid and a compound containing choline as a head group. It is widely present in animals, plants, yeast, and molds, and is also called lecithin. It is a membrane phospholipid of mammals and is mainly contained in brain water, nerves, blood cells, and egg yolk. In plants, it is contained in soybeans, sunflower seeds, and wheat germ. In general, glycerol often has a saturated fatty acid bound to the 1st position and an unsaturated fatty acid bound to the 2nd position.
  • phosphatidylcholine may be a mixture of phosphatidylcholine of various structures, may be isolated from natural products (e.g., egg yolk, soybeans, etc.), or may be chemically synthesized.
  • the lipid nanoparticles of the present invention may contain DOTAP in an amount of 10 mol% to 70 mol% based on the entire lipid nanoparticle, for example, 15 mol% to 70 mol%, 20 mol%. It may be from 70 mol%, 15 mol% to 65 mol%, 20 mol% to 65 mol%, 20 mol% to 60 mol%, or 20 mol% to 50 mol%, but is not limited thereto.
  • the lipid nanoparticles of the present invention contain 10 mol% to 50 mol%, 15 mol% to 45 mol%, or 20 mol% to 60 mol% of phosphatidylcholine based on the total lipid nanoparticles.
  • 10 mol% to 50 mol% 15 mol% to 45 mol%
  • 20 mol% to 60 mol% of phosphatidylcholine based on the total lipid nanoparticles.
  • lipid nanoparticles when lipid nanoparticles are prepared using 244-cis ionizable lipids, they are generally mostly delivered to the liver ( Figure 4), but when they contain cationic lipids such as DOTAP or phosphatidylcholine, It was confirmed that mRNA was expressed through lung-specific delivery ( Figures 5 to 11). Therefore, lipid nanoparticles containing ionizable lipids and cationic lipids represented by Formula 1 (or Formula 2) of the present invention are specifically delivered to lung tissue, and thus deliver anionic drugs specifically to the lungs with high efficiency. It can be passed on.
  • Formula 1 or Formula 2
  • the lipid nanoparticles of the present invention may further include structural lipids or PEG-lipids.
  • the structural lipid maintains the particle shape within the lipid nanoparticle and is dispersed on the core and surface of the nanoparticle to improve the stability of the nanoparticle.
  • the structural lipid is, for example, cholesterol, cholesterol, spinasterol, fecosterol, sitosterol, ergosterol, ergostenol, campesterol, stigmasterol, brassicasterol, tomatidine, ursolic acid, alpha-tocopherol or these. It may be a mixture of, but is not limited thereto.
  • the PEG-lipid refers to a conjugated form of lipid and PEG, and refers to a lipid to which polyethylene glycol polymer, a hydrophilic polymer, is bound.
  • the PEG-lipid within lipid nanoparticles contributes to particle stability in serum and plays a role in preventing aggregation between nanoparticles.
  • PEG-lipids protect nucleic acids from degrading enzymes, enhance the stability of nucleic acids in the body, and can increase the half-life of drugs encapsulated in nanoparticles.
  • the PEG-lipid may be, for example, PEG-ceramide, PEG-DMG, PEG-c-DOMG, PEG-DLPE, PEG-DMPE, PEG-DPPC, PEG-DSPE or mixtures thereof, specifically C16-PEG2000 ceramide. However, it is not limited to this.
  • the molar ratio of ionizable lipid: DOTAP: structural lipid: PEG-lipid is 10 to 40. : 5 to 80 : 5 to 80 : 1 to 5. More specifically, the molar ratio may be 20 to 30: 10 to 70: 10 to 70: 1 to 5, and more specifically, 20 to 30: 15 to 70: 15 to 70: 1 to 3. Not limited.
  • the molar ratio of ionizable lipid: phosphatidylcholine: structural lipid: PEG-lipid is 10 to 10. It may be 40:5 to 80:5 to 80:1 to 5. More specifically, the molar ratio may be 20 to 30:10 to 50:30 to 60:1 to 5, and more specifically, 20 to 30:20 to 40:30 to 60:1 to 3. Not limited.
  • the lipid nanoparticles of the present invention exhibit a positive charge under acidic pH conditions, they can easily form a complex with the drug through electrostatic interaction with therapeutic agents such as nucleic acids and drugs that exhibit a negative charge, allowing the encapsulation of anionic drugs with high efficiency. It can be used as an intracellular or in vivo drug delivery composition. Therefore, the lipid nanoparticles of the present invention can be useful for the delivery of not only nucleic acids but also all types of drugs with anionic properties. That is, the lipid nanoparticles of the present invention can ultimately be manufactured in a form (encapsulated form) that additionally contains an anionic drug, and the encapsulated anionic drug can be specifically delivered to the lungs.
