Composition for use in treatment of acne

Abstract

The present invention relates to compositions (e.g., immunogenic compositions) useful for vaccination against dermatobacterium acnes. These compositions comprise dermatobacterium acnes antigens and combinations of antigens for use in the form of nucleic acids (e.g., mRNA) encoding antigen proteins or in the form of recombinant protein antigens.

Description

Technical Field

[0001] This invention belongs to the following field: treatment and prevention of acne-causing bacteria ( Cutibacterium acnes , C. acnes Infections such as acne vulgaris. In particular, the present invention relates to antigens and combinations of antigens that can be used for immunization against Propionibacterium acnes, in the form of nucleic acids (e.g., mRNA) encoding antigen proteins or in the form of recombinant protein antigens. Background Technology

[0002] *Propionibacterium acnes* (formerly known as *Propionibacterium acnes*) is a Gram-positive bacterium best known for its role in skin conditions such as acne vulgaris. It is a symbiotic bacterium of the skin, primarily found within sebaceous follicles, which provide a unique, lipid-rich environment due to sebum secretion. In addition to the skin, *Propionibacterium acnes* is also found in the conjunctiva, respiratory tract, urogenital tract, and gastrointestinal tract of humans and other animals.

[0003] Acne is a skin disorder affecting the hair follicles and sebaceous glands, impacting over 85% of adolescents, with over 20% of the population still experiencing symptoms after adolescence. Acne vulgaris presents with varying degrees of severity: mild, moderate, and severe. Moderate and severe acne account for more than one-third of all cases and require medication. Acne can also develop after puberty, as an adult-onset condition often associated with hormonal fluctuations, which are more prevalent in women. *Propionibacterium acnes* is also believed to play a role in other severe types of acne, such as acne conglobata, acne fulminans, and cystic acne. In addition to dermatopathology, *Propionibacterium acnes* is also present in corneal ulcers and is a common cause of chronic endophthalmitis after cataract surgery. Various other inflammatory diseases have been associated with *Propionibacterium acnes*, including post-operative implant and device-related infections (implant-associated infections), endocarditis, sarcoidosis, osteomyelitis, allergic alveolitis, pulmonary vasculitis, SAPHO syndrome (synovitis, acne, impetigo, osteoproliferative disorders, osteitis), and lumbar nerve root inflammation leading to sciatica. Recent studies suggest that *Propionibacterium acnes* also has a potential pathogenic role in non-infectious diseases such as prostate cancer, as its ability to persist intracellularly may lead to alterations in gene expression.

[0004] Phylogenetic studies based on multilocus gene sequencing and whole-genome analysis of Human Microbiome Project (HMP) isolates have provided valuable insights into the genetic population structure of *Propionibacterium acnes*. Multilocus sequence typing (MLST) generation is defined as the generation of sequence types (STs) for each *Propionibacterium acnes* isolate using different sequences of different alleles. Two different MLST typing schemes have been developed, classifying *Propionibacterium acnes* into closely related clusters IA1, IA2, IB, IC, II, and III (McDowell et al., 2012) or I-1a, I-1b, I-2, II, and III (Lomholt and Kilian, 2010) according to the MLST8 or MLST9 typing schemes, respectively. Some have suggested that treatment should target only IA1 strains (McLaughlin et al. 2019 – p. 23; O'Neill and Gallo 2018 – p. 4, second column, end of the first long paragraph), as these are the most commonly isolated from the skin of acne patients, while others can be found on acne-prone and healthy skin. However, other phyla have also been shown to be associated with acne (WO 2021 / 165543). For example, strains of *Propionibacterium acnes* from phyla IA2, IC, or II have been isolated from inflammatory lesions in human subjects (WO 2021 / 165543).

[0005] Propionibacterium acnes strains can be grouped using genome sequencing of 16S rDNA sequences called ribotypes (RTs). This system allows for comparison of populations of Propionibacterium acnes strains within individuals based on 16S rDNA sequences. The top 10 major ribotypes were highly abundant, and a large number of rare ribotypes were also identified. Analysis of the top 10 ribotypes can identify disease-specific and health-specific associations (Fitz-Gibbon et al. 2013; Tomida). et al. 2013; McLaughlin et al. 2019). The three most abundant ribotypes (RT1, RT2, and RT3) were fairly evenly distributed in acne-prone and normal individuals. However, ribotypes RT4, RT5, RT7, RT8, RT9, and RT10 were predominantly found in acne-prone individuals, while RT6 was closely associated with normal skin. A phylogenetic tree based on unique single nucleotide polymorphism positions in the core genome obtained from these 71 *Propionibacterium acnes* genomes showed that 16S rDNA ribotypes largely represent lineage relationships and indicated that 16S rDNA sequences are useful molecular markers for distinguishing major *Propionibacterium acnes* lineages (Fitz-Gibbon et al. 2013; Tomida). et al. (2013).

[0006] Current treatments target one or two steps in the pathogenesis of acne, including benzoyl peroxide, topical retinoids, and topical or oral antibiotics. However, antibiotic resistance is a problem for acne. Furthermore, oral isotretinoin is recommended for more severe cases, but it can cause adverse reactions, the most significant of which is teratogenicity. An effective vaccine against Propionibacterium acnes is needed. Summary of the Invention

[0007] The inventors have provided *Dermatophyte acnes* antigens and combinations thereof that can be used to provide immune resistance to *Dermatophyte acnes*.

[0008] In particular, the inventors have demonstrated that antigens CAMP2, DsA1, DsA2 and / or PITP peptides derived from *Propionibacterium acnes*, individually or in combination, elicit an antibody response when delivered by mRNA encoding the relevant antigens or in the form of recombinant peptides.

[0009] Therefore, the present invention provides *Propionibacterium acnes* CAMP2 polypeptide, modified *Propionibacterium acnes* CAMP2 polypeptide, *Propionibacterium acnes* DsA1 polypeptide, *Propionibacterium acnes* DsA2 polypeptide, *Propionibacterium acnes* PITP polypeptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide, and chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide, as well as nucleic acids comprising nucleotide sequences encoding such polypeptides.

[0010] The polypeptide antigens described herein may be delivered, for example, in the form of nucleic acids (e.g., mRNA) containing nucleotide sequences encoding the polypeptide, or, for example, in the form of recombinant proteins.

[0011] CAMP2 The Christie–Atkins–Munch–Petersen assay (CAMP) factor 2, or CAMP2, is a polypeptide belonging to the CAMP factor superfamily, whose members are known for their co-hemolytic and cytotoxic properties. *Propionibacterium acnes* CAMP factors 1–5 (or CAMP1–5) are considered pathogenic agents in the co-hemolytic reactions of *Propionibacterium acnes* with both sheep and human erythrocytes (Choudhury, 1978, Sörensen et al., Journal of Microbiological Methods (2010) 83(2):211–216). The amino acid sequences of *Propionibacterium acnes* CAMP factors 1–5 exhibit a low degree of sequence identity (e.g., approximately 50% for CAMP2 and CAMP4, and even lower for other amino acid sequences). The native *Propionibacterium acnes* CAMP2 polypeptide is a secreted protein.

[0012] CAMP2 has been identified as a putative acne-associated virulence factor (Holland et al. (2010)). The expression and secretion of CAMP protein by different *Propionibacterium acnes* strains are considered to be one of the factors contributing to *Propionibacterium acnes*' resistance to opsonization and phagocytosis. CAMP2 has been reported to induce phagocytosis in in vitro assays involving co-culture with *Propionibacterium acnes* (Wang et al. 2018). Furthermore, in a mouse ear model, anti-CAMP2 neutralizing antibodies showed a significant reduction in *Propionibacterium acnes*-induced inflammation (Liu et al. 2011, *Vaccine*, 29: 3230-3238).

[0013] The inventors have recognized that the *Propionibacterium acnes* CAMP2 peptide can be used to elicit an immune response against *Propionibacterium acnes* infection. Specifically, the inventors have demonstrated that the *Propionibacterium acnes* CAMP2 peptide elicits an antibody (e.g., IgG) response. Notably, the *Propionibacterium acnes* CAMP2 peptide elicits an antibody (e.g., IgG) response when delivered in the form of mRNA encoding the antigen as well as a recombinant protein. Furthermore, the inventors have demonstrated that when the mRNA encoding the *Propionibacterium acnes* CAMP2 peptide is delivered in lipid nanoparticles (LNPs), an antibody (e.g., IgG) response is elicited. The antibodies induced by the *Propionibacterium acnes* CAMP2 peptide reduce the co-hemolytic activity of CAMP2. Therefore, the *Propionibacterium acnes* CAMP2 peptide can elicit antibodies in subjects that can, for example, neutralize the biological activity of the CAMP2 peptide, such as antibodies that neutralize the inflammatory activity of CAMP2.

[0014] Therefore, the inventors have demonstrated that the *Propionibacterium acnes* CAMP2 peptide (e.g., delivered as mRNA encoding the antigen or as a recombinant protein) is a suitable vaccine antigen that can be used alone, in combination with other *Propionibacterium acnes* antigens described herein (e.g., one or more of the *Propionibacterium acnes* DsA1 peptide, *Propionibacterium acnes* DsA2 peptide, *Propionibacterium acnes* PITP peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide, and chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide (e.g., delivered as mRNA encoding the antigen or as a recombinant protein)), or as a chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide). The CAMP2 peptide can elicit antibodies that neutralize the inflammatory activity of CAMP2, while immunization with one or more of these other *Propionibacterium acnes* antigens can elicit antibodies that bind to the surface of *Propionibacterium acnes* bacteria and recruit effector cells (e.g., effector cells of the immune system, such as phagocytes).

[0015] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a *Propionibacterium acnes* CAMP2 polypeptide. Typically, the nucleic acid is messenger RNA (mRNA). In a preferred embodiment, the nucleic acid is a non-naturally occurring nucleic acid, such as non-naturally occurring mRNA. Therefore, the nucleic acid can be a synthetic nucleic acid (e.g., synthetic mRNA). In another aspect, the present invention provides a polypeptide comprising the amino acid sequence of a *Propionibacterium acnes* CAMP2 polypeptide. The *Propionibacterium acnes* CAMP2 polypeptide used in the present invention, delivered, for example, as mRNA and / or in an LNP, can induce antibodies in a subject. Such antibodies can neutralize the biological activity of the CAMP2 polypeptide, such as CAMP2 inflammatory activity. Such antibodies can neutralize the co-hemolytic activity of the CAMP2 polypeptide. Neutralization of CAMP2 inflammatory activity can be determined by a reduction in pro-inflammatory cytokines such as IL-1β and / or IL-6. Neutralization of co-hemolytic activity can be determined by an in vitro co-hemolysis assay.

[0016] The amino acid sequence of the natural *Propionibacterium acnes* CAMP2 antigen is provided in SEQ ID NO: 202:

[0017] The natural signal peptide sequence corresponds to residues 1-28 in the amino acid sequence of *Propionibacterium acnes* CAMP2 (SEQ ID NO: 202), and is shown underlined. Residues 29-267 of SEQ ID NO: 202 correspond to the mature form of the full-length *Propionibacterium acnes* CAMP2 polypeptide (without the natural signal peptide sequence).

[0018] The N-terminal and C-terminal domains of *Propionibacterium acnes* CAMP2 are shown in bold (N-terminal domain) and bold underlined (C-terminal domain) text. The N-terminal domain of the *Propionibacterium acnes* CAMP2 polypeptide corresponds to amino acid residues 29-176 of the *Propionibacterium acnes* CAMP2 polypeptide in SEQ ID NO: 202. The C-terminal domain of the *Propionibacterium acnes* CAMP2 polypeptide corresponds to amino acid residues 189-267 of the *Propionibacterium acnes* CAMP2 polypeptide in SEQ ID NO: 202. The linker domain is located between the N-terminal and C-terminal domains. The linker domain corresponds to amino acid residues 177-188 of SEQ ID NO: 202.

[0019] "Propionibacterium acnes CAMP2 polypeptide" includes the mature form of the full-length natural Propionibacterium acnes CAMP2 polypeptide without its natural signal peptide sequence and its immunogenic variants. Immunogenic variants of the natural Propionibacterium acnes CAMP2 polypeptide are capable of eliciting an immune response in subjects (e.g., an antigen-specific immune response), such as an antibody response. Immunogenic variants of the natural Propionibacterium acnes CAMP2 polypeptide include immunogenic fragments of the natural Propionibacterium acnes CAMP2 polypeptide. Immunogenic fragments of Propionibacterium acnes CAMP2 include natural Propionibacterium acnes CAMP2 polypeptides at least 50, 75, 100, 125, 150, 175, 200, or 225 amino acids in length.

[0020] Exemplary *Propionibacterium acnes* CAMP2 polypeptide sequences include the native *Propionibacterium acnes* CAMP2 polypeptide sequence in SEQ ID NO: 203, which lacks the native secretion signal peptide sequence of the *Propionibacterium acnes* CAMP2 polypeptide. Other exemplary *Propionibacterium acnes* polypeptide sequences are listed in SEQ ID NOs 313-338. These are naturally occurring CAMP2 polypeptides from different *Propionibacterium acnes* strains and include the native secretion signal peptide sequence of the *Propionibacterium acnes* CAMP2 polypeptide. Corresponding sequences of CAMP2 polypeptides are provided in SEQ ID NOs 339-363, but without the native secretion signal peptide sequence.

[0021] Analysis of 430 CAMP2 polypeptide sequences from naturally occurring *Dermatophyte acnes* strains revealed only 27 distinct CAMP2 sequences (SEQ ID NO 202 and 313 to 338) when the native secretion signal peptide sequence was included in the alignment, and only 26 distinct CAMP2 sequences (SEQ ID NO 203 and 339 to 363) when the native secretion signal peptide sequence was excluded. Sequence variations among these CAMP2 polypeptides were concentrated at certain residues (Figure 44 and Table 1).

[0022] Table 1. Analysis of naturally occurring cAMP2 peptide variants

[0023]

[0024] * SEQ ID NO: 203 is the CAMP2 sequence of strain KPA171202, which does not contain a secreted signal peptide sequence.

[0025] If multiple CAMP2 variants are found at a specific location, all substituted residues are indicated by " / ", and their corresponding frequencies are also indicated by " / " in the last two columns.

[0026] In some embodiments, the *Propionibacterium acnes* CAMP2 polypeptide comprises amino acid substitutions at one or more positions corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221, and / or 223 of SEQ ID NO: 203. In some embodiments, the *Propionibacterium acnes* CAMP2 polypeptide comprises amino acid substitutions at one or more positions corresponding to the following: SEQ ID NO: Residues of 203 include T6 (e.g., T6I), A9 (e.g., A9T), S11 (e.g., S11A), S18 (e.g., S18N), D19 (e.g., D19E or D19Y), R21 (e.g., R21H), I24 (e.g., I24M, I24L, or I24T), A29 (e.g., A29P), H30 (e.g., H30R), V37 (e.g., V37A), D48 (e.g., D48N), R61 (e.g., R61H), E65 (e.g., E65D), A68 (e.g., A68T), D76 (e.g., D76N), V87 (e.g., V87A), I91 (e.g., I91V), D92 (e.g., D92G), and T98 (…). For example, T98K), T100 (e.g., T100I), R106 (e.g., R106S), K118 (e.g., K118N), S128 (e.g., S128T), A138 (e.g., A138T), R143 (e.g., R143H), E145 (e.g., E145D or E145K), T154 (e.g., T154A), K169 (e.g., K169R), N177 (e.g., N177D), D179 (e.g., D179N or D179H), A189 (e.g., A189E), N207 (e.g., N207D or N207A), E221 (e.g., E221K), and / or L223 (e.g., L223F).

[0027] In some embodiments, the dermatitis acnes CAMP2 polypeptide comprises a sequence of any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16 or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203) or a sequence having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. Typically, sequence variations are located at the positions listed above, that is, the positions corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221 and / or 223 of SEQ ID NO: 203. Therefore, in some embodiments, the *Dermatitis acnes* CAMP2 polypeptide comprises amino acid substitutions at one or more positions corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221, and / or 223 of SEQ ID NO: 203. In some embodiments, the *Dermatitis acnes* CAMP2 polypeptide comprises a sequence having at least 90% identity with any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203). In some embodiments, the dermatitis acnes CAMP2 polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16 or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203).Typically, the *Propionibacterium acnes* CAMP2 polypeptide comprises a sequence having at least 85% identity with any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203). In some embodiments, the *Propionibacterium acnes* CAMP2 polypeptide comprises, is substantially composed of, or is composed of (e.g., includes) the amino acid sequence according to SEQ ID NO: 203.

[0028] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the *Dermatitis acnes* CAMP2 polypeptide comprises a nucleotide sequence according to any one of SEQ ID NO: 153-167, SEQ ID NO: 87-89, or SEQ ID NO: 95-101, or a sequence having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprises a nucleotide sequence at least 75% identical to any one of SEQ ID NO: 153-167, SEQ ID NO: 87-89, or SEQ ID NO: 95-101.

[0029] Modified CAMP2 The inventors have produced a modified *Propionibacterium acnes* CAMP2 polypeptide comprising a *Propionibacterium acnes* CAMP2 polypeptide sequence and a heterologous transmembrane domain (TMB) sequence. Including the heterologous TMB sequence is advantageous because such sequences can localize the antigen to the cell membrane when the antigen is expressed in cells (e.g., in eukaryotic cells, such as cells in the subject). Containing the heterologous TMB sequence reduces intracellular antigen localization compared to antigens without the TMB sequence. The inventors have demonstrated that such modified *Propionibacterium acnes* CAMP2 polypeptides containing the TMB domain sequence elicit an antibody (e.g., IgG) response. Antibodies induced by the modified *Propionibacterium acnes* CAMP2 polypeptide reduce the co-hemolytic activity of the *Propionibacterium acnes* CAMP2 polypeptide. The *Propionibacterium acnes* CAMP2 polypeptide can elicit antibodies in the subject that can, for example, neutralize the biological activity of the CAMP2 polypeptide, such as antibodies that neutralize the inflammatory activity of CAMP2.

[0030] Furthermore, the modified *Propionibacterium acnes* CAMP2 peptide elicited a higher antibody titer compared to the *Propionibacterium acnes* CAMP2 peptide lacking the TMB domain. Therefore, including the TMB sequence can enhance the immunogenicity (e.g., antibody response) of the *Propionibacterium acnes* CAMP2 peptide in subjects. The inclusion of the TMB sequence in the modified *Propionibacterium acnes* CAMP2 peptide can elicit a stronger antibody response, for example, a higher antibody titer in subjects compared to the CAMP2 peptide without the TMB sequence. Surface-expressed antigens (e.g., the modified *Propionibacterium acnes* CAMP2 peptide) can enhance B cell activation in subjects, resulting in enhanced antigen-specific B cell responses (e.g., enhanced antigen-specific antibody responses, such as higher antibody titers).

