Multivalent Streptococcus Vaccines

Abstract

The invention is directed to immunogenic compositions, including vaccines, containing multivalent immunogenic composition comprising 25 different serotypes of capsular polysaccharides of S. pneumoniae. Compositions are preferably liquid and thermo stable for periods of time that allow for distribution and use. The invention is also directed to method for the manufacture and methods for the administration of 25 valent immunogenic compositions of S. pneumoniae.

Claims

1. An immunogenic composition comprising at least 25 different serotypes of polysaccharides of S. pneumoniae serotypes coupled to carrier protein.

2. The immunogenic composition of claim 1, wherein one or more of the capsular polysaccharides are from about 10 kDa to about 50 kDa.

3. The immunogenic composition of claim 1, wherein one or more of the capsular polysaccharides from about 30 KDa to about 100 KDa.

4. The immunogenic composition of claim 1, wherein one or more of the capsular polysaccharides are from about 100 KDa to about 300 KDa.

5. The immunogenic composition of claim 1, wherein the carrier protein comprises CRM, purified CRM197, recombinantly produced CRM, tetanus toxoid fragments (TTHc), tetanus toxin, tetanus toxin heavy chain proteins, diphtheria toxoid, tetanus toxoid, Pseudomonas exoprotein A, Pseudomonas aeruginosa toxoid, Bordetella pertussis toxoid (PT), Clostridium perfringens toxoid, Escherichia coli heat-labile toxin B subunit, Neisseria meningitidis outer membrane complex, protein PorB, Hemophilus influenzae protein D, Flagellin Fli C, Horseshoe crab Haemocyanin, RSV virus proteins, adenylate cyclase toxin (ACT), 69 KDa protein and Human Papilloma viral protein antigens or its VLP form, Hepatitis B core antigen or its VLP form or derivatives of HBsAg, and/or and fragments, derivatives, and modifications thereof.

6. The immunogenic composition of claim 1, wherein one or more of the polysaccharides are covalently coupled to a linker and the linker is coupled to the carrier protein.

7. The immunogenic composition of claim 1, which is a liquid.

8. The immunogenic composition of claim 1, which comprises from about 0.25 ml to about 1.0 ml per dose.

9. The immunogenic composition of claim 1, which comprises about 0.5 ml per dose.

10. The immunogenic composition of claim 1, which comprises 10 micrograms or less of total polysaccharides and protein per dose.

11. The immunogenic composition of claim 1, which comprises 4 micrograms or less of total polysaccharides and protein per dose.

12. The immunogenic composition of claim 1, wherein the carrier protein comprise from about 0.5% to about 0.7%, by weight, per dose.

13. The immunogenic composition of claim 1, which comprises about equal amount by weight of capsular polysaccharides to total carrier protein.

14. The immunogenic composition of claim 1, which comprises a greater amount by weight of capsular polysaccharides to total carrier protein.

15. The immunogenic composition of claim 1, further comprising of at least one adjuvant.

16. The immunogenic composition of claim 15, wherein the at least one adjuvant is selected from the group consisting of aluminum salt, calcium phosphate, a liposome of monophosphoryl lipid A (MPLA), saponin QS-21, a TLR7/8 agonist, and combinations thereof.

17. The immunogenic composition of claim 16, wherein the aluminum salt is selected from the group consisting of aluminum phosphate, aluminum sulfate and/or aluminum hydroxide.

18. The immunogenic composition of claim 1, wherein the at least 25 different serotypes of capsular polysaccharides of S. pneumoniae comprise serotypes: 1, 2, 3, 4, 5, 6B, 6C, 7F, 8, 9N, 9V, 10A, 12F, 14, 15A, 15B, 15C, 16F, 18C, 19A, 22F, 23F, 24F, 33F, and 35B.

19. The immunogenic composition of claim 1, which, upon administration to a subject, generates a minimal immune response to carrier protein as compared to the immune response to polysaccharide.

20. The immunogenic composition of claim 1, which provides effective treatment or prevention of infection by S. pneumoniae bacteria.

