CN110256321B - Preparation method and application of (2R,3S,4S)-4-amino-2-tetradecylpyrrolidin-3-ol - Google Patents
Preparation method and application of (2R,3S,4S)-4-amino-2-tetradecylpyrrolidin-3-ol Download PDFInfo
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- CN110256321B CN110256321B CN201910504257.3A CN201910504257A CN110256321B CN 110256321 B CN110256321 B CN 110256321B CN 201910504257 A CN201910504257 A CN 201910504257A CN 110256321 B CN110256321 B CN 110256321B
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- IBZWJKPWFKTHKK-KSZLIROESA-N (2R,3S,4S)-4-amino-2-tetradecylpyrrolidin-3-ol Chemical compound N[C@@H]1[C@@H]([C@H](NC1)CCCCCCCCCCCCCC)O IBZWJKPWFKTHKK-KSZLIROESA-N 0.000 title claims description 31
- 238000002360 preparation method Methods 0.000 title abstract description 19
- -1 p-methoxybenzyl Chemical group 0.000 claims abstract description 32
- WGQKYBSKWIADBV-UHFFFAOYSA-N benzylamine Chemical compound NCC1=CC=CC=C1 WGQKYBSKWIADBV-UHFFFAOYSA-N 0.000 claims abstract description 26
- 150000001875 compounds Chemical class 0.000 claims abstract description 24
- WJKHJLXJJJATHN-UHFFFAOYSA-N triflic anhydride Chemical compound FC(F)(F)S(=O)(=O)OS(=O)(=O)C(F)(F)F WJKHJLXJJJATHN-UHFFFAOYSA-N 0.000 claims abstract description 24
- VQOXUMQBYILCKR-UHFFFAOYSA-N 1-Tridecene Chemical compound CCCCCCCCCCCC=C VQOXUMQBYILCKR-UHFFFAOYSA-N 0.000 claims abstract description 18
- 229910052739 hydrogen Inorganic materials 0.000 claims abstract description 10
- 239000001257 hydrogen Substances 0.000 claims abstract description 10
- 238000000034 method Methods 0.000 claims abstract description 8
- 125000001797 benzyl group Chemical group [H]C1=C([H])C([H])=C(C([H])=C1[H])C([H])([H])* 0.000 claims abstract description 4
- NGXSWUFDCSEIOO-UHFFFAOYSA-N pyrrolidin-3-amine Chemical compound NC1CCNC1 NGXSWUFDCSEIOO-UHFFFAOYSA-N 0.000 claims abstract description 4
- 238000005865 alkene metathesis reaction Methods 0.000 claims abstract description 3
- YMWUJEATGCHHMB-UHFFFAOYSA-N Dichloromethane Chemical compound ClCCl YMWUJEATGCHHMB-UHFFFAOYSA-N 0.000 claims description 54
- 238000006243 chemical reaction Methods 0.000 claims description 45
- IJGRMHOSHXDMSA-UHFFFAOYSA-N Atomic nitrogen Chemical compound N#N IJGRMHOSHXDMSA-UHFFFAOYSA-N 0.000 claims description 42
- WEVYAHXRMPXWCK-UHFFFAOYSA-N Acetonitrile Chemical compound CC#N WEVYAHXRMPXWCK-UHFFFAOYSA-N 0.000 claims description 33
- 229910052757 nitrogen Inorganic materials 0.000 claims description 23
- 239000012043 crude product Substances 0.000 claims description 21
- 239000003814 drug Substances 0.000 claims description 19
- KDLHZDBZIXYQEI-UHFFFAOYSA-N Palladium on carbon Substances [Pd] KDLHZDBZIXYQEI-UHFFFAOYSA-N 0.000 claims description 18
- 239000002904 solvent Substances 0.000 claims description 18
- VEXZGXHMUGYJMC-UHFFFAOYSA-N Hydrochloric acid Chemical compound Cl VEXZGXHMUGYJMC-UHFFFAOYSA-N 0.000 claims description 16
- XEKOWRVHYACXOJ-UHFFFAOYSA-N Ethyl acetate Chemical compound CCOC(C)=O XEKOWRVHYACXOJ-UHFFFAOYSA-N 0.000 claims description 15
- OKKJLVBELUTLKV-UHFFFAOYSA-N Methanol Chemical compound OC OKKJLVBELUTLKV-UHFFFAOYSA-N 0.000 claims description 15
- VYPSYNLAJGMNEJ-UHFFFAOYSA-N Silicium dioxide Chemical compound O=[Si]=O VYPSYNLAJGMNEJ-UHFFFAOYSA-N 0.000 claims description 15
- RMVRSNDYEFQCLF-UHFFFAOYSA-N thiophenol Chemical compound SC1=CC=CC=C1 RMVRSNDYEFQCLF-UHFFFAOYSA-N 0.000 claims description 15
- 206010009944 Colon cancer Diseases 0.000 claims description 14
- 238000001914 filtration Methods 0.000 claims description 13
- 201000007270 liver cancer Diseases 0.000 claims description 13
- 208000014018 liver neoplasm Diseases 0.000 claims description 13
- 238000010898 silica gel chromatography Methods 0.000 claims description 13
- JUJWROOIHBZHMG-UHFFFAOYSA-N Pyridine Chemical compound C1=CC=NC=C1 JUJWROOIHBZHMG-UHFFFAOYSA-N 0.000 claims description 12
- 208000029742 colonic neoplasm Diseases 0.000 claims description 12
- 201000001441 melanoma Diseases 0.000 claims description 11
- XLYOFNOQVPJJNP-UHFFFAOYSA-N water Substances O XLYOFNOQVPJJNP-UHFFFAOYSA-N 0.000 claims description 9
- 239000012295 chemical reaction liquid Substances 0.000 claims description 8
- 238000001035 drying Methods 0.000 claims description 8
- UFHFLCQGNIYNRP-UHFFFAOYSA-N Hydrogen Chemical compound [H][H] UFHFLCQGNIYNRP-UHFFFAOYSA-N 0.000 claims description 7
- 206010058467 Lung neoplasm malignant Diseases 0.000 claims description 6
- CSNNHWWHGAXBCP-UHFFFAOYSA-L Magnesium sulfate Chemical compound [Mg+2].[O-][S+2]([O-])([O-])[O-] CSNNHWWHGAXBCP-UHFFFAOYSA-L 0.000 claims description 6
- 239000005457 ice water Substances 0.000 claims description 6
- 201000005202 lung cancer Diseases 0.000 claims description 6
- 208000020816 lung neoplasm Diseases 0.000 claims description 6
- 125000001736 nosyl group Chemical group S(=O)(=O)(C1=CC=C([N+](=O)[O-])C=C1)* 0.000 claims description 6
- 239000000047 product Substances 0.000 claims description 6
- UMJSCPRVCHMLSP-UHFFFAOYSA-N pyridine Natural products COC1=CC=CN=C1 UMJSCPRVCHMLSP-UHFFFAOYSA-N 0.000 claims description 6
- 206010028980 Neoplasm Diseases 0.000 claims description 5
- UIIMBOGNXHQVGW-UHFFFAOYSA-M Sodium bicarbonate Chemical class [Na+].OC([O-])=O UIIMBOGNXHQVGW-UHFFFAOYSA-M 0.000 claims description 5
- FJDQFPXHSGXQBY-UHFFFAOYSA-L caesium carbonate Chemical compound [Cs+].[Cs+].[O-]C([O-])=O FJDQFPXHSGXQBY-UHFFFAOYSA-L 0.000 claims description 5
- 229910000024 caesium carbonate Inorganic materials 0.000 claims description 5
- JRZJOMJEPLMPRA-UHFFFAOYSA-N olefin Natural products CCCCCCCC=C JRZJOMJEPLMPRA-UHFFFAOYSA-N 0.000 claims description 5
