CN103980358A - Preparation method of liraglutide - Google Patents

Preparation method of liraglutide Download PDF

Info

Publication number
CN103980358A
CN103980358A CN201410001671.XA CN201410001671A CN103980358A CN 103980358 A CN103980358 A CN 103980358A CN 201410001671 A CN201410001671 A CN 201410001671A CN 103980358 A CN103980358 A CN 103980358A
Authority
CN
China
Prior art keywords
fmoc
palmitoyl
resin
gly
otbu
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.)
Granted
Application number
CN201410001671.XA
Other languages
Chinese (zh)
Other versions
CN103980358B (en
Inventor
路杨
杨东晖
方晨
周亮
刘少华
Current Assignee (The listed assignees may be inaccurate. Google has not performed a legal analysis and makes no representation or warranty as to the accuracy of the list.)
HANGZHOU SINOPEP PHARMACEUTICAL Inc
Original Assignee
HANGZHOU SINOPEP PHARMACEUTICAL Inc
Priority date (The priority date is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the date listed.)
Filing date
Publication date
Application filed by HANGZHOU SINOPEP PHARMACEUTICAL Inc filed Critical HANGZHOU SINOPEP PHARMACEUTICAL Inc
Priority to CN201410001671.XA priority Critical patent/CN103980358B/en
Priority to PCT/CN2014/075113 priority patent/WO2015100876A1/en
Publication of CN103980358A publication Critical patent/CN103980358A/en
Application granted granted Critical
Publication of CN103980358B publication Critical patent/CN103980358B/en
Active legal-status Critical Current
Anticipated expiration legal-status Critical

Links

Classifications

    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/04—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length on carriers
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K14/00—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof
    • C07K14/435—Peptides having more than 20 amino acids; Gastrins; Somatostatins; Melanotropins; Derivatives thereof from animals; from humans
    • C07K14/575—Hormones
    • C07K14/605—Glucagons
    • C—CHEMISTRY; METALLURGY
    • C07—ORGANIC CHEMISTRY
    • C07K—PEPTIDES
    • C07K1/00—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length
    • C07K1/107—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by chemical modification of precursor peptides
    • C07K1/1072—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by chemical modification of precursor peptides by covalent attachment of residues or functional groups
    • C07K1/1077—General methods for the preparation of peptides, i.e. processes for the organic chemical preparation of peptides or proteins of any length by chemical modification of precursor peptides by covalent attachment of residues or functional groups by covalent attachment of residues other than amino acids or peptide residues, e.g. sugars, polyols, fatty acids

Landscapes

  • Chemical & Material Sciences (AREA)
  • Organic Chemistry (AREA)
  • Health & Medical Sciences (AREA)
  • Life Sciences & Earth Sciences (AREA)
  • Medicinal Chemistry (AREA)
  • Proteomics, Peptides & Aminoacids (AREA)
  • Molecular Biology (AREA)
  • Biochemistry (AREA)
  • Biophysics (AREA)
  • General Health & Medical Sciences (AREA)
  • Genetics & Genomics (AREA)
  • Analytical Chemistry (AREA)
  • Chemical Kinetics & Catalysis (AREA)
  • General Chemical & Material Sciences (AREA)
  • Medicines That Contain Protein Lipid Enzymes And Other Medicines (AREA)
  • Gastroenterology & Hepatology (AREA)
  • Zoology (AREA)
  • Toxicology (AREA)
  • Endocrinology (AREA)
  • Peptides Or Proteins (AREA)

Abstract

The invention relates to a synthesis technology of liraglutide, and solves the problems of long synthesis period, high cost, low yield and a large amount of impurity in the prior art. The method provided by the invention comprises the specific steps of: A) synthesizing a Fmoc-Lys-(Glu(N alpha-Palmitoyl)-OtBu)-OH fragment by a liquid phase; B) in the presence of an activating agent system, coupling a resin solid phase carrier and Fmoc-Gly-OH to obtain Fmoc-Gly-resin; C) successively coupling amino acids with N-terminal Fmoc protection and side chain protection according to peptide sequence of liraglutide backbone by a solid phase synthesis method, wherein the lysine tripeptide fragment employs Fmoc-Lys-(Glu(N alpha-Palmitoyl)-OtBu)-OH; and D) cracking, purifying and freeze-drying to obtain liraglutide. The invention provides a liraglutide synthesis technology with short synthesis cycle, low cost, high yield and suitability for scale production.

