WO2018189515A1

WO2018189515A1 - Novel pneumococcal vaccine

Info

Publication number: WO2018189515A1
Application number: PCT/GB2018/050935
Authority: WO
Inventors: Stephen D. BENTLEY; Nicholas J. CROUCHER
Original assignee: Imperial Innovations Ltd; Genome Research Ltd
Current assignee: Genome Research Ltd; Ip2ipo Innovations Ltd
Priority date: 2017-04-10
Filing date: 2018-04-09
Publication date: 2018-10-18
Anticipated expiration: 2019-10-10
Also published as: GB201705733D0

Abstract

The invention relates to a pneumococcal vaccine, to pharmaceutical compositions comprising said vaccine and to their uses in vaccination against pathogenic pneumococcal strains.

Description

NOVEL PNEUMOCOCCAL VACCINE

FIELD OF THE INVENTION

BACKGROUND OF THE INVENTION

Streptococcus pneumoniae, or pneumococcus, is a Gram-positive, alpha-hemolytic (under aerobic conditions) or beta-hemolytic (under anaerobic conditions), facultative anaerobic member of the genus Streptococcus. As a significant human pathogenic bacterium S.

pneumoniae was recognized as a major cause of pneumonia in the late 19th century, and is the subject of many humoral immunity studies. S. pneumoniae resides asymptomatically in healthy carriers typically colonizing the respiratory tract, sinuses, and nasal cavity. However, the bacterium may become pathogenic and spread to other locations to cause disease, particularly in susceptible individuals with weaker immune systems, such as the elderly and young children. S. pneumoniae is the main cause of community acquired pneumonia and meningitis in children and the elderly, and of septicemia in those infected with HIV. The organism also causes many types of pneumococcal infections other than pneumonia. These invasive pneumococcal diseases include bronchitis, rhinitis, acute sinusitis, otitis media,

conjunctivitis, meningitis, sepsis, osteomyelitis, septic arthritis, endocarditis, peritonitis, pericarditis, cellulitis, and brain abscess.

Due to the importance of disease caused by S. pneumoniae several vaccines have been developed to protect against invasive infection. The World Health Organization recommend routine childhood pneumococcal vaccination; it is incorporated into the childhood

immunization schedule in a number of countries including the United Kingdom, United States, and South Africa.

There are two different types of pneumococcal vaccine used, namely pneumococcal conjugate vaccine (PCV) and pneumococcal polysaccharide vaccine (PPV). PCV is given to all children as part of the childhood vaccination programme. PPV is given to people aged 65 years or over, and others who are at high risk. PCVs protects against up to 13 types of S. pneumoniae bacteria, and PPV protects against 23 types. It is thought that PPV is around 50-70% effective at preventing more serious types of invasive pneumococcal infection.

However, the circulating pneumococcal population is able to evade vaccination regimes, therefore, there is a need to provide alternative and effective vaccines against S.

pneumoniae.

SUMMARY OF THE INVENTION

According to a first aspect of the invention, there is provided a pneumococcal vaccine comprising one or more proteins, or one or more nucleic acid molecules encoding said proteins, which are present on capsular pneumococcal strains and absent on non-capsular pneumococcal strains.

According to a further aspect of the invention there is provided a pharmaceutical composition comprising a pneumococcal vaccine as described herein.

According to a further aspect of the invention, there is provided a method of preventing pneumococcal infection in a mammal which comprises administering to the mammal a therapeutically effective amount of a vaccine composition as described herein.

According to a further aspect of the invention, there is provided a method of inducing an immune response in a mammal, wherein the method includes administering to the mammal, an effective amount of a vaccine composition described herein. According to a further aspect of the invention, there is provided a kit of parts comprising a vaccine composition as described herein, a medical instrument or other means for administering the vaccine composition and instructions for use.

DETAILED DESCRIPTION OF THE INVENTION

According to a first aspect of the invention, there is provided a pneumococcal vaccine comprising one or more proteins, or one or more nucleic acid molecules encoding said proteins, which are present on capsular pneumococcal strains and absent on non-capsular pneumococcal strains. S. pneumoniae is only thought to be able to cause invasive disease when expressing a polysaccharide capsule. Strains which lack the polysaccharide capsule are often referred to as nontypable since the typing method used to categorise members of the species relies on binding of antibodies with sera specific for different polysaccharide capsule structures.