- 238000010992 reflux Methods 0.000 claims description 5
- 206010030155 Oesophageal carcinoma Diseases 0.000 claims description 4
- 238000001704 evaporation Methods 0.000 claims description 4
- JHHZLHWJQPUNKB-UHFFFAOYSA-N pyrrolidin-3-ol Chemical compound OC1CCNC1 JHHZLHWJQPUNKB-UHFFFAOYSA-N 0.000 claims description 4
- 208000000461 Esophageal Neoplasms Diseases 0.000 claims description 3
- 201000011510 cancer Diseases 0.000 claims description 3
- 238000001816 cooling Methods 0.000 claims description 3
- 201000004101 esophageal cancer Diseases 0.000 claims description 3
- 229910052763 palladium Inorganic materials 0.000 claims description 3
- 238000002390 rotary evaporation Methods 0.000 claims description 3
- 238000005406 washing Methods 0.000 claims description 3
- QGZKDVFQNNGYKY-UHFFFAOYSA-O Ammonium Chemical compound [NH4+] QGZKDVFQNNGYKY-UHFFFAOYSA-O 0.000 claims description 2
- 229910002651 NO3 Inorganic materials 0.000 claims description 2
- NHNBFGGVMKEFGY-UHFFFAOYSA-N Nitrate Chemical compound [O-][N+]([O-])=O NHNBFGGVMKEFGY-UHFFFAOYSA-N 0.000 claims description 2
- 239000003054 catalyst Substances 0.000 claims description 2
- 238000010438 heat treatment Methods 0.000 claims description 2
- 150000003839 salts Chemical class 0.000 claims description 2
- 229920006395 saturated elastomer Polymers 0.000 claims description 2
- 206010006187 Breast cancer Diseases 0.000 claims 1
- 208000026310 Breast neoplasm Diseases 0.000 claims 1
- 208000005718 Stomach Neoplasms Diseases 0.000 claims 1
- 206010017758 gastric cancer Diseases 0.000 claims 1
- 201000011549 stomach cancer Diseases 0.000 claims 1
- 229940125782 compound 2 Drugs 0.000 abstract description 19
- 239000002246 antineoplastic agent Substances 0.000 abstract description 3
- 229940041181 antineoplastic drug Drugs 0.000 abstract description 3
- 125000004217 4-methoxybenzyl group Chemical group [H]C1=C([H])C(=C([H])C([H])=C1OC([H])([H])[H])C([H])([H])* 0.000 abstract 1
- 230000003449 preventive effect Effects 0.000 abstract 1
- 230000001225 therapeutic effect Effects 0.000 abstract 1
- 210000004027 cell Anatomy 0.000 description 46
- RTZKZFJDLAIYFH-UHFFFAOYSA-N Diethyl ether Chemical compound CCOCC RTZKZFJDLAIYFH-UHFFFAOYSA-N 0.000 description 18
- 239000000243 solution Substances 0.000 description 16
- IAZDPXIOMUYVGZ-UHFFFAOYSA-N Dimethylsulphoxide Chemical compound CS(C)=O IAZDPXIOMUYVGZ-UHFFFAOYSA-N 0.000 description 11
- 229940125904 compound 1 Drugs 0.000 description 11
- 208000010507 Adenocarcinoma of Lung Diseases 0.000 description 9
- XMPZTFVPEKAKFH-UHFFFAOYSA-P ceric ammonium nitrate Chemical compound [NH4+].[NH4+].[Ce+4].[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O.[O-][N+]([O-])=O XMPZTFVPEKAKFH-UHFFFAOYSA-P 0.000 description 9
- 125000001495 ethyl group Chemical group [H]C([H])([H])C([H])([H])* 0.000 description 9
- 230000002401 inhibitory effect Effects 0.000 description 9
- 201000005249 lung adenocarcinoma Diseases 0.000 description 9
- 239000003208 petroleum Substances 0.000 description 9
- 239000007858 starting material Substances 0.000 description 9
- 230000005764 inhibitory process Effects 0.000 description 8
- 230000015572 biosynthetic process Effects 0.000 description 7
- 238000003786 synthesis reaction Methods 0.000 description 7
- 238000012258 culturing Methods 0.000 description 6
- 230000008034 disappearance Effects 0.000 description 6
- 229940079593 drug Drugs 0.000 description 6
- 230000000694 effects Effects 0.000 description 6
- 230000004663 cell proliferation Effects 0.000 description 5
- 238000002474 experimental method Methods 0.000 description 5
- FAPWRFPIFSIZLT-UHFFFAOYSA-M Sodium chloride Chemical class [Na+].[Cl-] FAPWRFPIFSIZLT-UHFFFAOYSA-M 0.000 description 4
- 230000000259 anti-tumor effect Effects 0.000 description 4
- 238000012658 bimolecular nucleophilic substitution Methods 0.000 description 4
- 239000001963 growth medium Substances 0.000 description 4
- 239000003112 inhibitor Substances 0.000 description 4
- QJGQUHMNIGDVPM-UHFFFAOYSA-N nitrogen(.) Chemical compound [N] QJGQUHMNIGDVPM-UHFFFAOYSA-N 0.000 description 4
- 230000035484 reaction time Effects 0.000 description 4
- 230000002194 synthesizing effect Effects 0.000 description 4
- 210000004881 tumor cell Anatomy 0.000 description 4
- 239000006227 byproduct Substances 0.000 description 3
- 238000004113 cell culture Methods 0.000 description 3
- WGLUMOCWFMKWIL-UHFFFAOYSA-N dichloromethane;methanol Chemical compound OC.ClCCl WGLUMOCWFMKWIL-UHFFFAOYSA-N 0.000 description 3
- 239000011541 reaction mixture Substances 0.000 description 3
- 239000007787 solid Substances 0.000 description 3
- RWRDLPDLKQPQOW-UHFFFAOYSA-N tetrahydropyrrole Substances C1CCNC1 RWRDLPDLKQPQOW-UHFFFAOYSA-N 0.000 description 3
- 238000001644 13C nuclear magnetic resonance spectroscopy Methods 0.000 description 2
- 238000005160 1H NMR spectroscopy Methods 0.000 description 2
- 108091003079 Bovine Serum Albumin Proteins 0.000 description 2
- UIIMBOGNXHQVGW-DEQYMQKBSA-M Sodium bicarbonate-14C Chemical class [Na+].O[14C]([O-])=O UIIMBOGNXHQVGW-DEQYMQKBSA-M 0.000 description 2
- HEDRZPFGACZZDS-MICDWDOJSA-N Trichloro(2H)methane Chemical compound [2H]C(Cl)(Cl)Cl HEDRZPFGACZZDS-MICDWDOJSA-N 0.000 description 2
- 230000001464 adherent effect Effects 0.000 description 2
- 239000006285 cell suspension Substances 0.000 description 2
- 239000012091 fetal bovine serum Substances 0.000 description 2
- 206010073071 hepatocellular carcinoma Diseases 0.000 description 2
- 239000007788 liquid Substances 0.000 description 2
- 239000000203 mixture Substances 0.000 description 2
