Rhinovirus vaccine

Abstract

The present invention relates to immunogenic compositions, and in particular, to immunogenic compositions for preventing, treating or ameliorating human rhinovirus (RV) infections. The invention is especially concerned with RV VP0 peptides (or proteins) and polynucleotides encoding such peptides, and their use in immunogenic compositions for eliciting an immune response and preventing rhinovirus infections.

Description

[0001] Rhinovirus Vaccine

[0002] The present invention relates to immunogenic compositions, and in particular, to immunogenic compositions for preventing, treating or ameliorating human rhinovirus (RV) infections. The invention is especially concerned with RV VPO peptides (or proteins) and polynucleotides encoding such peptides, and their use in immunogenic compositions for eliciting an immune response and preventing rhinovirus infections.

[0003] Rhinoviruses (RVs) are picornaviruses, which are small single stranded RNA viruses with a genome size of approximately 6.8-7.2 Kb. RVs are the major cause of common colds, of virus-induced wheezing illnesses in early childhood and of acute attacks (exacerbations) of lung diseases such as asthma, bronchiectasis, chronic obstructive pulmonary disease (COPD), cystic fibrosis and chronic fibrosing lung diseases. RVs cause around 75% to 90% of such illnesses.

[0004] There are currently no effective anti-RV treatments and no anti-RV vaccines. A major obstacle to the development of a RV vaccine has been the large number of genetically and antigenically distinct circulating strains. A recent study estimated that approximately 180 genetically distinct strains of RV are in circulation. These strains are classified based on their genome identity into three species: RV-A, RV-B and RV-C. RV-A and RV-C are the largest species numerically and are most important in terms of clinical illness, with RV-A and RV-Cs responsible for most exacerbations of asthma and COPD and childhood wheezing illnesses resulting in hospitalisation. However, RV-Bs are also associated with an increased risk of wheezing illnesses.

[0005] There is, therefore, a need to provide an immunogenic composition that is capable of protecting individuals against many, if not all, RV strains.

[0006] The inventors previous work identified RV VPO proteins from single representative strains that, when used as vaccine immunogens, surprisingly evoke cellular immunity against all the other members of that RV species. In particular, the inventors discovered that combinations of VPO peptides used together in a single vaccine are particularly effective for immunising humans against infections with all RV strains.

[0007] However, the inventors have now discovered that a single RV VPO protein, RV-C04 VPO, is particularly effective at eliciting a broad cellular immune response, when adjuvanted with alum and CpG 7909. Therefore, according to a first aspect of the invention, there is provided an immunogenic composition comprising : (i) a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof, or a polynucleotide encoding the peptide, variant or fragment thereof; (ii) a CpG oligonucleotide; and (iii) an aluminium salt.

[0008] As shown in the examples, the inventors surprisingly discovered that the RV-C04 VPO peptide, when adjuvanted with aluminium salt and CpG 7909, is able to elicit broad cellular immune responses that cross-react with other RV strains from the same species. For example, as shown in Figure 1, animals immunized with recombinant RV- C04 VPO alone produce greater than 100-fold lower vaccine-specific IgG than animals immunized with RV-C04 VPO plus alum and CpG 7909.

[0009] It is well-documented that there are three different species of rhinoviruses: rhinovirus A (RV-A), which is also called type A rhinovirus, rhinovirus B (RV-B), which is also called type B rhinovirus, and rhinovirus C (RV-C), which is also called type C rhinovirus. RVs are further classified into strains according to their nucleotide sequence homologies. As used herein, the term "strain" refers to a subdivision within a species of rhinoviruses and relies on the VP1 gene sequence of the rhinovirus. For example, the term RV-C04 refers to the RV strain. RVs have been classified according to several other parameters, including receptor specificity and antiviral susceptibility.

[0010] RVs have a 25 nm capsid of icosahedral symmetry, made up of 60 copies of each of four virus-coded proteins (VP1, VP2, VP3 and VP4) and enclosing a single-stranded RNA genome of approximately 7,500 nucleotides. The RNA is of positive polarity, is polyadenylated at its 3' terminus and is covalently bound at its 5' terminal end to a small protein, VPg. The primary translational product of this RNA is a single, "large" polyprotein, divided into three smaller polyproteins called, Pl, P2 and P3, which are subsequently processed by proteolytic cleavage to yield the mature virus proteins. The Pl polyprotein is composed of four peptides (1A or VP4, IB or VP2, 1C or VP3, and ID or VP1), the P2 polyprotein is composed of three peptides (2A, 2B and 2C) and the P3 polyprotein is composed of four peptides (3A, 3B, 3C and 3D). 2A and 3C are viral proteases, while 3D corresponds to the viral RNA polymerase. The Pl polyprotein is the precursor that gives rise to the four structural proteins of the nucleocapsid. The Pl polyprotein is first cleaved to produce the VPO polyprotein, which contains the amino acid sequence of VP4 and VP2 peptides, the VP3 peptide and the VP1 peptide. The VPO polyprotein is then cleaved into the VP4 peptide and the VP2 peptide once the virus has assembled. It is well-documented that the term "VPO polyprotein", "VPO peptide" or "peptide 1AB" refers to the protein precursor derived from the RV Pl polyprotein and which consists of the amino acid sequence of VP4 and VP2 peptides. VPO polyprotein is typically about 330 amino acids long. The amino acid sequence of the VPO polyprotein slightly varies according to the RV strain or species.

[0011] Accordingly, a fragment of the RV VPO peptide may comprise or consist of the corresponding VP2 peptide and / or VP4 peptide. Thus, a fragment of the RV VPO may have at least 1, 2, 3, 4, 5, 10, 20, 25, 30, 45, 50, 60, 70, 80, 90 or 100 amino acids fewer than the full-length RV VPO described herein.

[0012] In one embodiment, the peptide, variant or fragment thereof, comprises or consists of at least 40 amino acids, at least 45 amino acids, at least 50 amino acids, at least 55 amino acids, at least 60 amino acids, at least 65 amino acids, at least 66 amino acids, at least 67 amino acids, at least 68 amino acids, or at least 69 amino acids.

[0013] In another embodiment, the peptide, variant or fragment thereof, comprises or consists of at least 70 amino acids, at least 80 amino acids, at least 90 amino acids, at least 100 amino acids, at least 110 amino acids, at least 120 amino acids, at least 130 amino acids, at least 140 amino acids, at least 150 amino acids, at least 160 amino acids, at least 170 amino acids, at least 180 amino acids, at least 190 amino acids, at least 200 amino acids, at least 210 amino acids, at least 220 amino acids, at least 230 amino acids, at least 240 amino acids, at least 250 amino acids, at least 255 amino acids, at least 260 amino acids, at least 261 amino acids, at least 262 amino acids, at least 263 amino acids, at least 264 amino acids, at least 265 amino acids, at least 266 amino acids, or at least 267 amino acids.

[0014] In another embodiment, the peptide, variant or fragment thereof, comprises or consists of at least 280 amino acids, at least 290 amino acids, at least 300 amino acids, at least 310 amino acids, at least 320 amino acids, or at least 330 amino acids.

[0015] In one embodiment, the human rhinovirus (RV)-C04 VPO peptide, or variant or fragment thereof is isolated. In one embodiment, the polynucleotide encoding the peptide, variant or fragment thereof is isolated. Accordingly, in one embodiment, there is provided an immunogenic composition comprising: (i) an isolated human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof, or an isolated polynucleotide encoding the peptide, variant or fragment thereof; (ii) a CpG oligonucleotide; and (iii) an aluminium salt. As used herein, the term "isolated" means removed from the natural environment, i.e. from rhinoviruses or cells infected by a rhinovirus. Usually, it refers to a peptide or a nucleic acid substantially free of cellular material, bacterial material, viral material, or culture medium when produced by recombinant DNA techniques, or chemical precursors, or other chemicals when chemically synthesized.

[0016] The present invention also relates to an immunogenic composition comprising a polynucleotide encoding the human rhinovirus (RV)-C04 VPO peptide.

[0017] In one embodiment, the polynucleotide comprises a nucleic acid sequence encoding the peptide, or a variant or fragment thereof. In one embodiment, the nucleic acid sequence is placed under the control of the elements necessary for its expression in a mammalian cell, in particular in human cells. The nucleic acid sequence may be incorporated in a plasmid, which can be further formulated in a delivery vehicle such as liposomes to facilitate its introduction into the host cell.

[0018] As used herein, the term "nucleic acid" includes DNA and RNA, such as messenger RNA (mRNA) or self-amplifying RNA (saRNA), and can be either double-stranded or single-stranded. In one embodiment, the nucleic acid is mRNA.

[0019] As used herein, the expression "elements necessary for expression in a mammalian cell" is understood to mean all the elements which allow the transcription of a DNA or DNA fragment into mRNA or saRNA and the translation of the latter into protein, inside a mammalian cell, such as a human cell. Typically, the elements necessary for the expression of a nucleic acid in a mammalian cell include a promoter that is functional in the selected mammalian cell and can be constitutive or inducible; a ribosome binding site; a start codon (ATG) if necessary; a region encoding a signal peptide (e.g., a lipidation signal peptide); a stop codon; and a 3' terminal region (translation and / or transcription terminator). Other transcription control elements, such as enhancers, operators, and repressors can be also operatively associated with the polynucleotide to direct transcription and / or translation into the cell. The signal peptide-encoding region may be adjacent to the nucleic acid included in the immunogenic composition of the invention and placed in proper reading frame. The signal peptide-encoding region can be homologous or heterologous to the DNA molecule encoding the mature peptide or fusion peptide of the invention and can be specific to the secretion apparatus of the host used for expression. The open reading frame constituted by the nucleic acid included in the immunogenic composition of the invention, solely or together with the signal peptide, is placed under the control of the promoter so that transcription and translation occur in the host system. Promoters, (and signal peptide encoding regions) are widely known and available to those skilled in the art.