  • the term "encapsulation” refers to encapsulating the delivery material to surround it and efficiently incorporate it into the body
  • the drug encapsulation efficiency refers to the lipid content relative to the total drug content used in manufacturing. It refers to the content of drug encapsulated in nanoparticles.
  • the anionic drug may be a nucleic acid, low molecular weight compound, peptide, protein, protein-nucleic acid structure, or anionic biopolymer-drug conjugate, but may be stably and efficiently delivered by forming lipid nanoparticles together with the ionizable lipid of the present invention. It is not limited to this as long as possible.
  • the nucleic acids include small interfering ribonucleic acid (siRNA), ribosomal ribonucleic acid (rRNA), deoxyribonucleic acid (DNA), complementary deoxyribonucleic acid (cDNA), aptamer, messenger ribonucleic acid (mRNA), and transport ribonucleic acid. It may be (tRNA), sgRNA, antisense oligonucleotide, shRNA, miRNA, ribozyme, PNA, and DNAzyme, or a mixture thereof, but is not limited thereto.
  • siRNA small interfering ribonucleic acid
  • rRNA ribosomal ribonucleic acid
  • DNA deoxyribonucleic acid
  • cDNA complementary deoxyribonucleic acid
  • aptamer messenger ribonucleic acid
  • mRNA messenger ribonucleic acid
  • mRNA messenger ribonucleic acid
  • transport ribonucleic acid It
  • the weight ratio of total lipid/nucleic acid in the lipid nanoparticles may be 1 to 30, specifically 5 to 27, and more specifically 10 to 23, but is not limited thereto.
  • Another aspect of the present invention is a pharmaceutical composition for preventing or treating lung disease, comprising lipid nanoparticles containing the ionizable lipid and cationic lipid of Formula 1 above, and an anionic drug as an active ingredient.
  • Another aspect of the present invention is a method of treating lung disease, comprising administering the composition to an individual in need thereof.
  • Another aspect of the present invention is a pharmaceutical composition for use in the prevention or treatment of lung disease, or use of the composition for the prevention or treatment of lung disease.
  • Lipid nanoparticles and anionic drugs are as described above.
  • the lipid nanoparticles of the present invention form a stable complex with anionic drugs such as nucleic acids and exhibit low cytotoxicity and effective cell absorption, so they are effective in delivering anionic drugs.
  • anionic drugs such as nucleic acids
  • the lipid nanoparticles can be specifically delivered to the lungs when administered, they can be useful in preventing or treating lung diseases.
  • the lipid nanoparticles of the present invention can specifically deliver drugs by targeting the lungs, and for the purpose of the present invention, prevent or treat various lung diseases depending on the type of anionic drug, type of nucleic acid, and nucleic acid sequence used. Effects can be expected. That is, in the present invention, the type of lung disease is not limited. Therefore, the lung diseases include, for example, emphysema, asthma, pneumonia, tuberculosis, pulmonary hypertension, lung cancer, neonatal bronchopulmonary dysplasia, chronic obstructive pulmonary disease, acute bronchitis, chronic bronchitis, bronchiolitis, bronchiectasis, hypersensitivity, lung parenchyma.
  • the lung diseases include, for example, emphysema, asthma, pneumonia, tuberculosis, pulmonary hypertension, lung cancer, neonatal bronchopulmonary dysplasia, chronic obstructive pulmonary disease, acute bronchitis, chronic bronchit
  • the term “treatment” refers to intervention to alter the natural processes of an individual or cell with a disease, which may be performed during the progression of the pathology or to prevent it.
