Lyme disease vaccine containing adjuvant and BORRELIA cell surface protein A antigen

Abstract

This disclosure relates to immunogenic compositions comprising Borrelia cell surface protein A (OspA) antigen or RNA polynucleotide encoding the OspA antigen, and an adjuvant. In some embodiments, the immunogenic compositions are suitable for stimulating an immune response to Borrelia in a subject. This disclosure also relates to kits of immunogenic compositions, their use, and methods of using them.

Description

[Technical Field]

[0001] Cross-references to related applications This application claims priority from U.S. Provisional Application No. 63 / 512,872, filed on 10 July 2023, the contents of which are incorporated in their entirety by reference.

[0002] Submission of sequence listings as XML files This application is filed together with an electronic sequence listing. The contents of the electronic sequence listing (377882009240SEQLIST.xml; size: 86,006 bytes; and creation date: July 9, 2024) are incorporated herein by reference in their entirety.

[0003] field This disclosure relates to immunogenic compositions comprising Borrelia cell surface protein A (OspA) antigen or RNA polynucleotide encoding the OspA antigen, and an adjuvant. In some embodiments, the immunogenic compositions are suitable for stimulating an immune response to Borrelia in a subject. This disclosure also relates to manufactured articles comprising the immunogenic compositions, their use, and methods of using them. [Background technology]

[0004] background Lyme borreliosis, or Lyme disease, is the most commonly reported tick-borne disease in Europe and North America. The disease is caused by infection with spirochete bacteria of the genus Borrelia. A monovalent serotype 1-OspA-based vaccine (LYMErix®) was approved and marketed in the United States for the prevention of Lyme disease caused by B. burgdorferi sensu stricto(ss), but this vaccine is no longer available. Currently, there are no commercially available preventative medications for Lyme disease. Therefore, there is an unmet need for a safe and effective Lyme disease vaccine. [Overview of the project] [Means for solving the problem]

[0005] overview In some embodiments, immunogenic compositions are provided herein that comprise (i) a Borrelia cell surface protein A (OspA) antigen or an RNA polynucleotide encoding an OspA antigen, and (ii) an adjuvant selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsions comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0006] In some embodiments, immunogenic compositions are also provided herein that include (i) a Borrelia cell surface protein A (OspA) antigen, and (ii) an adjuvant selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsions comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0007] In some of the embodiments, the immunogenic composition does not contain RNA polynucleotides encoding non-OspA Borrelia cell surface protein antigens.

[0008] In some of the embodiments, the immunogenic composition does not contain non-OspA Borrelia cell surface protein antigens.

[0009] In some of the embodiments, the adjuvant is an aluminum salt adjuvant. In some of the embodiments, the immunogenic composition further comprises a second adjuvant. In some of the embodiments, the second adjuvant is selected from TLR9 agonists; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsions comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0010] In some of the embodiments, the adjuvant is a TLR9 agonist. In some of the embodiments, the immunogenic composition further comprises a second adjuvant. In some of the embodiments, the second adjuvant is selected from aluminum salt adjuvants; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0011] In some of the embodiments, the adjuvant is a liposome-based adjuvant comprising lipopolysaccharides and saponins. In some of the embodiments, the immunogenic composition further comprises a second adjuvant. In some of the embodiments, the second adjuvant is selected from aluminum salt adjuvants; TLR9 agonists; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0012] In some of the embodiments, the adjuvant is an oil-in-water emulsion-based adjuvant comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters. In some of the embodiments, the immunogenic composition further comprises a second adjuvant. In some of the embodiments, the second adjuvant is selected from aluminum salt adjuvants; TLR9 agonists; and liposome-based adjuvants comprising lipopolysaccharides and saponins.

[0013] In some of the embodiments, the TLR9 agonist is an oligonucleotide containing a non-methylated cytidine-phosphoguanosine (CpG) motif. In some of the embodiments, the oligonucleotide is 10 to 35 nucleotides long.

[0014] In some of the embodiments, the oligonucleotide comprises the sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2). In some of the embodiments, the oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1).

[0015] In some embodiments, immunogenic compositions comprising a 10-35 nucleotide length Toll-like receptor 9 (TLR9) agonist oligonucleotide, as well as a Borrelia cell surface protein A (OspA) antigen, and a Toll-like receptor 9 (TLR9) agonist oligonucleotide containing the sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2) and a non-methylated cytidine-phospho-guanosine (CpG) motif, are also provided herein.

[0016] In some of the embodiments, the oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1).

[0017] In some embodiments, immunogenic compositions comprising an oligonucleotide, such as Borrelia cell surface protein A (OspA) antigen and Toll-like receptor 9 (TLR9) agonist oligonucleotide 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1), wherein the oligonucleotide comprises a non-methylated cytidine-phospho-guanosine (CpG) motif, are also provided herein.

[0018] In some embodiments of any aspect, the immunogenic composition further comprises a second adjuvant. In some embodiments of any aspect, the second adjuvant is selected from aluminum salt adjuvants; liposome-based adjuvants comprising lipopolysaccharide and saponin; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0019] In some embodiments of any aspect, the second adjuvant is an aluminum salt adjuvant.

[0020] In some embodiments, there is also provided herein an immunogenic composition comprising (i) Borrelia outer surface protein A (OspA) antigen, (ii) the toll-like receptor 9 (TLR9) agonist oligonucleotide 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1), wherein the oligonucleotide comprises an unmethylated cytidine-phospho-guanosine (CpG) motif, and (iii) an aluminum salt adjuvant.

[0021] In some embodiments of any aspect, the aluminum salt adjuvant comprises one or more of amorphous aluminum hydroxyphosphate sulfate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate. In some embodiments of any aspect, the aluminum salt adjuvant comprises aluminum hydroxide.

[0022] In some embodiments of any aspect, the immunogenic composition comprises from about 0.25 to about 1.25 mg of Al 3+ , from about 0.25 mg to about 0.50 mg of Al 3+ , or from about 0.30 mg to about 0.40 mg of Al 3+ .

[0023] In some of the embodiments, the oligonucleotide is a single-stranded oligodeoxynucleotide. In some of the embodiments, the oligonucleotide is a complete RNA. In some of the embodiments, the oligonucleotide is an RNA / DNA chimera.

[0024] In some of the embodiments, the oligonucleotide comprises at least one phosphorothioate linkage. In some of the embodiments, the oligonucleotide comprises only a phosphorothioate linkage. In some of the embodiments, the oligonucleotide comprises a combination of one or more phosphodiester linkages and one or more phosphorothioate linkages.

[0025] In some of the embodiments, the immunogenic composition comprises about 375 μg to about 6000 μg of TLR9 agonist or about 750 μg to about 3000 μg of TLR9 agonist. In some of the embodiments, the immunogenic composition comprises about 375 μg, about 750 μg, about 1000 μg, about 1500 μg, about 3000 μg, or about 6000 μg of TLR9 agonist.

[0026] In some of the embodiments, the lipopolysaccharide is lipid A or a derivative thereof. In some of the embodiments, the lipopolysaccharide is monophosphoryl lipid A or a derivative thereof. In some of the embodiments, the lipopolysaccharide is 3-O-desacyl-4'-monophosphoryl lipid A. In some of the embodiments, the lipopolysaccharide is 3-deacyl-phosphorylated hexaacyl disaccharide.

[0027] In some of the embodiments, the immunogenic composition contains about 1 μg to about 100 μg, about 10 μg to about 50 μg, about 20 μg to about 30 μg, about 21 μg to about 29 μg, about 22 μg to about 28 μg, about 23 μg to about 27 μg, or about 24 μg to about 26 μg of lipopolysaccharide. In some of the embodiments, the immunogenic composition contains about 1 μg to about 30 μg, about 5 μg to about 15 μg, about 6 μg to about 14 μg, about 7 μg to about 13 μg, about 8 μg to about 12 μg, or about 9 μg to about 11 μg of lipopolysaccharide. In some of the embodiments, the immunogenic composition contains about 1 μg to about 9 μg, about 2 μg to about 8 μg, about 3 μg to about 7 μg, or about 4 μg to about 6 μg of lipopolysaccharide.

[0028] In some of the embodiments, the saponin is derived from the bark of Quillaja saponaria Molina. In some of the embodiments, the saponin is Quill A or a derivative thereof. In some of the embodiments, the saponin is QS-17. In some of the embodiments, the saponin is QS-21.

[0029] In some of the embodiments, the immunogenic composition contains about 1 μg to about 100 μg, about 10 μg to about 50 μg, about 20 μg to about 30 μg, about 21 μg to about 29 μg, about 22 μg to about 28 μg, about 23 μg to about 27 μg, or about 24 μg to about 26 μg of saponins. In some of the embodiments, the immunogenic composition contains about 1 μg to about 30 μg, about 5 μg to about 15 μg, about 6 μg to about 14 μg, about 7 μg to about 13 μg, about 8 μg to about 12 μg, or about 9 μg to about 11 μg of saponins. In some of the embodiments, the immunogenic composition contains about 1 μg to about 9 μg, about 2 μg to about 8 μg, about 3 μg to about 7 μg, or about 4 μg to about 6 μg of saponins.

[0030] In some of the embodiments, the liposome-based adjuvant further comprises a sterol. In some of the embodiments, the sterol is β-sitosterol, stigmasterol, ergosterol, ergocalciferol, or cholesterol. In some of the embodiments, the sterol is cholesterol.

[0031] In some of the embodiments, the ratio of saponins to sterols is 1:1 to 1:100 w / w or 1:1 to 1:5 w / w. In some of the embodiments, the ratio of saponins to sterols is approximately 1:1 w / w.

[0032] In some of the embodiments, the liposome-based adjuvant further comprises a neutral lipid. In some of the embodiments, the neutral lipid is phosphatidylcholine. In some of the embodiments, the phosphatidylcholine is egg yolk phosphatidylcholine, dioleoyl phosphatidylcholine (DOPC), or dilauryl phosphatidylcholine. In some of the embodiments, the phosphatidylcholine is DOPC.

[0033] In some of the embodiments, the sorbitan ester is sorbitan ester 20. In some of the embodiments, the sorbitan ester is sorbitan ester 85.

[0034] In some of the embodiments, the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 20. In some of the embodiments, the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 80.

[0035] In some of the embodiments, the immunogenic composition contains squalene in an amount of about 2% w / v to about 7% w / v, about 3% w / v to about 6% w / v, or about 4% w / v to about 5% w / v.

[0036] In some of the embodiments, the immunogenic composition comprises sorbitan esters in concentrations of about 0.2% w / v to about 0.8% w / v, about 0.3% w / v to about 0.7% w / v, or about 0.4% w / v to about 0.6% w / v.

[0037] In some of the embodiments, the immunogenic composition comprises polyoxyethylene sorbitan ester in concentrations of about 0.2% w / v to about 0.8% w / v, about 0.3% w / v to about 0.7% w / v, or about 0.4% w / v to about 0.6% w / v.

[0038] In some of the embodiments, the OspA antigen is two or more OspA antigens. In some of the embodiments, the two or more OspA antigens are a first OspA antigen and a second OspA antigen. In some of the embodiments, the two or more OspA antigens are a first OspA antigen, a second OspA antigen, and a third OspA antigen. In some of the embodiments, each of the two or more OspA antigens has a non-identical amino acid sequence.

[0039] In some parts of any embodiment, the OspA antigen (e.g., the first, second, and / or third OspA antigen) is independently the OspA antigen of B. burgdorferi, B. afzelii, B. bavariensis, B. garinii, B. mayonii, B. lusitaniae, B. bissettii, B. valasiana, or B. spielmanii. In some parts of any embodiment, the OspA antigen is the OspA antigen of B. burgdorferi.

[0040] In some parts of any embodiment, an OspA antigen (e.g., a first, second, and / or third OspA antigen) independently comprises one or more epitopes derived from an OspA protein. In some parts of any embodiment, one or more epitopes comprises at least two, three, four, or five epitopes derived across one or more OspA proteins. In some parts of any embodiment, one or more epitopes comprises at least two, three, four, or five epitopes derived across at least two, three, four, or five OspA serotypes.

[0041] In some of the embodiments, one or more epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 4-49. In some of the embodiments, one or more epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4-49.

[0042] In some of the embodiments, one or more epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49.

[0043] In any embodiment, an OspA antigen (e.g., a first, second, and / or third OspA antigen) independently has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any of SEQ ID NOs.

[0044] In any embodiment, the OspA antigen (e.g., the first, second, and / or third OspA antigen) independently comprises at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids of the amino acid sequence shown in any of SEQ ID NOs. 50-62.

[0045] In some part of any embodiment, the OspA antigen (e.g., the first, second, and / or third OspA antigen) independently comprises the amino acid sequence shown in any of SEQ ID NOs. 50-62.

[0046] In some parts of any embodiment, the OspA antigen (e.g., the first, second, and / or third OspA antigen) is independently an OspA chimeric antigen comprising a number of epitopes derived from one or more OspA proteins. In some parts of any embodiment, the number of epitopes comprises at least two, three, four, five, or six epitopes derived across at least two, three, four, five, or six OspA serotypes.

[0047] In some of the embodiments, the numerous epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 4-49. In some of the embodiments, the numerous epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4-49.

[0048] In some of the embodiments, the numerous epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49.

[0049] In any embodiment, an OspA antigen (e.g., a first, second, and / or third OspA antigen) independently comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 63-82.

[0050] In any part of the embodiment, an OspA antigen (e.g., a first, second, and / or third OspA antigen) independently comprises at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids of the amino acid sequence shown in any one of SEQ ID NOs. 63-82.

[0051] In some part of any embodiment, the OspA antigen (e.g., the first, second, and / or third OspA antigen) independently comprises the amino acid sequence shown in any one of SEQ ID NOs. 63-82.

[0052] In any embodiment, an OspA antigen (e.g., a first, second, and / or third OspA antigen) independently comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 79-81.

[0053] In any embodiment, the immunogenic composition comprises a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 79, and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80. In some of the embodiments, the immunogenic composition comprises a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79 and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80.

[0054] In any embodiment, the immunogenic composition comprises a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 89, and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some of the embodiments, the immunogenic composition comprises a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79 and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 81.

[0055] In some part of any embodiment, the immunogenic composition comprises a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some of the embodiments, the immunogenic composition comprises a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80 and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 81.

[0056] In some of the embodiments, the immunogenic composition comprises a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence represented by SEQ ID NO: 80, and at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88% sequence identity with the amino acid sequence represented by SEQ ID NO: 80. The immunogenic composition comprises a first OspA antigen comprising an amino acid sequence having 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity, and a third OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some parts of any embodiment, the immunogenic composition comprises a first OspA antigen comprising an amino acid sequence shown in SEQ ID NO: 79, a second OspA antigen comprising an amino acid sequence shown in SEQ ID NO: 80, and a third OspA antigen comprising an amino acid sequence shown in SEQ ID NO: 81.

[0057] In some of the embodiments, the OspA antigens (e.g., the first, second, and / or third OspA antigens) are independently not lipidized.

[0058] In some of the embodiments, the OspA antigens (e.g., the first, second, and / or third OspA antigens) are independently lipidized.

[0059] In some of the embodiments, the immunogenic composition is in a dose of 0.3 mL to 1.5 mL.

[0060] In some of the embodiments, the immunogenic composition is intended to stimulate an immune response to Borrelia in a subject by administering an effective amount of the immunogenic composition to the subject. In some of the embodiments, the immunogenic composition is intended to protect a subject from infection by Borrelia by administering an effective amount of the immunogenic composition to the subject. In some of the embodiments, the immunogenic composition is intended to prevent a subject from contracting Lyme disease by administering an effective amount of the immunogenic composition to the subject.

[0061] In some of the embodiments, the subject is a human subject.

[0062] In some of the embodiments, the human subjects are at least 18 years of age. In some of the embodiments, the human subjects are between 18 and 70 years of age.

[0063] In some of the embodiments, the human subjects are under 18 years of age. In some of the embodiments, the human subjects are between 2 and 5 years of age. In some of the embodiments, the human subjects are between 5 and 18 years of age.

[0064] In some embodiments, methods for stimulating an immune response to Borrelia in a subject are also provided herein, which include administering an effective amount of any of the provided immunogenic compositions to the subject.

[0065] In some embodiments, methods for protecting a subject from infection by Borrelia are also provided herein, which include administering an effective amount of one of the provided immunogenic compositions to the subject.

[0066] In some embodiments, methods for preventing a subject from contracting Lyme disease are also provided herein, which include administering an effective amount of any of the provided immunogenic compositions to the subject.

[0067] In some of the embodiments, the immunogenic composition is administered by intramuscular injection.

[0068] In some of the embodiments, a first dose and a second dose of the immunogenic composition are administered to the subject. In some of the embodiments, the second dose of the immunogenic composition is administered about 2 weeks to about 10 weeks after the administration of the first dose of the immunogenic composition. In some of the embodiments, the second dose of the immunogenic composition is administered about 1 month or about 2 months after the administration of the first dose of the immunogenic composition.

[0069] In some of the optional embodiments, a third dose of the immunogenic composition is administered to the subject. In some of the optional embodiments, the third dose of the immunogenic composition is administered approximately 4 to 8 months after the administration of the first dose of the immunogenic composition. In some of the optional embodiments, the third dose of the immunogenic composition is administered approximately 6 months after the administration of the first dose of the immunogenic composition.

[0070] In some of the embodiments, a booster dose of the immunogenic composition is administered to the subject. In some of the embodiments, the booster dose is administered at least about one year or at least about two years after the administration of the first dose.

[0071] In some of the embodiments, multiple booster doses are administered to the subject. In some of the embodiments, a first booster dose is administered at least about one year or at least about two years after the administration of the first dose. In some of the embodiments, the booster dose is administered once a year.

[0072] In some of the embodiments, the subject is a human subject.

[0073] In some of the embodiments, the human subjects are at least 18 years of age. In some of the embodiments, the human subjects are between 18 and 70 years of age.

[0074] In some of the embodiments, the human subjects are under 18 years of age. In some of the embodiments, the human subjects are between 2 and 5 years of age. In some of the embodiments, the human subjects are between 5 and 18 years of age.

[0075] In some embodiments, the use of any of the provided immunogenic compositions to stimulate an immune response to Borrelia in a subject by administering an effective amount of the immunogenic composition to the subject is also provided herein.

[0076] In some embodiments, the use of any of the immunogenic compositions provided is also provided herein for protecting a subject from infection by Borrelia by administering an effective amount of the immunogenic composition to the subject.

[0077] In some embodiments, the use of any of the immunogenic compositions provided is also provided herein for preventing a subject from contracting Lyme disease by administering an effective amount of the immunogenic composition to the subject.

[0078] In some embodiments, the use of any of the immunogenic compositions provided in the manufacture of a pharmaceutical product for stimulating an immune response to Borrelia in a subject is also provided herein.

[0079] In some embodiments, the use of any of the immunogenic compositions provided in the manufacture of a pharmaceutical product for protecting a subject from infection by Borrelia is also provided herein.

[0080] In some embodiments, the use of any of the immunogenic compositions provided in the manufacture of a pharmaceutical product for preventing a subject from contracting Lyme disease is also provided herein.

[0081] In some of the embodiments, the immunogenic composition is administered by intramuscular injection.

[0082] In some of the embodiments, a first dose and a second dose of the immunogenic composition are administered to the subject. In some of the embodiments, the second dose of the immunogenic composition is administered about 2 weeks to about 10 weeks after the administration of the first dose of the immunogenic composition. In some of the embodiments, the second dose of the immunogenic composition is administered about 1 month or about 2 months after the administration of the first dose of the immunogenic composition.

[0083] In some of the optional embodiments, a third dose of the immunogenic composition is administered to the subject. In some of the optional embodiments, the third dose of the immunogenic composition is administered approximately 4 to 8 months after the administration of the first dose of the immunogenic composition. In some of the optional embodiments, the third dose of the immunogenic composition is administered approximately 6 months after the administration of the first dose of the immunogenic composition.

[0084] In some of the embodiments, a booster dose of the immunogenic composition is administered to the subject. In some of the embodiments, the booster dose is administered at least about one year or at least about two years after the administration of the first dose.

[0085] In some of the embodiments, multiple booster doses are administered to the subject. In some of the embodiments, a first booster dose is administered at least about one year or at least about two years after the administration of the first dose. In some of the embodiments, the booster dose is administered once a year.

[0086] In some of the embodiments, the subject is a human subject.

[0087] In some of the embodiments, the human subjects are at least 18 years of age. In some of the embodiments, the human subjects are between 18 and 70 years of age.

[0088] In some of the embodiments, the human subjects are under 18 years of age. In some of the embodiments, the human subjects are between 2 and 5 years of age. In some of the embodiments, the human subjects are between 5 and 18 years of age.

[0089] In some embodiments, vials containing any of the immunogenic compositions provided are also provided herein.

[0090] In some of the embodiments, the immunogenic composition is in liquid form. In some of the embodiments, the immunogenic composition is in lyophilized form.

