Mycobacterium tuberculosis OS-tb oligosaccharide conjugate, and preparation method and application thereof

A technology of Mycobacterium tuberculosis and conjugates, applied in the field of Mycobacterium tuberculosis OS-tb oligosaccharide conjugates and its preparation, can solve problems such as unification, achieve wide application value, solve low immune protection, and avoid bacterial drug resistance problem effect

Active Publication Date: 2017-09-22
CANSINO BIOLOGICS INC
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, despite the rapid development of chemical synthesis of oligosaccharides in recent years, there is still no unified and effective universal method
The core oligosaccharide fragment OS of Mycobacterium tuberculosis with a single structure is used as an antigen, coupled with a carrier to prepare an oligosaccharide conjugate vaccine, and there are no related reports

Method used

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  • Mycobacterium tuberculosis OS-tb oligosaccharide conjugate, and preparation method and application thereof
  • Mycobacterium tuberculosis OS-tb oligosaccharide conjugate, and preparation method and application thereof
  • Mycobacterium tuberculosis OS-tb oligosaccharide conjugate, and preparation method and application thereof

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preparation example Construction

[0055] The general synthetic method of following Mycobacterium tuberculosis OS-tb oligosaccharide conjugates, the steps are as follows:

[0056] Synthesis of oligosaccharide conjugates

[0057] Activation of oligosaccharides: Dissolving oligosaccharides in ethanol (EtOH) and water (H 2 O) in the mixture (EtOH: H 2 (0=1:1), diethyl squarylate (5 equivalents) was added, and then saturated sodium carbonate solution was added dropwise every 5 minutes until the pH of the reaction solution = 8, and the reaction was stirred at room temperature for 1.5 hours. TLC detection showed that the reaction was complete, and it was separated and purified by silica gel column chromatography (the eluent used was dichloromethane (DCE) / methanol (MeOH)). After dissolving the product in water, use Sephadex LH 2 0 column for further purification, followed by freeze-drying to obtain the target product.

[0058] Synthesis of oligosaccharide-protein conjugates: According to the molar ratio of 20:1, t...

Embodiment 1

[0060] OS-tb-CRM 197 Synthesis of glycoconjugates (OS-CRM 197 )

[0061]

[0062] Synthesize OS-CRM by the method of general synthesis method steps 197 Conjugate, take 716mg CRM 197 Powder (5% CRM 197 , 95% sucrose) dissolved in 4mL buffer solution (Na 2 B 4 o 7 0.07mol / L, KHCO 3 0.035mol / L, pH9.0), then add 10mg compound OS (NMR such as figure 1 ) and diethyl squarylate, let stand at room temperature for 72 hours. Afterwards, the reaction solution was added to a 15mL ultrafiltration centrifuge tube (molecular weight cut-off 30KDa), centrifuged at 4000 rpm for 30 minutes, then deionized water was added, and centrifuged again, and this was repeated 7 times. After lyophilization, 31.4 mg of the sample was obtained. Mass spectrum: MALDI-TOF-MS (m / z) 63947.

[0063] OS-CRM 197 Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of the conjugate, see figure 2 , the resulting characterization showed that, with unconjugated CRM 197 carrier protein...

Embodiment 2

[0065] Synthesis of OS-BSA glycoconjugates

[0066]

[0067] Take OS (5mg), BSA (20mg) and OS-CRM 197 Synthesize in a similar way: the amount of activated oligosaccharide OS in the synthesis is 20eq., mass spectrum: MALDI-TOF-MS (m / z) 72327, the result is as follows image 3 .

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Abstract

The invention provides a mycobacterium tuberculosis OS-tb oligosaccharide conjugate, and a preparation method and application thereof. In the mycobacterium tuberculosis OS-tb oligosaccharide conjugate, the chemical structure of oligosaccharide is clear and single, is not a mixture, and can be abundantly synthesized by a chemical method; through the design of a connecting body to be coupled with a carrier for preparing the antigen, the problems of instability and low immune protection force of BCG vaccine can be solved; a relatively good immune effect can also be generated on some special people groups with low immunity. Wide application values are realized in the preparation of the tuberculosis vaccine.

Description

technical field [0001] The invention relates to the technical field of biological products, in particular to a Mycobacterium tuberculosis OS-tb oligosaccharide conjugate and its preparation method and application. Background technique [0002] Tuberculosis is a chronic infectious disease caused by Mycobacterium tuberculosis (Mtb) infection. About 1 / 3 of the people in the world are infected with MTB, and 5% to 10% of the infected people develop into TB patients, and about 2 million tuberculosis patients die every year. Tuberculosis chemotherapy is a relatively complete and effective method for the treatment and control of tuberculosis. However, with the increase of primary and acquired multidrug-resistant tuberculosis and extensively drug-resistant tuberculosis, current chemotherapy for tuberculosis is facing various challenges. Although chemotherapy can improve the patient's condition and prolong life, it brings disadvantages such as great pain, strong side effects and ser...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): A61K39/385A61K39/04A61P31/06
CPCA61K39/04A61K39/385A61K2039/60A61K2039/6031A61K2039/6037A61K2039/6081
Inventor 王浩猛莘春林朱涛常云松邵忠琦毛慧华宇学峰
Owner CANSINO BIOLOGICS INC
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