- 239000012074 organic phase Substances 0.000 description 2
- 230000035755 proliferation Effects 0.000 description 2
- 125000006239 protecting group Chemical group 0.000 description 2
- 239000002994 raw material Substances 0.000 description 2
- FBQCDJRSCUVUFL-BZSNNMDCSA-N (2s,3s,4s)-4-amino-2-tetradecyloxolan-3-ol Chemical compound CCCCCCCCCCCCCC[C@@H]1OC[C@H](N)[C@@H]1O FBQCDJRSCUVUFL-BZSNNMDCSA-N 0.000 description 1
- FBQCDJRSCUVUFL-UHFFFAOYSA-N 4-amino-3-hydroxy-2-tetradecyltetrahydrofuran Natural products CCCCCCCCCCCCCCC1OCC(N)C1O FBQCDJRSCUVUFL-UHFFFAOYSA-N 0.000 description 1
- 208000017897 Carcinoma of esophagus Diseases 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
- LFQSCWFLJHTTHZ-UHFFFAOYSA-N Ethanol Chemical compound CCO LFQSCWFLJHTTHZ-UHFFFAOYSA-N 0.000 description 1
- WQZGKKKJIJFFOK-GASJEMHNSA-N Glucose Natural products OC[C@H]1OC(O)[C@H](O)[C@@H](O)[C@@H]1O WQZGKKKJIJFFOK-GASJEMHNSA-N 0.000 description 1
- 108010019160 Pancreatin Proteins 0.000 description 1
- 239000012980 RPMI-1640 medium Substances 0.000 description 1
- 235000005811 Viola adunca Nutrition 0.000 description 1
- 240000009038 Viola odorata Species 0.000 description 1
- 235000013487 Viola odorata Nutrition 0.000 description 1
- 235000002254 Viola papilionacea Nutrition 0.000 description 1
- 150000001335 aliphatic alkanes Chemical class 0.000 description 1
- 150000001336 alkenes Chemical class 0.000 description 1
- 230000003698 anagen phase Effects 0.000 description 1
- 230000009286 beneficial effect Effects 0.000 description 1
- 125000000319 biphenyl-4-yl group Chemical group [H]C1=C([H])C([H])=C([H])C([H])=C1C1=C([H])C([H])=C([*])C([H])=C1[H] 0.000 description 1
- 238000007664 blowing Methods 0.000 description 1
- 244000309464 bull Species 0.000 description 1
- 230000001413 cellular effect Effects 0.000 description 1
- DQLATGHUWYMOKM-UHFFFAOYSA-L cisplatin Chemical compound N[Pt](N)(Cl)Cl DQLATGHUWYMOKM-UHFFFAOYSA-L 0.000 description 1
- 229960004316 cisplatin Drugs 0.000 description 1
- 229940126214 compound 3 Drugs 0.000 description 1
- 238000005138 cryopreservation Methods 0.000 description 1
- 238000000354 decomposition reaction Methods 0.000 description 1
- 238000007865 diluting Methods 0.000 description 1
- 230000009982 effect on human Effects 0.000 description 1
- 201000005619 esophageal carcinoma Diseases 0.000 description 1
- 239000000706 filtrate Substances 0.000 description 1
- 239000012467 final product Substances 0.000 description 1
- 238000007710 freezing Methods 0.000 description 1
- 230000008014 freezing Effects 0.000 description 1
- 239000008103 glucose Substances 0.000 description 1
- 230000012010 growth Effects 0.000 description 1
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 1
- 238000002372 labelling Methods 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 239000002609 medium Substances 0.000 description 1
- 239000012982 microporous membrane Substances 0.000 description 1
- VMGAPWLDMVPYIA-HIDZBRGKSA-N n'-amino-n-iminomethanimidamide Chemical compound N\N=C\N=N VMGAPWLDMVPYIA-HIDZBRGKSA-N 0.000 description 1
- 229930014626 natural product Natural products 0.000 description 1
- 239000013642 negative control Substances 0.000 description 1
- 229940055695 pancreatin Drugs 0.000 description 1
- 230000000144 pharmacologic effect Effects 0.000 description 1
- 239000011148 porous material Substances 0.000 description 1
- 239000013641 positive control Substances 0.000 description 1
- 239000000843 powder Substances 0.000 description 1
- 125000002924 primary amino group Chemical group [H]N([H])* 0.000 description 1
- 239000000376 reactant Substances 0.000 description 1
- BOLDJAUMGUJJKM-LSDHHAIUSA-N renifolin D Natural products CC(=C)[C@@H]1Cc2c(O)c(O)ccc2[C@H]1CC(=O)c3ccc(O)cc3O BOLDJAUMGUJJKM-LSDHHAIUSA-N 0.000 description 1
- 238000000926 separation method Methods 0.000 description 1
- 235000002639 sodium chloride Nutrition 0.000 description 1
- HPALAKNZSZLMCH-UHFFFAOYSA-M sodium;chloride;hydrate Chemical class O.[Na+].[Cl-] HPALAKNZSZLMCH-UHFFFAOYSA-M 0.000 description 1
- 239000010902 straw Substances 0.000 description 1
- 125000000472 sulfonyl group Chemical group *S(*)(=O)=O 0.000 description 1
- 239000006228 supernatant Substances 0.000 description 1
- 230000001502 supplementing effect Effects 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- AYPRHVFFCLKSBU-UHFFFAOYSA-N trifluoromethanesulfonoperoxoic acid Chemical compound OOS(=O)(=O)C(F)(F)F AYPRHVFFCLKSBU-UHFFFAOYSA-N 0.000 description 1
- 238000005303 weighing Methods 0.000 description 1
Classifications
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- A—HUMAN NECESSITIES
- A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
- A61P—SPECIFIC THERAPEUTIC ACTIVITY OF CHEMICAL COMPOUNDS OR MEDICINAL PREPARATIONS
- A61P35/00—Antineoplastic agents
-
- C—CHEMISTRY; METALLURGY
- C07—ORGANIC CHEMISTRY
- C07D—HETEROCYCLIC COMPOUNDS
- C07D207/00—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom
- C07D207/02—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom
- C07D207/04—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members
- C07D207/10—Heterocyclic compounds containing five-membered rings not condensed with other rings, with one nitrogen atom as the only ring hetero atom with only hydrogen or carbon atoms directly attached to the ring nitrogen atom having no double bonds between ring members or between ring members and non-ring members with hetero atoms or with carbon atoms having three bonds to hetero atoms with at the most one bond to halogen, e.g. ester or nitrile radicals, directly attached to ring carbon atoms