[0020] The nucleic acid sequences may be codon optimized such that the transcription of the DNA encoding the peptide of the invention is enhanced and / or the translation of the mRNA encoding the peptide is prolonged.

[0021] In one embodiment, the amino acid sequence of the RV-C04 VPO peptide is provided herein as SEQ ID No: 1, as follows:

[0022] MGAQVSKQNVGSHENSVSATGGSVIKYFNINYYKDSASSGLTKQDFSQDPSKFTQPLAEALTNPALMSPTVEA CGMSDRLKQITIGNSTITTQDTLNSILAYGEWPKYLSDLDASSVDKPTHPETSSDRFYTLTSVDWTTTSKGWW WKLPDCLKDMGIFGQNLYHHALGRSGYIIHTQCNATKFNSGCLIVAWPEHQLAYIGEANVNVGYDHTHPGEG GHVIGSNVRRDNKQPDEDPFFNCNGTLLGNITIFPHQLINLRTNNSSTIWPYINCVPMDNMLRHNNLSLVII PIVPLRAANGVTKVPITISIAPDKSEFSGARQSVKQ

[0023] [SEQ ID No: 1]

[0024] Accordingly, in one embodiment, the RV-C04 VPO peptide comprises an amino acid sequence substantially as set out in SEQ ID No: 1, or a variant or fragment thereof.

[0025] The VP4 peptide of RV-C04 constitutes amino acid residues 1-67 of SEQ ID No: 1 and the VP2 peptide of RV-C04 constitutes amino acid residues 68-328 of SEQ ID No: 1.

[0026] Thus, in embodiments in which the immunogenic composition comprises a fragment of RV-C04 VPO, the fragment may comprise or consist of the amino acids forming the

[0027] VP2 and / or VP4 peptides.

[0028] In one embodiment, the RV-C04 VPO peptide is encoded by the nucleotide sequence (DNA) of SEQ ID No: 2, as follows:

[0029] ATGGGCGCACAGGTCAGCAAGCAAAATGTCGGCTCGCATGAAAACTCAGTCTCAGCCACAGGTGGATCCGTGA TTAAGTATTTCAACATCAATTACTACAAGGATTCTGCTAGCTCTGGCTTGACTAAACAAGATTTTTCCCAAGA CCCATCGAAATTCACACAACCTCTAGCAGAAGCACTTACAAATCCAGCTTTAATGTCACCAACTGTTGAAGCA TGTGGGATGTCCGATAGGCTTAAACAAATTACTATCGGGAATTCCACTATAACAACACAAGATACACTAAACT CTATACTGGCATATGGGGAGTGGCCCAAATACTTGAGTGACCTGGACGCTTCCTCAGTGGATAAACCTACCCA CCCAGAAACATCATCTGATAGATTTTACACATTAACTAGTGTAGATTGGACCACTACGTCTAAAGGTTGGTGG TGGAAGTTGCCTGATTGCCTTAAAGATATGGGCATCTTCGGGCAAAATCTGTACCATCATGCATTGGGTAGGT CAGGGTACATAATACACACCCAATGTAATGCCACAAAATTCAATAGTGGTTGTCTAATAGTGGCTGTTGTACC AGAACACCAGCTAGCTTACATAGGTGAAGCAAATGTCAATGTTGGTTATGATCACACACACCCTGGTGAGGGA GGACATGTAATTGGTTCAAATGTTAGGAGAGATAACAAGCAACCTGATGAAGACCCCTTCTTTAATTGTAATG GGACCCTGCTTGGTAACATCACTATATTCCCACACCAGCTCATAAACTTGAGGACAAACAATTCCAGCACAAT TGTTGTACCATACATTAATTGTGTACCTATGGACAACATGCTCAGGCACAACAACCTATCTCTAGTTATTATT CCAATCGTTCCTCTCAGAGCCGCAAATGGTGTCACCAAGGTCCCCATTACAATCTCAATAGCACCAGATAAGT CAGAGTTCTCAGGGGCTAGACAGTCTGTAAAACAG [SEQ ID No: 2]

[0030] Accordingly, in one embodiment, the RV-C04 VPO peptide is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 2, or a variant or fragment thereof. Alternatively, the immunogenic composition comprises a polynucleotide comprising the nucleotide sequence substantially as set out in SEQ ID No: 2, or a variant or fragment thereof.

[0031] In one embodiment, the RNA sequence corresponding to the DNA sequence of SEQ ID

[0032] No: 2 is provided herein as SEQ ID No: 3, as follows:

[0033] AUGGGCGCACAGGUCAGCAAGCAAAAUGUCGGCUCGCAUGAAAACUCAGUCUCAGCCACAGGUGGAUCCGUGA UUAAGUAUUUCAACAUCAAUUACUACAAGGAUUCUGCUAGCUCUGGCUUGACUAAACAAGAUUUUUCCCAAGA CCCAUCGAAAUUCACACAACCUCUAGCAGAAGCACUUACAAAUCCAGCUUUAAUGUCACCAACUGUUGAAGCA UGUGGGAUGUCCGAUAGGCUUAAACAAAUUACUAUCGGGAAUUCCACUAUAACAACACAAGAUACACUAAACU CUAUACUGGCAUAUGGGGAGUGGCCCAAAUACUUGAGUGACCUGGACGCUUCCUCAGUGGAUAAACCUACCCA CCCAGAAACAUCAUCUGAUAGAUUUUACACAUUAACUAGUGUAGAUUGGACCACUACGUCUAAAGGUUGGUGG UGGAAGUUGCCUGAUUGCCUUAAAGAUAUGGGCAUCUUCGGGCAAAAUCUGUACCAUCAUGCAUUGGGUAGGU CAGGGUACAUAAUACACACCCAAUGUAAUGCCACAAAAUUCAAUAGUGGUUGUCUAAUAGUGGCUGUUGUACC AGAACACCAGCUAGCUUACAUAGGUGAAGCAAAUGUCAAUGUUGGUUAUGAUCACACACACCCUGGUGAGGGA GGACAU GUAAUU G GUU CAAAU GUU AG GAGAGAUAACAAG CAAC CU GAU GAAGAC C C CUU CUUUAAUU GUAAU G GGACCCUGCUUGGUAACAUCACUAUAUUCCCACACCAGCUCAUAAACUUGAGGACAAACAAUUCCAGCACAAU UGUUGUACCAUACAUUAAUUGUGUACCUAUGGACAACAUGCUCAGGCACAACAACCUAUCUCUAGUUAUUAUU CCAAUCGUUCCUCUCAGAGCCGCAAAUGGUGUCACCAAGGUCCCCAUUACAAUCUCAAUAGCACCAGAUAAGU CAGAGUUCUCAGGGGCUAGACAGUCUGUAAAACAGCAG

[0034] [SEQ ID No: 3]

[0035] Accordingly, in one embodiment, the immunogenic composition comprises a polynucleotide comprising the nucleotide sequence (RNA) substantially as set out in SEQ ID No: 3, or a variant or fragment thereof.

[0036] In one embodiment, the immunogenic composition comprises or encodes between 15 and 150 pg, between 20 and 125 pg, or between 25 and 100 pg of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0037] In another embodiment, the immunogenic composition comprises or encodes between 25 and 140 pg, between 25 and 130 pg, between 25 and 120 pg, between 25 and 110 pg, or between 25 and 100 pg of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0038] In another embodiment, the immunogenic composition comprises or encodes between 20 and 100 pg, between 25 and 100 pg, between 30 and 100 pg, between 35 and 100 pg, between 40 and 100 pg, between 45 and 100 pg, or between 50 and 100 pg of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof. In some embodiments, the immunogenic composition comprises or encodes 25 pg, 50 pg, or 100 pg of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0039] In one embodiment, the immunogenic composition comprises or encodes between 5 and 550 pg / mL, between 10 and 525 pg / mL, between 20 and 500 pg / mL, between 30 and 475 pg / mL, between 40 and 450 pg / mL, or between 50 and 425 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof. In one embodiment, the immunogenic composition comprises or encodes between 60 and 400 pg / mL, between 70 and 375 pg / mL, between 80 and 350 pg / mL, between 90 and 325 pg / mL, or between 100 and 300 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0040] In one embodiment, the immunogenic composition comprises or encodes between 50 and 550 pg / mL, between 50 and 525 pg / mL, between 50 and 500 pg / mL, between 50 and 475 pg / mL, between 50 and 450 pg / mL, between 50 and 425 pg / mL, between 50 and 400 pg / mL, between 50 and 375 pg / mL, between 50 and 350 pg / mL, between 50 and 325 pg / mL, or between 50 and 300 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0041] In another embodiment, the immunogenic composition comprises or encodes between 25 and 550 pg / mL, between 50 and 550 pg / mL, between 75 and 550 pg / mL, between 100 and 550 pg / mL, between 150 and 550 pg / mL, between 200 and 550 pg / mL, between 250 and 550 pg / mL, between 300 and 550 pg / mL, between 350 and 550 pg / mL, between 400 and 550 pg / mL, or between 450 and 550 pg / mL, of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0042] In one embodiment, the immunogenic composition comprises or encodes between 5 and 250 pg / mL, between 10 and 240 pg / mL, between 20 and 230 pg / mL, between 30 and 220 pg / mL, between 40 and 210 pg / mL, or between 50 and 200 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

[0043] In another embodiment, the immunogenic composition comprises or encodes between 50 and 240 pg / mL, between 50 and 230 pg / mL, between 50 and 220 pg / mL, between

[0044] 50 and 210 pg / mL, between 50 and 200 pg / mL, between 50 and 190 pg / mL, between

[0045] 50 and 180 pg / mL, between 50 and 170 pg / mL, between 50 and 160 pg / mL, or between 50 and 150 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof. In another embodiment, the immunogenic composition comprises or encodes between

[0046] 20 and 250 pg / mL, between 30 and 250 pg / mL, between 40 and 250 pg / mL, between

[0047] 50 and 250 pg / mL, between 60 and 250 pg / mL, between 70 and 250 pg / mL, between

[0048] 80 and 250 pg / mL, between 90 and 250 pg / mL, or between 100 and 250 pg / mL of a human rhinovirus (RV)-C04 VP0 peptide, or a variant or fragment thereof.