Unencapsulated strains therefore produce no positive tests so are designated as

nontypable. The inventors of the present invention believe that complete eradication of pneumococcal bacteria from the human microbiota will have an unknown (and potentially harmful) effect upon the natural ecosystem. Therefore, the invention provides a vaccine which is able to specifically target only the pathogenic, capsular pneumococcal strains. Therefore, the nonpathogenic, non-capsular pneumococcal strains will be unaffected by treatment with the vaccine of the present invention.

Therefore the vaccine of the invention would not aim to use the immune response alone to remove targeted pneumococci, as previous designs have done, but instead put them at a competitive disadvantage with divergent unencapsulated pneumococci (also termed classic non-typeable pneumococci¹ ; these may also be regarded as pneumococci-like streptococci), such that they were eliminated from the population through the combination of vaccine- induced immunity and displacement through competition with the divergent unencapsulated pneumococci or pneumococcus-like streptococci. There are several publications which support the claimed invention. For example, Jomaa et al (2006) Vaccine 24, 5133-5139 studied the effects of immunising mice with PiaA, an ABC transporter iron-binding protein within the scope of the present invention, in combination with PiuA. Priming doses were administered via an intranasal route, a potential route of administration claimed herein, as allowing for the possibility of being combined with a dose of a probiotic. These priming doses were shown to induce an antibody response in both the bloodstream and the respiratory tract, following booster intranasal and systemic booster doses. These adaptive immune responses were sufficient to protect mice against pulmonary infection progressing from an inoculum of pneumococci in the nasopharynx. Hence this study indicates there is the potential for the vaccine of the present invention to induce an adaptive immune response at the mucosal surface, which could be protective against carriage and non-invasive, as well as invasive, pneumococcal disease, particularly if a probiotic were provided to the recipient at the same time.

In addition, Anderson et al (2016) Vaccine 34, 6292-6300 studied the effects of systemically immunising mice with the surface-exposed degradative enzyme Eng, a surface anchored endo- alpha-N-acetylgalactosaminidase within the scope of the present invention. This was found to elicit a humoral immune response that protected against progression to pulmonary disease from nasopharyngeal inoculation. Hence this study indicates the potential for the Eng component of the vaccine of the present invention to stimulate a response that protects against non-invasive, as well as invasive, pneumococcal disease.

In one embodiment, the one or more proteins are selected from one or more of:

transcriptional regulator PIcR, putative; sucrose-6-phosphate hydrolase; L-fucose isomerase; carbamate kinase; surface anchored beta-galactosidase; capsule biosynthesis integral membrane regulatory protein Wzg; surface anchored endo-alpha-N- acetylgalactosaminidase; and ABC transporter iron-binding protein. References herein to "transcriptional regulator PIcR, putative" include the product of the plcR gene which is a transcriptional regulator and has a cluster of orthologous genes (COG) designation of CLS01686, using the scheme defined in Croucher et al (2015) Sci. Data 2: 150058. References herein to "sucrose-6-phosphate hydrolase" include the product of the sacC gene which is involved in carbohydrate metabolism and has a cluster of orthologous genes (COG) designation of CLS01545, using the scheme defined in Croucher et al (2015) Sci. Data 2. 150058. References herein to "L-fucose isomerase" include the product of the fuel gene which is involved in carbohydrate metabolism and has a cluster of orthologous genes (COG) designation of CLS01869, using the scheme defined in Croucher et al (2015) Sci. Data 2: 150058. References herein to "carbamate kinase" include the product of the arcC gene which is involved in carbohydrate metabolism and has a cluster of orthologous genes (COG) designation of CLS01862, using the scheme defined in Croucher et al (2015) Sci. Data 2. 150058. References herein to "surface anchored beta-galactosidase" include the product of the bgaA gene which is a surface-associated degradative enzyme and has a cluster of orthologous genes (COG) designation of CLS00596, using the scheme defined in Croucher et al (2015) Sci. Data 2. 150058. References herein to "capsule biosynthesis integral membrane regulatory protein Wzg" include the product of the wzg gene which is a surface-associated enzyme involved in regulation of capsule production and has a cluster of orthologous genes (COG) designation of CLS00362, using the scheme defined in Croucher et al (2015) Sci. Data 2: 150058.