21. The immunogenic composition of claim 1, comprising a a pharmacologically acceptable carrier.

22. The method for manufacture of the immunogenic composition of claim 1, comprising: activating carrier proteins to form activated carrier proteins; reducing a disulfide of each carrier protein to create a sulfhydryl group; and coupling capsular polysaccharides to the activated carrier proteins.

23. The method of claim 18, wherein the activated carrier proteins are selected from the group consisting of cross-reactive material (CRM197) obtained or derived from C. diptheriae, and recombinant CRM197 obtained or derived from P. fluorescens or E. coli.

24. The method of claim 18, further comprising coupling PEG spacers to the activated carrier proteins.

25. An immunogenic composition comprising one or more polysaccharides of one of more different serotypes of S. pneumoniae coupled to carrier protein via adipic acid dihydrazide (ADH) linkers, wherein the one or more polysaccharides have a molecular weight of from about 100 KDa to about 300 KDa.

26. The immunogenic composition of claim 25, wherein the one or more of the ADH linkers are pegylated dihydrazide (HZ-PEG-HZ) linkers.

27. The immunogenic composition of claim 25, wherein the carrier protein comprises CRM, recombinantly produced CRM, or a domain of CRM.

28. A method of manufacture of the immunogenic composition of claim 25, comprising: providing the one or more polysaccharides of one of more serotypes of S. pneumoniae and carrier protein; activating the one or more polysaccharides and/or the carrier protein with pegylated-ADH linkers; and linking the polysaccharides to carrier protein via carbodiimide chemistry.

29. The method of claim 28, wherein the carrier protein comprises CRM, recombinantly produced CRM, tetanus toxoid or toxoid fragments (TTHc), tetanus toxin, tetanus toxin heavy chain proteins, diphtheria toxoid, tetanus toxoid, Pseudomonas exoprotein A, Pseudomonas aeruginosa toxoid, Bordetella pertussis toxoid (PT), Clostridium perfringens toxoid, or a combination thereof.

30. The method of claim 28, wherein the one or more serotypes comprise serotypes: 1, 2, 3, 4, 5, 6B, 6C, 7F, 8, 9N, 9V, 10A, 12F, 14, 15A, 15B, 15C, 16F, 18C, 19A, 22F, 23F, 24F, 33F, and 35B.

Description

DESCRIPTION OF THE INVENTION

[0042] Streptococcus pneumoniae is a Gram-positive bacterium which can cause diseases including pneumonia, bacteremia, meningitis, and acute otitis media. The microorganisms are encapsulated with a variety of polysaccharides which produces serotype specificity. At least 90 pneumococcal serotypes are known of which about 23 account for 90% of invasive diseases. The protection against invasive disease is directly related to the ability to generate an antibody response that is specific to a particular capsular polysaccharide associated with the microorganisms of the infection, otherwise referred to as serotype specificity.

[0043] A multivalent S. pneumoniae immunogenic composition was surprisingly created comprising polysaccharides of at least 25 different capsular polysaccharide serotypes. The presence of additional serotypes over available vaccines Prevnar-13) is predicted to cover an additional about 21-25% more of invasive pneumococcal diseases (IPDs). The serotypes selected, and those not selected, were determined by the presence or absence of the invasive disease-causing isolates post pneumococcal conjugate vaccine (PCV) introduction as determined globally by region, the likely invasive serotypes in children less than about 5 years of age, the regional needs in low and middle-income countries, and determination of newly emerging serotypes in children less than about 5 years of age since, all prioritization based on frequency in Gavi countries, potential for epidemics and known characterization of polysaccharides. There is also a rational for exclusion of certain known serotypes as the not likely to be invasive serotypes in children less than about 5 years of age, as not likely to meet the regional needs in low and middle-income countries, and as not newly emerging serotypes in children less than about 5 years of age since, as not prioritization based on frequency in Gavi countries, as having little to no potential for epidemics, and as not known to be properly characterized. Important criteria for making the selection of serotypes are listed in Table 1.