Description

A kind of method of preparing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]
Technical field
The present invention relates to a kind of preparation method of polypeptide drug, is a kind of preparation method for the treatment of specifics-Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] of the synthetic long-acting type ii diabetes with glucagon-like-peptide-1 (GLP-1) receptor stimulant.
Background technology
Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], illustrious name is: Liraglutide, structural formula is as follows:
Peptide sequence is:
H-His-Ala-Glu-Gly-Thr-Phe-Thr-Ser-Asp-Val-Ser-Ser-Tyr-Leu-Glu-Gly-Gln-Ala-Ala-Lys(N-ε-(N-α-Palmitoyl-L-γ-glutamyl))-Glu-Phe-Ile-Ala-Trp-Leu-Val-Arg-Gly-Arg-Gly-OH
Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] be first that developed by Novo Nordisk Co.,Ltd of Denmark be also at present unique one long-acting gLP-1 analogue, there is the effect of GLP-1 receptor stimulant, similar to GLP-1 at aspects such as molecular structure, biological activity, action target spot and immunogenicities.The homology of the molecular structure of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] and GLP-1 (7-37) reaches 97%, and textural difference shows Lys 34substituted Lys by Arg 26via glutamate-induced generation palmitoylation; fatty acid side chain can make Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] reversibly be combined with albumin in blood night; extended the action time of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]; and strengthen the opposing to DPP-4 enzyme liberating; fatty acid side chain can also make Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] molecule, and in injection site, selfing is unified into heptamer, thereby delays it from subcutaneous attraction, is its action time and approaches 24 hours; inject once and can inject at any time every day, and hypoglycemia occurrence risk is little.In addition, this product can also reduce the secretion of hyperglycemic-glycogenolytic factor in blood sugar dependency mode, and postpones stomach emptying.
The Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] of Novo Nordisk is prepared by biological methods such as genetically engineereds, and technical difficulty is large, and production cost is high, is unfavorable for the scale operation of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37].US6268343B1 and US6458924B2 have reported the solid-liquid synthesis method of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], and intermediate GLP-1 (7-37)-OH all needs reversed-phase HPLC purifying, then under liquid-phase condition with N α-Palmitoyl-Glu (OSu)-OtBu reaction, this method needs twice purifying, and synthesis cycle is long, and waste liquid is many, and cost costliness is unfavorable for the shortcoming of scale operation.
WO2013037266A1 discloses a kind of preparation method of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]; concrete steps are: by Fmoc solid-phase synthesis; there is the amino acid of the Fmoc protection of N end and side chain protected according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order successively coupling; wherein Methionin adopts Fmoc-Lys (Alloc)-OH; slough Alloc; by solid-phase synthesis coupling Palmitoyl-Glu-Offiu on lysine side-chain amino, after cracking, obtain product.This method, owing to using tetrakis triphenylphosphine palladium to slough Alloc, not only makes high expensive, is unfavorable for scale operation, also can make metal residual cause heavy metal content to exceed standard, and causes quality product and content not high.
In sum, in the solid phase synthesis process of existing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], because synthesis cycle is long, cost is high, and yield is low, and impurity is many, is not suitable for suitability for industrialized production.
The inventor uses existing synthetic method, prepares Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], finds that synthesis step is more, and synthesis cycle is long, and purity and yield are not high, are unsuitable for commercial scale production.For this reason, the inventor is studied the synthetic method of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], thereby has obtained technical scheme of the present invention.
Summary of the invention
The object of this invention is to provide a kind of solid phase synthesis process of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37].The technical issues that need to address of the present invention are: synthesis cycle is long, and cost is high, and yield is low, and impurity is many, are not suitable for suitability for industrialized production.
Synthetic route of the present invention is as shown in Figure 1: first by liquid phase process synthetic lysine tripeptide fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH; secondly under the existence of activator system; obtain Fmoc-Gly-resin by resin solid phase carrier and Fmoc-Gly-OH coupling; then pass through solid-phase synthesis; the amino acid according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order successively coupling with the Fmoc protection of N end and side chain protected, wherein Methionin tripeptide fragment adopts Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, last cracking, purifying, freeze-drying, obtains Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37].
In the present invention, some conventional abbreviations have following implication;
Fmoc: fluorenylmethyloxycarbonyl
Fmoc-AA: the amino acid of fluorenylmethyloxycarbonyl protection
DIC: N, N '-di-isopropyl carbodiimide
DCC: N, N '-dicyclohexylcarbodiimide
PyBOP: phosphofluoric acid benzotriazole-1-base-oxygen base tripyrrole alkyl phosphorus
HATU: 2-(7-azo benzotriazole)-N, N, N', N'-tetramethyl-urea phosphofluoric acid ester
HOBt: 1-hydroxybenzene a pair of horses going side by side triazole
HOSu: N-hydroxy-succinamide
TBu: the tertiary butyl
Trt: trityl
Boc: tertbutyloxycarbonyl
Palmitoyl: palmitoyl
Pbf: 2,2,4,6,7-pentamethyl-Dihydrobenzofuranes-5-alkylsulfonyl
Tyr: tyrosine
Ile: Isoleucine
Gln: glutamine
Asn: l-asparagine
Cys: halfcystine
Pro: proline(Pro)
Leu: leucine
Gly: glycine
Arg: arginine
Val: α-amino-isovaleric acid
Trp: tryptophane
Ala: L-Ala
Phe: phenylalanine
Glu: L-glutamic acid
Lys: Methionin
Ser: Serine
Asp: aspartic acid
Thr: Threonine
His: Histidine
DMF: N, N '-dimethyl formamide
MeOH: methyl alcohol
DCM: methylene dichloride
NMP: N-Methyl pyrrolidone
DMSO: dimethyl sulfoxide (DMSO)
TFA: trifluoracetic acid
EDT: dithioglycol
Piperidine: hexahydropyridine
DMAP:4-Dimethylamino pyridine
DIEA: N, N '-diisopropylethylamine
TMP: 2，4，6-trimethylpyridine.
The invention provides a kind of synthetic method of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], its step is as follows for this reason:
Step 1, by liquid phase process synthetic lysine tripeptide fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH;
Step 2, under the existence of activator system, obtains Fmoc-Gly-resin by resin solid phase carrier and Fmoc-Gly-OH coupling;
Step 3, by solid-phase synthesis, has the amino acid of the Fmoc protection of N end and side chain protected according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order successively coupling, wherein Methionin tripeptide fragment adopts Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH;
Step 4, cracking, purifying, freeze-drying, obtains Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37].
Wherein, the solid phase synthesis process described in step 1, described fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu) liquid phase of-OH synthesizes: Palmiticacid, HOSu, DCC coupling obtain Palmitoyl-OSu activation fat, and then reaction obtains two peptide fragment Palmitoyl-Glu-OtBu with H-Glu-OtBu; Palmitoyl-Glu-OtBu, HOSu, DCC coupling obtain Palmitoyl-Glu (OSu)-OtBu and activate fat, and then reaction obtains Methionin tripeptide fragment Fmoc-Lys-(Glu (N with Fmoc-Lys-OH α-Palmitoyl)-OtBu)-OH.
Wherein, the solid phase synthesis process described in step 2, described resin solid carrier adopts 2-CTC resin, and described activator system is selected from DIEA, TMP or NMM, the Fmoc-Gly-CTC resin that described Fmoc-Gly-resin is 0.10 ~ 0.35mmol/g substitution value.
Wherein, the solid phase synthesis process described in step 2, described resin solid carrier adopts king's resin, and described activator system is made up of DIC, HOBt and DMAP, Fmoc-Gly-king's resin that described Fmoc-Gly-king's resin is 0.10 ~ 0.35mmol/g substitution value.
Wherein, the solid phase synthesis process described in step 3,
1) what the piperidines that employing is 1:4 by volume ratio and DMF formed goes to protect liquid to remove the Fmoc protecting group on Fmoc-Gly-resin, obtains H-Gly-resin;
2), under the existence of coupling agent system, the arginine coupling of H-Gly-resin and Fmoc protection and side chain protected obtains Fmoc-Arg (Pbf)-Gly-resin;
3) repeating step 1), 2), carry out successively amino acid whose coupling according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order, wherein Methionin adopts Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, coupling amino acid order is:
Fmoc-Gly-OH、Fmoc-Arg(Pbf)-OH?、Fmoc-Val-OH、Fmoc-Leu-OH、Fmoc-Trp(Boc)-OH、Fmoc-Ala-OH、Fmoc-Ile-OH、Fmoc-Phe-OH、Fmoc-Glu(OtBu)-OH、Fmoc-?Lys-(Glu(N α-Palmitoyl)-OtBu)-OH、Fmoc-Ala-OH、Fmoc-Ala-OH、Fmoc-Gln(Trt)-OH、Fmoc-Gly-OH、Fmoc-Glu(OtBu)-OH、Fmoc-Leu-OH、Fmoc-Tyr(tBu)-OH、Fmoc-Ser(tBu)-OH、Fmoc-Ser(tBu)-OH、Fmoc-Val-OH、Fmoc-Asp(OtBu)-OH、Fmoc-Ser(tBu)-OH、Fmoc-Thr(tBu)-OH、Fmoc-Phe-OH、Fmoc-Thr(tBu)-OH、Fmoc-Gly-OH、Fmoc-Glu(OtBu)-OH、Fmoc-Ala?-OH、Boc-His(Trt)-OH；