- C07D207/14—Nitrogen atoms not forming part of a nitro radical
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- Chemical & Material Sciences (AREA)
- Organic Chemistry (AREA)
- Health & Medical Sciences (AREA)
- General Chemical & Material Sciences (AREA)
- Medicinal Chemistry (AREA)
- Nuclear Medicine, Radiotherapy & Molecular Imaging (AREA)
- Chemical Kinetics & Catalysis (AREA)
- Pharmacology & Pharmacy (AREA)
- Life Sciences & Earth Sciences (AREA)
- Animal Behavior & Ethology (AREA)
- General Health & Medical Sciences (AREA)
- Public Health (AREA)
- Veterinary Medicine (AREA)
- Pharmaceuticals Containing Other Organic And Inorganic Compounds (AREA)
- Pyrrole Compounds (AREA)
Abstract
本发明涉及药物化学领域，具体涉及一种(2R,3S,4S)‑4‑氨基‑2‑十四烷基吡咯烷‑3‑醇的制备方法及其应用。该方法包含如下步骤：(3R,4R,5R)‑5‑烯丙基‑4‑((4‑甲氧基苄基)氧基)‑1‑((4‑硝基苯基)磺酰基)吡咯烷‑3‑醇，与三氟甲磺酸酐反应后，经卞胺处理得(3S,4S,5R)‑5‑烯丙基‑N‑苄基‑4‑((4‑甲氧基苄基)氧基)‑1‑((2‑硝基苯基)磺酰基)吡咯烷‑3‑胺（化合物2），化合物2与1‑十三烯发生烯烃复分解反应后，脱去邻硝基苯磺酰基，氢气双键还原，脱除苄基，继而脱除对甲氧基苄基，得化合物P1。(2R,3S,4S)‑4‑氨基‑2‑十四烷基吡咯烷‑3‑醇在制备预防和/或治疗抗癌药物中的应用。The invention relates to the field of medicinal chemistry, in particular to a preparation method of (2R, 3S, 4S)-4-amino-2-tetradecylpyrrolidine-3-alcohol and application thereof. The method comprises the steps of: (3R, 4R, 5R)-5-allyl-4-((4-methoxybenzyl)oxy)-1-((4-nitrophenyl)sulfonyl) Pyrrolidine-3-alcohol, after reacting with trifluoromethanesulfonic anhydride, is treated with benzylamine to obtain (3S,4S,5R)-5-allyl-N-benzyl-4-((4-methoxybenzyl base)oxy)-1-((2-nitrophenyl)sulfonyl)pyrrolidine-3-amine (compound 2), after compound 2 and 1-tridecene undergo olefin metathesis reaction, remove the o-nitro group Benzenesulfonyl, hydrogen double bond reduction, removal of benzyl, and then removal of p-methoxybenzyl to obtain compound P1. Application of (2R,3S,4S)-4-amino-2-tetradecylpyrrolidin-3-alcohol in the preparation of preventive and/or therapeutic anticancer drugs.
Description
Technical Field
The invention relates to the field of medicinal chemistry, in particular to a preparation method and application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol.
Background
A method for preparing (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (P1) is disclosed in the literature "Practical Synthesis of Pachamissamine (Jaspine B), 2-epi-Pachamissamine, and the 2-epi-Pyrrolidine analog" (chem.Pharm.Bull.2016,64, 179-188), and the specific method is as follows:
the compounds represented by the above structural formula are hereinafter referred to as compound 1, compound 2 (or intermediate 2), compound 3 (or intermediate 3), compound 4 (or intermediate 4), and compound P1.
The method comprises the steps of taking a 2,3, 4-trisubstituted allyl pyrrolidine compound (3R,4R,5R) -5-allyl-4- ((4-methoxybenzyl) oxy) -1- ((4-nitrophenyl) sulfonyl) pyrrolidine-3-alcohol (compound 1) as a raw material, synthesizing an intermediate 2 through hydroxyl triflate and bimolecular nucleophilic substitution reaction, carrying out olefin double decomposition reaction on the intermediate 2 and 1-tridecene, and finally removing protecting groups on amino and hydroxyl and reducing double bonds to obtain the (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol (a target compound P1).
This route suffers mainly from the following disadvantages:
1) in the synthesis of an intermediate compound 2, the reaction condition of a compound 1 and trifluoromethanesulfonic anhydride is-20 ℃ to 0 ℃, the subsequent bimolecular nucleophilic substitution reaction takes benzylamine as a solvent and reacts for 40 hours at 50 ℃, the yield of 2 steps is only 50%, and a byproduct is generated
And (4) generating.
2) The reaction for synthesizing the target compound P1 from the intermediate 2 is a separate 4-step reaction reported in the literature, wherein 3 steps need to be separated and purified by silica gel column chromatography, and the operation is complex. In addition, p-methoxybenzyl (PMB) is difficult to remove, the hydrogen reduction time takes 4 days or more, the reaction time is long, the yield is 44%, and the reproducibility is low, and when the experiment is repeated, the total yield of 4 steps is 10%.
Disclosure of Invention
The invention aims to provide a preparation method of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol.
The invention also aims to provide the application of the (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-ol in preparing anti-cancer drugs.
The invention is realized by the following technical scheme:
a preparation method of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-ol (P1) is provided, wherein the structural formula of the (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-ol (P1) is shown in the specification
Comprises the following steps:
the preparation method of the (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (P1) comprises the following steps:
s1: (3R,4R,5R) -5-allyl-4- ((4-methoxybenzyl) oxy) -1- ((4-nitrophenyl) sulfonyl) pyrrolidin-3-ol (compound 1), reacted with trifluoromethanesulfonic anhydride, and then treated with benzylamine to give 4-amino substituted compound (3S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidin-3-amine (compound 2);
s2: after the (3S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidine-3-amine (compound 2) and 1-tridecene are subjected to olefin metathesis reaction, o-nitrobenzenesulfonyl is removed, hydrogen double bond reduction is carried out, benzyl is removed, and then P-methoxybenzyl is removed, so that (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-ol (compound P1) is obtained.