  • the desired therapeutic effects include preventing the occurrence or recurrence of the disease, alleviating symptoms, reducing all direct or indirect pathological consequences of the disease, preventing metastasis, reducing the rate of disease progression, and alleviating the disease state. Or it includes temporary relief, remission, or improvement of prognosis.
  • the present invention includes all actions to improve the course of lung-related diseases by administering a composition containing lipid nanoparticles containing ionizable lipids and cationic lipids and anionic drugs as active ingredients.
  • prevention refers to all actions that suppress or delay the onset of a disease by administering the lipid nanoparticles.
  • the lipid nanoparticles of the present invention are used for treatment or prevention purposes, they are administered to an individual in a therapeutically effective amount.
  • therapeutically effective amount refers to an effective amount of anionic drug-containing lipid nanoparticles.
  • therapeutically effective amount means an amount sufficient to treat the disease with a reasonable benefit/risk ratio applicable to medical treatment, and the effective dose level is determined by the type and severity of the individual, age, gender, type of disease, It can be determined based on factors including the activity of the drug, sensitivity to the drug, time of administration, route of administration and excretion rate, duration of treatment, drugs used simultaneously, and other factors well known in the medical field.
  • the pharmaceutical composition of the present invention may be administered as an individual therapeutic agent or in combination with other therapeutic agents, and may be administered sequentially or simultaneously with commercially available therapeutic agents. And it can be administered single or multiple times.
  • the administered dose of the pharmaceutical composition of the present invention may be determined by an expert depending on various factors such as the patient's condition, age, gender, and complications. Since the active ingredient of the composition of the present invention has excellent safety, it can be used beyond the determined administration dose.
  • composition containing the lipid nanoparticles may be administered orally, intramuscularly, intravenously, arterially, subcutaneously, intraperitoneally, pulmonaryly, and intranasally, and may specifically be administered intravenously, but is not limited thereto.
  • composition of the present invention may further include one or more pharmaceutically acceptable carriers for administration.
  • Pharmaceutically acceptable carriers can be saline solution, sterile water, Ringer's solution, buffered saline solution, dextrose solution, maltodextrin solution, glycerol, ethanol, and a mixture of one or more of these ingredients, and if necessary, antioxidants, buffer solutions, Other common additives such as bacteriostatic agents can be added.
  • Example 1 Preparation of lipid nanoparticles for transpulmonary delivery
  • 244-cis (see WO2023/136689) was prepared as an ionizable lipid and synthesized according to Scheme 1 below.
  • lipid nanoparticles containing 244-cis were named '244-cis lipid nanoparticles'.
  • SM-102 or MC3 was used as a control.
  • Ionizable lipids cationic lipids
  • DOTAP 1,2-dioleoyl-3-trimethylammonium propane
  • DOPE 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine
  • EPC egg phosphatidylcholine (Avanti, 890704P)
  • cholesterol Cholesterol powder, BioReagent, for cell culture, ⁇ 99%, Sigma, Korea
  • C16-PEG2000 ceramide (Avanti, USA) were dissolved in ethanol at a molar ratio of 26.5:10 to 60:12 to 62:1.5 (Table 2). I ordered it.
  • ingredient Content mol% ionizable lipids 26.5 26.5 26.5 26.5 cationic lipids 10 20 40 60 cholesterol 62 52 32 12 PEG-lipid 1.5 1.5 1.5 1.5 1.5
  • Ionizable lipids, cholesterol, cationic lipids, and PEG-ceramide conjugates were dissolved in ethanol and nucleic acids (mRNA) were dissolved in aqueous phase (sodium acetate or sodium citrate) at a volume ratio of 1:3 at a flow rate of 12 ml/min.
  • mRNA nucleic acids