[0091] In some of the embodiments, the vial contains a single dose of the immunogenic composition. The single dose is between 0.3 mL and 1.5 mL.

[0092] In some embodiments, pre-filled syringes containing any of the immunogenic compositions provided are also provided herein.

[0093] In some of the embodiments, the syringe contains a single dose of the immunogenic composition. In some of the embodiments, the single dose is 0.3 mL to 1.5 mL.

[0094] In some embodiments, a kit is also provided herein, comprising: a) a first composition comprising an adjuvant selected from an aluminum salt adjuvant; a Toll-like receptor 9 (TLR9) agonist; a liposome-based adjuvant comprising lipopolysaccharides and saponins; and an oil-in-water emulsion-based adjuvant comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters; b) a second composition comprising a Borrelia cell surface protein A (OspA) antigen; and c) instructions for preparing any of the immunogenic compositions provided by combining the first and second compositions.

[0095] In some of the embodiments, the second composition is in liquid form. In some of the embodiments, the second composition is in lyophilized form.

[0096] In some of the embodiments, the instructions are for preparing a single-dose immunogenic composition. In some of the embodiments, the kit is for a single-dose immunogenic composition. In some of the embodiments, the single dose is 0.3 mL to 1.5 mL.

[0097] In some of the embodiments, the first and / or second composition includes a second adjuvant. In some of the embodiments, the first composition includes a second adjuvant. In some of the embodiments, the second composition includes a second adjuvant. In some of the embodiments, the first and second compositions each include a second adjuvant.

[0098] In some of the embodiments, the second adjuvant is an aluminum salt adjuvant. In some of the embodiments, the aluminum salt adjuvant comprises one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate. In some of the embodiments, the aluminum salt adjuvant comprises aluminum hydroxide.

[0099] In any embodiment, the kit further includes d) a further set of instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject.

[0100] In some embodiments, kits are also provided herein that include a) any of the immunogenic compositions provided; and b) instructions for administering the immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject.

[0101] In some of the embodiments, the kit further includes a syringe for intramuscular injection of the immunogenic composition. In some of the embodiments, the syringe is pre-filled with the immunogenic composition.

[0102] In some embodiments, kits are also provided herein, including a) any of the vials provided; and b) instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject.

[0103] In some of the embodiments, the kit further includes a syringe for intramuscular injection of an immunogenic composition.

[0104] In some embodiments, kits are also provided herein that include a) any of the pre-filled syringes provided; and b) instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject.

[0105] In some of the embodiments, the syringe is for intramuscular injection of the immunogenic composition.

[0106] In some of the embodiments, administration is carried out according to one of the methods provided.

[0107] In some of the embodiments, the subject is a human subject.

[0108] In some of the embodiments, the human subjects are at least 18 years of age. In some of the embodiments, the human subjects are between 18 and 70 years of age.

[0109] In some of the embodiments, the human subjects are under 18 years of age. In some of the embodiments, the human subjects are between 2 and 5 years of age. In some of the embodiments, the human subjects are between 5 and 18 years of age. [Brief explanation of the drawing]

[0110] [Figure 1]Figure 1 shows schematic diagrams of three chimeric OspA antigens, each of which combines two different serotypes derived from OspA. The top diagram is a schematic of the OspA antigen, which is divided into a C-terminal portion (top; plain) and an N-terminal portion (bottom; hatched pattern). The bottom diagrams show the three chimeric OspA antigens: S1 / 2 (left), S6 / 4 (center), and S5 / 3 (right), which contain different N-terminal and C-terminal portions derived from the indicated serotypes. [Figure 2A]Figures 2A-2G show antibody responses using ELISA against different indicated OspA antigens after administration of one of three OspA antigens or a mixture of OspA antigens (non-lipidized OspA chimeric antigen, non-lipidized OspA full-length serotype 1 (S1) antigen, or lipidized OspA chimeric antigen) using either no adjuvant (φ) or with one of the following adjuvant formulations: aluminum hydroxide (alum), CpG 1018® adjuvant (Dynavax Technologies Corporation, Emeryville, CA; CpG), or a combination of CpG and alum (CpG + alum). Figure 2A shows the antibody response to OspA S5 / 3 after administration of the indicated non-lipidized OspA chimeric antigen using different adjuvant formulations. Figure 2B shows the antibody response to OspA S6 / 4 after administration of the indicated non-lipidized OspA chimeric antigen using different adjuvant formulations. Figure 2C shows the antibody response to OspA S1 / 2 after administration of the indicated non-lipidized OspA antigen using different adjuvant formulations. Figure 2D shows the antibody response to the full-length OspA S1 antigen after administration of the indicated non-lipidized OspA antigen using different adjuvant formulations. Figure 2E shows the antibody response to OspA S1 / 2 after administration of the indicated lipidized OspA chimeric antigen using different adjuvant formulations. Figure 2F shows the antibody response to OspA S5 / 3 after administration of the indicated lipidized OspA chimeric antigen using different adjuvant formulations. Figure 2G shows the antibody response to OspA S6 / 4 after administration of the indicated lipidized OspA antigen using different adjuvant formulations. The multiplier increase (FI) of titer levels between certain immunogenic compositions is also shown above the graphs. In addition, the median antibody titer is also shown above each condition for Figures 2E-2G. [Figure 2B] Same as above. [Figure 2C] Same as above. [Figure 2D] Same as above. [Figure 2E] Same as above. [Figure 2F] Same as above. [Figure 2G] Same as above. [Figure 3A] Figures 3A–3C show antibody responses using ELISA against OspA chimeric antigens S1 / 2 (Figure 3A), S5 / 3 (Figure 3B), and S6 / 4 (Figure 3C) after the first injection (marked "1") or second injection (marked "2") of the lipid-modified OspA chimeric antigens and the indicated adjuvant formulations. The multiplicative increase (FI) in titer levels between the first and second injections is also shown above each immunogenic composition. [Figure 3B] Same as above. [Figure 3C] Same as above. [Modes for carrying out the invention]

[0111] Detailed explanation This disclosure relates to immunogenic compositions comprising Borrelia cell surface protein A (OspA) antigen or RNA polynucleotide encoding the OspA antigen, and an adjuvant. In some embodiments, the immunogenic compositions are suitable for stimulating an immune response to Borrelia in a subject. This disclosure also relates to products comprising the immunogenic compositions, their use, and methods of using them.

[0112] All publications, including patent documents, scientific articles, and databases, referenced in this application are incorporated by reference in their entirety for all purposes to the same extent as each individual publication is incorporated by reference individually. If any definition provided herein contradicts or otherwise conflicts with any definition provided herein in a patent, application, published application, or other publication incorporated herein by reference, the definition provided herein shall prevail over the definition incorporated herein by reference.

[0113] Section headings used herein are for organizational purposes only and should not be construed as limiting the subject matter described herein.

[0114] General techniques and definitions The practices described herein, unless otherwise indicated, utilize conventional techniques of molecular biology (including recombinant techniques), microbiology, cell biology, biochemistry, and immunology, which are within the scope of skills in the art.

[0115] As used herein and in the appended claims, the singular forms "a," "an," and "the" refer to multiple subjects unless otherwise indicated. For example, "an" excipient includes one or more excipients.

[0116] As used herein, the phrase “comprising” is open-ended and indicates that such an embodiment may include additional elements. In contrast, the phrase “consisting of” is closed and indicates that such an embodiment does not include additional elements (except for trace amounts of impurities). The phrase “consisting essentially of” is partially closed and indicates that such an embodiment may further include elements that do not substantially alter the essential features of such an embodiment.

[0117] As used herein with respect to values, the term "approximately" encompasses 90% to 110% of that value (for example, approximately 3000 μg of CpG adjuvant refers to 2700 μg to 3300 μg of CpG adjuvant).

[0118] Where used interchangeably herein, the terms “polynucleotide” and “oligonucleotide” include single-stranded DNA (ssDNA), double-stranded DNA (dsDNA), single-stranded RNA (ssRNA), and double-stranded RNA (dsRNA); modified oligonucleotides and oligonucleosides; or combinations thereof. Oligonucleotides may consist of linear or cyclic segments, or they may contain both linear and cyclic segments. Oligonucleotides are generally polymers of linked nucleosides through phosphodiester linkages, although alternative linkages such as phosphorothioate esters may also be used in oligonucleotides. Nucleosides consist of a sugar-bonded purine (adenine (A) or guanine (G), or a derivative thereof) or pyrimidine (thymine (T), cytosine (C), or uracil (U), or a derivative thereof) base. The four nucleoside units (or bases) in DNA are called deoxyadenosine, deoxyguanosine, thymidine, and deoxycytidine. A nucleotide is a phosphate ester of a nucleoside.

[0119] As used herein, the terms “CpG,” “CpG motif,” and “cytosine-phosphate-guanosine” refer to unmethylated cytidine-phospho-guanosine dinucleotide, which, when present in oligonucleotides, contributes to measurable immune responses in vitro, in vivo, and / or ex vivo. Examples of measurable immune responses include antigen-specific antibody production, cytokine secretion, and activation or proliferation of lymphocyte populations, such as NK cells, CD4+ T lymphocytes, CD8+ T lymphocytes, or B lymphocytes. Preferably, CpG-containing oligonucleotides preferentially activate Th1-type responses.

[0120] The term “antigen” refers to a substance that is specifically recognized and bound to an antibody or T cell antigen receptor. Antigens may include peptides, polypeptides, proteins, glycoproteins, polysaccharides, complex carbohydrates, sugars, gangliosides, lipids, and phospholipids; parts thereof; and combinations thereof. In the context of this disclosure, the term “antigen” typically refers to a polypeptide or protein antigen having a length of at least eight amino acid residues, which may include one or more post-translational modifications.

[0121] The terms “polypeptide” and “protein” are used interchangeably herein to refer to polymers of amino acid residues and are not limited to any particular length unless otherwise specified. Polypeptides may comprise native amino acid residues or combinations of native and non-native amino acid residues. The term also includes post-expression modifications of polypeptides, such as glycosylation, sialylation, acetylation, and phosphorylation. In some embodiments, polypeptides may include modifications to the native or native sequence, as long as the protein maintains the desired activity (e.g., antigenicity).

[0122] As used herein, the terms “isolated” and “purified” refer to material that has been removed from at least one naturally occurring and related component (e.g., removed from its original environment). When used in reference to recombinant proteins, the term “isolated” refers to a protein removed from the culture medium of the host cell that produced the protein. In some embodiments, the isolated protein (e.g., Borrelia antigen) is at least 75%, 90%, 95%, 96%, 97%, 98%, or 99% pure, as determined by high-performance liquid chromatography (HPLC).

[0123] An "effective amount" or "sufficient amount" of a substance is the amount sufficient to produce a beneficial or desired outcome, including clinical results, and therefore, the "effective amount" depends on the context in which it is applied. In the context of administering an immunogenic composition, an effective amount contains sufficient antigen and adjuvant to stimulate an immune response (preferably, a seroprotection level of the antibody and antigen).

[0124] In this disclosure, the terms “individual” and “subject” refer to mammals. Mammals include humans, non-human primates (e.g., monkeys), domesticated animals, sporting animals, rodents (e.g., mice and rats), and pets (e.g., dogs and cats).

[0125] As used herein with respect to immunogenic compositions, the term “dose” refers to the measured portion of the immunogenic composition ingested by (administered to or received by) the subject at any given time.

[0126] "Stimulation" of a response or parameter includes inducing and / or enhancing that response or parameter when compared to otherwise identical conditions except for the parameter of interest, or alternatively, when compared to a different condition. For example, "stimulation" of an immune response means an increase in the response. Depending on the parameter being measured, the increase may be 2 to 200 times or more, 5 to 500 times or more, 10 to 1000 times or more, or 2, 5, 10, 50, or 100 to 200, 500, 1,000, 5,000, or 10,000 times.

[0127] Conversely, “inhibition” of a response or parameter includes reducing and / or suppressing that response or parameter when compared to otherwise identical conditions except for the parameter of interest, or alternatively, when compared to a different condition. For example, “inhibition” of an immune response means a reduction in the response. Depending on the parameter being measured, the reduction may be 2 to 200 times, 5 to 500 times or more, 10 to 1000 times or more, or 2, 5, 10, 50, or 100 to 200, 500, 1,000, 2,000, 5,000, or 10,000 times.

[0128] The relative terms “higher” and “lower” refer to a measurable increase or decrease in a response or parameter, respectively, compared to otherwise identical conditions except for the parameter of interest, or alternatively, compared to a different condition. For example, “higher antibody titer” refers to an antigen-reactive antibody titer resulting from administration of the immunogenic composition of this disclosure that is at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 times higher than the antigen-reactive antibody titer resulting from control conditions. Similarly, “lower antibody titer” refers to an antigen-reactive antibody titer resulting from control conditions that is at least 2, 3, 4, 5, 6, 7, 8, 9, or 10 times lower than the antigen-reactive antibody titer resulting from administration of the immunogenic composition of this disclosure.

[0129] As used herein, the term “immunization” refers to a process that increases the response of an object to an antigen and thus improves its ability to resist or overcome infection and / or resist disease.

[0130] As used herein, the term "vaccination" refers to the introduction of a vaccine into the subject's body.

[0131] An "adjuvant" refers to a substance that, when added to a composition containing an antigen, enhances or strengthens the immune response to the antigen in a subject upon exposure.

[0132] As used herein, “amino acid sequence identity percentage (%)”, “identity percentage”, and “sequence identity” are defined, when used in relation to an amino acid sequence (reference polypeptide sequence), as the percentage of amino acid residues in a candidate sequence (e.g., target antigen) that are identical to the amino acid residues in the reference polypeptide sequence, without considering any conservative substitutions as part of the sequence identity, after sequence alignment and gap introduction as necessary to achieve the maximum sequence identity percentage. Alignment for the purpose of determining the amino acid sequence identity percentage can be achieved in various ways within the skill of the art, for example, using publicly available computer software such as BLAST, BLAST-2, ALIGN, or Megalign (DNASTAR) software. Those skilled in the art can determine appropriate parameters for aligning sequences, including any algorithm necessary to achieve the maximum alignment over the entire length of the sequences being compared.

[0133] Amino acid substitutions may include replacing one amino acid in a polypeptide with another amino acid. Amino acid substitutions may be introduced into the antigen of interest, as well as into products screened for desired activity, such as increased stability and / or immunogenicity.

[0134] Amino acids can generally be classified according to the following common side-chain characteristics: (1) Hydrophobic: norleucine, Met, Ala, Val, Leu, Ile; (2) Neutral hydrophilic: Cys, Ser, Thr, Asn, Gln; (3) Acidic: Asp, Glu; (4) Basicity: His, Lys, Arg; (5) Residues that affect chain orientation: Gly, Pro; and (6) Aromatic: Trp, Tyr, Phe.

[0135] Conservative amino acid substitutions involve replacing one member of these classes with another member of the same class. Non-conservative amino acid substitutions involve replacing one member of these classes with a member of another class.

[0136] I. Immunogenic composition This disclosure relates to immunogenic compositions containing Borrelia cell surface protein A (OspA) antigen or RNA polynucleotide encoding the OspA antigen, and an adjuvant. In some embodiments, the immunogenic composition contains Borrelia OspA antigen and an adjuvant. In some embodiments, the adjuvant is any of those described herein, for example, any of those described in Section IA. In some embodiments, the OspA antigen is any of those described herein, for example, any of those described in Section IB. In some embodiments, the immunogenic composition contains additional components, for example, any of those described in Section IC.

[0137] In some embodiments, the immunogenic composition is intended to stimulate an immune response to Borrelia in a subject. In some embodiments, the antigen and adjuvant are present in the immunogenic composition in amounts effective for stimulating an immune response to Borrelia in the subject. In some embodiments, stimulation is achieved by administering an effective amount of the immunogenic composition to the subject.

[0138] In some embodiments, the immunogenic composition is intended to protect a subject from infection by Borrelia. In some embodiments, the antigen and adjuvant are present in the immunogenic composition in amounts effective to stimulate an immune response against Borrelia in the subject. In some embodiments, protection is achieved by administering an effective amount of the immunogenic composition to the subject.

[0139] In some embodiments, the immunogenic composition is intended to prevent a subject from contracting Lyme disease. In some embodiments, the antigen and adjuvant are present in the immunogenic composition in amounts effective to stimulate an immune response to Borrelia in the subject. In some embodiments, prevention is achieved by administering an effective amount of the immunogenic composition to the subject.

[0140] In some embodiments, the subjects are human subjects. In some embodiments, the human subjects are at least 18 years old. In some embodiments, the human subjects are approximately 18 to 70 years old. In some embodiments, the human subjects are approximately 18 to 55 years old. In some embodiments, the human subjects are under 18 years old. In some embodiments, the human subjects are approximately 2 to 5 years old. In some embodiments, the human subjects are approximately 5 to 18 years old.

[0141] In some embodiments, the immunogenic composition is in a dose of 0.3 mL to 1.5 mL. In some embodiments, the immunogenic composition is in a dose of approximately 0.25 mL, 0.50 mL, 0.75 mL, or 1.0 mL. In some embodiments, the immunogenic composition is in a dose of approximately 0.50 mL. In some embodiments, the immunogenic composition is in a dose of approximately 0.75 mL. In some embodiments, the immunogenic composition is in a unit dose.

[0142] A. Adjuvant Adjuvants that may be used in this immunogenic composition include inorganic salts, e.g., aluminum salts, e.g., aluminum hydroxide, aluminum hydroxide gel (e.g., Rehydragel®), aluminum phosphate, and calcium phosphate; surfactants and microparticles, e.g., nonionic block polymer surfactants, cholesterol, virosoms, saponins (e.g., Quill A, QS-21, and GPI-0100 in Matrix-M®, etc. (Lovgren et al., Biotechnol Appl Biochem, 10(2):161~172, 1988)), proteasomes, immunostimulatory complexes, cochleates, quaternary amines (dimethyldioctadecylammonium bromide (DDA)), pyridine, vitamin A, vitamin E, and poly(lactide-coglycolide) microparticles (Shah et al., Methods Mol). Biol, 1494:1~14, 2017); bacterial products, e.g., RIBI adjuvant system (Ribi Inc.), cell wall skeleton of Mycobacterium phlei (Detox®), muramyl dipeptide (MDP) and tripeptide (MTP), monophosphoryl lipid A, Bacillus Calmete-Guerin, heat-unstable E. coli enterotoxin, cholera toxin, trehalose dimycolate, and CpG oligodeoxynucleotide; cytokines and hormones, e.g., interleukins (IL-1, IL-2, IL-6, IL-12, IL-15, and IL-18), granulocyte-macrophage colony-stimulating factor, dehydroepiandrosterone, and 1,25-dihydroxyvitamin D3; polyanions, e.g., dextran; polyacrylic acids (e.g., polymethyl methacrylate and Carbopol 934P); Carriers, e.g., tetanus toxoid, diphtheria toxoid, cholera toxin B subunit, enterotoxogenic E. coli mutant heat-unstable enterotoxin (rmLT), and heat shock proteins; oil-in-water emulsions, e.g., AMPHIGEN® (Hydronics, USA); water-in-oil emulsions, e.g., Freund's complete and incomplete adjuvants; and liposome-based adjuvants.Further adjuvants that may be used in this immunogenic composition are described in Pulendran et al., Nat Rev Drug Discov 20:454~475, 2021; and Facciola et al., Vaccines (Basel) 10(5):819, 2022.

[0143] In some embodiments, the adjuvant is selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants; and oil-in-water emulsion-based adjuvants. In some embodiments, the adjuvant is selected from the group consisting of aluminum salt adjuvants; TLR9 agonists which are oligonucleotides containing a non-methylated cytidine-phospho-guanosine (CpG) motif; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0144] In some embodiments, the adjuvant is an aluminum salt adjuvant, for example, any of those described herein, e.g., in Section IA-1. In some embodiments, the adjuvant is a TLR9 agonist, for example, a TLR9 agonist which is an oligonucleotide containing a non-methylated CpG motif, for example, any of those described herein, e.g., in Section IA-2. In some embodiments, the adjuvant is a liposome-based adjuvant, for example, a liposome-based adjuvant containing lipopolysaccharides and saponins, for example, any of those described herein, e.g., in Section IA-3. In some embodiments, the adjuvant is an oil-in-water emulsion-based adjuvant, for example, an oil-in-water emulsion-based adjuvant containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters, for example, any of those described herein, e.g., in Section IA-4.

