CN118290282A - Ionizable lipid compound, preparation method and application thereof - Google Patents
Ionizable lipid compound, preparation method and application thereof Download PDFInfo
- Publication number
- CN118290282A CN118290282A CN202410324191.0A CN202410324191A CN118290282A CN 118290282 A CN118290282 A CN 118290282A CN 202410324191 A CN202410324191 A CN 202410324191A CN 118290282 A CN118290282 A CN 118290282A
- Authority
- CN
- China
- Prior art keywords
- compound
- pharmaceutically acceptable
- lipid
- tautomer
- stereoisomer
- Prior art date
- Legal status (The legal status 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 status listed.)
- Pending
Links
- -1 lipid compound Chemical class 0.000 title claims description 23
- 238000002360 preparation method Methods 0.000 title claims description 14
- 150000002632 lipids Chemical class 0.000 claims abstract description 78
- 150000001875 compounds Chemical class 0.000 claims abstract description 67
- 239000002105 nanoparticle Substances 0.000 claims abstract description 47
- 150000003839 salts Chemical class 0.000 claims abstract description 26
- 230000000155 isotopic effect Effects 0.000 claims abstract description 17
- 239000008194 pharmaceutical composition Substances 0.000 claims abstract description 13
- 125000000217 alkyl group Chemical group 0.000 claims description 24
- 208000037265 diseases, disorders, signs and symptoms Diseases 0.000 claims description 24
- 125000000304 alkynyl group Chemical group 0.000 claims description 15
- 238000000034 method Methods 0.000 claims description 15
- 201000010099 disease Diseases 0.000 claims description 14
- 229910052739 hydrogen Inorganic materials 0.000 claims description 14
- 239000001257 hydrogen Substances 0.000 claims description 14
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 11
- 239000003814 drug Substances 0.000 claims description 10
- 238000011282 treatment Methods 0.000 claims description 8
- 125000002947 alkylene group Chemical group 0.000 claims description 7
- 230000007935 neutral effect Effects 0.000 claims description 7
- 125000000739 C2-C30 alkenyl group Chemical group 0.000 claims description 6
- 230000000069 prophylactic effect Effects 0.000 claims description 6
- 230000001225 therapeutic effect Effects 0.000 claims description 6
- 239000002671 adjuvant Substances 0.000 claims description 5
- 125000004450 alkenylene group Chemical group 0.000 claims description 5
- 125000004419 alkynylene group Chemical group 0.000 claims description 5
- 239000004480 active ingredient Substances 0.000 claims description 4
- 239000000032 diagnostic agent Substances 0.000 claims description 3
- 229940039227 diagnostic agent Drugs 0.000 claims description 3
- 150000002431 hydrogen Chemical class 0.000 claims description 3
- 238000004519 manufacturing process Methods 0.000 claims description 3
- 230000002265 prevention Effects 0.000 claims description 3
- 230000008569 process Effects 0.000 claims description 3
- 125000002091 cationic group Chemical group 0.000 claims description 2
- 238000003745 diagnosis Methods 0.000 claims description 2
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 claims description 2
- 229910052736 halogen Inorganic materials 0.000 claims description 2
- 150000002367 halogens Chemical class 0.000 claims description 2
- 125000002496 methyl group Chemical group [H]C([H])([H])* 0.000 claims description 2
- 125000000843 phenylene group Chemical group C1(=C(C=CC=C1)*)* 0.000 claims description 2
- 125000001436 propyl group Chemical group [H]C([*])([H])C([H])([H])C([H])([H])[H] 0.000 claims description 2
- 239000000203 mixture Substances 0.000 abstract description 14
- 150000007523 nucleic acids Chemical class 0.000 abstract description 10
- 108020004707 nucleic acids Proteins 0.000 abstract description 9
- 102000039446 nucleic acids Human genes 0.000 abstract description 9
- 239000000126 substance Substances 0.000 abstract description 5
- 238000005538 encapsulation Methods 0.000 abstract description 4
- 230000000975 bioactive effect Effects 0.000 abstract description 2
- 238000006243 chemical reaction Methods 0.000 description 22
- 108020004999 messenger RNA Proteins 0.000 description 19
- 235000002639 sodium chloride Nutrition 0.000 description 17
- YMWUJEATGCHHMB-UHFFFAOYSA-N dichloromethane Natural products ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 description 14
- 125000001424 substituent group Chemical group 0.000 description 14
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Natural products CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 description 13
- 125000004432 carbon atom Chemical group C* 0.000 description 13
- 210000004027 cell Anatomy 0.000 description 12
- HVYWMOMLDIMFJA-DPAQBDIFSA-N cholesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CCCC(C)C)[C@@]1(C)CC2 HVYWMOMLDIMFJA-DPAQBDIFSA-N 0.000 description 12
- 125000003342 alkenyl group Chemical group 0.000 description 11
- 229920002477 rna polymer Polymers 0.000 description 11
- 229920001223 polyethylene glycol Polymers 0.000 description 10
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 9
- 239000002202 Polyethylene glycol Substances 0.000 description 9
- OILXMJHPFNGGTO-UHFFFAOYSA-N (22E)-(24xi)-24-methylcholesta-5,22-dien-3beta-ol Natural products C1C=C2CC(O)CCC2(C)C2C1C1CCC(C(C)C=CC(C)C(C)C)C1(C)CC2 OILXMJHPFNGGTO-UHFFFAOYSA-N 0.000 description 8
- VHYFNPMBLIVWCW-UHFFFAOYSA-N 4-Dimethylaminopyridine Chemical compound CN(C)C1=CC=NC=C1 VHYFNPMBLIVWCW-UHFFFAOYSA-N 0.000 description 8
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical group N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 description 8
- 208000035475 disorder Diseases 0.000 description 8
- 239000012074 organic phase Substances 0.000 description 8
- 239000000243 solution Substances 0.000 description 8
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 description 8
- 241000282414 Homo sapiens Species 0.000 description 7
- 241000699670 Mus sp. Species 0.000 description 7
- 238000002156 mixing Methods 0.000 description 7
- MZSAMHOCTRNOIZ-UHFFFAOYSA-N 3-[4-(aminomethyl)-6-(trifluoromethyl)pyridin-2-yl]oxy-N-phenylaniline Chemical compound NCC1=CC(=NC(=C1)C(F)(F)F)OC=1C=C(NC2=CC=CC=C2)C=CC=1 MZSAMHOCTRNOIZ-UHFFFAOYSA-N 0.000 description 6
- OQMZNAMGEHIHNN-UHFFFAOYSA-N 7-Dehydrostigmasterol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)C=CC(CC)C(C)C)CCC33)C)C3=CC=C21 OQMZNAMGEHIHNN-UHFFFAOYSA-N 0.000 description 6
- 108060001084 Luciferase Proteins 0.000 description 6
- 239000005089 Luciferase Substances 0.000 description 6
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 6
- ZEEBGORNQSEQBE-UHFFFAOYSA-N [2-(3-phenylphenoxy)-6-(trifluoromethyl)pyridin-4-yl]methanamine Chemical compound C1(=CC(=CC=C1)OC1=NC(=CC(=C1)CN)C(F)(F)F)C1=CC=CC=C1 ZEEBGORNQSEQBE-UHFFFAOYSA-N 0.000 description 6
- LGJMUZUPVCAVPU-UHFFFAOYSA-N beta-Sitostanol Natural products C1CC2CC(O)CCC2(C)C2C1C1CCC(C(C)CCC(CC)C(C)C)C1(C)CC2 LGJMUZUPVCAVPU-UHFFFAOYSA-N 0.000 description 6
- 239000012267 brine Substances 0.000 description 6
- 239000011203 carbon fibre reinforced carbon Substances 0.000 description 6
- 235000012000 cholesterol Nutrition 0.000 description 6
- 150000002430 hydrocarbons Chemical group 0.000 description 6
- 239000011734 sodium Substances 0.000 description 6
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical compound O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 6
- 238000012360 testing method Methods 0.000 description 6
- HJUGFYREWKUQJT-UHFFFAOYSA-N tetrabromomethane Chemical compound BrC(Br)(Br)Br HJUGFYREWKUQJT-UHFFFAOYSA-N 0.000 description 6
- CFOAUYCPAUGDFF-UHFFFAOYSA-N tosmic Chemical compound CC1=CC=C(S(=O)(=O)C[N+]#[C-])C=C1 CFOAUYCPAUGDFF-UHFFFAOYSA-N 0.000 description 6
- RIOQSEWOXXDEQQ-UHFFFAOYSA-N triphenylphosphine Chemical compound C1=CC=CC=C1P(C=1C=CC=CC=1)C1=CC=CC=C1 RIOQSEWOXXDEQQ-UHFFFAOYSA-N 0.000 description 6
- LMDZBCPBFSXMTL-UHFFFAOYSA-N 1-Ethyl-3-(3-dimethylaminopropyl)carbodiimide Substances CCN=C=NCCCN(C)C LMDZBCPBFSXMTL-UHFFFAOYSA-N 0.000 description 5
- 241001465754 Metazoa Species 0.000 description 5
- OAICVXFJPJFONN-UHFFFAOYSA-N Phosphorus Chemical group [P] OAICVXFJPJFONN-UHFFFAOYSA-N 0.000 description 5
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 description 5
- SAHIZENKTPRYSN-UHFFFAOYSA-N [2-[3-(phenoxymethyl)phenoxy]-6-(trifluoromethyl)pyridin-4-yl]methanamine Chemical compound O(C1=CC=CC=C1)CC=1C=C(OC2=NC(=CC(=C2)CN)C(F)(F)F)C=CC=1 SAHIZENKTPRYSN-UHFFFAOYSA-N 0.000 description 5
- 125000004429 atom Chemical group 0.000 description 5
- 230000008901 benefit Effects 0.000 description 5
- 239000003153 chemical reaction reagent Substances 0.000 description 5
- 229940125904 compound 1 Drugs 0.000 description 5
- 239000012043 crude product Substances 0.000 description 5
- 238000001727 in vivo Methods 0.000 description 5
- 108091070501 miRNA Proteins 0.000 description 5
- 239000000741 silica gel Substances 0.000 description 5
- 229910002027 silica gel Inorganic materials 0.000 description 5
- 239000004055 small Interfering RNA Substances 0.000 description 5
- 208000024891 symptom Diseases 0.000 description 5
- FPQQSJJWHUJYPU-UHFFFAOYSA-N 3-(dimethylamino)propyliminomethylidene-ethylazanium;chloride Chemical compound Cl.CCN=C=NCCCN(C)C FPQQSJJWHUJYPU-UHFFFAOYSA-N 0.000 description 4
- BBYDXOIZLAWGSL-UHFFFAOYSA-N 4-fluorobenzoic acid Chemical compound OC(=O)C1=CC=C(F)C=C1 BBYDXOIZLAWGSL-UHFFFAOYSA-N 0.000 description 4
- 108020004414 DNA Proteins 0.000 description 4
- 102000053602 DNA Human genes 0.000 description 4
- DHMQDGOQFOQNFH-UHFFFAOYSA-N Glycine Chemical compound NCC(O)=O DHMQDGOQFOQNFH-UHFFFAOYSA-N 0.000 description 4
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 description 4
- 108020004459 Small interfering RNA Proteins 0.000 description 4
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical compound [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- NINIDFKCEFEMDL-UHFFFAOYSA-N Sulfur Chemical group [S] NINIDFKCEFEMDL-UHFFFAOYSA-N 0.000 description 4
- WYURNTSHIVDZCO-UHFFFAOYSA-N Tetrahydrofuran Chemical compound C1CCOC1 WYURNTSHIVDZCO-UHFFFAOYSA-N 0.000 description 4