References herein to "surface anchored endo-alpha-N-acetylgalactosaminidase" include the product of the eng gene which is a surface-associated degradative enzyme and has a cluster of orthologous genes (COG) designation of CLS00380, using the scheme defined in Croucher et al (2015) Sci. Data 2: 150058.

References herein to "ABC transporter iron-binding protein" include the product of the piaA gene which is the substrate binding protein from an iron transporter and is involved in iron acquisition and has a cluster of orthologous genes (COG) designation of CLS00926, using the scheme defined in Croucher et al (2015) Sci. Data 2: 150058.

In one embodiment, the pneumococcal vaccine comprises two or more of said proteins. In a further embodiment, the pneumococcal vaccine comprises three or more of said proteins.

In a further embodiment, the pneumococcal vaccine comprises four or more of said proteins. In a further embodiment, the pneumococcal vaccine comprises five or more of said proteins. In a further embodiment, the pneumococcal vaccine comprises six or more of said proteins. In a further embodiment, the pneumococcal vaccine comprises seven or more of said proteins.

In a further embodiment, the pneumococcal vaccine comprises all eight of said proteins.

In one embodiment, the vaccine composition comprises one or more proteins which are present on capsular pneumococcal strains and absent on non-capsular pneumococcal strains.

In an alternative embodiment, the vaccine composition comprises one or more nucleic acid molecules encoding said one or more proteins described herein. References herein to "nucleic acid molecules" typically refer to DNA or RNA. In a further embodiment, the one or more nucleic acid molecules comprise one or more oligonucleotides encoding said one or more proteins. Without being bound by theory, it will be appreciated that said one or more proteins function as antigens within the vaccine composition and provide the function of being immunogenic (i.e. generating an immune response within the individual). It is envisaged that either the full length proteins or nucleic acids described herein will be present within the vaccine composition or fragments of said proteins or nucleic acids may suitably be employed.

In one embodiment, at least about 1 μg and up to about 200 μg of each of said proteins is present within the vaccine composition. In one embodiment, the vaccine composition additionally comprises one or more adjuvants. References herein to the term "adjuvant" refer to a compound that, when used in

combination with a specific immunogen in a formulation, will augment or otherwise alter or modify the resultant immune response. Modification of the immune response can include intensification or broadening the specificity of either or both antibody and cellular immune responses. Modification of the immune response can also mean decreasing or suppressing certain antigen-specific immune responses.

In a further embodiment, at least about 1 μg and up to about 20 μg adjuvant is present within the vaccine composition. Examples of suitable adjuvants include: alum; aluminum hydroxide; aluminum phosphate; calcium phosphate hydroxide; paraffin oil; killed bacteria such as

Bordetella pertussis, Mycobacterium bovis and toxoids; squalene, detergents; plant saponins from quillaja, soybean, polygala senega; cytokines such as IL-1 , IL-2, IL-12; Freund's complete adjuvant; and Freund's incomplete adjuvant. In one embodiment, the vaccine composition additionally comprises one or more capsular polysaccharides. In a further embodiment, the capsular polysaccharides are selected from serotypes: 1 , 2, 3, 4, 5, 6B, 7F, 8, 9N, 9V, 10A, 11A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F and 33F of S. pneumoniae. In a yet further embodiment, the capsular

polysaccharides are selected from serotypes: 1 , 3, 4, 5, 6B, 7F, 9N, 9V, 14, 15B, 18C, 19A, 19F, 22F, 23F and 33F of S. pneumoniae.

In one embodiment, the vaccine composition additionally comprises a pharmaceutically acceptable carrier, diluent, excipient, or combination thereof, in which the immunogen (i.e. one or more proteins as defined herein) is/are suspended or dissolved.

Pharmaceutically acceptable carriers are known, and include but are not limited to, water for injection, saline solution, buffered saline, dextrose, water, glycerol, sterile isotonic aqueous buffer, and combinations thereof. For parenteral administration, such as subcutaneous injection, the carrier may include water, saline, alcohol, a fat, a wax, a buffer or combinations thereof. Pharmaceutically acceptable carriers, diluents, and other excipients are described in detail in Remington's Pharmaceutical Sciences (Mack Pub. Co. N.J. current edition). The formulation should suit the mode of administration. In a preferred embodiment, the formulation is suitable for administration to humans, preferably is sterile, non-particulate and/or non-pyrogenic.

In other embodiments, the vaccine composition can include one or more diluents, preservatives, solubilizers and/or emulsifiers. For example, the vaccine composition can include minor amounts of wetting or emulsifying agents, or pH buffering agents to improve vaccine efficacy. The composition can be a solid form, such as a lyophilized powder suitable for reconstitution, a liquid solution, suspension, emulsion, tablet, pill, capsule, sustained release formulation, or powder. Oral formulation can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium

saccharine, cellulose, magnesium carbonate, etc.

It may also be desirable to include other components in a vaccine composition, such as delivery vehicles including but not limited to aluminum salts, water-in-oil emulsions, biodegradable oil vehicles, oil-in-water emulsions, biodegradable microcapsules, and liposomes. In other embodiments, the vaccine composition can include antibacterial agents such as benzyl alcohol or methyl paraben; antioxidants such as ascorbic acid or sodium bisulfite; chelating agents such as ethylenediaminetetraacetic acid; buffers such as acetates, citrates or phosphates and agents for the adjustment of tonicity such as sodium chloride or dextrose.

Administration of the vaccine composition can be systemic or local. Methods of

administering a vaccine composition include, but are not limited to, parenteral administration (e.g., intradermal, intramuscular, intravenous and subcutaneous), epidural, and mucosal (e.g., intranasal and oral or pulmonary routes or by suppositories). In a specific embodiment, compositions described herein are administered intramuscularly, intravenously,

subcutaneously, transdermal^ or intradermally. The compositions may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucous, colon, conjunctiva, nasopharynx, oropharynx, vagina, urethra, urinary bladder and intestinal mucosa, etc.) and may be administered together with other biologically active agents. In some embodiments, intranasal or other mucosal routes of administration of a composition may induce an antibody or other immune response that is substantially higher than other routes of administration. In another embodiment, intranasal or other mucosal routes of administration of a composition described herein may induce an antibody or other immune response at the site of immunization. In one embodiment, the vaccine composition has a volume of between about 50 μΙ and about 500 μΙ.

References herein to "pneumococcal infection" refer to infection by Streptococcus pneumoniae. References herein to "effective amount" refer to a dose which is sufficient or most likely to elicit antibodies such that the immunized subject has reduced severity of infection.

According to a further aspect of the invention, there is provided a method of inducing an immune response in a mammal, wherein the method includes administering to the mammal, an effective amount of a vaccine composition described herein.

In one embodiment, the mammal is a human. In another embodiment, the mammal is selected from: an adult such as an elderly adult greater than 65 years old; an infant less than 1 year old; a toddler between 1 and 2 years old; or a young child between 2 and 5 years old.

In one embodiment, the vaccine composition is administered in a single dose regimen. In another embodiment, the vaccine composition is administered in a two dose regimen that includes a first and a second dose. In one embodiment, the second dose is administered at least about 1 week, 2 weeks, 3 weeks, 1 month or 1 year after the first dose. In another embodiment, the vaccine composition is administered in a three dose regimen.

According to a further aspect of the invention, there is provided a kit of parts comprising a vaccine composition as described herein, a medical instrument or other means for administering the vaccine composition and instructions for use.

In one embodiment, the vaccine composition is packaged in a hermetically sealed container such as an ampoule or sachette indicating the quantity of composition. In one embodiment, the composition is supplied as a liquid. In another embodiment, the composition is supplied as a dry sterilized lyophilized powder or water free concentrate in a hermetically sealed container, wherein the composition can be reconstituted, for example, with water or saline, to obtain an appropriate concentration for administration to a subject.

When the vaccine composition is systemically administered, for example, by subcutaneous or intramuscular injection, a needle and syringe, or a needle-less injection device can be used. The vaccine formulation can be enclosed in ampoules, disposable syringes or multiple dose vials made of glass or plastic.

The following study illustrates the invention described herein:

The genetic diversity of pneumococci isolated from cases of asymptomatic carriage has been previously characterised^2,3. Following sequencing with lllumina technology, contigs were assembled using Velvet⁴. Protein coding sequences were then predicted using

Prodigal⁵ and Glimmer⁶, and the protein sequences translated and aligned using BLAT⁷. These were grouped into 'clusters of orthologous genes' (COGs) using COGtriangles⁸ and custom scripts². Concatenation of the 'core' COGs allowed a phylogeny to be constructed using RAxML⁹, and a population clustering to be generated with BAPS¹⁰.