[0145] In some embodiments, the immunogenic composition contains a second adjuvant distinct from a first adjuvant. In some embodiments, the first and second adjuvants are, individually, any of the adjuvants described herein. In some embodiments, the first and second adjuvants are selected from the group consisting of aluminum salt adjuvants; TLR9 agonists; liposome-based adjuvants; and oil-in-water emulsion-based adjuvants. In some embodiments, the first and second adjuvants are selected from the group consisting of aluminum salt adjuvants; TLR9 agonists which are oligonucleotides containing a non-methylated CpG motif; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

[0146] In some embodiments, the second adjuvant is a polycationic carrier. In some embodiments, the first adjuvant is a TLR9 agonist, e.g., an oligonucleotide containing a non-methylated CpG motif, and the second adjuvant is a polycationic carrier. In some embodiments, the first adjuvant, e.g., the TLR9 agonist, forms a cationic complex with the polycationic carrier. In some embodiments, the formation of the cationic complex results in an increased immune response compared to the immune response induced by the first adjuvant alone. In some embodiments, the polycationic carrier is any one described in Dong et al. ACS Appl. Mater. Interfaces 14(5):6331~6342, 2022; and Fu et al. European Polymer Journal 113:115~132, 2019. In some embodiments, the polycationic carrier is polycationic polyethyleneimine (PEI). In some embodiments, the polycationic carrier is an aluminum salt adjuvant, for example, any of those described herein, for example, in Section IA-1.

[0147] In some embodiments, the second adjuvant is an aluminum salt adjuvant, which may be any of the amounts described herein. In some embodiments, the first adjuvant is a TLR9 agonist, e.g., an oligonucleotide containing a non-methylated CpG motif, and the second adjuvant is an aluminum salt adjuvant. In some embodiments, the first adjuvant is a liposome-based adjuvant, e.g., a liposome-based adjuvant containing lipopolysaccharides and saponins, and the second adjuvant is an aluminum salt adjuvant. In some embodiments, the first adjuvant is an oil-in-water emulsion-based adjuvant, e.g., an oil-in-water emulsion-based adjuvant containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters, and the second adjuvant is an aluminum salt adjuvant. In some embodiments, the aluminum salt adjuvant is one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate. In some embodiments, the aluminum salt adjuvant is one or both of aluminum hydroxide and aluminum phosphate. In some embodiments, the aluminum salt adjuvant is aluminum hydroxide. In some embodiments, the aluminum salt adjuvant is potassium aluminum sulfate.

[0148] In some embodiments, the first adjuvant is a TLR9 agonist, for example, an oligonucleotide containing a non-methylated CpG motif, and the second adjuvant is potassium aluminum sulfate. In some embodiments, the first adjuvant is a liposome-based adjuvant, for example, a liposome-based adjuvant containing lipopolysaccharides and saponins, and the second adjuvant is potassium aluminum sulfate. In some embodiments, the first adjuvant is an oil-in-water emulsion-based adjuvant, for example, an oil-in-water emulsion-based adjuvant containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters, and the second adjuvant is potassium aluminum sulfate.

[0149] In some embodiments, the first adjuvant is a lipopolysaccharide, and the second adjuvant is an aluminum salt adjuvant. In some embodiments, the lipopolysaccharide is adsorbed onto the aluminum salt adjuvant. In some embodiments, the lipopolysaccharide is lipid A or a derivative thereof. In some embodiments, the lipopolysaccharide is monophosphoryl lipid A or a derivative thereof. In some embodiments, the lipopolysaccharide is 3-O-desacyl-4'-monophosphoryl lipid A. In some embodiments, the lipopolysaccharide is 3-deacyl-phosphorylated hexaacyl disaccharide. In some embodiments, the aluminum salt adjuvant is one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and aluminum potassium sulfate. In some embodiments, the aluminum salt adjuvant is one or both of aluminum hydroxide and aluminum phosphate. In some embodiments, the aluminum salt adjuvant is aluminum hydroxide. In some embodiments, the aluminum salt adjuvant is aluminum potassium sulfate. In some embodiments, the first adjuvant is 3-O-desacyl-4'-monophosphoryl lipid A, and the second adjuvant is aluminum hydroxide. In some embodiments, the first and second adjuvants together constitute adjuvant AS04. In some embodiments, the adjuvant is AS04.

[0150] In some embodiments, the adjuvant is a synthetic TLR7 / 8 agonist. In some embodiments, the adjuvant is 3M-052 (Kasturi et al., Sci Immunol 5(48):eabb1025, June 19, 2020). In some embodiments, 3M-052 is formulated in poly(lactic acid-coglycolic acid) or PLGA nanoparticles. In some embodiments, 3M-052 is formulated with an aluminum salt adjuvant. In some embodiments, 3M-052 is formulated with potassium aluminum sulfate. In some embodiments, the first adjuvant is 3M-052 and the second adjuvant is any of those described herein. In some embodiments, the first adjuvant is 3M-052 and the second adjuvant is a TLR4 agonist. In some embodiments, the TLR4 agonist is GLA.

[0151] In some embodiments, the adjuvant is a saponin-based adjuvant. In some embodiments, the adjuvant consists of nanoparticles containing saponins. In some embodiments, the saponins are extracted from the Quillaja saponaria tree. In some embodiments, the nanoparticles contain saponins, cholesterol, and phospholipids. In some embodiments, the adjuvant is Matrix-M®.

[0152] 1. Aluminum salt adjuvant In some embodiments, the adjuvant is an aluminum salt adjuvant. In some embodiments, the OspA antigen is adsorbed to the aluminum salt adjuvant. In some embodiments, the aluminum salt adjuvant is one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and aluminum potassium sulfate. In some embodiments, the aluminum salt adjuvant is one or both of aluminum hydroxide and aluminum phosphate. In some embodiments, the aluminum salt adjuvant is aluminum hydroxide. In some embodiments, the aluminum salt adjuvant is aluminum potassium sulfate.

[0153] In some embodiments, the immunogenic composition contains about 0.25 to about 1.25 mg of Al 3+ It contains. In some embodiments, the immunogenic composition contains about 0.5 to about 1 mg of Al 3+ It contains. In some embodiments, the immunogenic composition contains about 0.25 mg to about 0.50 mg of Al 3+ It contains. In some embodiments, the immunogenic composition contains about 0.30 mg to about 0.40 mg of Al 3+ It contains. In some embodiments, the immunogenic composition contains about 0.25 mg, about 0.50 mg, about 0.75 mg, about 1.00 mg, or about 1.25 mg of Al 3+ It contains.

[0154] 2. Oligonucleotide toll-like receptor 9 (TLR9) agonists Toll-like receptors (TLRs) are expressed in and on dendritic cells and other innate immune cells, and are among the most important receptors for stimulating responses to the presence of invading pathogens. Humans have multiple types of TLRs that recognize different parts of viruses or bacteria but have similar structures. By activating specific TLRs, it is possible to stimulate and control specific types of innate immune responses that can be used to enhance adaptive responses.

[0155] In some embodiments, the adjuvant in this immunogenic composition is an oligonucleotide TLR9 agonist. In some embodiments, the oligonucleotide contains an unmethylated cytidine-phosphoguanosine (CpG) motif. TLR9 (CD289) recognizes the unmethylated CpG motif found in microbial DNA, which can be mimicked using synthetic CpG-containing oligodeoxynucleotides (CpG-ODNs). CpG-ODNs are known to enhance antibody production and stimulate helper T1 (Th1) cell responses (Coffman et al., Immunity, 33:492-503, 2010). Based on structure and biological function, CpG-ODNs are divided into three common classes: CpG-A, CpG-B, and CpG-C (Campbell, Methods Mol Biol, 1494:15-27, 2017). The degree of B cell activation varies among classes, with CpG-A ODN being a weak B cell activator, CpG-C ODN being a good B cell activator, and CpG-B ODN being a strong B cell activator. The oligonucleotide TLR9 agonist in this immunogenic composition is preferably a good B cell activator (CpG-C ODN), or more preferably a strong B cell activator (CpG-B ODN).

[0156] Oligonucleotide TLR9 agonists often contain a palindromic sequence following the general formula 5'-purine-purine-CG-pyrimidine-pyrimidine-3' or 5'-purine-purine-CG-pyrimidine-pyrimidine-CG-3' (U.S. Patent No. 6,589,940). TLR9 agonism is also observed with certain non-palindromic CpG-enriched phosphorothioate oligonucleotides, but may not be affected by changes in the nucleotide sequence. In addition, TLR9 agonism can be neutralized by methylation of cytosine within the CpG dinucleotide. In some embodiments, the TLR9 agonist in this immunogenic composition is an oligonucleotide of 8 to 35 nucleotides in length. In some embodiments, the TLR9 agonist in this immunogenic composition is an oligonucleotide of 8 to 35 nucleotides in length containing the sequence 5'-AACGTTCG-3'. In some embodiments, the oligonucleotide is longer than 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides, and shorter than 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, or 24 nucleotides. In some embodiments, the TLR9 agonist is an oligonucleotide of 10 to 35 nucleotides in length. In some embodiments, the TLR9 agonist is an oligonucleotide of 10 to 35 nucleotides in length containing the sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2). In some embodiments, the oligonucleotide is longer than 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 nucleotides, and shorter than 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, or 24 nucleotides. In some embodiments, the oligonucleotide does not contain the sequence 5'-TCGTCGTTTT GTCGTTTTGT CGTT-3' (as shown in ODN 2006 or ODN 7909, SEQ ID NO: 3).

[0157] Researchers at Dynavax Technologies Corporation (Emeryville, CA) have identified CpG 1018® adjuvant, a 22-mer phosphorothioate-linked oligodeoxynucleotide, which contains a specific sequence capable of substantially enhancing the immune response to co-administered antigens across species (Campbell, Methods Mol Biol, 1494:15~27, 2017). CpG 1018® adjuvant (5'-TGACTGTGAA CGTTCGAGAT GA-3', shown in SEQ ID NO: 1) was selected after screening a broad panel of oligonucleotides for their immunostimulatory activity in vitro and in vivo. CpG 1018® adjuvant is a CpG-B ODN that is active in mouse, rabbit, dog, baboon, cynomolgus monkey, and human. In some embodiments, the oligonucleotide is 22–35 nucleotides long and contains the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1). In some embodiments, provided that the longest length is less than the minimum length, the oligonucleotide is longer than 22, 23, 24, 25, 26, 27, 28, 29, 30, 31, 32, or 33 nucleotides, and shorter than 35, 34, 33, 32, 31, 30, 29, 28, 27, 26, 25, or 24 nucleotides.

[0158] The exemplary oligonucleotide CpG 1018® adjuvant is CpG-ODN, but oligonucleotides that are not complete DNA molecules can also be used in this immunogenic composition. In some embodiments, the oligonucleotide is a DNA / RNA chimeric molecule, where CpG and the palindromic sequence are deoxyribonucleic acid, and one or more nucleic acids outside these regions are ribonucleic acid. In some embodiments, the oligonucleotide is linear. In other embodiments, the oligonucleotide is circular or contains a hairpin loop. In some embodiments, the oligonucleotide is single-stranded. In some embodiments, the oligonucleotide is double-stranded.

[0159] In some embodiments, the oligonucleotide may contain modifications. These modifications may be modifications of the 3'OH or 5'OH group, modifications of nucleotide bases, modifications of sugar components, and modifications of phosphate groups. The modified bases may contain a palindromic sequence of the oligonucleotide, insofar as the modified bases maintain the same specificity to their natural complement through Watson-Crick base pairing (e.g., the palindromic portion remains self-complementary). In some embodiments, the oligonucleotide contains a non-standard base. In some embodiments, the oligonucleotide contains a modified nucleoside. In some embodiments, the modified nucleoside is selected from the group consisting of 2'-deoxy-7-deazaguanosine, 2'-deoxy-6-thioguanosine, arabinoguanosine, 2'-deoxy-2'-substituted arabinoguanosine, and 2'-O-substituted arabinoguanosine.

[0160] Oligonucleotides may contain phosphate group modifications. For example, in addition to phosphodiester linkages, phosphate modifications include methylphosphonates, phosphorothioates, phosphoramidates (crosslinked or uncrosslinked), phosphotryesters, and phosphorodithioates, and may be used in any combination. Other non-phosphate linkages may also be used. In some embodiments, oligonucleotides contain only a phosphorothioate skeleton. In some embodiments, oligonucleotides contain only a phosphodiester skeleton. In some embodiments, oligonucleotides contain a combination of phosphate linkages in the phosphate skeleton, for example, a combination of phosphodiester and phosphorothioate linkages. Oligonucleotides having a phosphorothioate skeleton may be more immunogenic and more resistant to degradation after injection into the host than those having a phosphodiester skeleton (Braun et al., J Immunol, 141:2084~2089, 1988; and Latimer et al., Mol Immunol, 32:1057~1064, 1995). In some embodiments, the oligonucleotide comprises at least one, two, or three internucleotide phosphorothioate ester links. In some embodiments, when a number of oligonucleotide molecules are present in a pharmaceutical composition containing at least one excipient, both stereoisomers of the phosphorothioate ester links are present in the number of oligonucleotide molecules. In some embodiments, all internucleotide links of the oligonucleotide are phosphorothioate links, or in other words, the oligonucleotide has a phosphorothioate backbone.

[0161] In some embodiments, the immunogenic composition contains about 375 μg to about 6000 μg of oligonucleotides. In some embodiments, the immunogenic composition contains about 750 μg to about 6000 μg of oligonucleotides. In some embodiments, the immunogenic composition contains about 750 μg to about 3000 μg of oligonucleotides. In some embodiments, the immunogenic composition contains about 750 μg to about 1500 μg of oligonucleotides. In some embodiments, the immunogenic composition contains about 1500 μg to about 3000 μg of oligonucleotides. In some embodiments, the immunogenic composition contains about 3000 μg to about 6000 μg of oligonucleotides. In some embodiments, the immunogenic composition contains about 750, 1000, 1250, or 1500 μg or more of oligonucleotides, and about 6000, 5000, 4000, or 3000 μg or less of oligonucleotides. In some embodiments, the immunogenic composition contains about 375, 750, 1500, 3000, or 6000 μg of oligonucleotide. In some embodiments, the immunogenic composition contains about 375 μg of oligonucleotide. In some embodiments, the immunogenic composition contains about 750 μg of oligonucleotide. In some embodiments, the immunogenic composition contains about 1000 μg of oligonucleotide. In some embodiments, the immunogenic composition contains about 1500 μg of oligonucleotide. In some embodiments, the immunogenic composition contains about 3000 μg of oligonucleotide. In some embodiments, the immunogenic composition contains about 6000 μg of oligonucleotide.

[0162] In some embodiments, the oligonucleotide is in the form of its pharmaceutically acceptable salt. Exemplary basic salts include ammonium salts, alkali metal salts, e.g., sodium salts, lithium salts, and potassium salts; alkaline earth metal salts, e.g., calcium salts, magnesium salts, and zinc salts; organic bases (e.g., organic amines), e.g., salts with N-Me-D-glucamine, N-[1-(2,3-dioleoyloxy)propyl]-N,N,N-trimethylammonium chloride, choline, tromethamine, dicyclohexylamine, and t-butylamine; and amino acids, e.g., salts with arginine and lysine. In some embodiments, the oligonucleotide is in the form of an ammonium salt, sodium salt, lithium salt, or potassium salt. In some embodiments, the oligonucleotide is in the form of a sodium salt.

[0163] 3. Liposome-based adjuvants In some embodiments, the adjuvant in the immunogenic composition is a liposome-based adjuvant, for example, any of those described in Tretiakova et al., Biochem (Mosc) Suppl Ser A Membr Cell Biol, 16(1):1~20, 2022; or Alving et al., Expert Rev Vaccines 19(3):279~292, 2020. In some embodiments, the liposome-based adjuvant contains a lipopolysaccharide. In some embodiments, the liposome-based adjuvant contains a saponin. In some embodiments, the liposome-based adjuvant contains both a lipopolysaccharide and a saponin.

[0164] In some embodiments, the lipopolysaccharide is lipid A or a derivative thereof. In some embodiments, the lipopolysaccharide is monophosphoryl lipid A or a derivative thereof. In some embodiments, the lipopolysaccharide is naturally occurring. In some embodiments, the lipopolysaccharide is synthetic. In some embodiments, the lipopolysaccharide is 3-O-desacyl-4'-monophosphoryl lipid A. In some embodiments, the lipopolysaccharide is 3-deacyl-phosphorylated hexaacyl disaccharide.

[0165] In some embodiments, the immunogenic composition contains about 1 μg to about 100 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 10 μg to about 50 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 20 μg to about 30 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 21 μg to about 29 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 22 μg to about 28 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 23 μg to about 27 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 24 μg to about 26 μg of lipopolysaccharide.

[0166] In some embodiments, the immunogenic composition contains about 1 μg to about 30 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 5 μg to about 15 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 6 μg to about 14 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 7 μg to about 13 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 8 μg to about 12 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 9 μg to about 11 μg of lipopolysaccharide.

[0167] In some embodiments, the immunogenic composition contains about 1 μg to about 9 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 2 μg to about 8 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 3 μg to about 7 μg of lipopolysaccharide. In some embodiments, the immunogenic composition contains about 4 μg to about 6 μg of lipopolysaccharide.

[0168] In some embodiments, the saponin is naturally occurring. In some embodiments, the saponin is synthetic. In some embodiments, the saponin is derived from the bark of Quillaja saponaria Molina. In some embodiments, the saponin is Quill A or a derivative thereof. In some embodiments, the saponin is QS-17. In some embodiments, the saponin is QS-21.

[0169] In some embodiments, the immunogenic composition contains about 1 μg to about 100 μg of saponins. In some embodiments, the immunogenic composition contains about 10 μg to about 50 μg of saponins. In some embodiments, the immunogenic composition contains about 20 μg to about 30 μg of saponins. In some embodiments, the immunogenic composition contains about 21 μg to about 29 μg of saponins. In some embodiments, the immunogenic composition contains about 22 μg to about 28 μg of saponins. In some embodiments, the immunogenic composition contains about 23 μg to about 27 μg of saponins. In some embodiments, the immunogenic composition contains about 24 μg to about 26 μg of saponins.

[0170] In some embodiments, the immunogenic composition contains about 1 μg to about 30 μg of saponins. In some embodiments, the immunogenic composition contains about 5 μg to about 15 μg of saponins. In some embodiments, the immunogenic composition contains about 6 μg to about 14 μg of saponins. In some embodiments, the immunogenic composition contains about 7 μg to about 13 μg of saponins. In some embodiments, the immunogenic composition contains about 8 μg to about 12 μg of saponins. In some embodiments, the immunogenic composition contains about 9 μg to about 11 μg of saponins.

[0171] In some embodiments, the immunogenic composition contains about 1 μg to about 9 μg of saponins. In some embodiments, the immunogenic composition contains about 2 μg to about 8 μg of saponins. In some embodiments, the immunogenic composition contains about 3 μg to about 7 μg of saponins. In some embodiments, the immunogenic composition contains about 4 μg to about 6 μg of saponins.

[0172] In some embodiments, the liposome-based adjuvant further contains a sterol. In some embodiments, the sterol is β-sitosterol, stigmasterol, ergosterol, ergocalciferol, or cholesterol. In some embodiments, the sterol is cholesterol.

[0173] In some embodiments, the ratio of saponins to sterols is 1:1 to 1:100 w / w. In some embodiments, the ratio of saponins to sterols is 1:1 to 1:5 w / w.

[0174] In some embodiments, the liposome-based adjuvant further contains a neutral lipid. In some embodiments, the neutral lipid is phosphatidylcholine. In some embodiments, the phosphatidylcholine is egg yolk phosphatidylcholine, dioleoyl phosphatidylcholine (DOPC), or dilauryl phosphatidylcholine. In some embodiments, the phosphatidylcholine is DOPC.

[0175] In some embodiments, the liposome-based adjuvant contains 3-O-desacyl-4'-monophosphoryl lipid A and QS-21. In some embodiments, the liposome-based adjuvant contains 3-O-desacyl-4'-monophosphoryl lipid A, QS-21, and cholesterol. In some embodiments, the liposome-based adjuvant contains 3-O-desacyl-4'-monophosphoryl lipid A, QS-21, cholesterol, and DOPC. In some embodiments, the liposome-based adjuvant is AS01, AS01B, or AS01E, for example, as described in U.S. Patent No. 10,039,823. In some embodiments, the liposome-based adjuvant is AS01. In some embodiments, the liposome-based adjuvant is AS01B. In some embodiments, the liposome-based adjuvant is AS01E.

[0176] In some embodiments, the liposome-based adjuvant contains 3-deacyl-phosphorylated hexaacyl disaccharide and QS-21. In some embodiments, the liposome-based adjuvant is ALFQ.

[0177] 4. Oil-in-water emulsion-based adjuvants In some embodiments, the adjuvant in the immunogenic composition is an oil-in-water emulsion-based adjuvant, for example, any of those described in Zhang et al., BMC Vet Res 14:415, 2018. In some embodiments, the oil-in-water emulsion-based adjuvant contains squalene. In some embodiments, the oil-in-water emulsion-based adjuvant contains a sorbitan ester (also known as Span). In some embodiments, the oil-in-water emulsion-based adjuvant contains a polyoxyethylene sorbitan ester (also known as Tween). In some embodiments, the oil-in-water emulsion-based adjuvant contains squalene, a sorbitan ester, and a polyoxyethylene sorbitan ester.

