Solid-phase synthesis method of ziconotide by segment process

A technology of ziconotide and solid-phase synthesis, which is applied in the preparation methods of peptides, chemical instruments and methods, peptides, etc., can solve the problems of long synthesis cycle, many intermediate steps, complicated operations, etc., and is beneficial to industrial large-scale production. , the effect of shortening the synthesis cycle and reducing the difficulty of purification

Inactive Publication Date: 2015-10-14
JINAN KANGHE MEDICAL TECH
View PDF7 Cites 12 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Regarding the method of oxidation to form disulfide bonds in the synthesis step of ziconotide, patents CN200710301598.8, CN201110139661.9 and CN201310202216.1 all disclose the use of Cys with three different sulfhydryl protecting groups to synthesize linear peptides, and utilize the self-stabilization of the protecting groups properties, the preparation strategy of stepwise oxidation to form three pairs of disulfide bonds is adopted; although the disulfide bonds formed have good orientation, this strategy has many intermediate steps and is cumbersome to operate
Patent CN200910188686.0 adopts the preparation strategy of one-step oxidation to form three pairs of disulfide bonds at the same time. Although the process is simplified, the m

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Solid-phase synthesis method of ziconotide by segment process
  • Solid-phase synthesis method of ziconotide by segment process
  • Solid-phase synthesis method of ziconotide by segment process

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0068] Example 1: Synthesis of Fmoc-Cys(Trt)-Rink Amide Resins

[0069] Accurately weigh 20.0g of Rink Amide resin (sub=0.33mmol / g) into a synthesis column, wash twice with 160mL DMF, add 160mL DCM to swell for 30min, filter out DCM, wash twice with 160mL DMF; add 20 %piperidine / DMF solution 160ml deprotected twice, reacted for 10min and 15min respectively; then washed twice with 100ml DMF, DCM, DMF respectively; filtered off DMF, added Fmoc-Cys(Trt)-OH / DIC / HOBT Mix the DMF solution [weigh 7.73g (13.2mmol) Fmoc-Cys(Trt)-OH and 1.96g (14.52mmol) HOBT in a conical flask, add 100mL DMF solution and stir to dissolve, add 2.25 ml (14.52mmol) DIC, activated for 3min]; react for 2h, remove the reaction liquid, wash twice with 160mL DMF, add 120mL end-capping reagent (24ml acetic anhydride, 20.4ml pyridine, 75.6mL DCM) to react for 2h, filter off with suction The reaction solution was washed 3 times with DMF, DCM and methanol respectively, and after vacuum drying, 22.06g of Fmoc-Cys...

Embodiment 2

[0070] Example 2: Synthesis of Fmoc-Cys(Trt)-Rink Amide-AM Resins

[0071] Accurately weigh 20.0g of Rink Amide-AM resin (sub=0.32mmol / g) into the synthesis column, wash twice with 160mL DMF, add 160mL DCM to swell for 30min, filter out the DCM, and wash twice with 160mL DMF; Add 160ml of 20% piperidine / DMF solution for deprotection twice, react for 10min and 15min respectively; then wash twice with 100ml DMF, DCM, DMF respectively; filter off DMF, add Fmoc-Cys(Trt)-OH / DIC / Mixed DMF solution of HOBT [weigh 3.75g (6.4mmol) Fmoc-Cys(Trt)-OH and 0.95g (7.04mmol) HOBT in a conical flask, add 100mL DMF solution and stir to dissolve, at low temperature (0°C) Add 1.09ml (7.04mmol) DIC, activate for 3min]; react for 2h, remove the reaction solution, wash twice with 160mL DMF, add 120mL end-capping reagent (24ml acetic anhydride, 20.4ml pyridine, 75.6mL DCM) to react for 2h, pump out The reaction solution was filtered off, washed three times with DMF, DCM, and methanol, and dried i...

Embodiment 3

[0072] Example 3: Synthesis of Fmoc-Gly-CTC Resins

[0073] Weigh 70.0g (sub=1.00mmol / g) of CTC resin and place it in a synthesis column, wash twice with 420mL DMF, add 420mL DCM to swell for 30min; after filtering off DCM, add 41.62g of Fmoc-Gly-OH DCM solution 250ml, add DIPEA 25.45ml at low temperature (0°C), react for 60min, remove the reaction solution, add DCM / CH 3 420ml of OH / DIPEA (volume ratio 17:2:1) mixed solution was capped for 30min; then washed 3 times with DMF, DCM, and methanol respectively, and vacuum-dried to obtain 86.50g of Fmoc-Gly-CTC Resins; the degree of substitution was 0.80mmol / g.

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

PUM

No PUM Login to view more

Abstract

The invention belongs to the field of polypeptide synthesis, and relates to a solid-phase synthesis method of ziconotide by a segment process. The method aims to solve the technical problems of long synthesis period and low crude peptide purity in the existing preparation method, and the technical problems of complex operation, low total yield and the like in the disulfide-bond formation process. The technical scheme is as follows: the method comprises the following steps: synthesizing four segment peptides [19-25]A, [12-18]B, [6-11]C and [1-5]D of ziconotide by a solid-phase process; sequentially connecting the all-protected peptide fragments B, C and D to the peptide resin A, and cracking to obtain a ziconotide linear peptide; and finally, carrying out liquid-phase one-step oxidization, which has the advantages of mild conditions, simplified operation, easy after-treatment and environment friendliness, to obtain the ziconotide. The technical scheme can greatly shorten the synthesis period, lower the synthesis and purification difficulty and the production cost, and enhance the purity of the linear peptide and the total yield of the product. The method is beneficial to industrial large-scale production.

Description

technical field [0001] The invention relates to the field of polypeptide synthesis, in particular to a method for solid-phase synthesis of ziconotide by a fragment method. Background technique [0002] Ziconotide (Ziconotide), trade name Prialt, was developed by Elan Company of Ireland. It was first approved for marketing in the United States on January 11, 2005. It was approved by the European Union in February 2005. In 2007, the special group of the Joint Analgesia Conference recommended Qi Conotide as a first-line intrathecal analgesic. It is an N-type calcium channel blocker, a synthetic form of the omega-conotoxin MVIIA isolated from the conus snail mollusk, and can be used as a non-opioid analgesic drug for intrathecal administration. Ziconotide specifically and effectively binds to N-VSCC (N-type voltage-sensitive calcium channels) to block the release of pain-causing transmitters, and then realizes its pharmacological effects. It is suitable for intrathecal inject...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to view more

Application Information

Patent Timeline
no application Login to view more
IPC IPC(8): C07K14/435C07K1/06C07K1/04
CPCC07K14/43504
Inventor 张颖杨慧李同金石鑫磊王仁友
Owner JINAN KANGHE MEDICAL TECH
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap