CN102949717A - Novel hepatitis B vaccine preparation containing poly I:C adjuvant - Google Patents
Novel hepatitis B vaccine preparation containing poly I:C adjuvant Download PDFInfo
- Publication number
- CN102949717A CN102949717A CN2012102322373A CN201210232237A CN102949717A CN 102949717 A CN102949717 A CN 102949717A CN 2012102322373 A CN2012102322373 A CN 2012102322373A CN 201210232237 A CN201210232237 A CN 201210232237A CN 102949717 A CN102949717 A CN 102949717A
- Authority
- CN
- China
- Prior art keywords
- hepatitis
- vaccine
- adjuvant
- preparation
- poly
- 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.)
- Pending
Links
Images
Landscapes
- Medicines Containing Antibodies Or Antigens For Use As Internal Diagnostic Agents (AREA)
Abstract
The invention relates to a novel hepatitis B vaccine preparation containing poly I:C adjuvant, and belongs to the technical fields of biotechnology and genetic engineering vaccines. The invention relates to a new formula of hepatitis B vaccine, and the vaccine comprises the main ingredients: genetic engineering hepatitis B virus surface antigens, aluminum adjuvant, poly I:C and the like. The immunogenicity of the hepatitis B vaccine prepared by the formula is superior to pure aluminum adjuvant-containing vaccine used currently, the hepatitis B vaccine can be used for immunizing the adult, kidney transplant patients, renal dialysis patients and the like, so as to prevent infection of hepatitis B virus. The hepatitis B vaccine can also be used for immune therapy of chronic hepatitis B patients.
Description
Technical field the present invention relates to a kind of new Hepatitis B virus vaccine preparation, for adopting hepatitis B (hereinafter to be referred as the hepatitis B) vaccine of Poly I:C and aluminium adjuvant, belongs to biological technical field.
Background technology China has the population infection of 50-70% to cross hepatitis B virus (HBV), and 10% crowd (approximately 1.3 hundred million people) is the virus carrier.The effective ways that also lack at present the radical cure hepatitis B, HB vaccination are the most cost-effective measures of prevention hepatitis.To going deep into that therapeutic vaccine is studied, the Hepatitis B virus vaccine of control combination will have more wide prospect [1-4] along with in recent years.
Hepatitis B virus (HBV) is a kind of DNA viruses of biting liver property that has, and only to people and orangutan susceptible, can cause liver tissue injury, causes hepatitis B.The HBV virus structure is comprised of shell and core, the contained hepatitis B surface antigen (HBsAg) of shell comprises again S antigen, front S1 and preS 2 antigen, as virus identification with invade the main in conjunction with albumen of host cell, HBsAg be present HBV vaccine research and development for main target spot.Clinical employed recombinant vaccine and haematogenous vaccine all form [5,6] by hepatitis B surface antigen master albumen (S) at present.Extensively since the inoculation, produced very good control and prevention of disease effect in China from 1985, infection and the sickness rate of HBV be effectively controlled [4,7].Yet huge population base has determined that the still sternness, the particularly antiviral therapy of HBV of prevention and control task of China HBV is with high costs and curative effect is limited, and easy drug resistance is so the HBV vaccine very necessary [8] that research and development have therapeutic effect.
In a large amount of Hepatitis B virus vaccines that use of China, mainly adopt HbsAg as antigen at present, the aluminium adjuvant of 0.5-1mg strengthens, and the humoral immunity level that this compatibility brings out is lower, and has defective aspect cellular immunization.Such vaccine exist the crowd that approximately has about 10% to it without immunoreation or hyporeactive, and shortcomings such as mutant [9] that exist partial immunity to escape.
Polyinosinic acid-polycytidylicacid (polyinosine-polycytidylic acid, Poly I:C) is a kind of double-stranded RNA of synthetic, can be by Toll sample receptor-3 (the Toll-like receptor 3 on the immunocyte, TLR-3) identify, thereby induce host cell to produce interferon-ALPHA and β, have antiviral and regulate immune multi-efficiency [10-12].Poly I:C is used for chronic hepatitis B, epidemic hemorrhagic fever, epidemic encephalitis type B, viral keratitis, herpes zoster, various wart class and respiratory tract infection etc. mainly as immunostimulant, anti-tumor medicine at present.Clinical adult's consumption is each 1-2mg intramuscular injection, and major side effects is crossed the property low grade fever for the rear a few patients of injection can occur one, and its application as Human vaccine adjuvant exists the probability of safety problem lower [13-14].
There is no at present the report that the two adjuvant compatibilities that adopt hepatitis B surface antigen to add Poly I:C and aluminium adjuvant are applied to hepatitis B vaccine.
Summary of the invention:
The invention provides a kind of new hepatitis B vaccine preparation, the composition of this vaccine also comprises Poly I:C except comprising HbsAg and aluminium adjuvant.