The first step was to use this analysis of population structure to identify a set of

unencapsulated streptococcal isolates that were highly divergent from the rest of the population in terms of their core genome sequence and gene content. The inventors hypothesised that these genetic differences were likely to make them antigenically distinct³. The inventors used this analysis to define a set of protein orthologues that are present in 5% or less of the divergent unencapsulated isolates (originally defined as "sequence cluster 12" (SC12)) and present in 95% or more of the rest of the population (defined as being outside SC12); the proteins presented in Table 1 are a subset of this set. Table 1 : Antigenic proteins very common in encapsulated pneumococci but absent from 'divergent' or 'classic' non-typeable pneumococci (SC12).

transcriptional regulator

CLS01686 1.03 0 1.00 plcR PlcR

sucrose-6-phosphate

CLS01545 1.05 0 1.00 sacC hydrolase

CLS01869 1.13 0 1.00 fuel L-fucose isomerase

CLS01862 1.14 0 1.00 arcC carbamate kinase surface anchored beta-

CLS00596 2.65 0 1.00 bgaA galactosidase

capsule biosynthesis integral membrane

CLS00362 2.84 0 0.98 wzg regulatory protein Wzg surface anchored endo- alpha-N- acetylgalactosaminidas

CLS00380 4.03 0 1.00 eng e

ABC transporter iron-

CLS00926 4.51 0 1.00 piaA binding protein

The second step was confirming the antigenic distinctiveness of SC12 through analysis of immunoglobulin G binding to pneumococcal proteins expressed in ^'fro¹¹. Across a sample of 35 clinical trial participants, a threshold of a mean normalised log2 immunoglobulin binding value of at least one, indicating a doubling relative to background, was used to identify antigens. This study can be used to identify eight antigenic proteins that were ubiquitous in encapsulated pneumococci, but absent from SC12 streptococci (Table 1). One is involved in iron acquisition: PiaA, the substrate binding protein from an iron transporter. Two are surface-associated degradative enzymes: Eng, a cell-wall associated endo-alpha-/V- acetylgalactosaminidase, and BgaA, a beta galactosidase. One is a surface-associated enzyme involved in regulation of capsule production: Wzg, an integral membrane protein. Three are involved in carbohydrate metabolism: ArcC, a carbamate kinase; Fuel, an L- fucose isomerase; and SacC, a sucrose-6-phosphate hydrolase. Finally, the list also includes a transcriptional regulator, PlcR.

The third step was identifying that encapsulated and unencapsulated pneumococci frequently co-circulate in the same populations, and therefore are likely to compete with one another. This has been inferred from large-scale genomics projects such as the survey of the Maela refugee camp, where unencapsulated pneumococci were very common¹². The following description illustrates the general principles of the invention described herein:

The identification of antigens that might facilitate the replacement of the encapsulated pneumococcal population by unencapsulated pneumococci or pneumococci-like streptococci would first require a collection of genetic data on a set of pneumococci and pneumococci- like organisms. Protein coding sequences could be predicted in an automated manner using software such as Glimmer⁶ or Prodigal⁵, or determined manually¹³, based on the genetic data. The encapsulation status of these bacteria may be inferred from the presence of protein coding sequences associated with the synthesis of pneumococcal capsules¹⁴, else determined experimentally using antisera, polymerase chain reactions, microarrays, or other genotyping method^15,16. The proteins encoded by protein coding sequences can be predicted through conventional nucleic acid to amino acid sequence translation software¹⁷. The predicted proteins can be clustered into groups likely to perform similar functions based on similarity of protein sequence following pairwise alignment using algorithms such as

BLAST¹⁸ or BLAT⁷, and subsequent analysis with clustering software such as

GET_HOMOLOGUES¹⁹, COGtriangles⁸ or TribeMCL²⁰. Those proteins that are unique to, or enriched in, the unencapsulated pneumococci or pneumococci-like streptococci can then be identified by quantifying the distribution of sequences encoding orthologous proteins in the genetic data associated with encapsulated and unencapsulated bacteria. The subset of those proteins likely to be viable vaccine antigens can then be identified on the basis of in silico immunogenicity prediction²⁴ and/or direct experimental evidence of surface exposure, such as proteomics²¹ or immunological work in mammals^22,23, or identification of particular functional motifs associated with secretion, bacterial surface association, or immunogenicity in mammals¹¹.