TABLE-US-00001 TABLE 1 Serotypes listed from highest to lowest frequency All Antibiotic ages More Less resistant Number of IPD <5 GAVI High invasive invasive common additional non IPD <5 and PCV than than emerging serotypes GAVI GAVI non-GAVI Africa <5 coverage 19A 19A genotype 1 15B/C  2 22F  8 22F  8 6C 35B 2 12F 12F 8 12F 12F 12F 15A 15B/C 3  8  8 12F 35B 24F 24F 15B/C 15A 4 22F 35B 6C 15B/C 10A 33F 16F 24F 5 24F 23B 15A 16F 23B 22F 23B 9N 6 10A 10A 9N 15A  8 38 35B 9L 7 !5A 45 33F 10A 15A 10A 12F 8 35B 20 23A 13 11A 9 23B 10F 10A 17F 15B/C 10 6C 24F 11A 7C 9N replace 6A 11 33F 16F 24F, 9N 38 15B/C, 16F, 35B Number of 10  1 Additional strains

[0044] As shown in Table 1, 24 valent with 6C to replace 6A to include 11 serotypes that are bolded—2, 8, 10A, 12F, 15A, 15B/C, 22F, 23B, 24F, 33F, 35B. 16F is believed to be prevalent in Africa, but less invasive. 9N is also prevalent in Africa with a high PCV. Global Pneumococcal Sequence Complex 16 has 65% MDR with 38 in high PCV and invasive. 9N protection from 9V is believed weak. Added in the adult data raises the priority for 9N, 23A and 11A. The remaining strains are very low invasiveness 11A, or relatively rare with little to no AMR or genotype signal 7C, 10F, 13, 17F, 20, 45.

[0045] The 25 serotypes identified include: serotypes 1, 2, 3, 4, 5, 6B, 6C, 7F, 8, 9N, 9V, 10A, 12F, 14, 15A, 15B/C, 16F, 18C, 19A, 19F, 22F, 23F, 24F, 33F, and 35B. The rational for inclusion of these serotypes is listed in Table 2.

TABLE-US-00002 TABLE 2 Serotypes Rationale for Inclusion 1, 3, 4, 5, 6B, Serotypes most likely to cause invasive disease 7F, 9V, 14, 18C, globally and included in currently licensed 19A, 19F, 23F Prevenar 13 22F, 33F Serotypes known to invasive and included in the investigational PCV15  6C Due to demonstrated cross-protection between serotype 6B and 6A, serotype 6B in Prevenar 13, is substituted with serotype 6C, an increasing cause of invasive, antimicrobial resistant disease  2 A highly invasive serotype recently associated with invasive disease in several LMIC, specifically in Bangladesh, Guatemala and Israel  8 Invasive disease in children <5 years of age in post 2010 era in non-Gavi and Gavi countries  9N Invasive serotype in children <5 years of age for which there is not demonstrated cross-protection by serotype 9V in the currently licensed vaccines 10A In the most common invasive serotypes in pediatric and adult populations, both in high income countries as well as LMICs 12F A highly invasive serotype known to cause epidemic disease 15A Common serotype found in invasive disease in children <5 years of age especially in non-Gavi countries 15B/C Common serotype found in invasive disease in children <5 years of age especially in non-Gavi countries 16F Increasingly cause of invasive disease in children <5 years of age, in Gavi and non-Gavi countries 24F Emerging invasive serotype recently described as the most common cause of meningitis in France, as well as reports from Argentina, Germany, Peru and Papua New Guinea 35B Emerging invasive serotypes

[0046] Preferably the immunogenic composition of the disclosure contains at least 25 different serotypes of bacterial capsular polysaccharides of S. pneumonia wherein the polysaccharides (PS) are conjugated and/or coupled to carrier proteins (e.g., PCV formulations). Preferably the PS are sufficiently purified from the desired serotypes of S. pneumonia and conjugated or otherwise coupled to carrier proteins.

[0047] Formulations of multivalent immunogenic compositions can be designed for the treatment of specific populations treatment contain mut

[0048] Preferred carrier proteins include, for example, cross reactive materials or CRM (e.g., purified CRM 197 or recombinantly produced CRM), tetanus toxoid fragments (TTHc), tetanus toxin, tetanus toxin heavy chain proteins, diphtheria toxoid, tetanus toxoid, Pseudomonas exoprotein A, Pseudomonas aeruginosa toxoid, Bordetella pertussis toxoid (PT), Clostridium perfringens toxoid, Escherichia coli heat-labile toxin B subunit, Neisseria meningitidis outer membrane complex (e.g., protein PorB), Hemophilus influenzae protein D, Flagellin Fli C, Horseshoe crab Haemocyanin, RSV virus proteins, adenylate cyclase toxin (ACT), 69 KDa protein and Human Papilloma viral protein antigens or its VLP form, Hepatitis B core antigen or its VLP form or derivatives of HBsAg, and/or and fragments, derivatives, and modifications thereof.

