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WO2018080213A1 - Composition immunogène multivalente ayant un titre d'igg augmenté, et son utilisation - Google Patents

Composition immunogène multivalente ayant un titre d'igg augmenté, et son utilisation Download PDF

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Publication number
WO2018080213A1
WO2018080213A1 PCT/KR2017/011959 KR2017011959W WO2018080213A1 WO 2018080213 A1 WO2018080213 A1 WO 2018080213A1 KR 2017011959 W KR2017011959 W KR 2017011959W WO 2018080213 A1 WO2018080213 A1 WO 2018080213A1
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serotype
composition
capsular
serotypes
derived
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Korean (ko)
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김태현
제훈성
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LG Chem Ltd
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LG Chem Ltd
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    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K39/02—Bacterial antigens
    • A61K39/09—Lactobacillales, e.g. aerococcus, enterococcus, lactobacillus, lactococcus, streptococcus
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/555—Medicinal preparations containing antigens or antibodies characterised by a specific combination antigen/adjuvant
    • A61K2039/55505—Inorganic adjuvants
    • A—HUMAN NECESSITIES
    • A61—MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61K—PREPARATIONS FOR MEDICAL, DENTAL OR TOILETRY PURPOSES
    • A61K39/00—Medicinal preparations containing antigens or antibodies
    • A61K2039/60—Medicinal preparations containing antigens or antibodies characteristics by the carrier linked to the antigen
    • A61K2039/6031—Proteins

Definitions

  • the present invention relates to a polyimmunogenic composition having improved IgG titer and its use, specifically to a vaccine composition for preventing pneumococcal disease comprising a capsular polysaccharide-carrying protein conjugate and immunogenicity against pneumococcus It relates to a composition.
  • Streptococcus pneumonia is a leading cause of meningitis, pneumonia and severely invasive diseases in infants and children around the world. More than 1.6 million people die each year from pneumococcal disease (2008 International Health Organization), and the incidence of invasive infectious diseases caused by pneumococcal in children under 5 years old and elderly people 65 years or older with low immunity high.
  • Pneumococci are classified into more than 90 serotypes according to the structural and immunological characteristics of the capsular polysaccharides, the main pathogenic factors surrounding them, of which 20 to 80% in humans It is known to be associated with pathogenicity.
  • the only host of pneumococci is humans, and they usually exist in colonies in the nasopharynx of healthy normal people (20-40% in infants, 5-10% in adults).
  • the US Centers for Disease Control and Prevention (CDC) estimated that approximately 2.1 million children under 5 years of age died from pneumonia, and 1.2 million of them died from developing countries alone annually.
  • pneumococcal meningitis and sepsis are reported to be about 3,000 and 50,000 cases per year, respectively (Peters TR, Poehling KA et al . JAMA 2007; 297: 1825-6; Invasive pneumococcal disease). .
  • pneumoACTION which is a database of pneumococcal disease, showed that 24,047 cases of pneumococcal infections occurred in Korean children in 2000 and 47 of them died (www.pneumoadip.org).
  • pneumococci are the most common cause of invasive infections (43.7%) in infants aged 3 to 59 months. appear.
  • multidrug resistant bacteria that are resistant to not only penicillin but also to three or more drugs are increasing, further increasing the difficulty of treating pneumococcal infectious diseases. Therefore, the need for pneumococcal vaccination for children and the elderly, which is a high risk group of pneumococcal infectious diseases, has been continuously raised.
  • multivalent pneumococcal polysaccharide vaccines have been developed and approved since 1977, and these capsular polysaccharide vaccines have proven useful in preventing pneumococcal disease in elderly and high-risk patients.
  • the immune system since the maturity of the immune system is lower than that of adults, when only the polysaccharide vaccine is received, the immune system does not recognize the polysaccharide antigen as an external invading factor, so it is difficult to expect a role as a vaccine.
  • a 7-valent pneumococcal conjugate vaccine a capsular polysaccharide-protein conjugated vaccine conjugated with a carrier protein that increases immunogenicity to polysaccharide antigens ( 7vPnC, Prevenar ® (Prevenar ®)) has been used in the development, has been reported to be effective for the prevention of invasive disease and otitis media in infants and children in many materials.
  • 7vPnC Prevenar ®
  • Prevenar ® Prevenar ®
  • the use of the 7-valent vaccine induced a decrease in invasive disease caused by the vaccine serotypes used in the vaccine, but together with the relative pneumococcus caused by some non-vaccine serotypes due to serotype replacement.
  • the capsular polysaccharide in order to provide a wider coverage - a protein conjugate vaccine in renal flow Rix ® (Synflorix ®), and the free the base serotypes of vena ®
  • a 13 add serotypes 6 kinds of pneumococcal conjugate vaccine Prevenar 13 ® (Prevenar13 ®) one of these has been developed, but presently on the market, resulting tried monitoring pneumococcal disease onset into after market, also 10 a and 13 a serotype other than the non-vaccine serotypes included in the vaccine Serotype replacement was observed (Weinberger DM et al . 2011; Lancet, 378: 9807,1962; Serotype replacement in disease following pneumococcal vaccination: A discussion of the evidence).
  • the pneumococcal conjugate vaccine of the present invention comprises 15 or more capsular polysaccharide antigens, specifically 17 capsular polysaccharide antigens, and may have broader serotype coverage than previously known pneumococcal conjugate vaccines. It can have an excellent effect in preventing.
  • One object of the present invention is to provide novel multivalent immunogenic compositions.
  • one object of the present invention includes capsular polysaccharide-carrying protein conjugates, wherein the conjugates are Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B.
  • Vaccine composition for the prevention of pneumococcal disease wherein each of 15 capsular polysaccharides derived from 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F and 23F is covalently conjugated to a carrier protein, and It is to provide an immunogenic composition for pneumococcal.
  • one object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F,
  • a vaccine composition for preventing pneumococcal disease wherein each of 15 capsular polysaccharides derived from 14, 15B, 18C, 19A, 19F, and 23F is covalently conjugated to a carrier protein, wherein the composition is derived from serotype 1
  • Capsular polysaccharides derived from serotypes 6A, 14, 19A and 19F each had a content ratio of greater than 0.25 and 0.95 or less, and capsular polysaccharides from serotype 6B exceeded 0.5 and a content ratio of 1.9 or less.
  • Eggplant is to provide a vaccine composition for the prevention of pneumococcal disease, and an immunogenic composition against pneumococcal.
  • Another object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C.
  • Vaccine composition for the prevention of pneumococcal disease and immunogenic composition for pneumococcal, wherein each of 17 capsular polysaccharides derived from 19A, 19F, 22F, 23F and 33F is covalently conjugated to a carrier protein.
  • another object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14
  • a vaccine composition for preventing pneumococcal disease wherein each of 17 capsular polysaccharides from 15B, 18C, 19A, 19F, 22F, 23F, and 33F is covalently conjugated to a carrier protein, wherein
  • the capsular polysaccharides derived from serotypes 6A, 14, 19A and 19F each had a content ratio of greater than 0.25 and 0.95 or less, and the capsular polysaccharides derived from serotype 6B were greater than 0.5 and 1.9 or less, compared to the capsular polysaccharides derived from type 1, respectively.
  • Still another object of the present invention is to provide a method for preventing pneumococcal disease by administering the vaccine composition or immunogenic composition to a subject in need thereof.
  • Another object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 15 kinds of capsular polysaccharides from 18C, 19A, 19F and 23F are covalently conjugated to a carrier protein, to provide a use for the preparation of a vaccine composition for the prevention of pneumococcal disease.
  • the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 15 kinds of capsular polysaccharides from 18C, 19A, 19F and 23F are covalently conjugated to a carrier protein, to provide a use for the preparation of a vaccine composition for the prevention of pneumococcal disease.
  • another object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F,
  • a vaccine composition for preventing pneumococcal disease wherein each of 15 capsular polysaccharides derived from 14, 15B, 18C, 19A, 19F, and 23F is covalently conjugated to a carrier protein, wherein the composition is derived from serotype 1
  • the capsular polysaccharides derived from serotypes 6A, 14, 19A and 19F each had a content ratio of greater than 0.25 and 0.95 or less, and the capsular polysaccharides from serotype 6B exceeded 0.5 and a content ratio of 1.9 or less.
  • Another object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 17 types of capsular polysaccharides derived from 18C, 19A, 19F, 22F, 23F and 33F are each covalently conjugated to a carrier protein, providing a use for the preparation of a vaccine composition for the prevention of pneumococcal disease It is.
  • another object of the present invention includes a capsular polysaccharide-carrying protein conjugate, wherein the conjugate is Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F,
  • a vaccine composition for the prevention of pneumococcal disease wherein each of 17 capsular polysaccharides from 14, 15B, 18C, 19A, 19F, 22F, 23F, and 33F is covalently conjugated to a carrier protein, wherein
  • the capsular polysaccharides derived from serotypes 6A, 14, 19A and 19F, respectively had a content ratio of greater than 0.25 and 0.95 or less, and the capsular polysaccharides derived from serotype 6B were greater than 0.5, compared to the capsular polysaccharides derived from serotype 1, respectively. It is to provide a use for use in the manufacture of a vaccine composition for the prevention of pneumococcal disease, having the following content ratio.
  • One aspect of the present invention for achieving the above object is a novel multivalent immunogenic composition.
  • one embodiment of the present invention is a vaccine composition for the prevention of pneumococcal disease comprising 15 species of capsular polysaccharide-carrying protein conjugates.
  • Another aspect of the present invention for achieving the above object is a vaccine composition for the prevention of pneumococcal disease comprising 17 species of capsular polysaccharide-carrying protein conjugate.
  • the 15 conjugates are 15 capsular polysaccharides derived from Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F and 23F. It refers to each covalently conjugated to the carrier protein.
  • each of the capsular polysaccharides derived from Streptococcus pneumoniae serotypes 22F and 33F is covalently conjugated to each carrier protein. It may further comprise a conjugate of species.
  • the composition is a vaccine composition comprising 15 different polysaccharide-protein conjugates, each conjugate comprising a capsular polysaccharide from Streptococcus pneumoniae of different serotypes conjugated to a carrier protein, wherein the capsular polysaccharide is Is prepared from serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F and 23F.
  • composition is also a vaccine composition comprising 17 different polysaccharide-protein conjugates, in addition to the 15 conjugates described above, capsular polysaccharides derived from Streptococcus pneumoniae serotypes 22F and 33F covalently bound to their respective transport proteins. It is prepared by further comprising a conjugate to the conjugate.
  • the overall IgG titer was confirmed as the increase in the number of valent valences, 15 or 17 valences, and 15 or 17 valent immunogenic compositions were prepared.
  • the IgG titer can be actually increased by simultaneously controlling the conjugate content of serotypes 6A, 6B, 14, 19A and 19F.
  • Capsular polysaccharides derived from serotypes 6A, 14, 19A, and 19F as compositions of the present invention may each have a content ratio of greater than 0.25 and less than or equal to 0.95 relative to the capsular polysaccharides derived from serotype 1, specifically 0.3 to 0.95, 0.3 to It can represent a content ratio of 0.9, 0.4 to 0.95, 0.4 to 0.9, 0.45 to 0.95, 0.45 to 0.9, 0.5 to 0.95 or 0.5 to 0.9.
  • serotype 1-derived capsular polysaccharide serotype 6A, 14, 19A, or 19F-derived capsular polysaccharide was 1: 0.3 to 0.95, 1: 0.3 to 0.9, 1: 0.4 to 0.95, 1: 0.4 to 0.9, 1: 0.45 to It may represent a content ratio of 0.95, 1: 0.45 to 0.9, 1: 0.5 to 0.95 or 1: 0.5 to 0.9.
  • the capsular polysaccharide derived from serotype 6B may have a content ratio of more than 0.5 and 1.9 or less than the capsular polysaccharide derived from serotype 1.
  • the content ratio may be 0.6 to 1, 9, 0.6 to 1.8, 0.8 to 1.9, 0.8 to 1.8, 0.9 to 1.9, 0.9 to 1.8, 1.0 to 1.9, or 1.0 to 1.8. That is, the capsular polysaccharide derived from serotype 1: the capsular polysaccharide derived from serotype 6B is 1: 0.6 to 1.9, 1: 0.6 to 1.8, 1: 0.8 to 1.9, 1: 0.8 to 1.8, 1: 0.9 to 1.9, 1: 0.9 It may represent a content ratio of ⁇ 1.8, 1: 1 ⁇ 1.9, or 1: 1: 1.8.
  • polysaccharides other than the above-mentioned capsular polysaccharides are equivalent to serotype 1 derived capsular polysaccharides (1: 1 ratio).
  • Immunogenic compositions provided by the present invention comprising the capsular polysaccharides in the same amount as described above, by the specific content ratio of the five serotypes, serotypes 1, 3, 4, 5, 6A, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F and 23F (additionally, 22F and 33F for 17 valent) may exhibit superior immunogenicity compared to immunogenic compositions in the same amount.