[0178] In some embodiments, the sorbitan ester is sorbitan ester 20. In some embodiments, the sorbitan ester is sorbitan ester 85.

[0179] In some embodiments, the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 20. In some embodiments, the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 80.

[0180] In some embodiments, the immunogenic composition contains about 2% w / v to about 7% w / v squalene. In some embodiments, the immunogenic composition contains about 3% w / v to about 6% w / v squalene. In some embodiments, the immunogenic composition contains about 4% w / v to about 5% w / v squalene.

[0181] In some embodiments, the immunogenic composition contains about 0.2% w / v to about 0.8% w / v of sorbitan ester. In some embodiments, the immunogenic composition contains about 0.3% w / v to about 0.7% w / v of sorbitan ester. In some embodiments, the immunogenic composition contains about 0.4% w / v to about 0.6% w / v of sorbitan ester.

[0182] In some embodiments, the immunogenic composition contains about 0.2% w / v to about 0.8% w / v of polyoxyethylene sorbitan ester. In some embodiments, the immunogenic composition contains about 0.3% w / v to about 0.7% w / v of polyoxyethylene sorbitan ester. In some embodiments, the immunogenic composition contains about 0.4% w / v to about 0.6% w / v of polyoxyethylene sorbitan ester.

[0183] In some embodiments, the oil-in-water emulsion-based adjuvant contains squalene, sorbitan ester 85, and polyoxyethylene sorbitan ester 80. In some embodiments, the immunogenic composition contains about 2% w / v to about 7% w / v squalene; about 0.2% w / v to about 0.8% w / v sorbitan ester 85; and about 0.2% w / v to about 0.8% w / v polyoxyethylene sorbitan ester 80. In some embodiments, the oil-in-water emulsion-based adjuvant is MF59. In some embodiments, the oil-in-water emulsion-based adjuvant is AddaVax®.

[0184] In some embodiments, the oil-in-water emulsion-based adjuvant contains additional immunoenhancing components. In some embodiments, the oil-in-water emulsion-based adjuvant contains vitamins. In some embodiments, the oil-in-water emulsion-based adjuvant contains vitamin E.

[0185] In some embodiments, the oil-in-water emulsion-based adjuvant contains squalene, vitamin E, and polyoxyethylene sorbitan ester 80. In some embodiments, the oil-in-water emulsion-based adjuvant is AS03. In some embodiments, the immunogenic composition contains about 11 mg of squalene, about 12 mg of vitamin E, and about 5 mg of polyoxyethylene sorbitan ester 80.

[0186] B. Borrelia cell surface protein A (OspA) antigen and RNA polynucleotide In some embodiments, the immunogenic composition contains Borrelia cell surface protein A (OspA) antigen or RNA polynucleotide encoding OspA antigen. In some embodiments, the immunogenic composition contains Borrelia OspA antigen. In some embodiments, the immunogenic composition does not contain non-OspA Borrelia cell surface protein antigen.

[0187] In some embodiments, the immunogenic composition contains an RNA polynucleotide encoding an OspA antigen. In some embodiments, the immunogenic composition does not contain an RNA polynucleotide encoding a non-OspA Borrelia cell surface protein antigen. In some embodiments, the RNA polynucleotide further comprises a 5' untranslated region (UTR) and a 3'UTR. In some embodiments, the RNA polynucleotide further comprises a poly(A) tail. In some embodiments, the RNA polynucleotide further comprises a 5' cap analog. In some embodiments, the RNA polynucleotide is a modified polynucleotide, for example, those described in U.S. Patents 10,898,574, 10,703,789, and 10,577,403.

[0188] RNA polynucleotides may be formulated with one or more excipients to (1) increase stability; (2) increase cell transfection; (3) enable sustained or delayed release; (4) alter in vivo distribution to specific tissues or cell types; (5) increase translation of encoded proteins in vivo; and / or (6) alter the release profile of encoded proteins in vivo. Excipients may include solvents, dispersion media, diluents, other liquid vehicles, dispersion or suspension aids, surfactants, isotonic agents, thickeners or emulsifiers, preservatives, lipidoids, liposomes, lipid nanoparticles, polymers, lipoplexes, core-shell nanoparticles, peptides, proteins, hyaluronidases, nanoparticle mimics, and combinations thereof. Exemplary formulations for RNA polynucleotides are described in U.S. Patents 10,898,574, 10,703,789, and 10,577,403.

[0189] In some embodiments, RNA polynucleotides are formulated in lipid nanoparticles. Examples of lipid nanoparticle compositions and methods for producing them are described, for example, in Semple et al. (2010) Nat. Biotechnol. 28:172~176; Jayarama et al. (2012), Angew. Chem. Int. Ed., 51:8529~8533; and Maier et al. (2013) Molecular Therapy 21, 1570~1578. In some embodiments, the lipid nanoparticles contain cationic lipids. In some embodiments, the lipid nanoparticles contain sterols. In some embodiments, the lipid nanoparticles contain PEG-modified lipids. In some embodiments, the lipid nanoparticles contain neutral lipids. In some embodiments, the lipid nanoparticles contain cationic lipids, sterols, and PEG-modified lipids. In some embodiments, the lipid nanoparticles contain cationic lipids, neutral lipids, sterols, and PEG-modified lipids.

[0190] Examples of lipids include DLin-DMA, DLin-K-DMA, 98N12-5, C12-200, DLin-MC3-DMA, DLin-KC2-DMA, DODMA, PLGA, PEG, PEG-DMG, PEGylated lipids, and amino alcohol lipids. In another embodiment, the lipids may be cationic lipids, such as DLin-DMA, DLin-D-DMA, DLin-MC3-DMA, DLin-KC2-DMA, DODMA, and amino alcohol lipids. The amino alcohol cationic lipids may be lipids described in and / or produced by the methods described in U.S. Patent Application Publication US20130150625. For example, cationic lipids include 2-amino-3-[(9Z,12Z)-octadeca-9,12-diene-1-yloxy]-2-{[(9Z,2Z)-octadeca-9,12-diene-1-yloxy]methyl}propan-1-ol (compound 1 of US20130150625); 2-amino-3-[(9Z)-octadeca-9-en-1-yloxy]-2-{[(9Z)-octadeca-9-en-1-yloxy]methyl}propan-1-ol (compound 1 of US20130150625); 2-amino-3-[(9Z, [12Z)-octadeca-9,12-diene-1-yloxy]-2-[(octyloxy)methyl]propan-1-ol (compound 3 of US20130150625); and 2-(dimethylamino)-3-[(9Z,12Z)-octadeca-9,12-diene-1-yloxy]-2-{[(9Z,12Z)-octadeca-9,12-diene-1-yloxy]methyl}propan-1-ol (compound 4 of US20130150625); or any pharmaceutically acceptable salt or stereoisomer thereof.

[0191] In some embodiments, the lipid nanoparticles contain ionic cationic lipids, such as 2,2-dilinoleyl-4-dimethylaminoethyl-[1,3]-dioxolane (DLin-KC2-DMA), dilinoleyl-methyl-4-dimethylaminobutyrate (DLin-MC3-DMA), or di((Z)-nonano-2-en-1-yl)9-((4-(dimethylamino)butanoyl)oxy)heptadecanedioate (L319), and further contain neutral lipids, sterols, and molecules that can reduce particle aggregation, such as PEG or PEG-modified lipids.

[0192] In some embodiments, the lipid nanoparticle formulation essentially consists of (i) at least one lipid selected from the group consisting of 2,2-dilinoleyl-4-dimethylaminoethyl-[1,3]-dioxolane (DLin-KC2-DMA), dilinoleyl-methyl-4-dimethylaminobutyrate (DLin-MC3-DMA), and di((Z)-nonano-2-en-1-yl)9-((4-(dimethylamino)butanoyl)oxy)heptadecanedioate (L319), in a molar ratio of approximately 20-60% cationic lipids: 5-25% neutral lipids: 25-55% sterols; and 0.5-15% PEG lipids; (ii) neutral lipids selected from DSPC, DPPC, POPC, DOPE, and SM; (iii) sterols, e.g., cholesterol; and (iv) PEG lipids, e.g., PEG-DMG or PEG-cDMA. In some embodiments, the lipid nanoparticles contain 50% ionic cationic lipids, 10% neutral lipids, 38.5% sterols, and 1.5% PEG-modified lipids. In some embodiments, the neutral lipid is 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC), the sterol is cholesterol, and the PEG-modified lipid is 1,2-dimiristoyl racalycero-3-methoxypolyethylene glycol-2000 (PEG-DMG) or PEG-cDMA.

[0193] Seven major OspA serotypes have been recognized in European isolates (designated as serotypes 1-7; see Wilske et al., J. Clin. Microbiol. 31:340-50, 1993). In some aspects, OspA serotypes tend to correlate with species; for example, serotype 1 corresponds to B. burgdorferi ss, serotype 2 corresponds to B. afzelii, and serotypes 3-7 correspond to B. garinii. Serotype 4 is thought to correspond, in some aspects, alternatively to B. bavariensis (Margos et al., Appl. Environ. Microbiol. 75:5410-6, 2009).

[0194] In some embodiments, the OspA antigen is referred to in International Publication No. 2021207615, International Publication No. 2021205022, International Publication No. 2010132758, U.S. Patent No. 9,975,927, U.S. Patent No. 10,766,931, U.S. Patent No. 11,572,392, U.S. Patent No. 9,023,367, U.S. Patent No. 8,623,375, U.S. Patent No. 8,623,376, U.S. Patent No. 9,303,073, and U.S. Patent No. 9,334,31 It is any of the OspA antigens described in U.S. Patent No. 1, U.S. Patent No. 9,895,434, U.S. Patent No. 11,305,000, U.S. Patent No. 9,562,079, U.S. Patent No. 9,534,021, U.S. Patent No. 10,266,571, U.S. Patent No. 8,821,893, U.S. Patent No. 7,887,815, U.S. Patent No. 7,582,304, U.S. Patent No. 7,605,248, and U.S. Patent Publication No. 20090324638.

[0195] In some embodiments, the OspA antigen comprises one or more OspA antigens, for example, 1, 2, 3, 4, 5, or 6 OspA antigens. References to OspA antigens in this disclosure are understood to independently describe any one embodiment of the OspA antigen of one or more OspA antigens.

[0196] In some embodiments, the OspA antigen is a single OspA antigen. In some embodiments, the OspA antigen comprises two or more OspA antigens. For example, each OspA antigen has a non-identical amino acid sequence, which may be a duplicate sequence or a non-duplicate sequence overall.

[0197] In some embodiments, the OspA antigen is the OspA antigen of B. burgdorferi. In some embodiments, the OspA antigen is the OspA antigen of B. afzelii. In some embodiments, the OspA antigen is the OspA antigen of B. bavariensis. In some embodiments, the OspA antigen is the OspA antigen of B. garinii. In some embodiments, the OspA antigen is the OspA antigen of B. mayonii. In some embodiments, the OspA antigen is the OspA antigen of B. lusitaniae. In some embodiments, the OspA antigen is the OspA antigen of B. bissettii. In some embodiments, the OspA antigen is the OspA antigen of B. spielmanii. In some embodiments, the OspA antigen is the OspA antigen of B. valasiana.

[0198] In some embodiments, each of the different OspA antigens is an OspA antigen derived from a different Borrelia species. In some embodiments, the OspA antigen comprises a first OspA antigen and a second OspA antigen. In some embodiments, the first OspA antigen and the second OspA antigen have non-identical amino acid sequences. In some embodiments, two or more OspA antigens further comprise a third OspA antigen. In some embodiments, the first OspA antigen, the second OspA antigen, and the third OspA antigen have non-identical amino acid sequences.

[0199] In some embodiments, the OspA antigen is a naturally occurring OspA protein or a fragment thereof. In some embodiments, the OspA antigen is a recombinant OspA protein.

[0200] In some embodiments, the OspA antigen contains one or more epitopes derived from the OspA protein. In some embodiments, one or more epitopes include at least one linear epitope. In some embodiments, one or more epitopes are linear epitopes. In some embodiments, one or more epitopes include at least one structural epitope. In some embodiments, one or more epitopes are structural epitopes.

[0201] In some embodiments, one or more epitopes contain at least two, three, four, five, or six epitopes derived from one or more OspA proteins. In some embodiments, one or more epitopes are derived from the same OspA protein. In some embodiments, one or more epitopes are derived from the same OspA serotype. In some embodiments, one or more epitopes are derived from multiple OspA proteins. In some embodiments, one or more epitopes contain at least two, three, four, five, or six epitopes derived from at least two, three, four, five, or six OspA proteins. In some embodiments, one or more epitopes are derived from multiple OspA serotypes. In some embodiments, one or more epitopes contain at least two, three, four, five, or six epitopes derived from at least two, three, four, five, or six OspA serotypes.

[0202] In some embodiments, one or more epitopes comprise OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, one or more epitopes are independently selected from OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, one or more epitopes comprise any of the linear OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, one or more epitopes comprise any of the structural OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, one or more epitopes comprise linear and structural OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 82.

[0203] In some embodiments, one or more epitopes comprise OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 62. In some embodiments, one or more epitopes are independently selected from OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 62. In some embodiments, one or more epitopes comprise any of the linear OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50 to 62. In some embodiments, one or more epitopes comprise any of the structural OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50 to 62. In some embodiments, one or more epitopes comprise linear and structural OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 62.

[0204] In some embodiments, one or more epitopes comprise OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, one or more epitopes are independently selected from OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, one or more epitopes comprise any of the linear OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, one or more epitopes comprise any of the structural OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, one or more epitopes comprise linear and structural OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 63 to 82.

[0205] In some embodiments, one or more epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49. In some embodiments, one or more epitopes are independently selected from any of the linear epitopes whose amino acid sequences are shown in SEQ ID NOs: 4 to 49.

[0206] In some embodiments, the OspA antigen is an OspA protein containing one or more three-dimensional epitopes. In some embodiments, the OspA antigen contains one or more three-dimensional epitopes or has one of the amino acid sequences shown in SEQ ID NOs. 50 to 82, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with it. In some embodiments, the OspA antigen contains one or more structural epitopes or has any of the amino acid sequences shown in SEQ ID NOs. 50 to 82, having at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 of its adjacent amino acids. In some embodiments, the OspA antigen contains one or more structural epitopes and has any of the amino acid sequences shown in SEQ ID NOs. 50 to 82.

[0207] In some embodiments, the OspA antigen is an OspA protein containing one or more three-dimensional epitopes. In some embodiments, the OspA antigen contains one or more three-dimensional epitopes, or has one of the amino acid sequences shown in SEQ ID NOs. 50 to 62, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with it. In some embodiments, the OspA antigen contains one or more structural epitopes, or has any of the amino acid sequences shown in SEQ ID NOs. 50-62, having at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 of its adjacent amino acids. In some embodiments, the OspA antigen contains one or more structural epitopes, or has any of the amino acid sequences shown in SEQ ID NOs. 50-62.

[0208] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any of SEQ ID NOs.

[0209] In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in any of SEQ ID NOs. 50-62.

[0210] In some embodiments, the OspA antigen comprises the amino acid sequence shown in any of SEQ ID NOs. 50-62.

[0211] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 50. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 50. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 50.

[0212] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 51. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 51. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 51.

[0213] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 52. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 52. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 52.

[0214] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 53. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 53. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 53.

[0215] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 54. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 54. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 54.

[0216] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 55. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 55. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 55.

[0217] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 56. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 56. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 56.

[0218] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 57. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 57. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 57.

[0219] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 58. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 58. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 58.

[0220] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 59. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 59. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 59.

[0221] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 60. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 60. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 60.

[0222] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 61. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 61. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 61.

[0223] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 62. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 62. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 62.

[0224] In some embodiments, the OspA antigen is an OspA chimeric antigen. In some embodiments, the OspA chimeric antigen contains multiple epitopes derived from one or more OspA proteins. In some embodiments, the OspA antigen is an OspA chimeritope.

[0225] In some embodiments, the numerous epitopes are derived from the same OspA protein. In some embodiments, the numerous epitopes are derived from the same OspA serotype. In some embodiments, the numerous epitopes are derived from multiple OspA proteins. In some embodiments, the numerous epitopes contain at least 2, 3, 4, or 5 epitopes derived from at least 2, 3, 4, 5, or 6 OspA proteins. In some embodiments, the numerous epitopes are derived from multiple OspA serotypes. In some embodiments, the numerous epitopes contain at least 2, 3, 4, 5, or 6 epitopes derived from at least 2, 3, 4, 5, or 6 OspA serotypes.

[0226] In some embodiments, the numerous epitopes include OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, the numerous epitopes are independently selected from OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, the numerous epitopes include any of the linear OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, the numerous epitopes include any of the structural OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50 to 82. In some embodiments, the numerous epitopes include linear and structural OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50 to 82.

[0227] In some embodiments, the numerous epitopes include OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50-62. In some embodiments, the numerous epitopes are independently selected from OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50-62. In some embodiments, the numerous epitopes include any of the linear OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50-62. In some embodiments, the numerous epitopes include any of the structural OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 50-62. In some embodiments, the numerous epitopes include linear and structural OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 50-62.

[0228] In some embodiments, the numerous epitopes include OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, the numerous epitopes are independently selected from OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, the numerous epitopes include any of the linear OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, the numerous epitopes include any of the structural OspA epitopes contained in the amino acid sequences shown in SEQ ID NOs. 63 to 82. In some embodiments, the numerous epitopes include linear and structural OspA epitopes contained in any of the amino acid sequences shown in SEQ ID NOs. 63 to 82.

[0229] In some embodiments, the numerous epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4-49. In some embodiments, the numerous epitopes are independently selected from any of the linear epitopes whose amino acid sequences are shown in SEQ ID NOs: 4-49.

[0230] In some embodiments, the OspA antigen is an OspA protein containing one or more linear epitopes. In some embodiments, the OspA antigen contains one or more linear epitopes or has one of the amino acid sequences shown in SEQ ID NOs. 50 to 82, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the OspA antigen. In some embodiments, the OspA antigen contains one or more linear epitopes or has any of the amino acid sequences shown in SEQ ID NOs. 50 to 82, having at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 of its adjacent amino acids. In some embodiments, the OspA antigen contains one or more linear epitopes and has any of the amino acid sequences shown in SEQ ID NOs. 50 to 82.

[0231] In some embodiments, the OspA antigen is an OspA protein containing one or more linear epitopes. In some embodiments, the OspA antigen contains one or more linear epitopes or has one of the amino acid sequences shown in SEQ ID NOs. 63-82, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the OspA antigen. In some embodiments, the OspA antigen contains one or more linear epitopes or has any of the amino acid sequences shown in SEQ ID NOs. 63-82, having at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 of its adjacent amino acids. In some embodiments, the OspA antigen contains one or more linear epitopes and has any of the amino acid sequences shown in SEQ ID NOs. 63-82.

[0232] In some embodiments, the OspA antigen is an OspA chimeric antigen containing a number of epitopes derived from one or more OspA proteins. In some embodiments, the number of epitopes are linear epitopes. In some embodiments, the OspA antigen is an OspA chimeritope. In some embodiments, the OspA antigen is an OspA chimeric antigen containing a number of linear epitopes, or has one of the amino acid sequences shown in SEQ ID NOs. 50 to 82, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with it. In some embodiments, the OspA antigen is an OspA chimeric antigen containing a large number of linear epitopes, or has any of the amino acid sequences shown in SEQ ID NOs. 50-82, having at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 of its adjacent amino acids. In some embodiments, the OspA antigen is an OspA chimeric antigen containing a large number of linear epitopes, and has any of the amino acid sequences shown in SEQ ID NOs. 50-82.

[0233] In some embodiments, the OspA antigen is an OspA chimeric antigen containing a number of epitopes derived from one or more OspA proteins. In some embodiments, the number of epitopes are linear epitopes. In some embodiments, the OspA antigen is an OspA chimeritope. In some embodiments, the OspA antigen is an OspA chimeric antigen containing a number of linear epitopes, or has one of the amino acid sequences shown in SEQ ID NOs. 63-82, having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with it. In some embodiments, the OspA antigen is an OspA chimeric antigen containing numerous linear epitopes, or has any of the amino acid sequences shown in SEQ ID NOs. 63-82, having at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 of its adjacent amino acids. In some embodiments, the OspA antigen is an OspA chimeric antigen containing numerous linear epitopes, having any of the amino acid sequences shown in SEQ ID NOs. 63-82.

[0234] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any of SEQ ID NOs: 63 to 82.

[0235] In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in any of SEQ ID NOs. 63-82.

[0236] In some embodiments, the OspA antigen comprises an amino acid sequence shown in any of SEQ ID NOs. 63-82.

[0237] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 63. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 63. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 63.

[0238] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 64. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 64. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 64.

[0239] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 65. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 65. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 65.

[0240] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 66. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 66. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 66.

[0241] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 67. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 67. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 67.

[0242] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 68. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 68. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 68.

[0243] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 69. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 69. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 69.

[0244] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 70. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 70. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 70.

[0245] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 71. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 71. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 71.