References

1. Hilty, M. et al. Global phylogenomic analysis of nonencapsulated Streptococcus

pneumoniae reveals a deep-branching classic lineage that is distinct from multiple sporadic lineages. Genome Biol. Evol. 6, 3281-3294 (2014).

2. Croucher, N. J. et al. Population genomics of post-vaccine changes in pneumococcal epidemiology. Nat. Genet. 45, 656-663 (2013).

3. Croucher, N. J. et al. Diversification of bacterial genome content through distinct

mechanisms over different timescales. Nat. Commun. 5, 5471 (2014).

4. Zerbino, D. R. & Birney, E. Velvet: algorithms for de novo short read assembly using de Bruijn graphs. Genome Res 18, 821-829 (2008).

5. Hyatt, D. et al. Prodigal: prokaryotic gene recognition and translation initiation site identification. BMC Bioinformatics 11 , 1 19 (2010).

6. Delcher, A. L, Bratke, K. A., Powers, E. C. & Salzberg, S. L. Identifying bacterial genes and endosymbiont DNA with Glimmer. Bioinformatics 23, 673-679 (2007).

7. Kent, W. J. BLAT-the BLAST-like alignment tool. Genome Res 12, 656-664 (2002). 8. Kristensen, D. M. et al. A low-polynomial algorithm for assembling clusters of

orthologous groups from intergenomic symmetric best matches. Bioinformatics 26, 1481-1487 (2010).

9. Stamatakis, A., Ludwig, T. & Meier, H. RAxML-lll: a fast program for maximum

likelihood-based inference of large phylogenetic trees. Bioinformatics 21 , 456-463 (2005).

10. Cheng, L, Connor, T. R., Siren, J., Aanensen, D. M. & Corander, J. Hierarchical and spatially explicit clustering of DNA sequences with BAPS software. Mol. Biol. Evol. 30, 1224-1228 (2013).

1 1. Croucher, H. J. et al. Diverse evolutionary patterns of pneumococcal antigens

identified by pangenome-wide immunological screening. Proc. Natl. Acad. Sci.

(2017) . doi: 10.1073/pnas.1613937114

12. Chewapreecha, C. et al. Dense genomic sampling identifies highways of

pneumococcal recombination. Nat. Genet. 46, 305-309 (2014).

13. Carver, T. et al. Artemis and ACT: viewing, annotating and comparing sequences stored in a relational database. Bioinformatics 24, 2672-2676 (2008).

14. Croucher, N. J. et al. Population genomic datasets describing the post-vaccine

evolutionary epidemiology of Streptococcus pneumoniae. Sci. Data 2, 150058 (2015).

15. Turner, P. et al. Improved detection of nasopharyngeal cocolonization by multiple pneumococcal serotypes by use of latex agglutination or molecular serotyping by microarray. J. Clin. Microbiol. 49, 1784-1789 (2011).

16. Salter, S. J. et al. Variation at the capsule locus, cps, of mistyped and non-typable Streptococcus pneumoniae isolates. Microbiology 158, 1560-1569 (2012).

17. Rice, P., Longden, I. & Bleasby, A. EMBOSS: the European Molecular Biology Open Software Suite. Trends Genet 16, 276-277 (2000).

18. Camacho, C. et al. BLAST+: architecture and applications. BMC Bioinformatics 10,

421 (2009).

19. Contreras-Moreira, B. & Vinuesa, P. GET_HOMOLOGUES, a versatile software

package for scalable and robust microbial pangenome analysis. Appl. Environ.

Microbiol. 79, 7696-7701 (2013).

20. Enright, A. J., Van Dongen, S. & Ouzounis, C. A. An efficient algorithm for large-scale detection of protein families. Nucleic Acids Res. 30, 1575-1584 (2002).