Hepatitis B vaccine of the present invention utilizes polyinosinic acid-polycytidylicacid (polyinosine-polycytidylic acid, be abbreviated as Poly I:C) effect of immunological adjuvant, Poly I:C is immune cell activated effectively, induce to produce take a type interferon as main panimmunity activity factor, in experimental animal, can induce high-caliber antigenic specificity body fluid and cell immune response.Adopt the immunological adjuvant of this kind particular type can carry out the immunization therapy of various types of HBV infection and the immunosuppressive drug user is carried out effective immunity, present more refractory asymptomatic HBsAg (HBsAg) carrier can be turned out cloudy and occurred resisting-HBs, also might make impaired liver obtain to a certain degree Regeneration and Repair.
The hbs antigen of using among the present invention is as genetic engineering recombinant antigen or its fragment or contain the antigen of pre S1 composition, rather than derives from the hbs antigen of hepatitis B human plasma.Genetic engineering surface antigen by yeast or expressing cho cell is comprised of 226 aminoacid.The HBsAg of recombinant yeast or expressing cho cell all can form the VLP granule of 22nm, is applied to community immunity for many years, and clinical proof has good preventive effect and safety.
The following Poly I:C of content range 0.2-1.2mg, the HBsAg5-125ug of each main constituent, aluminium adjuvant (pressing aluminium ion calculates) 0.2-1.2mg in every dose of the hepatitis B vaccine of the present invention.
Preferred scope is as follows:
Poly I:C 0.5-1.0mg, HBsAg 10-100ug, aluminum hydroxide adjuvant (are pressed Al
3+Calculate 1mg/ml) 0.5-1.0mg.
Most preferred scope is as follows:
Poly I:C 0.5-1.0mg, HBsAg 25-75ug, aluminum hydroxide adjuvant (are pressed Al
3+Calculate 1mg/ml) 0.5-1.0mg.
More than narrate every dose and refer to each bacterin preparation, such as each injection, or every injection, or the injection medicament of each use amount.
Vaccine using method of the present invention is identical with existing hepatitis B vaccine using method.
Hepatitis B vaccine of the present invention can prepare by the following method:
Form:
Polyinosinic acid-polycytidylicacid (Poly I:C)
Hepatitis B virus surface antigen (vaccine)
Aluminum hydroxide adjuvant
Preparation:
Add aluminum hydroxide adjuvant in the hepatitis B virus surface antigen, behind the mixing, add Poly I:C solution, mixing namely gets vaccine liquid, and this vaccine liquid can be made into the bacterin preparation of the injection of unit dose through packing.
Also can contain other human body that can improve immunity of organism or microprotein, polysaccharide or micromolecular compound in the bacterin preparation of the present invention.
Bacterin preparation of the present invention can be used as the medicine of adult, kidney dialysis patient, organ transplantation patient and hepatitis B high-risk group's immunization therapy.Also can be used as the medicine of the immunization therapy of chronic hepatitis B patients.
Vaccine of the present invention also can form combined vaccine with other bacterin preparations that contains aluminium adjuvant.
The difference of hepatitis B vaccine of the present invention and existing hepatitis B vaccine is, added immunostimulant PolyI:C, Poly I:C is joined in the vaccine, can effectively stimulate body to produce high-level humoral immunization and cellular immunization, this vaccine brings bright prospect will for the treatment of chronic hepatitis B.
Zooperal result shows, vaccine of the present invention obviously is better than existing hepatitis B vaccine.Be applied to behind the human body greatly humoral immunization and the cellular immunization of the raising human body anti-hepatitis B virus of spoke degree, can be used for the treatment of chronic hepatitis B patients.
Animal experiment shows that this novel hepatitis B vaccine is safe, to experimental animal without visible toxicity.Effectively stimulating animal produces the specific antibody of high titre.
The prescription that hepatitis B vaccine of the present invention adopts can induce body to produce stronger immunoreation.This vaccine can be used for being grown up, other type hepatitis b vaccination loser, kidney dialysis patient, organ transplantation patient's immunity, and the immunization therapy of chronic hepatitis B patients.
Description of drawings
Fig. 1 is the anti-S specific IgG antibodies of mice serum figure after the immunity of ELISA detection proteantigen.Utilizing ELISA to detect 1.25 μ g hepatitis B virus surface antigens cooperates adjuvant to stimulate BALB/c mouse to say the result who produces the S specific IgG.F1 represents simple vaccine group, and F2 represents vaccine hydro-oxidation aluminium adjuvant group, and F3 represents vaccine and adds Poly I:C adjuvant group, and F4 represents the vaccine adding aluminum hydroxide and adds the two adjuvants of Poly I:C, and NS represents simple normal saline group.The 2nd week after for the first time immunity, obvious antibody male rotary all appears in the mice that employing contains Poly I:C adjuvant group, the adjuvant group is better than and does not add adjuvant group (p＜0.05) when detecting in the 2nd week (6week) after the immunity for the second time, and detecting aluminum hydroxide adjuvant group titre the 8th week (12week) after the immunity for the second time is 10
3.38 ± 0.31, two adjuvant group antibody titers are respectively 10
3.78 ± 0.48, all be better than other two groups (p＞0.05).The two adjuvants of this presentation of results are inducing the effect that produces the S specific antibody suitable with the collaborative proteantigen of simple aluminium hydroxide, and can both keep for a long time.