  • composition is specifically serotype 6A when serotype 1, 3, 4, 5, 7F, 9V, 12F, 15B, 18C and 23F derived capsular polysaccharides are respectively 4.4 ⁇ g / mL (eg, 2.2 ⁇ g / dose), respectively.
  • 14, 19A and 19F are greater than 1.1 ⁇ g / mL, respectively, 4.18 ⁇ g / mL, 1.32 ⁇ g / mL to 4.18 ⁇ g / mL, 1.32 ⁇ g / mL to 3.96 ⁇ g / mL, 1.76 ⁇ g / mL to 4.18 ⁇ g / mL, 1.76 ⁇ g, respectively.
  • concentrations of / mL to 3.96 ⁇ g / mL, 1.98 ⁇ g / mL to 4.18 ⁇ g / mL, 1.98 ⁇ g / mL to 3.96 ⁇ g / mL, 2.2 ⁇ g / mL to 4.18 ⁇ g / mL, or 2.2 ⁇ g / mL to 3.96 ⁇ g / mL Serotype 6B may be present in an amount greater than 2.2 ⁇ g / mL, 8.36 ⁇ g / mL, 2.64 ⁇ g / mL to 8.36 ⁇ g / mL, 2.64 ⁇ g / mL to 7.92 ⁇ g / mL, 3.52 ⁇ g / mL to 8.36 ⁇ g / mL, 3.52 ⁇ g / mL to 7.92 ⁇ g / mL, 3.96 ⁇ g / mL to 8.36 ⁇ g / mL, 3.
  • the composition may additionally include the capsular polysaccharide derived from serotype 22F and the capsular polysaccharide derived from 33F in the same amount as the capsular polysaccharide derived from serotype 1.
  • pneumococcal pneumonia accounts for about 50% of all pneumonia, severe chills, fever, cough and chest pain, sputum is often bloody, complications that can cause pleurisy, meningitis, endocarditis, peritonitis ( Stein GE et al. 2001; Diagn. Microbiol.Infect, Dis 39: 181; Comparative serum bactericidal activity of clarithromycin and azithromycin against macrolide-sensitive and resistant strains of Streptococcus pneumoniae ).
  • pneumococcus in the present invention refers to Streptococcus pneumoniae and is generally a commensal organism that colonizes the mucosal surface of the human nasopharynx. If the host's factor allows access to the lower respiratory tract of the organism, then a vigorous inflammatory response follows, which causes dense consolidation when the alveolar space fills the exudate, resulting in pneumonia May cause.
  • the pneumococci can synthesize more than 90 structurally unique capsular polysaccharides, and the serotypes of pneumococci are classified according to the structural and immunological characteristics of these capsular polysaccharides.
  • the immune response may be different depending on the type of capsular polysaccharide, that is, the serotype of pneumococcal from which capsular polysaccharide is derived.
  • the vaccine composition of the present invention specifically contains 15 capsular polysaccharides derived from Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F and 23F. It can be prepared using.
  • 17-valent forms can be prepared, including capsular polysaccharides derived from serotypes 22F and 33F.
  • the capsular polysaccharide is recognized as an antigen when administered in the body, so that it can produce an antibody against it, thereby preparing a vaccine composition for preventing pneumococci.
  • the term “antigen” refers to a substance that can specifically induce an immune response when the substance invades the body.
  • 15 kinds of capsular polysaccharides derived from Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F, and 23F or
  • 17 capsular polysaccharides may act as antigens, respectively.
  • the capsular polysaccharide can be prepared by standard techniques known to those skilled in the art, and are not particularly limited in its method.
  • the capsular polysaccharide can be reduced in size through hydrolysis to reduce viscosity and induce effective immunogenicity.
  • 17 different serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F, 22F, 23F and 33F) Streptococcus pneumoniae with lysates were dissolved using sodium deoxycholate, respectively, and the polysaccharides bound to the cells were released.
  • serotypes 1, 3, 4, 5, 6A, 6B, 9V, 12F, 15B, 18C, 19A, 19F, 22F, and 23F are capable of ion bonding with CTAB (cetyltrimethylammonium bromide) to perform CTAB process 3 serotypes 7F, 14 and 33F that do not react with CTAB were purified using an aluminum phosphate gel (Algel) solution.
  • CTAB cetyltrimethylammonium bromide
  • the conjugate protein and the capsular polysaccharide are conjugated. It was prepared and used.
  • carrier protein refers to a protein that can be covalently conjugated with the capsular polysaccharide to increase the immunogenicity of the polysaccharide antigen.
  • the carrier protein may be conjugated with the capsular polysaccharide through a standard conjugation method, and the capsular polysaccharide-carrying protein conjugate formed therefrom may be one or a plurality of capsular polysaccharides conjugated to one carrier protein.
  • the carrier protein may be, but is not limited to, a protein that is specifically nontoxic, nonreactive, and obtainable in sufficient amount and purity.
  • the carrier protein in the present invention is not limited thereto, but may be, for example, CRM197.
  • CRM197 in the present invention is a non-toxic variant of diphtheria toxin (ie, toxoid) isolated from the culture of Corynebacterium diphtheriae strain C7 ( ⁇ 197).
  • CRM197 can be purified via ultrafiltration, ammonium sulphate precipitation and ion exchange chromatography.
  • the CRM197 may be recombinantly prepared according to US Pat. No. 5,614,382.
  • the carrier protein is also inactivated, such as tetanus toxoid, pertussis toxoid, cholera toxoid (WO2004 / 083251), E. coli LT, E. coli ST or Pseudomonas aeruginosa .
  • Bacterial toxins may be a variant of diphtheria toxins such as CRM173, CRM228, CRM45, but is not limited thereto.
  • OMPC outer complex c
  • porin porin
  • transferrin binding protein pneumolysin
  • pneumococcal surface protein A PspA
  • pneumococcal adhesin protein PsaA
  • group C5a peptidase from A or Group B streptococci or purified protein derivatives of Haemophilus influenzae protein D, ovalbumin, keyhole limpet hemocyanin (KLH), bovine serum albumin (BSA), tuberculin (PPD)
  • KLH keyhole limpet hemocyanin
  • BSA bovine serum albumin
  • PPD tuberculin
  • diphtheria toxins such as CRM173, CRM228, CRM45, but is not limited thereto.
  • the conjugate may have a structure in which the capsular polysaccharide and the carrier protein are connected by -O-C (NH) -NH- group using a cyanylation method.