  • Lipid nanoparticles were prepared by mixing using a microfluidic mixing device (Benchtop Nanoassemblr; PNI, Canada). The weight ratio of mRNA:ionizable lipid was 1:20.
  • hEPO mRNA (SEQ ID NO: 1) was encapsulated, and lipid nanoparticles containing 244-cis, DOPE (20 mol%), cholesterol, and PEG-lipid were administered intravenously to 7-week-old Balb/c mice at a dose of 0.1 mg/kg based on mRNA. Injected. Blood was collected after 3, 6, 9, 24, and 48 hours, and hEPO protein expression was confirmed using ELISA (R&D systems, USA) ( Figure 1).
  • lipid nanoparticles containing 244-cis effectively delivered and expressed nucleic acids in vivo, and hEPO was detected at a significant level in the blood.
  • Lipid nanoparticles containing 244-cis showed a significantly higher level than the control MC3 LNP, and showed a level equivalent to that of SM-102 LNP.
  • the lipid nanoparticles of Experimental Example 1-1 were injected intravenously into 7-week-old Balb/c mice at a dose of 0.5 mg/kg based on mRNA, and blood was collected 3 hours later. MCP-1 protein expression was confirmed using ELISA (Invitrogen, USA) ( Figure 2).
  • Lipid nanoparticles in which DOPE was replaced with DOTAP in the 244-cis lipid nanoparticles of Experimental Example 1-1 above were injected intravenously into 7-week-old Balb/c mice at a dose of 0.5 mg/kg based on mRNA, and blood was collected 6 hours later. did. MCP-1 protein expression was confirmed using ELISA (Invitrogen, USA) ( Figure 3).
  • fLuciferase mRNA (SEQ ID NO: 2) was encapsulated and lipid nanoparticles containing 244-cis, DOTAP, cholesterol, and PEG-lipid were prepared according to the contents in Table 2, and administered to 7-week-old C57BL/6 at a dose of 0.2 mg/kg based on mRNA. It was injected intravenously into mice. Six hours later, 0.25 mg/kg of luciferin was administered intraperitoneally, and bioluminescence was confirmed through ex vivo organ images using IVIS (PerkinElmer, USA) equipment (FIGS. 4 to 8). As a control, DOPE-containing lipid nanoparticles were used.
  • Lipid nanoparticles containing 244-cis and phosphatidylcholine (EPC) were prepared according to the contents in Table 2, and 0.1 mg/kg of mRNA was injected intravenously into 7-week-old C57BL/6 mice. Six hours later, 0.25 mg/kg of luciferin was administered intraperitoneally, and bioluminescence was confirmed through ex vivo organ images using IVIS (PerkinElmer, USA) equipment (FIGS. 10 and 11).
  • 244-cis lipid nanoparticles containing phosphatidylcholine showed high lung-specific luminescence intensity.
  • LSL-tdTomato mice (The Jackson Laboratory #: 007914), which express Tomato protein using Cre protein, were used.
  • Cre mRNA SEQ ID NO: 3
  • lipid nanoparticles containing 244-cis, DOTAP (20 mol%), cholesterol, and PEG-lipid were administered to 7-week-old LSL-tdTomato mice at a dose of 0.3 mg/kg based on mRNA. It was administered intravenously twice at daily intervals.
  • lung tissue cells were isolated and tdTomato fluorescence expression in each lung cell (endothelial cells, fibroblasts, epithelial cells, immune cells) was confirmed using flow cytometry (LSRFortessa, BD) ( Figure 12).
  • APC anti-mouse CD31 antibody BioLegend; 102409) for staining endothelial cells
  • FITC anti-mouse CD45.2 antibody BioLegend; 109805
  • PE/Cyanine7 anti-mouse CD326 antibody BioLegend; 109805
  • lung tissue cells were isolated and tdTomato fluorescence expression was confirmed in each lung cell (endothelial cells, fibroblasts, epithelial cells, immune cells) using flow cytometry (LSRFortessa, BD) ( Figures 13 and 14 ).
  • the lipid nanoparticles of the present invention showed overwhelmingly higher expression than the control MC3 LNP in all types of lung cells.
  • endothelial cells showed over 80% tdTomato expression
  • epithelial cells, immune cells, and fibroblasts also showed tdTomato expression at a level of about 30% (Figure 13).
  • tdTomato expression was shown by more than 40%, especially in Sca-1 + fibroblasts ( Figure 14).
  • the 244-cis LNP of the present invention can deliver and express mRNA specifically to the lung compared to the MC3 LNP, and can specifically deliver mRNA to fibroblasts in a lung fibrosis model.

Landscapes

  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Chemical & Material Sciences (AREA)
  • Public Health (AREA)
  • General Health & Medical Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Veterinary Medicine (AREA)
  • Pharmacology & Pharmacy (AREA)
  • Animal Behavior & Ethology (AREA)
  • Engineering & Computer Science (AREA)
  • Epidemiology (AREA)
  • Bioinformatics & Cheminformatics (AREA)
  • Molecular Biology (AREA)
  • Biomedical Technology (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
  • Optics & Photonics (AREA)
  • Biotechnology (AREA)
  • Genetics & Genomics (AREA)
  • General Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Physics & Mathematics (AREA)
  • Pulmonology (AREA)
  • Nanotechnology (AREA)
  • Biochemistry (AREA)
  • Medicinal Preparation (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)

Abstract

La présente invention concerne une composition pour l'administration transpulmonaire de médicaments et son utilisation. Des nanoparticules lipidiques contenant des lipides ionisables et des lipides cationiques selon la présente invention sont spécifiquement administrées au tissu pulmonaire et à des cellules spécifiques de celui-ci, ont une excellente biocompatibilité, et peuvent administrer des médicaments anioniques avec une efficacité élevée, et peuvent ainsi être utiles dans des domaines techniques pertinents tels que la thérapie génique médiée par des nanoparticules lipidiques.
PCT/KR2023/016792 2022-10-27 2023-10-26 Formulation d'administration de médicament à nanoparticules lipidiques biodégradables ciblant les poumons WO2024091037A1 (fr)

Applications Claiming Priority (2)

Application Number Priority Date Filing Date Title
KR10-2022-0140249 2022-10-27
KR20220140249 2022-10-27

Publications (1)

Publication Number Publication Date
WO2024091037A1 true WO2024091037A1 (fr) 2024-05-02

Family

ID=90831413

Family Applications (1)

Application Number Title Priority Date Filing Date
PCT/KR2023/016792 WO2024091037A1 (fr) 2022-10-27 2023-10-26 Formulation d'administration de médicament à nanoparticules lipidiques biodégradables ciblant les poumons

Country Status (2)

Country Link
KR (1) KR20240083166A (fr)
WO (1) WO2024091037A1 (fr)

Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20140308304A1 (en) * 2011-12-07 2014-10-16 Alnylam Pharmaceuticals, Inc. Lipids for the delivery of active agents
US20150166462A1 (en) * 2012-03-29 2015-06-18 Shire Human Genetic Therapies. Inc. Ionizable cationic lipids
KR20190093816A (ko) * 2016-10-26 2019-08-26 큐어백 아게 지질 나노입자 mRNA 백신
US20210107861A1 (en) * 2014-06-25 2021-04-15 Acuitas Therapeutics, Inc. Lipids and lipid nanoparticle formulations for delivery of nucleic acids
WO2023136689A1 (fr) * 2022-01-17 2023-07-20 에스티팜 주식회사 Lipide ionisable contenant une liaison ester biodégradable et nanoparticules lipidiques le comprenant