Described coupling agent system comprises condensing agent and reaction solvent, and described condensing agent is selected from DIC/HOBt, PyBOP/HOBt/DIEA or HATU/HOBt/DIEA; Described reaction solvent is selected from DMF, DCM, NMP, DMSO or the arbitrary combination between them.
Method of the present invention obtains through screening, and screening process is as follows:
1) selection of mol ratio: H-Glu (OtBu)-Phe-Ile-Ala-Trp (Boc)-Leu-Val-Arg (Pbf)-Gly-Arg (Pbf)-Gly-resin: Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu) mol ratio of-OH:HATU:HOBt:DIEA is: 1:3:3:3:3 and 1:5:5:5:5;
2) selection of temperature of reaction:
25 oc and 35 oc
3) selection in reaction times:
Reaction times is: 2 hours and 3 hours.
8 kinds of experiment conditions have been proposed for this reason:
Experiment condition 1: get 3.43g (1.0mmol) H-Glu (OtBu)-Phe-Ile-Ala-Trp (Boc)-Leu-Val-Arg (Pbf)-Gly-Arg (Pbf)-Gly-resin, 2.16g (3.0 mmol) Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, 0.41g (3.0 mmol) HOBt and 1.14g (3.0 mmol) HATU add stirring and dissolving in 20ml DMF, is cooled to 0 oc, adds 0.5ml (3.0 mmol) DIEA in above-mentioned solution, 25 oc reaction 2 hours, then the remaining amino acid of coupling successively, coupling amino acid order is: Fmoc-Ala-OH, Fmoc-Ala-OH, Fmoc-Gln (Trt)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Leu-OH, Fmoc-Tyr (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Val-OH, Fmoc-Asp (OtBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Thr (tBu)-OH, Fmoc-Phe-OH, Fmoc-Thr (tBu)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Ala-OH, Boc-His (Trt)-OH, cracking, purifying, freeze-drying, obtain Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] essence peptide,
Experiment condition 2-8, experimental implementation is as shown in experiment condition 1, and different experiment condition and experimental result thereof are as follows:
Experiment condition Mol ratio Temperature Time Total recovery Purity
Experiment condition 1 1:3:3:3 25℃ 2 hours 23% 99.10%
Experiment condition 2 1:5:5:5 25℃ 2 hours 27% 99.11%
Experiment condition 3 1:3:3:3 35℃ 2 hours 27% 99.23%
Experiment condition 4 1:5:5:5 35℃ 2 hours 31% 99.75%
Experiment condition 5 1:3:3:3 25℃ 3 hours 28% 99.54%
Experiment condition 6 1:5:5:5 25℃ 3 hours 29% 99.64%
Experiment condition 7 1:3:3:3 35℃ 3 hours 28% 99.13%
Experiment condition 8 1:5:5:5 35℃ 3 hours 26% 99.26%
Above result shows, the purification effect optimum of experiment condition 4.
Compared to the prior art method of the present invention has obvious advantage, and relevant contrast experiment is as follows:
Patent Total recovery Purity
The technology of the present invention 31% 99.75%
WO2013037266A1 15.87% 99.17%
US6458924B2 22% NA
The invention has the beneficial effects as follows: select fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu) the direct solid phase synthesis Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] of-OH, the synthesis cycle that has solved prior art existence is long, and cost is high, and purity is low, and impurity is many, is not suitable for the problem of suitability for industrialized production; The invention provides that a kind of synthesis cycle is short, cost is low, yield is higher, be applicable to the synthesis technique of the Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] of large-scale production.
Brief description of the drawings
Fig. 1 synthetic route of the present invention;
The HPLC spectrogram of Fig. 2 Methionin tripeptide fragment;
The HPLC spectrogram of the thick peptide of Fig. 3 Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37];
The HPLC spectrogram of Fig. 4 Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] essence peptide;
Fig. 5 embodiment 12 Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] essence peptide mass spectrograms.
Embodiment
Further illustrate by the following examples the present invention.
Embodiment mono-: Palmitoyl-OSu activates the synthetic of fat
Take 256.42g Palmiticacid (1.0mol), 138.10g HOSu(1.2mol) add in 2000ml THF, under ice-water bath, add 247.56g DCC(1.2mol), react 1 hour, be warmed up to room temperature reaction 3 hours, reacting liquid filtering, mother liquor is spin-dried for, and adds DCM and dissolves, and filters, saturated sodium bicarbonate is washed 3 times, pure water 2 times, back extraction 2 times, merges organic phase, dried over anhydrous sodium carbonate, be spin-dried for, 0-5 DEG C of ice ethyl alcohol recrystallization 3 times, filters, solid oil pump draws the dry 314.62g of obtaining Palmitoyl-OSu activation fat, yield 89%.
Embodiment bis-: Palmitoyl-Glu-OtBu's is synthetic
Take 101.62g H-Glu-OtBu(0.5mol) and 79.50g Na 2cO 3(0.75mol) join in the mixing solutions of 500ml water and 500ml THF and dissolve, take 176.75g Palmitoyl-OSu(0.5mol) join 500ml THF, after dissolving, drip in above-mentioned mixing solutions, under room temperature, reaction is spent the night, with 10% dilute hydrochloric acid adjusting PH to 7, revolve to steam and remove THF, regulate afterwards PH to 3.Obtain a large amount of white precipitates, filter.By the 0-5 DEG C of ice ethyl alcohol recrystallization for white precipitate obtaining.Solid oil pump draws the dry 192.11g of obtaining Palmitoyl-Glu-OtBu, yield 87%.
Embodiment tri-: Palmitoyl-Glu (OSu)-OtBu's is synthetic
Take 88.33g Palmitoyl-Glu-OtBu(0.2mol), 27.62g HOSu(0.24mol) add in 1000ml THF, under ice-water bath, add 49.51g DCC(0.24mol), react 1 hour, be warmed up to room temperature reaction 3 hours, reacting liquid filtering, mother liquor is spin-dried for, adding DCM dissolves, filter, saturated sodium bicarbonate is washed 3 times, pure water 2 times, back extraction 2 times, merge organic phase, dried over anhydrous sodium carbonate, be spin-dried for, 0-5 DEG C of ice ethyl alcohol recrystallization 3 times, filter, solid oil pump draws the dry 94.81g of obtaining Palmitoyl-Glu (OSu)-OtBu to activate fat, yield 88%.
Embodiment tetra-: Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH synthetic
Take 36.74g Fmoc-Lys-OH(0.1mol) and 15.90g Na 2cO 3(0.15mol) join in the mixing solutions of 100ml water and 100ml THF and dissolve, take 53.87g Palmitoyl-Glu (OSu)-OtBu(0.1mol) join 100ml THF, after dissolving, drip in above-mentioned mixing solutions, under room temperature, reaction is spent the night, with 10% dilute hydrochloric acid adjusting PH to 7, revolve to steam and remove THF, regulate afterwards PH to 3.Obtain a large amount of white precipitates, filter.By the 0-5 DEG C of ice ethyl alcohol recrystallization for white precipitate obtaining.Solid oil pump draws the dry 67.24g of obtaining Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, HPLC purity is 97.40%, yield 85%.
Embodiment five: the Fmoc-Gly-CTC resin that substitution value is 0.10mmol/g synthetic
Taking substitution value is the 2-CTC resin 20g of 0.4mmol/g, join in solid state reaction post, join in solid state reaction post, with DMF washing 1 time,, after 30 minutes, get 13.37g Fmoc-Gly-OH DMF and dissolve with the swelling resin of DMF, under ice-water bath, add after 7.5ml DIEA activation, add in the above-mentioned reaction column that resin is housed, react after 2 hours, add 100ml anhydrous methanol sealing 1 hour.With DMF washing 3 times, DCM washing 3 times, with anhydrous methanol sealing 30 minutes, methyl alcohol shrank dry, obtains 22.34g Fmoc-Gly-CTC resin, and detection substitution degree is 0.10mmol/g.
Embodiment six: the Fmoc-Gly-CTC resin that substitution value is 0.25mmol/g synthetic
Taking substitution value is the 2-CTC resin 10g of 0.95mmol/g, join in solid state reaction post, join in solid state reaction post, with DMF washing 1 time,, after 30 minutes, get 14.11g Fmoc-Gly-OH DMF and dissolve with the swelling resin of DMF, under ice-water bath, add after 8.0ml DIEA activation, add in the above-mentioned reaction column that resin is housed, react after 2 hours, add 100ml anhydrous methanol sealing 1 hour.With DMF washing 3 times, DCM washing 3 times, with anhydrous methanol sealing 30 minutes, methyl alcohol shrank dry, obtains Fmoc-Gly-CTC resin, and detection substitution degree is 0.25mmol/g.
Embodiment seven: Fmoc-Gly-king's resin that substitution value is 0.10mmol/g synthetic
Taking substitution value is king's resin 20g of 0.45mmol/g, join in solid state reaction post, join in solid state reaction post, with DMF washing 1 time, with the swelling resin of DMF after 30 minutes, get 13.37g Fmoc-Gly-OH, 6.01g HOBt dissolves with DMF, under ice-water bath, add after 6.0ml DIC activation, add in the above-mentioned reaction column that resin is housed, after 5 minutes, add 2.75g DMAP, react after 2 hours, with DMF washing 3 times, DCM washing 3 times, spend the night with 100ml acetic anhydride/pyridine end-blocking, methyl alcohol shrinks dry, obtain Fmoc-Gly-king's resin, detection substitution degree is 0.10mmol/g.
Embodiment eight: Fmoc-Gly-king's resin that substitution value is 0.25mmol/g synthetic
Taking substitution value is king's resin 20g of 0.75mmol/g, join in solid state reaction post, join in solid state reaction post, with DMF washing 1 time, with the swelling resin of DMF after 30 minutes, get 22.28g Fmoc-Gly-OH, 10.13g HOBt dissolves with DMF, under ice-water bath, add after 8.0ml DIC activation, add in the above-mentioned reaction column that resin is housed, after 5 minutes, add 4.5g DMAP, react after 2 hours, with DMF washing 3 times, DCM washing 3 times, spend the night with 100ml acetic anhydride/pyridine end-blocking, methyl alcohol shrinks dry, obtain 22.54g Fmoc-Gly-king resin, detection substitution degree is 0.25mmol/g.