In the preparation method of the (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-ol, the reaction temperature of the benzylamine treatment in the step S1 is 30-40 ℃, and the reaction time is 0.5-1.5 h. Preferably, the reaction temperature of the benzylamine treatment in the step S1 is 35 ℃, and the reaction time is 1 h.
In the above preparation method of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol, S1 comprises the following steps:
dissolving (3R,4R,5R) -5-allyl-4- ((4-methoxybenzyl) oxy) -1- ((4-nitrophenyl) sulfonyl) pyrrolidine-3-ol (compound 1) in dichloromethane, adding pyridine, dropwise adding trifluoromethanesulfonic anhydride at-20 ℃ under the protection of nitrogen, reacting for 20min at-20 ℃, washing an organic phase with hydrochloric acid (1M), a saturated sodium bicarbonate solution, ice water and saturated common salt water, drying anhydrous magnesium sulfate, filtering, evaporating the solvent under reduced pressure, directly dissolving the obtained crude product in benzylamine, reacting for 0.5-1.5h at 30-40 ℃ under the protection of nitrogen, and directly separating and purifying the reaction solution by using a silica gel column chromatography to obtain (3S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidin-3-amine (Compound 2).
Preferably, the S1 includes the following steps: dissolving (3R,4R,5R) -5-allyl-4- ((4-methoxybenzyl) oxy) -1- ((4-nitrophenyl) sulfonyl) pyrrolidine-3-ol (compound 1) in dichloromethane, adding pyridine, dropwise adding trifluoromethanesulfonic anhydride at-20 ℃ under the protection of nitrogen, reacting for 20min at-20 ℃, washing an organic phase with hydrochloric acid (1M), a saturated sodium bicarbonate solution, ice water and saturated salt water, drying anhydrous magnesium sulfate, filtering, evaporating the solvent under reduced pressure, directly dissolving the obtained crude product in benzylamine, reacting for 1h at 35 ℃ under the protection of nitrogen, and directly separating and purifying the reaction liquid by using a silica gel column chromatography to obtain (3S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitre) Phenylphenyl) sulfonyl) pyrrolidin-3-amine (compound 2).
In the above preparation method of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol, ceric ammonium nitrate is used as a reactant in the step of removing p-methoxybenzyl in step S2.
In the above preparation method of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol, S2 comprises the following steps: dissolving (3S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidin-3-amine (Compound 2) in dichloromethane, and adding 1-tridecene, Grubbs under nitrogen2ndHeating and refluxing the catalyst for 7 hours to obtain a reaction solutionCooling to room temperature, filtering through diatomite, eluting with ethyl acetate, and rotary evaporating to remove solvent to obtain crude olefin compound; dissolving the crude olefin compound in acetonitrile, adding cesium carbonate and thiophenol under the protection of nitrogen, reacting for 3h at 50 ℃, cooling the reaction liquid to room temperature, filtering through diatomite, eluting with dichloromethane, and removing the solvent by rotary evaporation to obtain the crude compound without o-nitrobenzenesulfonyl; the crude compound obtained is dissolved in methanol and 20% Pd (OH) is added2Reacting 30 wt% of/C with 10 wt% of Pd/C (10 wt%) for 24 hours in a hydrogen atmosphere, filtering the reaction liquid by using kieselguhr, and spin-drying the solvent to obtain a crude product of the reduction product without benzyl; dissolving the benzyl-removed crude product of the reduction product in acetonitrile/water (1:4), adding ammonium ceric nitrate, reacting for 3h at room temperature, and directly separating and purifying the reaction liquid by using a silica gel column chromatography to obtain (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol (compound P1).
In the present invention, the temperature of the hydrochloric acid, the saturated sodium bicarbonate solution, and the saturated brine is 0 ℃.
The concentration of the hydrochloric acid is 1M.
The ratio of acetonitrile/water (1:4) is volume ratio.
The 20% Pd (OH)2The amount of the/C is 30 wt%, and the amount of the 10% Pd/C is 10 wt%.
Application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol (P1) in preparing anticancer drugs. The cancer includes lung cancer, colon cancer, melanoma, and liver cancer.
The application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing a medicament for treating lung cancer.
Application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing medicine for treating colon cancer.
Application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing medicine for treating melanoma.
The application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing medicine for treating liver cancer.
Application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing medicines for inhibiting human lung cancer A549 cell proliferation.
Application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparation of medicines for inhibiting human colon cancer LOVO cell proliferation.
The application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing a medicament for inhibiting the cell proliferation of a human melanoma cell A375.
The application of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing a medicament for inhibiting the cell proliferation of human liver cancer HepG 2.
The invention has the beneficial effects that:
(1) the preparation method provided by the invention has the characteristics of no need of intermediate separation, mild reaction conditions, gram-scale achievement, simple operation, high reproducibility and the like.
(2) In the synthesis of intermediate compound 2, the reaction conditions of compound 1 and trifluoromethanesulfonic anhydride are-20 ℃ to 0 ℃ as reported in the literature, and the subsequent bimolecular nucleophilic substitution reaction takes benzylamine as solvent and takes reaction at 50 ℃ for 40 hours, so that the yield of 2 steps is only 50%, and side products are generated (24%). The inventor finds that the reaction condition of the compound 1 and the trifluoromethanesulfonic anhydride is-20 ℃, the subsequent bimolecular nucleophilic substitution reaction only needs to be heated to 35 ℃, the raw material point completely disappears in 1 hour, no by-product is generated due to overhigh temperature, the reaction yield is 78%, and the reaction yield is improved by nearly 30% compared with that reported in the literature.
(3) The reaction for synthesizing the target compound P1 from the intermediate 2 is a separate 4-step reaction reported in the literature, wherein 3 steps need to be separated and purified by silica gel column chromatography, and the operation is complex. In addition, p-methoxybenzyl (PMB) was difficult to remove, the hydrogen reduction time required 4 days or more, the reaction time was long, the yield was 44%, and the reproducibility was low, and the average yield of 4 steps was 10% when the experiment was repeated. Although the method for synthesizing P1 from 2 is completed through 4 steps, only the final product P1 needs to be purified, when the P-methoxybenzyl (PMB) protecting group is removed, the reaction is shortened to 3 hours by using Ceric Ammonium Nitrate (CAN), the operation is simple, the total yield of 4 steps is 41%, and the repeatability is high.