[0031] Therefore, the inventors have demonstrated that these modified *Dermatitis acnes* CAMP2 peptides are suitable vaccine antigens that can be used alone or in combination with other *Dermatitis acnes* antigens described herein, such as *Dermatitis acnes* DsA1 peptide, *Dermatitis acnes* DsA2 peptide, *Dermatitis acnes* PITP peptide, chimeric *Dermatitis acnes* DsA1 / DsA2 peptide, and chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP peptide. These modified *Dermatitis acnes* CAMP2 peptides, when used alone or in combination with one or more other antigens described herein, can elicit an immune response against infection. The modified *Dermatitis acnes* CAMP2 peptides described herein can be delivered via nucleic acids containing a nucleotide sequence encoding the modified *Dermatitis acnes* CAMP2 peptide.

[0032] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a modified *Propionibacterium acnes* CAMP2 polypeptide, wherein the modified *Propionibacterium acnes* CAMP2 polypeptide comprises an amino acid sequence including the *Propionibacterium acnes* CAMP2 polypeptide sequence and a heterologous transmembrane domain (TMB) sequence. When an antigen is expressed in a cell (e.g., a eukaryotic cell, such as a cell in a subject), the heterologous TMB sequence is able to localize the antigen to the cell membrane. This can enhance the immunogenicity (e.g., antibody response, such as antibody titer) induced in a subject, for example, in a subject lacking an antigen with a TMB sequence.

[0033] In another aspect, the present invention provides a polypeptide comprising the amino acid sequence of a modified dermatitis acnes CAMP2 polypeptide.

[0034] In some embodiments, the heterologous transmembrane domain sequence is a TMB sequence of a eukaryotic transmembrane polypeptide (eukaryotic transmembrane sequence), a TMB sequence of a prokaryotic transmembrane polypeptide (prokaryotic transmembrane sequence), or a TMB sequence of a viral transmembrane protein (viral transmembrane domain sequence). In some embodiments, the heterologous transmembrane domain is a TMB sequence of a viral transmembrane protein.

[0035] In some embodiments, the heterologous TMB sequence is located at the N-terminus of the modified *Propionibacterium acnes* CAMP2 polypeptide. In other embodiments, the heterologous TMB sequence is located at the C-terminus of the modified *Propionibacterium acnes* CAMP2 polypeptide.

[0036] Typically, the nucleic acid encoding the modified *Dermatophyte acnes* CAMP2 polypeptide containing a transmembrane domain, as described herein, also contains a nucleotide sequence encoding a secretion signal peptide sequence.

[0037] Further details regarding the TMB structural domains applicable to this invention are provided below. Exemplary TMB sequences are described below.

[0038] In some embodiments, the heterologous transmembrane domain comprises a hemagglutinin (HA) transmembrane domain sequence from influenza A virus or influenza B virus (preferably from influenza A virus).

[0039] In some embodiments, the transmembrane domain (TMB) sequence is directly fused to the *Propionibacterium acnes* CAMP2 peptide described herein (i.e., in the modified *Propionibacterium acnes* CAMP2 peptide described herein, there is no linker, such as an amino acid linker, connecting the TMB sequence to the *Propionibacterium acnes* CAMP2 peptide). In other embodiments of the modified *Propionibacterium acnes* CAMP2 peptide described herein, the disclosed TMB sequence is attached to the *Propionibacterium acnes* CAMP2 peptide described herein via a linker. Preferred linkers are listed in the “Linkers” section below.

[0040] In some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises a *Propionibacterium acnes* CAMP2 polypeptide sequence comprising a sequence according to any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203), or a sequence having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. Typically, variations in the *Propionibacterium acnes* CAMP2 polypeptide sequence may be located at the positions listed above. Therefore, the CAMP2 polypeptide sequence variation of *Propionibacterium acnes* can typically be located at positions corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221 and / or 223 of SEQ ID NO: 203 as listed above herein. In some embodiments, the CAMP2 polypeptide sequence of *Dermatitis acnes* comprises amino acid substitutions at one or more positions corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221 and / or 223 of SEQ ID NO: 203.In some embodiments, the *Dermatitis acnes* CAMP2 polypeptide comprises amino acid substitutions at one or more positions corresponding to the following: residues of SEQ ID NO:203: T6 (e.g., T6I), A9 (e.g., A9T), S11 (e.g., S11A), S18 (e.g., S18N), D19 (e.g., D19E or D19Y), R21 (e.g., R21H), I24 (e.g., I24M, I24L or I24T), A29 (e.g., A29P), H30 (e.g., H30R), V37 (e.g., V37A), D48 (e.g., D48N), R61 (e.g., R61H), E65 (e.g., E65D), A68 (e.g., A68T), D76 (e.g., D76N), V87 (e.g., V87A), I91 (e.g., I91V), D92 (e.g., D92G), T98 (e.g., T98K), T100 (e.g., T100I), R106 (e.g., R106S), K118 (e.g., K118N), S128 (e.g., S128T), A138 (e.g., A138T), R143 (e.g., R143H), E145 (e.g., E145D or E145K), T154 (e.g., T154A), K169 (e.g., K169R), N177 (e.g., N177D), D179 (e.g., D179N or D179H), A189 (e.g., A189E), N207 (e.g., N207D or N207A), E221 (e.g., E221K), and / or L223 (e.g., L223F). In some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises a *Propionibacterium acnes* CAMP2 polypeptide sequence having at least 90% identity with any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203). In some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises a *Propionibacterium acnes* CAMP2 polypeptide sequence having at least 95% identity with any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203).Typically, the modified *Propionibacterium acnes* CAMP2 polypeptide contains the *Propionibacterium acnes* CAMP2 polypeptide sequence, which contains a sequence having at least 85% identity with any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16 or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203).

[0041] In some embodiments, the modified *Dermatitis acnes* CAMP2 polypeptide comprises a sequence according to any one of SEQ ID NO: 207 or SEQ ID NO: 5-9 (e.g., SEQ ID NO: 207) or a sequence having at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. Typically, sequence variations are located at the positions described above, corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221 and / or 223 of SEQ ID NO: 207. Therefore, in some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises amino acid substitutions at one or more positions corresponding to residues 6, 9, 11, 18, 19, 21, 24, 29, 30, 37, 48, 61, 65, 68, 76, 87, 91, 92, 98, 100, 106, 118, 128, 138, 143, 145, 154, 169, 177, 179, 189, 207, 221, and / or 223 in SEQ ID NO: 207. In some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises a sequence having at least 90% identity with any one of SEQ ID NO: 203, SEQ ID NO: 207, SEQ ID NO: 43-58, SEQ ID NO: 1-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 207). In some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO: 203, SEQ ID NO: 207, SEQ ID NO: 43-58, SEQ ID NO: 1-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 207). In some embodiments, the modified *Propionibacterium acnes* CAMP2 polypeptide comprises a sequence having at least 85% identity with any one of SEQ ID NO: 203, SEQ ID NO: 207, SEQ ID NO: 43-58, SEQ ID NO: 1-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 207).

[0042] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the modified dermatitis acnes CAMP2 polypeptide comprises a nucleotide sequence according to any one of SEQ ID NO: 153-167, SEQ ID NO: 87-89 or SEQ ID NO: 95-101 or having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the modified *Dermatitis acnes* CAMP2 polypeptide comprises a nucleotide sequence according to any one of SEQ ID NO: 90-94 or SEQ ID NO: 391-392 (e.g., SEQ ID NO: 91) or having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprises a nucleotide sequence that is at least 75% or 85% (e.g., 75%) identical to any one of SEQ ID NO: 153-167, SEQ ID NO: 391-392, or SEQ ID NO: 87-101 (e.g., SEQ ID NO: 91).

[0043] The sequences of SEQ ID NO: 90, 91, 391, and 392 were compared. Variations were found at the positions shown in SEQ ID NO: 398 and 399.In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the modified *Dermatitis acnes* CAMP2 polypeptide comprises the nucleotide sequence according to SEQ ID NO: 398, wherein the nucleotides at positions 66; 78; 81; 87; 102; 120; 150; 222; 232; 234; 261; 298; 321; 339; 351; 367; 420; 423; 435; 438; 456; 462; 468; 478; 516; 528; 541; 546; 574; 583; 648; 664; 684; 706; 711; 735; 834 are each independently selected from A or C; the nucleotides at positions 480; 660; 849 are each independently selected from A, C, or G; and the nucleotides at positions 171; 282; 318; 375 are each independently selected from A, C, or G. The nucleotides at positions 432, 675, and 738 are each independently selected from A, C, or T; the nucleotides at positions 6, 57, 93, 114, 168, 246, 300, 363, 384, 396, 405, 447, 459, 543, 585, 591, 609, 666, 708, 714, and 857 are each independently selected from A or G; the nucleotides at positions 73, 97, 103, 118, 169, 181, 190, 283, 433, 493, 553, 745, 769, 775, 793, 805, 808, 826, 841, and 850 are each independently selected from A or T; The nucleotides at positions 18; 27; 54; 98; 119; 153; 162; 170; 182; 191; 210; 213; 240; 264; 284; 312; 369; 390; 408; 434; 441; 483; 494; 554; 573; 603; 690; 746; 770; 774; 776; 794; 806; 809; 827; 842; 851; 855 are each independently selected from C or G; and the nucleotides at positions 30; 33; 36; 48; 51; 108; 111; 117; 123; 135; 177; 183; 186; 189; 207; ... The nucleotides at positions 219; 225; 243; 255; 258; 267; 270; 276; 279; 285; 297; 327; 372; 378; 402; 426; 429; 448; 453; 465; 477; 501; 507; 519; 522; 531; 537; 549; 564; 582; 594; 615; 624; 636; 672; 693; 696; 702; 729; 741; 750; 753; 771; 780; 795; 804; 807; 810; 828; 837; 840; and 852 are independently selected from C or T.In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the modified *Propionibacterium acnes* CAMP2 polypeptide comprises the nucleotide sequence according to SEQ ID NO: 399, wherein the nucleotides at positions 66; 81; 87; 102; 222; 232; 234; 261; 298; 321; 339; 351; 420; 438; 456; 462; 468; 478; 516; 528; 541; 546; 574; 583; 648; 660; 664; 684; 706; 711; 735; 834 are each independently selected from A or C; the nucleotide at position 849 is selected from A, C, or G; and the nucleotides at positions 318; 432; 675; 834 are each independently selected from A or C. The nucleotides at positions 6, 57, 93, 168, 300, 363, 384, 396, 405, 447, 459, 480, 543, 585, 609, 666, 708, 714, and 857 are each independently selected from A or G; the nucleotides at positions 73, 97, 103, 118, 169, 181, 283, 433, 493, 745, 769, 793, 805, 808, 826, 841, and 850 are each independently selected from A, C, or T. A or T; the nucleotides at positions 18; 27; 54; 98; 119; 153; 170; 182; 213; 264; 284; 312; 369; 390; 408; 434; 483; 494; 573; 603; 746; 770; 774; 794; 806; 809; 827; 842; 851; 855 are each independently selected from C or G; and the nucleotides at positions 30; 33; 36; 48; 117; 123; 135; 171; 177; 183; 186; 189; 2 The nucleotides at positions 25; 243; 255; 258; 267; 270; 276; 282; 285; 297; 327; 372; 375; 378; 426; 453; 465; 477; 501; 519; 522; 531; 549; 564; 582; 624; 636; 672; 693; 696; 738; 741; 750; 753; 771; 780; 795; 807; 810; 828; 837; 840; and 852 are each independently selected from C or T. Typically, the nucleic acid containing the nucleotide sequence according to SEQ ID NO: 398 or 399 encodes the sequence according to SEQ ID NO: 6.

[0044] In some embodiments, the nucleic acid comprises at least 75% or 85% (e.g., 75%) the same nucleotide sequence as any one of SEQ ID NO: 153-167, SEQ ID NO: 391-392 or SEQ ID NO: 87-101 (e.g., SEQ ID NO: 91).

[0045] In one embodiment, the nucleic acid of the present invention is an mRNA comprising or composed of the following structural elements (e.g., an mRNA composed of the following structural elements): (i) A 5' cap with the following structure:

[0046] (ii) Having a 5' untranslated region (5' UTR) of the nucleic acid sequence according to SEQ ID NO: 265; (iii) A protein-coding region having a nucleic acid sequence according to SEQ ID NO: 90, SEQ ID NO: 391-392 or SEQ ID NO: 91 (e.g., SEQ ID NO: 91); (iv) Possessing a 3' untranslated region (3' UTR) according to the nucleic acid sequence of SEQ ID NO: 266; and (v) A poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), for example, wherein the poly-A tail comprises at least 100 adenosine nucleotides.

[0047] In some embodiments, the mRNA is chemically modified, and the chemical modification includes replacing each uridine with N1-methylpseudouridine. The mRNA may be encapsulated in LNPs.

[0048] DsA1 Dermatobacterium acnes sulfated skin adhesin 1 (DsA1), also known as P22 or P022, has been identified as a putative virulence factor of Dermatobacterium acnes (Lodes et al., 2006 (Microbiology). Dec. 2006; 152(Pt 12):3667-3681; McDowell et al., 2011). This protein is abundant in both acne-affected and healthy hair follicle samples (Bek-Thomsen). et al. (2014). The DsA1 polypeptide, its derivatives and fragments of *Propionibacterium acnes* are described in WO 2021 / 165543.

[0049] The inventors have demonstrated that mRNA encoding different *Propionibacterium acnes* DsA1 peptides and recombinant *Propionibacterium acnes* DsA1 protein elicits antibody (e.g., IgG) responses. Antibodies induced by *Propionibacterium acnes* DsA1 peptides have been shown to bind to the surface of *Propionibacterium acnes* bacteria and recruit effector cells (e.g., effector cells of the immune system, such as phagocytes). Antibodies induced by *Propionibacterium acnes* DsA1 peptides have been shown to induce opsonization and phagocytosis in vitro.

[0050] The DsA1 polypeptide of the present invention can induce an immune response (cross-reactive immune response) against a variety of different strains, phylogens and disease-causing variants of the dermatitis acnes.

[0051] The inventors have demonstrated that the *Dermatitis acnes* DsA1 peptide is a suitable vaccine antigen that can be used in combination with one or more other *Dermatitis acnes* antigens that can elicit an antibody response in subjects, such as the *Dermatitis acnes* DsA2 peptide, the *Dermatitis acnes* PITP peptide, and the *Dermatitis acnes* CAMP2 peptide described herein. The *Dermatitis acnes* CAMP2 peptide can elicit antibodies in subjects, for example, antibodies that can neutralize the biological activity of the CAMP2 peptide, such as antibodies that exhibit the inflammatory activity of CAMP2.

[0052] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a *Propionibacterium acnes* DsA1 polypeptide. In another aspect, the present invention provides a polypeptide comprising an amino acid sequence of a *Propionibacterium acnes* DsA1 polypeptide. The *Propionibacterium acnes* DsA1 polypeptide used in the present invention, delivered, for example, as mRNA or a recombinant protein, can induce antibodies in a subject. Such antibodies can opsonize *Propionibacterium acnes* bacteria. Opsonization may target *Propionibacterium acnes* bacteria with immune effector cells (e.g., phagocytes). This may result in such immune effector cells killing *Propionibacterium acnes* bacteria (e.g., phagocytosis). The binding of antibodies to the cell surface of *Propionibacterium acnes* can be measured, for example, using an in vitro surface binding assay. An in vitro opsonization-phagocytosis assay can be used to assess the ability of antibodies to induce phagocytes to opsonize and kill *Propionibacterium acnes* strains of different genetic types.

[0053] The amino acid sequence (without signal peptide sequence) of the natural mature form of the full-length *Dermatophyte acnes* DsA1 polypeptide is provided in SEQ ID NO: 204: SSNRPRSVAQAAIATDGKGIIDKDCRDAVINDAKLRAAIAGALVKAGFSSADAVALAPRIAKEMAKEGV LLINHHKLKALIGAQLGLLTDAKIQRAAAAVDLGIKATLAATIIPNALHSAAFKDAVVANLVAAGVDKKLAKATAVA IAATALNPALGPIAKTEAIKAEIAAQAALLVGRGVHLKKAAIEHIIGRSFDAAVATAIVSSPILNARIVTHLVRAGI DKSLAVQIAPRIIDRLAKEPLLALNTAKLMKNITRQIVDVITADKAIKTAEQLEKELPALDDLVKKACSCPKPTPTP TPTPTPTPKPTPTPTPKPTPTPKPKPTPAPAPTSGATSDESTSRSGGHSQGGSGTHYIHHGVAPVLTHSSDLPSTGF Naturally matured *Dermatitis acnes* DsA1 polypeptide typically comprises an N-terminal exchange region (“NSR”), a first conserved subdomain (“CSD1”), a first exchange region (“SR1”), a second conserved subdomain (“CSD2”), a second exchange region (“SR2”), a third conserved subdomain (“CSD3”), a region containing a Pro-Thr repeat (“PT repeat region”—shown in bold above), and a C-terminal region (“CTR”; typically with an LXTG motif near the C-terminus). The C-terminal LXTG motif is considered essential for cell wall anchoring of the protein.

[0054] For example, in the native *Dermatobacterium acnes* DsA1 polypeptide of SEQ ID NO: 204, the N-terminal exchange region (“NSR”) corresponds to amino acid residues 1 to 20, the first conserved subdomain (“CSD1”) corresponds to amino acid residues 21 to 102, the first exchange region (“SR1”) corresponds to amino acid residues 103 to 119, the second conserved subdomain (“CSD2”) corresponds to amino acid residues 120 to 239, the second exchange region (“SR2”) corresponds to amino acid residues 240 to 249, the third conserved subdomain (“CSD3”) corresponds to amino acid residues 250 to 295, the region containing Pro-Thr repeats (“PT repeat region”) corresponds to amino acid residues 296 to 333, and the C-terminal region (“CTR”) corresponds to amino acid residues 334 to 377 (with an LXTG motif corresponding to amino acid residues 372 to 376).

[0055] The sequences of naturally occurring DsA1 peptides are largely highly identical, and differ only in the length and composition of the PT repeat region (with the rare exception of point mutations and, in particular, the presence of terminal LPTTG motifs). A more detailed sequence analysis of DsA1 has been described elsewhere (WO 2021 / 165543). As is known in the art, the positions of NSR, CSD1, SR1, CSD2, SR2, CSD3, PT repeat region, and CTR within other naturally occurring *Dermatophyte acnes* DsA1 peptides can be determined by aligning the peptide sequence of a given DsA1 peptide with that of SEQ ID NO: 204 and identifying regions in the DsA1 peptide that have high sequence identity with the sequences of NSR, CSD1, SR1, CSD2, SR2, CSD3, PT repeat region, and CTR of SEQ ID NO: 204, respectively.

[0056] "Propionibacterium acnes DsA1 polypeptide" includes the mature form of the full-length natural Propionibacterium acnes DsA1 polypeptide without its natural signal peptide sequence and its immunogenic variants. The immunogenic variants of the natural Propionibacterium acnes DsA1 polypeptide are capable of eliciting an immune response (e.g., an antigen-specific immune response), such as an antibody response, in a subject. The immunogenic variants of the natural Propionibacterium acnes DsA1 polypeptide comprise an immunogenic fragment of the natural Propionibacterium acnes DsA1 polypeptide. The immunogenic Propionibacterium acnes DsA1 fragment comprises a natural Propionibacterium acnes DsA1 polypeptide of at least 50, at least 75, at least 100, at least 125, at least 150, at least 175, at least 200, at least 225, at least 250, at least 275, at least 300, at least 325, or at least 350 amino acids in length. In some embodiments, the immunogenic Propionibacterium acnes DsA1 fragment contains a CSD2 domain of the Propionibacterium acnes DsA1 polypeptide. In some embodiments, immunogenic variants exclude sequence motifs found in the subject's (e.g., human) proteome, such as sequence motifs of 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, or 15 or more (e.g., 8 or more) amino acids in length. The excluded subject proteome sequence motifs are typically 8 or more amino acids in length. This helps to minimize unwanted cross-reactivity due to homology between the antigen and its own proteins.

[0057] An exemplary *Dermatophyte acnes* DsA1 polypeptide sequence includes the native *Dermatophyte acnes* DsA1 polypeptide sequence in SEQ ID NO: 204, which lacks the native secretion signal peptide sequence of the *Dermatophyte acnes* DsA1 polypeptide.

[0058] In some embodiments, the *Propionibacterium acnes* DsA1 polypeptide comprises a sequence of any one of SEQ ID NO: 204, SEQ ID NO: 17-19, or SEQ ID NO: 59-61, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the *Propionibacterium acnes* DsA1 polypeptide comprises a sequence having at least 90% identity with any one of SEQ ID NO: 204, SEQ ID NO: 17-19, or SEQ ID NO: 59-61. In some embodiments, the *Propionibacterium acnes* DsA1 polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO: 204, SEQ ID NO: 17-19, or SEQ ID NO: 59-61.

[0059] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the DsA1 polypeptide of *Propionibacterium acnes* comprises a nucleotide sequence according to any one of SEQ ID NO: 102-104 or SEQ ID NO: 168-170, or a sequence having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprises a nucleotide sequence at least 75% identical to any one of SEQ ID NO: 102-104 or SEQ ID NO: 168-170.

[0060] DsA^2 Dermatitis acnes sulfate dermatin-adhesion 2 (DsA2), also known as P27 or P027, has been identified as a putative virulence factor for *Propionibacterium acnes* (Lodes et al., 2006 (Microbiology). Dec. 2006; 152(Pt 12):3667-3681; McDowell et al., 2011). This protein has been found in both acne-affected and healthy hair follicle samples (Bek-Thomsen). et al.(2014). DsA1 and DsA2 proteins are homologs (paralogs), with typical sequence identity between 60% and 71%, depending on the aligned protein region. Excluding PT region length polymorphism, high (typically >90%) sequence identity can be observed in both intact DsA1 and DsA2 proteins. The DsA2 polypeptide, its derivatives, and fragments from *Propionibacterium acnes* are described in WO 2021 / 165543.

[0061] The inventors have demonstrated that mRNA encoding different *Propionibacterium acnes* DsA2 peptides and recombinant *Propionibacterium acnes* DsA2 protein elicits antibody (e.g., IgG) responses. Antibodies induced by *Propionibacterium acnes* DsA2 peptides have been shown to bind to the surface of *Propionibacterium acnes* bacteria and recruit effector cells (e.g., effector cells of the immune system, such as phagocytes). Antibodies induced by *Propionibacterium acnes* DsA2 peptides have been shown to induce opsonization and phagocytosis in vitro.

[0062] The DsA2 polypeptide of the present invention can induce an immune response (cross-reactive immune response) against a variety of different strains, phylogens and disease-causing variants of the dermatitis acnes.

[0063] The inventors have demonstrated that the *Dermatitis acnes* DsA2 peptide is a suitable vaccine antigen that can be used in combination with one or more other *Dermatitis acnes* antigens that can elicit an antibody response in subjects (e.g., the *Dermatitis acnes* DsA1 peptide, the *Dermatitis acnes* PITP peptide, and the *Dermatitis acnes* CAMP2 peptide described herein). The *Dermatitis acnes* CAMP2 peptide can elicit antibodies in subjects, for example, antibodies that can neutralize the biological activity of the CAMP2 peptide, such as antibodies that have inflammatory activity against CAMP2.

[0064] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* DsA2 polypeptide. In another aspect, the present invention provides a polypeptide comprising an amino acid sequence of a *Dermatitis acnes* DsA2 polypeptide. The *Dermatitis acnes* DsA2 polypeptide used in the present invention, delivered, for example, as mRNA or a recombinant protein, can induce antibodies in a subject. Such antibodies can opsonize *Dermatitis acnes* bacteria. Opsonization may target *Dermatitis acnes* bacteria with immune effector cells (e.g., phagocytes). This may result in such immune effector cells killing *Dermatitis acnes* bacteria (e.g., phagocytosis). The binding of antibodies to the cell surface of *Dermatitis acnes* can be measured, for example, using an in vitro surface binding assay. In vitro opsonization-phagocytosis assays can be used to assess the ability of antibodies to induce phagocytes to opsonize and kill *Dermatitis acnes* strains of different genetic types.

[0065] The amino acid sequence (without signal peptide sequence) of the natural mature form of the full-length *Dermatophyte acnes* DsA2 polypeptide is provided in SEQ ID NO: 205: ASNGNSSITQSAAFSPRATTKISEDCRKAIINDLKLRGAIVGALVKAGLSAADAAALAPRIAAEMAAEG TLTINHHRLKVLVASQLGLVADAAVQHAAAAIDLSFKAILGASIIPNALGSAAFKNAVIANLVAAGIDKHLARATAV AIVATALNPALGPIAKFELIKAEIAAQAALLIRRGVHLQKAAIEHVIGRAFDAAVATAIISSPILSARIVTHLVRAG IDKSIAISLAPHIVKRLAKEPLLAFNTAKLVKDIARQIVDIRNTQEAIAVYKQLKAELPTLDGLVQKACTPEPTPTP TPTPTPTPTPAPTPTPAPTPTPAPTPAPTPTPAPTPTPTPTPTPTPTPTHGATTTTPISRTTDRHNLGSHHTRIAAP Naturally matured *Dermatitis acnes* DsA2 polypeptide typically comprises an N-terminal exchange region (“NSR”), a first conserved subdomain (“CSD1”), a first exchange region (“SR1”), a second conserved subdomain (“CSD2”), a second exchange region (“SR2”), a third conserved subdomain (“CSD3”), a region containing a Pro-Thr repeat (“PT repeat”), and a C-terminal region (“CTR”; typically with an LXTG motif near the C-terminus). The C-terminal LXTG motif is considered essential for cell wall anchoring of the protein.

[0066] For example, in the native Dermatobacterium acnes DsA2 polypeptide of SEQ ID NO: 205, the N-terminal exchange region (“NSR”) corresponds to amino acid residues 1 to 21, the first conserved subdomain (“CSD1”) corresponds to amino acid residues 22 to 103, the first exchange region (“SR1”) corresponds to amino acid residues 104 to 120, the second conserved subdomain (“CSD2”) corresponds to amino acid residues 121 to 240, the second exchange region (“SR2”) corresponds to amino acid residues 241 to 250, the third conserved subdomain (“CSD3”) corresponds to amino acid residues 251 to 295, the region containing Pro-Thr repeats (“PT repeat region”) corresponds to amino acid residues 296 to 349, and the C-terminal region (“CTR”) corresponds to amino acid residues 350 to 392 (with an LXTG motif corresponding to amino acid residues 385 to 389).

[0067] The sequences of naturally occurring DsA2 peptides are largely highly identical, and differ only in the length and composition of the PT repeat region (except for rare exceptions such as point mutations and, in particular, the presence of terminal LPTTG motifs). A more detailed sequence analysis of DsA2 has been described elsewhere (WO2021 / 165543). As is known in the art, the positions of NSR, CSD1, SR1, CSD2, SR2, CSD3, the PT repeat region, and the CTR within other naturally occurring *Dermatophyte acnes* DsA2 peptides can be determined by aligning the peptide sequence of a given DsA2 peptide with the peptide sequence of SEQ ID NO: 205 and identifying regions in the DsA2 peptide that have high sequence identity with the sequences of NSR, CSD1, SR1, CSD2, SR2, CSD3, the PT repeat region, and the CTR of SEQ ID NO: 205, respectively.

[0068] "Propionibacterium acnes DsA2 polypeptide" includes the mature form of the full-length natural Propionibacterium acnes DsA2 polypeptide without its natural signal peptide sequence and its immunogenic variants. Immunogenic variants of the natural Propionibacterium acnes DsA2 polypeptide are capable of eliciting an immune response (e.g., an antigen-specific immune response), such as an antibody response, in a subject. Immunogenic variants of the natural Propionibacterium acnes DsA2 polypeptide include an immunogenic fragment of the natural Propionibacterium acnes DsA2 polypeptide. The immunogenic Propionibacterium acnes DsA2 fragment comprises a natural Propionibacterium acnes DsA2 polypeptide at least 50, 75, 100, 125, 150, 175, 200, 225, 250, 275, 300, 325, 350, or 375 amino acids in length. In some embodiments, the immunogenic Propionibacterium acnes DsA2 fragment contains a CSD2 domain of the Propionibacterium acnes DsA2 polypeptide. In some embodiments, immunogenic variants exclude sequence motifs found in the subject's (e.g., human) proteome, such as sequence motifs of 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, or 15 or more (e.g., 8 or more) amino acids in length. The excluded subject proteome sequence motifs are typically 8 or more amino acids in length. As further explained above, this helps to minimize unwanted cross-reactivity due to homology between the antigen and its own proteins.

[0069] An exemplary *Dermatophyte acnes* DsA2 polypeptide sequence includes the native *Dermatophyte acnes* DsA2 polypeptide sequence in SEQ ID NO: 205, which lacks the native secretion signal peptide sequence of the *Dermatophyte acnes* DsA2 polypeptide.

[0070] In some embodiments, the *Propionibacterium acnes* DsA2 polypeptide comprises a sequence of any one of SEQ ID NO: 205, SEQ ID NO: 20-27, or SEQ ID NO: 62-69, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the *Propionibacterium acnes* DsA2 polypeptide comprises a sequence having at least 90% identity with any one of SEQ ID NO: 205, SEQ ID NO: 20-27, or SEQ ID NO: 62-69. In some embodiments, the *Propionibacterium acnes* DsA2 polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO: 205, SEQ ID NO: 20-27, or SEQ ID NO: 62-69.

[0071] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the DsA2 polypeptide of *Propionibacterium acnes* comprises a nucleotide sequence according to any one of SEQ ID NO: 105-112 or SEQ ID NO: 171-178, or a sequence having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprises a nucleotide sequence at least 75% identical to any one of SEQ ID NO: 105-112 or SEQ ID NO: 171-178.

[0072] ALIHAKALPATGTGA The putative iron transporter protein (PITP) of *Propionibacterium acnes*, also known as P28 or P028, is a protein involved in the iron uptake mechanism of *Propionibacterium acnes* (Lodes et al., 2006). Iron uptake plays an important role in the survival of bacteria in host tissues. Under iron-limited conditions, the expression of PITP on the cell surface increases (Lodes et al., 2006). PITP (2006). The PITP polypeptide, its derivatives and fragments of *Propionibacterium acnes* are described in WO 2021 / 165543.

[0073] The *Propionibacterium acnes* PITP peptide of the present invention can induce immune responses against a variety of different *Propionibacterium acnes* strains, serotypes, and disease-causing variants. The *Propionibacterium acnes* PITP peptide exhibits the ability to elicit antibodies that bind to the surface of *Propionibacterium acnes* bacteria and recruit effector cells (e.g., effector cells of the innate immune system, such as phagocytes). Compared to using a single antigen, the combination of the *Propionibacterium acnes* PITP peptide of the present invention with *Propionibacterium acnes* DsA1 and / or DsA2 peptides (delivered as mRNA or as a recombinant protein) can elicit immune responses against a larger number of *Propionibacterium acnes* strains or serotypes.

[0074] The inventors have demonstrated that mRNA encoding different *Propionibacterium acnes* PITP peptides and recombinant *Propionibacterium acnes* PITP proteins elicits antibody (e.g., IgG) responses. Antibodies induced by *Propionibacterium acnes* PITP peptides show binding to the surface of *Propionibacterium acnes* bacteria. Antibodies induced by *Propionibacterium acnes* PITP peptides show in vitro induction of opsonization and phagocytosis. Antibodies induced by *Propionibacterium acnes* PITP peptides also show cross-reactivity across a range of *Propionibacterium acnes* strains.

[0075] The inventors have demonstrated that the *Propionibacterium acnes* PITP peptide is a suitable vaccine antigen that can be used in combination with one or more other *Propionibacterium acnes* antigens that can elicit an antibody response in subjects (e.g., the *Propionibacterium acnes* DsA1 peptide, the *Propionibacterium acnes* DsA2 peptide, and the *Propionibacterium acnes* CAMP2 peptide described herein). The *Propionibacterium acnes* CAMP2 peptide can elicit antibodies in subjects, for example, antibodies that can neutralize the biological activity of the CAMP2 peptide, such as antibodies that have inflammatory activity against CAMP2.

[0076] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a *Propionibacterium acnes* PITP polypeptide. In another aspect, the present invention provides a polypeptide comprising an amino acid sequence of a *Propionibacterium acnes* PITP polypeptide. The *Propionibacterium acnes* PITP polypeptide used in the present invention, delivered, for example, as mRNA or a recombinant protein, can induce antibodies in a subject. Such antibodies can opsonize *Propionibacterium acnes* bacteria. Opsonization may target *Propionibacterium acnes* bacteria with immune effector cells (e.g., phagocytes). This may result in such immune effector cells killing *Propionibacterium acnes* bacteria (e.g., phagocytosis). The binding of antibodies to the cell surface of *Propionibacterium acnes* can be measured, for example, using an in vitro surface binding assay. In vitro opsonization-phagocytosis assays can be used to assess the ability of antibodies to induce phagocytes to opsonize and kill *Propionibacterium acnes* strains of different genetic types.

[0077] The amino acid sequence (without signal peptide sequence) of the natural mature form of the full-length *Dermatophyte acnes* PITP polypeptide is provided in SEQ ID NO: 206: et al., AGPTVTVIPVGREGGDITISGKGFSTTGFGVYVAVAPASVPEFYGNSDKFYGYDPSKDTTESPSTIWVY TPSQKAIGSRFAQGRPMNNDGSFTITMKAPPFEQGKDFVVLTTKAHGVGKTDHSDDTRTPVTYREATPAPTGPKTPI Note: In the translation, for the sake of clarity, the superscript in "DsA^2" is retained as it is likely a specific identifier in the original text. If it's a typo in the original, it may need to be corrected according to the actual situation. APSKQPSKQAAPSKQVKPSKQAGPNKQSTTPQQKTAEHRSQTPAAHRTMTKQVCTIGASKVTSGSLTWGIRTSFTSY LRGPIANGSWKLSGGANWNGSAFTFPLTSGSFDPATKSGSLKYSGSVHMTGHHGILDMTLAEPSLQIKGSTGHLYLD VKSSSMDGKKTNYGRVDFATFGVSVSGNAAIKGSPVKLTATGAKAFAGFYRAGEPMNPLSTNLTLSAEKVCHNVTVD AVTGKVIGDDSGKGAGRGLPVTGAEGPSSDEIDLGIVGGLALTAVVSTVVVCRRYAARI Natural, mature *Propionibacterium acnes* PITP polypeptides typically contain an extended neo-oncocin family domain (“ENFD”), a first exchange region (“SR1”), a heme-binding domain (“HbD”), a second exchange region (“SR2”; which includes the first four N-terminal residues of the LPXTG motif (i.e., including LPXT but not G)) and a hydrophobic C-terminal region (“hLAR”; which includes the C-terminal glycine residue of the LPXTG motif). The LPXTG motif is considered essential for protein cell wall anchoring.

[0078] For example, in the native *Dermatitis acnes* PITP polypeptide of SEQ ID NO: 206, the extended novel tumor suppressor protein family domain (“ENFD”) corresponds to amino acid residues 1 to 133, the first exchange region (“SR1”) corresponds to amino acid residues 134 to 206, the heme-binding domain (“HbD”) corresponds to amino acid residues 207 to 365, the second exchange region (“SR2”) corresponds to amino acid residues 366 to 399, and the hydrophobic C-terminal region (“hLAR”) corresponds to amino acid residues 400 to 436. The LPXTG motif corresponds to amino acid residues 396-400.

[0079] The sequences of naturally occurring PITP peptides are largely highly identical, differing only at the N-terminus and C-terminus (except for point mutations). A more detailed sequence analysis of DsA1 has been described elsewhere (WO 2021 / 165543). As is known in the art, the positions of ENFD, SR1, HbD, SR2, and hLAR within other naturally occurring *Dermatophyte acnes* PITP peptides can be determined by aligning the peptide sequence of a given PITP peptide with that of SEQ ID NO: 206 and identifying regions within the PITP peptide that exhibit high sequence identity with the sequences of ENFD, SR1, HbD, SR2, and hLAR, respectively, of SEQ ID NO: 206.

[0080] "Propionibacterium acnes PITP polypeptide" includes the mature form of a full-length natural PITP polypeptide of *Propionibacterium acnes* without its natural signal peptide sequence and its immunogenic variants. Immunogenic variants of the natural *Propionibacterium acnes* PITP polypeptide are capable of eliciting an immune response (e.g., an antigen-specific immune response), such as an antibody response, in a subject. Immunogenic variants of the natural *Propionibacterium acnes* PITP polypeptide include immunogenic fragments of the natural *Propionibacterium acnes* PITP polypeptide. Immunogenic *Propionibacterium acnes* PITP fragments comprise natural *Propionibacterium acnes* PITP polypeptides of at least 50, at least 75, at least 100, at least 125, at least 150, at least 175, at least 200, at least 225, at least 250, at least 275, at least 300, at least 325, at least 350, at least 375, at least 400, or at least 425 amino acids in length. In some embodiments, the immunogenic *Propionibacterium acnes* PITP fragment comprises the ENFD and / or HbD (e.g., ENFD) domains of the *Propionibacterium acnes* PITP polypeptide. In some embodiments, immunogenic variants exclude sequence motifs found in the subject's (e.g., human) proteome, such as sequence motifs of 8 or more, 9 or more, 10 or more, 11 or more, 12 or more, 13 or more, 14 or more, or 15 or more (e.g., 8 or more) amino acids in length. The excluded subject proteome sequence motifs are typically 8 or more amino acids in length. As further explained above, this helps to minimize unwanted cross-reactivity due to homology between the antigen and its own proteins.

[0081] An exemplary *Propionibacterium acnes* PITP polypeptide sequence includes the natural *Propionibacterium acnes* PITP polypeptide sequence in SEQ ID NO: 206, which lacks the natural secretion signal peptide sequence of the *Propionibacterium acnes* PITP polypeptide.

[0082] In some embodiments, the *Propionibacterium acnes* PITP polypeptide comprises a sequence of any one of SEQ ID NO: 206, SEQ ID NO: 31-37, or SEQ ID NO: 73-79, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the *Propionibacterium acnes* PITP polypeptide comprises a sequence having at least 90% identity with any one of SEQ ID NO: 206, SEQ ID NO: 31-37, or SEQ ID NO: 73-79. In some embodiments, the *Propionibacterium acnes* PITP polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO: 206, SEQ ID NO: 31-37, or SEQ ID NO: 73-79. In some embodiments, the *Propionibacterium acnes* PITP polypeptide comprises, is composed of, or is substantially composed of (e.g., comprises) the amino acid sequence according to SEQ ID NO: 73. Typically, the *Propionibacterium acnes* PITP polypeptide comprises the amino acid sequence according to SEQ ID NO: 73 or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it.

[0083] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the PITP polypeptide of *Propionibacterium acnes* comprises a nucleotide sequence according to any one of SEQ ID NO: 115, SEQ ID NO: 117-121, SEQ ID NO: 182-188 (e.g., SEQ ID NO: 182) or having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprises a nucleotide sequence that is at least 75% identical to any one of SEQ ID NO: 115, SEQ ID NO: 117-121, SEQ ID NO: 182-188 (e.g., SEQ ID NO: 182 or SEQ ID NO: 115, such as SEQ ID NO: 182).

[0084] In one embodiment, the nucleic acid of the present invention is an mRNA comprising or composed of the following structural elements (e.g., composed of the following structural elements): (i) A 5' cap with the following structure:

[0085] (ii) Having a 5' untranslated region (5' UTR) of the nucleic acid sequence according to SEQ ID NO: 265; (iii) Having a protein-coding region according to the nucleic acid sequence of SEQ ID NO: 115; (iv) Possessing a 3' untranslated region (3' UTR) according to the nucleic acid sequence of SEQ ID NO: 266; and (v) A poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), for example, wherein the poly-A tail comprises at least 100 adenosine nucleotides.

[0086] In some embodiments, the mRNA is chemically modified, and the chemical modification includes replacing each uridine with N1-methylpseudouridine. The mRNA may be encapsulated in LNPs.

[0087] Chimeric DsA1 / DsA2 The chimeric DsA1 / DsA2 polypeptide of Propionibacterium acnes has been previously described (WO 2021 / 165543).

[0088] The DsA1 and DsA2 proteins of *Propionibacterium acnes* are homologs (paralogs), with typical sequence identity between 60% and 71%, depending on the aligned protein region. Excluding PT region length polymorphisms, high (typically >90%) sequence identity is observed in both intact DsA1 and DsA2 proteins. *Propionibacterium acnes* DsA1 and DsA2 are differentially expressed in strains within phylogenetic types IA1, IC, and II. Some strains express more DsA1 peptide than DsA2 peptide, while others express more DsA2 peptide than DsA1 peptide. Providing both *Propionibacterium acnes* DsA1 and DsA2 peptides may be advantageous, as this allows targeting a wider range of *Propionibacterium acnes* strains. If DsA1 and DsA2 peptides are targeted individually, they may displace each other's functions. Therefore, providing both DsA1 and DsA2 peptides may reduce the chance of immune defense evasion. For example, providing both DsA1 and DsA2 peptides can elicit an immune (e.g., antibody) response against a larger number of *D. acnes* strains compared to using either DsA1 or DsA2 peptides alone (e.g., it can elicit an antibody response against *D. acnes* strains expressing lower levels of one of the two peptides). Therefore, providing both DsA1 and DsA2 peptides can increase immunogenicity. Providing a chimeric *D. acnes* DsA1 / DsA2 peptide as a single molecule, rather than providing DsA1 and DsA2 peptides as separate peptides, can facilitate the manufacture of peptides and / or nucleic acids used in this invention.

[0089] The chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide of the present invention can be used to induce an immune response against *Propionibacterium acnes* infection. Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide show binding to the surface of *Propionibacterium acnes* bacteria. Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide show the ability to induce opsonization and killing in vitro. These antibodies can exhibit superior cross-reactivity compared to antibodies induced by individual (non-chimeric) *Propionibacterium acnes* DsA1 peptides or *Propionibacterium acnes* DsA2 peptides. The chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide can induce antibodies that specifically bind to both the full-length *Propionibacterium acnes* DsA1 peptide and the *Propionibacterium acnes* DsA2 peptide, for example, as determined in ELISA assays. Antibodies induced by either DsA1 or DsA2 peptides can be cross-reactive against various *Propionibacterium acnes* serotypes. Furthermore, antibodies induced by the chimeric DsA1 / DsA2 peptide are cross-reactive against various *Propionibacterium acnes* serotypes. The inventors have demonstrated that the chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide elicits antibody (e.g., IgG) responses when delivered in the form of mRNA encoding the antigen and recombinant protein.

[0090] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide, wherein the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises: (i) (a) CSD2 of the *Dermatitis acnes* DsA1 polypeptide and (b) CSD1 and / or CSD3 of the *Dermatitis acnes* DsA2 polypeptide; or (ii) (a) CSD2 of the *Dermatitis acnes* DsA2 polypeptide and (b) CSD1 and / or CSD3 of the *Dermatitis acnes* DsA1 polypeptide. The nucleic acid may be messenger RNA (mRNA). On the other hand, the present invention provides a chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide having an amino acid sequence comprising: (i) (a) CSD2 of the *Propionibacterium acnes* DsA1 polypeptide and (b) CSD1 and / or CSD3 of the *Propionibacterium acnes* DsA2 polypeptide; or (ii) (a) CSD2 of the *Propionibacterium acnes* DsA2 polypeptide and (b) CSD1 and / or CSD3 of the *Propionibacterium acnes* DsA1 polypeptide. The chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide used in the present invention, delivered, for example, as mRNA or recombinant protein, can elicit an immune response (e.g., an antibody response) in a subject. Such antibodies can be cross-reactive antibodies that specifically bind to both the natural *Propionibacterium acnes* DsA1 polypeptide and the natural *Propionibacterium acnes* DsA2 polypeptide. Such antibody responses can be cross-reactive against various *Propionibacterium acnes* serotypes (e.g., serotypes IA1, IA2, IB, IC, II, and III).

[0091] In some embodiments, the chimeric dermatophyte DsA1 / DsA2 polypeptide comprises: (i) NSR of DsA1 peptide or NSR of DsA2 peptide of Dermatitis acnes (e.g. NSR of DsA1 peptide of Dermatitis acnes). (ii) CSD1 of the DsA1 peptide of Propionibacterium acnes or CSD1 of the DsA2 peptide of Propionibacterium acnes (e.g., CSD1 of the DsA1 peptide of Propionibacterium acnes). (iii) Optionally, SR1 (or a portion thereof) of the Dermatitis acnes DsA1 polypeptide and / or SR1 (or a portion thereof) of the Dermatitis acnes DsA2 polypeptide. (iv) CSD2 of the DsA1 peptide of Propionibacterium acnes or CSD2 of the DsA2 peptide of Propionibacterium acnes (e.g., CSD2 of the DsA2 peptide of Propionibacterium acnes). (v) Optionally, SR2 (or a portion thereof) of the DsA1 polypeptide of *Propionibacterium acnes* and / or SR2 (or a portion thereof) of the DsA2 polypeptide of *Propionibacterium acnes*, and (vi) CSD3 of the DsA1 peptide of Propionibacterium acnes or CSD3 of the DsA2 peptide of Propionibacterium acnes (e.g., CSD3 of the DsA1 peptide of Propionibacterium acnes).

[0092] As described in WO 2021 / 165543, the SRs within the *D. acnes* DsA1 and DsA2 peptides are typically regions enriched with essentially disordered peptide sequences (i.e., they are largely unstructured). The SRs share the property of connecting (“spacers”) two domains (e.g., CSDs (e.g., CSD1, CSD2, and CSD3)). Therefore, the CSDs from the *D. acnes* DsA1 and DsA2 peptides can be combined by appropriately engineering the SRs such that their lengths (number of amino acid residues) are conserved. Advantageously, this design strategy can preserve the structural integrity of the individual domains (e.g., CSDs) in the resulting chimeric DsA1 / DsA2 peptides.

[0093] In some embodiments, the chimeric dermatophyte DsA1 / DsA2 polypeptide of the present invention comprises (e.g., from the N-terminus to the C-terminus) amino acid residues 21-102 of SEQ ID NO: 204, amino acid residues 121-240 of SEQ ID NO: 205, and amino acid residues 250-295 of SEQ ID NO: 204.

[0094] In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises a sequence according to any one of SEQ ID NO: 28-30, SEQ ID NO: 39, or SEQ ID NO: 70-72, or SEQ ID NO: 81, or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO: 28-30, SEQ ID NO: 39, or SEQ ID NO: 70-72, or SEQ ID NO: 81. Preferably, the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises the amino acid sequence according to SEQ ID NO: 70 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it.

[0095] In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises, is composed of, or is substantially composed of (e.g., includes) the amino acid sequence according to SEQ ID NO: 70.

[0096] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises a nucleotide sequence according to SEQ ID NO: 113-114, SEQ ID NO: 179, SEQ ID NO: 123, or SEQ ID NO: 190 (e.g., SEQ ID NO: 179) or having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises a nucleotide sequence that is at least 85% identical to SEQ ID NO: 113-114, SEQ ID NO: 179, SEQ ID NO: 123, or SEQ ID NO: 190 (e.g., SEQ ID NO: 179). In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acne DsA1 / DsA2 polypeptide comprises or consists of the following: a nucleotide sequence encoding a secretion signal peptide sequence (e.g., the viral secretion signal peptide sequence described herein) and a nucleotide sequence according to SEQ ID NO: 179. In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acne DsA1 / DsA2 polypeptide comprises or consists of the following: a nucleotide sequence according to SEQ ID NO: 113.

[0097] In one embodiment, the nucleic acid of the present invention is an mRNA comprising or composed of the following structural elements (e.g., composed of the following structural elements): (i) A 5' cap with the following structure:

[0098] (ii) Having a 5' untranslated region (5' UTR) of the nucleic acid sequence according to SEQ ID NO: 265; (iii) Having a protein-coding region according to the nucleic acid sequence of SEQ ID NO: 113; (iv) Possessing a 3' untranslated region (3' UTR) according to the nucleic acid sequence of SEQ ID NO: 266; and (v) A poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), for example, wherein the poly-A tail comprises at least 100 adenosine nucleotides.

[0099] In some embodiments, the mRNA is chemically modified, and the chemical modification includes replacing each uridine with N1-methylpseudouridine. The mRNA may be encapsulated in LNPs.

[0100] Chimeric DsA1 / DsA2 / PITP The chimeric DsA1 / DsA2 / PITP polypeptide of *Dermatitis acnes* has been previously described (WO 2021 / 165543).

[0101] The DsA1 and DsA2 peptides can be expressed on *Propionibacterium acnes* serotypes IA1, IA2, IC, and II. Antibodies induced by the DsA1 and DsA2 peptides may not bind to *Propionibacterium acnes* MLST serotypes IB and III. On the other hand, antibodies against the *Propionibacterium acnes* PITP peptide may be able to bind to *Propionibacterium acnes* serotypes IB and III. Therefore, PITP may be an antigen that can complement the immune response induced by other *Propionibacterium acnes* antigens, such as DsA1 and / or DsA2. Compared with using individual antigens, the combination of *Propionibacterium acnes* PITP peptides with chimeric *Propionibacterium acnes* DsA1 / DsA2 peptides (delivered as mRNA or as recombinant protein) can elicit an immune response against a larger number of *Propionibacterium acnes* strains or serotypes. The combination of DsA1 and / or DsA2, delivered as mRNA or as a recombinant protein, with PITP can provide a cross-reactive immune response (e.g., antibody response) against a larger number of *D. acnes* strains or phylogens compared to the use of only DsA1 and / or DsA2. The *D. acnes* PITP peptide can be provided as a single peptide molecule (delivered as mRNA or as a recombinant protein) or as part of a chimeric *D. acnes* DsA1 / DsA2 / PITP peptide (delivered as mRNA or as a recombinant protein).

[0102] The chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide of the present invention (delivered as mRNA or recombinant protein) can be used to elicit an immune response against *Dermatitis acnes* infection. The chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide (delivered as mRNA or recombinant protein) can elicit an immune response against a larger number of *Dermatitis acnes* strains compared to using a single antigen (e.g., DsA1, DsA2, or PITP), such as a cross-reactive immune response. Providing the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide, instead of providing the *Dermatitis acnes* DsA1 polypeptide, *Dermatitis acnes* DsA2 polypeptide, and *Dermatitis acnes* PITP polypeptide as separate polypeptides (or the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide and *Dermatitis acnes* PITP polypeptide), facilitates the manufacture of the polypeptides and / or nucleic acids used in the present invention.

[0103] The chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide can elicit cross-reactive antibody responses against various *Propionibacterium acnes* serotypes. The inventors have demonstrated that the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide induces antibody (e.g., IgG) responses when delivered in the form of mRNA encoding the antigen and a recombinant protein. Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide show binding to the surface of *Propionibacterium acnes* bacteria.

[0104] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide, wherein the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide comprises the ENFD of the *Dermatitis acnes* PITP polypeptide and: (i)(a) CSD2 of the DsA1 polypeptide of *Propionibacterium acnes* and (b) CSD1 and / or CSD3 of the DsA2 polypeptide of *Propionibacterium acnes*; or (ii)(a) CSD2 of the DsA2 polypeptide of *Propionibacterium acnes* and (b) CSD1 and / or CSD3 of the DsA1 polypeptide of *Propionibacterium acnes*.

[0105] Nucleic acids can be messenger RNA (mRNA).

[0106] In another aspect, the present invention provides a chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide comprising the ENFD of the dermatitis acnes PITP polypeptide and: (i)(a) CSD2 of the DsA1 polypeptide of *Propionibacterium acnes* and (b) CSD1 and / or CSD3 of the DsA2 polypeptide of *Propionibacterium acnes*; or (ii)(a) CSD2 of the DsA2 polypeptide of *Propionibacterium acnes* and (b) CSD1 and / or CSD3 of the DsA1 polypeptide of *Propionibacterium acnes*.

[0107] The *Propionibacterium acnes* DsA1 / DsA2 / PITP peptides used in this invention, delivered, for example, as mRNA or recombinant protein, can elicit an immune response (e.g., antibody response) in a subject. Such antibodies may include antibodies that specifically bind to the natural *Propionibacterium acnes* DsA1 peptide, antibodies that specifically bind to the natural *Propionibacterium acnes* DsA2 peptide, and / or antibodies that specifically bind to the natural *Propionibacterium acnes* PITP peptide. These antibodies may be cross-reactive against various *Propionibacterium acnes* serotypes (e.g., serotypes IA1, IA2, IB, IC, II, and III).

[0108] In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises CSD2 of the *Propionibacterium acnes* DsA2 polypeptide, CSD1 of the *Propionibacterium acnes* DsA1 polypeptide, CSD3 of the *Propionibacterium acnes* DsA1 polypeptide, and ENFD of the *Propionibacterium acnes* PITP polypeptide. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide comprises: (a)(i) NSR of the DsA1 peptide of Propionibacterium acnes or NSR of the DsA2 peptide of Propionibacterium acnes (e.g., NSR of the DsA1 peptide of Propionibacterium acnes). (ii) CSD1 of the DsA1 peptide of Propionibacterium acnes or CSD1 of the DsA2 peptide of Propionibacterium acnes (e.g., CSD1 of the DsA1 peptide of Propionibacterium acnes). (iii) Optionally, SR1 (or a portion thereof) of the Dermatitis acnes DsA1 polypeptide and / or SR1 (or a portion thereof) of the Dermatitis acnes DsA2 polypeptide. (iv) CSD2 of the DsA1 peptide of Propionibacterium acnes or CSD2 of the DsA2 peptide of Propionibacterium acnes (e.g., CSD2 of the DsA2 peptide of Propionibacterium acnes). (v) Optionally, SR2 (or a portion thereof) of the DsA1 polypeptide of *Propionibacterium acnes* and / or SR2 (or a portion thereof) of the DsA2 polypeptide of *Propionibacterium acnes*, and (vi) CSD3 of the *Propionibacterium acnes* DsA1 peptide or CSD3 of the *Propionibacterium acnes* DsA2 peptide (e.g., CSD3 of the *Propionibacterium acnes* DsA1 peptide); and (b) ENFD of PITP polypeptide of Propionibacterium acnes.

[0109] As described in WO 2021 / 165543, the SRs within the *Dermatologic acnes* DsA1 and DsA2 peptides and PITP are typically regions enriched with essentially disordered polypeptide sequences (i.e., they are largely unstructured). The SRs share the property of connecting (“spacers”) two domains (e.g., CSDs (e.g., CSD1, CSD2, and CSD3 of the *Dermatologic acnes* DsA1 and DsA2 peptides), the ENFD of the *Dermatologic acnes* PITP peptide, or the HbD of the *Dermatologic acnes* PITP peptide). Therefore, the CSDs of the *Dermatologic acnes* DsA1 and DsA2 peptides, the ENFD of the *Dermatologic acnes* PITP peptide, and / or the HbD of the *Dermatologic acnes* PITP peptide can be combined by scattering SRs of similar length (number of amino acid residues) within them to those SRs in the native *Dermatologic acnes* peptide. Advantageously, this design strategy can preserve the structural integrity of each domain (e.g., CSDs, ENFD, and HbD) in the resulting chimeric DsA1 / DsA2 / PITP peptide.

[0110] In some embodiments, the chimeric dermatophyte DsA1 / DsA2 / PITP polypeptide comprises an amino acid sequence according to any one of SEQ ID NO:38, SEQ ID NO:40-41, SEQ ID NO:80, SEQ ID NO:82-83 or SEQ ID NO:367-368 (e.g. SEQ ID NO:80, 82, 83 or 368) or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity with it. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide comprises a sequence having at least 90% identity with any one of SEQ ID NO: 38, SEQ ID NO: 40-41, SEQ ID NO: 80, SEQ ID NO: 82-83, or SEQ ID NO: 367-368 (e.g., SEQ ID NO: 80, 82, 83, or 368). In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide comprises a sequence having at least 95% identity with any one of SEQ ID NO: 38, SEQ ID NO: 40-41, SEQ ID NO: 80, SEQ ID NO: 82-83, or SEQ ID NO: 367-368 (e.g., SEQ ID NO: 80, 82, 83, or 368).

[0111] In some embodiments, the chimeric dermatophyte DsA1 / DsA2 / PITP polypeptide consists of the amino acid sequence of SEQ ID NO: 80, 82, 83 or 368.

[0112] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide comprises a nucleotide sequence according to any one of SEQ ID NO: 122, SEQ ID NO 124-125, SEQ ID NO: 189 or SEQ ID NO: 191-192 (e.g., SEQ ID NO: 189 or 192) or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98% or at least 99% identity with it. In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatophyte DsA1 / DsA2 / PITP polypeptide comprises at least 85% identical nucleotide sequences to any one of SEQ ID NO: 122, SEQ ID NO 124-125, SEQ ID NO: 189 or SEQ ID NO: 191-192 (e.g., SEQ ID NO: 189 or 192).

[0113] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatophyte DsA1 / DsA2 polypeptide consists of the following: a nucleotide sequence encoding a secretion signal peptide sequence (e.g., the viral secretion signal peptide sequence described herein) and a sequence according to any one of SEQ ID NO: 189 or SEQ ID NO: 191-192.

[0114] Chimeric DsA1 / DsA2 / PITP / CAMP2 The inventors have designed a chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide (also referred to herein as a chimeric *Dermatitis acnes* CAMP2 / DsA1 / DsA2 / PITP polypeptide). This chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide can elicit an immune (e.g., antibody) response. This response can be a protective immune (e.g., antibody) response. The chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide can elicit an immune response (e.g., antibody) against a natural *Dermatitis acnes* DsA1 polypeptide, an immune response (e.g., antibody) against a natural *Dermatitis acnes* DsA2 polypeptide, an immune response (e.g., antibody) against a natural *Dermatitis acnes* PITP polypeptide, and / or an immune response (e.g., antibody) against a natural *Dermatitis acnes* CAMP2 polypeptide. Such antibodies are cross-reactive against various *Dermatitis acnes* MLST serotypes (e.g., serotypes IA1, IA2, IB, IC, II, and III). Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide can neutralize the biological activities of the CAMP2 peptide, such as its inflammatory activity. Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide can opsonize *Propionibacterium acnes* bacteria. Opsonization may target *Propionibacterium acnes* bacteria with immune effector cells (e.g., phagocytosis). This may lead to the killing of *Propionibacterium acnes* bacteria by these immune effector cells (e.g., phagocytosis). The chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide can induce antibodies that neutralize the biological activities of the CAMP2 peptide, such as its inflammatory activity. Therefore, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide can elicit (a) antibodies that neutralize the biological activity (e.g., inflammatory activity) of the *Propionibacterium acnes* CAMP2 peptide, and (b) antibodies that make *Propionibacterium acnes* susceptible to opsonin action and recruit immune effector cells (e.g., phagocytes). Thus, immunization with the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide of the present invention (e.g., as mRNA or as a recombinant protein delivery) can provide a more effective immune response (e.g., more effective killing of *Propionibacterium acnes* bacteria in the subject) than immunization with the chimeric DsA1 / DsA2 / PITP peptide or with the DsA1, DsA2, or PITP antigen alone or with the CAMP2 peptide itself.

[0115] The inventors have demonstrated that the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide, when delivered as mRNA encoding the peptide, elicits an antibody (e.g., IgG) response against the antigens DsA1, DsA2, PITP, and CAMP2. Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide show binding to the surface of *Propionibacterium acnes* bacteria. Antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide show in vitro opsonization and killing. The inventors have also shown that antibodies induced by the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide reduce the co-hemolytic activity of the *Propionibacterium acnes* CAMP2 peptide.

[0116] Combining DsA1, DsA2, PITP, and CAMP2 antigens into a single chimeric peptide can advantageously reduce vaccine reactivity, for example, when the chimeric peptide is delivered as a single mRNA (e.g., because a single mRNA vaccine formulation can use less cationic lipids or LNPs than a vaccine formulation containing several different mRNAs). Combining DsA1, DsA2, PITP, and CAMP2 antigens into a single chimeric peptide can also facilitate vaccine manufacturing, as it is easier, faster, and less costly to prepare a single nucleic acid or peptide combining these four antigens rather than separate nucleic acids or peptides targeting different antigens.

[0117] As described above, the design strategy can leverage the structural integrity of individual structured domains (e.g., CSDs of DsA1 and DsA2 and ENFD of PITP) in the resulting chimeric DsA1 / DsA2 / PITP / CAMP2 peptide. The design strategy can also leverage the flexible linker domain of the *D. acnes* CAMP2 peptide and / or the flexible linker-like sequence at the C-terminus of the chimeric *D. acnes* DsA1 / DsA2 / PITP peptide.

[0118] Therefore, in one aspect, the present invention provides a nucleic acid comprising a nucleotide sequence encoding a chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide, wherein the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises dermatitis acnes DsA1 or an immunogenic fragment thereof, dermatitis acnes DsA2 or an immunogenic fragment thereof, a dermatitis acnes PITP polypeptide or an immunogenic fragment thereof, and a dermatitis acnes CAMP2 polypeptide or an immunogenic fragment thereof. Typically, the nucleic acid is messenger RNA (mRNA).

[0119] In another aspect, the present invention provides a chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprising dermatitis acnes DsA1 or an immunogenic fragment thereof, dermatitis acnes DsA2 or an immunogenic fragment thereof, dermatitis acnes PITP polypeptide or an immunogenic fragment thereof, and dermatitis acnes CAMP2 polypeptide or an immunogenic fragment thereof.

[0120] In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* DsA1 polypeptide is the CSD2 of the *Propionibacterium acnes* DsA1 polypeptide. In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* DsA2 polypeptide is the CSD2 of the *Propionibacterium acnes* DsA2 polypeptide.

[0121] In some embodiments, the second conserved subdomain (“CSD2”) of the *Dermatitis acnes* DsA1 polypeptide corresponds to amino acid residues 120-239 of SEQ ID NO: 204. A description of the other domains and their boundaries of the *Dermatitis acnes* DsA1 polypeptide is given in the “DsA1” section above. As is known in the art, the positions of CSD2 and other domains in other native *Dermatitis acnes* DsA1 polypeptides can be determined by comparing the polypeptide sequence of a given DsA1 polypeptide with the polypeptide sequence of SEQ ID NO: 204 and identifying DsA1 polypeptide regions that have high sequence identity with CSD2 and other domains of SEQ ID NO: 204, respectively.

[0122] In some embodiments, the second conserved subdomain (“CSD2”) of the *Dermatitis acnes* DsA2 polypeptide corresponds to amino acid residues 121-240 of SEQ ID NO: 205. A description of the other domains and their boundaries of the *Dermatitis acnes* DsA2 polypeptide is given in the “DsA2” section above. As is known in the art, the positions of CSD2 and other domains in other native *Dermatitis acnes* DsA2 polypeptides can be determined by comparing the polypeptide sequence of a given DsA2 polypeptide with the polypeptide sequence of SEQ ID NO: 205 and identifying DsA2 polypeptide regions that have high sequence identity with CSD2 and other domains of SEQ ID NO: 205, respectively.

[0123] In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* PITP peptide comprises the ENFD of the *Propionibacterium acnes* PITP peptide.

[0124] In some embodiments, the extended neo-oncocin family domain (“ENFD”) of *Propionibacterium acnes* PITP corresponds to amino acid residues 1-133 of SEQ ID NO: 206. A description of the other domains and boundaries of the *Propionibacterium acnes* PITP polypeptide is given in the “PITP” section above. As is known in the art, the positions of ENFD and other domains in other native *Propionibacterium acnes* PITP polypeptides can be determined by comparing the polypeptide sequence of a given PITP polypeptide with the polypeptide sequence of SEQ ID NO: 206 and identifying PITP polypeptide regions that have high sequence identity with ENFD and other domains of SEQ ID NO: 206, respectively.

[0125] In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* CAMP2 peptide includes the N-terminal domain of the *Propionibacterium acnes* CAMP2 peptide. In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* CAMP2 peptide includes both the N-terminal domain and a linker domain of the *Propionibacterium acnes* CAMP2 peptide. In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* CAMP2 peptide includes the C-terminal domain of the *Propionibacterium acnes* CAMP2 peptide. In some embodiments, the immunogenic fragment of the *Propionibacterium acnes* CAMP2 peptide includes both the linker domain and the C-terminal domain of the *Propionibacterium acnes* CAMP2 peptide.

[0126] In some embodiments, the N-terminal domain of the *Dermatitis acnes* CAMP2 polypeptide corresponds to residues 29-176 of SEQ ID NO: 202. In some embodiments, the C-terminal domain of the *Dermatitis acnes* CAMP2 polypeptide corresponds to residues 189-267 of SEQ ID NO: 202. In some embodiments, the linker domain of the *Dermatitis acnes* CAMP2 polypeptide corresponds to residues 177-188 of SEQ ID NO: 202. As is known in the art, the positions of the N-terminal domain, C-terminal domain, and linker domain in other naturally occurring *Dermatitis acnes* CAMP2 polypeptides can be determined by comparing the polypeptide sequence of a given CAMP2 polypeptide with the polypeptide sequence of SEQ ID NO: 202 and identifying CAMP2 polypeptide regions with high sequence identity to the N-terminal domain, C-terminal domain, and linker domain of SEQ ID NO: 202. Analysis of 430 CAMP2 polypeptide sequences from naturally occurring *Dermatitis acnes* strains showed a high degree of sequence conservation among CAMP2 sequences (see the “CAMP2” section above). Figure 44 shows an exemplary alignment of the CAMP2 polypeptide sequence from a naturally occurring strain of *Dermatophyte acnes* relative to SEQ NO: 202 (i.e., KPA171202_REF in Figure 44). For example, the alignment in Figure 44 can be used to identify the locations of the N-terminal domain, C-terminal domain, and adapter domain in sequences other than SEQ ID NO: 202.

[0127] In some embodiments, any two or more of (1) the *Dermatitis acnes* DsA1 polypeptide or its immunogenic fragment, (2) the *Dermatitis acnes* DsA2 polypeptide or its immunogenic fragment, (3) the *Dermatitis acnes* PITP polypeptide or its immunogenic fragment, and (4) the *Dermatitis acnes* CAMP2 polypeptide or its immunogenic fragment are attached via a connector (e.g., a connector described in the “Connectors” section below). In other embodiments, (1) the *Dermatitis acnes* DsA1 polypeptide or its immunogenic fragment, (2) the *Dermatitis acnes* DsA2 polypeptide or its immunogenic fragment, (3) the *Dermatitis acnes* PITP polypeptide or its immunogenic fragment, and (4) the *Dermatitis acnes* CAMP2 polypeptide or its immunogenic fragment are directly fused to each other (i.e., there is no connector, such as an amino acid connector, linking the DsA1, DsA2, PITP, and CAMP2 polypeptides or their immunogenic fragments to each other). In some embodiments, such chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 peptides contain a linker domain of the dermatitis acnes CAMP2 peptide.

[0128] In one embodiment, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises: (a) Chimeric DsA1 / DsA2 peptides of Propionibacterium acnes, such as those described herein; (b) An immunogenic fragment of the *Propionibacterium acnes* PITP polypeptide, comprising the ENFD of the *Propionibacterium acnes* PITP polypeptide; and (c) CAMP2 polypeptide of Propionibacterium acnes or its immunogenic fragment.

[0129] In some embodiments, fitting Propionibacterium acnes The DsA1 / DsA2 polypeptide comprises (i) (1) CSD2 of the *Dermatophyte acnes* DsA1 polypeptide and (2) CSD1 and / or CSD3 of the *Dermatophyte acnes* DsA2 polypeptide; or (ii) (1) CSD2 of the *Dermatophyte acnes* DsA2 polypeptide and (2) CSD1 and / or CSD3 of the *Dermatophyte acnes* DsA1 polypeptide. See above and the descriptions of the domains and boundaries of the *Dermatophyte acnes* DsA1 and DsA2 polypeptides in the “DsA1” and “DsA2” sections, respectively. In some embodiments, the chimeric *Dermatophyte acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide is as defined in (ii). Typically, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises CSD1 of the dermatitis acnes DsA1 polypeptide, CSD2 of the dermatitis acnes DsA2 polypeptide, and CSD3 of the dermatitis acnes DsA1 polypeptide.

[0130] In some embodiments, (a) the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 70, or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it.

[0131] In some embodiments, the immunogenic fragment of the ENFD containing the *Dermatitis acnes* PITP polypeptide in (b) comprises a sequence corresponding to amino acid residues 1 to 133 of SEQ ID NO: 73 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sequence of amino acid residues 1 to 133 of SEQ ID NO: 73. In some embodiments, the immunogenic fragment of the ENFD containing the *Dermatitis acnes* PITP polypeptide in (b) comprises a sequence corresponding to amino acid residues 1 to 146 of SEQ ID NO: 73 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sequence of amino acid residues 1 to 146 of SEQ ID NO: 73. Typically, the immunogenic fragment of the ENFD containing the dermatitis acnes PITP polypeptide in (b) contains the sequence of amino acid residues 1-146 corresponding to SEQ ID NO: 73.

[0132] In some embodiments, (a) and (b) comprise SEQ ID NO: 80 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, (a) and (b) comprise SEQ ID NO: 82 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, (a) and (b) comprise SEQ ID NO: 83 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. Typically, (a) and (b) comprise SEQ ID NO: 80.

[0133] In some embodiments, (c) is a *Propionibacterium acnes* CAMP2 polypeptide, such as the full-length *Propionibacterium acnes* CAMP2 polypeptide, typically the mature form of the full-length *Propionibacterium acnes* CAMP2 polypeptide (without a natural signal peptide sequence). In some embodiments, (c) comprises SEQ ID NO: 203 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. Typically, (c) comprises SEQ ID NO: 203.

[0134] In some embodiments, the immunogenic fragment of the *Dermatitis acnes* CAMP2 polypeptide in (c) comprises the N-terminal domain of the *Dermatitis acnes* CAMP2 polypeptide. In some embodiments, (c) comprises amino acid residues 29-176 of SEQ ID NO: 202 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sequence of amino acid residues 29-176 of SEQ ID NO: 202.

[0135] In some embodiments, the immunogenic fragment of the *Dermatitis acnes* CAMP2 polypeptide in (c) comprises the N-terminal domain and the linker domain of the *Dermatitis acnes* CAMP2 polypeptide. In some embodiments, (c) comprises amino acid residues 29-188 of SEQ ID NO: 202 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sequence of amino acid residues 29-188 of SEQ ID NO: 202.

[0136] In some embodiments, the immunogenic fragment of the *Dermatitis acnes* CAMP2 polypeptide in (c) comprises the C-terminal domain of the *Dermatitis acnes* CAMP2 polypeptide. In some embodiments, (c) comprises amino acid residues 189-267 of SEQ ID NO: 202 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sequence of amino acid residues 189-267 of SEQ ID NO: 202.

[0137] In some embodiments, the immunogenic fragment of the *Dermatitis acnes* CAMP2 polypeptide in (c) comprises a linker domain and a C-terminal domain of the *Dermatitis acnes* CAMP2 polypeptide. In some embodiments, (c) comprises amino acid residues 177-267 of SEQ ID NO: 202 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with the sequence of amino acid residues 177-267 of SEQ ID NO: 202.

[0138] In some embodiments, any two or more of (a), (b), and (c) are attached via a linker (e.g., a linker described in the “Linkers” section below). Typically, (a), (b), and (c) are directly fused to each other (i.e., without a linker, such as an amino acid linker connecting (a), (b), and (c)), for example, in the order specified below. In some embodiments, (c) comprises a linker domain of the *Dermatophyte acnes* CAMP2 polypeptide.

[0139] In some embodiments, the chimeric dermatitis acne DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises, from the N-terminus to the C-terminus, (a), (b), and (c). In other embodiments, the chimeric dermatitis acne DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises, from the N-terminus to the C-terminus, (c), (a), and (b). Typically, the chimeric dermatitis acne DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises, from the N-terminus to the C-terminus, (c), (a), and (b).

[0140] In some embodiments, the chimeric dermatitis acne-derived DsA1 / DsA2 / PITP / CAMP2 polypeptide includes a transmembrane domain (TMB) sequence, such as the TMB as defined herein (see the "Heterologous Transmembrane Domain (TMB)" section below). In some embodiments, the TMB sequence is located at the N-terminus of the chimeric dermatitis acne-derived DsA1 / DsA2 / PITP / CAMP2 polypeptide. In other embodiments, the TMB sequence is located at the C-terminus of the chimeric dermatitis acne-derived DsA1 / DsA2 / PITP / CAMP2 polypeptide. Typically, the TMB sequence is located at the C-terminus of the chimeric dermatitis acne-derived DsA1 / DsA2 / PITP / CAMP2 polypeptide.

[0141] In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to any one of SEQ ID NO: 373-376 (e.g., SEQ ID NO: 373 or 374) or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 373 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it, and a TMB sequence (e.g., the sequence according to SEQ ID NO: 84). Typically, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises (1) the sequence according to SEQ ID NO: 374 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it; or (2) the sequence according to SEQ ID NO: 373 or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it, and the TMB sequence.

[0142] In some embodiments, the nucleic acid comprising a nucleotide sequence encoding a chimeric dermatophyte DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises a nucleotide sequence encoding a secretion signal peptide sequence as described herein (e.g., the secretion signal peptide sequence described in the “Secretion Signal Peptide (SS) Sequence” section below).

[0143] In some embodiments, encoding integration is included. Propionibacterium acnesThe nucleic acid of the DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to any one of SEQ ID NO: 386-390 or SEQ ID NO: 393 (e.g., SEQ ID NO: 387) or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises the sequence according to SEQ ID NO: 384 or SEQ ID NO: 385 or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it, and the nucleotide sequence encoding the TMB sequence (e.g., the nucleotide sequence encoding SEQ ID NO: 84). In some embodiments, the nucleotide sequence encoding SEQ ID NO: 84 is as defined in SEQ ID NO: 395 or 396. Typically, a nucleic acid containing a nucleotide sequence encoding a chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises (1) the sequence according to SEQ ID NO: 387, or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it; or (2) the sequence according to SEQ ID NO: 384 or SEQ ID NO: 385, or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it, and a nucleotide sequence encoding a TMB sequence.

[0144] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acne DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises a nucleotide sequence according to any one of SEQ ID NO: 377-383 or SEQ ID NO: 394 (e.g., any one of SEQ ID NO: 377-380) or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identity with it. In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acne DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises a nucleotide sequence having at least 75% identity with any one of SEQ ID NO: 377-380.

[0145] In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises the sequence of the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide as described herein. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises the sequence of the chimeric dermatitis acnes DsA1 / DsA2 polypeptide as described herein.

[0146] Variants of the polypeptide of the present invention Mutations of cysteine ​​residues One or more cysteine ​​residues in the polypeptide described herein can be mutated by substitution of a single amino acid (e.g., substitution with a serine residue). Cysteine ​​residues are involved in disulfide bond formation, and therefore, cysteine ​​mutations can restrict polypeptide polymerization.

[0147] In some embodiments, the *Dermatitis acnes* DsA1 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA1 polypeptide. In some embodiments, the *Dermatitis acnes* DsA1 polypeptide comprises one or more single amino acid substitutions at positions corresponding to C25 (e.g., C25S), C291 (e.g., C291S or C291M), and / or C293 (e.g., C293S or C293P) of SEQ ID NO: 204.

[0148] In some embodiments, the *Dermatitis acnes* DsA2 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA2 polypeptide. In some embodiments, the *Dermatitis acnes* DsA2 polypeptide comprises one or more single amino acid substitutions at positions corresponding to C26 (e.g., C26S) and / or C292 (e.g., C292S) of SEQ ID NO: 205.

[0149] In some embodiments, the *Propionibacterium acnes* PITP polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Propionibacterium acnes* PITP polypeptide. In some embodiments, the *Propionibacterium acnes* PITP polypeptide comprises one or more single amino acid substitutions at positions corresponding to C200 (e.g., C200S), C371 (e.g., C371S), and / or C429 (e.g., C429S) of SEQ ID NO: 206. In some embodiments, the *Propionibacterium acnes* PITP polypeptide comprises a single amino acid substitution at one or both (e.g., both) positions corresponding to C200 (e.g., C200S) and C371 (e.g., C371S) of SEQ ID NO: 206. In a preferred embodiment, the *Propionibacterium acnes* PITP polypeptide having the sequence of SEQ ID NO: 73 comprises serine residues at positions 200 and 371. In other embodiments, the *Propionibacterium acnes* PITP polypeptide having the sequence of SEQ ID NO: 77 comprises serine residues at positions 200 and 371.

[0150] In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA1 polypeptide and / or at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA2 polypeptide. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to C25 (e.g., C25S), C291 (e.g., C291S or C291M), and / or C293 (e.g., C293S or C293P) of SEQ ID NO: 204. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises one or more (e.g., all three) amino acid substitutions selected from C25S, C291S, and / or C293P, wherein C25, C291, and / or C293 correspond to residues in SEQ ID NO: 204. In a preferred embodiment, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide having the sequence of SEQ ID NO: 70 comprises serine residues at positions 25 and 291 and a proline residue at position 293. In other embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide having the sequence of SEQ ID NO: 72 comprises serine residues at positions 25 and 291 and a proline residue at position 293. In other embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide having the sequence of SEQ ID NO: 81 comprises a serine residue at position 18 and a proline residue at position 286. In other embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide having the sequence of SEQ ID NO: 81 comprises a serine residue at position 18, a methionine residue at position 284, and a proline residue at position 286, wherein these residues correspond to positions 25, 291, and 293 in SEQ ID NO: 204, respectively.

[0151] In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA1 polypeptide, one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA2 polypeptide, and / or one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* PITP polypeptide. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide comprises a single amino acid substitution at one or more positions corresponding to C25 (e.g., C25S), C291 (e.g., C291S or C291M), and / or C293 (e.g., C293S or C293P) of SEQ ID NO: 204. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide comprises one or more (e.g., all three) amino acid substitutions selected from C25S, C291S, and / or C293P, wherein C25, C291, and / or C293 correspond to residues in SEQ ID NO: 204. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide having the sequence of SEQ ID NO: 80 or SEQ ID NO: 83 (e.g., SEQ ID NO: 83) comprises serine residues at positions 25 and 291 and a proline residue at position 293. In other embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide having the sequence of SEQ ID NO: 82 comprises a serine residue at position 25 and a proline residue at position 293. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide having the sequence of SEQ ID NO: 82 comprises a serine residue at position 25, a methionine residue at position 291, and a proline residue at position 293. In other embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide having the sequence of SEQ ID NO: 368 comprises a serine residue at position 18, a methionine residue at position 284, and a proline residue at position 286. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide having the sequence of SEQ ID NO: 368 comprises a serine residue at position 18, a methionine residue at position 284, and a proline residue at position 286, wherein these residues correspond to positions 25, 291, and 293 in SEQ ID NO: 204, respectively.

[0152] In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA1 polypeptide, one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* DsA2 polypeptide, and / or one or more (e.g., all) positions corresponding to cysteine ​​residues in the native *Dermatitis acnes* PITP polypeptide. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide comprises a single amino acid substitution at one or more positions corresponding to C25 (e.g., C25S), C291 (e.g., C291S or C291M), and / or C293 (e.g., C293S or C293P) of SEQ ID NO: 204. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises one or more (e.g., all three) amino acid substitutions selected from C25S, C291S, and / or C293P, wherein C25, C291, and / or C293 correspond to residues in SEQ ID NO: 204. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide having the sequence of SEQ ID NO: 373 or SEQ ID NO: 374 comprises serine residues at positions 264 and 530 and a proline residue at position 532. In other embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide having the sequence of SEQ ID NO: 375 comprises serine residues at positions 185 and 451 and a proline residue at position 453. In other embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide having the sequence SEQ ID NO: 376 contains serine residues at positions 25 and 291 and a proline residue at position 293.

[0153] Mutations at glycosylation sites Glycosylation can occur in eukaryotic cells, but typically not in prokaryotic cells. As used herein, “glycosylation” refers to the addition of a sugar unit to a protein. Specifically, N-linked glycosylation is the attachment of a glycan to the amide nitrogen of an asparagine (Asn; N) residue in a protein. N-glycosylation can occur at any asparagine residue in a protein that is accessible and recognized by glycosylation enzymes after translation, and is most common at accessible asparagine residues that are part of an NXS / T motif, where the second amino acid residue following asparagine is a serine (Ser; S) or threonine (Thr; T). O-linked glycosylation is the attachment of a glycan to the oxygen atom of a serine (Ser) or threonine (Thr) residue in a protein. The attachment process produces a glycosylated protein. This glycan can be a polysaccharide. Non-human glycosylation patterns can cause peptides to exhibit undesirable reactivity when used to initiate antibodies. Furthermore, glycosylation of peptides that are normally non-glycosylated (such as the peptides described herein, which are naturally occurring peptides in prokaryotic cells or derived from such peptides) can alter their immunogenicity. For example, glycosylation can mask important immunogenic epitopes within proteins. Therefore, to reduce or eliminate glycosylation, asparagine residues or serine / threonine residues can be modified, for example, by substitution with another amino acid.

[0154] In some embodiments, one or more of the following peptides, such as the *Dermatitis acnes* CAMP2 peptide, the modified *Dermatitis acnes* CAMP2 peptide, the *Dermatitis acnes* DsA1 peptide, the *Dermatitis acnes* DsA2 peptide, the *Dermatitis acnes* PITP peptide, the chimeric *Dermatitis acnes* DsA1 / DsA2 peptide, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP peptide, and the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 peptide, contain at least one mutated glycosylation site (preferably at least one mutated N-linked glycosylation site) and / or at least one O-linked glycosylation site. In some embodiments, one or more of the following peptides, such as the *Dermatitis acnes* CAMP2 peptide, the modified *Dermatitis acnes* CAMP2 peptide, the *Dermatitis acnes* DsA1 peptide, the *Dermatitis acnes* DsA2 peptide, the *Dermatitis acnes* PITP peptide, the chimeric *Dermatitis acnes* DsA1 / DsA2 peptide, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP peptide, and the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 peptide, are removed. Removing glycosylation sites reduces the glycosylation of the peptide. In some embodiments, one or more of the following: *Propionibacterium acnes* CAMP2 peptide, modified *Propionibacterium acnes* CAMP2 peptide, *Propionibacterium acnes* DsA1 peptide, *Propionibacterium acnes* DsA2 peptide, *Propionibacterium acnes* PITP peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide, and chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide have reduced glycosylation relative to the corresponding native peptide. Removal of the glycosylation site can eliminate the glycosylation of the peptide.

[0155] In some embodiments, the modification includes the substitution of one or more of the N, S, and T amino acids (e.g., in an NXS / T sequence motif), where X corresponds to any amino acid. In some embodiments, the modification includes the substitution of one or more serine (Ser) or threonine (Thr) residues in the protein. In some embodiments, the N, S, or T amino acid is substituted with a conserved amino acid. Typically, the N amino acid can be substituted with a Q, S, K, or A amino acid.

[0156] In some embodiments, the *Dermatitis acnes* CAMP2 polypeptide and / or modified *Dermatitis acnes* CAMP2 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Dermatitis acnes* CAMP2 polypeptide. The N-glycosylation site in the *Dermatitis acnes* CAMP2 polypeptide and / or modified *Dermatitis acnes* CAMP2 polypeptide may be present at the position corresponding to residue N166 of SEQ ID NO: 203. The N-glycosylation site may include N166, F167, and S168 relative to the residue number in SEQ ID NO: 203, wherein glycosylation is present at N166. In some embodiments, the *Dermatitis acnes* CAMP2 polypeptide and / or modified *Dermatitis acnes* CAMP2 polypeptide of the present invention comprises an amino acid substitution at a position corresponding to N166 (e.g., N166S) relative to the residue number in SEQ ID NO: 203. The *Dermatitis acnes* CAMP2 polypeptide and / or modified *Dermatitis acnes* CAMP2 polypeptide may contain the sequence of SEQ ID NO: 50, 51, or 54, wherein a serine residue is present at position 166. The O-glycosylation site in the *Dermatitis acnes* CAMP2 polypeptide and / or modified *Dermatitis acnes* CAMP2 polypeptide may be present at a position corresponding to any S and / or T residue in SEQ ID NO: 203.

[0157] In some embodiments, the *Propionibacterium acnes* DsA1 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* DsA1 polypeptide. The N-glycosylation site in the *Propionibacterium acnes* DsA1 polypeptide may be present at the position corresponding to residue N255 of SEQ ID NO: 204. The N-glycosylation site may include N255, I256, and T257 relative to the residue number in SEQ ID NO: 204, wherein glycosylation is present at N255. In some embodiments, the *Propionibacterium acnes* DsA1 polypeptide described herein comprises an amino acid substitution at the position corresponding to N255 (e.g., N255Q or N255Q) relative to the residue number in SEQ ID NO: 204. The O-glycosylation site in the *Propionibacterium acnes* DsA1 polypeptide described herein may be present at the position corresponding to any S and / or T residue (e.g., residue S292) in SEQ ID NO: 204.

[0158] In some embodiments, the *Propionibacterium acnes* DsA2 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* DsA2 polypeptide. The N-glycosylation site in the *Propionibacterium acnes* DsA2 polypeptide may be present at the position corresponding to residue N5 of SEQ ID NO: 205. The N-glycosylation site may include N5, S6, and S7 relative to the residue number in SEQ ID NO: 205, wherein glycosylation is present at N5. In some embodiments, the *Propionibacterium acnes* DsA2 polypeptide described herein comprises an amino acid substitution at the position corresponding to N5 (e.g., N5Q) relative to the residue number in SEQ ID NO: 205. The *Propionibacterium acnes* DsA2 polypeptide may comprise the sequence of SEQ ID NO: 68 or 69, wherein glutamine is present at position 5. The O-glycosylation site in the *Propionibacterium acnes* DsA2 polypeptide described herein may be present at the position corresponding to any S and / or T residue in SEQ ID NO: 205.

[0159] In some embodiments, the *Propionibacterium acnes* PITP polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* PITP polypeptide. The N-glycosylation sites in the *Propionibacterium acnes* PITP polypeptide may be present at positions corresponding to residues N230, N242, N362, and N373 of SEQ ID NO: 206. The N-glycosylation sites may include (i) N230, G231, and S232 (glycosylation at N230); (ii) N242, G243, and S244 (glycosylation at N242); (iii) N362, L363, and T364 (glycosylation at N362); and / or (iv) N373, V374, and T375 (glycosylation at N373), wherein the amino acid numbers are relative to the residue numbers in SEQ ID NO: 206. In some embodiments, the *Propionibacterium acnes* PITP polypeptide of the present invention comprises one or more (e.g., all) amino acid substitutions at positions corresponding to N230 (e.g., N230K), S244 (e.g., S244G), N362 (e.g., N362Q), and N373 (e.g., N373S), wherein the amino acid numbers are relative to the residue numbers in SEQ ID NO: 206. In some embodiments, the *Propionibacterium acnes* PITP polypeptide described herein comprises one or more (e.g., all) amino acid substitutions at positions corresponding to N230 (e.g., N230K), N242 (e.g., N242G), N362 (e.g., N362Q), and N373 (e.g., N373S), wherein the amino acid numbers are relative to the residue numbers in SEQ ID NO: 206. The O-glycosylation site in the *Propionibacterium acnes* PITP polypeptide may be present at positions corresponding to any S and / or T residues in SEQ ID NO: 206. In some embodiments, the PITP polypeptide of *Dermatophyte acnes* as described herein comprises the sequence of SEQ ID NO: 76, 77, or 79, having lysine at position 230, glycine at position 244, glutamine at position 362, and serine at position 373.

[0160] In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* DsA1 polypeptide and / or at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* DsA2 polypeptide. The N-glycosylation site in the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide may be present at the position corresponding to residue N255 of SEQ ID NO: 70. The N-glycosylation site may include N255, I256, and T257 relative to the residue number in SEQ ID NO: 70, wherein glycosylation is present at N255. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises a substitution of N255Q or N255A (e.g., N255Q) from SEQ ID NO: 70. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide of the present invention contains an amino acid substitution at a position corresponding to N255 (e.g., N255Q or N255A) relative to the residue number in SEQ ID NO: 70. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 72, wherein glutamine is present at position 255. O-glycosylation sites in the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide may be present at positions corresponding to any S and / or T residues in SEQ ID NO: 70. O-glycosylation sites in the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide may be present at positions corresponding to residues S291 and / or S292 in SEQ ID NO: 70. O-glycosylation sites may include S291 and S292 relative to residue numbers in SEQ ID NO: 70, wherein glycosylation is present at S291 and / or S292. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises one or more (e.g., all three) substitutions selected from N255A, S291M, and S292G. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises one or more (e.g., all three) amino acid substitutions at positions corresponding to N255 (e.g., N255A), S291 (e.g., S291M), and S292 (e.g., S292G), relative to the residue numbers in SEQ ID NO: 70. The chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide may comprise the sequence of SEQ ID NO: 81, with alanine at position 248, methionine at position 284, and glycine at position 285.In some embodiments, the chimeric dermatophyte DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 81, having alanine at position 248, methionine at position 284, and glycine at position 285, wherein these residues correspond to positions 255, 291, and 292 in SEQ ID NO: 70, respectively.

[0161] In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide described herein comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* DsA1 polypeptide, one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* DsA2 polypeptide, and / or one or more (e.g., all) positions corresponding to glycosylation sites in the native *Propionibacterium acnes* PITP polypeptide. The N-glycosylation site in the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide may be present at the position corresponding to residue N255, relative to the residue number in SEQ ID NO: 80 or 83. The N-glycosylation site may include N255, I256, and T257, with glycosylation at N255. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide comprises a substitution of N255Q or N255A (e.g., N255A) as in SEQ ID NO: 80 or 83. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide of the present invention comprises an amino acid substitution at a position corresponding to N255 (e.g., N255Q or N255A) relative to the residue number in SEQ ID NO: 80 or 83. The O-glycosylation site in the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide may be present at a position corresponding to any S and / or T residue in SEQ ID NO: 80 or 83. Chimeric Propionibacterium acnesThe O-glycosylation site in the DsA1 / DsA2 / PITP polypeptide may be present at one or more (e.g., all) positions corresponding to residues S291, S292, T299, T301, T303, T305, T425, S428, and T436 of SEQ ID NO: 80 or 83. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide as described herein comprises a substitution of the PTPTPTPT region located between positions 298 and 305 of SEQ ID NO: 80 or 83 with a GGGGG linker. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide comprises one or more (e.g., all) substitutions selected from S291M, S292G, P298G, T299G, P300G, T301G, P302G, T425G, S428G, and T436G. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide comprises one or more (e.g., all) substitutions selected from S291M, S292G, T425G, S428G, and T436G relative to residue numbers in SEQ ID NO: 80 or 83. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide comprises one or more (e.g., all) amino acid substitutions at positions corresponding to residue numbers in SEQ ID NO: 80 or 83, such as S291 (e.g., S291M), S292 (e.g., S292G), T425 (e.g., T425G), S428 (e.g., S428G), and T436 (e.g., T436G). The chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide may contain the sequence of SEQ ID NO: 82, with methionine at position 291 and glycine at positions 292, 299, 301, 422, 425, and 433. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide may contain the sequence of SEQ ID NO: 82, with methionine at position 291 and glycine at positions 292, 298, 299, 300, 301, 302, 422, 425, and 433. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP polypeptide may comprise the sequence of SEQ ID NO: 368, having alanine at position 248, methionine at position 284, and glycine at positions 285, 291, 292, 293, 294, 295, 415, 418, and 426.

[0162] In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein contains a single amino acid substitution at one or more (e.g., all) positions corresponding to glycosylation sites in the native dermatitis acnes DsA1 polypeptide, one or more (e.g., all) positions corresponding to glycosylation sites in the native dermatitis acnes DsA2 polypeptide, one or more (e.g., all) positions corresponding to glycosylation sites in the native dermatitis acnes PITP polypeptide, and / or one or more (e.g., all) positions corresponding to glycosylation sites in the native dermatitis acnes CAMP2 polypeptide.

[0163] As described in the "Chimeric DsA1 / DsA2 / PITP / CAMP2" section above, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may contain the *Dermatitis acnes* CAMP2 polypeptide or an immunogenic fragment thereof. The N-glycosylation site in the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may be present at the position corresponding to residue N166 of SEQ ID NO: 203 (i.e., corresponding to N194 of SEQ ID NO: 202), provided that such residue is present in the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide. The N-glycosylation sites in the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may be present at positions N166, F167, and S168 (where glycosylation is at N166) (i.e., corresponding to positions N194, F195, and S196, where glycosylation is at N194), provided that these residues are present in the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide. In some embodiments, the *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide of the present invention contains amino acid substitutions at positions corresponding to N166 (e.g., N166S) (i.e., corresponding to N194, if present in the polypeptide), where the residues are present. The O-glycosylation site in the *Dermatobacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may be present at the location corresponding to any S and / or T residues in SEQ ID NO: 203 (i.e., any S and / or T residues corresponding to residues 29 to 267 in SEQ ID NO: 202) if present in the chimeric *Dermatobacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide.

[0164] As described in the "Chimeric DsA1 / DsA2 / PITP / CAMP2" section above, the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may contain SEQ ID NO: 80, 82, or 83. The N-glycosylation site in the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may be located at the position corresponding to residue N255, relative to the residue number in SEQ ID NO: 80 or 83. The N-glycosylation site in the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may include N255, I256, and T257, with glycosylation at N255. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide of the present invention contains an amino acid substitution at a position corresponding to N255 (e.g., N255Q or N255A), relative to the residue number in SEQ ID NO: 80 or 83. The O-glycosylation site in the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may be present at a position corresponding to any S and / or T residue in SEQ ID NO: 80 or 83. The O-glycosylation site in the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may be present at one or more (e.g., all) positions corresponding to residues S291, S292, T299, T301, T303, T305, T425, S428, and T436 in SEQ ID NO: 80 or 83. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide as described herein comprises a substitution of the PTPTPTPT region located between positions 298 and 305 of SEQ ID NO: 80 or 83 with a GGGGG linker. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises one or more (e.g., all) substitutions selected from S291M, S292G, P298G, T299G, P300G, T301G, P302G, T425G, S428G, and T436G. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises one or more (e.g., all) substitutions selected from S291M, S292G, T425G, S428G, and T436G with respect to residue numbers in SEQ ID NO: 80 or 83.In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide comprises one or more (e.g., all) amino acid substitutions at positions corresponding to residue numbers S291 (e.g., S291M), S292 (e.g., S292G), T425 (e.g., T425G), S428 (e.g., S428G), and T436 (e.g., T436G) in SEQ ID NO: 80 or 83. The chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide may comprise the sequence of SEQ ID NO: 82, with methionine at position 291 and glycine at positions 292, 299, 301, 422, 425, and 433. In some embodiments, the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide may contain the sequence of SEQ ID NO: 82, having methionine at position 291 and glycine at positions 292, 298, 299, 300, 301, 302, 422, 425 and 433.

[0165] Secretion signal peptide (SS) sequence The *Propionibacterium acnes* CAMP2 polypeptide, *Propionibacterium acnes* DsA1 polypeptide, *Propionibacterium acnes* DsA2 polypeptide, *Propionibacterium acnes* PITP polypeptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide, and / or chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein may contain a secretion signal peptide (SS) sequence. The secretion signal peptide can be cleaved during post-translational processing of the *Propionibacterium acnes* polypeptides described herein. Therefore, the mature form of the *Propionibacterium acnes* polypeptide may not contain the secretion signal peptide sequence. However, the nucleotide sequence encoding the secretion signal peptide sequence may be present in the nucleic acid described herein that encodes the *Propionibacterium acnes* polypeptide described herein.

[0166] The *Propionibacterium acnes* CAMP2 polypeptide, *Propionibacterium acnes* DsA1 polypeptide, *Propionibacterium acnes* DsA2 polypeptide, *Propionibacterium acnes* PITP polypeptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide, and / or chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein may contain the secretion signal peptide sequence of the corresponding native *Propionibacterium acnes* polypeptide. This may be advantageous if the *Propionibacterium acnes* polypeptide is expressed as a recombinant protein in prokaryotic cells.

[0167] In some embodiments, the *Dermatitis acnes* CAMP2 polypeptide, *Dermatitis acnes* DsA1 polypeptide, *Dermatitis acnes* DsA2 polypeptide, *Dermatitis acnes* PITP polypeptide, chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide, chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide, and / or chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein may contain a viral or eukaryotic (e.g., human) secretory signal peptide sequence. Using a viral or eukaryotic secretory signal peptide sequence attached to the polypeptides described herein can provide numerous advantages for immunogenic compositions. When expressed from mRNA, particularly in eukaryotic cells, polypeptides of the present invention containing the SS sequence can have increased extracellular expression relative to polypeptides without the SS sequence. Increased extracellular expression can promote higher immunogenicity, thereby enhancing vaccine efficacy.

[0168] Viral SS sequences can be found in publicly accessible databases (such as NCBI or UniProt databases), which include annotated viral peptide sequences and identify experimentally validated start and end positions of the SS.

[0169] In some embodiments, the SignalP algorithm can be used to predict the SS sequence of a given known input polypeptide sequence and the location of the SS sequence cleavage site. The SignalP algorithm (more specifically SignalP v6.0) is described in more detail in Armenteros et al. (Nature Biotechnology. 37: 420-423. 2019), Teufel et al. (Nature Biotechnology. 40: 1023-1025. 2022) and https: / / services.healthtech.dtu.dk / services / SignalP-6.0 / , each of which is incorporated herein by reference in its entirety. The strength of the prediction is assessed based on a cumulative ranking score that considers the likelihood of detecting typical features of the signal sequence (SS probability score) and the probability of cleavage at the cleavage site (cleavage probability score). In some embodiments, the viral secretion signal peptide has a SignalP cleavage probability score of at least 0.8, at least 0.85, at least 0.90, or at least 0.95, as determined using SignalP 6.0. In some embodiments, the viral secretion signal peptide has a SignalP cleavage probability score of at least 0.8, at least 0.85, at least 0.90, or at least 0.95, as determined using SignalP 6.0. 6.0 The SignalP signal peptide probability score is determined to be at least 0.8, at least 0.85, at least 0.90, or at least 0.95.

[0170] In some embodiments, the SS sequence is a viral SS sequence. In some embodiments, the viral secretion signal peptide sequence is derived from a viral sequence of a virus capable of infecting humans. The descriptions preceding "secretion signal peptide sequence" such as "influenza," "SARS-CoV-2," "varicella-zoster virus (VZV)," "measles," "rubella," "rabies," "Ebola," and "smallpox" indicate that the secretion signal peptide is derived from the virus corresponding to that name.

[0171] In some embodiments, the viral secretion signal peptide is derived from viral sequences selected from the group consisting of: influenza secretion signal peptide sequences, SARS-CoV-2 secretion signal peptide sequences, varicella-zoster virus (VZV) secretion signal peptide sequences, measles secretion signal peptide sequences, rubella secretion signal peptide sequences, mumps secretion signal peptide sequences, Ebola secretion signal peptide sequences, rabies secretion signal peptide sequences, and smallpox secretion signal peptide sequences. These specific signal peptides are derived from viral sequences in viruses that have been administered to humans as vaccines (live attenuated vaccines, inactivated vaccines, or mRNA vaccines) and have demonstrated a strong safety profile.

[0172] In some embodiments, the viral secretion signal peptide is selected from the group consisting of: influenza hemagglutinin (HA) secretion signal peptide sequence, SARS CoV-2 spike secretion signal peptide sequence, VZV gB secretion signal peptide sequence, VZV gE secretion signal peptide sequence, VZV gI secretion signal peptide sequence, VZV gK secretion signal peptide sequence, measles F-protein secretion signal peptide sequence, rubella E1 protein secretion signal peptide sequence, rubella E2 protein secretion signal peptide sequence, mumps F-protein secretion signal peptide sequence, Ebola GP protein secretion signal peptide sequence, rabies virus glycoprotein (rabies G) secretion signal peptide sequence, and smallpox 6kDa IC protein secretion signal peptide sequence.

[0173] In some embodiments, the viral secretion signal peptide comprises an HA secretion signal peptide sequence from influenza A or influenza B, preferably from influenza A.

[0174] The exemplary viral secretion signal peptide amino acid sequences disclosed herein are shown in Table 2 below. The exemplary viral secretion signal peptide amino acid sequences derived from influenza A or B are shown in Table 3 below.

[0175] Table 2 – Amino acid sequences of viral secretion signal peptide (SS) Name of protein organism strain SS sequence HA (H1N1) Influenza virus A / New Caledonia / 20 / 1999 MKAKLLVLLCTFTATYA (SEQ ID NO: 210) HA (H1N1pdm) Influenza virus A / California / 7 / 2009 MKAILVVLLYTFATANA (SEQ ID NO: 211) HA (H3N2) Influenza virus A / Moscow / 10 / 1999 MKTIIALSYILCLVFA (SEQ ID NO: 212) HA B Influenza virus B / Phuket / 3073 / 2013 MKAIIVLLMVVTSNA (SEQ ID NO: 213) spike SARSCoV-2 Wuhan-1 MFVFLVLLPLVS (SEQ ID NO: 214) spike SARSCoV-2 Wuhan-1 (Long Version) MFLLTTKRTMFVFLVLLPLVS (SEQ ID NO: 215) gB VZV Oka strain MSPCGYYSKWRNRDRPEYRRNLRFRRFFSSIHPNAAAGSGFNGPGVFITSVTGVWLCFLCIFSMFVTAVVS (SEQ ID NO: 216) gE VZV Oka strain MGTVNKPVVGVLMGFGIITGTLRITNPVRA (SEQ ID NO: 217) gI VZV Oka strain MFLIQCLISAVIFYIQVTNA (SEQ ID NO: 218) gK VZV Oka strain MQALGIKTEHFIIMCLLSGHA (SEQ ID NO: 219) F measles Edmonston-Zagreb strain MGLKVNVSAIFMAVLLTLQTPTG (SEQ ID NO: 220) E1 rubella RA27 / 3 strain MGAAALTAVVLQGYNPPAYG (SEQ ID NO: 221) E2 rubella RA27 / 3 strain MGAPQAFLAGLLLAAVAVGTARA (SEQ ID NO: 222) F mumps Miyahara strain MKVFLVTCLGFAVFSSSSVC (SEQ ID NO: 223) GP Ebola virus Mayinga-76 strain MGVTGILQLPRDRFKRTSFFLWVIILFQRTFS (SEQ ID NO: 224) 6 kDa IC smallpox Germany 91-3 strain MRSLIIFLLFPSIIYS (SEQ ID NO: 225) Rabies G Rabies Rabies Pasteur strain MVPQALLFVPLLVFPLCFG (SEQ ID NO: 226) Table 3 – Amino acid sequences of influenza A and B specific viral secretion signal peptides (SS) strain name or ID SS sequence A / Beijing / 262 / 95 (H1N1)-like virus MKAKLLVLLCTFTATYA (SEQ ID NO: 210) A / Brisbane / 02 / 2018 (H1N1) pdm09-like virus MKAILVVLLYTFTTANA (SEQ ID NO: 227) A / Brisbane / 59 / 2007 (H1N1)-like virus MKVKLLVLLCTFTATYA (SEQ ID NO: 228) A / California / 7 / 2009 (H1N1)-like virus MKAILVVLLYTFATANA (SEQ ID NO: 211) A / Guangdong-Maonan / SWL1536 / 2019 (H1N1)pdm09-like virus MKAILVVLLYTFTTANA (SEQ ID NO: 227) A / Hawaii / 70 / 2019 (H1N1) pdm09-like virus MKAILVVLLYTFTTANA (SEQ ID NO: 227) A / Michigan / 45 / 2015 (H1N1) pdm09-like virus MKAILVVLLYTFTTANA (SEQ ID NO: 227) A / New Caledonia / 20 / 99 (H1N1)-like virus MKAKLLVLLCTFTATYA (SEQ ID NO: 210) A / Solomon Islands / 3 / 2006 (H1N1)-like virus MKVKLLVLLCTFTATYA (SEQ ID NO: 228) A / Sydney / 5 / 2021 (H1N1) pdm09-like virus MKAILVVMLYTFTTANA (SEQ ID NO: 229) A / Victoria / 2570 / 2019 (H1N1) pdm09-like virus MKAILVVMLYTFTTANA (SEQ ID NO: 229) A / Victoria / 4897 / 2022 (H1N1)pdm09-like virus MKAILVVMLYTFTTANA (SEQ ID NO: 229) A / Wisconsin / 588 / 2019 (H1N1)pdm09-like virus MKAILVVMLYTFTTANA (SEQ ID NO: 229) A / Wisconsin / 67 / 2022 (H1N1)pdm09-like virus MKAILVVMLYTFTTANA (SEQ ID NO: 229) H1N1 has a total of SEQ #1 (without substitution). MKAILVVLLYTFTTANA (SEQ ID NO: 227) A / Brisbane / 10 / 2007 (H3N2)-like virus MKTIIALSYILCLVFT (SEQ ID NO: 230) A / California / 7 / 2004 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Cambodia / e0826360 / 2020 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Darwin / 6 / 2021 (H3N2)-like virus MKTIIALSNILCLVFA (SEQ ID NO: 231) A / Darwin / 9 / 2021 (H3N2)-like virus MKTIIALSNILCLVFA (SEQ ID NO: 231) A / Fujian / 411 / 2002 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Hong Kong / 2671 / 2019 (H3N2)-like virus MKAIIALSNILCLVFA (SEQ ID NO: 232) A / Hong Kong / 45 / 2019 (H3N2)-like virus MKAIIALSNILCLVFA (SEQ ID NO: 232) A / Hong Kong / 4801 / 2014 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Kansas / 14 / 2017 (H3N2)-like virus MKTIIALSCILCLVFA (SEQ ID NO: 233) A / Moscow / 10 / 99 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Perth / 16 / 2009 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Singapore / INFIMH-16-0019 / 2016 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / South Australia / 34 / 2019 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Switzerland / 8060 / 2017 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Switzerland / 9715293 / 2013 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Sydney / 5 / 97 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Texas / 50 / 2012 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Victoria / 361 / 2011 (H3N2)-like virus MKTIIALSHILCLVFA (SEQ ID NO: 234) A / Wellington / 1 / 2004 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) A / Wisconsin / 67 / 2005 (H3N2)-like virus MKTIIALSYILCLVFA (SEQ ID NO: 212) H3N2 has SEQ #1 (without substitution). MKTIIALSYILCLVFA (SEQ ID NO: 212) B / Austria / 1359417 / 2021 (B / Victoria lineage) sample MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Brisbane / 60 / 2008-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Colorado / 06 / 2017-like virus (B / Victoria / 2 / 87 lineage) MKAIIVLLMVVTSSA (SEQ ID NO: 235) B / Hong Kong / 330 / 2001-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Malaysia / 2506 / 2004-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Washington / 02 / 2019 (B / Victoria lineage)-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Beijing / 184 / 93-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Florida / 4 / 2006-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Massachusetts / 2 / 2012-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Phuket / 3073 / 2013-like virus MKAIIVLLMVVTSNA (SEQ ID NO: 213) B / Sichuan / 379 / 99-like virus MEAIIVLLMVVTSNA (SEQ ID NO: 236) The common sequence of influenza B (VICTORIA / YAMAGATA) (without substitutions) MKAIIVLLMVVTSNA (SEQ ID NO: 213) H1N1 has SEQ #2 (which has substitution). <![CDATA[MKX1X2LX3VX4LX5TFX6X7X8X9AX1 is selected from A and V; X2 is selected from I and K; X3 is selected from V and L; X4 is selected from L and M; X5 is selected from Y and C; X6 is selected from T and A; X7 is selected from T and A; X8 is selected from A and T; X9 is selected from N and Y (SEQ ID NO: 237)]]> H3N2 has SEQ #2 (which has substitution). <![CDATA[MKX1IIALSX2ILCLVFX3X1 is selected from T and A; X2 is selected from Y, N, C, and H; X3 is selected from T and A (SEQ ID NO: 238)]]> Influenza B (VICTORIA) has a total of SEQ (with substitutions). <![CDATA[MKAIIVLLMVVTSX1AX1 is selected from S and N (SEQ ID NO: 239)]]> Influenza B (YAMAGATA) has a total of SEQ (with substitutions). <![CDATA[MX1AIIVLLMVVTSNAX1 is selected from K and E (SEQ ID NO: 240)]]> The secretion signal peptide sequence may be located at the N-terminus or C-terminus of the polypeptide described herein (e.g., at the N-terminus).

[0176] In some embodiments, the SS amino acid sequence is encoded by a codon-optimized polynucleotide sequence.

[0177] In some embodiments, the viral secretion signal peptide is derived from a viral sequence in a virus capable of infecting humans.

[0178] In some embodiments, the viral secretion signal peptide is derived from viral sequences selected from the group consisting of: influenza secretion signal peptide sequences; and non-influenza secretion signal peptide sequences selected from the group consisting of: SARS CoV-2 secretion signal peptide sequences, varicella-zoster virus (VZV) secretion signal peptide sequences, measles secretion signal peptide sequences, rubella secretion signal peptide sequences, mumps secretion signal peptide sequences, Ebola secretion signal peptide sequences, smallpox secretion signal peptide sequences, and rabies secretion signal peptide sequences.