- HZYXFRGVBOPPNZ-UHFFFAOYSA-N UNPD88870 Natural products C1C=C2CC(O)CCC2(C)C2C1C1CCC(C(C)=CCC(CC)C(C)C)C1(C)CC2 HZYXFRGVBOPPNZ-UHFFFAOYSA-N 0.000 description 4
- REAYFGLASQTHKB-UHFFFAOYSA-N [2-[3-(1H-pyrazol-4-yl)phenoxy]-6-(trifluoromethyl)pyridin-4-yl]methanamine Chemical compound N1N=CC(=C1)C=1C=C(OC2=NC(=CC(=C2)CN)C(F)(F)F)C=CC=1 REAYFGLASQTHKB-UHFFFAOYSA-N 0.000 description 4
- QVGXLLKOCUKJST-UHFFFAOYSA-N atomic oxygen Chemical group [O] QVGXLLKOCUKJST-UHFFFAOYSA-N 0.000 description 4
- 239000003795 chemical substances by application Substances 0.000 description 4
- 235000019439 ethyl acetate Nutrition 0.000 description 4
- 238000000338 in vitro Methods 0.000 description 4
- 239000000463 material Substances 0.000 description 4
- DILRJUIACXKSQE-UHFFFAOYSA-N n',n'-dimethylethane-1,2-diamine Chemical compound CN(C)CCN DILRJUIACXKSQE-UHFFFAOYSA-N 0.000 description 4
- 229910052757 nitrogen Inorganic materials 0.000 description 4
- 229910052760 oxygen Inorganic materials 0.000 description 4
- 239000001301 oxygen Chemical group 0.000 description 4
- 239000002245 particle Substances 0.000 description 4
- 229910052698 phosphorus Inorganic materials 0.000 description 4
- 239000011574 phosphorus Chemical group 0.000 description 4
- 229920002523 polyethylene Glycol 1000 Polymers 0.000 description 4
- 229920000642 polymer Polymers 0.000 description 4
- NLQLSVXGSXCXFE-UHFFFAOYSA-N sitosterol Natural products CC=C(/CCC(C)C1CC2C3=CCC4C(C)C(O)CCC4(C)C3CCC2(C)C1)C(C)C NLQLSVXGSXCXFE-UHFFFAOYSA-N 0.000 description 4
- 229910000033 sodium borohydride Inorganic materials 0.000 description 4
- 239000012279 sodium borohydride Substances 0.000 description 4
- 229910000104 sodium hydride Inorganic materials 0.000 description 4
- HCXVJBMSMIARIN-PHZDYDNGSA-N stigmasterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)/C=C/[C@@H](CC)C(C)C)[C@@]1(C)CC2 HCXVJBMSMIARIN-PHZDYDNGSA-N 0.000 description 4
- 235000016831 stigmasterol Nutrition 0.000 description 4
- 229940032091 stigmasterol Drugs 0.000 description 4
- BFDNMXAIBMJLBB-UHFFFAOYSA-N stigmasterol Natural products CCC(C=CC(C)C1CCCC2C3CC=C4CC(O)CCC4(C)C3CCC12C)C(C)C BFDNMXAIBMJLBB-UHFFFAOYSA-N 0.000 description 4
- 229910052717 sulfur Inorganic materials 0.000 description 4
- 239000011593 sulfur Chemical group 0.000 description 4
- 229940124597 therapeutic agent Drugs 0.000 description 4
- GVJHHUAWPYXKBD-IEOSBIPESA-N α-tocopherol Chemical compound OC1=C(C)C(C)=C2O[C@@](CCC[C@H](C)CCC[C@H](C)CCCC(C)C)(C)CCC2=C1C GVJHHUAWPYXKBD-IEOSBIPESA-N 0.000 description 4
- IKYKEVDKGZYRMQ-PDBXOOCHSA-N (9Z,12Z,15Z)-octadecatrien-1-ol Chemical compound CC\C=C/C\C=C/C\C=C/CCCCCCCCO IKYKEVDKGZYRMQ-PDBXOOCHSA-N 0.000 description 3
- 229960000549 4-dimethylaminophenol Drugs 0.000 description 3
- ZOXJGFHDIHLPTG-UHFFFAOYSA-N Boron Chemical compound [B] ZOXJGFHDIHLPTG-UHFFFAOYSA-N 0.000 description 3
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 description 3
- JGFZNNIVVJXRND-UHFFFAOYSA-N N,N-Diisopropylethylamine (DIPEA) Chemical compound CCN(C(C)C)C(C)C JGFZNNIVVJXRND-UHFFFAOYSA-N 0.000 description 3
- XUIMIQQOPSSXEZ-UHFFFAOYSA-N Silicon Chemical compound [Si] XUIMIQQOPSSXEZ-UHFFFAOYSA-N 0.000 description 3
- KEAYESYHFKHZAL-UHFFFAOYSA-N Sodium Chemical compound [Na] KEAYESYHFKHZAL-UHFFFAOYSA-N 0.000 description 3
- 102000036639 antigens Human genes 0.000 description 3
- 108091007433 antigens Proteins 0.000 description 3
- 230000015572 biosynthetic process Effects 0.000 description 3
- 229910052796 boron Inorganic materials 0.000 description 3
- 230000001413 cellular effect Effects 0.000 description 3
- 239000003937 drug carrier Substances 0.000 description 3
- 125000005842 heteroatom Chemical group 0.000 description 3
- UVNRLSCOYBEJTM-UHFFFAOYSA-N linolenic alcohol Natural products CCCCCCCCC=C/CC=C/CC=C/CCO UVNRLSCOYBEJTM-UHFFFAOYSA-N 0.000 description 3
- 239000000546 pharmaceutical excipient Substances 0.000 description 3
- WTJKGGKOPKCXLL-RRHRGVEJSA-N phosphatidylcholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCC=CCCCCCCCC WTJKGGKOPKCXLL-RRHRGVEJSA-N 0.000 description 3
- 150000003904 phospholipids Chemical class 0.000 description 3
- 239000002504 physiological saline solution Substances 0.000 description 3
- 239000000651 prodrug Substances 0.000 description 3
- 229940002612 prodrug Drugs 0.000 description 3
- 229930195734 saturated hydrocarbon Natural products 0.000 description 3
- 229910052710 silicon Inorganic materials 0.000 description 3
- 239000010703 silicon Substances 0.000 description 3
- 239000012312 sodium hydride Substances 0.000 description 3
- 150000003431 steroids Chemical class 0.000 description 3
- 239000000758 substrate Substances 0.000 description 3
- KZJWDPNRJALLNS-VPUBHVLGSA-N (-)-beta-Sitosterol Natural products O[C@@H]1CC=2[C@@](C)([C@@H]3[C@H]([C@H]4[C@@](C)([C@H]([C@H](CC[C@@H](C(C)C)CC)C)CC4)CC3)CC=2)CC1 KZJWDPNRJALLNS-VPUBHVLGSA-N 0.000 description 2
- CSVWWLUMXNHWSU-UHFFFAOYSA-N (22E)-(24xi)-24-ethyl-5alpha-cholest-22-en-3beta-ol Natural products C1CC2CC(O)CCC2(C)C2C1C1CCC(C(C)C=CC(CC)C(C)C)C1(C)CC2 CSVWWLUMXNHWSU-UHFFFAOYSA-N 0.000 description 2
- RQOCXCFLRBRBCS-UHFFFAOYSA-N (22E)-cholesta-5,7,22-trien-3beta-ol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)C=CCC(C)C)CCC33)C)C3=CC=C21 RQOCXCFLRBRBCS-UHFFFAOYSA-N 0.000 description 2
- WCGUUGGRBIKTOS-GPOJBZKASA-N (3beta)-3-hydroxyurs-12-en-28-oic acid Chemical compound C1C[C@H](O)C(C)(C)[C@@H]2CC[C@@]3(C)[C@]4(C)CC[C@@]5(C(O)=O)CC[C@@H](C)[C@H](C)[C@H]5C4=CC[C@@H]3[C@]21C WCGUUGGRBIKTOS-GPOJBZKASA-N 0.000 description 2
- NRJAVPSFFCBXDT-HUESYALOSA-N 1,2-distearoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCCCCCCCCCCC NRJAVPSFFCBXDT-HUESYALOSA-N 0.000 description 2
- KLEXDBGYSOIREE-UHFFFAOYSA-N 24xi-n-propylcholesterol Natural products C1C=C2CC(O)CCC2(C)C2C1C1CCC(C(C)CCC(CCC)C(C)C)C1(C)CC2 KLEXDBGYSOIREE-UHFFFAOYSA-N 0.000 description 2
- 108020000948 Antisense Oligonucleotides Proteins 0.000 description 2
- 108020005544 Antisense RNA Proteins 0.000 description 2
- OILXMJHPFNGGTO-NRHJOKMGSA-N Brassicasterol Natural products O[C@@H]1CC=2[C@@](C)([C@@H]3[C@H]([C@H]4[C@](C)([C@H]([C@@H](/C=C/[C@H](C(C)C)C)C)CC4)CC3)CC=2)CC1 OILXMJHPFNGGTO-NRHJOKMGSA-N 0.000 description 2
- 108091079001 CRISPR RNA Proteins 0.000 description 2
- SGNBVLSWZMBQTH-FGAXOLDCSA-N Campesterol Natural products O[C@@H]1CC=2[C@@](C)([C@@H]3[C@H]([C@H]4[C@@](C)([C@H]([C@H](CC[C@H](C(C)C)C)C)CC4)CC3)CC=2)CC1 SGNBVLSWZMBQTH-FGAXOLDCSA-N 0.000 description 2
- OKTJSMMVPCPJKN-UHFFFAOYSA-N Carbon Chemical group [C] OKTJSMMVPCPJKN-UHFFFAOYSA-N 0.000 description 2
- CURLTUGMZLYLDI-UHFFFAOYSA-N Carbon dioxide Chemical compound O=C=O CURLTUGMZLYLDI-UHFFFAOYSA-N 0.000 description 2
- LPZCCMIISIBREI-MTFRKTCUSA-N Citrostadienol Natural products CC=C(CC[C@@H](C)[C@H]1CC[C@H]2C3=CC[C@H]4[C@H](C)[C@@H](O)CC[C@]4(C)[C@H]3CC[C@]12C)C(C)C LPZCCMIISIBREI-MTFRKTCUSA-N 0.000 description 2
- ARVGMISWLZPBCH-UHFFFAOYSA-N Dehydro-beta-sitosterol Natural products C1C(O)CCC2(C)C(CCC3(C(C(C)CCC(CC)C(C)C)CCC33)C)C3=CC=C21 ARVGMISWLZPBCH-UHFFFAOYSA-N 0.000 description 2
- DNVPQKQSNYMLRS-NXVQYWJNSA-N Ergosterol Natural products CC(C)[C@@H](C)C=C[C@H](C)[C@H]1CC[C@H]2C3=CC=C4C[C@@H](O)CC[C@]4(C)[C@@H]3CC[C@]12C DNVPQKQSNYMLRS-NXVQYWJNSA-N 0.000 description 2
- 239000004471 Glycine Substances 0.000 description 2
- 108020005004 Guide RNA Proteins 0.000 description 2
- BTEISVKTSQLKST-UHFFFAOYSA-N Haliclonasterol Natural products CC(C=CC(C)C(C)(C)C)C1CCC2C3=CC=C4CC(O)CCC4(C)C3CCC12C BTEISVKTSQLKST-UHFFFAOYSA-N 0.000 description 2
- 108091026898 Leader sequence (mRNA) Proteins 0.000 description 2
- 108010087230 Sincalide Proteins 0.000 description 2
- 108091027967 Small hairpin RNA Proteins 0.000 description 2
- VMHLLURERBWHNL-UHFFFAOYSA-M Sodium acetate Chemical compound [Na+].CC([O-])=O VMHLLURERBWHNL-UHFFFAOYSA-M 0.000 description 2
- 229930182558 Sterol Natural products 0.000 description 2
- 108091046869 Telomeric non-coding RNA Proteins 0.000 description 2
- 108091036066 Three prime untranslated region Proteins 0.000 description 2
- OILXMJHPFNGGTO-ZRUUVFCLSA-N UNPD197407 Natural products C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)C=C[C@H](C)C(C)C)[C@@]1(C)CC2 OILXMJHPFNGGTO-ZRUUVFCLSA-N 0.000 description 2
- 230000002378 acidificating effect Effects 0.000 description 2
- 229940087168 alpha tocopherol Drugs 0.000 description 2
- 239000000074 antisense oligonucleotide Substances 0.000 description 2
- 238000012230 antisense oligonucleotides Methods 0.000 description 2
- 239000008346 aqueous phase Substances 0.000 description 2
- MJVXAPPOFPTTCA-UHFFFAOYSA-N beta-Sistosterol Natural products CCC(CCC(C)C1CCC2C3CC=C4C(C)C(O)CCC4(C)C3CCC12C)C(C)C MJVXAPPOFPTTCA-UHFFFAOYSA-N 0.000 description 2
- NJKOMDUNNDKEAI-UHFFFAOYSA-N beta-sitosterol Natural products CCC(CCC(C)C1CCC2(C)C3CC=C4CC(O)CCC4C3CCC12C)C(C)C NJKOMDUNNDKEAI-UHFFFAOYSA-N 0.000 description 2
- 230000004071 biological effect Effects 0.000 description 2
- OILXMJHPFNGGTO-ZAUYPBDWSA-N brassicasterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)/C=C/[C@H](C)C(C)C)[C@@]1(C)CC2 OILXMJHPFNGGTO-ZAUYPBDWSA-N 0.000 description 2
- 235000004420 brassicasterol Nutrition 0.000 description 2
- SGNBVLSWZMBQTH-PODYLUTMSA-N campesterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CC[C@@H](C)C(C)C)[C@@]1(C)CC2 SGNBVLSWZMBQTH-PODYLUTMSA-N 0.000 description 2
- 235000000431 campesterol Nutrition 0.000 description 2
- 239000000969 carrier Substances 0.000 description 2
- 238000010609 cell counting kit-8 assay Methods 0.000 description 2
- 238000012512 characterization method Methods 0.000 description 2
- 239000000460 chlorine Chemical group 0.000 description 2
- 229940107161 cholesterol Drugs 0.000 description 2
- 239000003184 complementary RNA Substances 0.000 description 2
- 230000000694 effects Effects 0.000 description 2
- DNVPQKQSNYMLRS-SOWFXMKYSA-N ergosterol Chemical compound C1[C@@H](O)CC[C@]2(C)[C@H](CC[C@]3([C@H]([C@H](C)/C=C/[C@@H](C)C(C)C)CC[C@H]33)C)C3=CC=C21 DNVPQKQSNYMLRS-SOWFXMKYSA-N 0.000 description 2