  • the vaccine compositions of the present invention are all compared to vaccine compositions conjugated by known reductive amination.
  • the serotype may show significantly better IgG titers.
  • the cyanation method may be appropriately performed by those skilled in the art through a known method, for example, may be performed using CDAP (1-cyano-4-dimethylaminopyridinium tetrafluoroborate) or CNBr, but is not limited thereto.
  • purified capsular polysaccharides can be chemically activated and each chemically activated capsular polysaccharide can be conjugated to the carrier protein one by one to form a glycoconjugate.
  • Cyanation activity by treatment with CDAP (1-cyano-4-dimethylaminopyridinium tetrafluoroborate) converts the hydroxyl group of the capsular polysaccharide to a cyanate group, thereby covalently binding to the amino group of the carrier protein CRM197.
  • CDAP 1-cyano-4-dimethylaminopyridinium tetrafluoroborate
  • the cyanation reaction by the CDAP may be specifically terminated by adding 3 molar equivalents of glycine (glycine) solution to 1 molar equivalent of CDAP and adjusting the pH to 9.0, but is not limited thereto.
  • the reaction solution and reaction conditions can be adjusted accordingly.
  • the capsular polysaccharide-carrying protein conjugates obtained can be purified by various methods. Examples of these methods include concentration / dialysis filtration processes, column chromatography and multilayer filtration. Purified polysaccharide-protein conjugates can be mixed and formulated into vaccine compositions of the invention and used respectively. Formulations of vaccine compositions of the invention can be carried out using methods known in the art. For example, compositions may be prepared by formulating individual capsular polysaccharide-carrying protein conjugates with a physiologically acceptable vehicle. Examples of such vehicles may include, but are not limited to, water, buffered saline, polyols (eg, glycerol, propylene glycol, liquid polyethylene glycols) or dextrose solutions.
  • a physiologically acceptable vehicle examples of such vehicles may include, but are not limited to, water, buffered saline, polyols (eg, glycerol, propylene glycol, liquid polyethylene glycols) or dextrose solutions.
  • the term “vaccine” refers to a biological agent containing an antigen that immunizes a living body, and refers to an immunogen or antigenic substance that immunizes the living body by administering it to a human or an animal to prevent infection.
  • the vaccine composition may further include one or more selected from the group consisting of an adjuvant, a preservative, a buffer, a cryoprotectant, a salt, a divalent cation, a nonionic detergent, and a free radical oxidation inhibitor.
  • adjuvant in the present invention refers to a substance used to increase the immunogenicity of the immunogenic composition of the present invention.
  • the adjuvant is often provided to enhance the immune response, which is well known to those skilled in the art.
  • adjuvants suitable for increasing the effectiveness of the vaccine composition of the present invention include, but are not limited to:
  • aluminum salts eg, aluminum hydroxide, aluminum phosphate, aluminum sulfate, etc.
  • Oil-in-water emulsion formulations (with or without muramyl peptide (defined below) or other specific immune stimulants such as bacterial cell wall components), for example (a) MF59 (WO 90/14837): Contains 5% Squalene, 0.5% Tween 80 and 0.5% Span 85 (optionally contains varying amounts of MTP-PE (although not required, see below)), Model 110Y Micro Formulated into submicron particles using a microfluidizer such as a microfluidizer (Microfluidics, Newton, Mass.), (B) SAF: 10% squalene, 0.4% Tween 80, 5% Pluronic -Block polymer L121 and thr-MDP (see below), microfluidized with a submicron emulsion, or vortexed to form a large particle size emulsion, and (c) Ribi TM adjuvant System (RAS) (Corixa, Hamilton, MT): 2% squalene, 0.
  • Tween 80 and from the group consisting of 3-O-deacylated monophosphoryl lipid A (MPL TM) (Corixa), trehalose dimicholate (TDM) and cell wall backbone (CWS) described in US Pat. No. 4,912,094. Contains at least one bacterial cell wall component, preferably MPL + CWS (Detox TM);
  • Saponin adjuvant such as Quill A or STIMULON TM QS-21 (Antigenics, Framingham, MA, US Pat. No. 5,057,540), may be used or produced from particles such as ISCOM ( Immunostimulatory complexes));
  • cytokines such as interleukins (eg, IL-1, IL-2, IL-4, IL-5, IL-6, IL-7, IL 12, IL-15, IL-18, etc.) Interferon (eg gamma interferon), granulocyte macrophage colony stimulating factor (GM-CSF), macrophage colony stimulating factor (MCSF), tumor necrosis factor (TNF), costimulatory molecules B7-1 and B7-2, and the like;
  • interleukins eg, interleukins (eg, IL-1, IL-2, IL-4, IL-5, IL-6, IL-7, IL 12, IL-15, IL-18, etc.)
  • Interferon eg gamma interferon
  • GM-CSF granulocyte macrophage colony stimulating factor
  • MCSF macrophage colony stimulating factor
  • TNF tumor necrosis factor
  • costimulatory molecules B7-1 and B7-2 costimulatory molecules
  • Wild type cholera toxin (CT) or mutant cholera toxin eg WO2002 / 098368
  • glutamic acid at amino acid position 29 according to WO2000 / 18434 is substituted with another amino acid, specifically histidine.
  • WO2002 / 098369 pertussis toxin (PT), or E. coli heat-labile toxin (LT), in particular LT-K63, LT-R72, CT-S109, PTK9 / G129 (WO93 / 13302 and WO92) / 19265), detoxified mutants of bacterial ADP-ribosylated toxins;
  • PT pertussis toxin
  • LT E. coli heat-labile toxin
  • LT-K63, LT-R72, CT-S109, PTK9 / G129 WO93 / 13302 and WO92
  • the muramyl peptides include N-acetyl-muramil-L-threonyl-D-isoglutamine (thr-MDP), N-acetyl-normuramyl-L-alanine-2- (1'-2 'dipalmityl) -sn-glycero-3-hydroxyphosphoryloxy) -ethylamine (MTP-PE) and the like, but is not limited thereto.
  • the aluminum salt adjuvant may be an aluminum-precipitated vaccine or an aluminum-adsorbed vaccine.
  • Aluminum salts include hydrated alumina, alumina hydrate, alumina trihydrate (ATH), aluminum hydrate, aluminum trihydrate, alhydrogel, Superfos, amphogel, aluminum hydroxide, aluminum hydroxyphosphate adjuvant (APA), Amorphous alumina, and the like, but is not limited thereto.
