CN-115737793-B - Echinococci granulosa and echinococci canadensis bivalent subunit vaccine and preparation method thereof

CN115737793BCN 115737793 BCN115737793 BCN 115737793BCN-115737793-B

Abstract

The invention discloses a echinococcosis granulosa and echinococcosis canadensis bivalent subunit vaccine and a preparation method thereof, wherein the bivalent subunit vaccine comprises an immunogen dEG95/dEC antigen protein and a pharmaceutically acceptable carrier, the amino acid sequence of the dEG95 protein is shown as SEQ ID No.1, and the amino acid sequence of the dEC protein is shown as SEQ ID No. 2. At present, no vaccine for simultaneously resisting echinococcosis granulosa infection and Canadian echinococcosis infection exists, and the invention adopts dEG95-dEC95 heterodimer protein as antigen, and the prepared double subunit vaccine for resisting the echinococcosis and Canadian echinococcosis has the advantages of low production cost, simple production process, safety, high efficiency, low cost, high antigen purity and the like, and can simultaneously prevent the infection of the echinococcosis and Canadian echinococcosis.

Inventors

Nie dongsheng
ZHANG ZHEN
ZHONG CONGHAO
WANG JIANGHUI
DU XIANGYUE

Assignees

申联生物医药（上海）股份有限公司

Dates

Publication Date: 20260505
Application Date: 20210903

Claims (7)

1. A echinococcosis granulosa and echinococcosis canadensis bivalent subunit vaccine is characterized by comprising an immunogen dEG95 antigen protein, dEC antigen protein and a pharmaceutically acceptable carrier, wherein the amino acid sequence of the dEG95 antigen protein is shown as SEQ ID No.1, and the amino acid sequence of the dEC antigen protein is shown as SEQ ID No. 2; the two are dEG95 antigen protein and dEC antigen protein which are connected in series by polypeptide and expressed in the same vector, and the formed single chain dimer protein dEG95-dEC95.
2. The echinococci granulosa and echinococci canadensis bivalent subunit vaccine according to claim 1, characterized in that the content of the dEG95 antigen protein and dEC antigen protein in the bivalent subunit vaccine is 40-100 μg/ml.
3. The echinococci granulosa and echinococci canadensis bivalent subunit vaccine according to claim 1, wherein the pharmaceutically acceptable carrier comprises an adjuvant.
4. A echinococci granulosa and echinococci canadensis bivalent subunit vaccine according to claim 3, characterized in that the adjuvant content in the bivalent subunit vaccine is 5% -60% V/V.
5. The echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine according to claim 1, wherein the pharmaceutically acceptable carrier comprises one or several of antioxidants, surfactants, colorants, volatile oils, buffers, dispersants, propellants and preservatives.
6. A method of preparing a echinococci granulosa and echinococci canadensis bivalent subunit vaccine according to claim 1, characterized in that the method comprises the steps of: S1, modifying genes of dEG95 antigen protein in echinococcus granulosus and dEC antigen protein in echinococcus canadensis, carrying out tandem connection, synthesizing genes to obtain dEG95-dEC95 genes, cloning the genes into an expression vector, and obtaining a recombinant expression vector containing the dEG95-dEC antigen protein genes; S2, transforming or transducing the recombinant expression vector containing the dEG95-dEC antigen protein gene obtained in the step S1 into a host to obtain a recombinant containing the recombinant expression vector; s3, culturing the recombinant obtained in the step S2, and expressing dEG95-dEC antigen protein; S4, purifying the dEG95-dEC antigen protein obtained in the step S3, and adding a pharmaceutically acceptable carrier to obtain the bivalent subunit vaccine.
7. The method for preparing a echinococci granulosa and echinococci canadensis bivalent subunit vaccine according to claim 6, wherein the dEG95-dEC antigen protein is an intracellular soluble protein.