- 238000002474 experimental method Methods 0.000 description 2
- 239000012634 fragment Substances 0.000 description 2
- 125000000524 functional group Chemical group 0.000 description 2
- XGVJWXAYKUHDOO-UHFFFAOYSA-N galanthidine Natural products C1CN2CC3=CC=4OCOC=4C=C3C3C2C1=CC(O)C3O XGVJWXAYKUHDOO-UHFFFAOYSA-N 0.000 description 2
- 239000011521 glass Substances 0.000 description 2
- 238000004128 high performance liquid chromatography Methods 0.000 description 2
- 230000007062 hydrolysis Effects 0.000 description 2
- 238000006460 hydrolysis reaction Methods 0.000 description 2
- 230000003301 hydrolyzing effect Effects 0.000 description 2
- 230000031700 light absorption Effects 0.000 description 2
- 238000004895 liquid chromatography mass spectrometry Methods 0.000 description 2
- XGVJWXAYKUHDOO-DANNLKNASA-N lycorine Chemical compound C1CN2CC3=CC=4OCOC=4C=C3[C@H]3[C@H]2C1=C[C@H](O)[C@H]3O XGVJWXAYKUHDOO-DANNLKNASA-N 0.000 description 2
- KQAOMBGKIWRWNA-UHFFFAOYSA-N lycorine Natural products OC1C=C2CCN3C2C(C1O)c4cc5OCOc5cc34 KQAOMBGKIWRWNA-UHFFFAOYSA-N 0.000 description 2
- 239000012528 membrane Substances 0.000 description 2
- 230000004048 modification Effects 0.000 description 2
- 238000012986 modification Methods 0.000 description 2
- 239000013642 negative control Substances 0.000 description 2
- 230000000144 pharmacologic effect Effects 0.000 description 2
- 239000008055 phosphate buffer solution Substances 0.000 description 2
- 238000011321 prophylaxis Methods 0.000 description 2
- 108090000623 proteins and genes Proteins 0.000 description 2
- 229920006395 saturated elastomer Polymers 0.000 description 2
- IZTQOLKUZKXIRV-YRVFCXMDSA-N sincalide Chemical compound C([C@@H](C(=O)N[C@@H](CCSC)C(=O)NCC(=O)N[C@@H](CC=1C2=CC=CC=C2NC=1)C(=O)N[C@@H](CCSC)C(=O)N[C@@H](CC(O)=O)C(=O)N[C@@H](CC=1C=CC=CC=1)C(N)=O)NC(=O)[C@@H](N)CC(O)=O)C1=CC=C(OS(O)(=O)=O)C=C1 IZTQOLKUZKXIRV-YRVFCXMDSA-N 0.000 description 2
- KZJWDPNRJALLNS-VJSFXXLFSA-N sitosterol Chemical compound C1C=C2C[C@@H](O)CC[C@]2(C)[C@@H]2[C@@H]1[C@@H]1CC[C@H]([C@H](C)CC[C@@H](CC)C(C)C)[C@@]1(C)CC2 KZJWDPNRJALLNS-VJSFXXLFSA-N 0.000 description 2
- 235000015500 sitosterol Nutrition 0.000 description 2
- 229950005143 sitosterol Drugs 0.000 description 2
- 239000001632 sodium acetate Substances 0.000 description 2
- 235000017281 sodium acetate Nutrition 0.000 description 2
- 239000011780 sodium chloride Substances 0.000 description 2
- 150000003432 sterols Chemical class 0.000 description 2
- 235000003702 sterols Nutrition 0.000 description 2
- YLQBMQCUIZJEEH-UHFFFAOYSA-N tetrahydrofuran Natural products C=1C=COC=1 YLQBMQCUIZJEEH-UHFFFAOYSA-N 0.000 description 2
- 229960000984 tocofersolan Drugs 0.000 description 2
- 231100000419 toxicity Toxicity 0.000 description 2
- 230000001988 toxicity Effects 0.000 description 2
- 238000000108 ultra-filtration Methods 0.000 description 2
- PLSAJKYPRJGMHO-UHFFFAOYSA-N ursolic acid Natural products CC1CCC2(CCC3(C)C(C=CC4C5(C)CCC(O)C(C)(C)C5CCC34C)C2C1C)C(=O)O PLSAJKYPRJGMHO-UHFFFAOYSA-N 0.000 description 2
- 229940096998 ursolic acid Drugs 0.000 description 2
- 239000003981 vehicle Substances 0.000 description 2
- 239000002076 α-tocopherol Substances 0.000 description 2
- 235000004835 α-tocopherol Nutrition 0.000 description 2
- 125000000008 (C1-C10) alkyl group Chemical group 0.000 description 1
- 125000003161 (C1-C6) alkylene group Chemical group 0.000 description 1
- CITHEXJVPOWHKC-UUWRZZSWSA-N 1,2-di-O-myristoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCCCCCCC CITHEXJVPOWHKC-UUWRZZSWSA-N 0.000 description 1
- KILNVBDSWZSGLL-KXQOOQHDSA-N 1,2-dihexadecanoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCCCCCCCCCCCCCC KILNVBDSWZSGLL-KXQOOQHDSA-N 0.000 description 1
- SLKDGVPOSSLUAI-PGUFJCEWSA-N 1,2-dihexadecanoyl-sn-glycero-3-phosphoethanolamine zwitterion Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP(O)(=O)OCCN)OC(=O)CCCCCCCCCCCCCCC SLKDGVPOSSLUAI-PGUFJCEWSA-N 0.000 description 1
- UVBYMVOUBXYSFV-XUTVFYLZSA-N 1-methylpseudouridine Chemical compound O=C1NC(=O)N(C)C=C1[C@H]1[C@H](O)[C@H](O)[C@@H](CO)O1 UVBYMVOUBXYSFV-XUTVFYLZSA-N 0.000 description 1
- IIZPXYDJLKNOIY-JXPKJXOSSA-N 1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphocholine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP([O-])(=O)OCC[N+](C)(C)C)OC(=O)CCC\C=C/C\C=C/C\C=C/C\C=C/CCCCC IIZPXYDJLKNOIY-JXPKJXOSSA-N 0.000 description 1
- 238000005160 1H NMR spectroscopy Methods 0.000 description 1
- CHHHXKFHOYLYRE-UHFFFAOYSA-M 2,4-Hexadienoic acid, potassium salt (1:1), (2E,4E)- Chemical compound [K+].CC=CC=CC([O-])=O CHHHXKFHOYLYRE-UHFFFAOYSA-M 0.000 description 1
- QKNYBSVHEMOAJP-UHFFFAOYSA-N 2-amino-2-(hydroxymethyl)propane-1,3-diol;hydron;chloride Chemical compound Cl.OCC(N)(CO)CO QKNYBSVHEMOAJP-UHFFFAOYSA-N 0.000 description 1
- ZXIATBNUWJBBGT-JXOAFFINSA-N 5-methoxyuridine Chemical compound O=C1NC(=O)C(OC)=CN1[C@H]1[C@H](O)[C@H](O)[C@@H](CO)O1 ZXIATBNUWJBBGT-JXOAFFINSA-N 0.000 description 1
- LRSASMSXMSNRBT-UHFFFAOYSA-N 5-methylcytosine Chemical compound CC1=CNC(=O)N=C1N LRSASMSXMSNRBT-UHFFFAOYSA-N 0.000 description 1
- HBAQYPYDRFILMT-UHFFFAOYSA-N 8-[3-(1-cyclopropylpyrazol-4-yl)-1H-pyrazolo[4,3-d]pyrimidin-5-yl]-3-methyl-3,8-diazabicyclo[3.2.1]octan-2-one Chemical class C1(CC1)N1N=CC(=C1)C1=NNC2=C1N=C(N=C2)N1C2C(N(CC1CC2)C)=O HBAQYPYDRFILMT-UHFFFAOYSA-N 0.000 description 1
- 108010088751 Albumins Proteins 0.000 description 1
- 102000009027 Albumins Human genes 0.000 description 1
- 235000007319 Avena orientalis Nutrition 0.000 description 1
- 235000007558 Avena sp Nutrition 0.000 description 1
- 108010017384 Blood Proteins Proteins 0.000 description 1
- 102000004506 Blood Proteins Human genes 0.000 description 1
- 241000283690 Bos taurus Species 0.000 description 1
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 1
- 125000006374 C2-C10 alkenyl group Chemical group 0.000 description 1
- 238000011740 C57BL/6 mouse Methods 0.000 description 1
- 108091033409 CRISPR Proteins 0.000 description 1
- 238000010354 CRISPR gene editing Methods 0.000 description 1
- 241000283707 Capra Species 0.000 description 1
- OKTJSMMVPCPJKN-NJFSPNSNSA-N Carbon-14 Chemical compound [14C] OKTJSMMVPCPJKN-NJFSPNSNSA-N 0.000 description 1
- 108090000994 Catalytic RNA Proteins 0.000 description 1
- 102000053642 Catalytic RNA Human genes 0.000 description 1
- ZAMOUSCENKQFHK-UHFFFAOYSA-N Chlorine atom Chemical group [Cl] ZAMOUSCENKQFHK-UHFFFAOYSA-N 0.000 description 1
- YZCKVEUIGOORGS-OUBTZVSYSA-N Deuterium Chemical compound [2H] YZCKVEUIGOORGS-OUBTZVSYSA-N 0.000 description 1
- 239000006144 Dulbecco’s modified Eagle's medium Substances 0.000 description 1
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 1
- 241000283073 Equus caballus Species 0.000 description 1
- VGGSQFUCUMXWEO-UHFFFAOYSA-N Ethene Chemical compound C=C VGGSQFUCUMXWEO-UHFFFAOYSA-N 0.000 description 1
- 239000005977 Ethylene Substances 0.000 description 1
- 241000282326 Felis catus Species 0.000 description 1
- PXGOKWXKJXAPGV-UHFFFAOYSA-N Fluorine Chemical group FF PXGOKWXKJXAPGV-UHFFFAOYSA-N 0.000 description 1
- 102000006395 Globulins Human genes 0.000 description 1
- 108010044091 Globulins Proteins 0.000 description 1
- JZNWSCPGTDBMEW-UHFFFAOYSA-N Glycerophosphorylethanolamin Natural products NCCOP(O)(=O)OCC(O)CO JZNWSCPGTDBMEW-UHFFFAOYSA-N 0.000 description 1
- 102000003886 Glycoproteins Human genes 0.000 description 1
- 108090000288 Glycoproteins Proteins 0.000 description 1
- 108091006905 Human Serum Albumin Proteins 0.000 description 1
- 102000008100 Human Serum Albumin Human genes 0.000 description 1
- PIWKPBJCKXDKJR-UHFFFAOYSA-N Isoflurane Chemical compound FC(F)OC(Cl)C(F)(F)F PIWKPBJCKXDKJR-UHFFFAOYSA-N 0.000 description 1
- 229930194542 Keto Natural products 0.000 description 1
- 239000004166 Lanolin Substances 0.000 description 1
- 102000004895 Lipoproteins Human genes 0.000 description 1
- 108090001030 Lipoproteins Proteins 0.000 description 1
- 241000282567 Macaca fascicularis Species 0.000 description 1
- 241000282560 Macaca mulatta Species 0.000 description 1
- 241000124008 Mammalia Species 0.000 description 1
- 102000006833 Multifunctional Enzymes Human genes 0.000 description 1
- 108010047290 Multifunctional Enzymes Proteins 0.000 description 1
- 241000699666 Mus <mouse, genus> Species 0.000 description 1
- 108700001237 Nucleic Acid-Based Vaccines Proteins 0.000 description 1
- 108700026244 Open Reading Frames Proteins 0.000 description 1
- 229910019142 PO4 Inorganic materials 0.000 description 1
- 241001494479 Pecora Species 0.000 description 1
- 241000288906 Primates Species 0.000 description 1
- 102000007327 Protamines Human genes 0.000 description 1
- 108010007568 Protamines Proteins 0.000 description 1
- 229930185560 Pseudouridine Natural products 0.000 description 1
- PTJWIQPHWPFNBW-UHFFFAOYSA-N Pseudouridine C Natural products OC1C(O)C(CO)OC1C1=CNC(=O)NC1=O PTJWIQPHWPFNBW-UHFFFAOYSA-N 0.000 description 1
- 241000283984 Rodentia Species 0.000 description 1
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical compound [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 1
- 241000282898 Sus scrofa Species 0.000 description 1
- 239000007984 Tris EDTA buffer Substances 0.000 description 1
- YZCKVEUIGOORGS-NJFSPNSNSA-N Tritium Chemical compound [3H] YZCKVEUIGOORGS-NJFSPNSNSA-N 0.000 description 1
- 229920004890 Triton X-100 Polymers 0.000 description 1
- 239000013504 Triton X-100 Substances 0.000 description 1
- ONVRFFJRASWLTG-UHFFFAOYSA-N [(1-amino-2-naphthalen-1-ylethylidene)amino] morpholine-4-carboxylate Chemical compound C=1C=CC2=CC=CC=C2C=1CC(N)=NOC(=O)N1CCOCC1 ONVRFFJRASWLTG-UHFFFAOYSA-N 0.000 description 1
- NRLNQCOGCKAESA-KWXKLSQISA-N [(6z,9z,28z,31z)-heptatriaconta-6,9,28,31-tetraen-19-yl] 4-(dimethylamino)butanoate Chemical compound CCCCC\C=C/C\C=C/CCCCCCCCC(OC(=O)CCCN(C)C)CCCCCCCC\C=C/C\C=C/CCCCC NRLNQCOGCKAESA-KWXKLSQISA-N 0.000 description 1
- ABRVLXLNVJHDRQ-UHFFFAOYSA-N [2-pyridin-3-yl-6-(trifluoromethyl)pyridin-4-yl]methanamine Chemical compound FC(C1=CC(=CC(=N1)C=1C=NC=CC=1)CN)(F)F ABRVLXLNVJHDRQ-UHFFFAOYSA-N 0.000 description 1