[0049] In some embodiments, the immunogenic composition comprises or encodes 125 pg / mL, 250 pg / mL, or 500 pg / mL of a human rhinovirus (RV)-C04 VP0 peptide, or a variant or fragment thereof. Typically, this concentration refers to the final drug substance antigen concentrations (i.e., what has been manufactured).

[0050] In some embodiments, the immunogenic composition comprises or encodes 50 pg / mL, 100 pg / mL, or 200 pg / mL of a human rhinovirus (RV)-C04 VP0 peptide, or a variant or fragment thereof. Typically, this concentration refers to the final drug product antigen concentrations (i.e., what has been created at a pharmacy and injected into patients).

[0051] The CpG oligonucleotide is a DNA fragment containing an unmethylated cytosine- guanine dinucleotide (CpG) motif. It is known to activate B cells, NK cells, dendritic cells, and induce the release of IL-12 and IFN, thereby inducing a strong Thl-type response and cellular immunity.

[0052] In one embodiment, the CpG oligonucleotide is a class A, class B or class C CpG oligonucleotide (ODN). Class B CpG molecules include CpG 1018 (ODN 1018), ODN D- SL01, CpG 7909 (ODN 2006). Class A CpG molecules include ODN 2216 and ODN 2336. Class C CpG molecules include ODN 2395 and ODN M362. Accordingly, in one embodiment, the CpG oligonucleotide is selected from the group consisting of: CpG 1018, ODN D-SL01, CpG 7909, ODN 2216, ODN 2336, ODN 2395 and ODN M362.

[0053] In one embodiment, the CpG oligonucleotide is a Class B CpG oligonucleotide.

[0054] Accordingly, in one embodiment, the CpG oligonucleotide may be selected from the group consisting of: CpG 7909, ODN D-SL01 and CpG 1018.

[0055] In one embodiment, the CpG oligonucleotide is CpG 7909. It is well documented that CpG 7909 (ODN 2006) is an immunostimulatory TLR9 agonist oligodeoxynucleotide. It is a short synthetic single-stranded DNA molecule containing unmethylated CpG dinucleotides (CpG motifs). These unmethylated CpG motifs mimic microbial DNA and act as immunostimulants via TLR9.

[0056] In one embodiment, the nucleotide sequence of CpG 7909 is provided herein as SEQ ID No: 4, as follows: tcgtcgttttgtcgttttgt cgtt

[0057] [SEQ ID No: 4]

[0058] Accordingly, in one embodiment, CpG 7909 comprises the nucleotide sequence substantially as set out in SEQ ID No: 4, or a variant or fragment thereof.

[0059] In one embodiment, the bases of CpG 7909 are phosphorothioate (nuclease resistant).

[0060] In one embodiment, the composition comprises between 50 and 750 pg, between 75 and 725 pg, between 100 and 700 pg, between 125 and 675 pg, between 150 and 650 pg, between 175 and 625 pg, between 200 and 600 pg, between 225 and 575 pg, or between 250 and 550 pg of CpG oligonucleotide.

[0061] In another embodiment, the composition comprises between 50 and 500 pg, between 75 and 500 pg, between 100 and 500 pg, between 125 and 500 pg, between 150 and 500 pg, between 175 and 500 pg, between 200 and 500 pg, between 225 and 500 pg, or between 250 and 500 pg of CpG oligonucleotide.

[0062] In another embodiment, the composition comprises between 250 and 750 pg, between 250 and 725 pg, between 250 and 700 pg, between 250 and 675 pg, between 250 and 650 pg, between 250 and 625 pg, between 250 and 600 pg, between 250 and 575 pg, between 250 and 550 pg, between 250 and 525 pg, or between 250 and 500 pg of CpG oligonucleotide. Typically, the composition comprises between 250 and 500 pg of CpG oligonucleotide.

[0063] In some embodiments, the composition comprises 500 pg of CpG oligonucleotide.

[0064] The presence of aluminium salt induces a Th2 response by improving the attraction and uptake of antigen by antigen-presenting cells (APCs). It can also activate innate immunity pathways triggered by pattern recognition receptors (PRRs). The aluminium salt may be selected from the group consisting of: aluminium hydroxide, aluminium phosphate, aluminium hydroxyphosphate, aluminium orthophosphate, and amorphous aluminium hydroxyphosphate sulfate.

[0065] In one embodiment, the aluminium salt is aluminium hydroxide. In one embodiment, the aluminium hydroxide is aluminium oxyhydroxide. The salt may take any suitable form (e.g. gel, crystalline, amorphous etc.).

[0066] In one embodiment, the composition comprises between 10 and 1000 pg, between 20 and 900 pg, between 30 and 800 pg, between 40 and 700 pg, between 50 and 600 pg, or between 60 and 500 pg of aluminium salt. In another embodiment, the composition comprises between 50 and 750 pg of aluminium salt.

[0067] In one embodiment, the composition comprises between 10 and 950 pg, between 10 and 900 pg, between 20 and 850 pg, between 10 and 800 pg, between 10 and 750 pg, between 10 and 700 pg, between 10 and 650 pg, between 10 and 600 pg, between 10 and 550 pg, or between 10 and 500 pg of aluminium salt.

[0068] In another embodiment, the composition comprises between 10 and 1000 pg, between 20 and 1000 pg, between 30 and 1000 pg, between 40 and 1000 pg, or between 50 and 1000 pg. In another embodiment, the composition comprises between 100 and 1000 pg, between 200 and 1000 pg, between 300 and 1000 pg, between 400 and 1000 pg, between 500 and 1000 pg, between 600 and 1000 pg, between 700 and 1000 pg, between 800 and 1000 pg, or between 900 and 1000 pg of aluminium salt.

[0069] In another embodiment, the composition comprises between 50 and 750 pg, between 100 and 700 pg, between 150 and 650 pg, between 200 and 600 pg, or between 250 and 550 pg of aluminium salt.

[0070] In some embodiments, the composition comprises 500 pg of aluminium salt.

[0071] In one embodiment, the composition comprises CpG 7909 and aluminium hydroxide.

[0072] In one embodiment, the CpG oligonucleotide may be absorbed onto the aluminium salt, and thus used as a combination adjuvant that induces both Thl and Th2 responses. In one embodiment, the aluminium salt:CpG oligonucleotide weight / weight (w / w) ratio is between 1:0.1 and 1 : 1. In another embodiment, the aluminium salt:CpG oligonucleotide weight / weight (w / w) ratio is between 1 :0.2 and 1 : 1, between 1 :0.3 and 1 : 1, between 1 :0.4 and 1 : 1, between 1 :0.5 and 1 : 1, between 1 :0.6 and 1 : 1, between 1 :0.7 and 1 : 1, between 1 :0.8 and 1 : 1, or between 1:0.9 and 1: 1. Typically, the aluminium salt:CpG oligonucleotide weight / weight (w / w) ratio is between 1 :0.2 and 1 : 1.

[0073] The immunogenic composition may comprise a further adjuvant. In some embodiments, the adjuvant is a Thl adjuvant. Examples of Thl adjuvants promoting a Thl immune response include TLR-9 agonists, or TLR-4 agonists. Alternatively in another embodiment, the adjuvant is Incomplete Freund's Adjuvant (IFA).

[0074] Further examples of adjuvants may include a synthetic form of DNA, a carbohydrate, a tablet binder, an ion exchange resin, a preservative, a polymer, an emulsion and / or a lipid. Examples of adjuvants may include monosodium glutamate, sucrose, dextrose, aluminium bovine, human serum albumin, cytosine phosphoguanine, potassium phosphate, plasdone C, anhydrous lactose, cellulose, polacrilin potassium, glycerine, asparagine, citric acid, potassium phosphate magnesium sulfate, iron ammonium citrate, 2-phenoxyethanol, aluminium, beta-propiolactone, bovine extract, DOPC, EDTA, formaldehyde, thimerosal, phenol, potassium aluminium sulfate, potassium glutamate, sodium borate, sodium metabisulphite, urea, PLGA, PVA, PLA, PVP, cyclodextrin-based stabilisers, oil in water emulsion adjuvants and / or lipid-based adjuvants.

[0075] The term "immunogenic composition" as used throughout, refers to a composition of matter (intended to be administered to a subject) that comprises at least one antigen or induces the expression of at least one antigen of a rhinovirus (in the case of nucleic acid immunisation), which has the capability to elicit an immunological response in the subject to which it is administered. Such an immune response can be a cellular and / or antibody-mediated immune response directed at least against the antigen of the composition.

[0076] The immunogenic composition described herein provides an effective means of vaccinating a subject against RV infection. Accordingly, in one embodiment, the immunogenic composition is a vaccine. The immunogenic composition according to the first aspect is particularly suitable for therapy or prophylaxis (i.e. vaccination) against rhinovirus infections.

[0077] Hence, in a second aspect of the invention, there is provided the immunogenic composition according to the first aspect, for use in therapy or prophylaxis.

[0078] In a third aspect, there is provided the immunogenic composition according to the first aspect, for use in eliciting an immune response.

[0079] In a fourth aspect, there is provided a method of eliciting an immune response in a subject, the method comprising administering, to a subject in need thereof, a therapeutically effective amount of an immunogenic composition according to the first aspect.