  • APA refers to a suspension of aluminum hydroxyphosphate.
  • Aluminum chloride and sodium phosphate are mixed in a ratio of 1: 1, aluminum hydroxyphosphate sulfate is precipitated, and the precipitate is made to be 2 to 8 ⁇ m using a high shear mixer, and then prepared by dialysis and sterilization with physiological saline solution.
  • commercially available Al (OH) 3 eg Alhydrogel or Superfos
  • OH Alhydrogel or Superfos
  • 50 to 200 g of protein can be adsorbed per mg of aluminum hydroxide, and this ratio is dependent on the pH of the protein and the pH of the solvent.
  • Low pI proteins bind more strongly than proteins with high pi.
  • Aluminum salts can form antigen reservoirs that slowly release antigens for two to three weeks to nonspecifically activate macrophages, complement, and innate immune mechanisms.
  • preservative means an anti-viral and / or antimicrobial agent that inhibits the growth of microorganisms in the vaccine composition, for example, chimerosal, phenoxyethanol, 2-phenoxyethanol. It may be, but is not limited to, formaldehyde, or mixtures thereof, all conventional preservatives used in the art may be used.
  • the vaccine composition may comprise one or more physiologically acceptable buffers.
  • the buffer may have buffering capacity at pH 4.0 to 10.0, specifically, pH 5.0 to 9.0, more specifically pH 6.0 to 8.0.
  • the buffer may be selected from the group consisting of TRIS, acetate, glutamate, lactate, maleate, tartrate, phosphate, citrate, carbonate, glycinate, histidine, glycine, succinate, triethanolamine buffer.
  • the buffer may be selected from buffers suitable for USP.
  • buffers include monobasic acids such as acetic acid, benzoic acid, gluconic acid, glyceric acid, lactic acid; Dibasic acids such as aconitic acid, adipic acid, ascorbic acid, carbonic acid, glutamic acid, malic acid, succinic acid, tartaric acid; Polybasic acids such as citric acid and phosphoric acid; It may be selected from the group consisting of ammonia, diethanolamine, glycine, triethanolamine, TRIS and the like.
  • the vaccine composition of the present invention may include a nonionic detergent.
  • a nonionic detergent for example, polysorbate 20 and polysorbate 80 in polyoxyethylene sorbitan esters (commonly called Tweens); Copolymers of ethylene oxide (EO), propylene oxide (PO), butylene oxide (BO) (eg DOWFAX TM); Oxoxynols having a different repeating number of oxy-1,2-ethanediyl groups, especially ostoxynol-9 (Triton-100); Ethylphenoxypolyethoxyethanol (IGEPAL CA-630 / NP-40); Phospholipids such as lecithin; Nonylphenol ethoxylates such as NP series; Polyoxyethylene fatty acid ethers (Brij surfactants) derived from lauryl, cetyl, stearyl, oleyl alcohols, in particular triethyleneglycol monolauryl ether (Brij 30); Sorbitan ethers, also known
  • Tween 80 may be included in the emulsion and a mixture of nonionic detergents such as Tween 80 / Span 85 may be used.
  • Combinations of polyoxyethylene sorbitan esters such as Tween 80 with octocinols such as Triton X-100 are also suitable, and combinations of Laureth 9 with Tween and or octosinol are also useful.
  • polyoxyethylene sorbitan esters such as Tween 80 may be used in an amount of 0.01% to 1% (w / v), in particular 0.1%; Octylphenoxy polyoxyethanol or nonylphenoxy polyoxyethanol (such as Triton X-100) may range from 0.001% to 0.1%, in particular from 0.005% to 0.02%;
  • the polyoxyethylene ether eg laureth 9 may comprise 0.1% to 20%, preferably 0.1% to 10%, in particular 0.1% to 1% or about 0.5%.
  • composition of the present invention may be formulated in a single dose dose vial, multiple dose dose vial or prefilled syringe form, and may further comprise a physiologically acceptable carrier.
  • Physiologically acceptable carriers used in liquid formulations include aqueous or non-aqueous solvents, suspensions, emulsions, oils. Examples of non-aqueous solvents are propylene glycol, polyethylene glycol, ethyl oleate.
  • Aqueous carriers include water, alcohol / aqueous solvents, emulsions or suspensions, physiological saline, buffer solutions.
  • oils include vegetable or animal oils, peanut oils, soybean oils, olive oils, sunflower oils, synthetic oils such as liver oils, marine oils, and lipids derived from milk or eggs.
  • Vaccine compositions of the present invention may be isotonic, hypertonic or hypotonic, and pharmaceutical compositions administered by infusion or injection are preferably but is not limited to isotonicity. On the other hand, isotonicity or hypertension may be advantageous for storage of the composition. If the vaccine composition is hypertonic, it can be diluted to isotonic prior to administration. Isotonic agents for dilution can be ionic tonicity agents such as salts or nonionic tonicity agents such as carbohydrates. Ionic isotonic agents include, but are not limited to, sodium chloride, calcium chloride, potassium chloride, magnesium chloride, and the like. Nonionic isotonic agents include, but are not limited to, sorbitol, glycerol, and the like.
  • the vaccine composition may further include aluminum element and sodium chloride, but is not limited thereto.
  • the vaccine composition may or may not contain a preservative depending on the purpose and use thereof.
  • the vaccine composition according to the present invention can be used to protect a subject susceptible to pneumococcal and to prevent pneumococcal disease by administering a pharmaceutically effective amount in a systemic or mucosal route.
  • prevention of the present invention refers to any action that inhibits or delays the infection caused by pneumococcal by administration of the vaccine composition of the present invention.
  • a “pharmaceutically effective amount” refers to a dosage required to elicit an antibody that is capable of significantly reducing the probability of infection or the severity of infection.
  • administration of the present invention refers to the introduction of certain substances into an individual in any suitable way.
  • the vaccine composition of the present invention may be administered by inhalation route through oral, nasal, rectal, transdermal or aerosol, but may be administered by bolus or infused slowly, but is not limited thereto.
  • the administration may be by injection via an intramuscular, intraperitoneal, intradermal or subcutaneous route; Or mucosal administration to the oral / digestive tract, airway or urogenital tract.
  • intranasal administration can be used for the treatment of pneumonia or otitis media, in which case more effective prevention of nasopharyngeal carriers of pneumococci can attenuate the infection at an early stage.
  • the term “individual” of the present invention means a living organism to which a pathogen can be infected, and in particular, may be a higher vertebrate, and more specifically, a mammal, but is not particularly limited thereto.