Patent Citations (5)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
US20140308304A1 (en) * 2011-12-07 2014-10-16 Alnylam Pharmaceuticals, Inc. Lipids for the delivery of active agents
US20150166462A1 (en) * 2012-03-29 2015-06-18 Shire Human Genetic Therapies. Inc. Ionizable cationic lipids
US20210107861A1 (en) * 2014-06-25 2021-04-15 Acuitas Therapeutics, Inc. Lipids and lipid nanoparticle formulations for delivery of nucleic acids
KR20190093816A (ko) * 2016-10-26 2019-08-26 큐어백 아게 지질 나노입자 mRNA 백신
WO2023136689A1 (fr) * 2022-01-17 2023-07-20 에스티팜 주식회사 Lipide ionisable contenant une liaison ester biodégradable et nanoparticules lipidiques le comprenant

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
KEJIN ZHOU: "Modular degradable dendrimers enable small RNAs to extend survival in an aggressive liver cancer model", PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES, NATIONAL ACADEMY OF SCIENCES, vol. 113, no. 3, 19 January 2016 (2016-01-19), pages 520 - 525, XP093163988, ISSN: 0027-8424, DOI: 10.1073/pnas.1520756113 *

Also Published As

Publication number Publication date
KR20240083166A (ko) 2024-06-11

Similar Documents

Publication Publication Date Title
Antimisiaris et al. Overcoming barriers by local drug delivery with liposomes
KR102198736B1 (ko) 생체 내 약물 전달을 위한 지질 나노입자 및 이의 용도
EP2567693B1 (fr) ARN interférents encapsulés dans des lipides
EP2430168B1 (fr) Compositions comprenant des amphiphiles et des colipides cationiques pour administrer des molécules thérapeutiques
KR101476167B1 (ko) 약물 송달 제어용 경폐 투여 리포좀
JP2012509258A (ja) 核酸送達系のための分岐カチオン性脂質
WO2013070010A1 (fr) Nouvelle composition pour l'administration d'un gène
CN104582691A (zh) 脂膜白蛋白纳米颗粒组合物以及制备和使用其的方法
KR20010101752A (ko) 적응성이 높은 운반체에 의한 경비운반／면역화
JP2012509366A (ja) 核酸送達系のための放出可能ポリマー脂質
JP5382682B2 (ja) 薬物送達複合体
WO2019212288A1 (fr) Composition de nanoparticules polymères pour l'administration d'arn messager, et son procédé de préparation
KR20130062953A (ko) 안티센스 올리고뉴클레오티드를 포함하는 약학 조성물 및 이의 사용 방법
JP3804452B2 (ja) 肝炎治療剤
KR20220092363A (ko) 지질 나노입자를 포함하는 암 예방 또는 치료용 조성물
WO2023136689A1 (fr) Lipide ionisable contenant une liaison ester biodégradable et nanoparticules lipidiques le comprenant
WO2024091037A1 (fr) Formulation d'administration de médicament à nanoparticules lipidiques biodégradables ciblant les poumons
KR20240082389A (ko) 생분해성 이황화 결합을 포함하는 이온화 가능한 지질 및 이를 포함하는 지질나노입자
EP1029544A1 (fr) Activateur de nuclease de cellules intracancereuses
WO2024096516A1 (fr) Formulation de nanoparticules lipidiques pour administration transpulmonaire de médicaments à base d'acides nucléiques et son utilisation
Betker et al. Nonadditive effects of repetitive administration of lipoplexes in immunocompetent mice
WO2024177282A1 (fr) Lipide ionisable comprenant du sulfure, et nanoparticules lipidiques le comprenant
WO2024177426A1 (fr) Nanoparticules lipidiques pour l'administration in vivo de médicaments
WO2023190176A1 (fr) Nanoparticules lipidiques pour l'administration d'acide nucléique à un tissu splénique, et procédé d'administration d'acide nucléique à un tissu splénique l'utilisant
EP4410275A1 (fr) Nanoparticule lipidique utilisée pour l'administration d'un acide nucléique au tissu cérébral

Legal Events

Date Code Title Description
121 Ep: the epo has been informed by wipo that ep was designated in this application

Ref document number: 23883142

Country of ref document: EP

Kind code of ref document: A1