[0080] It will be appreciated that the use and method of the invention comprises vaccination.

[0081] In some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against a rhinovirus infection.

[0082] In one embodiment, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against at least one, at least two, or at least three species of rhinoviruses, more particularly, against RV-A, RV-B, and / or RV-C.

[0083] Advantageously, the immune response that is induced by the immunogenic composition according to the first aspect, is a specific cell-mediated immune response not only directed to the homologous strain(s) of rhinovirus from which the immunogenic composition is derived but also to other (heterologous) strains of rhinoviruses of the same species of rhinoviruses, which can extend to strains of rhinoviruses of another group of rhinoviruses.

[0084] In some embodiments, therefore, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against at least one strain of RV-A, selected from a group consisting of: RV- 1A, RV-A1, RV-A10, RV-A100, RV-A101, RV-A103, RV-A106, RV-A107, RV-A108, RV- Al l, RV-A12, RV-A13, RV-A15, RV-A16, RV-A18, RV-A19, RV-A1B, RV-A2, RV-A20, RV-A21, RV-A22, RV-A23, RV-A24, RV-A25, RV-A28, RV-A29, RV-A30, RV-A31, RV- A32, RV-A33, RV-A34, RV-A36, RV-A38, RV-A39, RV-A40, RV-A41, RV-A43, RV-A44, RV-A45, RV-A46, RV-A47, RV-A49, RV-A50, RV-A51, RV-A53, RV-A54, RV-A55, RV- A56, RV-A57, RV-A58, RV-A59, RV-A60, RV-A61, RV-A62, RV-A63, RV-A64, RV-A65, RV-A66, RV-A67, RV-A68, RV-A7, RV-A71, RV-A73, RV-A74, RV-A75, RV-A76, RV- A77, RV-A78, RV-A8, RV-A80, RV-A81, RV-A82, RV-A85, RV-A88, RV-A89, RV-A9, RV- A90, RV-A94, RV-A95, RV-A96, and RV-A98. In some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against all strains of RV-A.

[0085] In some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against at least one strain of RV-B, selected from a group consisting of: RV-B103, RV- B102, RV-B101, RV-B100, RV-B27, RV-B26, RV-B4, RV-B97, RV-B35, RV-B84, RV- B93, RV-B92, RV-B91, RV-B17, RV-B5, RV-B42, RV-B6, RV-B37, RV-B48, RV-B69, RV- B52, RV-B72, RV-B3, RV-B14, RV-B99, RV-B86, RV-B83, RV-B79, and RV-B70. In some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against all strains of RV-B.

[0086] In some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against at least one strain of RV-C, selected from a group consisting of: RV-C58, RV- C50, RV-C48, RV-C46, RV-C13, RV-C51, RV-C55, RV-C56, RV-C45, RV-C44, RV-C41, RV-C4, RV-C38, RV-C34, RV-C33, RV-C31, RV-C30, RV-C29, RV-C27, RV-C26, RV- C24, RV-C23, RV-C21, RV-C16, RV-C14, RV-C47, RV-C53, RV-C54, RV-C22, RV-C37, RV-C49, RV-C25, RV-C18, RV-C12, RV-C20, RV-C19, RV-C28, RV-C36, RV-C39, RV- C15, RV-C5, RV-C11, RV-C9, RV-C10, RV-C1, RV-C43, RV-C42, RV-C40, RV-C32, RV- C17, RV-C8, RV-C7, RV-C6, RV-C3, RV-C2, and RV-C35. In some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against all strains of RV-C.

[0087] In one embodiment, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against RV-A16, RV-B27 and / or RV-C04.

[0088] Furthermore, it is well-known that rhinoviruses are the major cause of common colds, virus-induced wheezing illnesses in early childhood and acute attacks (exacerbations) of lung diseases such as asthma, bronchiectasis, chronic obstructive pulmonary disease (COPD), cystic fibrosis and chronic fibrosing lung diseases.

[0089] Accordingly, in some embodiments, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, elicits an immune response against common colds, virus-induced wheezing illnesses, acute attacks of lung diseases such as asthma, exacerbations of asthma, bronchiectasis, chronic obstructive pulmonary disease (COPD), cystic fibrosis and / or chronic fibrosing lung disease.

[0090] As used herein, the expression "eliciting an immune response" may involve inducing a specific cell-mediated immune response. This means the generation of a specific T lymphocyte response following the administration of an immunogenic composition in a subject. The two main cellular effectors of the specific T lymphocyte response are the helper T-cells and the cytotoxic T lymphocytes (CTLs).

[0091] CD4+ "helper" T-cells or helper T-cells, are immune response mediators, and play an important role in establishing and maximizing the capabilities of the adaptive immune response. These cells can have to some extent a direct cytotoxic activity, but, in essence "manage" the immune response, by directing other cells involved in the protection of organisms against pathogens. The activation of a naive helper T-cell causes it to release cytokines, which influences the activity of many cell types such as B lymphocytes, CTLs, and APCs (Antigen Presenting Cells) that activated it. Helper T- cells require a much milder activation stimulus than cytotoxic T-cells. Helper T-cells can provide extra signals that "help" activate cytotoxic cells. Two types of effector CD4+ helper T cell responses can be induced by a professional APC, designated Thl and Th2. The measure of cytokines associated with Thl or Th2 responses will give a measure of successful immunisation. This can be achieved by specific ELISA or ELISPOT designed for measurement of Thl-cytokines such as IFN-y, IL-2, and others, or Th2- cytokines such as IL-4, IL-5, IL-13 among others.

[0092] As used herein, the expression "helper T-cell-mediated immune response" refers to an immune response wherein CD4+T-cells or helper T-cells are activated and secrete lymphokines to stimulate both cell-mediated and antibody-mediated branches of the immune system. As known from the skilled person, helper T-cell activation promotes lymphokine secretion, immunoglobulin isotype switching, affinity maturation of the antibody response, macrophage activation and / or enhanced activity of natural killer and cytotoxic T-cells. Lymphokines are proteins secreted by lymphocytes that affect their own activity and / or the activity of other cells. Lymphokines include, but are not limited to, interleukins and cytokines, e.g., IL-2, IL-4, IL-5, IL-6, IL-10, IL-12, or IFN- -

[0093] It is well documented that helper T-cells differentiate into two major subtypes of cells known as Thl and Th2 cells (also known as Type 1 and Type 2 helper T cells, respectively).

[0094] Additionally, it is well known to the skilled person that Thl cells mainly secrete IL-2 and IFN-y. They promote cellular immune response by maximizing the killing efficacy of macrophages and the proliferation of cytotoxic CD8+T-cells. Additionally, the type 1 cytokine IFN-y increases the production of IL-12 by dendritic cells and macrophages, and, via positive feedback, IL-12 stimulates the production of IFN-y in helper T-cells, thereby promoting the Thl profile. IFN-y also inhibits the production of cytokines such as IL-4, an important cytokine associated with the Type 2 response, and thus it also acts to preserve its own response.

[0095] On the contrary, Th2 cells mainly secrete IL-4, IL-5 and IL-13, and promote humoral immune response by stimulating B cells into proliferation, inducing B-cell antibody class switching. The Type 2 response further promotes its own profile using two different cytokines. IL-4 acts on helper T-cells to promote the production of Th2 cytokines (including itself), while IL-10 inhibits a variety of cytokines including IL-2 and IFN-y in helper T-cells and IL-12 in dendritic cells and macrophages.

[0096] In one embodiment, the cell-mediated immune response induced by the immunogenic composition of the invention is primarily a Thl cell-mediated immune response. In one embodiment, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, induces an immune response through the secretion of IFN-y.

[0097] A critical component of protective cellular immunity elicited by natural infection with respiratory viruses is the formation of tissue resident memory T cells (TRM) in the airway mucosa and lungs. These TRM cells rapidly expand and clear subsequent infections. Accordingly, in another embodiment, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, induces an immune response through the formation of tissue resident memory T cells (TRM). In another embodiment, the immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, induces an immune response through the production of RV VPO-specific IgG antibodies.

[0098] Alternatively, in another embodiment, immunogenic composition for use according to the third aspect, or the method according to the fourth aspect, induces a cytotoxic T cell immune response.

[0099] Cytotoxic T cells (also known as Tc, killer T cell, or cytotoxic T-lymphocyte (CTL)), which express generally the CD8 marker, are a sub-group of T cells and may also be involved in the T cell-mediated immune response. They induce the death of cells that are infected with viruses (and other pathogens). These CTLs directly attack other cells carrying certain foreign or abnormal molecules on their surface. The ability of such cellular cytotoxicity can be detected using in vitro cytolytic assays (chromium release assay). Thus, induction of a specific cellular immunity can be demonstrated by the presence of such cytotoxic T cells, when antigen-loaded target cells are lysed by specific CTLs that are generated in vivo following vaccination or infection.

[0100] Similarly to helper T-cells, CD8+T-cells include distinct subsets, which were termed, analogously to the Thl / Th2 terminology, Tel and Tc2.

[0101] The Tel immune response involves specific IFN-y-producing CD8+T-cells which are activated, proliferate and produce IFN-y upon specific antigen stimulation. The level of IFN-y-producing CD8+T-cells can be measured by ELISPOT and by flow cytometry measurement of intracellular IFN-y in these cells.

[0102] Naive cytotoxic T cells are activated when their T-cell receptor (TCR) strongly interacts with a peptide-bound MHO class I molecule. This affinity depends on the type and orientation of the antigen / MHC complex, and is what keeps the CTL and infected cell bound together. Once activated the CTL undergoes a process called clonal expansion in which it gains functionality, and divides rapidly, to produce an army of "armed" effector cells. Activated CTL will then travel throughout the body in search of cells bearing that unique MHC Class I + peptide. This could be used to identify such CTLs in vitro by using peptide-MHC Class I tetramers in flow cytometric assays.

[0103] When exposed to these infected cells, effector CTL release perforin and granulysin, cytotoxins which form pores in the target cell's plasma membrane, allowing ions and water to flow into the infected cell, and causing it to burst or lyse. CTL release granzyme, a serine protease that enters cells via pores to induce apoptosis (cell death). Release of these molecules from CTL can be used as a measure of successful induction of cellular immune response following vaccination. This can be done by enzyme linked immunosorbant assay (ELISA) or enzyme linked immunospot assay (ELISPOT) where CTLs can be quantitatively measured. Since CTLs are also capable of producing important cytokines such as IFN-y, quantitative measurement of IFN-y- producing CD8 cells can be achieved by ELISPOT and by flow cytometric measurement of intracellular IFN-y in these cells.

[0104] In another aspect, there is provided a method of preparing the immunogenic composition according to the first aspect, the method comprising combining : (i) a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof, or a polynucleotide encoding the peptide, variant or fragment thereof; (ii) a CpG oligonucleotide; and (iii) an aluminium salt.

[0105] It will be appreciated that the immunogenic composition according to the invention, may be used in a medicament, which may be used as a monotherapy (i.e. use of the immunogenic composition alone), for vaccination against an RV infection.

[0106] Alternatively, the immunogenic composition according to the invention may be used as an adjunct to, or in combination with, known therapies for treating, ameliorating, or preventing an RV infection.

[0107] The immunogenic composition of the invention may be combined in compositions having a number of different forms depending, in particular, on the manner in which the composition is to be used. Thus, for example, the composition may be in the form of a powder, tablet, capsule, liquid, ointment, cream, gel, hydrogel, aerosol, spray, micellar solution, transdermal patch, liposome suspension, polyplex, emulsion, lipid nanoparticles (e.g. with peptide, DNA or RNA on the surface or encapsulated) or any other suitable form that may be administered to a person or animal in need of vaccination. The lipid nanoparticle may comprise one or more components selected from a group consisting of: a cationic lipid (which may be ionisable); phosphatidylcholine; cholesterol; and polyethylene glycol (PEG)-lipid. It will be appreciated that the vehicle of medicaments according to the invention should be one which is well-tolerated by the subject to whom it is given.

[0108] Medicaments comprising the immunogenic composition of the invention may be used in a number of ways. For instance, oral administration may be required, in which case the agents may be contained within a composition that may, for example, be ingested orally in the form of a tablet, capsule or liquid. Compositions comprising agents and medicaments of the invention may be administered by inhalation (e.g. intranasally). Compositions may also be formulated for topical use. For instance, creams or ointments may be applied to the skin.

[0109] The immunogenic composition of the invention may also be incorporated within a slow- or delayed-release device. Such devices may, for example, be inserted on or under the skin, and the medicament may be released over weeks or even months. The device may be located at least adjacent the treatment site. Such devices may be particularly advantageous when long-term treatment with the immunogenic composition is required and which would normally require frequent administration (e.g. at least daily injection).

[0110] In some embodiments, however, medicaments according to the invention may be administered to a subject by injection into the blood stream, muscle, skin or directly into a site requiring treatment. Injections may be intravenous (bolus or infusion) or subcutaneous (bolus or infusion), or intradermal (bolus or infusion), or intramuscular (bolus or infusion).

[0111] It will be appreciated that the amount of immunogenic composition that is required is determined by its biological activity and bioavailability, which in turn depends on the mode of administration, the physiochemical properties of the immunogenic composition and whether it is being used as a monotherapy or in a combined therapy. The frequency of administration will also be influenced by the half-life of the active agent within the subject being treated. Optimal dosages to be administered may be determined by those skilled in the art, and will vary with the particular the immunogenic composition in use, the strength of the composition, the mode of administration, and the type and advancement of the viral infection. Additional factors depending on the particular subject being treated will result in a need to adjust dosages, including subject age, weight, gender, diet, and time of administration.

[0112] Generally, a daily dose of between O.OOlpg / kg of body weight and lOOmg / kg of body weight of the immunogenic composition of the invention may be used for the immunisation, depending upon the agent used. In one embodiment, the daily dose of agent is between Ipg / kg of body weight and lOOmg / kg of body weight, or between lOpg / kg and lOmg / kg body weight, or between approximately lOOpg / kg and lOmg / kg body weight. Daily doses may be given as a single administration (e.g. a single daily injection or inhalation of a nasal spray). Alternatively, the immunogenic composition may require administration twice or more times during a day. As an example, the immunogenic composition may be administered as an initial primer and a subsequent boost(s), or two boosts administered at between a week or monthly intervals. In one embodiment, the immunogenic composition may be administered as an initial primer and a subsequent boost, administered between two to six weeks apart. In some embodiments, subsequent boosts may then be administered yearly to susceptible patients, such as those with weakened immune systems. Known procedures, such as those conventionally employed by the pharmaceutical industry (e.g. in vivo experimentation, clinical trials, etc.), may be used to form specific formulations of the immunogenic composition according to the invention and precise therapeutic regimes (such as daily doses of the agents and the frequency of administration).

[0113] A "subject" may be a vertebrate, mammal, or domestic animal. Hence, compositions and medicaments according to the invention may be used to treat any mammal, for example livestock (e.g. a horse), pets, or may be used in other veterinary applications. In one embodiment, the subject is a human being.

[0114] A "therapeutically effective amount" of the immunogenic composition is any amount which, when administered to a subject, is the amount of the aforementioned that is needed to ameliorate, prevent or treat any given disease, optionally prophylactically.

[0115] For example, the immunogenic composition of the invention may be used from about 0.001 pg to about 1 mg, and optionally from about 0.001 pg to about 500 pg. In one embodiment, the amount of the immunogenic composition is an amount from about 0.01 pg to about 250 pg, or from about 0.1 pg to about 100 pg. In one embodiment, the immunogenic composition according to the invention is administered at a dose of l-50pg.

[0116] The immunogenic compositions of the invention may further comprise a pharmaceutically acceptable vehicle. A "pharmaceutically acceptable vehicle" as referred to herein, is any known compound or combination of known compounds that are known to those skilled in the art to be useful in formulating pharmaceutical compositions.

[0117] In one embodiment, the pharmaceutically acceptable vehicle may be a solid, and the composition may be in the form of a powder or tablet. A solid pharmaceutically acceptable vehicle may include one or more substances which may also act as flavouring agents, lubricants, solubilisers, suspending agents, dyes, fillers, glidants, compression aids, inert binders, sweeteners, preservatives, dyes, coatings, or tabletdisintegrating agents. The vehicle may also be an encapsulating material. In powders, the vehicle is a finely divided solid that is in admixture with the finely divided active agents according to the invention. In tablets, the active agent (e.g. immunogenic composition according to the invention) may be mixed with a vehicle having the necessary compression properties in suitable proportions and compacted in the shape and size desired. The powders and tablets may contain up to 99% of the active agents. Suitable solid vehicles include, for example calcium phosphate, magnesium stearate, talc, sugars, lactose, dextrin, starch, gelatin, cellulose, polyvinylpyrrolidine, low melting waxes and ion exchange resins. In another embodiment, the pharmaceutical vehicle may be a gel and the composition may be in the form of a cream or the like.

[0118] However, the pharmaceutical vehicle may be a liquid, and the pharmaceutical composition is in the form of a solution. Liquid vehicles are used in preparing solutions, suspensions, emulsions, syrups, elixirs and pressurized compositions. The immunogenic composition according to the invention may be dissolved or suspended in a pharmaceutically acceptable liquid vehicle such as water, an organic solvent, a mixture of both or pharmaceutically acceptable oils or fats. The liquid vehicle can contain other suitable pharmaceutical additives such as solubilisers, emulsifiers, buffers, preservatives, sweeteners, flavouring agents, suspending agents, thickening agents, colours, viscosity regulators, stabilizers or osmo-regulators. Suitable examples of liquid vehicles for oral and parenteral administration include water (partially containing additives as above, e.g. cellulose derivatives, optionally sodium carboxymethyl cellulose solution), alcohols (including monohydric alcohols and polyhydric alcohols, e.g. glycols) and their derivatives, and oils (e.g. fractionated coconut oil and arachis oil). For parenteral administration, the vehicle can also be an oily ester such as ethyl oleate and isopropyl myristate. Sterile liquid vehicles are useful in sterile liquid form compositions for parenteral administration. The liquid vehicle for pressurized compositions can be a halogenated hydrocarbon or other pharmaceutically acceptable propellant.

[0119] Liquid pharmaceutical compositions, which are sterile solutions or suspensions, can be utilized by, for example, subcutaneous, intradermal, intrathecal, epidural, intraperitoneal, intravenous and particularly intramuscular injection. The nucleic acid sequence, or expression cassette of the invention may be prepared as a sterile solid composition that may be dissolved or suspended at the time of administration using sterile water, saline, or other appropriate sterile injectable medium.

[0120] The immunogenic composition of the invention may be administered orally in the form of a sterile solution or suspension containing other solutes or suspending agents (for example, enough saline or glucose to make the solution isotonic), bile salts, acacia, gelatin, sorbitan monoleate, polysorbate 80 (oleate esters of sorbitol and its anhydrides copolymerized with ethylene oxide) and the like. The immunogenic composition according to the invention can also be administered orally either in liquid or solid composition form. Compositions suitable for oral administration include solid forms, such as pills, capsules, granules, tablets, and powders, and liquid forms, such as solutions, syrups, elixirs, and suspensions. Forms useful for parenteral administration include sterile solutions, emulsions, and suspensions.