- 238000002835 absorbance Methods 0.000 description 1
- 238000010521 absorption reaction Methods 0.000 description 1
- DPXJVFZANSGRMM-UHFFFAOYSA-N acetic acid;2,3,4,5,6-pentahydroxyhexanal;sodium Chemical compound [Na].CC(O)=O.OCC(O)C(O)C(O)C(O)C=O DPXJVFZANSGRMM-UHFFFAOYSA-N 0.000 description 1
- UGAPHEBNTGUMBB-UHFFFAOYSA-N acetic acid;ethyl acetate Chemical compound CC(O)=O.CCOC(C)=O UGAPHEBNTGUMBB-UHFFFAOYSA-N 0.000 description 1
- 239000002253 acid Substances 0.000 description 1
- 230000009471 action Effects 0.000 description 1
- 230000003213 activating effect Effects 0.000 description 1
- 230000003044 adaptive effect Effects 0.000 description 1
- 230000002776 aggregation Effects 0.000 description 1
- 238000004220 aggregation Methods 0.000 description 1
- 208000026935 allergic disease Diseases 0.000 description 1
- PNEYBMLMFCGWSK-UHFFFAOYSA-N aluminium oxide Inorganic materials [O-2].[O-2].[O-2].[Al+3].[Al+3] PNEYBMLMFCGWSK-UHFFFAOYSA-N 0.000 description 1
- CEGOLXSVJUTHNZ-UHFFFAOYSA-K aluminium tristearate Chemical compound [Al+3].CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O.CCCCCCCCCCCCCCCCCC([O-])=O CEGOLXSVJUTHNZ-UHFFFAOYSA-K 0.000 description 1
- 229940063655 aluminum stearate Drugs 0.000 description 1
- 230000003444 anaesthetic effect Effects 0.000 description 1
- 238000003556 assay Methods 0.000 description 1
- 238000011914 asymmetric synthesis Methods 0.000 description 1
- WGDUUQDYDIIBKT-UHFFFAOYSA-N beta-Pseudouridine Natural products OC1OC(CN2C=CC(=O)NC2=O)C(O)C1O WGDUUQDYDIIBKT-UHFFFAOYSA-N 0.000 description 1
- 230000008512 biological response Effects 0.000 description 1
- 238000003390 bioluminescence detection Methods 0.000 description 1
- 239000000872 buffer Substances 0.000 description 1
- 229910052799 carbon Inorganic materials 0.000 description 1
- 239000001569 carbon dioxide Substances 0.000 description 1
- 229910002092 carbon dioxide Inorganic materials 0.000 description 1
- 239000001768 carboxy methyl cellulose Substances 0.000 description 1
- 230000003833 cell viability Effects 0.000 description 1
- 239000001913 cellulose Substances 0.000 description 1
- 229920002678 cellulose Polymers 0.000 description 1
- 235000010980 cellulose Nutrition 0.000 description 1
- 150000001783 ceramides Chemical class 0.000 description 1
- 238000007385 chemical modification Methods 0.000 description 1
- 229910052801 chlorine Chemical group 0.000 description 1
- 230000008878 coupling Effects 0.000 description 1
- 238000010168 coupling process Methods 0.000 description 1
- 238000005859 coupling reaction Methods 0.000 description 1
- 238000002425 crystallisation Methods 0.000 description 1
- 230000008025 crystallization Effects 0.000 description 1
- 231100000135 cytotoxicity Toxicity 0.000 description 1
- 230000003013 cytotoxicity Effects 0.000 description 1
- 238000007405 data analysis Methods 0.000 description 1
- DEZRYPDIMOWBDS-UHFFFAOYSA-N dcm dichloromethane Chemical compound ClCCl.ClCCl DEZRYPDIMOWBDS-UHFFFAOYSA-N 0.000 description 1
- 238000001514 detection method Methods 0.000 description 1
- 229910052805 deuterium Inorganic materials 0.000 description 1
- 238000011161 development Methods 0.000 description 1
- 150000001982 diacylglycerols Chemical class 0.000 description 1
- 125000005265 dialkylamine group Chemical class 0.000 description 1
- 150000001985 dialkylglycerols Chemical class 0.000 description 1
- 235000014113 dietary fatty acids Nutrition 0.000 description 1
- GXGAKHNRMVGRPK-UHFFFAOYSA-N dimagnesium;dioxido-bis[[oxido(oxo)silyl]oxy]silane Chemical compound [Mg+2].[Mg+2].[O-][Si](=O)O[Si]([O-])([O-])O[Si]([O-])=O GXGAKHNRMVGRPK-UHFFFAOYSA-N 0.000 description 1
- ZPWVASYFFYYZEW-UHFFFAOYSA-L dipotassium hydrogen phosphate Chemical compound [K+].[K+].OP([O-])([O-])=O ZPWVASYFFYYZEW-UHFFFAOYSA-L 0.000 description 1
- 229910000396 dipotassium phosphate Inorganic materials 0.000 description 1
- 235000019797 dipotassium phosphate Nutrition 0.000 description 1
- BNIILDVGGAEEIG-UHFFFAOYSA-L disodium hydrogen phosphate Chemical compound [Na+].[Na+].OP([O-])([O-])=O BNIILDVGGAEEIG-UHFFFAOYSA-L 0.000 description 1
- 239000006185 dispersion Substances 0.000 description 1
- 238000009826 distribution Methods 0.000 description 1
- 239000002552 dosage form Substances 0.000 description 1
- 239000003651 drinking water Substances 0.000 description 1
- 235000020188 drinking water Nutrition 0.000 description 1
- 229940079593 drug Drugs 0.000 description 1
- 238000001035 drying Methods 0.000 description 1
- 238000002296 dynamic light scattering Methods 0.000 description 1
- 239000003792 electrolyte Substances 0.000 description 1
- 238000002330 electrospray ionisation mass spectrometry Methods 0.000 description 1
- 150000002085 enols Chemical class 0.000 description 1
- BEFDCLMNVWHSGT-UHFFFAOYSA-N ethenylcyclopentane Chemical compound C=CC1CCCC1 BEFDCLMNVWHSGT-UHFFFAOYSA-N 0.000 description 1
- 238000000605 extraction Methods 0.000 description 1
- 239000000194 fatty acid Substances 0.000 description 1
- 229930195729 fatty acid Natural products 0.000 description 1
- 150000004665 fatty acids Chemical class 0.000 description 1
- 239000012091 fetal bovine serum Substances 0.000 description 1
- 238000011049 filling Methods 0.000 description 1
- 238000001914 filtration Methods 0.000 description 1
- GNBHRKFJIUUOQI-UHFFFAOYSA-N fluorescein Chemical compound O1C(=O)C2=CC=CC=C2C21C1=CC=C(O)C=C1OC1=CC(O)=CC=C21 GNBHRKFJIUUOQI-UHFFFAOYSA-N 0.000 description 1
- 238000000799 fluorescence microscopy Methods 0.000 description 1
- 238000001506 fluorescence spectroscopy Methods 0.000 description 1
- 239000011737 fluorine Chemical group 0.000 description 1
- 229910052731 fluorine Inorganic materials 0.000 description 1
- 238000001415 gene therapy Methods 0.000 description 1
- 125000005456 glyceride group Chemical group 0.000 description 1
- 229960002449 glycine Drugs 0.000 description 1
- 230000036541 health Effects 0.000 description 1
- 125000004404 heteroalkyl group Chemical group 0.000 description 1
- 125000004435 hydrogen atom Chemical group [H]* 0.000 description 1
- 238000005286 illumination Methods 0.000 description 1
- 230000028993 immune response Effects 0.000 description 1
- 238000011503 in vivo imaging Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000001939 inductive effect Effects 0.000 description 1
- 230000002401 inhibitory effect Effects 0.000 description 1
- 239000007924 injection Substances 0.000 description 1
- 238000002347 injection Methods 0.000 description 1
- 150000007529 inorganic bases Chemical class 0.000 description 1
- 239000007928 intraperitoneal injection Substances 0.000 description 1
- 150000002500 ions Chemical class 0.000 description 1
- 230000007794 irritation Effects 0.000 description 1
- 229960002725 isoflurane Drugs 0.000 description 1
- 125000000468 ketone group Chemical group 0.000 description 1
- 210000003292 kidney cell Anatomy 0.000 description 1
- 229940039717 lanolin Drugs 0.000 description 1
- 235000019388 lanolin Nutrition 0.000 description 1
- 239000000787 lecithin Substances 0.000 description 1
- 229940067606 lecithin Drugs 0.000 description 1
- 235000010445 lecithin Nutrition 0.000 description 1
- 230000000670 limiting effect Effects 0.000 description 1
- 239000002502 liposome Substances 0.000 description 1
- 238000003670 luciferase enzyme activity assay Methods 0.000 description 1
- 239000000391 magnesium silicate Substances 0.000 description 1
- 229940099273 magnesium trisilicate Drugs 0.000 description 1
- 229910000386 magnesium trisilicate Inorganic materials 0.000 description 1
- 235000019793 magnesium trisilicate Nutrition 0.000 description 1
- 230000001404 mediated effect Effects 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- COTNUBDHGSIOTA-UHFFFAOYSA-N meoh methanol Chemical compound OC.OC COTNUBDHGSIOTA-UHFFFAOYSA-N 0.000 description 1
- 230000002503 metabolic effect Effects 0.000 description 1
- 125000001570 methylene group Chemical group [H]C([H])([*:1])[*:2] 0.000 description 1
- 150000007522 mineralic acids Chemical class 0.000 description 1
- PEECTLLHENGOKU-UHFFFAOYSA-N n,n-dimethylpyridin-4-amine Chemical compound CN(C)C1=CC=NC=C1.CN(C)C1=CC=NC=C1 PEECTLLHENGOKU-UHFFFAOYSA-N 0.000 description 1
- VWBWQOUWDOULQN-UHFFFAOYSA-N nmp n-methylpyrrolidone Chemical compound CN1CCCC1=O.CN1CCCC1=O VWBWQOUWDOULQN-UHFFFAOYSA-N 0.000 description 1
- 231100000252 nontoxic Toxicity 0.000 description 1
- 230000003000 nontoxic effect Effects 0.000 description 1
- 229940023146 nucleic acid vaccine Drugs 0.000 description 1
- 239000003921 oil Substances 0.000 description 1
- 230000003287 optical effect Effects 0.000 description 1
- 150000007524 organic acids Chemical class 0.000 description 1
- 235000005985 organic acids Nutrition 0.000 description 1
- 150000007530 organic bases Chemical class 0.000 description 1
- AICOOMRHRUFYCM-ZRRPKQBOSA-N oxazine, 1 Chemical compound C([C@@H]1[C@H](C(C[C@]2(C)[C@@H]([C@H](C)N(C)C)[C@H](O)C[C@]21C)=O)CC1=CC2)C[C@H]1[C@@]1(C)[C@H]2N=C(C(C)C)OC1 AICOOMRHRUFYCM-ZRRPKQBOSA-N 0.000 description 1
- 230000036961 partial effect Effects 0.000 description 1
- 235000021317 phosphate Nutrition 0.000 description 1
- 150000008103 phosphatidic acids Chemical class 0.000 description 1
- 150000008104 phosphatidylethanolamines Chemical class 0.000 description 1
- 150000003013 phosphoric acid derivatives Chemical class 0.000 description 1
- 229950004354 phosphorylcholine Drugs 0.000 description 1
- 230000000704 physical effect Effects 0.000 description 1
- 229920000058 polyacrylate Polymers 0.000 description 1
- 230000008488 polyadenylation Effects 0.000 description 1
- 229920001184 polypeptide Polymers 0.000 description 1
- 239000001267 polyvinylpyrrolidone Substances 0.000 description 1
- 229920000036 polyvinylpyrrolidone Polymers 0.000 description 1
- 235000013855 polyvinylpyrrolidone Nutrition 0.000 description 1
- 239000004302 potassium sorbate Substances 0.000 description 1
- 235000010241 potassium sorbate Nutrition 0.000 description 1
- 229940069338 potassium sorbate Drugs 0.000 description 1
- 230000003449 preventive effect Effects 0.000 description 1