(4) The result of the experiment on the proliferation activity of the anti-tumor cells shows that (2R,3S,4S)) IC of 4-amino-2-tetradecylpyrrolidin-3-ol on human Lung cancer cells (A549), human Colon cancer cells (LOVO), human melanoma cells (A375), human hepatoma cells (HepG2)50The values are all less than 10 mu M, and the synthetic drug IC meets the requirements specified in the preclinical research guiding principle of new traditional Chinese medicine50The inhibitor is considered to have certain inhibition effect when the concentration is less than or equal to 10 mu M. The compound (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol has certain inhibiting effect on human lung cancer cells (A549), human colon cancer cells (LOVO), human melanoma cells (A375) and human liver cancer cells (HepG2), wherein the inhibiting effect on IC of the human lung adenocarcinoma cells (A549), the human colon cancer cells (LOVO) and the human liver cancer cells (HepG2)50Are all lower than 0.1 mu M, which shows that the compound (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol has strong inhibition effect on the proliferation of human lung adenocarcinoma (A549), human colon cancer (LOVO) and human liver cancer (HepG 2).
Detailed Description
The present invention will be further described with reference to specific embodiments so that those skilled in the art may better understand the present invention, but the present invention is not limited thereto.
EXAMPLE 1 Synthesis of intermediate Compound 2
Compound 1(1.08g,2.4mmol) was dissolved in dichloromethane (9.6mL), pyridine (396 μ L,4.8mmol) was added, trifluoromethanesulfonic anhydride (605 μ L,3.6mmol) was added dropwise at-20 ℃ under nitrogen, the reaction was carried out for 20min, and the disappearance of the starting material spot was detected by TLC (ethyl acetate-petroleum ether ═ 1: 2). The reaction solution was diluted with dichloromethane, washed with 0 ℃ 1M hydrochloric acid, 0 ℃ saturated sodium bicarbonate, ice water, 0 ℃ saturated common salt in this order, dried over anhydrous magnesium sulfate, filtered, and the solvent was removed under reduced pressure, the crude product was directly dissolved in benzylamine (7.2mL), reacted at 35 ℃ for 1 hour under nitrogen protection, and the reaction solution was separated and purified by silica gel column chromatography (ethyl acetate-dichloromethane-petroleum ether ═ 3:3:4) to obtain compound 2(0.787g, 61%).1H NMR(CDCl3,600MHz)δ(ppm):7.94(dd,J＝8.1,1.2Hz,1H),7.56–7.61(m,1H),7.48– 7.54(m,2H),7.19–7.32(m,5H),7.00(d,J＝8.6Hz,2H),6.77–6.83(m,2H),5.73–5.82(m, 1H),5.05–5.15(m,2H),4.33(d,J＝11.2Hz,1H),4.09–4.16(m,2H),3.80(s,3H),3.72(dd,J＝ 9.1,7.1Hz,1H),3.63–3.68(m,2H),3.60(d,J＝13.1Hz,1H),3.40(ddd,J＝9.7,7.1,4.0Hz,1H), 3.12(t,J＝9.3Hz,1H),2.56–2.63(m,1H),2.14–2.22(m,1H),1.85(s,1H).13C NMR(CDCl3, 150MHz)δ(ppm):159.39,148.72,139.72,133.56,133.27,131.15,130.64,130.39,129.46, 129.28,128.49,128.05,127.24,123.79,118.70,113.76,78.42,70.42,63.38,57.61,55.32,51.96, 51.62,38.86.HRMS(ESI):m/zcalcd for C28H32N3O6S(M+H)+:538.2012.found:538.2009.
Example 2 Synthesis of intermediate Compound 2
Compound 1(0.36g,0.8mmol) was dissolved in dichloromethane (5.0mL), pyridine (132 μ L,1.6mmol) was added, trifluoromethanesulfonic anhydride (202 μ L,1.2mmol) was added dropwise at-20 ℃ under nitrogen, the reaction was carried out for 20min, and the disappearance of the starting material spot was detected by TLC (ethyl acetate-petroleum ether ═ 1: 2). The reaction solution was diluted with dichloromethane, washed with 1M hydrochloric acid at 0 ℃, saturated sodium bicarbonate at 0 ℃, ice water, and saturated common salt at 0 ℃, dried over anhydrous magnesium sulfate, filtered, and the solvent was removed under reduced pressure, the crude product was directly dissolved in benzylamine (3.0mL), reacted at 35 ℃ for 1 hour under nitrogen protection, and the reaction solution was separated and purified by silica gel column chromatography (ethyl acetate-dichloromethane-petroleum ether ═ 3:3:4) to obtain compound 2(0.335g, 78%).
EXAMPLE 3 Synthesis of intermediate Compound 2
Compound 1(1.44g,3.2mmol) was dissolved in dichloromethane (20.0ml), pyridine (528 μ L,6.4mmol) was added, trifluoromethanesulfonic anhydride (808 μ L,4.8mmol) was added dropwise at-20 ℃ under nitrogen, the reaction was carried out for 20min, and the disappearance of the starting material spot was detected by TLC (ethyl acetate-petroleum ether ═ 1: 2). The reaction solution was diluted with dichloromethane, washed with 0 ℃ 1M hydrochloric acid, 0 ℃ saturated sodium bicarbonate, ice water, 0 ℃ saturated common salt in this order, dried over anhydrous magnesium sulfate, filtered, and the solvent was removed under reduced pressure, the crude product was directly dissolved in benzylamine (12.0mL), reacted at 35 ℃ for 1 hour under nitrogen protection, and the reaction solution was separated and purified by silica gel column chromatography (ethyl acetate-dichloromethane-petroleum ether ═ 3:3:4) to give compound 2(1.308g, 76%).
EXAMPLE 4 preparation of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (P1)
Compound 2(100mg,0.186mmol) was dissolved in dichloromethane (6mL) and 1-tridecene (136mg,0.744mmol), Grubbs, added under nitrogen2nd(31mg,0.037mmol) and the reaction was heated to reflux for 7h and the starting material disappeared by TLC (3: 2:5 ethyl acetate-dichloromethane-petroleum ether). The reaction was cooled to room temperature and filtered through celite, eluting with ethyl acetate to give a brown crude. The crude product was dissolved in acetonitrile (6mL), and cesium carbonate (121mg,0.372mmol), thiophenol (31mg,0.28mmol) were added under nitrogen protection, reacted at 50 ℃ for 3h, and the disappearance of the starting material was detected by TLC (methanol-dichloromethane ═ 7: 93). The reaction was cooled to room temperature and filtered through celite, eluting with dichloromethane, to give a brown crude. Dissolving the obtained crude product in methanol (6mL), adding 20% palladium on carbon (30mg) and 10% palladium on carbon (10 mg), reacting for 24h under hydrogen atmosphere, filtering the reaction solution with diatomite, and spin-drying the solvent to obtain colorless oily crude product. The resulting crude product was dissolved in acetonitrile/water (1:4), and cerium ammonium nitrate (174mg,0.372mmol) was added to react at room temperature for 3 hours. The reaction mixture was directly subjected to silica gel column chromatography (methanol-ethanol-dichloromethane-aqueous ammonia: 6:12:77:5) to give (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (23mg, 41%) as a white solid.