[0121] It will be appreciated that the invention extends to any nucleic acid or peptide or variant, derivative or analogue thereof, which comprises substantially the amino acid or nucleic acid sequences of any of the sequences referred to herein, including variants or fragments thereof. The terms "substantially the amino acid / nucleotide / peptide sequence", "variant" and "fragment", can be a sequence that has at least 40% sequence identity with the amino acid / nucleotide / peptide sequences of any one of the sequences referred to herein, for example 40% identity with the sequence identified as SEQ ID Nos: 1-4 and so on.

[0122] Amino acid / polynucleotide / polypeptide sequences with a sequence identity which is greater than 65%, or greater than 70%, or greater than 75%, or greater than 80% sequence identity to any of the sequences referred to are also envisaged. In some embodiments, the amino acid / polynucleotide / polypeptide sequence has at least 85% identity with any of the sequences referred to, or at least 90% identity, or at least 92% identity, or at least 95% identity, or at least 97% identity, or at least 98% identity, or at least 99% identity with any of the sequences referred to herein.

[0123] The skilled technician will appreciate how to calculate the percentage identity between two amino acid / polynucleotide / polypeptide sequences. In order to calculate the percentage identity between two amino acid / polynucleotide / polypeptide sequences, an alignment of the two sequences must first be prepared, followed by calculation of the sequence identity value. The percentage identity for two sequences may take different values depending on :- (i) the method used to align the sequences, for example, ClustalW, BLAST, FASTA, Smith-Waterman (implemented in different programs), or structural alignment from 3D comparison; and (II) the parameters used by the alignment method, for example, local vs global alignment, the pair-score matrix used (e.g. BLOSUM62, PAM250, Gonnet etc.), and gap-penalty, e.g. functional form and constants.

[0124] Having made the alignment, there are many different ways of calculating percentage identity between the two sequences. For example, one may divide the number of identities by: (i) the length of shortest sequence; (ii) the length of alignment; (iii) the mean length of sequence; (iv) the number of non-gap positions; or (v) the number of equivalenced positions excluding overhangs. Furthermore, it will be appreciated that percentage identity is also strongly length dependent. Therefore, the shorter a pair of sequences is, the higher the sequence identity one may expect to occur by chance.

[0125] Hence, it will be appreciated that the accurate alignment of protein or DNA sequences is a complex process. The popular multiple alignment program ClustalW (Thompson et al., 1994, Nucleic Acids Research, 22, 4673-4680; Thompson et al., 1997, Nucleic Acids Research, 24, 4876-4882) is one way for generating multiple alignments of proteins or DNA in accordance with the invention. Suitable parameters for ClustalW may be as follows: For DNA alignments: Gap Open Penalty = 15.0, Gap Extension Penalty = 6.66, and Matrix = Identity. For protein alignments: Gap Open Penalty = 10.0, Gap Extension Penalty = 0.2, and Matrix = Gonnet. For DNA and Protein alignments: ENDGAP = -1, and GAPDIST = 4. Those skilled in the art will be aware that it may be necessary to vary these and other parameters for optimal sequence alignment.

[0126] In some embodiments, calculation of percentage identities between two amino acid / polynucleotide / polypeptide sequences may then be calculated from such an alignment as (N / T)*100, where N is the number of positions at which the sequences share an identical residue, and T is the total number of positions compared including gaps and either including or excluding overhangs. In some embodiments, overhangs are included in the calculation. Hence, one method for calculating percentage identity between two sequences comprises (i) preparing a sequence alignment using the ClustalW program using a suitable set of parameters, for example, as set out above; and (ii) inserting the values of N and T into the following formula :- Sequence Identity = (N / T)*100.

[0127] Alternative methods for identifying similar sequences will be known to those skilled in the art. For example, a substantially similar nucleotide sequence will be encoded by a sequence which hybridizes to DNA sequences or their complements under stringent conditions. By stringent conditions, the inventors mean the nucleotide hybridises to filter-bound DNA or RNA in 3x sodium chloride / sodium citrate (SSC) at approximately 45°C followed by at least one wash in 0.2x SSC / 0.1% SDS at approximately 20-65°C. Alternatively, a substantially similar polypeptide may differ by at least 1, but less than 5, 10, 20, 50 or 100 amino acids from the sequences shown in, for example, in those of SEQ ID Nos: 1 to 4 that are amino acid sequences.

[0128] Due to the degeneracy of the genetic code, it is clear that any nucleic acid sequence described herein could be varied or changed without substantially affecting the sequence of the protein encoded thereby, to provide a functional variant thereof. Suitable nucleotide variants are those having a sequence altered by the substitution of different codons that encode the same amino acid within the sequence, thus producing a silent (synonymous) change. Other suitable variants are those having homologous nucleotide sequences but comprising all, or portions of, sequence, which are altered by the substitution of different codons that encode an amino acid with a side chain of similar biophysical properties to the amino acid it substitutes, to produce a conservative change. For example, small non-polar, hydrophobic amino acids include glycine, alanine, leucine, isoleucine, valine, proline, and methionine. Large non-polar, hydrophobic amino acids include phenylalanine, tryptophan and tyrosine. The polar neutral amino acids include serine, threonine, cysteine, asparagine and glutamine. The positively charged (basic) amino acids include lysine, arginine and histidine. The negatively charged (acidic) amino acids include aspartic acid and glutamic acid. It will therefore be appreciated which amino acids may be replaced with an amino acid having similar biophysical properties, and the skilled technician will know the nucleotide sequences encoding these amino acids.

[0129] All of the features described herein (including any accompanying claims, abstract and drawings), and / or all of the steps of any method or process so disclosed, may be combined with any of the above aspects in any combination, except combinations where at least some of such features and / or steps are mutually exclusive.

[0130] For a better understanding of the invention, and to show how embodiments of the same may be carried into effect, reference will now be made, by way of example, to the accompanying Figures, in which :-

[0131] Figure 1 shows the geometric mean titers of vaccine-specific IgG in the plasma of rhesus macaques over time following immunization with RV-C04 VP0 alone or RV-C04 adjuvanted with alum and CpG 7909. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day -5, 0, 14, 28, 35, 42 and 49 and plasma prepared. The amount of vaccine-specific IgG at each timepoint was determined by end point titer ELISA assay.

[0132] Figure 2 shows PBMC IFN-y ELISpot responses over time after immunization of rhesus macaques with RV-C04 VPO alone or RV-C04 adjuvanted with alum and CpG 7909, against VPO from RV-A16. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day 0, 14, 35 and 49 and PBMCs prepared. PBMCs were stimulated with a peptide pool corresponding to full length RV-A16 VPO and IFN-y secretion measure by ELISpot assay. Data presented as ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control. ***p<0.001 ANOVA.

[0133] Figure 3 shows PBMC IFN-y ELISpot responses over time after immunization of rhesus macaques with RV-C04 VPO alone or RV-C04 adjuvanted with alum and CpG 7909 against VPO from RV-B27. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day 0, 14, 35 and 49 and PBMCs prepared. PBMCs were stimulated with a peptide pool corresponding to full length RV-B27 VPO and IFN-y secretion measure by ELISpot assay. Data presented as ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control. *p<0.05, **p<0.01 ANOVA.

[0134] Figure 4 shows PBMC IFN-y ELISpot responses over time after immunization of rhesus macaques with RV-C04 VPO alone or RV-C04 adjuvanted with alum and CpG 7909 against VPO from RV-C04. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day 0, 14, 35 and 49 and PBMCs prepared. PBMCs were stimulated with a peptide pool corresponding to full length RV-C04 VPO and IFN-y secretion measure by ELISpot assay. Data presented as ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control. ****p<0.0001 ANOVA.

[0135] Figure 5 shows PBMC IL-4 ELISpot responses over time after immunization of rhesus macaques with RV-C04 VPO alone or RV-C04 adjuvanted with alum and CpG 7909 against VPO from RV-A16. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day 0, 14, 35 and 49 and PBMCs prepared. PBMCs were stimulated with a peptide pool corresponding to full length RV-A16 VPO and IL-4 secretion measure by ELISpot assay. Data presented as ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control. ****p<0.0001 ANOVA.

[0136] Figure 6 shows PBMC IL-4 ELISpot responses over time after immunization of rhesus macaques with RV-C04 VPO alone or RV-C04 adjuvanted with alum and CpG 7909 against VPO from RV-B27. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day 0, 14, 35 and 49 and PBMCs prepared. PBMCs were stimulated with a peptide pool corresponding to full length RV-B27 VPO and IL-4 secretion measure by ELISpot assay. Data presented as ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control. ****p<0.0001 ANOVA.

[0137] Figure 7 shows PBMC IL-4 ELISpot responses over time after immunization of rhesus macaques with RV-C04 VPO alone or RV-C04 adjuvanted with alum and CpG 7909 against VPO from RV-C04. Rhesus macaques were immunized on Day 0 and Day 28. Blood was drawn on Day 0, 14, 35 and 49 and PBMCs prepared. PBMCs were stimulated with a peptide pool corresponding to full length RV-C04 VPO and IL-4 secretion measure by ELISpot assay. Data presented as ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control. ****p<0.0001 ANOVA.

[0138] Figure 8 shows geometric mean titres of RV-C04 VPO-binding IgGl and IgG2c in mice immunized with different doses of RV-C04 VPO adjuvanted with alum and CpG 7909. Mice were immunized twice on Day 0 and Day 21 and serum prepared from blood taken on Day 35. RV-C04 VPO-binding IgGl or IgG2c titres were determined by cut point ELISA. Data presented as geometric mean titres. ****p<0.0001 ANOVA.

[0139] Figure 9 shows geometric mean titres of RV-C04 VPO-binding IgGl and IgG2c in mice immunized with different doses of RV-C04 VPO adjuvanted with alum. Mice were immunized twice on Day 0 and Day 21 and serum prepared from blood taken on Day 35. RV-C04 VPO-binding IgGl or IgG2c titres were determined by cut point ELISA. Data presented as geometric mean titres. ***p<0.001, *p<0.05 ANOVA.