Fig. 2 is the anti-S specific IgG antibodies of mice serum figure after the immunity of ELISA detection proteantigen.Utilizing ELISA to detect 1.25 μ g hepatitis B virus surface antigens cooperates adjuvant to stimulate BALB/c mouse to say the result who produces the S1 specific IgG.F1 represents simple vaccine group, and F2 represents vaccine hydro-oxidation aluminium adjuvant group, and F3 represents vaccine and adds Poly I:C adjuvant group, and F4 represents the vaccine adding aluminum hydroxide and adds the two adjuvants of Poly I:C, and NS represents simple normal saline group.The 2nd week after for the first time immunity, each is organized mice and antibody male rotary all occurs, respectively organize antibody titer no significant difference (p＞0.05) when detecting in the 2nd week (6week) after the immunity for the second time, detecting aluminum hydroxide adjuvant group titre the 8th week (12week) after the immunity for the second time is 10
4.18 ± 0.38, two adjuvant group antibody titers are respectively 10
4.25 ± 0.75, all be better than simple proteins group (p＞0.05).The two adjuvants of this presentation of results are inducing the effect that produces the S1 specific antibody suitable with the collaborative proteantigen of simple aluminium hydroxide, and can both keep for a long time.
Fig. 3 is the anti-S specific IgG antibodies of mice serum figure behind the ELISA detection Adv booster immunization.Utilize adenovirus carrier vaccine (rAdv) that ELISA detect to express hepatitis B virus surface antigen to the reinforced effects of the S specific IgG that cooperates different adjuvant immunity mices through protein vaccine.F1+ represents simple vaccine+rAdv group, F2+ represents vaccine hydro-oxidation aluminium adjuvant group+rAdv group, F3+ represents vaccine and adds Poly I:C adjuvant group+rAdv group, and F4+ represents the vaccine adding aluminum hydroxide and adds the two adjuvants of Poly I:C+rAdv group, and rAdv represents simple normal saline group+rAdv group.In the 2nd week behind booster immunization, blood sampling detects anti-S specific antibody, and antibody titer is respectively F1+:10
2.99 ± 0.13, F2+:10
3.42 ± 0.59, F3+:10
3.15 ± 0.46, F4+:10
3.68 ± 0.56The F2+ group is obviously organized (p＜0.05) greater than F1+ with the F4+ group.This result again illustrates and adopts under this immunization protocol, and two adjuvants carry out early immune with the collaborative proteantigen of simple aluminium hydroxide, induce the effect that produces the S specific antibody suitable.
Fig. 4 is the anti-S specific IgG antibodies of mice serum figure behind the ELISA detection Adv booster immunization.Utilize adenovirus carrier vaccine (rAdv) that ELISA detect to express hepatitis B virus surface antigen to the reinforced effects of the S1 specific IgG that cooperates different adjuvant immunity mices through protein vaccine.F1+ represents simple vaccine+rAdv group, F2+ represents vaccine hydro-oxidation aluminium adjuvant group+rAdv group, F3+ represents vaccine and adds Poly I:C adjuvant group+rAdv group, and F4+ represents the vaccine adding aluminum hydroxide and adds the two adjuvants of Poly I:C+rAdv group, and rAdv represents simple normal saline group+rAdv group.In the 2nd week behind booster immunization, blood sampling detects anti-S1 specific antibody, and antibody titer is respectively F1+:10
3.59 ± 0.61, F2+:10
3.89 ± 0.54, F3+:10
3.89 ± 0.26, F4+:10
4.38 ± 0.66Only the F4+ group obviously is better than F1+ group (p＜0.05).This presentation of results adopts under this immunization protocol, and the collaborative proteantigen of two adjuvants carries out early immune after adenovirus carrier vaccine strengthens, and has some superiority aspect S 1 specific antibody inducing to produce.
Fig. 5 is that mice S Specific T cell immunity reacted after ELISPOT detected the Adv booster immunization.Utilize adenovirus carrier vaccine (rAdv) that ELISPOT detect to express hepatitis B virus surface antigen to the reinforced effects of the S Specific T cell immunity that cooperates different adjuvant immunity mices through protein vaccine.F1+ represents simple vaccine+rAdv group, F2+ represents vaccine hydro-oxidation aluminium adjuvant group+rAdv group, F3+ represents vaccine and adds Poly I:C adjuvant group+rAdv group, and F4+ represents the vaccine adding aluminum hydroxide and adds the two adjuvants of Poly I:C+rAdv group, and rAdv represents simple normal saline group+rAdv group.In the 2nd week behind booster immunization, the aseptic spleen of getting carries out S specificity ELIPOT detection, and the result shows counting obviously more than other each groups (p＜0.05) of two adjuvant generations that group is induced.This presentation of results adopts the collaborative proteantigen of two adjuvants to carry out early immune after adenovirus carrier vaccine strengthens, and can induce to produce than all the other with reference to the stronger S specificity cellular immunity response of adjuvant group, uses for the therapeutic of vaccine to lay the foundation.