[0049] Coupling of PS to carrier protein may be direct, PS to carrier protein, or indirect through one or more linkers. Preferred linkers include, for example, linkers of polyethylene glycol (PEG). Linkages may be monovalent or multivalent (e.g., bivalent, trivalent, etc.). Linkers are preferably coupled with polysaccharide and/or carrier proteins by connecting to PEG via two hydrazine functional groups cable of covalently compounding with both carrier protein as well as polysaccharides. This creates a class of covalently compounded PEG products that have the additional effect of PEG on their properties compared to conjugates made by conventional methods. PEG has an additional enhancing effect on the immunogenicity of polysaccharides compared to regular conjugates and a depressing effect on the Immune response of carrier proteins. This allows for an efficient and effective composition with large numbers of serotypes. This surprising and unexpected benefits observed are important considerations in developing immunogenic compounds such as vaccines.

[0050] The process of coupling can involve, for example, PS activation via either oxidation or cyanylation chemistry. The PS is oxidized by sodium periodate and introduced with either reactive aldehyde or isothiocyanate (—OCN) groups. Coupling strategies include, for example, short and long linker and/or short and long PS.

[0051] Preferably the immunogenic composition is prepared as and maintained as a liquid, although composition may be lyophilized and rehydrated before use. Preferably, the compositions, whether liquid or lyophilized, are suitable for storage at room temperatures for periods of time. Preferred periods include weeks, months and years. Preferably the compositions of the invention are thermo-stable at 30° C. for at least 2 years and at 50° C. for at least 3 months.

[0052] Also preferred, compositions comprise from about 0.25 ml to about 1.0 ml per dose, and more preferred are doses of about 0.5 ml. Preferably the immunogenic composition comprises 10 micrograms or less of total polysaccharides and total protein per dose. More preferred, immunogenic compositions comprise 4 micrograms or less of total polysaccharides per dose. Preferably carrier protein comprises from about 0.5% to about 0.7%, by weight, per dose. Also preferred are immunogenic compositions which comprise greater amount by weight of capsular polysaccharides to total carrier protein, although compositions may contain about equal amount by weight of capsular polysaccharides to total carrier protein.

[0053] Compositions of the invention may include stabilizers to maintain efficacy for long periods and over multiple temperatures. Stabilizers and protective agents include, for example, excipients, buffers (e.g., citrate, calcium carbonate), amino acids (e.g., lysine, arginine, glycine, etc.), salts, bulking agents, antioxidants and dispersants. Protectant agents include, for example, dextran and other lower molecular weight sugars such as sucrose, trehalose, mannitol, and/or medium-chain triglyceride (MCT) oil.

[0054] Protection against pneumococcal disease is obtained by the generation of an immunological response in the individual administered the composition. Suitable immunological response preferably includes the generation of protective antibodies against the different polysaccharide components. Preferably the antibodies observed after administration of the immunological composition fall slowly, more slowly that the rapid reductions observed with convention PCV vaccines. This eliminates a need for multiple injections, so that good protection is achieved after one or at most two injections, saving cost as well as pain to patients such as infants and children (e.g., greater than 3 months). In addition, have a reduced number of injections allows protection to be more widely available than conventional multiple injections, especially for those unable to return for repeated injections.