Embodiment nine: the preparation of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] CTC resin
Take the Fmoc-Gly-CTC resin that 4.46g (1mmol) substitution value is 0.10mmol/g, add in solid state reaction post, with DMF washing 1 time, with the swelling Fmoc-Gly-CTC resin of DMF after 30 minutes, with DMF: the mixing solutions that pyridine volume ratio is 4:1 is sloughed Fmoc protection, then with DMF washing 6 times, take 3.24g Fmoc-Arg (Pbf)-OH(5mmol), 0.68g HOBt(5mmol) to add volume ratio be DCM and the DMF mixing solutions of 1:1, under ice-water bath, add 0.8ml DIC(10mmol) activation after, add in the above-mentioned reaction column that resin is housed, under room temperature, react after 2 hours, detect and judge reaction end with ninhydrin method, if resin water white transparency, represent to react completely, resin colour developing, represents reaction not exclusively, need to react 1 hour again, and this judging criterion is applicable to detect and judge reaction end with ninhydrin method in follow-up amino acid coupling.Repeat the step that the above-mentioned Fmoc of removing protected and added corresponding amino acid coupling, according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order, complete successively Fmoc-Gly-OH, Fmoc-Arg (Pbf)-OH, Fmoc-Val-OH, Fmoc-Leu-OH, Fmoc-Trp (Boc)-OH, Fmoc-Ala-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, Fmoc-Ala-OH, Fmoc-Ala-OH, Fmoc-Gln (Trt)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Leu-OH, Fmoc-Tyr (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Val-OH, Fmoc-Asp (OtBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Thr (tBu)-OH, Fmoc-Phe-OH, Fmoc-Thr (tBu)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Ala-OH, the coupling of Boc-His (Trt)-OH.Wherein solvent is changed to when Fmoc-Leu-OH and Fmoc-Phe-OH coupling: selecting volume ratio is DMSO and the DMF mixing solutions of 1:4; When Fmoc-Asp (OtBu)-OH coupling, coupling reagent is changed to: PyBOP/HOBt/DIEA; When Boc-His (Trt)-OH coupling, coupling reagent is changed to: HATU/HOBt/DIEA, and coupling is complete, by DMF washing 3 times for Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] CTC resin, DCM washing 3 times, MeOH washing 3 times, DCM washing 3 times, MeOH washing 3 times, drains and obtains 9.67g Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] CTC resin.
Embodiment ten: the preparation of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] king resin
Take 4.57g(1mmol) the substitution value Fmoc-Gly-king's resin that is 0.10mmol/g, add in solid state reaction post, with DMF washing 1 time, with the swelling Fmoc-Gly-king's resin of DMF after 30 minutes, with DMF: the mixing solutions that pyridine volume ratio is 4:1 is sloughed Fmoc protection, then with DMF washing 6 times, take 3.24g Fmoc-Arg (Pbf)-OH(5mmol), 0.68g HOBt(5mmol) to add volume ratio be DCM and the DMF mixing solutions of 1:1, under ice-water bath, add 0.8ml DIC(10mmol) activation after, add in the above-mentioned reaction column that resin is housed, under room temperature, react after 2 hours, detect and judge reaction end with ninhydrin method, if resin water white transparency, represent to react completely, resin colour developing, represents reaction not exclusively, need to react 1 hour again, and this judging criterion is applicable to detect and judge reaction end with ninhydrin method in follow-up amino acid coupling.Repeat the step that the above-mentioned Fmoc of removing protected and added corresponding amino acid coupling, according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order, complete successively Fmoc-Gly-OH, Fmoc-Arg (Pbf)-OH, Fmoc-Val-OH, Fmoc-Leu-OH, Fmoc-Trp (Boc)-OH, Fmoc-Ala-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, Fmoc-Ala-OH, Fmoc-Ala-OH, Fmoc-Gln (Trt)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Leu-OH, Fmoc-Tyr (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Val-OH, Fmoc-Asp (OtBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Thr (tBu)-OH, Fmoc-Phe-OH, Fmoc-Thr (tBu)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Ala-OH, the coupling of Boc-His (Trt)-OH.
Wherein solvent is changed to when Fmoc-Leu-OH and Fmoc-Phe-OH coupling: selecting volume ratio is DMSO and the DMF mixing solutions of 1:4; When Fmoc-Asp (OtBu)-OH coupling, coupling reagent is changed to: PyBOP/HOBt/DIEA; When Boc-His (Trt)-OH coupling, coupling reagent is changed to: HATU/HOBt/DIEA, and coupling is complete, by DMF washing 3 times for Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] CTC resin, DCM washing 3 times, MeOH washing 3 times, DCM washing 3 times, MeOH washing 3 times, drains and obtains 9.78g Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] king resin.
Embodiment 11: the mass-producing preparation of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] king resin
Take 4570g(1mol) the substitution value Fmoc-Gly-king's resin that is 0.10mmol/g, add in solid state reaction post, with DMF washing 1 time, with the swelling Fmoc-Gly-king's resin of DMF after 30 minutes, with DMF: the mixing solutions that pyridine volume ratio is 4:1 is sloughed Fmoc protection, then with DMF washing 6 times, take 3240g Fmoc-Arg (Pbf)-OH(5mol), 682g HOBt(5mol) to add volume ratio be DCM and the DMF mixing solutions of 1:1, under ice-water bath, add 800ml DIC(5mol) activation after, add in the above-mentioned reaction column that resin is housed, under room temperature, react after 2 hours, detect and judge reaction end with ninhydrin method, if resin water white transparency, represent to react completely, resin colour developing, represents reaction not exclusively, need to react 1 hour again, and this judging criterion is applicable to detect and judge reaction end with ninhydrin method in follow-up amino acid coupling.Repeat the step that the above-mentioned Fmoc of removing protected and added corresponding amino acid coupling, according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order, complete successively Fmoc-Gly-OH, Fmoc-Arg (Pbf)-OH, Fmoc-Val-OH, Fmoc-Leu-OH, Fmoc-Trp (Boc)-OH, Fmoc-Ala-OH, Fmoc-Ile-OH, Fmoc-Phe-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH, Fmoc-Ala-OH, Fmoc-Ala-OH, Fmoc-Gln (Trt)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Leu-OH, Fmoc-Tyr (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Val-OH, Fmoc-Asp (OtBu)-OH, Fmoc-Ser (tBu)-OH, Fmoc-Thr (tBu)-OH, Fmoc-Phe-OH, Fmoc-Thr (tBu)-OH, Fmoc-Gly-OH, Fmoc-Glu (OtBu)-OH, Fmoc-Ala-OH, the coupling of Boc-His (Trt)-OH.Wherein solvent is changed to when Fmoc-Leu-OH and Fmoc-Phe-OH coupling: selecting volume ratio is DMSO and the DMF mixing solutions of 1:4; When Fmoc-Asp (OtBu)-OH coupling, coupling reagent is changed to: PyBOP/HOBt/DIEA; When Boc-His (Trt)-OH coupling, coupling reagent is changed to: HATU/HOBt/DIEA, and coupling is complete, by DMF washing 3 times for Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] CTC resin, DCM washing 3 times, MeOH washing 3 times, DCM washing 3 times, MeOH washing 3 times, drains and obtains 9795g Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] king resin.
Embodiment 12: the preparation of the thick peptide of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]
Take Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] CTC resin or the Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] king resin of 100g full guard, join in three mouthfuls of round-bottomed flasks of 1000mL, press TFA: thioanisole: methyl-phenoxide: the volume ratio configuration lysate 10L of EDT=90:5:3:2, lysate is added in above-mentioned resin, room temperature reaction 2 hours, filter, with the resin after a small amount of TFA washing cracking 3 times, merging filtrate, concentrated, liquid after concentrated is joined in ice ether and precipitates 1 hour, centrifugal, anhydrous diethyl ether centrifuge washing 6 times, vacuum-drying, obtain the thick peptide 34.13g of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], HPLC purity 83.03%, thick peptide yield 78%.
Embodiment 13: the preparation of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] essence peptide acetate
After the mixing solutions 30L that takes the thick peptide of 3413g Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] 50% acetonitrile+50% water dissolves, obtain target product by C18 or 2 purifying of C8 post, after turning salt, lyophilize.Purification condition for the first time: moving phase is: A phase: 0.1%TFA; B phase: acetonitrile, detect wavelength 220nm, collect object peak cut.Purification condition for the second time: moving phase is: A phase: 0.3% acetic acid; B phase: acetonitrile.Detect wavelength 220nm, collect object peak cut.Turn salt condition: moving phase: A phase: 20mM ammonium acetate-aqueous solution; B phase: acetonitrile; Detect wavelength 220nm.Collect object peak cut, concentrated by rotary evaporation, freeze-drying obtains Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] acetate essence peptide 11.24g, HPLC purity 99.75%, purifying total recovery 40%, total recovery 31%.
Above content is to select embodiment further description made for the present invention in conjunction with concrete repairing, and can not assert that specific embodiment of the invention is confined to these explanations.For general technical staff of the technical field of the invention, without departing from the inventive concept of the premise, can also make some simple deduction or replace, all should be considered as belonging to insured's scope of the present invention.