Fig. 6 is that mice S1 Specific T cell immunity reacted after ELISPOT detected the Adv booster immunization.Utilize adenovirus carrier vaccine (rAdv) that ELISPOT detect to express hepatitis B virus surface antigen to the reinforced effects of the S1 Specific T cell immunity that cooperates different adjuvant immunity mices through protein vaccine.F1+ represents simple vaccine+rAdv group, F2+ represents vaccine hydro-oxidation aluminium adjuvant group+rAdv group, F3+ represents vaccine and adds Poly I:C adjuvant group+rAdv group, and F4+ represents the vaccine adding aluminum hydroxide and adds the two adjuvants of Poly I:C+rAdv group, and rAdv represents simple normal saline group+rAdv group.In the 2nd week behind booster immunization, the aseptic spleen of getting carries out S1 specificity ELIPOT detection, and the result shows counting obviously more than other each groups (p＜0.05) of two adjuvant generations that group is induced.This presentation of results adopts the collaborative proteantigen of two adjuvants to carry out early immune after adenovirus carrier vaccine strengthens, and can induce to produce than all the other with reference to the stronger S1 specificity cellular immunity response of adjuvant group, uses for the therapeutic of vaccine to lay the foundation.
The specific embodiment:
Further specify by the following examples the present invention.
Embodiment 1
The preparation method of hepatitis B vaccine injection
Raw material and source:
Expressing cho cell hepatitis B virus surface antigen (vaccine dry powder): be Huabei Pharmaceutic Co., Ltd's product, be mixed with 200 μ g/ml with the injection normal saline.
Aluminum hydroxide adjuvant: 1.0mg/ml (presses Al
3+Calculate, 1mg/ml).
Polyinosinic acid-polycytidylicacid (Poly I:C): available from U.S. SIGMA company.Be mixed with 2mg/ml with the injection normal saline, with the degerming membrane filtration degerming of 0.22 μ m, frozen in-20 ℃.
The preparation method of vaccine: get the hepatitis B virus surface antigen 5.0ml of expressing cho cell in aseptic vial, behind the abundant mixing of adding aluminum hydroxide adjuvant 10ml, add Poly I:C solution 5.0ml, be the 20ml vaccine solution behind the mixing.Every milliliter of the vaccine liquid of preparing contains hbs antigen 50ug, aluminium adjuvant 0.5mg (presses Al
3+Calculate, 1mg/ml), Poly I:C 0.5mg, after this solution packing, make the vaccine injecta of 20 bottles of every bottle of 1ml.
Comparative study bacterin preparation of the present invention and the effect of aluminum hydroxide adjuvant compatibility agent in two pin simple proteins immunity commonly used,
Test as follows:
Prepare the raw material of vaccine and originate as follows:
Expressing cho cell hepatitis B virus surface antigen (vaccine): be Huabei Pharmaceutic Co., Ltd's product, be mixed with 200 μ g/ml with the injection normal saline.
Aluminum hydroxide adjuvant: 1.0mg/ml (presses Al
3+Calculate, 1mg/ml).
Polyinosinic acid-polycytidylicacid (Poly I:C): available from U.S. SIGMA company.Be mixed with 2mg/ml with the injection normal saline, with the degerming membrane filtration degerming of 0.22 μ m, frozen in-20 ℃.
The preparation method of vaccine: get the hepatitis B virus surface antigen 5.0ml of expressing cho cell in aseptic vial, behind the abundant mixing of adding aluminum hydroxide adjuvant 10ml, add Poly I:C solution 5.0ml, be the 20ml vaccine solution behind the mixing.Every milliliter of the vaccine liquid of preparing contains hbs antigen 50ug, aluminium adjuvant 0.5mg (presses Al
3+Calculate, 1mg/ml), Poly I:C0.5mg.
The prescription of other each matched group vaccines following (table 1), preparation method is the same.
Table 1, the zoopery preparation of hepatitis B vaccine
Contrast experiment's method:
Animal immune: female SPF BALB/c mouse, body weight 16-18 gram, 12 every group.In intramuscular injection in the 0th, 28 day, immunizing dose 0.1ml.
The 8th week get blood after two weeks and the second pin immunity after each immunity, get blood 0.1ml through eye socket at every turn, separation of serum, frozen in-20 ℃ before measuring.