  • composition of the present invention may be administered in a single inoculation, or two, three, four or more times at appropriate intervals, but is not limited thereto.
  • routine inoculation plans for infants and newborns for invasive diseases caused by Streptococcus pneumoniae can be 2, 4, 6 and 12 to 15 months of age.
  • composition may further comprise one or more proteins from Streptococcus pneumoniae.
  • Streptococcus pneumoniae proteins suitable for inclusion include not only the proteins described in WO2002 / 053761, but also the proteins identified in WO2002 / 083855, all within the scope of the present invention.
  • each polysaccharide in a total of 0.5 mL provided that 6B is 8.8 ⁇ g / mL; About 29.3 ⁇ g CRM197 transport protein; 0.5 mg of elemental aluminum (2 mg aluminum phosphate) adjuvant; About 4.25 mg sodium chloride (without preservatives) or about 3.5 mg (with preservatives); About 295 ⁇ g succinate buffer; About 3 mg of 2-phenoxyethanol and about 60 ⁇ g of formaldehyde were mixed (with preservatives) to prevent pneumococcal disease of the 13-valent (named 'LBVE013') and 17-valent (named 'LBVE017') Vaccine compositions were prepared. In addition, the content of the composition was 100%, and a vaccine composition was prepared separately by adjusting the content of some serotypes 6A, 6B, 14, 19A and 19F.
  • serum levels of rabbits inoculated with the LBVE013 vaccine composition confirmed higher serotype-specific IgG concentrations than prevena13 ® (Table 1), but other serotype-derived capsular polysaccharides were identified.
  • Table 1 serum levels of rabbits inoculated with the LBVE013 vaccine composition
  • the 17 transverse composition was prepared, it was confirmed that the IgG titer for each serotype was significantly decreased (FIG. 1).
  • serotypes 6A, 6B, 14, 19A and 19F conjugates at the same time and re-measuring the IgG titer for the entire serotype, it was confirmed that the IgG titer that decreased with the increase of the valence was restored again.
  • Another aspect of the invention is an immunogenic composition for pneumococci comprising capsular polysaccharide-carrying protein conjugates.
  • the conjugate and pneumococcus are as described above.
  • composition comprising the capsular polysaccharide-carrying protein conjugate of the present invention comprises Streptococcus pneumoniae-derived capsular polysaccharide having 15 or 17 different serotypes, and when administered to the body, it is recognized as an antigen It can be used as an immunogenic composition against pneumococcal to cause an immune response to produce an antibody.
  • Another aspect of the invention is a method of preventing pneumococcal disease by administering the vaccine composition or immunogenic composition to a subject in need thereof.
  • Streptococcus pneumoniae serotypes 1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B for use in the preparation of vaccine compositions for the prevention of pneumococcal disease
  • 15 capsular polysaccharides derived from, 18C, 19A, 19F, and 23F each provide a use of 15 capsular polysaccharide-carrier protein conjugates covalently conjugated to a carrier protein.
  • Another aspect of the present invention provides a capsular polysaccharide derived from Streptococcus pneumoniae serotypes 22F and 33F, in addition to the aforementioned 15 conjugates, for use in the preparation of a vaccine composition for the prevention of pneumococcal disease. It is to provide the use of 17 kinds of capsular polysaccharide-carrier protein conjugate further comprising two conjugates covalently conjugated.
  • the capsular polysaccharide derived from serotypes 6A, 14, 19A, and 19F of the serotype 1-derived capsular polysaccharide in the composition has a content ratio of more than 0.25 and 0.95 or less, and the capsular polysaccharide derived from serotype 6B is 0.5. It may exceed and have a content ratio of 1.9 or less.
  • the capsular polysaccharide derived from serotype 22F and the capsular polysaccharide derived from 33F may be present in the composition in the same amount as the capsular polysaccharide derived from serotype 1.
  • Vaccine compositions immunogenic compositions, and prevention of pneumococcal disease are as described above.
  • the present invention provides a composition comprising pneumococcal capsular polysaccharides having 15 or more different serotypes, specifically, capsular polysaccharides derived from pneumococcal having 17 different serotypes, and in particular, the composition comprises Optimal amounts of polysaccharides can be included to induce good serum IgG titers and functional antibody activity. Therefore, the vaccine composition and the immunogenic composition according to the present invention can be usefully used to prevent diseases caused by pneumococci in infants, infants, children, and adults.
  • FIG. 1 shows that IgG titers are generally lowered in most serotypes as the valence increases from 13-valent (LBVE013) to 17-valent (LBVE017), and 6A, 6B, 14, 19A, and 19F have high IgG titers at 13-valent.
  • LBVE013 13-valent
  • LBVE017 17-valent
  • 6A, 6B, 14, 19A, and 19F have high IgG titers at 13-valent.
  • Increasing the conjugate's dose to 50% simultaneously suggests an overall increase in IgG titer (recovery).
  • 22F and 33F the standard used is different from the rest of the serotypes, so it is drawn separately from other serotypes.
  • Figure 2 shows the amount of serotypes 6A, 6B, 14, 19A, 19F conjugates in the 17-valent composition (LBVE017) simultaneously changed to 100%, 90%, 75%, 50%, 25%, 10%, 1% 17 shows serotypic patterns of change in each of the serotypes.
  • the standard is different from the rest of the serotypes.
  • FIG. 3 is a diagram of dose threshold setting of serotypes 6A, 6B, 14, 19A, and 19F conjugates in 17-valent composition (LBVE017), when all 17-valent (LBVE017) serotypes were all received without dose reduction (100 When each GMT value of the 6A, 6B, 14, 19A, and 19F serotypes of%) is taken as the (lower limit), the range of conjugate amounts of each of the five conjugates above 40 can show higher immunogenicity than the reference. To 95%.
  • Figure 4 shows that in the case of 13-valent (LBVE013), unlike 17-valent compositions, immunogenicity is rather reduced in most serotypes when simultaneously reducing the amount of conjugates of serotypes 6A, 6B, 14, 19A, and 19F. Shows.
  • Streptococcus pneumoniae with 17 different serotypes (1, 3, 4, 5, 6A, 6B, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F, 22F, 23F and 33F) Obtained from the Center for Disease Control and Prevention (USC), a trustee, and prepared a cell bank in the following manner.
  • USC Center for Disease Control and Prevention
  • Streptococcus pneumoniae strains were plated on blood agar medium to isolate single colonies. Among the 10 or more single colonies, a single good growing colony was selected, inoculated and cultured in a liquid medium containing no animal-derived components, and a research cell bank (RCB) containing synthetic glycerol was prepared.