1H NMR (CDCl3,400MHz)δ(ppm):3.48(t,J＝5.6Hz,1H),3.29–3.43(m,2H),2.82(q,J＝5.6Hz, 1H),2.60(dq,J＝11.6,6.0,5.5Hz,1H),1.84(brs,4H),1.54–1.63(m,1H),1.25(s,25H),0.88(t, J＝6.7Hz,3H).13C NMR(CDCl3,100MHz)δ(ppm):75.72,65.00,53.15,51.73,34.02,31.85, 29.55,29.58(2C),29.62(4C),29.48,29.29,26.75,22.61,14.00.IR(KBr,cm-1):3348,3242,2918, 2850,1558,1469,1458,1377,1350,1118,1080,1028,956,908,894,869,825,719.HRMS(ESI): m/zcalcd for C18H38N2O(M+H)+:299.3062.found:299.3047.
Example 5 preparation of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (P1)
Compound 2(50mg,0.093mmol) was dissolved in dichloromethaneTo an alkane (5mL), 1-tridecene (68mg,0.372mmol), Grubbs, was added under nitrogen2nd(16mg,0.019mmol) and the reaction was heated under reflux for 7h and the starting material disappeared by TLC (ethyl acetate-dichloromethane-petroleum ether: 3:2: 5). The reaction was cooled to room temperature and filtered through celite, eluting with ethyl acetate to give a brown crude. The crude product was dissolved in acetonitrile (5mL), and cesium carbonate (60 mg,0.186mmol), thiophenol (16mg,0.14mmol) were added under nitrogen protection, reacted at 50 ℃ for 3h, and the disappearance of the starting material was detected by TLC (methanol-dichloromethane ═ 7: 93). The reaction was cooled to room temperature and filtered through celite, eluting with dichloromethane, to give a brown crude. Dissolving the obtained crude product in methanol (5mL), adding 20% palladium on carbon (15mg) and 10% palladium on carbon (5mg), reacting for 24h under hydrogen atmosphere, filtering the reaction solution with diatomite, and spin-drying the solvent to obtain colorless oily crude product. The crude product was dissolved in acetonitrile/water (1:4), and cerium ammonium nitrate (87mg,0.186mmol) was added to react at room temperature for 3 hours. The reaction mixture was directly subjected to silica gel column chromatography (methanol-ethanol-dichloromethane-aqueous ammonia: 6:12:77:5) to give (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (10mg, 36%) as a white solid.
Example 6 preparation of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol (P1)
Compound 2(150mg,0.279mmol) was dissolved in dichloromethane (15mL) and 1-tridecene (144mg,1.116mmol), Grubbs, added under nitrogen2nd(48mg,0.057mmol) and the reaction was heated to reflux for 7h and the starting material disappeared by TLC (3: 2:5 ethyl acetate-dichloromethane-petroleum ether). The reaction was cooled to room temperature and filtered through celite, eluting with ethyl acetate to give a brown crude. The crude product was dissolved in acetonitrile (15mL), and cesium carbonate (180mg,0.558mmol), thiophenol (48mg,0.42mmol) were added under nitrogen, reacted at 50 ℃ for 3h, and the disappearance of the starting material was detected by TLC (methanol-dichloromethane ═ 7: 93). The reaction was cooled to room temperature and filtered through celite, eluting with dichloromethane, to give a brown crude. Dissolving the obtained crude product in methanol (15mL), adding 20% palladium on carbon (45mg) and 10% palladium on carbon (15mg), reacting for 24h under hydrogen atmosphere, filtering the reaction solution with diatomite, and spin-drying the solvent to obtain colorless oily crude product. The crude product obtained is dissolved in acetonitrile/water (1:4),cerium ammonium nitrate (261mg,0.558mmol) was added thereto, and the mixture was reacted at room temperature for 3 hours. The reaction mixture was directly subjected to silica gel column chromatography (methanol-ethanol-dichloromethane-aqueous ammonia: 6:12:77:5) to give (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol as a white solid (28mg, 34%).
The pharmacological tests and results of the compounds of the invention are as follows:
experimental example 1: research on antitumor activity of compound of the invention
1.1 Experimental materials
Compound P1 was dissolved in dimethyl sulfoxide (DMSO, final concentration 0.4%) and prepared to 1mg/mL in 15% fetal bovine serum RPMI-1640 medium, and diluted to the desired concentration. Human lung adenocarcinoma cells (A549), human colon carcinoma cells (LOVO), human melanoma cells (A375), human hepatoma cells (HepG2) and human esophageal carcinoma cells (TE-1), all purchased from cell banks of Chinese academy of sciences, cultured in DMEM (high glucose) medium containing 15% fetal bovine serum, placed at 37 ℃ in 5% CO2Culturing in an incubator.
1.2 antitumor Activity test
1.2.1 MTT preparation method
Preparation of 5mg/mL MTT solution: weighing 500.0mg of MTT powder, dissolving in warm 100mL PBS, filtering with 0.22 μm pore size microporous membrane to obtain filtrate, subpackaging in autoclaved EP tube, and freezing at-20 deg.C in dark place.
1.2.2 cell culture and Experimental methods
Taking out the cryopreservation tube with the tumor cells from the liquid nitrogen, quickly putting the tube into a37 ℃ incubator, and shaking continuously until the tube is melted. After wiping the edge of the cryopreserving tube cover with 75% alcohol, sucking the cell suspension, transferring the cell suspension into a 10mL centrifuge tube, and supplementing 5mL culture medium. Centrifuging at low speed (25 deg.C, 3000r/min,5min), discarding supernatant, adding culture medium, and centrifuging once again. Diluting with appropriate amount of culture medium, blowing off cells with a straw to obtain suspension, transferring into a culture flask, and standing at 37 deg.C and 5% CO2Culturing in a cell culture box. The culture medium is replaced the next day, and the mixture is placed at 37 ℃ and 5% CO2And continuing culturing in the cell culture box.