[0055] A preferred formulation of the immunogenic composition comprises at least 25 different serotypes of bacterial capsular polysaccharides of Streptococcus pneumoniae covalently connected to carrier protein through, preferably, PEGylated linker compounds. The carrier protein is covalently connected to bacterial capsular polysaccharides through a number of multifunctional PEG linkers, which may be homo-multi-functional or hetero-multi-functional, and preferably of defined lengths. Preferred linkers include adipic acid di-hydrazide (ADH) and PEGylated-ADH linkers. Preferable, the linkers are from 1 KDa to 3.5 kDa, and may be greater than 3.5 KDa. Preferred hetero- and/or homo-linker sizes include, for example, 40 Å and less, 30 Å and less, 20 Å and less, 10 Å and less, 5 Å and less, 2 Å and less, and/or combinations of these sizes. Preferably the polysaccharide-protein covalent PEG compound is prepared wherein carrier protein reacts with cleaved and depolymerized polysaccharide fragments of optimum chain length. Polysaccharides are conjugated to carrier protein, either of which may be coupled via an activating agent, such as for example, carbodiimide (e.g. 1-ethyl-3(3-dimethylaminopropyl; EDC or EDAC), to give a derivatized carrier protein in presence of a 2-(N-morpholino) ethane sulphonic acid (MES buffer) or EDC/sNHS chemistry. Carbodiimide conjugation, as with CDI-mediated conjugation, works by activating carboxyl groups for direct reaction with primary amines via amide bond formation. EDC couples primary amines to carboxylic acids by creating activated ester leaving groups. Basically, the carbonyl of acid attacks the carbodiimide of EDC creating a proton transfer. The primary amine attacks the carbonyl carbon of the acid forming a tetrahedral intermediate which forms an amine and discards urea.

[0056] Preferably the immunological composition contains PS with low molecular weight and bifunctional linkers preferably that enhance immunogenicity. Preferred molecular sizes are 300 kDa and lower, 200 KDa and lower, 100 KDa and lower, 75 Kda and lower, 50 kDa and lower, 25 KDa and lower, 10 KDa and lower, and/or combinations of these molecular sizes. This provides higher immunogenicity and higher avidity of bivalent compounds as well as lower carrier protein immunogenicity.

[0057] Another embodiment of the invention is directed to methods for the administration of vaccines of the invention to patients in need thereof for treating or preventing an infection. The method comprises administering a therapeutically effective amount of the vaccine of the invention to a mammal, comprising determining the therapeutically effective amount of the vaccine to be administered that provides therapy to an infected patient and/or protection from infection. The therapeutically effective amount is typically determined by based on the weight of the mammal and the strength or responsiveness of the patient's immune system and can be determined by those skilled in the art. The therapeutically effective amount is administered to a patient in need thereof, which may be to treat an active or suspected infection or prevent an infection. The vaccine may have been obtained from a lyophilized powder and reconstituted to an aqueous or non-aqueous liquid prior to administration to the patient. Preferably the vaccine is administered as a liquid, which may be administer via intra-muscular, intra-peritoneal, or intra-venous injection, and the patient may be an infant, a toddler, an adolescent, an adult or a senior. Surprisingly, the compositions of the invention do not generate side effects such as redness or inflammation at the injection site, and does not generate a generalized fever or inflammation, or other unwanted side effects for the patient. Preferably an immunologically effective vaccine contains only the multivalent composition and nothing further such as, for example, no added adjuvants.

[0058] Preferably the immunogenic compositions of the invention comprises administering multivalent immunogenic compositions to an individual for the treatment or prevention of a Streptococcus infection, and preferably infection attributable to Streptococcus pneumoniae. Infections that are treatable with immunogenic compositions include, for example, pneumonia, bacteremia, meningitis, and acute Otitis media. Preferably administration comprises intramuscular injection, intraperitoneal injection, intravenous injection, intranasal, oral or transdermal. Preferably the patient is an infant, a toddler, a child, an adolescent, an adult or a senior. Preferred compositions include immunogenic compositions designed for the treatment and/or prevention of infection of infants, of individuals less than 3 years of age, of individuals less than 5 years of age, of individuals less than 15 years of age, in adults, and in individuals greater than 60 years of age.

[0059] Other embodiments and uses of the invention will be apparent to those skilled in the art from consideration of the specification and practice of the invention disclosed herein. All references cited herein, including all publications, U.S. and foreign patents and patent applications, are specifically and entirely incorporated by reference. It is intended that the specification and examples be considered exemplary only with the true scope and spirit of the invention indicated by the following claims. Furthermore, the term “comprising of” includes the terms “consisting of” and “consisting essentially of.”