Method for preparing telomerase polypeptide vaccine

A technology of polypeptide vaccine and telomerase, which is applied in the preparation methods of peptides, chemical instruments and methods, peptides, etc., and can solve the problems of unprovided technical process, unlisted product purity, yield, etc.

Inactive Publication Date: 2013-01-16
NANJING UNIV OF TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there is no detailed report on the synthesis process of GV1001 at home and abroad. Chinese patent CN100376290C proposes to prepare by methods such as solid-phase synthesis, enzymatic cleavage, and recombinant expression, and lists The peptide was synthesized by a continuous flow solid-phase peptide synthesis method, activated by pentafluorophenyl ester, TBTU or DIC, and 95% TFA was used as the scavenger, but the patent only briefly introduced a general solid-phase peptide synthesis method, and did not provide Detailed technical process, important data such as product purity and yield are not listed

Method used

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  • Method for preparing telomerase polypeptide vaccine

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Experimental program
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Effect test

Embodiment 1

[0055] Such as figure 1 As shown, the preparation method claimed in the present invention comprises the following steps:

[0056] step one,

[0057] 1. Put 10 g of CTC resin with a loading capacity of 0.6 mmol / g in the reactor, add NMP to wash twice, and then use NMP to swell for 1 hour.

[0058] 2. Weigh 2.81g of Fmoc-L-Lys(Boc)-OH, add 50 ml of NMP to dissolve, add 4.18 ml of DIPEA after complete dissolution, mix well and add to the reactor to start the reaction.

[0059] 4. After 6 hours of reaction, the reaction was terminated. Wash 3 times with NMP, 2 times with DCM, and 2 times with methanol, and directly use for the next step of amino acid coupling.

[0060] Step two,

[0061] 1. Add the piperidine NMP solution with a volume ratio of 1:4 to the Fmoc-L-Lys(Boc)-CTC resin for deprotection reaction. After the reaction, the product is washed 3 times with NMP, 2 times with methanol, and 2 times with DCM 2 times.

[0062] 2. Weigh 6.07g Fmoc-Pro-OH, 2.43g HOBt, 6.83g HB...

Embodiment 2

[0072] Such as figure 1 As shown, the preparation method claimed in the present invention comprises the following steps:

[0073] step one,

[0074] 1. Put 10 g of WANG resin with a loading capacity of 1.0 mmol / g in the reactor, add NMP to wash twice, and then use NMP to swell for 1 h.

[0075] 2. Weigh 28.8g Fmoc-L-Lys(Boc)-OH, add 80ml NMP to dissolve, measure 4.67mL DIC to react for 2 hours, add it to the reactor, add 62mg DMAP to start the reaction.

[0076] 4. React for 6 hours, finish the reaction, wash with NMP for three times, measure 3.8ml of acetic anhydride and 3.3ml of pyridine and mix them in 70ml of NMP, add them into the reactor and react for 2 hours, wash with NMP for 3 times, DCM for 2 times, methanol for 2 times, directly used in the next step of amino acid coupling.

[0077] Step two,

[0078] 1. Add the piperidine NMP solution with a volume ratio of 1:4 to the Fmoc-L-Lys(Boc)-WANG resin for deprotection reaction. After the reaction, wash with NMP for 3 ...

Embodiment 3

[0089] Compared with Example 2, the only difference is that the solid-phase synthetic resin in this example is HMPA resin with a loading amount of 0.1 mmol / g.

[0090] The coupling agent used in step 1 was DIC / HOBt; the reaction solvent was NMP / DMF, the amount of DMAP was 12.5 mg, and the reaction time was 1 h. The coupling agents used in step 2 were HOAt and HATU, the organic base was DIPEA, the amount of the corresponding protected amino acid and coupling agent was 1.5 times the amount of resin loading, and the reaction time was 10 h. In step 3, the lysis reagent is TFA / thioanisole / anisole / EDT with a volume ratio of 90 / 5 / 3 / 2.

[0091] The purity of the product obtained in this example is 63%, and the synthesis yield is 48%.

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Abstract

The invention discloses a method for preparing telomerase polypeptide vaccine GV1001. The method includes the steps of firstly, subjecting Fmoc-L-Lys(Boc)-OH and solid synthetic resin as starter materials to reaction so as to obtain Fmoc-L-Lys(Boc)-resin; secondly, subjecting the Fmoc-L-Lys(Boc)-resin to gradual coupling to obtain side-chains-protected GV1001-resin according to a polypeptide sequence of GV1001 by Fmoc solid-phase synthesis; thirdly, decomposing the side-chains-protected GV1001-resin by decomposing reagent, adding precipitating reagent to allow for precipitation so as to obtain crude peptide, and purifying and freeze-drying the crude peptide to obtain the telomerase polypeptide vaccine GV1001. The method for preparing the telomerase polypeptide vaccine GV1001 has practical industrial application prospect and considerable economic and practical values.

Description

technical field [0001] The invention belongs to the technical field of pharmacy, and in particular relates to a method for preparing telomerase polypeptide vaccine GV1001. Background technique [0002] Malignant tumor is a common disease that threatens human life and health. The current surgical treatment, radiotherapy, drug treatment and other methods have no significant therapeutic effects on some malignant tumors, and the side effects after treatment are large. Studies have confirmed that the activation of telomerase and the regulation of telomere length are closely related to tumor formation, and have a high degree of specificity in the process of tumor treatment. In recent years, there have been many reports on the relationship between telomerase expression and tumor formation and development, and good progress has been made in tumor therapy targeting telomerase. [0003] In 1999, the world patent WO00 / 02581 disclosed a series of antigenic polypeptides derived from te...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K7/08C07K1/06C07K1/04A61K39/00
Inventor 朱颐申周云隆韦萍欧阳平凯
Owner NANJING UNIV OF TECH
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