- 108090000765 processed proteins & peptides Proteins 0.000 description 1
- 102000004196 processed proteins & peptides Human genes 0.000 description 1
- 239000000047 product Substances 0.000 description 1
- QQONPFPTGQHPMA-UHFFFAOYSA-N propylene Natural products CC=C QQONPFPTGQHPMA-UHFFFAOYSA-N 0.000 description 1
- 125000004805 propylene group Chemical group [H]C([H])([H])C([H])([*:1])C([H])([H])[*:2] 0.000 description 1
- 229950008679 protamine sulfate Drugs 0.000 description 1
- 102000004169 proteins and genes Human genes 0.000 description 1
- PTJWIQPHWPFNBW-GBNDHIKLSA-N pseudouridine Chemical compound O[C@@H]1[C@H](O)[C@@H](CO)O[C@H]1C1=CNC(=O)NC1=O PTJWIQPHWPFNBW-GBNDHIKLSA-N 0.000 description 1
- 238000000746 purification Methods 0.000 description 1
- 238000010791 quenching Methods 0.000 description 1
- 230000000171 quenching effect Effects 0.000 description 1
- 230000002285 radioactive effect Effects 0.000 description 1
- 230000002829 reductive effect Effects 0.000 description 1
- 108091092562 ribozyme Proteins 0.000 description 1
- 239000012488 sample solution Substances 0.000 description 1
- 239000000344 soap Substances 0.000 description 1
- 235000019812 sodium carboxymethyl cellulose Nutrition 0.000 description 1
- 229920001027 sodium carboxymethylcellulose Polymers 0.000 description 1
- 239000004334 sorbic acid Substances 0.000 description 1
- 235000010199 sorbic acid Nutrition 0.000 description 1
- 229940075582 sorbic acid Drugs 0.000 description 1
- 229960005322 streptomycin Drugs 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 230000008685 targeting Effects 0.000 description 1
- WHRNULOCNSKMGB-UHFFFAOYSA-N tetrahydrofuran thf Chemical compound C1CCOC1.C1CCOC1 WHRNULOCNSKMGB-UHFFFAOYSA-N 0.000 description 1
- 125000000383 tetramethylene group Chemical group [H]C([H])([*:1])C([H])([H])C([H])([H])C([H])([H])[*:2] 0.000 description 1
- PHCBRBWANGJMHS-UHFFFAOYSA-J tetrasodium;disulfate Chemical compound [Na+].[Na+].[Na+].[Na+].[O-]S([O-])(=O)=O.[O-]S([O-])(=O)=O PHCBRBWANGJMHS-UHFFFAOYSA-J 0.000 description 1
- 238000002560 therapeutic procedure Methods 0.000 description 1
- 238000003354 tissue distribution assay Methods 0.000 description 1
- 231100000048 toxicity data Toxicity 0.000 description 1
- 229910052722 tritium Inorganic materials 0.000 description 1
- GPRLSGONYQIRFK-MNYXATJNSA-N triton Chemical compound [3H+] GPRLSGONYQIRFK-MNYXATJNSA-N 0.000 description 1
- 235000013311 vegetables Nutrition 0.000 description 1
- 238000010792 warming Methods 0.000 description 1
- 238000005406 washing Methods 0.000 description 1
- 239000001993 wax Substances 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
- 150000003751 zinc Chemical class 0.000 description 1
- FHQVHHIBKUMWTI-OTMQOFQLSA-N {1-hexadecanoyl-2-[(Z)-octadec-9-enoyl]-sn-glycero-3-phospho}ethanolamine Chemical compound CCCCCCCCCCCCCCCC(=O)OC[C@H](COP(O)(=O)OCCN)OC(=O)CCCCCCC\C=C/CCCCCCCC FHQVHHIBKUMWTI-OTMQOFQLSA-N 0.000 description 1
Landscapes
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
Abstract
The specification provides compounds of formula (I), or pharmaceutically acceptable salts, isotopic variations, tautomers or stereoisomers thereof, wherein each group is defined in the specification. Also provided are lipid nanoparticles and pharmaceutical compositions comprising the compounds, and the use of the compounds, lipid nanoparticles, and compositions in delivery of nucleic acids, the compounds being useful for delivery of a variety of bioactive substances, and having the characteristics of high encapsulation efficiency and high delivery efficiency.
Description
Technical Field
The invention belongs to the field of biological medicine, and in particular relates to an ionizable lipid compound, a preparation method and application thereof.
Background
The nucleic acid vaccine is prepared through introducing exogenous gene (DNA or RNA) encoding specific antigen protein directly into host cell, expressing the antigen protein in the host cell and inducing the host to produce immune response to the antigen protein for the purpose of preventing and treating diseases.
Moderna and BioNTech use Lipid Nanoparticles (LNP) as delivery systems, the LNP major components including Cationic Lipid molecules (Cationic Lipid), cholesterol, neutral lipids and polyethylene glycol conjugated lipids. The cationic lipid molecule is the core of the LNP delivery system, and the molecular structure plays a decisive role in the aspects of the delivery efficiency, targeting, preparation stability and the like of the whole liposome nano-particle.
Since specific delivery to achieve different kinds of nucleic acid substance delivery and different targets has different requirements for the delivery system, further development of new lipid molecules is required in order to meet different demands of gene therapy.
Disclosure of Invention
The invention aims to provide an ionizable lipid compound with a novel structure and application thereof, wherein the compound can be used for delivering various bioactive substances, has the characteristics of high encapsulation efficiency, high delivery efficiency and good biocompatibility, and provides more choices for delivering disease treatment or preventive agents.
In a first aspect of the invention, there is provided a compound of formula (I), or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof:
Wherein,
Each R 1、R2、R3 is independently selected from hydrogen or C 1-8 alkyl;
L 1 is selected from C 1-10 alkylene, C 2-10 alkenylene, or C 2-10 alkynylene;
G is phenylene;
Each R 4、R5 is independently selected from C 1-30 alkyl, C 2-30 alkenyl, or C 2-30 alkynyl.
In one embodiment, G is selected from
In one embodiment, the compound of formula (I) has the structure of formula (II):
Wherein R 1、R2、R3、R4、R5、L1 is as defined above.
In one embodiment, the total length of R 1, the total length of R 2, and the total length of R 3 are each independently 1,2, 3,4,5, 6, 7, or 8 carbon atoms.
In one embodiment, each R 1、R2、R3 is independently selected from hydrogen, methyl, ethyl, or propyl.
In one embodiment, R 1 is hydrogen; in another embodiment, R 1 is C 1 alkyl; in another embodiment, R 1 is C 2 alkyl; in another embodiment, R 1 is C 3 alkyl; in another embodiment, R 1 is C 4 alkyl; in another embodiment, R 1 is C 5 alkyl; in another embodiment, R 1 is C 6 alkyl; in another embodiment, R 1 is C 7 alkyl; in another embodiment, R 1 is C 8 alkyl.
In one embodiment, R 2 is hydrogen; in another embodiment, R 2 is C 1 alkyl; in another embodiment, R 2 is C 2 alkyl; in another embodiment, R 2 is C 3 alkyl; in another embodiment, R 2 is C 4 alkyl; in another embodiment, R 2 is C 5 alkyl; in another embodiment, R 2 is C 6 alkyl; in another embodiment, R 2 is C 7 alkyl; in another embodiment, R 2 is C 8 alkyl.
In one embodiment, R 3 is hydrogen; in another embodiment, R 3 is C 1 alkyl; in another embodiment, R 3 is C 2 alkyl; in another embodiment, R 3 is C 3 alkyl; in another embodiment, R 3 is C 4 alkyl; in another embodiment, R 3 is C 5 alkyl; in another embodiment, R 3 is C 6 alkyl; in another embodiment, R 3 is C 7 alkyl; in another embodiment, R 3 is C 8 alkyl.
In one embodiment, L 1 is selected from C 1-10 alkylene; in another embodiment, L 1 is selected from C 2-10 alkenylene; in another embodiment, L 1 is selected from C 2-10 alkynylene.
In one embodiment, the total length of L 1 is 1,2, 3, 4, 5, 6, 7, 8, 9, or 10 carbon atoms.
In one embodiment, L 1 is selected from C 1-6 alkylene; in another embodiment, L 1 is selected from C 1-4 alkylene; in another embodiment, L 1 is selected from C 1-2 alkylene.
In one embodiment, L 1 is selected from-CH 2-、-(CH2)2-、-(CH2)3 -or- (CH 2)4 -.
In one embodiment, L 1 is-CH 2 -; in another embodiment, L 1 is- (CH 2)2 -; in another embodiment, L 1 is- (CH 2)3 -, and in another embodiment, L 1 is- (CH 2)4 -.
In one embodiment, G is
In one embodiment, R 4 is selected from C 1-30 alkyl, C 2-30 alkenyl, or C 2-30 alkynyl.
In one embodiment, the total length of R 4 is 1,2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 carbon atoms.
In one embodiment, R 4 is selected from- (CH 2)m1CH＝CH(CH2)n1CH＝CH(CH2)m2CH3;
m1 and m2 are each independently selected from 2, 3,4, 5, 6, 7, 8, 9, 10 or 11;
n1 is selected from 0,1,2 or 3.
In one embodiment, R 4 is selected from- (CH 2)7CH＝CHCH2CH＝CH(CH2)4CH3; in another embodiment, R 4 is selected from- (CH 2)8CH＝CHCH2CH＝CH(CH2)4CH3), in another embodiment R 4 is selected from- (CH 2)9CH＝CHCH2CH＝CH(CH2)4CH3), in another embodiment R 4 is selected from- (CH 2)7CH＝CH(CH2)2CH＝CH(CH2)4CH3), in another embodiment R 4 is selected from- (CH 2)8CH＝CH(CH2)2CH＝CH(CH2)4CH3), and in another embodiment R 4 is selected from- (CH 2)9CH＝CH(CH2)2CH＝CH(CH2)4CH3.
In one embodiment, R 5 is selected from C 1-30 alkyl, C 2-30 alkenyl, or C 2-30 alkynyl.
In one embodiment, the total length of R 5 is 1,2, 3,4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25, 26, 27, 28, 29, or 30 carbon atoms.
In one embodiment, R 5 is selected from- (CH 2)m1CH＝CH(CH2)n1CH＝CH(CH2)m2CH3;
m1 and m2 are each independently selected from 6, 7, 8, 9, 10 or 11;
n1 is selected from 0,1,2 or 3.
In one embodiment, R 5 is selected from- (CH 2)7CH＝CHCH2CH＝CH(CH2)4CH3; in another embodiment, R 4 is selected from- (CH 2)8CH＝CHCH2CH＝CH(CH2)4CH3), in another embodiment R 5 is selected from- (CH 2)9CH＝CHCH2CH＝CH(CH2)4CH3), in another embodiment R 5 is selected from- (CH 2)7CH＝CH(CH2)2CH＝CH(CH2)4CH3), in another embodiment R 5 is selected from- (CH 2)8CH＝CH(CH2)2CH＝CH(CH2)4CH3), and in another embodiment R 5 is selected from- (CH 2)9CH＝CH(CH2)2CH＝CH(CH2)4CH3.
In a first aspect of the invention, the following compounds are provided:
in a second aspect of the invention, there is provided a process for the preparation of a compound of formula (II) above, or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, comprising:
reacting a compound of formula (IIa) with a compound of formula (IIb) to obtain a compound of formula (II);
Wherein X 1 is selected from halogen; r 1、R2、R3、R4、R5、L1 are each as defined above.
In a third aspect of the present invention, there is provided a process for the preparation of compound 1, comprising:
Reacting linolenic alcohol (compound 1-1) with carbon tetrabromide in the presence of triphenylphosphine to obtain compound 1-2;
reacting p-toluenesulfonyl methyl isonitrile (TosMIC) with compound 1-2 to obtain compound 1-3; further, the reaction is carried out under strongly basic conditions; still further, the reaction is carried out in the presence of sodium hydride;
Hydrolyzing the compound 1-3 to obtain a compound 1-4; further, hydrolysis is performed under acidic conditions; further, hydrolysis is performed in the presence of hydrochloric acid;
Reducing the compound 1-4 to obtain a compound 1-5; further, reducing the compound 1-4 by sodium borohydride to obtain a compound 1-5;
Reacting the compound 1-5 with 4-fluorobenzoic acid to obtain a compound 1-6; further, the compound 1-5, 4-fluorobenzoic acid is reacted in the presence of EDCI and DMAP (4-dimethylaminopyridine) to obtain the compound 1-6;
Reacting the compound 1-6 with N, N-dimethyl ethylenediamine to obtain a compound 1; further, reacting the compounds 1-6 with N, N-dimethylethylenediamine in the presence of DIEA to obtain a compound shown in a formula 1;
in one embodiment, the method for preparing compound 1 comprises:
s1: reacting linolenic alcohol with carbon tetrabromide in the presence of triphenylphosphine to obtain a compound 1-2;
s2: reacting p-toluenesulfonyl methyl isonitrile with a compound 1-2 under a strong alkaline condition to obtain a compound 1-3;
s3: hydrolyzing the compound 1-3 under an acidic condition to obtain a compound 1-4;
S4: reducing the compound 1-4 by NaBH 4 to obtain a compound 1-5;
S5: reacting the compounds 1-5 and 4-fluorobenzoic acid in the presence of EDCI and DMAP to obtain compounds 1-6;
S6: reacting the compounds 1-6 with N, N-dimethylethylenediamine in the presence of DIEA to obtain the compound 1.
In a fourth aspect of the invention, there is provided a pharmaceutical composition comprising a compound as described in the preceding first aspect, or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, and a pharmaceutically acceptable adjuvant.
In a fifth aspect of the invention, there is provided a lipid nanoparticle comprising a lipid component and optionally an active ingredient;
the lipid component comprises: ionizable cationic lipids, structural lipids, neutral lipids, and polymeric lipids;
Wherein the ionizable cationic lipid is a compound of the first aspect described above, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof.
In one embodiment, the neutral lipid is selected from one or more of DSPC, DMPC, DOPC, DPPC, POPC, DOPE, DMPE, POPE or DPPE.
In one embodiment, the structural lipid is selected from one or more of cholesterol, sitosterol, stigmasterol, rock-soap sterol, brassicasterol, ergosterol, lycorine, ursolic acid, alpha-tocopherol, stigmasterol, aveosterol, ergocalcitol, or campesterol.
In one embodiment, the polymeric lipid is a pegylated lipid.
In one embodiment, the pegylated lipid is selected from one or more of PEG-modified phosphatidylethanolamine, PEG-modified phosphatidic acid, PEG-modified ceramide, PEG-modified dialkylamine, PEG-modified diacylglycerol, PEG-modified dialkylglycerol.
In one embodiment, the pegylated lipid is selected from one or more of DMPE-PEG1000、DPPE-PEG1000、DSPE-PEG1000、DOPE-PEG1000、DMG-PEG2000、Ceramide-PEG2000、DMPE-PEG2000、DPPE-PEG2000、DSPE-PEG2000、Azido-PEG2000、DSPE-PEG2000-Mannose、Ceramide-PEG5000、DSPE-PEG5000.
In a sixth aspect of the present invention, there is provided a method for preparing lipid nanoparticles, comprising:
(1) Sequentially dissolving and mixing an ionizable lipid compound, cholesterol, phospholipid and polyethylene glycol lipid (in mol percent) in ethanol according to 50%, 38.5%, 10% and 1.5%, respectively;
(2) The mRNA active ingredient was dissolved in 25mM sodium acetate solution (ph=4.5);
(3) Mixing the organic phase dissolved with the lipid mixture and the aqueous phase dissolved with the mRNA component by using an automatic high-throughput microfluidic system according to a flow rate ratio ranging from 1:1 to 1:4, wherein the mixing speed is 10-18mL/min;
(4) The lipid nanoparticles (N/P6) were diluted with phosphate buffer solution and then the nanoparticle solution was ultrafiltered to the original preparation volume using an ultrafiltration tube (available from Millipore) with a molecular weight cut-off of 30 kDa;
(5) The obtained nano particles are filtered and sterilized by a sterile filter membrane with the diameter of 0.2 mu m and then stored in a sealed glass bottle at low temperature.
In a seventh aspect of the present invention, there is provided the use of a compound according to the first aspect, or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, or a pharmaceutical composition according to the fourth aspect, or a lipid nanoparticle according to the fifth aspect, for the manufacture of a medicament for the treatment, diagnosis or prophylaxis of a disease.
In an eighth aspect of the invention there is provided the use of a compound according to the first aspect of the invention, or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, or a pharmaceutical composition according to the fourth aspect, or a lipid nanoparticle according to the fifth aspect, for the manufacture of a medicament for the delivery of a load selected from one or more of a therapeutic agent, a prophylactic agent or a diagnostic agent.
In a ninth aspect of the invention there is provided a method of treating, diagnosing or preventing a disease in a subject comprising administering to the subject an effective amount of a pharmaceutical composition according to the fourth aspect of the foregoing, or a lipid nanoparticle according to the fifth aspect.
In one embodiment, the therapeutic, prophylactic or diagnostic agent is selected from one or more of a nucleic acid, a small molecule compound, a polypeptide or a protein.
In one embodiment, the nucleic acid is selected from one or more of RNA or DNA.
In one embodiment, the RNA is selected from one or more of small interfering RNA (siRNA), short hairpin RNA (shRNA), antisense RNA (aRNA), messenger RNA (mRNA), modified messenger RNA (mmRNA), long non-coding RNA (lncRNA), microrna (miRNA), small activating RNA (saRNA), poly-coding nucleic acid (MCNA), poly-coding nucleic acid (PCNA), guide RNA (gRNA), CRISPRRNA (CRRNA), or ribozyme.
In one embodiment, the RNA is mRNA.
In one embodiment, the RNA is a modified mRNA.
Detailed Description
I. Definition of the definition
In the present invention, unless otherwise indicated, scientific and technical terms used herein have the meanings commonly understood by one of ordinary skill in the art. Also, the relative terms and laboratory procedures used herein are terms and conventional procedures that are widely used in the corresponding arts. Meanwhile, in order to better understand the present invention, definitions and explanations of related terms are provided below.
As used herein and unless otherwise indicated, the terms "comprising," "including," "having," "containing," and their grammatical equivalents are generally understood to be open-ended and not to be limiting, e.g., not to exclude other, unrecited elements or steps.
Compounds are named herein using standard nomenclature. Compounds having asymmetric centers, it is to be understood (unless otherwise indicated) that all optical isomers and mixtures thereof are encompassed. Furthermore, unless otherwise specified, all isomeric compounds encompassed by the present invention may occur with carbon-carbon double bonds in the form of Z and E. Compounds that exist in different tautomeric forms, one of the compounds is not limited to any particular tautomer, but is intended to encompass all tautomeric forms.
The compounds of the invention may include one or more asymmetric centers and thus may exist in a variety of stereoisomeric forms, for example, enantiomeric and/or diastereomeric forms. For example, the compounds of the invention may be individual enantiomers, diastereomers, or geometric isomers (e.g., cis and trans isomers), or may be in the form of mixtures of stereoisomers, including racemic mixtures and mixtures enriched in one or more stereoisomers. The isomers may be separated from the mixtures by methods known to those skilled in the art, including: chiral High Pressure Liquid Chromatography (HPLC), formation and crystallization of chiral salts; alternatively, preferred isomers may be prepared by asymmetric synthesis.
The compounds of the present invention may exist in tautomeric forms. Tautomers are functional group isomers that result from the rapid movement of an atom in a molecule at two positions. Tautomers are a particular functional group isomer, and a pair of tautomers can be converted to each other, but usually take a relatively stable one as its predominant form. The most prominent examples are enol and keto tautomers.
The invention also includes isotopically-labeled compounds (isotopically-variant) which are identical to those recited in formula (I), formula (IA) or formula (IB), but for the fact that one or more atoms are replaced by an atom having an atomic mass or mass number different from the atomic mass or mass number usually found in nature. Examples of isotopes that can be incorporated into compounds of the invention include isotopes of hydrogen, carbon, nitrogen, oxygen, phosphorus, sulfur, fluorine, and chlorine, such as 2H、3H、13C、11C、14C、15N、18O、17O、31P、32P、35S、18F and 36 Cl, respectively. The compounds of the invention, prodrugs thereof, and pharmaceutically acceptable salts of the compounds or prodrugs thereof, which contain the isotopes described above and/or other isotopes of other atoms, are within the scope of this invention. Certain isotopically-labeled compounds of the present invention, for example those into which radioactive isotopes (e.g., 3 H and 14 C) are introduced, are useful in drug and/or substrate tissue distribution assays. Tritium, i.e., 3 H, and carbon-14, i.e., 14 C isotopes are particularly preferred because they are easy to prepare and detect. Further, substitution with heavier isotopes, such as deuterium, i.e., 2 H, may be preferred in some circumstances because greater metabolic stability may afford therapeutic benefits such as increased in vivo half-life or reduced dosage requirements. Isotopically-labelled compounds of formula (I), formula (IA) or formula (IB) of the present invention and prodrugs thereof are typically obtained by substituting a readily available isotopically-labelled reagent for a non-isotopically-labelled reagent during the course of the preparation of the compound.
The term "substituted" or "substituted" means that any one or more hydrogen atoms on a particular atom is replaced with a substituent.
When numerical ranges are listed, it is intended to include each and every value and subrange within the range.
The term "alkyl" refers to a straight or branched saturated hydrocarbon moiety. In one embodiment, the alkyl group is a straight chain saturated hydrocarbon. "C 1-30 alkyl" refers to a straight or branched saturated hydrocarbon group having 1 to 30 carbon atoms. The term "C 1-30 alkyl" also includes heteroalkyl groups in which one or more (e.g., 1,2,3, or 4) carbon atoms are replaced with a heteroatom (e.g., oxygen, sulfur, nitrogen, boron, silicon, phosphorus). The alkyl group may be optionally substituted with one or more substituents, for example, with 1 to 5 substituents, 1 to 3 substituents, or 1 substituent.
The term "alkenyl" refers to a straight or branched hydrocarbon moiety having one or more carbon-carbon double bonds. "C 2-30 alkenyl" refers to a straight or branched hydrocarbon group having 2 to 30 carbon atoms and at least one carbon-carbon double bond. In some embodiments, C 10-20 alkenyl, C 15-20 alkenyl, C 17-20 alkenyl, C 18 alkenyl, C 19 alkenyl, C 20 alkenyl are preferred. The term "C 2-30 alkenyl" also includes heteroalkenyl groups in which one or more (e.g., 1,2, 3, or 4) carbon atoms are replaced with heteroatoms (e.g., oxygen, sulfur, nitrogen, boron, silicon, phosphorus). The alkenyl group may be optionally substituted with one or more substituents, for example, with 1 to 5 substituents, 1 to 3 substituents, or 1 substituent.
The term "alkynyl" refers to a straight or branched hydrocarbon moiety having one or more carbon-carbon triple bonds. "C 2-30 alkynyl" refers to a straight or branched hydrocarbon group having 2 to 30 carbon atoms, at least one carbon-carbon triple bond, and optionally one or more carbon-carbon double bonds. In some embodiments, C 10-20 alkenyl, alkenyl C 15-20 alkynyl, C 17-20 alkynyl, C 18 alkynyl, C 19 alkynyl, C 20 alkynyl are preferred. The term "C 2-30 alkynyl" also includes heteroalkynyl groups in which one or more (e.g., 1,2,3, or 4) carbon atoms are replaced with heteroatoms (e.g., oxygen, sulfur, nitrogen, boron, silicon, phosphorus). Alkynyl groups may be optionally substituted with one or more substituents, for example, with 1 to 5 substituents, 1 to 3 substituents, or 1 substituent.
The term "alkylene" refers to a divalent group formed by removing another hydrogen of an alkyl group, which may be substituted or unsubstituted. "C 1-10 alkylene" refers to a divalent group formed by removing another hydrogen of a C 1-10 alkyl group, and may be substituted or unsubstituted. In some embodiments, methylene (-CH 2 -), ethylene (-CH 2CH2 -), propylene (-CH 2CH2CH2 -), butylene (-CH 2CH2CH2CH2 -) are preferred.
The term "alkenylene" refers to a divalent group formed by removal of another hydrogen of an alkenyl group, which may be substituted or unsubstituted. "C 2-10 alkenylene" refers to a divalent group formed by removal of another hydrogen of a C 2-10 alkenyl group, and may be substituted or unsubstituted.
The term "alkynylene" refers to a divalent group formed by the removal of another hydrogen of an alkynyl group, which may be substituted or unsubstituted. "C 2-10 alkynylene" refers to a divalent group formed by removal of another hydrogen of a C 2-10 alkynyl group, and may be substituted or unsubstituted.
The term "the total length of the variables a and B is x carbon atoms" means that the number of carbon atoms in the backbone of the group represented by the variable a and the number of carbon atoms in the backbone of the group represented by the variable B are x.
"Optionally substituted" means that the substituent may be designated, or may be unsubstituted.
"Nucleic acid" refers to a single-or double-stranded deoxyribonucleic acid (DNA) or ribonucleic acid (RNA) molecule, and hybrid molecules thereof. Examples of nucleic acid molecules include, but are not limited to, messenger RNAs (mrnas), micrornas (mirnas), small interfering RNAs (sirnas), self-amplifying RNAs (sarnas), antisense oligonucleotides (ASOs), and the like. The nucleic acid may be further chemically modified, the chemical modification being selected from one of pseudouridine, N1-methyl-pseudouridine, 5-methoxyuridine, 5-methylcytosine, or a combination thereof. The mRNA molecules contain protein coding regions and may further contain expression control sequences, typical expression control sequences include, but are not limited to, 5 'caps (5' caps), 5 'untranslated regions (5' utrs), 3 'untranslated regions (3' utrs), polyadenylation sequences (polyas), miRNA binding sites.
"Cationic lipid" refers to a lipid molecule capable of being positively charged under physiological pH conditions.
"Neutral lipids" refers to lipid molecules that are uncharged at specific pH conditions, such as physiological pH conditions. Examples of neutral lipids include, but are not limited to, 1, 2-distearoyl-sn-glycero-3-phosphorylcholine (DSPC), 1, 2-dimyristoyl-sn-glycero-3-phosphorylcholine (DMPC), 1, 2-dioleoyl-sn-glycero-3-phosphorylcholine (DOPC), 1, 2-dipalmitoyl-sn-glycero-3-phosphorylcholine (DPPC), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphorylcholine (POPC), 1, 2-dioleoyl-sn-glycero-3-phosphorylethanolamine (DOPE), 1, 2-dimyristoyl-sn-glycero-3-phosphorylethanolamine (DMPE), 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphorylethanolamine (DPPE), 1, 2-dipalmitoyl-sn-glycero-3-phosphorylethanolamine (DPPE).
"Structural lipids" refers to lipids, such as steroids, that are commonly used to enhance nanoparticle stability by filling in the interstices between the lipids. The steroid is a compound having a cyclopenta-polyhydrophenanthrene carbon skeleton, and in a preferred embodiment, the steroid is selected from cholesterol, sitosterol, stigmasterol, soapsterol, brassicasterol, ergosterol, lycorine, ursolic acid, alpha-tocopherol, stigmasterol, oat sterol, ergocalcitol, or campesterol.
"Polymer lipid" refers to a molecule that contains a polymer moiety and a lipid moiety. In some embodiments, the polymer lipid is a polyethylene glycol (PEG) lipid. Other lipids capable of reducing aggregation, such as products of coupling compounds having uncharged, hydrophilic, steric-blocking moieties to lipids, may also be used.
"Lipid nanoparticle" refers to particles having a nanoscale size that contain a lipid component.
The term "pharmaceutically acceptable" is intended to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.
The term "pharmaceutically acceptable salt" refers to pharmaceutically acceptable acid addition salts or base addition salts, including salts of compounds with inorganic or organic acids, and salts of compounds with inorganic or organic bases.
The term "pharmaceutically acceptable excipients" refers to those excipients which do not significantly stimulate the organism and which do not impair the biological activity and properties of the active compound. Suitable adjuvants are well known to those skilled in the art. Pharmaceutically acceptable excipients for use in the present invention refer to non-toxic carriers, adjuvants or vehicles that do not destroy the pharmacological activity of the co-formulated compounds. Pharmaceutically acceptable carriers, adjuvants or vehicles that may be used in the compositions of the invention include, but are not limited to, ion exchangers, alumina, aluminum stearate, lecithin, serum proteins (e.g., human serum albumin), buffer substances (e.g., phosphates), glycine, sorbic acid, potassium sorbate, partial glyceride mixtures of saturated vegetable fatty acids, water, salts or electrolytes (e.g., protamine sulfate), disodium hydrogen phosphate, potassium hydrogen phosphate, sodium chloride, zinc salts, silica gel, magnesium trisilicate, polyvinylpyrrolidone, cellulose-based substances, polyethylene glycol, sodium carboxymethyl cellulose, polyacrylates, waxes, polyethylene-polyoxypropylene-block polymers, polyethylene glycol, and lanolin.
Typically, physiological saline may be used as a pharmaceutically acceptable carrier. Other suitable carriers include, for example, water, buffered water, 0.9% saline, 0.3% glycine, and the like, including glycoproteins for improved stability, e.g., albumin, lipoproteins, globulins, and the like. In compositions comprising saline or other carrier-containing salts, the carrier is preferably added after the lipid particles are formed. Thus, after formation of the lipid-nucleic acid composition, the composition may be diluted in a pharmaceutically acceptable carrier such as physiological saline.
The term "treating" relates to reversing, alleviating, inhibiting the progression or prevention of a disorder or condition to which the term applies, or one or more symptoms of such disorder or condition. The term "treatment" as used herein relates to the action of a verb treatment, the latter as just defined.
The term "subject" includes, but is not limited to: a human (i.e., male or female of any age group, e.g., pediatric subjects (e.g., infants, children, adolescents) or adult subjects (e.g., young adults, middle aged adults, or senior adults)) and/or a non-human animal, e.g., a mammal, e.g., a primate (e.g., cynomolgus monkey, rhesus monkey), cow, pig, horse, sheep, goat, rodent, cat, and/or dog. In some embodiments, the subject is a human. In some embodiments, the subject is a non-human animal. The terms "human", "patient" and "subject" are used interchangeably herein.
The term "effective amount" refers to an amount sufficient to elicit a biological response of interest. As will be appreciated by those of ordinary skill in the art, the effective amount of the pharmaceutical composition, lipid nanoparticle of the present invention may vary depending on the following factors: for example, biological targets, pharmaceutical compositions, pharmacokinetics of lipid nanoparticles, the disease being treated, the mode of administration, and the age health and symptoms of the subject. The effective amount includes a therapeutically effective amount and a prophylactically effective amount.
As used herein, unless otherwise indicated, a "therapeutically effective amount" is an amount sufficient to provide a therapeutic benefit in the treatment of a disease, disorder or condition, or to delay or minimize one or more symptoms associated with a disease, disorder or condition. A therapeutically effective amount of a pharmaceutical composition, lipid nanoparticle refers to an amount of a therapeutic agent, alone or in combination with other therapies, that provides a therapeutic benefit in the treatment of a disease, disorder or condition. The term "therapeutically effective amount" may include an amount that improves overall treatment, reduces or avoids symptoms or causes of a disease or disorder, or enhances the therapeutic effect of other therapeutic agents.
As used herein, a "prophylactically effective amount" of a pharmaceutical composition, lipid nanoparticle, unless otherwise indicated, is an amount sufficient to prevent a disease, disorder, or condition, or to prevent one or more symptoms associated with a disease, disorder, or condition, or to prevent recurrence of a disease, disorder, or condition. A prophylactically effective amount of a pharmaceutical composition, lipid nanoparticle refers to an amount of therapeutic agent, alone or in combination with other agents, that provides a prophylactic benefit in the prevention of a disease, disorder or condition. The term "prophylactically effective amount" may include an amount that improves overall prophylaxis, or an amount that enhances the prophylactic effect of other prophylactic agents.
Examples II
The present invention will be further described in detail with reference to the following examples in order to make the technical solution of the present invention clearer and more specific. The following examples are presented only to illustrate specific embodiments of the invention so that those skilled in the art can understand the invention and are not intended to limit the scope of the invention. In the specific embodiment of the present invention, technical means, methods, and the like not specifically described are conventional technical means, methods, and the like in the art. Materials, reagents and the like used in the examples are commercially available unless otherwise specified.
TABLE 1
| English or abbreviations | Chinese character |
| THF | Tetrahydrofuran (THF) |
| DCM | Dichloromethane (dichloromethane) |
| MeOH | Methanol |
| NMP | N-methylpyrrolidone |
| EtOAc | Acetic acid ethyl ester |
| CDCl3 | Deuterated chloroform |
| TosMIC | P-toluenesulfonylmethyl isonitrile |
| DMAP | 4-Dimethylaminopyridine |
| EDCI | 1- (3-Dimethylaminopropyl) -3-ethylcarbodiimide |
| NaBH4 | Sodium borohydride |
| NaH | Sodium hydride |
| Na2SO4 | Sodium sulfate |
EXAMPLE 1 Synthesis of Compound 1
In a 250mL round bottom flask, linolenic alcohol 1-1 (20 g,75.0mmol,1.0 eq.) carbon tetrabromide (37.3 g,112.5mmol,1.5 eq.) and triphenylphosphine (29.5 g,112.5mmol,1.5 eq.) were dissolved in dichloromethane (200 mL) at room temperature. The reaction system was stirred at room temperature overnight for reaction. The reaction was monitored by LCMS, after completion of the reaction, the reaction was quenched by addition of water (250 mL), extracted 3 times with dichloromethane (100 mL), the organic phases were combined, washed with brine (200 mL), dried over anhydrous Na 2SO4, filtered, and concentrated. The crude product was purified by a silica gel column to give 14g of oily compound 1-2.
TosMIC (3.85 g,19.7mmol,0.5 eq.) was dissolved in anhydrous DMF (30 mL) in ice bath, then sodium hydride (60%, 2.36g,59.1mmol,1.5 eq.) was added in portions to the reaction system, allowed to react for 0.5 hours after warming to room temperature, then Compound 1-2 (13.0 g,39.4mmol,1.0 eq.) was added, and the reaction was continued at room temperature for 12 hours after completion of the addition. After the reaction was completed, the reaction system was quenched by adding water (200 mL), extracted 3 times with ethyl acetate (70 mL), the organic phases were combined, washed 3 times with brine (200 mL), dried over anhydrous Na 2SO4, filtered, and concentrated. The crude product was purified by a silica gel column to give 6.9g of the oily compound 1-3.
Compounds 1-3 (4.9 g,7.1mmol,1.0 eq.) were dissolved in tetrahydrofuran (30 mL) in a 100mL round bottom flask at room temperature, hydrochloric acid (6M, 5.0 mL) was added to the reaction system, and the reaction system was stirred at room temperature for 2 hours. The reaction was adjusted to pH neutral with saturated aqueous sodium bicarbonate, extracted 3 times with ethyl acetate (30 mL), the organic phases were combined, washed with brine (100 mL), dried over anhydrous Na 2SO4, filtered, and concentrated. The crude product was purified by a silica gel column to give 3.0g of compounds 1-4.
Compounds 1-4 (1.5 g,2.8mmol,1.0 eq.) were dissolved in 10mL of methanol in a 25mL round bottom flask, sodium borohydride (158.9 mg,4.2mmol,1.5 eq.) was added and reacted for 3 hours at room temperature. After completion of the reaction, 15mL of water was added, quenched, extracted 3 times with ethyl acetate (20 mL), the organic phases were combined, washed with brine (50 mL), dried over anhydrous Na 2SO4, filtered, and concentrated to give 1.2g of the oily compound 1-5, which was used directly in the next reaction without purification.
In a 50mL round bottom flask, 4-fluorobenzoic acid (300 mg,2.1mmol,1.0 eq.) compounds 1-5 (1.1 g,2.1mmol,1.0 eq.), EDCI (1.2 g,6.3mmol,3.0 eq.), DMAP (256.6 mg,2.1mmol,1.0 eq.) are dissolved in 20mL dichloromethane. The reaction was carried out at room temperature overnight. LCMS monitors the reaction, after the reaction is completed, 20mL of water is added for quenching, dichloromethane (20 mL) is used for extraction for 3 times, the organic phases are combined, brine (50 mL) is used for washing, anhydrous Na 2SO4 is used for drying, filtering and concentrating are carried out, and the crude product is purified by a silica gel column to obtain 410mg of oily compound 1-6.
Compounds 1-6 (300 mg,0.46mmol,1.0 eq.) and N, N-dimethylethylenediamine (405.5 mg,4.6mmol,10.0 eq.) were dissolved in DIEA (5 mL) and NMP (5 mL) in a 20mL sealed tube and the reaction was warmed to 150℃and stirred for 6 hours. The reaction was cooled to room temperature, quenched by pouring into 30mL of water, extracted 3 times with ethyl acetate (30 mL), the organic phases were combined, washed with brine (50 mL), dried over anhydrous Na 2SO4, filtered, concentrated, and the crude product purified by prep-TLC to give 80.02mg of compound 1 as an oil.