  • RBC research cell bank
  • Master cell bank was prepared by taking out one vial from the cell bank for which the expression of polysaccharides with unique serotypes was confirmed, proliferating the cells in a liquid medium containing no animal-derived components, and adding synthetic glycerol. One vial was taken out of the bank to proliferate cells in a liquid medium containing no animal-derived components, and then synthetic glycerol was added to prepare a cell bank for production. The prepared cell bank was stored at -70 ° C or below for use in the next step.
  • One vial in the cell bank for preparation was thawed and seeded in a liquid medium containing no animal-derived component to start species fermentation.
  • Species were cultured at 37 ⁇ 2 ° C. in an unstirred state until reaching a certain cell concentration (Optical Density, OD 600 ) and reaching the end point of the mid-exponential growth phase.
  • the culture broth obtained from the species culture was inoculated into a fermentor containing a liquid medium containing no animal-derived components to start main fermentation.
  • This culture was then carried out while adjusting the pH of the medium with potassium hydroxide solution at 37 ⁇ 2 ° C. Every two hours, the optimal cell density and glucose concentration in the medium were measured. Fermentation was terminated when glucose in the medium was depleted.
  • Phosphoric acid was added to the sample treated with sodium deoxycholate, and the supernatant was recovered by centrifugation. The recovered supernatant was passed through a depth filter and then buffer exchanged with concentrated and phosphate buffer. After the buffer exchange, the sample was passed through an active carbon filter, and then impurities were removed by the following two methods.
  • CTAB cetyltrimethylammonium bromide
  • the sample was subjected to a depth filter and ultrafiltration (UF / DF), and then the amount of ethanol and sodium chloride was adjusted and stored in a raw form.
  • UF / DF depth filter and ultrafiltration
  • the equilibrated complex was dissolved by incubation (37 ⁇ 2 ° C) in sodium phosphate (Na 3 PO 4 ) buffer solution at a rate of 0.1 M per 20 g of saccharides, and then cyanoborohydride (100 mg / mL) was added. The conjugation reaction was started by doing so. After incubation at 37 ⁇ 2 ° C. for about 44-52 hours, the temperature was lowered to 23 ⁇ 2 ° C. and 1 mL of 0.9% NaCl solution was added to the reactor.
  • Sodium borohydride solution (100 mg / mL) is added to 1.8 to 2.2 molar equivalents of sodium borohydride per mole of saccharide, and the mixture is incubated with stirring at 23 ⁇ 2 ° C. to any unreacted saccharide present in the saccharide. Aldehydes were reduced. The mixture was diluted with 5 mL of 0.9% aqueous sodium chloride solution and the diluted conjugate mixture was diafiltered using a 100 kDa MWCO membrane.
  • a 2M NaCl polysaccharide solution was prepared by adding sodium chloride powder to a 9V polysaccharide stock solution prepared without hydrolysis.
  • CDAP (1-cyano-4-dimethylaminopyridinium tetrafluoroborate) was dissolved at a ratio of 0.5 w / w% to polysaccharide, added to a 9V polysaccharide solution, and stirred for 15 minutes to induce a polysaccharide activation reaction.
  • the sodium hydroxide solution was then raised until the pH was 9.5 ⁇ 0.1 ° C. and then stirred for 3 minutes so that the hydroxyl groups of the polysaccharides could be sufficiently activated by CDAP.
  • CRM197 was added at a ratio of CRM197 1.0 w / w% to polysaccharide in the polysaccharide solution that had undergone polysaccharide activation, and the conjugation reaction was performed at room temperature for 1 hour.
  • the conjugation reaction was terminated by adding 2 M glycine solution in 3 molar equivalents to 1 molar equivalent of CDAP and incubating overnight at room temperature with pH adjusted to 9.0.
  • the terminated conjugate was concentrated and diafiltered into an ultrafiltration filter through a buffer containing 0.9% sodium chloride.
  • the original capsular polysaccharide originated from each serotype was dissolved in water for injection so that the final concentration range was within the range described below and filtered through a 0.45 ⁇ m filter.
  • serotypes 6A, 12F and 19A were dissolved at 8-12 mg / ml, and 2-4 mg / ml for serotypes 7F, 14 and 23F.
  • serotypes 15B, 22F, and 33F the solution was dissolved in the range of 2-5 mg / ml and filtered.
  • Each serotype was run by incubating the solution in the pH and temperature ranges described below. Specifically, phosphoric acid at 70-80 ° C. overnight for serotypes 1, 3, 5, 6B, 7F, 14 and 23F, 70-80 ° C. for 1-4 hours for serotypes 6A and 19F, for serotypes 9V and 18C. The solution was incubated at pH 2.0, 65-80 ° C. for 1-3 hours. Serotypes 22F and 33F were incubated overnight at 75-85 °C and serotype 12F at pH 2.0, 75-85 °C for 1-3 hours using phosphate solution.
  • w% was dissolved at a weight ratio of 4 w / w% for serotypes 5, 9V, 18C, 19F, 22F and 23F, and 5 w / w% for serotype 12F and added to each polysaccharide solution.
  • sodium hydroxide solution was added to raise the pH to 9.4 to 9.7, followed by stirring for 3 to 7 minutes so that the hydroxyl groups of the polysaccharides could be sufficiently activated by CDAP.
  • Conjugation reaction was performed for 1 hour to 4 hours by adding CRM197 0.5-1.0 w / w% to polysaccharide to each serotype polysaccharide solution. The reaction conversion was measured using HPLC-SEC, and CDAP was added as needed.
  • the reaction was terminated by adding 3-6 molar equivalents of glycine solution relative to 1 molar equivalent of CDAP added for all serotypes and adjusting the pH to 9.0.
  • the conjugation solution was stirred at 21-24 ° C. for 1 hour and then stored overnight at 2-8 ° C. low temperature.
  • the diluted conjugate mixture was concentrated and diafiltered into an ultrafiltration filter using at least 20 volumes of buffer.
  • the buffer was maintained in the range of pH 5.5 to 6.5, and a buffer containing 0.9% sodium chloride was used.
  • Fractional molecular weight of the ultrafiltration filter was carried out using 300 kDa in all serotypes, and the permeate was discarded.
  • Step 5 Sterile Filtration
  • the residue after diafiltration was diluted to less than 0.4 g / L based on polysaccharide concentration using a buffer and filtered through a 0.22 ⁇ m filter.