Immune protective effect detects:
1. the ELISA of S specific IgG measures in the mice serum: anti-HBs (anti-S) diagnostic kit is available from Shanghai Kehua Bio-engineering Co., Ltd.For dual-antigen sandwich method for determining anti--test kit of HBS titre.With 2.1 times of the meansigma methods of the aluminium adjuvant group Mus serum A450/650 value of 1: 25 times of dilution as (Cut off) dividing value, with resisting-HBs quantitative assay standard substance production standard curve that Nat'l Pharmaceutical ﹠ Biological Products Control Institute provides, the calculating serum that serum A450/650 value to be checked is higher than dividing value resists-the S antibody content simultaneously.The geometric mean titer of each organizes laboratory animal serum anti--S is relatively seen Fig. 1.
2. the ELISA of S1 specific IgG measures in the mice serum: hepatitis B virus front S 1 antibody (anti-S1) diagnostic kit is available from Shanghai Alpha Bioisystech Co., Ltd.For dual-antigen sandwich method for determining anti--test kit of HBS titre.With 2.1 times of the meansigma methods of the aluminium adjuvant group Mus serum A450/650 value of 1: 25 times of dilution as (Cut off) dividing value, with resisting-HBs quantitative assay standard substance production standard curve that Nat'l Pharmaceutical ﹠ Biological Products Control Institute provides, the calculating serum that serum A450/650 value to be checked is higher than dividing value resists-the S antibody content simultaneously.The geometric mean titer of each organizes laboratory animal serum anti--S 1 is relatively seen Fig. 2.
Comparative study bacterin preparation of the present invention and commonly used aluminum hydroxide adjuvant compatibility agent are just exempted from effect in the adenovirus booster immunization at albumen, test as follows:
The one or two pin immunity is with the raw material of vaccine and originate as follows:
Expressing cho cell hepatitis B virus surface antigen (vaccine): be Huabei Pharmaceutic Co., Ltd's product, be mixed with 200 μ g/ml with the injection normal saline.
Aluminum hydroxide adjuvant: 1.0mg/ml (presses Al
3+Calculate, 1mg/ml).
Polyinosinic acid-polycytidylicacid (Poly I:C): available from U.S. SIGMA company.Be mixed with 2mg/ml with the injection normal saline, with the degerming membrane filtration degerming of 0.22 μ m, frozen in-20 ℃.
The preparation method of vaccine: get the hepatitis B virus surface antigen 5.0ml of expressing cho cell in aseptic vial, behind the abundant mixing of adding aluminum hydroxide adjuvant 10ml, add Poly I:C solution 5.0ml, be the 20ml vaccine solution behind the mixing.Every milliliter of the vaccine liquid of preparing contains hbs antigen 50ug, aluminium adjuvant 0.5mg (presses Al
3+Calculate, 1mg/ml), Poly I:C0.5mg.
The prescription of other each matched group vaccines following (table 1), preparation method is the same.
Table 1, the zoopery preparation of hepatitis B vaccine
Raw material and the source of vaccine used in the immunity of the 3rd pin:
Adenovirus carrier vaccine rAdv: by our unit's self-control, comprise HBV envelope protein S and S1 gene, but effective expression S and S1.The injection normal saline is configured to 1x10
9Vp/ml
Contrast experiment's method:
Animal immune: female SPF BALB/c mouse, body weight 16-18 gram, 12 every group.In the 0th, 28 day intramuscular injection protein vaccine, immunizing dose 0.1ml.Adopt the Ad-SS1 immunity in the time of the 98th day, immunizing dose 0.1ml, intramuscular injection.
Get blood after the last immunity two weeks and pluck eyeball and get blood, separation of serum, frozen in-20 ℃ before measuring.Mice is taken off neck put to death the rear aseptic spleen of getting, grind and separate the acquisition splenocyte suspension, be used for immediately ELISPOT behind the counting adjustment cell concentration and detect.
Immune protective effect detects:
1. the ELISA of S specific IgG measures in the mice serum: anti-HBs (anti-S) diagnostic kit is available from Shanghai Kehua Bio-engineering Co., Ltd.For dual-antigen sandwich method for determining anti--test kit of HBS titre.With 2.1 times of the meansigma methods of the aluminium adjuvant group Mus serum A450/650 value of 1: 25 times of dilution as (Cut off) dividing value, with resisting-HBs quantitative assay standard substance production standard curve that Nat'l Pharmaceutical ﹠ Biological Products Control Institute provides, the calculating serum that serum A450/650 value to be checked is higher than dividing value resists-HBs content simultaneously.The geometric mean titer of each organizes laboratory animal serum anti--S is seen Fig. 3.
2. the ELISA of S1 specific IgG measures in the mice serum: hepatitis B virus front S 1 antibody (anti-S1) diagnostic kit is available from Shanghai Alpha Bioisystech Co., Ltd.For dual-antigen sandwich method for determining anti--test kit of HBS titre.With 2.1 times of the meansigma methods of the aluminium adjuvant group Mus serum A450/650 value of 1: 25 times of dilution as (Cut off) dividing value, with resisting-HBs quantitative assay standard substance production standard curve that Nat'l Pharmaceutical ﹠ Biological Products Control Institute provides, the calculating serum that serum A450/650 value to be checked is higher than dividing value resists-HBs content simultaneously.The geometric mean titer of each organizes laboratory animal serum anti--S1 is seen Fig. 4.