[0246] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 72. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 72. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 72.

[0247] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 73. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 73. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 73.

[0248] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 74. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 74. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 74.

[0249] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 75. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 75. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 75.

[0250] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 76. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 76. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 76.

[0251] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 77. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 77. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 77.

[0252] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 78. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 78. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 78.

[0253] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 79.

[0254] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 80.

[0255] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 81.

[0256] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 82. In some embodiments, the OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in SEQ ID NO: 82. In some embodiments, the OspA antigen includes the amino acid sequence shown in SEQ ID NO: 82.

[0257] In some embodiments, the OspA antigen has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any of SEQ ID NOs. In some embodiments, the OspA antigen contains at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids to the amino acid sequence shown in any of SEQ ID NOs. In some embodiments, the OspA antigen contains the amino acid sequence shown in any of SEQ ID NOs.

[0258] In some embodiments, the immunogenic composition comprises a first OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, and a second OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the first OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the first OspA antigen includes the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the second OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the second OspA antigen includes the amino acid sequence shown in SEQ ID NO: 80.

[0259] In some embodiments, the immunogenic composition comprises a first OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 89, and a second OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the first OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the first OspA antigen includes the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the second OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the second OspA antigen includes the amino acid sequence shown in SEQ ID NO: 81.

[0260] In some embodiments, the immunogenic composition comprises a first OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, and a second OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the first OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the first OspA antigen includes the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the second OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the second OspA antigen includes the amino acid sequence shown in SEQ ID NO: 81.

[0261] In some embodiments, the immunogenic composition is a first OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, and at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87% sequence identity with the amino acid sequence shown in SEQ ID NO: 80. The formulation includes a second OspA antigen having 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity, and a third OspA antigen having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. In some embodiments, the first OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the first OspA antigen includes the amino acid sequence shown in SEQ ID NO: 79. In some embodiments, the second OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the second OspA antigen includes the amino acid sequence shown in SEQ ID NO: 80. In some embodiments, the third OspA antigen includes at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids in the amino acid sequence shown in SEQ ID NO: 81.

[0262] In some embodiments, the OspA antigen is not lipid-modified. In some embodiments, the first OspA antigen is not lipid-modified. In some embodiments, the second OspA antigen is not lipid-modified. In some embodiments, the third OspA antigen is not lipid-modified.

[0263] In some embodiments, the OspA antigen is lipid-based. In some embodiments, the first OspA antigen is lipid-based. In some embodiments, the second OspA antigen is lipid-based. In some embodiments, the third OspA antigen is lipid-based. In some embodiments, the first and second OspA antigens are lipid-based. In some embodiments, the first and third OspA antigens are lipid-based. In some embodiments, the second and third OspA antigens are lipid-based. In some embodiments, the first, second, and third OspA antigens are lipid-based.

[0264] Lipidization can confer adjuvant properties or stabilize the OspA antigen. Protein lipidization can occur using many different known methods (see, for example, U.S. Patent No. 8,986,704, U.S. Patent No. 11,305,000, Bouchon et al., Anal Biochem, 1997, Hantke et al., Eur. J. Biochem, 1973, and Erdile et al., Infect Immun, 1993). In some embodiments, lipidization occurs through a chemical lipidization process. In some embodiments, lipidization occurs using an E. coli expression system. In some embodiments, lipidization is supported by the presence of a lipidization signal peptide sequence, e.g., one present on the OspA antigen. In some embodiments, the E. coli expression system is BL21 DE3. In some embodiments, the E. coli expression system uses one or all of the enzymes diacylglyceryltransferase, signal peptidase II, and transacylase. In some embodiments, the first step involves the transfer of a diacylglyceride to the sulfhydryl group of the cysteine ​​of an unmodified prolipoprotein, followed by cleavage of the signal peptide by signal peptidase II, and finally acylation of the α-amino group of the N-terminal cysteine ​​of the apolipoprotein. The result is the replacement of a glycerol group with a certain lipid and two further lipids on the N-terminal cysteine ​​residue of the polypeptide. The lipidation signal sequence cleaved during lipidation is not present in the final polypeptide sequence.

[0265] In some embodiments, the lipidized OspA antigen is purified. In some embodiments, purification is performed using surfactant extraction and / or ion exchange chromatography. In some embodiments, purification is performed using ion exchange chromatography and Triton X100 extraction. In some embodiments, the purification process reduces the endotoxin level. In some embodiments, purification reduces the endotoxin level to below 10 EU / mg.

[0266] In some embodiments, the OspA antigen is lipidated by the addition or presence of a lipid moiety to the OspA antigen. In some embodiments, the lipid moiety, such as a fatty acid, is added to the OspA antigen, for example, at the C-terminus and / or N-terminus. In some embodiments, glycerol is added to the OspA antigen, for example, at the C-terminus and / or N-terminus. In some embodiments, the lipid moiety, such as a fatty acid, is added to the OspA antigen at the N-terminus. In some embodiments, the lipid molecule, such as a fatty acid, and glycerol are added to the OspA antigen at the N-terminus. In some embodiments, the lipidated OspA antigen has three fatty acid groups and glycerol added to the N-terminus of the lipidated OspA antigen.

[0267] In some embodiments, the lipid moiety is one to three lipids, such as C 14~20 alkyl and / or C 14~20 alkenyl. In some embodiments, the lipid moiety includes, as major components, saturated fatty acids (C 14 to C 18 range), unsaturated fatty acids (C 16 / C<00****>), and / or palmitate (C16:0). In some embodiments, the lipid moiety is attached to glycerol. In some embodiments, the lipid moiety is attached to the N-terminal cysteine of the polypeptide of the OspA antigen. In some embodiments, the N-terminal cysteine is present on the signal peptide sequence. In some embodiments, the lipid moiety is attached at the amino-terminal cysteine residue of the signal peptide sequence. In some embodiments, the lipid moiety is attached via the sulfhydryl group of the cysteine residue of the signal peptide sequence. In some embodiments, the N-terminal cysteine can be lipidated at both the amino group (-HN2) and the sulfhydryl group (-SH). In some embodiments, the N-terminal cysteine is lipidated to form approximately one to three fatty acid modifications. In some embodiments, the lipid moiety attached to the N-terminal cysteine is S-[2,3-bis(palmitoyloxy)-(2R,S)-propyl]-N-palmitoylcysteine.

[0268] In some embodiments, the OspA antigen may further comprise a lipid-containing signal peptide sequence. Lipid-containing signal peptide sequences are known in the Art (see, for example, U.S. Patent Nos. 8,986,704 and 11,305,000). Exemplary lipid-containing signal peptide sequences include, but are not limited to, those shown in SEQ ID NOs. 83-85. In some embodiments, the OspA antigen further comprises the lipid-containing signal peptide sequence shown in SEQ ID NO. 83. In some embodiments, the OspA antigen further comprises the lipid-containing signal peptide sequence shown in SEQ ID NO. 84. In some embodiments, the OspA antigen further comprises the lipid-containing signal peptide sequence shown in SEQ ID NO. 85. In some embodiments, the OspA antigen comprises an OspB reader sequence. In some embodiments, the OspB reader sequence comprises the sequence shown in SEQ ID NO. 83.

[0269] The lipid-conjugated signal peptide sequence is cleaved during processing, and therefore, the lipid-conjugated OspA antigen does not need to contain the lipid-conjugated signal peptide sequence in the mature lipid-conjugated protein.

[0270] C. Additional Components The immunogenic compositions of this disclosure may comprise one or more additional components, such as one or more excipients, one or more stabilizers, one or more preservatives, and / or additional antigens.

[0271] Pharmaceutically acceptable excipients for this immunogenic composition include solvents, bulking agents, buffers, osmotic regulators, preservatives, surfactants, and emulsifiers (Pramanick et al., Pharma Times, 45:65-77, 2013). In some embodiments, the immunogenic composition contains an excipient that functions as one or more of a solvent, bulking agent, buffer, and osmotic regulator (for example, sodium chloride in physiological saline can function as both an aqueous vehicle and an osmotic regulator). In some embodiments, the immunogenic composition contains an excipient that functions as one or both a surfactant and an emulsifier (for example, the nonionic surfactant polysorbate 80).

[0272] In some embodiments, the immunogenic composition contains an aqueous vehicle as a solvent. Suitable vehicles include sterile water, physiological saline solution, phosphate-buffered saline, and Ringer's solution. In some embodiments, the immunogenic composition is isotonic.

[0273] In some embodiments, the immunogenic composition contains a buffer. The buffer controls the pH to inhibit the degradation of the active agent during processing, storage, and, optionally, reconstitution. Preferred buffers include acetates, citrates, phosphates, sulfates, or salts containing Tris. In some embodiments, the buffer is not a phosphate-containing buffer. In certain other embodiments, the buffer is a Tris buffer. Other preferred buffers include amino acids, such as arginine, glycine, histidine, and lysine. The buffer may further contain hydrochloric acid or sodium hydroxide. In some embodiments, the buffer maintains the pH of the immunogenic composition within the range of 6 to 9. In some embodiments, the pH is higher than 6, 7, or 8 (lower limit). In some embodiments, the pH is less than 9, 8, or 7 (upper limit). That is, the pH is in the range of about 6 to 9, with the lower limit being less than the upper limit. In some embodiments, the pH is about 6.5, about 7.0, or about 7.5.

[0274] In some embodiments, the immunogenic composition contains an osmotic regulator. Suitable osmotic regulators include dextrose, glycerol, sodium chloride, glycerin, and mannitol.

[0275] In some embodiments, the immunogenic composition contains a bulking agent. The bulking agent is particularly useful when the immunogenic composition is lyophilized before administration. In some embodiments, the bulking agent is a protective agent that aids in stabilizing the active agent and preventing its degradation during freezing or spray drying and / or during storage. Suitable bulking agents include sugars (e.g., monosaccharides, disaccharides, and polysaccharides), such as sucrose, lactose, trehalose, mannitol, sorbitol, glucose, and raffinose.

[0276] In some embodiments, the immunogenic composition contains a stabilizer. Suitable stabilizers include sugars, such as sucrose and lactose, amino acids, such as the sodium salt of glycine or glutamic acid, and proteins, such as human serum albumin or gelatin.

[0277] In some embodiments, the immunogenic composition contains a preservative. Suitable preservatives include antioxidants and antibacterial agents. In some embodiments, the immunogenic composition is prepared under aseptic conditions and is in a single-use container, and thus the inclusion of a preservative is not required.

[0278] II. Product The present disclosure also provides vials, syringes (e.g., prefilled syringes), and kits containing any of the immunogenic compositions described herein, e.g., in Section I. In some embodiments, the immunogenic composition is in liquid form. In some embodiments, the immunogenic composition is in lyophilized form. In some embodiments, the vials, syringes, or kits provided contain a single-dose of the immunogenic composition, e.g., a dose of 0.3 mL to 1.5 mL.

[0279] This disclosure also provides a kit containing the first composition and the second composition. In some embodiments, the first composition contains an adjuvant, which may be any adjuvant described herein, for example, in Section IA. In some embodiments, the second composition contains a Borrelia cell surface protein A (OspA) antigen, which may be any one described herein, for example, in Section IB. In some embodiments, the second composition is in liquid form. In some embodiments, the second composition is in lyophilized form.

[0280] In some embodiments, the kit further includes instructions for combining the first and second compositions to prepare any of the immunogenic compositions described herein, for example, in Section I. In some embodiments, the instructions are for preparing a single dose of the immunogenic composition. In some embodiments, the kit is for preparing a single dose of the immunogenic composition, for example, a dose of 0.3 mL to 1.5 mL.

[0281] In some embodiments, combining the first composition with the second composition includes mixing the first and second compositions. In some embodiments, combining the first composition with the second composition includes mixing the first and second compositions in a single vial. In some embodiments, combining the first composition with the second composition includes mixing the first and second compositions in a single vial by gentle inversion. In some embodiments, the mixing is done at room temperature for up to about 1 hour.

[0282] In some embodiments, the first composition contains a second adjuvant, which may be any of those described herein, for example, in Section IA.

[0283] In some embodiments, the kit further includes instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject. In some embodiments, administration follows any of the methods described herein, e.g., any of those described in Section III. In some embodiments, the subject is a human subject. In some embodiments, the human subject is at least 18 years old. In some embodiments, the human subject is about 18 to about 70 years old. In some embodiments, the human subject is about 18 to about 55 years old. In some embodiments, the human subject is under 18 years old. In some embodiments, the human subject is about 2 to about 5 years old. In some embodiments, the human subject is about 5 to about 18 years old.

[0284] In some embodiments, the kit contains a suitably packaged immunogenic composition. In some embodiments, the kit contains suitably packaged i) a first composition and ii) a second composition such that the first and second compositions are combined to form an immunogenic composition. For example, if the immunogenic composition is a lyophilized powder, a vial with an elastic stopper can be used so that the powder can be easily resuspended by injecting a liquid (e.g., sterile water, saline, etc.) through the elastic stopper. In some embodiments, the kit contains an administration device (e.g., a syringe). In some embodiments, the syringe is pre-filled with the immunogenic composition. In some embodiments, administration is by intramuscular injection. Instructions for the use of the immunogenic composition generally include information such as dosage, schedule, and route of administration for the intended method of use.

[0285] III. How to Use This disclosure also relates to a method for stimulating an immune response to Borrelia in a subject, comprising administering an effective amount of any of the immunogenic compositions described herein, for example, in Section I, to the subject. In some embodiments, the antigen and adjuvant are present in the immunogenic composition in an effective amount to stimulate an immune response to Borrelia in the subject.

[0286] This disclosure also relates to a method for protecting a subject from infection by Borrelia, comprising administering an effective amount of one of the immunogenic compositions described herein, for example, in Section I, to the subject. In some embodiments, the antigen and adjuvant are present in the immunogenic composition in an amount effective to stimulate an immune response to Borrelia in the subject.

[0287] This disclosure also relates to a method for preventing a subject from contracting Lyme disease, the method comprising administering an effective amount of one of the immunogenic compositions described herein, for example, in Section I, to the subject. In some embodiments, the antigen and adjuvant are present in the immunogenic composition in an amount effective to stimulate an immune response to Borrelia in the subject.

[0288] In some embodiments, the subjects are human subjects. In some embodiments, the human subjects are at least 18 years old. In some embodiments, the human subjects are approximately 18 to 70 years old. In some embodiments, the human subjects are approximately 18 to 55 years old. In some embodiments, the human subjects are under 18 years old. In some embodiments, the human subjects are approximately 2 to 5 years old. In some embodiments, the human subjects are approximately 5 to 18 years old.

[0289] In some embodiments, the methods of the present disclosure are suitable for increasing the immune response to Borrelia in a subject compared to the baseline immune response to Borrelia in the subject before administration of the immunogenic composition. Stimulating the immune response, meaning increasing the immune response, may result from inducing a new immune response (e.g., as a result of an initial vaccination regimen) or enhancing an existing immune response (e.g., as a result of a booster vaccination regimen). In some embodiments, stimulating the immune response includes one or more of the following: stimulating cytokine production; stimulating B lymphocyte proliferation; stimulating antibody production; stimulating interferon pathway-related gene expression; stimulating chemoattractant-related gene expression; and stimulating plasmacytoid dendritic cell maturation. In some embodiments, the immune response prevents the subject from becoming infected with Borrelia. In some embodiments, the immune response prevents the subject from contracting Lyme disease.

[0290] In some embodiments, a first dose and a second dose of the immunogenic composition are administered to the subject. In some embodiments, the second dose of the immunogenic composition is administered approximately 2 to 12 weeks after the administration of the first dose of the immunogenic composition. In some embodiments, the second dose of the immunogenic composition is administered approximately 2 to 10 weeks after the administration of the first dose of the immunogenic composition. In some embodiments, the second dose is administered approximately 2 weeks to 2 months after the administration of the first dose. In some embodiments, the second dose is administered approximately 2 to 6 weeks after the administration of the first dose. In some embodiments, the second dose is administered approximately 2 to 4 weeks after the administration of the first dose. In some embodiments, the second dose is administered approximately 3 weeks after the administration of the first dose.

[0291] In some embodiments, the second dose is administered approximately 2 to 6 weeks after the administration of the first dose. In some embodiments, the second dose is administered approximately 3 to 5 weeks after the administration of the first dose. In some embodiments, the second dose is administered approximately 1 month after the administration of the first dose.

[0292] In some embodiments, the second dose is administered approximately 6 to 10 weeks after the administration of the first dose. In some embodiments, the second dose is administered approximately 7 to 9 weeks after the administration of the first dose. In some embodiments, the second dose is administered approximately 2 months after the administration of the first dose.

[0293] In some embodiments, a third dose of the immunogenic composition is administered to the subject. In some embodiments, the third dose is administered approximately 4 to 8 months after the first dose. In some embodiments, the third dose is administered approximately 5 to 7 months after the first dose. In some embodiments, the third dose is administered approximately 22 to 26 weeks after the first dose. In some embodiments, the third dose is administered approximately 6 months after the first dose.

[0294] In some embodiments, a booster dose of the immunogenic composition is administered to the subject. In some embodiments, the booster dose is administered at least one year after the first dose. In some embodiments, the booster dose is administered at least two years after the first dose.

[0295] In some embodiments, the booster dose is administered approximately 10 months to 5 years after the first dose. In some embodiments, the booster dose is administered approximately 1 year to 5 years after the first dose. In some embodiments, the booster dose is administered approximately 20 months to 5 years after the first dose. In some embodiments, the booster dose is administered approximately 2 years to 5 years after the first dose. In some embodiments, the booster dose is administered approximately 2 years to 4 years after the first dose. In some embodiments, the booster dose is administered approximately 2 years to 3 years after the first dose.

[0296] In some embodiments, the booster dose is administered approximately 10 to 14 months after the first dose. In some embodiments, the booster dose is administered approximately 1 year after the first dose.

[0297] In some embodiments, the booster dose is administered about 20 to about 28 months after the first dose. In some embodiments, the booster dose is administered about 2 years after the first dose.

[0298] In some embodiments, multiple booster doses are administered to the subject. In some embodiments, multiple annual booster doses are administered to the subject. In some embodiments, the first booster dose is administered at least about 1 year or at least about 2 years after administration of the first dose. In some aspects of any embodiment, the booster dose is administered annually.

[0299] In some embodiments, the first and second doses of the immunogenic composition are administered to the subject, wherein the second dose is administered about 1 month after the first dose.

[0300] In some embodiments, the first, second, and booster doses of the immunogenic composition are administered to the subject, wherein the second dose is administered about 1 month after the first dose and the booster dose is administered at least 2 years after the first dose. In some embodiments, the first, second, and booster doses of the immunogenic composition are administered to the subject, wherein the second dose is administered about 1 month after the first dose and the booster dose is administered about 2 years after the first dose.

[0301] In some embodiments, the first and second doses of the immunogenic composition are administered to the subject, wherein the second dose is administered about 2 months after the first dose.

[0302] In some embodiments, first, second, and booster doses of the immunogenic composition are administered to the subject, where the second dose is administered about two months after the first dose, and the booster dose is administered at least one year after the first dose.

[0303] In some embodiments, first, second, and booster doses of the immunogenic composition are administered to the subject, where the second dose is administered about two months after the first dose, and the booster dose is administered at least two years after the first dose.

[0304] In some embodiments, first, second, and third doses of the immunogenic composition are administered to the subject, where the second dose is administered approximately two months after the first dose, and the third dose is administered approximately six months after the first dose.

[0305] In some embodiments, first, second, third, and booster doses of the immunogenic composition are administered to the subject, where the second dose is administered about two months after the first dose, the third dose is administered about six months after the first dose, and the booster dose is administered at least two years after the first dose.

[0306] In some embodiments, first, second, third, and multiple booster doses of the immunogenic composition are administered to the subject, where the second dose is administered about two months after the first dose, the third dose is administered about six months after the first dose, the first booster dose is administered about one year after the first dose, and the booster doses are administered annually.

[0307] In some embodiments, the immunogenic composition is administered by intramuscular injection. In some embodiments, the injection site is the arm of the subject. In some embodiments, the injection site is the deltoid muscle of the subject.

[0308] Exemplary Embodiments In particular, the embodiments provided are as follows: Embodiment 1. An immunogenic composition comprising (i) a Borrelia cell surface protein A (OspA) antigen or an RNA polynucleotide encoding an OspA antigen, and (ii) an adjuvant selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsions containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters. Embodiment 2. An immunogenic composition comprising (i) a Borrelia cell surface protein A (OspA) antigen, and (ii) an adjuvant selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsions containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters. Embodiment 3. An immunogenic composition according to Embodiment 1 or Embodiment 2, which does not contain an RNA polynucleotide encoding a non-OspA Borrelia cell surface protein antigen. Embodiment 4. An immunogenic composition according to any one of Embodiments 1 to 3, which does not contain a non-OspA Borrelia cell surface protein antigen. Embodiment 5. The immunogenic composition according to any one of Embodiments 1 to 4, wherein the adjuvant is an aluminum salt adjuvant. Embodiment 6. The immunogenic composition according to Embodiment 5, further comprising a second adjuvant. Embodiment 7. The immunogenic composition according to Embodiment 6, wherein the second adjuvant is selected from a TLR9 agonist; a liposome-based adjuvant comprising lipopolysaccharides and saponins; and an oil-in-water emulsion comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters. Embodiment 8. The immunogenic composition according to any one of Embodiments 1 to 4, wherein the adjuvant is a TLR9 agonist. Embodiment 9. The immunogenic composition according to Embodiment 8, further comprising a second adjuvant. Embodiment 10. The immunogenic composition according to Embodiment 9, wherein the second adjuvant is selected from aluminum salt adjuvants; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters. Embodiment 11. The immunogenic composition according to any one of Embodiments 1 to 4, wherein the adjuvant is a liposome-based adjuvant comprising a lipopolysaccharide and a saponin. Embodiment 12. The immunogenic composition according to Embodiment 11, further comprising a second adjuvant. Embodiment 13. The immunogenic composition according to Embodiment 12, wherein the second adjuvant is selected from aluminum salt adjuvants; TLR9 agonists; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters. Embodiment 14. The immunogenic composition according to any one of Embodiments 1 to 4, wherein the adjuvant is an oil-in-water emulsion-based adjuvant comprising squalene, sorbitan ester, and polyoxyethylene sorbitan ester. Embodiment 15. The immunogenic composition according to Embodiment 14, further comprising a second adjuvant. Embodiment 16. The immunogenic composition according to Embodiment 15, wherein the second adjuvant is selected from aluminum salt adjuvants; TLR9 agonists; and liposome-based adjuvants comprising lipopolysaccharides and saponins. Embodiment 17. An immunogenic composition according to any one of Embodiments 1-4, 7-10, 13, and 16, wherein the TLR9 agonist is an oligonucleotide containing a non-methylated cytidine-phospho-guanosine (CpG) motif. Embodiment 18. The immunogenic composition according to Embodiment 17, wherein the oligonucleotide is 10 to 35 nucleotides in length. Embodiment 19. An immunogenic composition according to Embodiment 17 or Embodiment 18, wherein the oligonucleotide comprises the sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2). Embodiment 20. An immunogenic composition according to any one of Embodiments 17 to 19, wherein the oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1). Embodiment 21. An immunogenic composition comprising a Borrelia cell surface protein A (OspA) antigen, and a Toll-like receptor 9 (TLR9) agonist oligonucleotide 10 to 35 nucleotides in length, comprising the sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2) and a non-methylated cytidine-phospho-guanosine (CpG) motif. Embodiment 22. The immunogenic composition according to Embodiment 21, wherein the oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1). Embodiment 23. An immunogenic composition comprising an oligonucleotide comprising Borrelia cell surface protein A (OspA) antigen and Toll-like receptor 9 (TLR9) agonist oligonucleotide 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1), wherein the oligonucleotide comprises a non-methylated cytidine-phospho-guanosine (CpG) motif. Embodiment 24. An immunogenic composition according to any one of Embodiments 21 to 23, further comprising a second adjuvant. Embodiment 25. The immunogenic composition according to Embodiment 24, wherein the second adjuvant is selected from aluminum salt adjuvants; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters. Embodiment 26. The immunogenic composition according to any one of Embodiments 9, 10, 12, 13, 15-20, 24, and 25, wherein the second adjuvant is an aluminum salt adjuvant. Embodiment 27. An immunogenic composition comprising (i) Borrelia cell surface protein A (OspA) antigen, (ii) Toll-like receptor 9 (TLR9) agonist oligonucleotide 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1), wherein the oligonucleotide comprises an unmethylated cytidine-phospho-guanosine (CpG) motif, and (iii) an aluminum salt adjuvant. Embodiment 28. An immunogenic composition according to any one of Embodiments 1-7, 10, 13, 16-20, and 25-27, wherein the aluminum salt adjuvant comprises one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate. Embodiment 29. An immunogenic composition according to any one of Embodiments 1-7, 10, 13, 16-20, and 25-28, wherein the aluminum salt adjuvant comprises aluminum hydroxide. Embodiment 30. Approximately 0.25 to 1.25 mg of Al 3+ Approximately 0.25 mg to 0.50 mg of Al 3+ , or approximately 0.30 mg to approximately 0.40 mg of Al 3+ An immunogenic composition according to any one of Embodiments 1 to 29, comprising [the specified element]. Embodiment 31. The immunogenic composition according to any one of Embodiments 17 to 30, wherein the oligonucleotide is a single-chain oligodeoxynucleotide. Embodiment 32. The immunogenic composition according to any one of Embodiments 17 to 30, wherein the oligonucleotide is a complete RNA. Embodiment 33. An immunogenic composition according to any one of Embodiments 17 to 30, wherein the oligonucleotide is an RNA / DNA chimera. Embodiment 34. An immunogenic composition according to any one of Embodiments 17 to 33, wherein the oligonucleotide comprises at least one phosphorothioate linkage. Embodiment 35. An immunogenic composition according to any one of Embodiments 17 to 34, wherein the oligonucleotide comprises only phosphorothioate linkages. Embodiment 36. An immunogenic composition according to any one of Embodiments 17 to 34, wherein the oligonucleotide comprises a combination of one or more phosphodiester linkages and one or more phosphorothioate linkages. Embodiment 37. An immunogenic composition according to any one of Embodiments 1-4, 7-10, 13, and 16-36, comprising approximately 375 μg to approximately 6000 μg of TLR9 agonist or approximately 750 μg to approximately 3000 μg of TLR9 agonist. Embodiment 38. An immunogenic composition according to any one of Embodiments 1-4, 7-10, 13, and 16-36, comprising approximately 375 μg, approximately 750 μg, approximately 1000 μg, approximately 1500 μg, approximately 3000 μg, or approximately 6000 μg of a TLR9 agonist. Embodiment 39. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-38, wherein the lipopolysaccharide is lipid A or a derivative thereof. Embodiment 40. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-39, wherein the lipopolysaccharide is monophosphoryl lipid A or a derivative thereof. Embodiment 41. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-40, wherein the lipopolysaccharide is 3-O-desacyl-4'-monophosphoryl lipid A. Embodiment 42. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-40, wherein the lipopolysaccharide is a 3-deacyl-phosphorylated hexaacyl disaccharide. Embodiment 43. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-42, comprising approximately 1 μg to approximately 100 μg, approximately 10 μg to approximately 50 μg, approximately 20 μg to approximately 30 μg, approximately 21 μg to approximately 29 μg, approximately 22 μg to approximately 28 μg, approximately 23 μg to approximately 27 μg, or approximately 24 μg to approximately 26 μg of lipopolysaccharide. Embodiment 44. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-42, comprising approximately 1 μg to approximately 30 μg, approximately 5 μg to approximately 15 μg, approximately 6 μg to approximately 14 μg, approximately 7 μg to approximately 13 μg, approximately 8 μg to approximately 12 μg, or approximately 9 μg to approximately 11 μg of lipopolysaccharide. Embodiment 45. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-42, comprising approximately 1 μg to approximately 9 μg, approximately 2 μg to approximately 8 μg, approximately 3 μg to approximately 7 μg, or approximately 4 μg to approximately 6 μg of lipopolysaccharide. Embodiment 46. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-45, wherein the saponin is derived from the bark of Quillaja saponaria Molina. Embodiment 47. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-46, wherein the saponin is quill A or a derivative thereof. Embodiment 48. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-47, wherein the saponin is QS-17. Embodiment 49. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-47, wherein the saponin is QS-21. Embodiment 50. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-49, comprising approximately 1 μg to approximately 100 μg, approximately 10 μg to approximately 50 μg, approximately 20 μg to approximately 30 μg, approximately 21 μg to approximately 29 μg, approximately 22 μg to approximately 28 μg, approximately 23 μg to approximately 27 μg, or approximately 24 μg to approximately 26 μg of saponins. Embodiment 51. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-49, comprising approximately 1 μg to approximately 30 μg, approximately 5 μg to approximately 15 μg, approximately 6 μg to approximately 14 μg, approximately 7 μg to approximately 13 μg, approximately 8 μg to approximately 12 μg, or approximately 9 μg to approximately 11 μg of saponins. Embodiment 52. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-49, comprising approximately 1 μg to approximately 9 μg, approximately 2 μg to approximately 8 μg, approximately 3 μg to approximately 7 μg, or approximately 4 μg to approximately 6 μg of saponins. Embodiment 53. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-52, wherein the liposome-based adjuvant further comprises a sterol. Embodiment 54. The immunogenic composition according to Embodiment 53, wherein the sterol is β-sitosterol, stigmasterol, ergosterol, ergocalciferol, or cholesterol. Embodiment 55. The immunogenic composition according to Embodiment 53 or Embodiment 54, wherein the sterol is cholesterol. Embodiment 56. An immunogenic composition according to any one of Embodiments 53 to 55, wherein the ratio of saponin to sterol is 1:1 to 1:100 w / w or 1:1 to 1:5 w / w, optionally about 1:1 w / w. Embodiment 57. An immunogenic composition according to any one of Embodiments 1-4, 7, 10-13, 16-20, 25, 26, and 28-56, wherein the liposome-based adjuvant further comprises a neutral lipid. Embodiment 58. The immunogenic composition according to Embodiment 57, wherein the neutral lipid is phosphatidylcholine. Embodiment 59. The immunogenic composition according to Embodiment 58, wherein the phosphatidylcholine is egg yolk phosphatidylcholine, dioleoyl phosphatidylcholine (DOPC), or dilauryl phosphatidylcholine. Embodiment 60. The immunogenic composition according to Embodiment 58 or Embodiment 59, wherein phosphatidylcholine is DOPC. Embodiment 61. An immunogenic composition according to any one of Embodiments 1-4, 7, 10, 13-20, 25, 26, and 28-60, wherein the sorbitan ester is sorbitan ester 20. Embodiment 62. An immunogenic composition according to any one of Embodiments 1-4, 7, 10, 13-20, 25, 26, and 28-60, wherein the sorbitan ester is sorbitan ester 85. Embodiment 63. An immunogenic composition according to any one of Embodiments 1-4, 7, 10, 13-20, 25, 26, and 28-62, wherein the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 20. Embodiment 64. An immunogenic composition according to any one of Embodiments 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 62, wherein the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 80. Embodiment 65. An immunogenic composition according to any one of Embodiments 1-4, 7, 10, 13-20, 25, 26, and 28-64, comprising squalene in an amount of about 2% w / v to about 7% w / v, about 3% w / v to about 6% w / v, or about 4% w / v to about 5% w / v. Embodiment 66. An immunogenic composition according to any one of Embodiments 1-4, 7, 10, 13-20, 25, 26, and 28-65, comprising a sorbitan ester in an amount of about 0.2% w / v to about 0.8% w / v, about 0.3% w / v to about 0.7% w / v, or about 0.4% w / v to about 0.6% w / v. Embodiment 67. An immunogenic composition according to any one of Embodiments 1-4, 7, 10, 13-20, 25, 26, and 28-66, comprising a polyoxyethylene sorbitan ester in an amount of about 0.2% w / v to about 0.8% w / v, about 0.3% w / v to about 0.7% w / v, or about 0.4% w / v to about 0.6% w / v. Embodiment 68. The immunogenic composition according to any one of Embodiments 1 to 67, wherein the OspA antigen is two or more OspA antigens. Embodiment 69. The immunogenic composition according to Embodiment 68, wherein two or more OspA antigens are a first OspA antigen and a second OspA antigen. Embodiment 70. The immunogenic composition according to Embodiment 68 or Embodiment 69, wherein two or more OspA antigens are a first OspA antigen, a second OspA antigen, and a third OspA antigen. Embodiment 71. An immunogenic composition according to any one of Embodiments 68 to 70, wherein each of two or more OspA antigens has a non-identical amino acid sequence. Embodiment 72. An immunogenic composition according to any one of Embodiments 1 to 71, wherein the OspA antigen is independently the OspA antigen of B. burgdorferi, B. afzelii, B. bavariensis, B. garinii, B. mayonii, B. lusitaniae, B. bissettii, B. valasiana, or B. spielmanii. Embodiment 73. An immunogenic composition according to any one of Embodiments 1 to 72, wherein the OspA antigen is independently the OspA antigen of B. burgdorferi. Embodiment 74. An immunogenic composition according to any one of Embodiments 1 to 73, wherein the OspA antigen independently comprises one or more epitopes derived from the OspA protein. Embodiment 75. The immunogenic composition according to Embodiment 74, wherein one or more epitopes comprise at least two, three, four, five, or six epitopes derived across one or more OspA proteins. Embodiment 76. An immunogenic composition according to Embodiment 74 or Embodiment 75, wherein one or more epitopes comprise at least two, three, four, five, or six epitopes derived from at least two, three, four, five, or six OspA serotypes. Embodiment 77. An immunogenic composition according to any one of Embodiments 74 to 76, wherein one or more epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49. Embodiment 78. An immunogenic composition according to any one of Embodiments 74 to 77, wherein one or more epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49. Embodiment 79. An immunogenic composition according to any one of Embodiments 74 to 78, wherein one or more epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49. Embodiment 80. An immunogenic composition according to any one of Embodiments 1 to 78, wherein the OspA antigen independently has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any of SEQ ID NOs. Embodiment 81. An immunogenic composition according to any one of Embodiments 1 to 78 and 80, wherein the OspA antigen independently comprises at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids of the amino acid sequence shown in any one of SEQ ID NOs. 50 to 62. Embodiment 82. An immunogenic composition according to any one of Embodiments 1 to 78, 80, and 81, wherein the OspA antigen independently comprises an amino acid sequence shown in any of SEQ ID NOs. 50 to 62. Embodiment 83. The immunogenic composition according to any one of Embodiments 1 to 78, wherein the OspA antigen is an OspA chimeric antigen comprising a number of epitopes derived from one or more OspA proteins, independently. Embodiment 84. The immunogenic composition according to Embodiment 83, comprising at least two, three, four, five, or six epitopes, the numerous epitopes originating from at least two, three, four, five, or six OspA serotypes. Embodiment 85. The immunogenic composition according to Embodiment 83 or Embodiment 84, wherein the numerous epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49. Embodiment 86. An immunogenic composition according to any one of Embodiments 83 to 85, wherein a number of epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49. Embodiment 87. An immunogenic composition according to any one of Embodiments 83 to 86, wherein the numerous epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49. Embodiment 88. An immunogenic composition according to any one of Embodiments 83 to 87, wherein a number of epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49. Embodiment 89. An immunogenic composition according to any one of Embodiments 1 to 78 and 83 to 86, wherein the OspA antigen independently comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 63 to 82. Embodiment 90. An immunogenic composition according to any one of Embodiments 1 to 78 and 83 to 89, wherein the OspA antigen independently comprises at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids of the amino acid sequence shown in any one of SEQ ID NOs. 63 to 82. Embodiment 91. An immunogenic composition according to any one of Embodiments 1 to 78 and 83 to 90, wherein the OspA antigen independently comprises the amino acid sequence shown in any one of SEQ ID NOs. 63 to 82. Embodiment 92. An immunogenic composition according to any one of Embodiments 1 to 78 and 83 to 89, wherein the OspA antigen independently comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 79 to 81. Embodiment 93. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, and 92, comprising a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 79, and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80. Embodiment 94. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, and 91 to 93, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79, and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80. Embodiment 95. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, and 92, comprising a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 89, and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. Embodiment 96. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, 91, 92, and 95, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79 and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 81. Embodiment 97. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, and 92, comprising a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. Embodiment 98. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, 91, 92, and 97, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80, and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 81. Embodiment 99. A first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, comprising at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, 92, and 93, comprising a second OspA antigen comprising an amino acid sequence having 96%, 97%, 98%, or 99% sequence identity, and a third OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 81. Embodiment 100. An immunogenic composition according to any one of Embodiments 1 to 78, 83 to 89, and 91 to 94, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79, a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80, and a third OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 81. Embodiment 101. An immunogenic composition according to any one of Embodiments 1 to 100, wherein the OspA antigen is independently not lipidized. Embodiment 102. An immunogenic composition according to any one of Embodiments 1 to 101, wherein the OspA antigen is independently lipidized. Embodiment 103. An immunogenic composition according to any one of Embodiments 1 to 102, in a dose of 0.3 mL to 1.5 mL. Embodiment 104. An immunogenic composition according to any one of Embodiments 1 to 103, for stimulating an immune response to Borrelia in a subject by administering an effective amount of the immunogenic composition to the subject. Embodiment 105. An immunogenic composition according to any one of Embodiments 1 to 104, for protecting a subject from infection by Borrelia by administering an effective amount of the immunogenic composition to the subject. Embodiment 106. An immunogenic composition according to any one of Embodiments 1 to 104, for preventing a subject from contracting Lyme disease by administering an effective amount of the immunogenic composition to the subject. Embodiment 107. An immunogenic composition according to any one of Embodiments 1 to 104, wherein the subject is a human subject. Embodiment 108. The immunogenic composition according to Embodiment 107, wherein the human subject is at least 18 years of age. Embodiment 109. The immunogenic composition according to Embodiment 107 or Embodiment 108, wherein the human subject is between 18 and 70 years of age. Embodiment 110. The immunogenic composition according to Embodiment 107, wherein the human subject is under 18 years of age. Embodiment 111. The immunogenic composition according to Embodiment 107 or Embodiment 110, wherein the human subject is 2 to 5 years of age. Embodiment 112. The immunogenic composition according to Embodiment 107 or Embodiment 110, wherein the human subject is between 5 and 18 years of age. Embodiment 113. A method for stimulating an immune response to Borrelia in a subject, comprising administering an effective amount of the immunogenic composition described in any one of Embodiments 1 to 112 to the subject. Embodiment 114. A method for protecting a subject from infection by Borrelia, comprising administering an effective amount of the immunogenic composition described in any one of Embodiments 1 to 112 to the subject. Embodiment 115. A method for preventing a subject from contracting Lyme disease, comprising administering an effective amount of the immunogenic composition described in any one of Embodiments 1 to 112 to the subject. Embodiment 116. The method according to any one of Embodiments 113 to 115, wherein the immunogenic composition is administered by intramuscular injection. Embodiment 117. The method according to any one of Embodiments 113 to 116, wherein a first dose and a second dose of the immunogenic composition are administered to a subject. Embodiment 118. The method according to Embodiment 117, wherein a second dose of the immunogenic composition is administered approximately 2 to 10 weeks after the administration of the first dose of the immunogenic composition. Embodiment 119. The method according to Embodiment 117 or Embodiment 118, wherein a second dose of the immunogenic composition is administered about one month or about two months after the administration of a first dose of the immunogenic composition. Embodiment 120. The method according to any one of Embodiments 117 to 119, wherein a third dose of the immunogenic composition is administered to a subject. Embodiment 121. The method according to Embodiment 120, wherein a third dose of the immunogenic composition is administered approximately 4 to 8 months after the administration of the first dose of the immunogenic composition. Embodiment 122. The method according to Embodiment 120 or Embodiment 121, wherein a third dose of the immunogenic composition is administered approximately six months after the administration of a first dose of the immunogenic composition. Embodiment 123. The method according to any one of Embodiments 113 to 122, wherein a booster dose of the immunogenic composition is administered to the subject. Embodiment 124. The method according to Embodiment 123, wherein the booster dose is administered at least about one year or at least about two years after the administration of the first dose. Embodiment 125. The method according to any one of Embodiments 113 to 124, wherein multiple booster doses are administered to the subject. Embodiment 126. The method according to Embodiment 125, wherein the first booster dose is administered at least about one year or at least about two years after the administration of the first dose. Embodiment 127. The method according to Embodiment 125 or Embodiment 126, wherein the booster dose is administered once a year. Embodiment 128. The method according to any one of Embodiments 113 to 127, wherein the subject is a human subject. Embodiment 129. The method according to Embodiment 128, wherein the human subject is at least 18 years of age. Embodiment 130. The method according to Embodiment 128 or Embodiment 129, wherein the human subject is between 18 and 70 years of age. Embodiment 131. The method according to Embodiment 128, wherein the human subject is under 18 years of age. Embodiment 132. The method according to Embodiment 128 or Embodiment 131, wherein the human subject is between 2 and 5 years of age. Embodiment 133. The method according to Embodiment 128 or Embodiment 131, wherein the human subject is between 5 and 18 years of age. Embodiment 134. Use of an immunogenic composition according to any one of Embodiments 1 to 112 to stimulate an immune response to Borrelia in a subject by administering an effective amount of the immunogenic composition to the subject. Embodiment 135. Use of an immunogenic composition according to any one of Embodiments 1 to 112 to protect a subject from infection by Borrelia by administering an effective amount of the immunogenic composition to the subject. Embodiment 136. Use of an immunogenic composition according to any one of Embodiments 1 to 112 to prevent a subject from contracting Lyme disease by administering an effective amount of the immunogenic composition to the subject. Embodiment 137. Use of an immunogenic composition according to any one of Embodiments 1 to 112 in the manufacture of a pharmaceutical product for stimulating an immune response to Borrelia in a subject. Embodiment 138. Use of an immunogenic composition according to any one of Embodiments 1 to 112 in the manufacture of a pharmaceutical product for protecting a subject from infection by Borrelia. Embodiment 139. Use of an immunogenic composition according to any one of Embodiments 1 to 112 in the manufacture of a pharmaceutical product for preventing a subject from contracting Lyme disease. Embodiment 140. The use according to any one of Embodiments 134 to 139, wherein the immunogenic composition is administered by intramuscular injection. Embodiment 141. The use according to any one of Embodiments 134 to 140, wherein a first dose and a second dose of the immunogenic composition are administered to the subject. Embodiment 142. The use according to Embodiment 141, wherein a second dose of the immunogenic composition is administered approximately 2 to 10 weeks after the administration of the first dose of the immunogenic composition. Embodiment 143. The use according to Embodiment 141 or Embodiment 142, wherein a second dose of the immunogenic composition is administered about one month or about two months after the administration of the first dose of the immunogenic composition. Embodiment 144. The use according to any one of Embodiments 141 to 143, wherein a third dose of the immunogenic composition is administered to the subject. Embodiment 145. The use according to Embodiment 144, wherein a third dose of the immunogenic composition is administered approximately 4 to 8 months after the administration of the first dose of the immunogenic composition. Embodiment 146. The use according to Embodiment 144 or Embodiment 145, wherein a third dose of the immunogenic composition is administered about six months after the administration of a first dose of the immunogenic composition. Embodiment 147. The use according to any one of Embodiments 134 to 146, wherein a booster dose of the immunogenic composition is administered to the subject. Embodiment 148. The use according to Embodiment 147, wherein the booster dose is administered at least about one year or at least about two years after the administration of the first dose. Embodiment 149. The use according to any one of Embodiments 134 to 148, wherein multiple booster doses are administered to the subject. Embodiment 150. The use according to Embodiment 149, wherein the first booster dose is administered at least about one year or at least about two years after the administration of the first dose. Embodiment 151. The use described in Embodiment 149 or Embodiment 150, wherein the booster dose is administered once a year. Embodiment 152. The use described in any one of Embodiments 133 to 151, wherein the subject is a human subject. Embodiment 153. The use described in Embodiment 152, wherein the human subject is at least 18 years of age. Embodiment 154. The use described in Embodiment 152 or Embodiment 153, wherein the human subject is between 18 and 70 years of age. Embodiment 155. The use described in Embodiment 152, wherein the human subject is under 18 years of age. Embodiment 156. The use described in Embodiment 152 or Embodiment 155, wherein the human subject is between 2 and 5 years of age. Embodiment 157. The use described in Embodiment 152 or Embodiment 155, wherein the human subject is between 5 and 18 years of age. Embodiment 158. A vial containing the immunogenic composition described in any one of Embodiments 1 to 112. Embodiment 159. The vial according to Embodiment 158, wherein the immunogenic composition is in liquid form. Embodiment 160. The vial according to Embodiment 158, wherein the immunogenic composition is in a lyophilized form. Embodiment 161. A vial according to any one of Embodiments 158 to 160, wherein the vial contains a single dose of an immunogenic composition. Embodiment 162. The vial according to Embodiment 161, wherein the single dose is 0.3 mL to 1.5 mL. Embodiment 163. A pre-filled syringe comprising the immunogenic composition described in any one of Embodiments 1 to 112. Embodiment 164. The pre-filled syringe according to Embodiment 163, wherein the syringe contains a single dose of the immunogenic composition. Embodiment 165. A pre-filled syringe according to Embodiment 164, wherein the single dose is 0.3 mL to 1.5 mL. Embodiment 166. a) A first composition comprising an adjuvant selected from aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters; b) A second composition comprising Borrelia cell surface protein A (OspA) antigen; and c) Instructions for preparing an immunogenic composition according to any one of embodiments 1 to 112 by combining the first composition and the second composition. A kit that includes this. Embodiment 167. The kit according to Embodiment 166, wherein the second composition is in liquid form. Embodiment 168. The kit according to Embodiment 166, wherein the second composition is in a freeze-dried form. Embodiment 169. A kit according to any one of Embodiments 166 to 168, wherein the instructions are for preparing a single-dose immunogenic composition. Embodiment 170. The kit according to any one of Embodiments 166 to 169, wherein the kit is for a single-dose immunogenic composition. Embodiment 171. The kit according to Embodiment 169 or Embodiment 170, wherein the single dose is 0.3 mL to 1.5 mL. Embodiment 172. A kit according to any one of Embodiments 166 to 171, wherein the first and / or second composition comprises a second adjuvant. Embodiment 173. The kit according to Embodiment 172, wherein the second adjuvant is an aluminum salt adjuvant. Embodiment 174. The kit according to Embodiment 173, wherein the aluminum salt adjuvant comprises one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate. Embodiment 175. The kit according to Embodiment 173 or Embodiment 174, wherein the aluminum salt adjuvant comprises aluminum hydroxide. Embodiment 176. d) A further set of instructions for administering immunogenic compositions to subjects to stimulate an immune response to Borrelia in the subjects. A kit according to any one of embodiments 166 to 175, further comprising: Embodiment 177. a) The immunogenic composition according to any one of Embodiments 1 to 112; and b) Instructions for administering immunogenic compositions to subjects to stimulate an immune response to Borrelia in the subjects. A kit that includes this. Embodiment 178. The kit according to Embodiment 176 or Embodiment 177, further comprising a syringe for intramuscular injection of an immunogenic composition. Embodiment 179. The kit according to Embodiment 178, wherein the syringe is pre-filled with an immunogenic composition. Embodiment 180. a) Vials according to any one of embodiments 158 to 162; and b) Instructions for administering immunogenic compositions to subjects to stimulate an immune response to Borrelia in the subjects. A kit that includes this. Embodiment 181. The kit according to Embodiment 180, further comprising a syringe for intramuscular injection of an immunogenic composition. Embodiment 182. a) A pre-filled syringe according to any one of embodiments 163 to 165; and b) Instructions for administering immunogenic compositions to subjects to stimulate an immune response to Borrelia in the subjects. A kit that includes this. Embodiment 183. The kit according to Embodiment 182, wherein the syringe is for intramuscular injection of an immunogenic composition. Embodiment 184. The kit according to any one of Embodiments 176 to 183, wherein administration follows the method described in any one of Embodiments 124 to 144. Embodiment 185. The kit according to any one of Embodiments 176 to 184, wherein the subject is a human subject. Embodiment 186. The kit according to Embodiment 185, wherein the human subject is at least 18 years of age. Embodiment 187. The kit according to Embodiment 185 or Embodiment 186, wherein the human subject is between 18 and 70 years of age. Embodiment 188. The kit according to Embodiment 185, wherein the human subject is under 18 years of age. Embodiment 189. The kit according to Embodiment 185 or Embodiment 188, wherein the human subject is between 2 and 5 years of age. Embodiment 190. The kit according to Embodiment 185 or Embodiment 188, wherein the human subject is between 5 and 18 years of age. [Examples]

[0309] Example 1 The effect of adjuvants on the immune response to Lyme disease immunogenic compositions.

[0310] Abbreviations: OD (optical density); OspA (cell surface protein A); alum (aluminum hydroxide); ELISA (enzyme-linked immunosorbent assay); mcg (microgram)

[0311] OspA serotypes: OspA serotype 1 is derived from Borrelia Burgdorferi. OspA serotype 2 is derived from Borrelia afzelii. OspA serotypes 3, 5, and 6 are derived from Borrelia garinii. OspA serotype 4 is derived from Borrelia bavariensis.

[0312] The experiment was conducted in mice to compare the effects of adjuvants administered with one of three different immunogenic compositions containing one of three different OspA antigens or antigen mixtures, formulated with or without adjuvants. The adjuvants tested included 1) CpG (whose nucleotide sequence is shown in SEQ ID NO: 1), 2) alum, or 3) a combination of CpG and alum.

[0313] Three different antigens or mixtures of antigens were tested in immunogenic compositions: (a) a non-lipidized full-length version of the OspA serotype 1 antigen (Borrelia Burgdorferi strain B31) (OspA S1) shown in SEQ ID NO: 50; (b) a mixture of non-lipidized OspA chimeric antigens containing three chimeric OspA antigens covering six serotypes (OspA chimeric antigens); and (c) a lipidized OspA chimeric antigen, which is the same as the three chimeric OspA chimeric antigens described in (b) but is further lipidized. The OspA chimeric antigen in the mixture of (b) above is illustrated below in Figure 1 and contains three different chimeric OspA antigens, each of which combines two different serotypes derived from OspA. The first chimeric antigen (S1 / 2), which covers the C-terminal portion of OspA serotype 2 and the N-terminal portion of OspA serotype 1, is shown in SEQ ID NO: 79. A second chimeric antigen (S6 / 4), covering the C-terminal portion of OspA serotype 4 and the N-terminal portion of OspA serotype 6, is shown in SEQ ID NO: 80. A third chimeric antigen (S5 / 3), covering the C-terminal portion of OspA serotype 3 and the N-terminal portion of OspA serotype 5, is shown in SEQ ID NO: 81. For the lipidized versions of the chimeric antigens in (c) above, the three aforementioned chimeric antigens contained additional fatty acids and glycerols added to the N-terminus of each OspA chimeric antigen. In addition, the His tag shown in SEQ ID NO: 87 was added to the C-terminus of each antigen. For example, the OspA S1 antigen with the His tag added is shown in SEQ ID NO: 86.

[0314] Female C3H / HeN mice were divided into 12 treatment groups, each containing 8 animals, and each group received one of 12 immunogenic compositions containing a given OspA antigen or mixture of OspA antigens and an adjuvant preparation, as shown in Table E1 below. Each type of antigen or mixture of antigens contained either no adjuvant, alum alone, CpG alone, or both CpG and alum. The immunogenic composition adjuvanted with alum alone contained antigen adsorbed to 50 micrograms of aluminum hydroxide (2% alpha hydrogel, Al(OH)3). The immunogenic composition adjuvanted with CpG alone contained antigen mixed with 10 micrograms of CpG. The immunogenic composition adjuvanted with both alum and CpG contained antigen adsorbed to 50 micrograms of alum and then mixed with 10 micrograms of CpG. All antigens and adjuvants were diluted in sterile saline.

[0315] [Table 1] JPEG2026525284000003.jpg245160

[0316] Female mice were injected intramuscularly on days 0 and 21. Blood was collected on days 21 and 35 for measurement of serum antibody levels by ELISA. The response was measured for each mouse. Antibody endpoint titer was defined as the reciprocal of the highest dilution from undiluted serum required to achieve 0.5 OD. A positive control was included for each ELISA plate to control for plate-to-plate variability. The positive control contained a mouse serum sample from a previous study with a defined antibody level. The negative control contained serum from unvaccinated mice.

[0317] Antibody levels in the serum of mice administered an immunogenic composition containing non-lipidized (non-lip) OspA chimeric antigen at day 35, i.e., after the second injection, were measured against OspA S5 / 3 (Figure 2A), OspA S6 / 4 (Figure 2B), OspA S1 / 2 (Figure 2C; left), or full-length OspA S1 (Figure 2D; left). Antibody levels in the serum of mice administered an immunogenic composition containing non-lip OspA S1 at day 35 were measured against either OspA S1 / 2 (Figure 2C; right) or full-length OspA S1 (Figure 2D; right). Finally, antibody levels in the serum of mice administered an immunogenic composition containing lipidized OspA chimeric antigen at day 35 were measured against OspA S1 / 2 (Figure 2E), OspA S5 / 3 (Figure 2F), or OspA S6 / 4 (Figure 2G). For each figure, the increase in antibody level ratio (FI) between the two shown conditions is also shown, measured by comparing the median antibody level from one shown condition to the other shown condition. In addition, for Figures 2E-2G, the median antibody level is also shown above each condition.

[0318] In mice administered immunogenic compositions containing non-lipidized OspA antigens, as shown in Figures 2A-2D, the use of any adjuvant, including alum, CpG, and CpG+alum, induced higher OspA antibody titers compared to the use of no adjuvant for each OspA antigen tested against it. Furthermore, the use of CpG+alum combinations formulated with any non-lipidized OspA antigen resulted in the highest OspA titers for each OspA antigen tested against it, with an FI 2.8–4.1 higher compared to the use of alum alone. In addition, as shown in Figure 2D, the use of either OspA chimeric antigen or full-length OspA S1 resulted in a similar antibody response to full-length OspA S1, indicating that both OspA chimeric antigen and S1 are similarly immunogenic to full-length OspA S1, and that antibodies against OspA S1 / 2 can recognize the main S1 epitope present on full-length OspA S1. In other words, the OspA S1 / 2 chimeric antigen retains the immunogenicity of the full-length OspA S1 antigen.

[0319] In contrast, as shown in Figures 2E–2G, in mice administered an immunogenic composition containing a lipidized OspA chimeric antigen, the use of alum resulted in only a limited increase in OspA antibody titer compared to the use without adjuvant. However, the use of either CpG or CpG + alum significantly enhanced OspA antibody titer compared to the use without adjuvant or with alum alone.

[0320] In summary, these results indicate that both non-lipidized and lipidized OspA chimeric antigens are immunogenic against OspA. Furthermore, the use of CpG-containing adjuvant formulations, with or without alum, enhanced the OspA antibody response to several OspA antigens when combined with several different OspA antigens or mixtures of OspA antigens.

[0321] In addition to evaluating antibody titers after the second injection, the antibody response dynamics between the first and second injections were also evaluated in serum from mice treated with lipid-modified OspA chimeric antigens. Figures 3A–3C show the antibody titers against OspA S1 / 2, OspA S5 / 3, and OspA S6 / 4 for each lipid-modified OspA formulation tested, in serum from day 21 after the first injection (1) and day 35 after the second injection (2). The increase in factorial change (FI) in antibody values, calculated by comparing the median antibody level induced by a given composition at day 35 to the median antibody level induced by the same composition at day 21, is also shown above each condition. The median antibody titer is also shown in Table E2.

[0322] [Table 2]

[0323] The results indicate that the first dose of the immunogenic composition yielded the highest antibody titer when formulated with both CpG and alum, compared to formulations without adjuvant, with alum alone, or with CpG alone. Therefore, the combination of OspA antigen, CpG, and alum has a synergistic effect that results in a greater immune response (i.e., after the first dose and before receiving the second dose). Furthermore, the second dose of the immunogenic composition, when formulated with both CpG and alum, increased the OspA antibody titer by between 5.6 and 8.8 times, depending on the serotype tested.

[0324] Overall, these results suggest that administration of CpG and OspA-based Lyme disease immunogenic compositions, with or without alum, may improve subsequent antibody responses to many different OspA serotypes, for example, by providing a greater immune response.

[0325] While the foregoing disclosure has been described in some detail by description and examples for the purposes of clarity and understanding, it will be apparent to those skilled in the art that certain changes and modifications may be made. Therefore, the examples should not be construed as limiting the scope of this disclosure, as this is further detailed by the appended claims.

[0326] array

[0327] [Table 3] JPEG2026525284000006.jpg254158JPEG2026525284000007.jpg255156JPEG2026525284000008.jpg254157 JPEG2026525284000009.jpg252156JPEG2026525284000010.jpg253156JPEG2026525284000011.jpg142154

Claims

1. An immunogenic composition comprising (i) a Borrelia cell surface protein A (OspA) antigen or an RNA polynucleotide encoding the OspA antigen, and (ii) an adjuvant selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsions containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

2. An immunogenic composition comprising (i) a Borrelia cell surface protein A (OspA) antigen, and (ii) an adjuvant selected from the group consisting of aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsions containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

3. The immunogenic composition according to claim 1 or claim 2, which does not contain RNA polynucleotides encoding non-OspA Borrelia cell surface protein antigens.

4. An immunogenic composition according to any one of claims 1 to 3, which does not contain non-OspA Borrelia cell surface protein antigens.

5. The immunogenic composition according to any one of claims 1 to 4, wherein the adjuvant is an aluminum salt adjuvant.

6. The immunogenic composition according to claim 5, further comprising a second adjuvant.

7. The immunogenic composition according to claim 6, wherein the second adjuvant is selected from a TLR9 agonist; a liposome-based adjuvant comprising lipopolysaccharide and saponin; and an oil-in-water emulsion comprising squalene, sorbitan ester, and polyoxyethylene sorbitan ester.

8. The immunogenic composition according to any one of claims 1 to 4, wherein the adjuvant is a TLR9 agonist.

9. The immunogenic composition according to claim 8, further comprising a second adjuvant.

10. The immunogenic composition according to claim 9, wherein the second adjuvant is selected from aluminum salt adjuvants; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

11. The immunogenic composition according to any one of claims 1 to 4, wherein the adjuvant is a liposome-based adjuvant containing lipopolysaccharide and saponin.

12. The immunogenic composition according to claim 11, further comprising a second adjuvant.

13. The immunogenic composition according to claim 12, wherein the second adjuvant is selected from aluminum salt adjuvants; TLR9 agonists; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

14. The immunogenic composition according to any one of claims 1 to 4, wherein the adjuvant is an oil-in-water emulsion-based adjuvant comprising squalene, sorbitan ester, and polyoxyethylene sorbitan ester.

15. The immunogenic composition according to claim 14, further comprising a second adjuvant.

16. The immunogenic composition according to claim 15, wherein the second adjuvant is selected from aluminum salt adjuvants; TLR9 agonists; and liposome-based adjuvants comprising lipopolysaccharides and saponins.

17. The immunogenic composition according to any one of claims 1 to 4, 7 to 10, 13, and 16, wherein the TLR9 agonist is an oligonucleotide containing a nonmethylated cytidine-phosphoguanosine (CpG) motif.

18. The immunogenic composition according to claim 17, wherein the oligonucleotide has a length of 10 to 35 nucleotides.

19. The immunogenic composition according to claim 17 or claim 18, wherein the oligonucleotide comprises sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2).

20. The immunogenic composition according to any one of claims 17 to 19, wherein the oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1).

21. An immunogenic composition comprising a Borrelia cell surface protein A (OspA) antigen, and a Toll-like receptor 9 (TLR9) agonist oligonucleotide 10 to 35 nucleotides in length, containing the sequence 5'-AACGTTCGAG-3' (SEQ ID NO: 2) and a non-methylated cytidine-phospho-guanosine (CpG) motif.

22. The immunogenic composition according to claim 21, wherein the oligonucleotide comprises the sequence 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1).

23. An immunogenic composition comprising an oligonucleotide, the Borrelia cell surface protein A (OspA) antigen and the Toll-like receptor 9 (TLR9) agonist oligonucleotide 5'-TGACTGTGAACGTTCGAGATGA-3' (SEQ ID NO: 1), wherein the oligonucleotide contains a non-methylated cytidine-phospho-guanosine (CpG) motif.

24. The immunogenic composition according to any one of claims 21 to 23, further comprising a second adjuvant.

25. The immunogenic composition according to claim 24, wherein the second adjuvant is selected from aluminum salt adjuvants; liposome-based adjuvants comprising lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants comprising squalene, sorbitan esters, and polyoxyethylene sorbitan esters.

26. The immunogenic composition according to any one of claims 9, 10, 12, 13, 15-20, 24, and 25, wherein the second adjuvant is an aluminum salt adjuvant.

27. An immunogenic composition comprising (i) Borrelia cell surface protein A (OspA) antigen, (ii) Toll-like receptor 9 (TLR9) agonist oligonucleotide 5'-TGACTGTGAA CGTTCGAGAT GA-3' (SEQ ID NO: 1), wherein the oligonucleotide contains a non-methylated cytidine-phospho-guanosine (CpG) motif, and (iii) an aluminum salt adjuvant.

28. The immunogenic composition according to any one of claims 1 to 7, 10, 13, 16 to 20, and 25 to 27, wherein the aluminum salt adjuvant comprises one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate.

29. The immunogenic composition according to any one of claims 1 to 7, 10, 13, 16 to 20, and 25 to 28, wherein the aluminum salt adjuvant comprises aluminum hydroxide.

30. Approximately 0.25 to 1.25 mg of Al ３＋ Approximately 0.25 mg to approximately 0.50 mg of Al ３＋ , or approximately 0.30 mg to approximately 0.40 mg of Al ３＋ An immunogenic composition according to any one of claims 1 to 29, comprising:

31. The immunogenic composition according to any one of claims 17 to 30, wherein the oligonucleotide is a single-stranded oligodeoxynucleotide.

32. The immunogenic composition according to any one of claims 17 to 30, wherein the oligonucleotide is complete RNA.

33. The immunogenic composition according to any one of claims 17 to 30, wherein the oligonucleotide is an RNA / DNA chimera.

34. The immunogenic composition according to any one of claims 17 to 33, wherein the oligonucleotide comprises at least one phosphorothioate linkage.

35. The immunogenic composition according to any one of claims 17 to 34, wherein the oligonucleotide comprises only phosphorothioate linkages.

36. The immunogenic composition according to any one of claims 17 to 34, wherein the oligonucleotide comprises a combination of one or more phosphodiester links and one or more phosphorothioate links.

37. An immunogenic composition according to any one of claims 1 to 4, 7 to 10, 13, and 16 to 36, comprising approximately 375 μg to approximately 6000 μg of TLR9 agonist or approximately 750 μg to approximately 3000 μg of TLR9 agonist.

38. An immunogenic composition according to any one of claims 1 to 4, 7 to 10, 13, and 16 to 36, comprising approximately 375 μg, approximately 750 μg, approximately 1000 μg, approximately 1500 μg, approximately 3000 μg, or approximately 6000 μg of a TLR9 agonist.

39. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 38, wherein the lipopolysaccharide is lipid A or a derivative thereof.

40. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 39, wherein the lipopolysaccharide is monophosphoryl lipid A or a derivative thereof.

41. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 40, wherein the lipopolysaccharide is 3-O-desacyl-4'-monophosphoryl lipid A.

42. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 40, wherein the lipopolysaccharide is 3-deacyl-phosphorylated hexaacyl disaccharide.

43. An immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 42, comprising approximately 1 μg to approximately 100 μg, approximately 10 μg to approximately 50 μg, approximately 20 μg to approximately 30 μg, approximately 21 μg to approximately 29 μg, approximately 22 μg to approximately 28 μg, approximately 23 μg to approximately 27 μg, or approximately 24 μg to approximately 26 μg of lipopolysaccharide.

44. An immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 42, comprising approximately 1 μg to approximately 30 μg, approximately 5 μg to approximately 15 μg, approximately 6 μg to approximately 14 μg, approximately 7 μg to approximately 13 μg, approximately 8 μg to approximately 12 μg, or approximately 9 μg to approximately 11 μg of lipopolysaccharide.

45. An immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 42, comprising approximately 1 μg to approximately 9 μg, approximately 2 μg to approximately 8 μg, approximately 3 μg to approximately 7 μg, or approximately 4 μg to approximately 6 μg of lipopolysaccharide.

46. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 45, wherein the saponin is derived from the bark of Quillaja saponaria molina.

47. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 46, wherein the saponin is quill A or a derivative thereof.

48. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 47, wherein the saponin is QS-17.

49. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 47, wherein the saponin is QS-21.

50. An immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 49, comprising approximately 1 μg to approximately 100 μg, approximately 10 μg to approximately 50 μg, approximately 20 μg to approximately 30 μg, approximately 21 μg to approximately 29 μg, approximately 22 μg to approximately 28 μg, approximately 23 μg to approximately 27 μg, or approximately 24 μg to approximately 26 μg of saponins.

51. An immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 49, comprising approximately 1 μg to approximately 30 μg, approximately 5 μg to approximately 15 μg, approximately 6 μg to approximately 14 μg, approximately 7 μg to approximately 13 μg, approximately 8 μg to approximately 12 μg, or approximately 9 μg to approximately 11 μg of saponins.

52. An immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 49, comprising approximately 1 μg to approximately 9 μg, approximately 2 μg to approximately 8 μg, approximately 3 μg to approximately 7 μg, or approximately 4 μg to approximately 6 μg of saponins.

53. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 52, wherein the liposome-based adjuvant further comprises sterols.

54. The immunogenic composition according to claim 53, wherein the sterol is β-sitosterol, stigmasterol, ergosterol, ergocalciferol, or cholesterol.

55. The immunogenic composition according to claim 53 or claim 54, wherein the sterol is cholesterol.

56. The immunogenic composition according to any one of claims 53 to 55, wherein the ratio of saponins to sterols is 1:1 to 1:100 w / w or 1:1 to 1:5 w / w, optionally about 1:1 w / w.

57. The immunogenic composition according to any one of claims 1 to 4, 7, 10 to 13, 16 to 20, 25, 26, and 28 to 56, wherein the liposome-based adjuvant further comprises a neutral lipid.

58. The immunogenic composition according to claim 57, wherein the neutral lipid is phosphatidylcholine.

59. The immunogenic composition according to claim 58, wherein the phosphatidylcholine is egg yolk phosphatidylcholine, dioleoyl phosphatidylcholine (DOPC), or dilauryl phosphatidylcholine.

60. The immunogenic composition according to claim 58 or claim 59, wherein phosphatidylcholine is DOPC.

61. The immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 60, wherein the sorbitan ester is sorbitan ester 20.

62. The immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 60, wherein the sorbitan ester is sorbitan ester 85.

63. The immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 62, wherein the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 20.

64. The immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 62, wherein the polyoxyethylene sorbitan ester is polyoxyethylene sorbitan ester 80.

65. An immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 64, comprising squalene in an amount of approximately 2% w / v to approximately 7% w / v, approximately 3% w / v to approximately 6% w / v, or approximately 4% w / v to approximately 5% w / v.

66. An immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 65, comprising a sorbitan ester in an amount of approximately 0.2% w / v to approximately 0.8% w / v, approximately 0.3% w / v to approximately 0.7% w / v, or approximately 0.4% w / v to approximately 0.6% w / v.

67. An immunogenic composition according to any one of claims 1 to 4, 7, 10, 13 to 20, 25, 26, and 28 to 66, comprising polyoxyethylene sorbitan ester in an amount of approximately 0.2% w / v to approximately 0.8% w / v, approximately 0.3% w / v to approximately 0.7% w / v, or approximately 0.4% w / v to approximately 0.6% w / v.

68. The immunogenic composition according to any one of claims 1 to 67, wherein the OspA antigen is two or more OspA antigens.

69. The immunogenic composition according to claim 68, wherein two or more OspA antigens are a first OspA antigen and a second OspA antigen.

70. The immunogenic composition according to claim 68 or claim 69, wherein two or more OspA antigens are a first OspA antigen, a second OspA antigen, and a third OspA antigen.

71. The immunogenic composition according to any one of claims 68 to 70, wherein each of two or more OspA antigens has a non-identical amino acid sequence.

72. The immunogenic composition according to any one of claims 1 to 71, wherein the OspA antigen is independently the OspA antigen of B. burgdorferi, B. afzelii, B. bavariensis, B. garinii, B. mayonii, B. lustaniae, B. bissettii, B. valasiana, or B. spielmanii.

73. The immunogenic composition according to any one of claims 1 to 72, wherein the OspA antigen is independently the OspA antigen of B. burgdorferi.

74. The immunogenic composition according to any one of claims 1 to 73, wherein the OspA antigen independently comprises one or more epitopes derived from the OspA protein.

75. The immunogenic composition according to claim 74, wherein one or more epitopes comprise at least two, three, four, five, or six epitopes derived across one or more OspA proteins.

76. The immunogenic composition according to claim 74 or 75, comprising at least two, three, four, five, or six epitopes, one or more of which originate from at least two, three, four, five, or six OspA serotypes.

77. The immunogenic composition according to any one of claims 74 to 76, wherein one or more epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49.

78. The immunogenic composition according to any one of claims 74 to 77, wherein one or more epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49.

79. The immunogenic composition according to any one of claims 74 to 78, wherein one or more epitopes comprises epitopes selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49.

80. The immunogenic composition according to any one of claims 1 to 78, wherein the OspA antigen independently has at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 50 to 62.

81. The immunogenic composition according to any one of claims 1 to 78 and 80, wherein the OspA antigen independently comprises at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids of the amino acid sequence shown in any one of SEQ ID NOs.

82. The immunogenic composition according to any one of claims 1 to 78, 80, and 81, wherein the OspA antigen independently comprises an amino acid sequence shown in any one of sequence numbers 50 to 62.

83. The immunogenic composition according to any one of claims 1 to 78, wherein the OspA antigen is an OspA chimeric antigen comprising a number of epitopes derived from one or more OspA proteins, independently.

84. The immunogenic composition according to claim 83, wherein the numerous epitopes comprise at least two, three, four, five, or six epitopes derived from at least two, three, four, five, or six OspA serotypes.

85. The immunogenic composition according to claim 83 or claim 84, wherein the numerous epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49.

86. The immunogenic composition according to any one of claims 83 to 85, wherein a number of epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 4 to 49.

87. The immunogenic composition according to any one of claims 83 to 86, wherein the numerous epitopes include epitopes selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49.

88. The immunogenic composition according to any one of claims 83 to 87, wherein a number of epitopes are independently selected from the amino acid sequences shown in SEQ ID NOs: 9, 10, 33, 34, 36, 47, and 49.

89. The immunogenic composition according to any one of claims 1 to 78 and 83 to 86, wherein the OspA antigen independently comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 63 to 82.

90. The immunogenic composition according to any one of claims 1 to 78 and 83 to 89, wherein the OspA antigen independently comprises at least 100, 110, 120, 130, 140, 150, 160, 170, 180, 190, 200, 210, 220, 230, 240, or 250 adjacent amino acids of the amino acid sequence shown in any one of sequence numbers 63 to 82.

91. The immunogenic composition according to any one of claims 1 to 78 and 83 to 90, wherein the OspA antigen independently comprises an amino acid sequence shown in any one of sequence numbers 63 to 82.

92. The immunogenic composition according to any one of claims 1 to 78 and 83 to 89, wherein the OspA antigen independently comprises an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in any one of SEQ ID NOs: 79 to 81.

93. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, and 92, comprising: a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 79; and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO:

80.

94. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, and 91 to 93, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79, and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO:

80.

95. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, and 92, comprising: a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 89; and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO:

81.

96. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, 91, 92, and 95, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79, and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO:

81.

97. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, and 92, comprising: a first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80; and a second OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO:

81.

98. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, 91, 92, and 97, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80, and a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO:

81.

99. A first OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO: 80, comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, or 96% sequence identity with the amino acid sequence shown in SEQ ID NO: 80 The immunogenic composition according to any one of claims 1 to 78, 83 to 89, 92, and 93, comprising a second OspA antigen comprising an amino acid sequence having 97%, 98%, or 99% sequence identity, and a third OspA antigen comprising an amino acid sequence having at least 80%, 81%, 82%, 83%, 84%, 85%, 86%, 87%, 88%, 89%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, or 99% sequence identity with the amino acid sequence shown in SEQ ID NO:

81.

100. An immunogenic composition according to any one of claims 1 to 78, 83 to 89, and 91 to 94, comprising a first OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 79, a second OspA antigen comprising the amino acid sequence shown in SEQ ID NO: 80, and a third OspA antigen comprising the amino acid sequence shown in SEQ ID NO:

81.

101. The immunogenic composition according to any one of claims 1 to 100, wherein the OspA antigen is independently not lipidized.

102. The immunogenic composition according to any one of claims 1 to 101, wherein the OspA antigen is independently lipidized.

103. An immunogenic composition according to any one of claims 1 to 102, wherein the dose is 0.3 mL to 1.5 mL.

104. An immunogenic composition according to any one of claims 1 to 103, for stimulating an immune response to Borrelia in a subject by administering an effective amount of the immunogenic composition to the subject.

105. An immunogenic composition according to any one of claims 1 to 104, for protecting a subject from infection by Borrelia by administering an effective amount of the immunogenic composition to the subject.

106. An immunogenic composition according to any one of claims 1 to 104, for the purpose of preventing a subject from contracting Lyme disease by administering an effective amount of the immunogenic composition to the subject.

107. The immunogenic composition according to any one of claims 1 to 104, wherein the target is a human subject.

108. The immunogenic composition according to claim 107, wherein the human subject is at least 18 years of age.

109. The immunogenic composition according to claim 107 or claim 108, wherein the human subject is between 18 and 70 years of age.

110. The immunogenic composition according to claim 107, wherein the human subject is under 18 years of age.

111. The immunogenic composition according to claim 107 or claim 110, wherein the human subject is between 2 and 5 years of age.

112. The immunogenic composition according to claim 107 or claim 110, wherein the human subject is between 5 and 18 years of age.

113. A method for stimulating an immune response to Borrelia in a subject, comprising administering an effective amount of the immunogenic composition described in any one of claims 1 to 112 to the subject.

114. A method for protecting a subject from infection by Borrelia, comprising administering an effective amount of the immunogenic composition described in any one of claims 1 to 112 to the subject.

115. A method for preventing a subject from contracting Lyme disease, comprising administering an effective amount of the immunogenic composition described in any one of claims 1 to 112 to the subject.

116. The method according to any one of claims 113 to 115, wherein the immunogenic composition is administered by intramuscular injection.

117. The method according to any one of claims 113 to 116, wherein a first dose and a second dose of the immunogenic composition are administered to a subject.

118. The method according to claim 117, wherein a second dose of the immunogenic composition is administered approximately two weeks to approximately ten weeks after the administration of a first dose of the immunogenic composition.

119. The method according to claim 117 or claim 118, wherein a second dose of the immunogenic composition is administered about one month or about two months after the administration of a first dose of the immunogenic composition.

120. The method according to any one of claims 117 to 119, wherein a third dose of the immunogenic composition is administered to a subject.

121. The method according to claim 120, wherein a third dose of the immunogenic composition is administered approximately four to eight months after the administration of a first dose of the immunogenic composition.

122. The method according to claim 120 or claim 121, wherein a third dose of the immunogenic composition is administered about six months after the administration of a first dose of the immunogenic composition.

123. The method according to any one of claims 113 to 122, wherein a booster dose of the immunogenic composition is administered to the subject.

124. The method according to claim 123, wherein the booster dose is administered at least about one year or at least about two years after the administration of the first dose.

125. The method according to any one of claims 113 to 124, wherein multiple booster doses are administered to the subject.

126. The method according to claim 125, wherein the first booster dose is administered at least about one year or at least about two years after the administration of the first dose.

127. The method according to claim 125 or claim 126, wherein the booster dose is administered once a year.

128. The method according to any one of claims 113 to 127, wherein the subject is a human subject.

129. The method according to claim 128, wherein the human subject is at least 18 years of age.

130. The method according to claim 128 or claim 129, wherein the human subject is between 18 and 70 years of age.

131. The method according to claim 128, wherein the human subject is under 18 years of age.

132. The method according to claim 128 or claim 131, wherein the human subject is between 2 and 5 years of age.

133. The method according to claim 128 or claim 131, wherein the human subject is between 5 and 18 years of age.

134. Use of the immunogenic composition according to any one of claims 1 to 112 for stimulating an immune response to Borrelia in a subject by administering an effective amount of the immunogenic composition to the subject.

135. Use of the immunogenic composition according to any one of claims 1 to 112 for protecting a subject from infection by Borrelia by administering an effective amount of the immunogenic composition to the subject.

136. Use of the immunogenic composition according to any one of claims 1 to 112 for preventing a subject from contracting Lyme disease by administering an effective amount of the immunogenic composition to the subject.

137. Use of an immunogenic composition according to any one of claims 1 to 112 in the manufacture of a pharmaceutical product for stimulating an immune response to Borrelia in a subject.

138. Use of the immunogenic composition according to any one of claims 1 to 112 in the manufacture of a pharmaceutical product for protecting a subject from infection by Borrelia.

139. Use of an immunogenic composition according to any one of claims 1 to 112 in the manufacture of a pharmaceutical product for preventing a subject from contracting Lyme disease.

140. The use according to any one of claims 134 to 139, wherein the immunogenic composition is administered by intramuscular injection.

141. The use according to any one of claims 134 to 140, wherein a first dose and a second dose of the immunogenic composition are administered to a subject.

142. The use according to claim 141, wherein a second dose of the immunogenic composition is administered approximately two weeks to approximately ten weeks after the administration of a first dose of the immunogenic composition.

143. The use according to claim 141 or claim 142, wherein a second dose of the immunogenic composition is administered about one month or about two months after the administration of a first dose of the immunogenic composition.

144. The use according to any one of claims 141 to 143, wherein a third dose of the immunogenic composition is administered to the subject.

145. The use according to claim 144, wherein a third dose of the immunogenic composition is administered approximately four to eight months after the administration of a first dose of the immunogenic composition.

146. The use according to claim 144 or claim 145, wherein a third dose of the immunogenic composition is administered about six months after the administration of a first dose of the immunogenic composition.

147. The use according to any one of claims 134 to 146, wherein a booster dose of the immunogenic composition is administered to the subject.

148. The use according to claim 147, wherein the booster dose is administered at least about one year or at least about two years after the administration of the first dose.

149. The use according to any one of claims 134 to 148, wherein multiple booster doses are administered to the subject.

150. The use according to claim 149, wherein the first booster dose is administered at least about one year or at least about two years after the administration of the first dose.

151. The use according to claim 149 or claim 150, wherein the booster dose is administered once a year.

152. The use according to any one of claims 133 to 151, wherein the subject is a human subject.

153. The use according to claim 152, wherein the human subject is at least 18 years of age.

154. The use according to claim 152 or claim 153, wherein the human subject is between 18 and 70 years of age.

155. The use according to claim 152, wherein the human subject is under 18 years of age.

156. The use according to claim 152 or claim 155, wherein the human subject is between 2 and 5 years of age.

157. The use according to claim 152 or claim 155, wherein the human subject is between 5 and 18 years of age.

158. A vial comprising the immunogenic composition according to any one of claims 1 to 112.

159. The vial according to claim 158, wherein the immunogenic composition is in liquid form.

160. The vial according to claim 158, wherein the immunogenic composition is in a lyophilized form.

161. A vial according to any one of claims 158 to 160, wherein the vial contains a single dose of an immunogenic composition.

162. The vial according to claim 161, wherein the single dose is 0.3 mL to 1.5 mL.

163. A pre-filled syringe comprising the immunogenic composition according to any one of claims 1 to 112.

164. The pre-filled syringe according to claim 163, wherein the syringe contains a single dose of an immunogenic composition.

165. A pre-filled syringe according to claim 164, wherein the single dose is 0.3 mL to 1.5 mL.

166. a) A first composition comprising an adjuvant selected from aluminum salt adjuvants; Toll-like receptor 9 (TLR9) agonists; liposome-based adjuvants containing lipopolysaccharides and saponins; and oil-in-water emulsion-based adjuvants containing squalene, sorbitan esters, and polyoxyethylene sorbitan esters; b) A second composition comprising Borrelia cell surface protein A (OspA) antigen; and c) Instructions for preparing the immunogenic composition according to any one of claims 1 to 112 by combining the first composition and the second composition. A kit that includes this.

167. The kit according to claim 166, wherein the second composition is in liquid form.

168. The kit according to claim 166, wherein the second composition is in a freeze-dried form.

169. The kit according to any one of claims 166 to 168, wherein the instructions are for preparing a single-dose immunogenic composition.

170. The kit according to any one of claims 166 to 169, wherein the kit is for a single-dose immunogenic composition.

171. The kit according to claim 169 or claim 170, wherein the single dose is 0.3 mL to 1.5 mL.

172. The kit according to any one of claims 166 to 171, wherein the first and / or second composition comprises a second adjuvant.

173. The kit according to claim 172, wherein the second adjuvant is an aluminum salt adjuvant.

174. The kit according to claim 173, wherein the aluminum salt adjuvant comprises one or more of amorphous aluminum hydroxyphosphate, aluminum hydroxide, aluminum phosphate, and potassium aluminum sulfate.

175. The kit according to claim 173 or claim 174, wherein the aluminum salt adjuvant comprises aluminum hydroxide.

176. d) A further set of instructions for administering immunogenic compositions to subjects to stimulate an immune response to Borrelia in the subjects. A kit according to any one of claims 166 to 175, further comprising:

177. a) The immunogenic composition according to any one of claims 1 to 112; and b) Instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject. A kit that includes this.

178. The kit according to claim 176 or claim 177, further comprising a syringe for intramuscular injection of an immunogenic composition.

179. The kit according to claim 178, wherein the syringe is pre-filled with an immunogenic composition.

180. a) the vial according to any one of claims 158 to 162; and b) Instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject. A kit that includes this.

181. The kit according to claim 180, further comprising a syringe for intramuscular injection of an immunogenic composition.

182. a) A pre-filled syringe according to any one of claims 163 to 165; and b) Instructions for administering an immunogenic composition to a subject to stimulate an immune response to Borrelia in the subject. A kit that includes this.

183. The kit according to claim 182, wherein the syringe is for intramuscular injection of an immunogenic composition.

184. The kit according to any one of claims 176 to 183, wherein administration is performed according to the method described in any one of claims 124 to 144.

185. A kit according to any one of claims 176 to 184, wherein the subject is a human subject.

186. The kit according to claim 185, wherein the human subject is at least 18 years of age.

187. The kit according to claim 185 or claim 186, wherein the human subject is between 18 and 70 years of age.

188. The kit according to claim 185, wherein the human subject is under 18 years of age.

189. The kit according to claim 185 or claim 188, wherein the human subject is between 2 and 5 years of age.

190. The kit according to claim 185 or claim 188, wherein the human subject is between 5 and 18 years of age.

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