Claims (6)

1. a synthetic method for Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37], described method steps is as follows:
Step 1, by liquid phase process synthetic lysine tripeptide fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH;
Step 2, under the existence of activator system, obtains Fmoc-Gly-resin by resin solid phase carrier and Fmoc-Gly-OH coupling;
Step 3, by solid-phase synthesis, has the amino acid of the Fmoc protection of N end and side chain protected according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order successively coupling, wherein Methionin tripeptide fragment adopts Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH;
Step 4, cracking, purifying, freeze-drying, obtains Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37].
2. method according to claim 1, is characterized in that:
Wherein, the solid phase synthesis process described in step 1, described fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu) liquid phase of-OH synthesizes: Palmiticacid, HOSu and DCC coupling obtain Palmitoyl-OSu, and then Palmitoyl-OSu and H-Glu-OtBu reaction obtain two peptide fragment Palmitoyl-Glu-OtBu; Palmitoyl-Glu-OtBu, HOSu and DCC coupling obtain Palmitoyl-Glu (OSu)-OtBu, and then Palmitoyl-Glu (OSu)-OtBu and Fmoc-Lys-OH reaction obtain Methionin tripeptide fragment Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH.
3. method according to claim 1, is characterized in that:
Wherein, the solid phase synthesis process described in step 2, described resin solid carrier adopts 2-CTC resin, and described activator system is selected from DIEA, TMP or NMM, the Fmoc-Gly-CTC resin that described Fmoc-Gly-resin is 0.10 ~ 0.35mmol/g substitution value.
4. method according to claim 1, is characterized in that:
Wherein, the solid phase synthesis process described in step 2, described resin solid carrier adopts king's resin, and described activator system is made up of DIC, HOBt and DMAP, Fmoc-Gly-king's resin that described Fmoc-Gly-king's resin is 0.10 ~ 0.35mmol/g substitution value.
5. method according to claim 1, is characterized in that:
Wherein, the solid phase synthesis process described in step 3,1) adopt going that the piperidines that is 1:4 by volume ratio and DMF form to protect liquid to remove the Fmoc protecting group on Fmoc-Gly-resin, obtain H-Gly-resin;
2), under the existence of coupling agent system, the arginine coupling of H-Gly-resin and Fmoc protection and side chain protected obtains Fmoc-Arg (Pbf)-Gly-resin;
3) repeating step 1), 2), carry out successively amino acid whose coupling according to Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] main chain peptide order, wherein Methionin tripeptide fragment adopts Fmoc-Lys-(Glu (N α-Palmitoyl)-OtBu)-OH.
6. method according to claim 5, is characterized in that:
Described coupling agent system comprises condensing agent and reaction solvent, and described condensing agent is selected from DIC/HOBt, PyBOP/HOBt/DIEA or HATU/HOBt/DIEA; Described reaction solvent is selected from DMF, DCM, NMP, DMSO or the arbitrary combination between them.
CN201410001671.XA 2014-01-03 2014-01-03 A kind of method preparing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37] Active CN103980358B (en)