Human lung adenocarcinoma cell (A549), human noduleIntestinal cancer cells (LOVO), human melanoma cells (A375), human liver cancer cells (HepG2) and human esophageal cancer cells (TE-1) are adherent cells, adherent tumor cells in logarithmic growth phase are washed according to the growth rate of the tumor cells, and the number of cells is adjusted to 1 × 10 by digesting with 0.25% EDTA pancreatin5Perml/mL in 96-well plates, 100. mu.L per well, at 37 ℃ in CO2Culturing in incubator, and administering after 24 h. The administration groups were dosed with different concentrations of drug (compound P1), each drug being provided in 5 dose groups of 100.00, 10.00, 1.00, 0.10, 0.01 μ M each, with three duplicate wells per concentration. A blank control, DMSO (0.8%) solvent control, and a cisplatin positive control were set. CO at 37 deg.C2After culturing in an incubator for 48 hours, the OD value was measured by the MTT method, and the cell inhibition ratio was calculated.
1.2.3 cellular IC50Calculation of values
After culturing human lung adenocarcinoma cells (A549), human colon cancer cells (LOVO), human melanoma cells (A375), human liver cancer cells (HepG2) and human esophageal cancer cells (TE-1) for 48h, the culture was terminated, and then 10. mu.L of 0.5% MTT solution was added to each well and placed in CO2In the incubator, after 4 hours, the liquid in each well was removed, then 0.2mL of DMSO solution was added, followed by sufficiently shaking with low frequency on a shaker to sufficiently dissolve formazan crystallized in blue-violet, placing in an enzyme-labeling instrument, recording OD at 490nm, calculating the average OD of three parallel wells of different concentrations, and calculating the cell inhibition ratio and IC of each test drug at different concentrations based on the average50Values (see table 1).
The inhibition rate (%) [ 1-test product OD value/negative control group OD value ] × 100%.
1.2.4 determination of results
The guiding principle of preclinical research of new Chinese medicine shows that the natural product IC50The inhibitor is considered to have certain inhibiting effect when the concentration is less than or equal to 30 mu M; synthesis of drug IC50The inhibitor is considered to have certain inhibition effect when the concentration is less than or equal to 10 mu M.
TABLE 1 IC of (2R,3S,4S) -4-amino-2-tetradecylpyrrolidin-3-ol against tumor cell proliferation50Value of
1.3 results and discussion
The result of the anti-tumor cell proliferation activity experiment shows that the IC of (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in human lung adenocarcinoma cells (A549), human colon carcinoma cells (LOVO), human melanoma cells (A375) and human liver cancer cells (HepG2)50The values are all less than 10 mu M, and the synthetic drug IC meets the requirements specified in the preclinical research guiding principle of new traditional Chinese medicine50The inhibitor is considered to have certain inhibition effect when the concentration is less than or equal to 10 mu M. The compound (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol has certain inhibiting effect on human lung adenocarcinoma cells (A549), human colon cancer cells (LOVO), human melanoma cells (A375) and human liver cancer cells (HepG2), wherein the inhibiting effect on IC of the human lung adenocarcinoma cells (A549), the human colon cancer cells (LOVO) and the human liver cancer cells (HepG2)50All are lower than 0.1 mu M, which shows that the compound (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-alcohol has strong inhibition effect on human lung adenocarcinoma (A549), human colon cancer (LOVO) and human liver cancer (HepG2) cells.
Claims (3)
1. A kind of (2)R,3S,4S) A process for producing (4-amino-2-tetradecylpyrrolidin-3-ol), (2)R,3S,4S) The structural formula of the (E) -4-amino-2-tetradecyl pyrrolidine-3-alcohol is shown in the specification
，
The method is characterized by comprising the following steps:
；
S1：(3R,4R,5R) -5-allyl-4- ((4-methoxybenzyl) oxy) -1- ((4-nitrophenyl) sulfonyl) pyrrolidine-3-ol is reacted with trifluoromethanesulfonic anhydride and then treated with benzylamine to obtain (3)S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrroleAlk-3-amines;
S2：(3S,4S,5R) -5-allyl-NAfter the-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidine-3-amine and 1-tridecene are subjected to olefin metathesis reaction, removing o-nitrobenzenesulfonyl, reducing a hydrogen double bond, removing benzyl, and then removing p-methoxybenzyl to obtain (2R,3S,4S) -4-amino-2-tetradecyl pyrrolidine-3-ol;
the S1 includes the following steps:
will (3)R,4R,5R) Dissolving-5-allyl-4- ((4-methoxybenzyl) oxy) -1- ((4-nitrophenyl) sulfonyl) pyrrolidine-3-alcohol in dichloromethane, adding pyridine, dropwise adding trifluoromethanesulfonic anhydride at-20 ℃ under the protection of nitrogen, reacting at-20 ℃ for 20min, washing with hydrochloric acid, saturated sodium bicarbonate solution, ice water and saturated salt water, drying with anhydrous magnesium sulfate, filtering, evaporating the solvent under reduced pressure, directly dissolving the obtained crude product in benzylamine, reacting at 30-40 ℃ for 0.5-1.5h under the protection of nitrogen, and directly separating and purifying the reaction solution by silica gel column chromatography to obtain (3)S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidin-3-amine;
the S2 includes the following steps:
will (3)S,4S,5R) -5-allyl-N-benzyl-4- ((4-methoxybenzyl) oxy) -1- ((2-nitrophenyl) sulfonyl) pyrrolidin-3-amine, dissolved in dichloromethane, under nitrogen protection 1-tridecene, Grubbs2ndHeating the catalyst for reflux reaction for 7h, cooling the reaction liquid to room temperature, filtering the reaction liquid by diatomite, eluting the reaction liquid by ethyl acetate, and removing the solvent by rotary evaporation to obtain a crude olefin compound; dissolving the crude olefin compound in acetonitrile, adding cesium carbonate and thiophenol under the protection of nitrogen, reacting for 3h at 50 ℃, cooling the reaction liquid to room temperature, filtering through diatomite, eluting with dichloromethane, and removing the solvent by rotary evaporation to obtain the crude compound without o-nitrobenzenesulfonyl; the crude compound obtained is dissolved in methanol and 20% Pd (OH) is added2Reacting the/C and 10% Pd/C for 24h in a hydrogen atmosphere, filtering the reaction solution by using diatomite, and spin-drying the solvent to obtain the debenzylationCrude product of reduction product of the base; dissolving the benzyl-removed crude product of the reduction product in acetonitrile/water (1:4), adding ammonium ceric nitrate, reacting at room temperature for 3h, and directly separating and purifying the reaction solution by silica gel column chromatography to obtain (2)R,3S4S) -4-amino-2-tetradecylpyrrolidin-3-ol.
2. The compound (2) according to claim 1R,3S,4S) Application of (E) -4-amino-2-tetradecyl pyrrolidine-3-alcohol in preparing medicine for preventing and/or treating cancer.
3. Compound (2) according to claim 2R,3S,4S) The application of the-4-amino-2-tetradecyl pyrrolidine-3-ol in preparing the medicine for preventing and/or treating the cancers is characterized in that the cancers are colon cancer, lung cancer, breast cancer, liver cancer, stomach cancer, melanoma and esophageal cancer.
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