Description

Echinococci granulosa and echinococci canadensis bivalent subunit vaccine and preparation method thereof Technical Field The invention belongs to the technical field of genetic engineering, relates to a subunit vaccine for resisting echinococci infection and a preparation method thereof, and in particular relates to a echinococci granulosa and Canadian echinococci bivalent subunit vaccine and a preparation method thereof. Background Echinococcosis (Echinococcosis) is a serious parasitic disease of human and veterinary species caused by the parasitic of echinococcosis (HYDATID CYST), a larva of echinococcosis, to tissues and organs such as lung and liver of humans and animals. Echinococcosis is widely spread and distributed in the world, and is classified by the world animal health organization (World Organization for ANIMAL HEALTH; french: office international des e pizooties, OIE) into a global reported infectious disease belonging to various zoonotics, and classified by the world health organization (World Health Organization, WHO) as one of diseases which are predicted and treated in advance by the global early warning system. Echinococcosis is also one of five major parasitic diseases planned to be prevented and treated by the ministry of health of China. Echinococcosis is mainly divided into echinococcosis granulosa (cystic echinococcosis, CE) and echinococcosis multiflorum according to the focus morphology and infection pathogen difference, wherein the echinococcosis granulosa is most widely distributed and the number of patients is the greatest. The causative agent of CE is currently composed of several Echinococcus granulosus (Echinococcus granuLosus), echinococcus Canadian (Echinococcus Canadensis), ma Jiqiu, and Echinococcus omutus, with CE caused by Echinococcus echinococcus type G1 exceeding 90% and CE caused by Echinococcus Canadensis type G6 exceeding 7%. The current research shows that the epidemic of echinococcosis is controlled mainly by cutting off the development link of echinococcosis, controlling the infection of intermediate hosts such as human beings, livestock and the like on the echinococcosis, preventing or expelling parasites to treat final hosts such as dogs and the like, and blocking the wide-range spreading of ova. Wherein the epidemic of echinococcosis granulosa can be effectively controlled for the vaccination of intermediate hosts. Lightowlers et al found EG95 to be one of the natural hexacercaria antigens present in EG and the most potent protective antigen among the numerous proteins screened, have been successful in developing vaccines against echinococcosis in sheep. However, the existing recombinant EG95 protein vaccine is prepared by renaturation of an escherichia coli inclusion body, has low purity, is difficult to maintain the spatial structure of the protein, has no obvious cross protection with the G6 type echinococcus canadensis, has a certain limitation, and is currently in urgent need of researching multivalent vaccines aiming at echinococcus granulosus and echinococcus canadensis for preventing and treating echinococcosis. Disclosure of Invention In view of the defects in the prior art, the invention aims to provide a echinococci granulosa and Canadian echinococci bivalent subunit vaccine and a preparation method thereof, which can simultaneously prevent the infection of the echinococci granulosa and Canadian echinococci, and have the advantages of safety, high efficiency, low cost, high antigen purity and the like. Wherein the anti-echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine comprises an immunogen dEG95/dEC antigen protein and a pharmaceutically acceptable carrier. As an implementation mode of the invention, in the anti-echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine, the amino acid sequence of the dEG95 protein is shown as SEQ ID No.1, and the amino acid sequence of the dEC protein is shown as SEQ ID No. 2. As one embodiment of the invention, in the anti-echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine, the dEG95 protein can be a monomer or a single-chain polymer connected in series by polypeptides. Examples are dimer 2dEG95, trimer 3dEG95, tetramer 4dEG95, pentamer 5dEG95. As one embodiment of the invention, the dEC protein in the anti-echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine can be a monomer or a single-chain polymer connected in series by polypeptides. Examples are dimer 2dEC95, trimer 3dEC95, tetramer 4dEC95, pentamer 5dEC95. As one embodiment of the invention, in the anti-echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine, the bivalent subunit can be a mixture of dEG95 protein and dEC protein formed by expressing the dEG95 protein and dEC protein respectively in different vectors. As one embodiment of the invention, in the anti-echinococcus granulosus and echinococcus canadensis bivalent subunit vaccine,