Priority Applications (2)

Application Number Priority Date Filing Date Title
CN201410001671.XA CN103980358B (en) 2014-01-03 2014-01-03 A kind of method preparing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]
PCT/CN2014/075113 WO2015100876A1 (en) 2014-01-03 2014-04-10 Method for preparing liraglutide

Applications Claiming Priority (1)

Application Number Priority Date Filing Date Title
CN201410001671.XA CN103980358B (en) 2014-01-03 2014-01-03 A kind of method preparing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]

Publications (2)

Publication Number Publication Date
CN103980358A true CN103980358A (en) 2014-08-13
CN103980358B CN103980358B (en) 2016-08-31

Family

ID=51272546

Family Applications (1)

Application Number Title Priority Date Filing Date
CN201410001671.XA Active CN103980358B (en) 2014-01-03 2014-01-03 A kind of method preparing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]

Country Status (2)

Country Link
CN (1) CN103980358B (en)
WO (1) WO2015100876A1 (en)

Cited By (14)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104356224A (en) * 2014-10-24 2015-02-18 杭州阿德莱诺泰制药技术有限公司 Preparation method of semaglutide
CN104497130A (en) * 2014-12-30 2015-04-08 杭州华津药业股份有限公司 Preparation method for lanreotide
CN105753964A (en) * 2014-12-16 2016-07-13 深圳翰宇药业股份有限公司 Preparation method of semaglutide and intermediate of semaglutide
WO2017162650A1 (en) 2016-03-23 2017-09-28 Bachem Holding Ag Method for preparing glucagon-like peptides
CN108026153A (en) * 2015-07-10 2018-05-11 赛诺菲 The alternatively new exendin-4 derivative of the dual GLP-1/ glucagon receptors activator of property peptide
CN110615836A (en) * 2018-06-20 2019-12-27 鲁南制药集团股份有限公司 Solid-phase synthesis method of liraglutide
CN110835369A (en) * 2019-12-02 2020-02-25 苏州天马医药集团天吉生物制药有限公司 Method for synthesizing liraglutide
CN110845600A (en) * 2018-08-21 2020-02-28 鲁南制药集团股份有限公司 Method for preparing liraglutide
WO2020182227A1 (en) * 2019-03-14 2020-09-17 美药星（南京）制药有限公司 Preparation method for high purity liraglutide side chain
CN111892650A (en) * 2020-07-06 2020-11-06 辽宁药联制药有限公司 Solid-phase synthesis method of liraglutide
WO2021007703A1 (en) * 2019-07-12 2021-01-21 Shanghai Space Peptides Pharmaceutical Co., Ltd. A method for preparing liraglutide via a solid phase peptide synthesis
WO2021017793A1 (en) * 2019-07-27 2021-02-04 深圳市健元医药科技有限公司 Method for preparing chemically synthesized acidic polypeptide
CN113748125A (en) * 2019-03-19 2021-12-03 恩泽生物科学有限公司 Glucagon-like peptide-1 (GLP-1) receptor agonists and analogs thereof
CN116730902A (en) * 2023-08-07 2023-09-12 杭州湃肽生化科技有限公司 Method for synthesizing liraglutide

Families Citing this family (16)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
UA116217C2 (en) 2012-10-09 2018-02-26 Санофі Exendin-4 derivatives as dual glp1/glucagon agonists
KR20150096684A (en) 2012-12-21 2015-08-25 사노피 Exendin-4 derivatives
EP3080149A1 (en) 2013-12-13 2016-10-19 Sanofi Dual glp-1/glucagon receptor agonists
WO2015086730A1 (en) 2013-12-13 2015-06-18 Sanofi Non-acylated exendin-4 peptide analogues
EP3080150B1 (en) 2013-12-13 2018-08-01 Sanofi Exendin-4 peptide analogues as dual glp-1/gip receptor agonists
WO2015086729A1 (en) 2013-12-13 2015-06-18 Sanofi Dual glp-1/gip receptor agonists
TW201625668A (en) 2014-04-07 2016-07-16 賽諾菲公司 Exendin-4 derivatives as peptidic dual GLP-1/glucagon receptor agonists
TW201625670A (en) 2014-04-07 2016-07-16 賽諾菲公司 Dual GLP-1/glucagon receptor agonists derived from EXENDIN-4
TW201625669A (en) 2014-04-07 2016-07-16 賽諾菲公司 Peptidic dual GLP-1/glucagon receptor agonists derived from Exendin-4
US9932381B2 (en) 2014-06-18 2018-04-03 Sanofi Exendin-4 derivatives as selective glucagon receptor agonists
AR105319A1 (en) 2015-06-05 2017-09-27 Sanofi Sa PROPHARMS THAT INCLUDE A DUAL AGONIST GLU-1 / GLUCAGON CONJUGATE HIALURONIC ACID CONNECTOR
WO2020019128A1 (en) * 2018-07-23 2020-01-30 深圳市健元医药科技有限公司 Alanine-lysine-glutamate tripeptide derivative and use thereof
WO2020127476A1 (en) 2018-12-19 2020-06-25 Krka, D.D., Novo Mesto Pharmaceutical composition comprising glp-1 analogue
CN110759990A (en) * 2019-10-31 2020-02-07 成都圣诺生物制药有限公司 Preparation method of linagliptin
WO2021123228A1 (en) 2019-12-18 2021-06-24 Krka, D.D., Novo Mesto Pharmaceutical composition comprising glp-1 analogue
EP4345104A1 (en) 2022-09-30 2024-04-03 Bachem Holding AG Method for preparing liraglutide

Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1999043705A1 (en) * 1998-02-27 1999-09-02 Novo Nordisk A/S N-terminally truncated glp-1 derivatives
CN102286092A (en) * 2011-09-14 2011-12-21 深圳翰宇药业股份有限公司 Solid-phase synthesis method of liraglutide
CN103275208A (en) * 2013-05-27 2013-09-04 成都圣诺生物制药有限公司 Preparation method for liraglutide
CN103288951A (en) * 2013-06-19 2013-09-11 深圳翰宇药业股份有限公司 Preparation method of liraglutide

Family Cites Families (1)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN103304660B (en) * 2013-07-12 2016-08-10 上海昂博生物技术有限公司 A kind of synthetic method of Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]

Patent Citations (4)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
WO1999043705A1 (en) * 1998-02-27 1999-09-02 Novo Nordisk A/S N-terminally truncated glp-1 derivatives
CN102286092A (en) * 2011-09-14 2011-12-21 深圳翰宇药业股份有限公司 Solid-phase synthesis method of liraglutide
CN103275208A (en) * 2013-05-27 2013-09-04 成都圣诺生物制药有限公司 Preparation method for liraglutide
CN103288951A (en) * 2013-06-19 2013-09-11 深圳翰宇药业股份有限公司 Preparation method of liraglutide

Non-Patent Citations (1)

* Cited by examiner, † Cited by third party
Title
张许: "降糖药艾塞那肽和利拉鲁肽的固相合成及艾塞那肽的药效学评价", 《中国优秀硕士学位论文全文数据库 医药卫生科技辑》, 15 July 2013 (2013-07-15), pages 079 - 48 *

Cited By (19)