3.S specific cellular immunity is measured: mice γ-IFN cytokine ELISPOT reagent is available from BD company.The by specification operation, behind the anti-γ of coated anti-usefulness-IFN antibody, every hole adds the spleen cell cultures suspension that 100ul contains the S antigen polypeptide, and every hole spleens cell number is 5X105, and 37 ℃ of 5%CO2 leave standstill and cultivate 24h; After discarding culture fluid, substep adds detection antibody and nitrite ion colour developing, Bioreader4000 spot-analysis instrument counting spot formation number (SFC).According to the quantity of how much judging specific killing T cell of speckle number, what of this quantity are proportionate with cellular immunization intensity.The results are shown in Figure 5.
4.S1 specific cellular immunity is measured: mice γ-IFN cytokine ELISPOT reagent is available from BD company.The by specification operation, behind the anti-γ of coated anti-usefulness-IFN antibody, every hole adds the spleen cell cultures suspension that 100ul contains the S1 antigen polypeptide, and every hole spleens cell number is 5X105, and 37 ℃ of 5%CO2 leave standstill and cultivate 24h; After discarding culture fluid, substep adds detection antibody and nitrite ion colour developing, Bioreader4000 spot-analysis instrument counting spot formation number (SFC).According to the quantity of how much judging specific killing T cell of speckle number, what of this quantity are proportionate with cellular immunization intensity.The results are shown in Figure 6.
List of references
[1] report [M] is comprehensively evaluated in Ministry of Public Health .2004 whole nation planned immunization. Beijing: People's Health Publisher, 2005.
[2]Leachy，D.Worldwide?epidemiology?of?HBV?infection，disease?burden，and?vaccine?prevention[J].J.Clin.Virol?2005，34：S1-S3.
[3]Kane?M.Global?programme?for?control?of?hepatitis?B?infection[J].Vaccine，1995，13：(S1)S47
[4] Dai Zhicheng, Qi Guoming. Chinese Seroepidemiological study of viral hepatitis (1992-1995) (upper volume) [M]. Beijing: scientific and technical literature publishing house, 1997.39-60.
[5]WHO.Hepatitis?B?surface?antigen?assay：operational?characteristics[R].Geneva：WHO/BCT/BTS/01.4，2001，5，1-15.
[6]Lok?A?S，McMahon?B?J.Chronic?hepatitis?B[J].Hepatology，2001，34：1225-1241.
[7] Liang Xiaofeng, Chen Yuansheng, Wang Xiaojun, etc. Chinese population aged over 3-years old serum of hepatitis B epidemic research [J]. Chinese epidemiology magazine, 2005,26 (9): 655-658.
[8] Tan Wenjie, combined strategy and the progress [J] of Ruan Li .HBV virus chronic infection vaccine therapy. viral journal, 2006, (5): 407-410.
[9] Liu Chongbai, Su Chongao. the problem of hepatitis b vaccination and existence [J]. Chinese planned immunization, 2004,10 (3): 159-162.
[10]Ngoi?S?M，Tovey?M?G，Vella?A?T.Targeting?poly(I:C)to?the?TLR3-independent?pathway?boosts?effector?CD8?Tcell?differentiation?through?IFN-alpha/beta[J].J?Immunol，2008，(11)：7670-80
[11]Rosenne?E，Shakhar?G，Melamed?R，et?al.Inducing?a?mode?of?NK-resistance?to?suppression?by?stress?and?surgery：a?potential?approach?based?on?low?dose?of?poly?I-C?to?reduce?postoperative?cancer?metastasis[J].Brain?Behav?Immun.2007，21(4)：395-408.
[12]He?X?S，Ansari?A?A，Ridgway?W?M，et?al.New?insights?to?the?immunopathology?and?autoimmune?responses?in?primary?biliary?cirrhosis[J].Cell?Immunol，2006.239：1-13.
[13] wear spedex. special medicine handbook [M] .2 of practical new drug version. Beijing: People's Medical Officer Press, 1997:273.
[14] Wang Li. pediatric pharmacology and pharmacotherapeutics [M]. Beijing: publishing house of Beijing Medical University, 2002:555-557.
Claims (6)
1. a hepatitis B vaccine preparation is characterized in that, contains hepatitis B surface antigen in the said preparation, aluminum hydroxide adjuvant, and polyinosinic acid-poly-cytidylic acid (polyinosine-polycytidylic acid, poly I:C).
2. the bacterin preparation of claim 1 is characterized in that, wherein hbs antigen is hepatitis B virus surface antigen or its fragment of recombinaant CHO cell manufacturing or the antigen that contains preS1.
3. the bacterin preparation of claim 1 is characterized in that, the content of PolyI:C is 0.2-1.2mg in every vaccinating agent, and the content of hepatitis B surface antigen is 5-125ug, aluminum hydroxide adjuvant (calculating with aluminum ions amount) 0.2-1.2mg.