1H NMR(400MHz,CDCl3):δppm 0.89(t,J＝6.8Hz,6H),1.19-1.36(m,36H),1.55-1.64(m,4H),2.03-2.08(m,8H),2.74-2.79(m,4H),2.87(s,6H),3.29(t,J＝5.2Hz,2H),3.62(t,J＝5.2Hz,2H),5.08(p,J＝6.4Hz,1H),5.32-5.42(m,8H),6.60(d,J＝8.4Hz,2H),7.90(d,J＝8.4Hz,2H),13.20(s,1H);ESI-MS m/z:719.7[M+H]+.
Pharmacological experiments
Experimental example 1 nanoparticle preparation
Materials used for lipid nanoparticle assembly are: (1) an ionizable lipid compound: the ionizable lipids synthesized as designed in the present invention or DLin-MC3-DMA (MC 3, purchased from AVT) as a control group; (2) structural lipids: such as Cholesterol (purchased from Sigma-Aldrich); (3) Phospholipids such as 1, 2-distearoyl-SN-glycero-3-phosphorylcholine (DSPC, available from AVT); (4) Polyethylene glycol lipid compounds such as dimyristoylglycerol-polyethylene glycol 2000 (DMG-PEG 2000 available from AVT); (5) The effective components of the nucleic acid fragment include Luciferase (Luciferase mRNA), siRNA, CRISPR Cas mRNA and the like. The component names and the structural formulas of the lipid nanoparticles are shown in Table 2.
TABLE 2
The preparation method of the lipid nanoparticle comprises the following steps: (1) Respectively dissolving and mixing 50%,38.5%,10% and 1.5% of ionizable lipid compound, cholesterol, phospholipid and polyethylene glycol lipid in ethanol; (2) The mRNA active ingredient was dissolved in 25mM sodium acetate solution (ph=4.5); (3) Mixing the organic phase dissolved with the lipid mixture and the aqueous phase dissolved with the mRNA component by using an automatic high-throughput microfluidic system according to a flow rate ratio ranging from 1:1 to 1:4, wherein the mixing speed is 10-18mL/min; (4) The lipid nanoparticles (N/P6) were diluted with phosphate buffer solution and then the nanoparticle solution was ultrafiltered to the original preparation volume using an ultrafiltration tube (available from Millipore) with a molecular weight cut-off of 30 kDa; (5) The obtained nano particles are filtered and sterilized by a sterile filter membrane with the diameter of 0.2 mu m and then stored in a sealed glass bottle at low temperature.
The preparation method of the lipid nanoparticle comprises a microfluidic mixing system, but is not limited to the method, and also comprises a T-type mixer, an ethanol injection method and the like.
Experimental example 2 characterization of physical Properties of lipid nanoparticles
Particle size and particle size dispersion coefficient (PDI) of the prepared lipid nanoparticles were measured using a Zetasizer Pro (available from Malvern Instruments Ltd) and DynaPro NanoStar (available from Wyatt) dynamic light scattering instrument. The extent of entrapment of the lipid nanoparticle to the RNA was characterized by the encapsulation efficiency (Encapsulation Efficiency%), a coefficient reflecting the extent of binding of the lipid nanoparticle to the RNA fragment. The coefficients were measured by the method of Quant-it TM RiboGreen RNA Assay (available from Invitrogen). Lipid nanoparticle samples were diluted in TE buffer (10 mM Tris-HCl,1mM EDTA,pH =7.5), and a portion of the sample solution was taken out and added to 0.5% Triton (Triton X-100) and allowed to stand at 37℃for 30 minutes. Adding inImmediately after the reaction, the fluorescence value was read by a Varioskan LUX multifunctional microplate reader (commercially available from Thermofisher) under the conditions of an absorption light band of 485nm and an emission light band of 528nm to obtain the entrapping rate value.
Experimental example 3 animal experiment
Assessment of the delivery efficacy and safety of nanoparticles loaded with luciferase mRNA (ex Trilink, L-7202) in mice. The test mice were SPF grade C57BL/6 mice, females, 6-8 weeks old, weighing 18-22g, purchased from Beijing Bei Fu Biotechnology Co. All animals are subjected to adaptive feeding for more than 7 days before the test, drinking water is fed freely during the test, the illumination is changed for 12/12h, the indoor temperature is 20-26 ℃, and the humidity is 40-70%. Mice were randomly grouped.
The lipid nanoparticle prepared above and having the luciferase mRNA contained therein was injected into mice intravenously at a single dose of 0.5mg/kg of mRNA, and the mice were subjected to in vivo bioluminescence detection using a small animal in vivo imaging system (IVIS LUMINA III, available from Perkinelmer) at 6 hours after administration. The specific operation steps of the detection are as follows: a15 mg/mL concentration of D-fluorescein solution was prepared in physiological saline, and each mouse was given the substrate by intraperitoneal injection. 10 minutes after substrate administration, mice were anesthetized with isoflurane at a concentration of 2.5% in an anesthetic box. The anesthetized mice are placed in IVIS for fluorescence imaging and data acquisition and data analysis are carried out on the parts with concentrated distribution of fluorescence.
The in vivo delivery efficiency of the lipid nanoparticle carrier is expressed as the average of the fluorescence intensity and total photon number of different animals within the same test group, as shown in table 3. The higher the values of fluorescence intensity and total photon number, the higher the in vivo delivery efficiency of the lipid nanoparticle for the mRNA fragment. Lipid nanoparticles comprising the ionizable lipids of the invention have good in vivo delivery efficiency.
TABLE 3 Table 3
Example 4 in vitro delivery efficiency and safety assessment
The effect of delivering nanoparticles loaded with luciferase mRNA (purchased from Trilink, L-7202) at the in vitro cellular level was evaluated as well as safety. The cells used in the test were human embryonic kidney cells 293 (HEK 293T cells) and cultured in DMEM Du modified Eagle medium (purchased from Thermofisher) containing 10% fetal bovine serum and 5% penicillin-streptomycin double antibody at room temperature of 37℃and carbon dioxide concentration of 5%. The cells were uniformly scattered and plated in a 48-well plate, and after 24 hours of stationary culture in an incubator, a lipid nanoparticle solution encapsulating fluorescein mRNA was added. After 24 hours incubation, cells were disrupted and the intensity and Relative Light Units (RLU) of each lipid nanoparticle-mediated luciferase expression was measured using luciferase assay reagents (available from Promega), with higher intensities of expression representing higher delivery efficiencies of the lipid material at the cellular level. Meanwhile, cytotoxicity was tested 24 hours later on a parallel lipid nanoparticle treated cell group using CCK-8 reagent (available from DOJINDO). The test was performed with the negative control of the cell group to which PBS alone was added. The method comprises the following specific steps: after CCK-8 solution is added into cells, the cells are kept stand in a cell incubator at 37 ℃ for 4 hours, and the light absorption value is read by a multifunctional enzyme-labeled instrument under the condition that the light absorption wave band is 450 nm. The ratio of absorbance read by the nanoparticle treated cells to that of the negative control was used as a characterization parameter for cell viability. The effect of nanoparticle delivery at the in vitro cellular level and toxicity data are shown in table 4. Lipid nanoparticles comprising the ionizable lipid compounds of the invention have good in vitro delivery efficiency and lower toxicity.
TABLE 4 Table 4
The foregoing descriptions of specific exemplary embodiments of the present invention are presented for purposes of illustration and description. It is not intended to limit the invention to the precise form disclosed, and obviously many modifications and variations are possible in light of the above teaching. The exemplary embodiments were chosen and described in order to explain the specific principles of the invention and its practical application to thereby enable one skilled in the art to make and utilize the invention in various exemplary embodiments and with various modifications as are suited to the particular use contemplated. It is intended that the scope of the invention be defined by the claims and their equivalents.
Claims (10)
1. A compound of formula (I), or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof:
Wherein,
Each R 1、R2、R3 is independently selected from hydrogen or C 1-8 alkyl;
L 1 is selected from C 1-10 alkylene, C 2-10 alkenylene, or C 2-10 alkynylene;
G is phenylene;
Each R 4、R5 is independently selected from C 1-30 alkyl, C 2-30 alkenyl, or C 2-30 alkynyl.
2. The compound of claim 1, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof, having a structure represented by formula (II):
wherein R 1、R2、R3、R4、R5、L1 is each as defined in claim 1.
3. The compound of claim 1 or 2, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof, wherein each R 1、R2、R3 is independently selected from hydrogen, methyl, ethyl, or propyl; or, L 1 is selected from-CH 2-、-(CH2)2-、-(CH2)3 -or- (CH 2)4 -.
4. The compound of claim 1 or 2, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof, wherein each R 4、R5 is independently selected from- (CH 2)m1CH＝CH(CH2)n1CH＝CH(CH2)m2CH3;
m1 and m2 are each independently selected from 2, 3,4, 5, 6, 7, 8, 9, 10 or 11;
n1 is selected from 0,1,2 or 3.
5. The compound of claim 1, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof, wherein the compound has the structure:
6. A process for the preparation of a compound of formula (II), or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, according to claim 2, which comprises:
reacting a compound of formula (IIa) with a compound of formula (IIb) to obtain a compound of formula (II);
Wherein X 1 is selected from halogen; r 1、R2、R3、R4、R5、L1 is each as defined in claim 2.
7. A pharmaceutical composition comprising a compound of any one of claims 1-5, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof, and a pharmaceutically acceptable adjuvant.
8. A lipid nanoparticle comprising a lipid component and optionally an active ingredient;
the lipid component comprises: ionizable cationic lipids, structural lipids, neutral lipids, and polymeric lipids;
Wherein the ionizable cationic lipid is a compound of any one of claims 1-5, or a pharmaceutically acceptable salt, isotopic variant, tautomer, or stereoisomer thereof.
9. Use of a compound according to any one of claims 1-5, or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, or a pharmaceutical composition according to claim 7, or a lipid nanoparticle according to claim 8, for the manufacture of a medicament for the treatment, diagnosis or prevention of a disease.
10. Use of a compound according to any one of claims 1-5, or a pharmaceutically acceptable salt, isotopic variant, tautomer or stereoisomer thereof, or a pharmaceutical composition according to claim 7, or a lipid nanoparticle according to claim 8, for the preparation of a medicament for delivering a load selected from one or more of a therapeutic, prophylactic or diagnostic agent.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410324191.0A CN118290282A (en) | 2024-03-20 | 2024-03-20 | Ionizable lipid compound, preparation method and application thereof |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN202410324191.0A CN118290282A (en) | 2024-03-20 | 2024-03-20 | Ionizable lipid compound, preparation method and application thereof |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN118290282A true CN118290282A (en) | 2024-07-05 |
Family
ID=91685384
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN202410324191.0A Pending CN118290282A (en) | 2024-03-20 | 2024-03-20 | Ionizable lipid compound, preparation method and application thereof |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN118290282A (en) |
-
2024
- 2024-03-20 CN CN202410324191.0A patent/CN118290282A/en active Pending
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| AU2010249881B2 (en) | Compositions comprising cationic amphiphiles and colipids for delivering therapeutics molecules | |
| CN115887674B (en) | lipid nanoparticles | |
| TWI828461B (en) | Ionizable lipid compounds | |
| JP2023505316A (en) | nanomaterials | |
| AU2013380825B2 (en) | Lipidosome preparation, preparation method and application thereof | |
| CN114989027A (en) | Cationic lipid compounds and compositions for delivery of nucleic acids and uses | |
| CN115850104B (en) | Ionizable lipid compounds | |
| CN118290282A (en) | Ionizable lipid compound, preparation method and application thereof | |
| CN118290339A (en) | Ionizable lipid compounds and uses thereof | |
| CN117924106A (en) | Ionizable lipid compounds | |
| WO2024149229A1 (en) | Lipid nanoparticle compositions and uses thereof | |
| CN116854606B (en) | Cationic lipid compound, composition and application thereof | |
| CN116082179B (en) | Ionizable lipid based on endogenous dicarboxylic acid, preparation method and application thereof | |
| EP4385523A1 (en) | Lipid-based topical injection formulations | |
| EP4332087A1 (en) | Lipid nanoparticles | |
| US20240245782A1 (en) | Ionizable lipid compounds | |
| CN116211831A (en) | Lipid matrix local injection preparation | |
| TWI847412B (en) | Lipid nanoparticles | |
| CN118105357A (en) | Intrathecal delivery preparation and application thereof | |
| TW202423452A (en) | Lipid-based topical injection formulations | |
| CN117534585A (en) | Novel ionizable cationic lipid compound, and preparation method and application thereof |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| PB01 | Publication | ||
| PB01 | Publication | ||
| SE01 | Entry into force of request for substantive examination |