[0140] Figure 10 shows geometric mean titres of RV-C04 VPO-binding IgGl and IgG2c in mice immunized with different doses of RV-C04 VPO alone. Mice were immunized twice on Day 0 and Day 21 and serum prepared from blood taken on Day 35. RV-C04 VPO- binding IgGl or IgG2c titres were determined by cut point ELISA. Data presented as geometric mean titres. **p<0.01 ANOVA. Figure 11 shows spleen IFN-y ELISpot responses after immunization of mice with RV- C04 adjuvanted with alum and CpG 7909. Mice were immunized on Day 0 and Day 21. Spleens were harvested on 35 and splenocytes prepared. Splenocytes were stimulated with a peptide pool corresponding to full length RV-C04 VPO or an irrelevant peptide control pool and IFN-y secretion measured by ELISpot assay. Data presented as mean spot forming units (SFU). ****p<0.0001, **p<0.001, *p<0.0001 ANOVA.

[0141] Figure 12 shows spleen IFN-y ELISpot responses after immunization of mice with RV- C04 adjuvanted with alum. Mice were immunized on Day 0 and Day 21. Spleens were harvested on 35 and splenocytes prepared. Splenocytes were stimulated with a peptide pool corresponding to full length RV-C04 VPO or an irrelevant peptide control pool and IFN-y secretion measured by ELISpot assay. Data presented as mean spot forming units (SFU). *p<0.0001 ANOVA.

[0142] Figure 13 shows spleen IFN-y ELISpot responses after immunization of mice with RV- C04 VPO alone. Mice were immunized on Day 0 and Day 21. Spleens were harvested on 35 and splenocytes prepared. Splenocytes were stimulated with a peptide pool corresponding to full length RV-C04 VPO or an irrelevant peptide control pool and IFN- y secretion measured by ELISpot assay. Data presented as mean spot forming units (SFU).

[0143] Figure 14 shows spleen IL-5 ELISpot responses after immunization of mice with RV- C04 adjuvanted with alum and CpG 7909. Mice were immunized on Day 0 and Day 21. Spleens were harvested on 35 and splenocytes prepared. Splenocytes were stimulated with a peptide pool corresponding to full length RV-C04 VPO or an irrelevant peptide control pool and IFN-y secretion measured by ELISpot assay. Data presented as mean spot forming units (SFU).

[0144] Figure 15 shows spleen IL-5 ELISpot responses after immunization of mice with RV- C04 adjuvanted with alum. Mice were immunized on Day 0 and Day 21. Spleens were harvested on 35 and splenocytes prepared. Splenocytes were stimulated with a peptide pool corresponding to full length RV-C04 VPO or an irrelevant peptide control pool and IFN-y secretion measured by ELISpot assay. Data presented as mean spot forming units (SFU). ****p<0.0001 ANOVA.

[0145] Figure 16 shows spleen IL-5 ELISpot responses after immunization of mice with RV- C04 alone. Mice were immunized on Day 0 and Day 21. Spleens were harvested on 35 and splenocytes prepared. Splenocytes were stimulated with a peptide pool corresponding to full length RV-C04 VPO or an irrelevant peptide control pool and IFN- y secretion measured by ELISpot assay. Data presented as mean spot forming units (SFU). ****p<0.0001 ANOVA.

[0146] Examples

[0147] The inventors previous work identified RV VPO proteins from single representative strains that, when used as vaccine immunogens, surprisingly evoke cellular immunity against all the other members of that RV species. In particular, the inventors discovered that combinations of VPO peptides used together in a single vaccine are particularly effective for immunising humans against infections with all RV strains.

[0148] As such, the inventors set out to continue this work and identify one single RV VPO protein, which is particularly effective at eliciting a broad cellular immune response.

[0149] Materials and Methods

[0150] Vaccine antigen production

[0151] Recombinant RV-C04 VPO was produced in E. Coli using standard methods that have been previously described (Glanville et al. 2013).

[0152] Rhesus macaque vaccine studies

[0153] All studies were performed under local laws and regulations and were reviewed and approved by the Institutional Animal Care and Use Committee (IACUC).

[0154] Vaccine preparation

[0155] Recombinant RV-C04 VPO was combined with alum (Alhydrogel 2%, Invivogen) and CpG 7909 (Invivogen) in Tris-buffered saline (TBS) or TBS alone to a final concentration of 200 pg / mL.

[0156] Rhesus macaque immunisation

[0157] Six animals (3 per group) were immunized with 100 pg of RV-C04 delivered with either alum / CpG or in TBS alone. Animals were immunized on Day 0 and Day 28.

[0158] Blood sampling and preparation of plasma and peripheral blood mononuclear cells Blood was collected on Days -5, 0, 14, 28, 35, 42 and 49 of the study from the saphenous or cephalic veins. The blood was incubated at room temperature for 30 minutes before being centrifuged and plasma and peripheral blood mononuclear cells (PBMCs) prepared by standard methods.

[0159] Plasma ELISA

[0160] 96-well plates were coated with 25 pg / well recombinant RV-C04 VPO protein diluted in PBS for detection of vaccine-specific IgG in monkey plasma samples. Plates were incubated overnight at 4°C. Plates were washed three times with PBS-Tween (PBS containing 0.05% Tween 20 (Sigma-Aldrich)). Non-specific binding was blocked by incubation with PBS 5% milk (Sigma-Aldrich) for 2 hours at room temperature. Plates were washed three times with PBS-Tween. 100 pL of serially diluted plasma sample were added per well, diluted in PBS-1% BSA and incubated for 2 hours at room temperature. Plates were washed three times with PBS-Tween. Bound antibody was detected with Peroxidase AffiniPure Goat Anti-Human IgG, Fey fragment specific (Jackson Immunoresearch, U.S.) diluted 1 :5000 in PBS 1% BSA (Sigma-Aldrich), incubated for 1 hour at room temperature. Plates were washed three times with PBS- Tween, then developed using 100 pL TMB substrate per well incubated for approximately 10 minutes, followed by 100 pL 0.18M H2SO4 per well to stop the reaction. Plates were read immediately at 450nm using a SpectraMax plate reader (Molecular Devices) and vaccine-specific IgG quantified by end point titer analysis.

[0161] PBMC ELISpot

[0162] 3xl05PBMCs were stimulated with peptide pools corresponding to full-length VPO from RV-A16, B27 or C04 (80 peptides per pool, 15 mer with 11 amino acid overlap) or media control. Each stimulation was performed in triplicate. IFN-y or IL-4 ELISpot was performed using Monkey IFN-y ELISpot PRO kit (ALP), strips (Mabtech-3421M-2AST- 10) and Human IL-4 ELISpot PRO kit (ALP) (Mabtech-3410-2APW-10), according to the manufacturer's instructions. The number of spot forming units (SFU) per 3xl05PBMCs was determined using an AID ELISpot machine (Model ELR088IFL) and data reported the ratio of geometric spot forming unit (SFU) means after stimulation with the peptide pool vs media only control.

[0163] Mouse vaccine studies

[0164] Mouse immunisation

[0165] Groups of 6 mice were immunized intramuscularly with 10, 5, 1, 0.1 or 0 pg RV-C04 VPO in a 50 pL volume. The doses of RV-C04 VPO were delivered alone, with 50 pg alum, or with alum and 50 pg CpG 7909. Animals were immunized on Day 0 and Day 21 and the study terminated on Day 35. Serum ELISA

[0166] 96-well ELISA microplates coated with 50 pL per well of 0.5 pg / mL APL-10456 at 4°C overnight. Plates were washed with, then blocked for 1 hour at room temperature with 100 pL / well PBS+2% BSA. Plates were washed with 200 pL / well PBS+0.05% Tween 20, then serum samples were added at a maximum concentration of 1 / 20 dilution in PBS 2% BSA, followed by a 1 / 3 dilution series of each sample to create an 8-point curve. All samples were run in duplicate. Serum samples were incubated on the plates for 2 hours at room temperature. Plates were washed with 200 pL / well PBS+0.05% Tween 20, then incubated for 20 minutes at room temperature in the dark with 50 uL of 1 pg / mL goat anti-mouse IgGl-HRP or IgG2c-HRP in PBS+ 1% BSA per well. Plates were washed with 200 pL / well PBS+0.05% Tween 20, then developed with 50 uL TMB substrate per well, followed by 50 uL 0.18M H2S04 and read at 450 nm using a SpectraMax ID3 analyser.

[0167] Spleen ELISPOT

[0168] Spleens were collected and placed in 2 mL RPMI 10 % FCS and 1 % penicillin / streptomycin and stored on ice until further processing. To isolate splenocytes, spleens were mashed through a 100 pm filter using the plunger from a sterile 2 mL syringe then washed with RPMI 10% FCS 1 % P / S. Red blood cells were lysed by incubating the cells in 2 mL ACK buffer for 2 minutes at room temperature. Spleen cells were then washed twice, once with PBS and once with RPMI 10% FBS 1% P / S, then placed on ice until used in ELISpot assays.