4. the bacterin preparation of claim 1 also can contain human body or microprotein and polysaccharide or the micromolecular compound of other known improved immunity of organism.
5. the bacterin preparation of claim 1 is for the preparation of the application in the immunotherapy medicaments of the medicine of adult, kidney dialysis patient, organ transplantation patient and hepatitis B high-risk group immunity and preparation chronic hepatitis B patients.
6. the bacterin preparation of claim 1 and other chronic hepatitis B patients are treated the chemical synthetic drug use in conjunction of usefulness.
Priority Applications (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012102322373A CN102949717A (en) | 2012-07-06 | 2012-07-06 | Novel hepatitis B vaccine preparation containing poly I:C adjuvant |
Applications Claiming Priority (1)
| Application Number | Priority Date | Filing Date | Title |
|---|---|---|---|
| CN2012102322373A CN102949717A (en) | 2012-07-06 | 2012-07-06 | Novel hepatitis B vaccine preparation containing poly I:C adjuvant |
Publications (1)
| Publication Number | Publication Date |
|---|---|
| CN102949717A true CN102949717A (en) | 2013-03-06 |
Family
ID=47759477
Family Applications (1)
| Application Number | Title | Priority Date | Filing Date |
|---|---|---|---|
| CN2012102322373A Pending CN102949717A (en) | 2012-07-06 | 2012-07-06 | Novel hepatitis B vaccine preparation containing poly I:C adjuvant |
Country Status (1)
| Country | Link |
|---|---|
| CN (1) | CN102949717A (en) |
Cited By (9)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103386128A (en) * | 2013-07-02 | 2013-11-13 | 中国食品药品检定研究院 | Tuberculosis subunit vaccine containing unite adjuvant |
| CN107456574A (en) * | 2017-08-14 | 2017-12-12 | 山东大学 | One kind is without adjuvant single dose HBsAg nanogel vaccines and preparation method thereof |
| CN107556166A (en) * | 2016-06-30 | 2018-01-09 | 中国科学院化学研究所 | Polyhydroxylated fullerene and preparation method thereof |
| CN109355334A (en) * | 2018-12-06 | 2019-02-19 | 江苏省农业科学院 | A method of preparing poly IC |
| CN109876140A (en) * | 2019-03-19 | 2019-06-14 | 何勇刚 | A kind of vaccine and its preparation method and application for treating chronic hepatitis B |
| CN113398261A (en) * | 2021-07-16 | 2021-09-17 | 山西医科大学 | Application of poly-sarcosine in preparing biological preparation for improving HBsAg positive mother infant hepatitis B vaccine response level |
| CN113423423A (en) * | 2018-11-13 | 2021-09-21 | 变异生物技术公司 | Immunogenic compositions for the treatment of hepatitis B |
| CN116115748A (en) * | 2023-01-17 | 2023-05-16 | 大连理工大学 | Al-Poly (I: C) composite adjuvant formed based on covalent interaction |
| CN117771361A (en) * | 2024-02-27 | 2024-03-29 | 天津中逸安健生物科技有限公司 | Lipid nanoadjuvant of polyinosinic acid-polycytidylic acid compound, and preparation method and application thereof |
Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1404875A (en) * | 2002-11-22 | 2003-03-26 | 北京绿竹生物技术有限责任公司 | B-type hepatitis vaccine |
| CN1997391A (en) * | 2005-06-08 | 2007-07-11 | 申益皮卡生物技术有限公司 | Polyinosinic acid-polycytidylic acid-based adjuvant |
| CN101883585A (en) * | 2007-12-07 | 2010-11-10 | 多见尔有限公司 | A powerful vaccine composition comprising a lipopeptide and poly I:C as an adjuvant |
-
2012
- 2012-07-06 CN CN2012102322373A patent/CN102949717A/en active Pending
Patent Citations (3)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN1404875A (en) * | 2002-11-22 | 2003-03-26 | 北京绿竹生物技术有限责任公司 | B-type hepatitis vaccine |
| CN1997391A (en) * | 2005-06-08 | 2007-07-11 | 申益皮卡生物技术有限公司 | Polyinosinic acid-polycytidylic acid-based adjuvant |
| CN101883585A (en) * | 2007-12-07 | 2010-11-10 | 多见尔有限公司 | A powerful vaccine composition comprising a lipopeptide and poly I:C as an adjuvant |
Cited By (12)
| Publication number | Priority date | Publication date | Assignee | Title |
|---|---|---|---|---|
| CN103386128A (en) * | 2013-07-02 | 2013-11-13 | 中国食品药品检定研究院 | Tuberculosis subunit vaccine containing unite adjuvant |
| CN103386128B (en) * | 2013-07-02 | 2017-02-08 | 中国食品药品检定研究院 | Tuberculosis subunit vaccine containing unite adjuvant |
| CN107556166A (en) * | 2016-06-30 | 2018-01-09 | 中国科学院化学研究所 | Polyhydroxylated fullerene and preparation method thereof |
| CN107456574A (en) * | 2017-08-14 | 2017-12-12 | 山东大学 | One kind is without adjuvant single dose HBsAg nanogel vaccines and preparation method thereof |