21. Cordwell, S. J. Technologies for bacterial surface proteomics. Current Opinion in Microbiology 9, 320-329 (2006).

22. Croucher, N. J. et al. Diverse evolutionary patterns of pneumococcal antigens

identified by pangenome-wide immunological screening. Proc. Natl. Acad. Sci. U. S. A. 114, E357-E366 (2017).

23. Wilson, R. et al. Naturally Acquired Human Immunity to Pneumococcus Is Dependent Journal of Biomedical Informatics on Antibody to Protein Antigens. PLOS Pathog. 13, 1-26 (2017)

24. Soria Guerra, R. et al. Volume 53, February 2015, Pages 405-414. An overview of bioinformatics tools for epitope prediction: Implications on vaccine development

Claims

1. A pneumococcal vaccine comprising one or more proteins, or one or more nucleic acid molecules encoding said proteins, which are present on capsular pneumococcal strains and absent on non-capsular pneumococcal strains.

2. The pneumococcal vaccine as defined in claim 1 , which comprises one or more proteins which are present on capsular pneumococcal strains and absent on non-capsular pneumococcal strains.

3. The pneumococcal vaccine as defined in claim 1 , which comprises one or more nucleic acid molecules encoding said one or more proteins which are present on capsular pneumococcal strains and absent on non-capsular pneumococcal strains.

4. The pneumococcal vaccine as defined in claim 3, wherein said one or more nucleic acid molecules comprise one or more oligonucleotides encoding said one or more proteins.

5. The pneumococcal vaccine as defined in any one of claims 1 to 4, wherein the one or more proteins are selected from one or more of: transcriptional regulator PlcR, putative; sucrose-6-phosphate hydrolase; L-fucose isomerase; carbamate kinase; surface anchored beta-galactosidase; capsule biosynthesis integral membrane regulatory protein Wzg; surface anchored endo-alpha-N-acetylgalactosaminidase; and ABC transporter iron-binding protein.

6. The pneumococcal vaccine as defined in any one of claims 1 to 5, which comprises; two or more of said proteins; or three or more of said proteins; or four or more of said proteins; or five or more of said proteins; or six or more of said proteins; or seven or more of said proteins; or all eight of said proteins.

7. A pharmaceutical composition comprising a pneumococcal vaccine as defined in any one of claims 1 to 6.

8. The pharmaceutical composition as defined in claim 7, which additionally comprises one or more adjuvants.

9. The pharmaceutical composition as defined in claim 7 or claim 8, which additionally comprises one or more capsular polysaccharides.

10. The pharmaceutical composition as defined in claim 9, wherein the capsular polysaccharides are selected from serotypes: 1 , 2, 3, 4, 5, 6B, 7F, 8, 9N, 9V, 10A, 1 1A, 12F, 14, 15B, 17F, 18C, 19A, 19F, 20, 22F, 23F and 33F of S. pneumoniae.

1 1. The pharmaceutical composition as defined in claim 10, wherein the capsular polysaccharides are selected from serotypes: 1 , 3, 4, 5, 6B, 7F, 9N, 9V, 14, 15B, 18C, 19A, 19F, 22F, 23F and 33F of S. pneumoniae.

12. The pharmaceutical composition as defined in any one of claims 7 to 1 1 , which additionally comprises a pharmaceutically acceptable carrier, diluent, excipient, or combination thereof.

13. The pharmaceutical composition as defined in any one of claims 7 to 12, which is adapted for parenteral administration (e.g., intradermal, intramuscular, intravenous and subcutaneous), epidural, and mucosal (e.g., intranasal and oral or pulmonary routes or by suppositories).

14. A method of preventing pneumococcal infection in a mammal which comprises administering to the mammal a therapeutically effective amount of a vaccine composition as defined in any one of claims 7 to 13.

15. A method of inducing an immune response in a mammal, wherein the method includes administering to the mammal, an effective amount of a vaccine composition as defined in any one of claims 7 to 13.

16. The method as defined in claim 14 or claim 15, wherein the mammal is a human.

17. The method as defined in claim 14 or claim 15, wherein the mammal is selected from: an adult such as an elderly adult greater than 65 years old; an infant less than 1 year old; a toddler between 1 and 2 years old; or a young child between 2 and 5 years old.

18. A kit of parts comprising a vaccine composition as defined in any one of claims 7 to 13, a medical instrument or other means for administering the vaccine composition and instructions for use.