  • the filtered product was subjected to in-process control (sugar content, residual DMAP). In-process controls were performed on the filtered residue to determine if further concentration, diafiltration and / or dilution were needed.
  • Aluminum salts (mainly aluminum phosphate) were added to the sterile filtrate so that the final concentration was 1 mg / mL based on aluminum ions, and extra salts were added to maintain a pH range of 5.5 to 6.5. After the adsorption, the stock solution was subjected to quality inspection to confirm quality suitability, and refrigerated at 2 to 8 ° C. until use.
  • the required amount of final bulk concentrate was calculated based on the batch volume and bulk polysaccharide concentration.
  • the required amount of 0.85% sodium chloride, succinate buffer, 2-phenoxyethanol and formaldehyde were added to a previously labeled formulation vessel, followed by the bulk concentrate. Then it was thoroughly mixed and filtered through a 0.22 ⁇ m filter.
  • the formulated bulk liquid was slowly mixed, then bulk aluminum phosphate was added and mixed well. pH was checked and adjusted if necessary.
  • the formulated bulk product was stored at 2-8 ° C.
  • the resulting vaccine composition contains 4.4 ⁇ g / mL of each polysaccharide (13, hereinafter, 'LBVE013') in 0.5 mL total, 1, 3, 4, 5, 6A, 7F, 9V, 14, 18C, 19A, 19F and 23F.
  • For 17 is (hereinafter 'LBVE017') 1, 3, 4, 5, 6A, 7F, 9V, 12F, 14, 15B, 18C, 19A, 19F, 22F, 23F and 33F), except that 6B is 8.8 ⁇ g / mL; About 29.3 ⁇ g CRM197 transport protein; 0.5 mg of elemental aluminum (2 mg aluminum phosphate) adjuvant; About 4.25 mg sodium chloride; About 295 ⁇ g succinate buffer; It contains about 3 mg of 2-phenoxyethanol and about 60 ⁇ g of formaldehyde (applicable when preservatives are added). The content is set as the reference amount of 100%, and in the following examples, the content of some serotypes is adjusted as needed (eg, 90%, 75%, 50%, 25%, 10%, 1%) for further analysis. Was performed.
  • Example 3 A study was conducted to evaluate whether each multivalent pneumococcal vaccine composition prepared in Example 3 has the ability to induce an immune response in rabbits. This immunogenicity was confirmed by measuring serum IgG concentration via antigen-specific ELISA.
  • a solution in which 13 or 17 polysaccharide antigens were conjugated to magnetic beads was attached to a 96-well plate. Each individual serum was adsorbed by reacting with 1 mg / mL CWPS multi solution (CWPS multi ® , Statens Serum Institute) for 30 minutes at room temperature to minimize non-specific antigen-antibody reactions, and then buffered for antibody dilution containing Tween 20. Was diluted to a suitable dilution multiple using. Plates with 13 or 17 polysaccharide antigen-conjugated magnetic beads were washed twice with a wash buffer, and 50 ⁇ l of previously adsorbed and diluted serum was added to the plates and allowed to react at room temperature for 30 minutes.
  • CWPS multi ® CWPS multi ®
  • the reacted plate was washed three times in the same manner, and each well was added with R-PE Ghycoerythrin goat anti-Rabbit IgG (1: 500), followed by reaction at room temperature for 30 minutes.
  • the plate was washed three times in the same manner as above, and 80 ⁇ l of the buffer solution was added to each well, and fluorescence was measured using a multiplex reader.
  • Example 13 is an immunogenic composition (LBVE013) prepared in the present invention are all relative to Prevenar 13 ® IgG concentration levels higher than prevena 13 ® were identified in the serotype.
  • serotypes 1, 6B, 7F, 9V 14 and 19F showed two to six times better effects than prevena 13 ® (Table 1).
  • the immunogenicity of LBVE017 prepared by adding four or more serotypes to LBVE013 was compared with LBVE013.
  • the immunogenic composition was confirmed that the decrease in the overall IgG titer occurs when the 17-valent valence is increased from 13 (FIG. 1).
  • the inventors of the present invention have all administered the doses of each of the 6A, 6B, 14, 19A and 19F conjugates, which had relatively high IgG titers at 13 valent relative to the 17-valent immunogenic composition. It was confirmed that the reduction to the 50% level at the same time. As a result, it was confirmed that an overall increase in immunogenicity occurs (Fig. 1).
  • serotypes 5, 6A, 6B, 14, 18C, and 19A IgG titers at levels higher than 13 were found.
  • the immunogenic composition according to the present invention can be very useful for developing pneumococcal conjugate vaccines by providing a practical way to overcome possible immune interference which can negatively affect the overall immunogenicity of the vaccine.
  • the inventors have sought to see if recovery of the immune response as seen in the 17-valent composition can be similarly observed in the 13-valent composition by simultaneously reducing the dose of each of the conjugates. Accordingly, even at 13-valent, the amount of the conjugate conjugate of the five serotypes was reduced to 50% or 25%, respectively, to prepare a 13-valent immunogenic composition, and its immunogenicity was 100% of the 13-valent immunogenic composition of the present invention (LBVE013). ) And Prevena 13 ® . As a result, the total immunogenicity did not recover (rather, decreased) in the thirteen, even though the doses of the five serotypes were reduced, unlike the results of the seventeen. This result means that the result of reducing the dose of some serotypes in the 17-valent immunogenic composition of the present invention is beyond the level predictable by those skilled in the art.

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Abstract

La présente invention concerne une composition immunogène multivalente présentant un titre d'IgG augmenté, et son utilisation et, en particulier: une composition vaccinale pour la prévention de maladies pneumococciques, contenant un conjugué polysaccharide capsulaire-protéine porteuse; et une composition immunogène pour le Streptococcus pneumoniae.
PCT/KR2017/011959 2016-10-28 2017-10-27 Composition immunogène multivalente ayant un titre d'igg augmenté, et son utilisation Ceased WO2018080213A1 (fr)

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US12097250B2 (en) 2017-12-06 2024-09-24 Merck Sharp & Dohme Llc Compositions comprising Streptococcus pneumoniae polysaccharide-protein conjugates and methods of use thereof
US11642406B2 (en) 2018-12-19 2023-05-09 Merck Sharp & Dohme Llc Compositions comprising Streptococcus pneumoniae polysaccharide-protein conjugates and methods of use thereof
US12016914B2 (en) 2018-12-19 2024-06-25 Merck Sharp & Dohme Llc Compositions comprising Streptococcus pneumoniae polysaccharide-protein conjugates and methods of use thereof

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