[0169] 96 well sterile plates were coated with 100 pL per well of 5 pg / mL purified anti-mouse IFN-y or 0.5 pg / mL purified anti-mouse IL-5 antibody overnight at 4°C. The plates were washed twice with 150 uL / well PBS and blocked for 5 hours at 37oC with 200uL / well of complete medium (RPMI supplemented with 10% fetal calf serum and 1% penicillin / streptomycin). The splenocytes were diluted in complete medium to a density of 1 x 107 cells / mL and 100 pL added to each well. The RV-C04 VP0 and control peptide pools were diluted in complete medium to 32 pg / mL and 100 pL added to each test well to give a final in well concentration of 16 pg / mL. The positive control used was PMA plus ionomycin and used at a final concentration of 50 ng / mL PMA and 500 ng / mL ionomycin. After addition of stimuli the plates were incubated at 37oC for 24 hours. After 24 hours, the cells and medium were tipped out, the plates washed twice with 200 pL / well PBS+0.05% Tween 20 and 150 pL / well distilled water added and incubated for 5 minutes to raise any remaining cells. After two washes with 200 pL / well PBS+0.05% Tween 20, 100 pL of detection antibody (2 pg / mL biotinylated anti-mouse IFN-y or 1 pg / mL biotinylated anti-mouse IL-5 antibody in PBS 1% w / v BSA (A7906, Sigma Aldrich) was added per well and the plate incubated for 90 minutes at room temperature. After three washes with 200 pL / well PBS-Tween 20, 100 pL / well of ExtrAvidin® Alkaline Phosphatase diluted in 1 :200 in PBS was added to each well and the plate incubated for 45 minutes at room temperature. The plate was washed twice with 200 pL / well PBS-Tween 20, twice with PBS and then lOOuL / well SIGMAFAST™ BCIP® / NBT substrate (B5655, Sigma-Aldrich), made up as per manufacturer's instructions, was added. The plate was incubated for 10 minutes in the dark at room temperature. The plates were washed three times with distilled water, dried and the number of spot forming units (SFU) per well counted the following day using an AID Classic EliSpot Reader (AID GmbH). The data was expressed as the number of SFU / 106 splenocytes.

[0170] Results

[0171] Adjuvants are reauired for recombinant RV-C04 immunogenicity

[0172] Figure 1 is a set of histograms that show the results from ELISA experiments to quantify the titer of vaccine-specific IgG in the plasma of rhesus macaques after immunization with recombinant RV-C04 VP0 delivered alone or adjuvanted with alum and CpG 7909. Animals immunized with recombinant RV-C04 VP0 alone produce much lower vaccine-specific IgG than animals immunized with RV-C04 VP0 plus alum and CpG 7909.

[0173] Cellular immunity evoked bv RV-C04 VP0 is adjuvant deoendent, cross-soecies reactive and Thl-polarized

[0174] Figures 2-7 are histograms that demonstrate the cellular immunity evoked by immunisation with recombinant RV-C04 VP0 against VP0 from heterotypic strains covering all three species of RV. Figures 2-4 show secretion of the canonical Thl cytokine IFN-y and Figures 5-7 show the secretion of the canonical Th2 cytokine IL-4 by PBMCs in response to stimulation with peptide pools corresponding to full-length VP0 from RV-A16, RV-B27 or RV-C04.

[0175] PBMCs from rhesus macaques immunized with RV-C04 VP0 plus alum / CpG 7909 produce significantly greater IFN-y when stimulated with all peptide pools (i.e. RV-A16 VP0, RV-B27 VP0 and RV-C04 VP0) compared to controls. However, PBMCs from animals immunised with RV-C04 alone do not produce significant amounts of IFN-y. In contrast, when IL-4 production was assessed, there was a significant difference between RV-C04 VPO peptide pool and control stimulated PBMCs from animals immunised with RV-C04 plus alum / CpG only. There was no significant increase in IL-4 production by PBMCs after stimulation with RV-A16 VPO or RV-B27 VPO peptide pools and there was no significant IL-4 production in response to stimulation with any peptide pool by PBMCs from animals immunised with RV-C04 VPO alone.

[0176] Thl- vs Th2 immune response

[0177] As shown by the IFN- (Thl) vs IL-5 (Th2) ELISpot (Figures 11-15) and the IgG2c (Thl) / IgGl (Th2) ratio (Figures 8-10), RV-C04 VPO alone, or with alum, elicits a Th2- biased immune response. However, when CpG 7909 is added, the immune response surprisingly switches from Th2 to Thl.

[0178] Conclusions

[0179] The inventors previous work identified RV VPO proteins from single representative strains that, when used as vaccine immunogens, surprisingly evoke cellular immunity against all the other members of that RV species. In particular, the inventors discovered that combinations of VPO peptides used together in a single vaccine are particularly effective for immunising humans against infections with all RV strains.

[0180] However, the inventors have now discovered that a single RV VPO protein, RV-C04 VPO, is surprisingly effective at eliciting a broad cellular immune response that crossreacts with other RV strains from the same species, when adjuvanted with aluminium salt and CpG 7909.

Claims

Claims1. An immunogenic composition comprising : (i) a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof, or a polynucleotide encoding the peptide, variant or fragment thereof; (ii) a CpG oligonucleotide; and (iii) an aluminium salt.

2. The immunogenic composition according to claim 1, wherein the RV-C04 VPO peptide comprises an amino acid sequence substantially as set out in SEQ ID No: 1, or a variant or fragment thereof.

3. The immunogenic composition according to either claim 1 or claim 2, wherein the RV-C04 VPO peptide is encoded by the nucleotide sequence substantially as set out in SEQ ID No: 2, or a variant or fragment thereof, or wherein the immunogenic composition comprises a polynucleotide comprising the nucleotide sequence substantially as set out in SEQ ID No: 2, or a variant or fragment thereof.

4. The immunogenic composition according to any preceding claim, wherein the immunogenic composition comprises a polynucleotide comprising the nucleotide sequence substantially as set out in SEQ ID No: 3, or a variant or fragment thereof.

5. The immunogenic composition according to any preceding claim, wherein the immunogenic composition comprises or encodes between 15 and 150 pg, between 20 and 125 pg, or between 25 and 100 pg of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

6. The immunogenic composition according to any preceding claim, wherein the immunogenic composition comprises or encodes: (i) between 5 and 550 pg / mL, between 10 and 525 pg / mL, between 20 and 500 pg / mL, between 30 and 475 pg / mL, between 40 and 450 pg / mL, or between 50 and 425 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof; or (ii) between 5 and 250 pg / mL, between 10 and 240 pg / mL, between 20 and 230 pg / mL, between 30 and 220 pg / mL, between 40 and 210 pg / mL, or between 50 and 200 pg / mL of a human rhinovirus (RV)-C04 VPO peptide, or a variant or fragment thereof.

7. The immunogenic composition according to any preceding claim, wherein the CpG oligonucleotide is a class A, class B or class C CpG oligonucleotide (ODN).

8. The immunogenic composition according to any preceding claim, wherein the CpG oligonucleotide is selected from the group consisting of: CpG 1018, ODN D-SL01, CpG 7909, ODN 2216, ODN 2336, ODN 2395 and ODN M362.

9. The immunogenic composition according to any preceding claim, wherein the CpG oligonucleotide is a Class B CpG oligonucleotide, optionally wherein the CpG oligonucleotide is selected from the group consisting of: CpG 7909, ODN D-SL01 and CpG 1018.

10. The immunogenic composition according to any preceding claim, wherein the CpG oligonucleotide is CpG 7909.

11. The immunogenic composition according to claim 10, wherein CpG 7909 comprises the nucleotide sequence substantially as set out in SEQ ID No: 4, or a variant or fragment thereof.

12. The immunogenic composition according to any preceding claim, wherein the composition comprises between 50 and 750 pg, between 75 and 725 pg, between 100 and 700 pg, between 125 and 675 pg, between 150 and 650 pg, between 175 and 625 pg, between 200 and 600 pg, between 225 and 575 pg, or between 250 and 550 pg of CpG oligonucleotide.

13. The immunogenic composition according to any preceding claim, wherein the aluminium salt is selected from the group consisting of: aluminium hydroxide, aluminium phosphate, aluminium hydroxyphosphate, aluminium orthophosphate, and amorphous aluminium hydroxyphosphate sulfate.

14. The immunogenic composition according to any preceding claim, wherein the aluminium salt is aluminium hydroxide, optionally wherein the aluminium hydroxide is aluminium oxyhydroxide.

15. The immunogenic composition according to any preceding claim, wherein the composition comprises between 10 and 1000 pg, between 20 and 900 pg, between 30 and 800 pg, between 40 and 700 pg, between 50 and 600 pg, or between 60 and 500 pg of aluminium salt.

16. The immunogenic composition according to any preceding claim, wherein the composition comprises between 50 and 750 pg, between 100 and 700 pg, between150 and 650 g, between 200 and 600 pg, or between 250 and 550 pg of aluminium salt.

17. The immunogenic composition according to any preceding claim, wherein the composition comprises CpG 7909 and aluminium hydroxide.

18. The immunogenic composition according to any preceding claim, wherein the aluminium salt:CpG oligonucleotide weight / weight (w / w) ratio is between 1 :0.2 and 1: 1.

19. The immunogenic composition according to any preceding claim, wherein the immunogenic composition is a vaccine.

20. The immunogenic composition according to any one of claims 1 to 19, for use in therapy or prophylaxis.

21. The immunogenic composition according to any one of claims 1 to 19, for use in eliciting an immune response, optionally against a rhinovirus infection.

22. The immunogenic composition for use according to claim 21, wherein the immunogenic composition elicits an immune response against at least one, at least two, or at least three species of rhinoviruses, more particularly, against RV-A, RV-B, and / or RV-C.

23. The immunogenic composition for use according to any one of claims 20 to 22, wherein the immunogenic composition elicits an immune response against common colds, virus-induced wheezing illnesses, asthma, exacerbations of asthma, bronchiectasis, chronic obstructive pulmonary disease (COPD), cystic fibrosis and / or chronic fibrosing lung disease.

24. A method of preparing the immunogenic composition according to any one of claims 1 to 19, the method comprising combining: (i) a human rhinovirus (RV)-C04 VP0 peptide, or a variant or fragment thereof, or a polynucleotide encoding the peptide, variant or fragment thereof; (ii) a CpG oligonucleotide; and (iii) an aluminium salt.

Images