* Cited by examiner, † Cited by third party
Publication number Priority date Publication date Assignee Title
CN104356224A (en) * 2014-10-24 2015-02-18 杭州阿德莱诺泰制药技术有限公司 Preparation method of semaglutide
CN105753964A (en) * 2014-12-16 2016-07-13 深圳翰宇药业股份有限公司 Preparation method of semaglutide and intermediate of semaglutide
CN104497130A (en) * 2014-12-30 2015-04-08 杭州华津药业股份有限公司 Preparation method for lanreotide
CN108026153B (en) * 2015-07-10 2022-05-27 赛诺菲 Novel exendin-4 derivatives as selective peptide dual GLP-1/glucagon receptor agonists
CN108026153A (en) * 2015-07-10 2018-05-11 赛诺菲 The alternatively new exendin-4 derivative of the dual GLP-1/ glucagon receptors activator of property peptide
US11117946B2 (en) 2016-03-23 2021-09-14 Bachem Holding Ag Method for preparing glucagon-like peptides
WO2017162650A1 (en) 2016-03-23 2017-09-28 Bachem Holding Ag Method for preparing glucagon-like peptides
CN110615836B (en) * 2018-06-20 2022-09-13 鲁南制药集团股份有限公司 Solid-phase synthesis method of liraglutide
CN110615836A (en) * 2018-06-20 2019-12-27 鲁南制药集团股份有限公司 Solid-phase synthesis method of liraglutide
CN110845600A (en) * 2018-08-21 2020-02-28 鲁南制药集团股份有限公司 Method for preparing liraglutide
CN110845600B (en) * 2018-08-21 2022-12-27 鲁南制药集团股份有限公司 Method for preparing liraglutide
WO2020182227A1 (en) * 2019-03-14 2020-09-17 美药星（南京）制药有限公司 Preparation method for high purity liraglutide side chain
CN113748125A (en) * 2019-03-19 2021-12-03 恩泽生物科学有限公司 Glucagon-like peptide-1 (GLP-1) receptor agonists and analogs thereof
WO2021007703A1 (en) * 2019-07-12 2021-01-21 Shanghai Space Peptides Pharmaceutical Co., Ltd. A method for preparing liraglutide via a solid phase peptide synthesis
WO2021017793A1 (en) * 2019-07-27 2021-02-04 深圳市健元医药科技有限公司 Method for preparing chemically synthesized acidic polypeptide
CN110835369A (en) * 2019-12-02 2020-02-25 苏州天马医药集团天吉生物制药有限公司 Method for synthesizing liraglutide
CN111892650A (en) * 2020-07-06 2020-11-06 辽宁药联制药有限公司 Solid-phase synthesis method of liraglutide
CN116730902A (en) * 2023-08-07 2023-09-12 杭州湃肽生化科技有限公司 Method for synthesizing liraglutide
CN116730902B (en) * 2023-08-07 2023-11-21 杭州湃肽生化科技有限公司 Method for synthesizing liraglutide

Also Published As

Publication number Publication date
CN103980358B (en) 2016-08-31
WO2015100876A1 (en) 2015-07-09

Similar Documents

Publication Publication Date Title
CN103980358B (en) A kind of method preparing Arg34Lys26-(N-EPSILON-(N-ALPHA-Palmitoyl-L-GAMMA-glutamyl))-GLP-1[7-37]
US11518794B2 (en) Synthesis method for liraglutide with low racemate impurity
EP3398960B1 (en) Method for preparing semaglutide
EP2757107B1 (en) Method for solid phase synthesis of liraglutide
CN104356224A (en) Preparation method of semaglutide
CN102875665B (en) Method for synthesizing liraglutide
CN103497245B (en) Method for synthesizing thymalfasin
CN103848910B (en) The solid phase synthesis process of a kind of Sa Molutai
CN103864918A (en) Solid-phase synthesis method for liraglutide
CN104650219A (en) Method for preparing liraglutide by convergent synthesis
CN103819553A (en) Method for preparing lixisenatide by using solid phase and liquid phase combined technology
CN112010961A (en) Solid-liquid synthesis method of somaglutide
CN107022021A (en) A kind of solid-phase synthesis of Liraglutide
WO2020199461A1 (en) Method for synthesizing polypeptide-derived compound
CN104211801A (en) Method for preparing lixisenatide
CN106478805A (en) A kind of preparation method of GLP-1 derivant
CN110950933B (en) Solid-liquid phase synthesis method of bremelanotide
CN104356221A (en) Preparation method of pexiganan
CN110615836B (en) Solid-phase synthesis method of liraglutide
CN103992401B (en) Method for preparing exenatide
CN103159850A (en) Preparation method of tetracosactide
CN110903353B (en) Method for preparing bremelanotide by solid-liquid phase combination
CN113173987B (en) Method for synthesizing linatide
CN105713082B (en) Method for preparing lixisenatide
CN104447979A (en) Method for preparing nesiritide

Legal Events

Date Code Title Description
C06 Publication
PB01 Publication
C10 Entry into substantive examination
SE01 Entry into force of request for substantive examination
CB02 Change of applicant information

Address after: 310018 Hangzhou economic and Technological Development Zone, Zhejiang, No. 6 Avenue, No. 452

Applicant after: ADLAI NORTYE BIOPHARMA Co.,Ltd.

Address before: 310018 Hangzhou economic and Technological Development Zone, Zhejiang, No. 6 Avenue, No. 452

Applicant before: HANGZHOU ADLAI NORTYE PHARMACEUTICAL TECHNOLOGY Co.,Ltd.

Address after: 310018 Hangzhou economic and Technological Development Zone, Zhejiang, No. 6 Avenue, No. 452

Applicant after: HANGZHOU ADLAI NORTYE PHARMACEUTICAL TECHNOLOGY Co.,Ltd.

Address before: 310018 Hangzhou economic and Technological Development Zone, Zhejiang, No. 6 Avenue, No. 452

Applicant before: Hangzhou Sinopep Pharmaceutical Inc.

COR Change of bibliographic data
C14 Grant of patent or utility model
GR01 Patent grant
PE01 Entry into force of the registration of the contract for pledge of patent right
PE01 Entry into force of the registration of the contract for pledge of patent right

Denomination of invention: Preparation method of liraglutide

Effective date of registration: 20170420

Granted publication date: 20160831

Pledgee: The Bank of Nanjing Limited by Share Ltd. of Hangzhou city Small and micro businesses franchise branch

Pledgor: ADLAI NORTYE BIOPHARMA Co.,Ltd.

Registration number: 2017330000031

PC01 Cancellation of the registration of the contract for pledge of patent right
PC01 Cancellation of the registration of the contract for pledge of patent right

Date of cancellation: 20171207

Granted publication date: 20160831

Pledgee: The Bank of Nanjing Limited by Share Ltd. of Hangzhou city Small and micro businesses franchise branch

Pledgor: ADLAI NORTYE BIOPHARMA Co.,Ltd.

Registration number: 2017330000031

PE01 Entry into force of the registration of the contract for pledge of patent right
PE01 Entry into force of the registration of the contract for pledge of patent right

Denomination of invention: Preparation method of liraglutide

Effective date of registration: 20171214

Granted publication date: 20160831

Pledgee: The Bank of Nanjing Limited by Share Ltd. of Hangzhou city Small and micro businesses franchise branch

Pledgor: ADLAI NORTYE BIOPHARMA Co.,Ltd.

Registration number: 2017330000290

PC01 Cancellation of the registration of the contract for pledge of patent right
PC01 Cancellation of the registration of the contract for pledge of patent right

Date of cancellation: 20201014

Granted publication date: 20160831

Pledgee: The Bank of Nanjing Limited by Share Ltd. of Hangzhou city Small and micro businesses franchise branch

Pledgor: ADLAI NORTYE BIOPHARMA Co.,Ltd.

Registration number: 2017330000290