| CN107456574B (en) * | 2017-08-14 | 2020-07-24 | 山东大学 | Adjuvant-free single-dose HBsAg nano gel vaccine and preparation method thereof |
| CN113423423A (en) * | 2018-11-13 | 2021-09-21 | 变异生物技术公司 | Immunogenic compositions for the treatment of hepatitis B |
| CN109355334A (en) * | 2018-12-06 | 2019-02-19 | 江苏省农业科学院 | A method of preparing poly IC |
| CN109876140A (en) * | 2019-03-19 | 2019-06-14 | 何勇刚 | A kind of vaccine and its preparation method and application for treating chronic hepatitis B |
| CN113398261A (en) * | 2021-07-16 | 2021-09-17 | 山西医科大学 | Application of poly-sarcosine in preparing biological preparation for improving HBsAg positive mother infant hepatitis B vaccine response level |
| CN116115748A (en) * | 2023-01-17 | 2023-05-16 | 大连理工大学 | Al-Poly (I: C) composite adjuvant formed based on covalent interaction |
| CN117771361A (en) * | 2024-02-27 | 2024-03-29 | 天津中逸安健生物科技有限公司 | Lipid nanoadjuvant of polyinosinic acid-polycytidylic acid compound, and preparation method and application thereof |
| CN117771361B (en) * | 2024-02-27 | 2024-06-07 | 天津中逸安健生物科技有限公司 | Lipid nanoadjuvant of polyinosinic acid-polycytidylic acid compound, and preparation method and application thereof |
Similar Documents
| Publication | Publication Date | Title |
|---|---|---|
| CN102949717A (en) | Novel hepatitis B vaccine preparation containing poly I:C adjuvant | |
| JPH085804B2 (en) | Hepatitis A and B mixed adjuvant vaccine | |
| CN105920599A (en) | Vaccine with cationic liposome DOTAP as adjuvant and preparation method thereof | |
| US10052378B2 (en) | Pharmaceutical compositions comprising CpG oligonucleotides | |
| US20190134191A1 (en) | ALHYDROGEL-SODIUM CHLORIDE COMPOUND IMMUNOLOGIC ADJUVANT, PREPARATION METHOD AND USE THEREOF (As Amended) | |
| CN107488235B (en) | A kind of preparation and application of new enhanced antigen joint polypeptid induction liver cancer-specific CTL cell | |
| CN104338132A (en) | Viral immunotherapy drug compound and purpose thereof | |
| KR20240128988A (en) | Aluminum-manganese composite nanocrystals, and method for producing the same and use thereof | |
| KR20070110513A (en) | Adjuvant composition comprising aluminium phosphate and 3d-mpl | |
| US20180246105A1 (en) | Immunological test kit for evaluating vaccine efficacy and storage method thereof | |
| CN105194668A (en) | Method for preparing conjugate of gp96 protein and PD1 antibody, and application of conjugate | |
| CN115252772A (en) | Application of artificially synthesized deoxyoligonucleotide containing CpG single strand in vaccine | |
| CN115120713A (en) | Aluminum hydroxide-CpG oligonucleotide-polypeptide composite adjuvant, vaccine, preparation method and application | |
| CN108210922A (en) | A kind of novel cellular immunity enhancing adjuvant | |
| CN108567977A (en) | A kind of immunopotentiator, immunotherapy medicaments composition and its preparation and purposes | |
| CN107693788B (en) | Pharmaceutical composition for preventing or treating hepatitis B and application thereof | |
| TWI827732B (en) | Pharmaceutical preparations for treating hepatitis B and preparation methods and uses thereof | |
| CN102349996A (en) | Human papilloma virus pharmaceutical composition and application thereof | |
| CN104338129B (en) | Application of Rapamycin as vaccine adjuvant and preparation method thereof | |
| CN109876140A (en) | A kind of vaccine and its preparation method and application for treating chronic hepatitis B | |
| CN101130074B (en) | Medicament for treating and/or preventing hepatitis B | |
| JP2013505971A (en) | Hepatitis B virus antigen preparation for cellular stimulation after therapeutic immunization | |
| CN101675994B (en) | Therapy vaccine preparation | |
| CN101422609A (en) | Use of lentinan as hepatic C virus nucleic acid vaccine immunologic adjavant | |
| JP2022513001A (en) | Immunogenic composition for the treatment of hepatitis B |
Legal Events
| Date | Code | Title | Description |
|---|---|---|---|
| C06 | Publication | ||
| PB01 | Publication | ||
| C10 | Entry into substantive examination | ||
| SE01 | Entry into force of request for substantive examination | ||
| C02 | Deemed withdrawal of patent application after publication (patent law 2001) | ||
| WD01 | Invention patent application deemed withdrawn after publication |
Application publication date: 20130306 |