[0179] In some embodiments, the viral secretion signal peptide is selected from the group consisting of: influenza hemagglutinin (HA) secretion signal peptide sequence, SARS CoV-2 spike secretion signal peptide sequence, VZV gB secretion signal peptide sequence, VZV gE secretion signal peptide sequence, VZV gI secretion signal peptide sequence, VZV gK secretion signal peptide sequence, measles F protein secretion signal peptide sequence, rubella E1 protein secretion signal peptide sequence, rubella E2 protein secretion signal peptide sequence, mumps F protein secretion signal peptide sequence, Ebola GP protein secretion signal peptide sequence, smallpox 6 kDa IC protein secretion signal peptide sequence, and rabies G protein secretion signal peptide sequence. Preferably, the viral secretion signal peptide comprises an HA secretion signal peptide sequence from influenza A or influenza B, more preferably an HA secretion signal peptide sequence from influenza A.

[0180] In some embodiments, the HA secretion signal peptide sequence comprises the amino acid sequence MKX1X2LX3VX4LX5TFX6X7X8X9A (SEQ ID NO: 237), wherein X1 is selected from A and V; X2 is selected from I and K; X3 is selected from V and L; X4 is selected from L and M; X5 is selected from Y and C; X6 is selected from T and A; X7 is selected from T and A; X8 is selected from A and T; and X9 is selected from N and Y.

[0181] In some embodiments, the HA secretion signal peptide sequence comprises an amino acid sequence selected from MKAKLLVLLCTFTATYA (SEQ ID NO: 210), MKAILVVLLYTFTTANA (SEQ ID NO: 227), MKVKLLVLLCTFTATYA (SEQ ID NO: 228), MKAILVVLLYTFATANA (SEQ ID NO: 211), and MKAILVVMLYTFTTANA (SEQ ID NO: 229).

[0182] In some embodiments, the HA secretion signal peptide sequence comprises the amino acid sequence MKX1IIALSX2ILCLVFX3 (SEQ ID NO: 238), wherein X1 is selected from T and A; X2 is selected from Y, N, C and H; and X3 is selected from T and A.

[0183] In some embodiments, the HA secretion signal peptide sequence comprises an amino acid sequence selected from MKTIIALSYILCLVFT (SEQ ID NO: 230), MKTIIALSYILCLVFA (SEQ ID NO: 212), MKTIIALSNILCLVFA (SEQ ID NO: 231), MKAIIALSNILCLVFA (SEQ ID NO: 232), MKTIIALSCILCLVFA (SEQ ID NO: 233), and MKTIIALSHILCLVFA (SEQ ID NO: 234).

[0184] In some embodiments, the HA secretion signal peptide sequence comprises the amino acid sequence MKAIIVLLMVVTSX1A (SEQ ID NO: 239), wherein X1 is selected from S and N.

[0185] In some embodiments, the HA secretion signal peptide sequence comprises the amino acid sequence MX1AIIVLLMVVTSNA (SEQ ID NO: 240), wherein X1 is selected from K and E.

[0186] In some embodiments, the HA secretion signal peptide sequence comprises an amino acid sequence selected from MKAIIVLLMVVTSNA (SEQ ID NO: 213), MKAIIVLLMVVTSSA (SEQ ID NO: 235), and MEAIIVLLMVVTSNA (SEQ ID NO: 236).

[0187] In some embodiments, the viral secretion signal peptide comprises an amino acid sequence selected from the group consisting of: MKAKLLVLLCTFTATYA (SEQ ID NO: 210); MKAILVVLLYTFATANA (SEQ ID NO: 211); MKTIIALSYILCLVFA (SEQ ID NO: 212); MKAIIVLLMVVTSNA (SEQ ID NO: 213); MFVFLVLLPLVS (SEQ ID NO: 214); MFLLTTKRTMFVFLVLLPLVS (SEQ ID NO: 215); MSPCGYYSKWRNRDRPEYRRNLRFRRFFSSIHPNAAAGSGFNGPGVFITSVTGVWLCFLCIFSMFVTAVVS (SEQ ID NO: 216); MGTVNKPVVGVLMGFGIITGTLRITNPVRA (SEQ ID NO: 217); MFLIQCLISAVIFYIQVTNA (SEQ ID NO: 218). 218); 222); MKVFLVTCLGFAVFSSSVC (SEQ ID NO: 223); MGVTGILQLPRDRFKRTSFFLWVIILFQRTFS (SEQ ID NO: 224); MRSLIIFLLFPSIIYS (SEQ ID NO: 225); and MVPQALLFVPLLVFPLCFG (SEQ ID NO: 226).

[0188] In some embodiments, the viral secretion signal peptide comprises the amino acid sequence of MKAKLLVLLCTFTATYA (SEQ ID NO: 210).

[0189] In some embodiments, the viral secretion signal peptide is located at the N-terminus of the polypeptide disclosed herein.

[0190] In some embodiments, the viral secretion signal peptide is located at the C-terminus of the polypeptide disclosed herein.

[0191] In some embodiments, the viral secretion signal peptide is linked to the polypeptide disclosed herein via a linker.

[0192] Heterogeneous transmembrane domain (TMB) The *Propionibacterium acnes* CAMP2 peptide, *Propionibacterium acnes* DsA1 peptide, *Propionibacterium acnes* DsA2 peptide, *Propionibacterium acnes* PITP peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide, and / or chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide described herein may contain a transmembrane domain (TMB), more specifically, a heterologous (non-natural) TMB. The inclusion of a heterologous TMB may be advantageous because it will allow the antigen to be localized to the cell membrane. Compared to antigens without a TMB sequence, this can reduce the intracellular localization of the antigen and further promote higher immunogenicity.

[0193] The *Dermatitis acnes* CAMP2 polypeptide, *Dermatitis acnes* DsA1 polypeptide, *Dermatitis acnes* DsA2 polypeptide, *Dermatitis acnes* PITP polypeptide, chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide, chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide, and / or chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide containing a heterologous transmembrane domain may also contain a secretion signal peptide sequence. Typically, the nucleic acid encoding the chimeric *Dermatitis acnes* CAMP2 polypeptide containing a heterologous transmembrane domain described herein also contains a nucleotide sequence encoding a secretion signal peptide sequence. Typically, the nucleic acid encoding the *Dermatitis acnes* DsA1 polypeptide containing a heterologous transmembrane domain described herein also contains a nucleotide sequence encoding a secretion signal peptide sequence. Typically, the nucleic acid encoding the *Dermatitis acnes* DsA2 polypeptide containing a heterologous transmembrane domain described herein also contains a nucleotide sequence encoding a secretion signal peptide sequence. Typically, the nucleic acid encoding a *Propionibacterium acnes* PITP polypeptide containing a heterologous transmembrane domain, as described herein, also contains a nucleotide sequence encoding a secretion signal peptide sequence. Typically, the nucleic acid encoding a chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide containing a heterologous transmembrane domain, as described herein, also contains a nucleotide sequence encoding a secretion signal peptide sequence. Typically, the nucleic acid encoding a chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide containing a heterologous transmembrane domain, as described herein, also contains a nucleotide sequence encoding a secretion signal peptide sequence. Typically, the nucleic acid encoding a chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide containing a heterologous transmembrane domain, as described herein, also contains a nucleotide sequence encoding a secretion signal peptide sequence.

[0194] TMBs can be derived from any known TMB in the art, including but not limited to TMBs derived from eukaryotic transmembrane proteins (e.g., mammalian transmembrane proteins, such as human transmembrane proteins), prokaryotic transmembrane proteins, and viral transmembrane proteins. TMBs can also be identified by computer simulation prediction algorithms, for example, in the TMHMM prediction methods described below: Krogh et al. (J Mol Biol. [Journal of Molecular Biology] 305(3): 567-580.2001) and https: / / services.healthtech.dtu.dk / services / TMHMM-2.0 / , each of which is incorporated herein by reference in its entirety. Some characteristics of TMBs are described in more detail in Albers et al. (Chapter 2 – cell membrane structures and functions. [Basic Neurochemistry eighth edition.] pp. 26-39. 2012), which is incorporated herein by reference. TMB is typically, but not exclusively, composed primarily of nonpolar (hydrophobic) amino acid residues and can cross the lipid bilayer once or multiple times. Methods for determining the hydrophobicity of amino acids are well known to those skilled in the art. See Simm et al. (2016), Biol Res., 49(1):31; Wimlet and White (1996), Nat Struct Biol., 3(10): 842-848; https: / / blanco.biomol.uci.edu / hydrophobicity_scales.html; and https: / / www.cgl.ucsf.edu / chimera / docs / UsersGuide / midas / hydrophob.html.

[0195] Transmembrane membranes (TMBs) typically contain α-helices, each containing 18-21 amino acids sufficient to span the lipid bilayer. Therefore, in some embodiments, the transmembrane domain comprises one or more α-helices. In some embodiments, the TMB: (a) comprises or consists of 15 to 50 amino acid residues, preferably 15 to 30 amino acid residues, more preferably 18 to 25 amino acid residues; and / or (b) comprises at least 50%, at least 55%, or at least 60% of hydrophobic amino acid residues, preferably selected from the group consisting of alanine, isoleucine, leucine, valine, phenylalanine, tryptophan, and tyrosine; and / or (c) comprises at least one α-helix.

[0196] In some embodiments, the transmembrane domain is derived from an integrated membrane protein, which, as further defined below and in Albers et al., is a membrane protein (also referred to as an intrinsic membrane protein) that is permanently attached to the lipid membrane. In some embodiments, the transmembrane domain is derived from an integrated polytopic protein. An integrated polytopic protein is a protein that spans the entire membrane. In some embodiments, the transmembrane domain is derived from a single-pass (transmembrane-crossing) protein, more particularly a bitopic membrane protein, such as type I or type II. A single-pass protein crosses the membrane only once (i.e., a bitopic membrane protein), while a bitopic protein shuttles in and out of the membrane several times. Single-pass proteins can be classified as type I (whose position causes the carboxyl terminus to face the cytoplasm) or type II (whose amino terminus faces the cytoplasm). In some embodiments, the transmembrane domain is derived from an integrated monotopic protein. A complete monotopic protein is a protein that binds to the membrane from only one side and does not completely span the lipid bilayer.

[0197] In some embodiments, the heterogeneous transmembrane domains are derived from non-human sequences.

[0198] In some embodiments, the heterologous transmembrane domain is derived from a viral sequence. The phrases preceding the phrase "transmembrane domain sequence" such as "influenza," "SARS-CoV-2," "varicella-zoster virus (VZV)," "measles," "rubella," "rabies," "Ebola," and "smallpox" indicate that the transmembrane domain sequence is derived from the virus corresponding to that name.

[0199] In some embodiments, the heterologous transmembrane domain is derived from viral transmembrane domain sequences selected from the group consisting of: influenza transmembrane domain sequences, SARS-CoV-2 transmembrane domain sequences, varicella-zoster virus (VZV) transmembrane domain sequences, measles transmembrane domain sequences, rubella transmembrane domain sequences, mumps transmembrane domain sequences, rabies transmembrane domain sequences, and Ebola transmembrane domain sequences. These specific transmembrane domains are derived from viral sequences in viruses administered to humans as vaccines (live attenuated vaccines, inactivated vaccines, or mRNA vaccines) that have demonstrated a robust safety profile.

[0200] In some embodiments, the heterologous transmembrane domain is selected from the group consisting of: influenza hemagglutinin (HA) transmembrane domain sequence, SARS CoV-2 spike transmembrane domain sequence, VZV gB transmembrane domain sequence, VZV gE transmembrane domain sequence, VZV gI transmembrane domain sequence, VZV gK transmembrane domain sequence, measles F-protein transmembrane domain sequence, rubella E1 protein transmembrane domain sequence, rubella E2 protein transmembrane domain sequence, mumps F-protein transmembrane domain sequence, rabies virus glycoprotein (rabies G) transmembrane domain sequence, and Ebola GP protein transmembrane domain sequence.

[0201] In some embodiments, the heterologous transmembrane domain comprises an HA transmembrane domain sequence derived from influenza A or influenza B, preferably from influenza A.

[0202] The exemplary viral transmembrane domain amino acid sequences disclosed herein are shown in Table 4 below.

[0203] Table 4 – Amino acid sequences of viral transmembrane domain (TMB) signals Name of protein organism strain TMB sequence HA (H1N1) Influenza virus A / New Caledonia / 20 / 1999 ILAIYSTVASSLVLLVSLGAISF (SEQ ID NO: 208) HA (H1N1pdm) Influenza virus A / California / 7 / 2009 ILAIYSTVASSLVLVVSLGAISF (SEQ ID NO: 209) HA (H3N2) Influenza virus A / Moscow / 10 / 1999 ILWISFAISCFLLCVVLLGFI (SEQ ID NO: 241) HA B Influenza virus B / Phuket / 3073 / 2013 STAASSLAVTLMLAIFIVYMV (SEQ ID NO: 242) spike SARS CoV-2 Wuhan-1 WYIWLGFIAGLIAIVMVTIML (SEQ ID NO: 243) gB VZV Oka strain FGALAVGLLVLAGLVAAFFAY (SEQ ID NO: 244) gE VZV Oka strain AAWTGGLAAVVLLCLVIFLIC (SEQ ID NO: 245) gI VZV Oka strain IIIPIVASVMILTAMVIVIVI (SEQ ID NO: 246) gK VZV Oka strain YFWCVQLKMIFFAWFVYGMYL (SEQ ID NO: 247) F measles Edmonston-Zagreb strain IVYILIAVCLGLIGIPALIC (SEQ ID NO: 248) E1 rubella RA27 / 3 strain LDHAFAAFVLLVPWVLIFMVC (SEQ ID NO: 249) E2 rubella RA27 / 3 strain WWQLTLGAICALLLAGLLACC (SEQ ID NO: 250) F mumps Miyahara strain IVALVLSILSIIISLLFCCW (SEQ ID NO: 251) GP Ebola Mayinga-76 strain WIPAGIGVTGVIIAVIALFCI (SEQ ID NO: 252) Rabies G Rabies Rabies Pasteur strain VLLSAGALTALMLIIFLMTCW (SEQ ID NO: 253) In some embodiments, the heterologous TMB sequence is located at the N-terminus or C-terminus (e.g., the C-terminus) of the peptide described herein.

[0204] In some embodiments, the TMB amino acid sequence is encoded by a codon-optimized polynucleotide sequence.

[0205] In some embodiments, one or more of the *Propionibacterium acnes* CAMP2 peptide, *Propionibacterium acnes* DsA1 peptide, *Propionibacterium acnes* DsA2 peptide, *Propionibacterium acnes* PITP peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide, and / or chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide contain the heterologous transmembrane domain described herein. In alternative embodiments, the *Propionibacterium acnes* CAMP2 peptide, *Propionibacterium acnes* DsA1 peptide, *Propionibacterium acnes* DsA2 peptide, *Propionibacterium acnes* PITP peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide, chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide, and / or chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide do not contain the heterologous transmembrane domain. Such peptides may contain a secretion signal peptide sequence. In further embodiments, the polypeptides of the present invention are secreted. In some embodiments, the *Dermatitis acnes* CAMP2 polypeptide described herein is a secreted polypeptide. In some embodiments, the *Dermatitis acnes* DsA1 polypeptide described herein is a secreted polypeptide. In some embodiments, the *Dermatitis acnes* DsA2 polypeptide described herein is a secreted polypeptide. In some embodiments, the *Dermatitis acnes* PITP polypeptide described herein is a secreted polypeptide. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein is a secreted polypeptide. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide described herein is a secreted polypeptide. In some embodiments, the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein is a secreted polypeptide. The secreted polypeptides described herein contain a secretion signal peptide sequence.

[0206] In some embodiments, the TMB: (a) comprises or consists of 15 to 50 amino acid residues, preferably 15 to 30 amino acid residues, more preferably 18 to 25 amino acid residues; and / or (b) comprises at least 50% hydrophobic amino acid residues, which are preferably selected from the group consisting of alanine, isoleucine, leucine, valine, phenylalanine, tryptophan, and tyrosine; and / or (c) comprises at least one α-helix.

[0207] In some embodiments, TMB is derived from an integrated membrane protein, preferably from a single-pass membrane protein, more preferably from a two-site membrane protein, and even more preferably from a type I two-site membrane protein.

[0208] In some embodiments, the TMB is derived from a non-human sequence.

[0209] In some embodiments, TMB is derived from a viral sequence.

[0210] In some embodiments, the TMB is derived from viral transmembrane domain sequences selected from the group consisting of: influenza transmembrane domain sequences; and non-influenza transmembrane domain sequences selected from the group consisting of: SARS CoV-2 transmembrane domain sequences, varicella-zoster virus (VZV) transmembrane domain sequences, measles transmembrane domain sequences, rubella transmembrane domain sequences, mumps transmembrane domain sequences, Ebola transmembrane domain sequences, and rabies transmembrane domain sequences.

[0211] In some embodiments, the TMB is selected from the group consisting of: influenza hemagglutinin (HA) transmembrane domain sequence, SARS CoV-2 spike transmembrane domain sequence, VZV gB transmembrane domain sequence, VZV gE transmembrane domain sequence, VZV gI transmembrane domain sequence, VZV gK transmembrane domain sequence, measles F protein transmembrane domain sequence, rubella E1 protein transmembrane domain sequence, rubella E2 protein transmembrane domain sequence, mumps F protein transmembrane domain sequence, Ebola GP protein transmembrane domain sequence, and rabies G protein transmembrane domain sequence, preferably wherein the TMB contains an HA transmembrane domain sequence from influenza A or influenza B, more preferably from influenza A.

[0212] In some embodiments, the TMB comprises an amino acid sequence selected from the group consisting of: ILAIYSTVASSLVLLVSLGAISF (SEQ ID NO: 208); ILAIYSTVASS LVLVVSLGAISF (SEQ ID NO: 209); ILWISFAISCFLLCVVLLGFI (SEQ ID NO: 241); STAASSLAVTLMLAIFIVYMV (SEQ ID NO: 242); WYIWLGFIAGLIAIVMVTIML (SEQ ID NO: 243); FGALAVGLLVLAGL VAAFFAY (SEQ ID NO: 244); AAWTGGLAAVVLLCLVIFLIC (SEQ ID NO: 245); IIIPIVASVMILTAMVIVIVI (SEQ ID NO: 246); YFWCVQ LKMIFFAWFVYGMYL (SEQ ID NO: 247); IVYILIAVCLGGLIGIPALIC (SEQ ID NO: 248). NO: 248); LDHAFAAFVLLVPWVLIFMVC (SEQ ID NO: 249); WWQLTLGAICALLLAGLLACC (SEQ ID NO: 250); IVAALVLSILSIIISL LFCCW (SEQ ID NO: 251); WIPAGIGVTGVIIAVIALFCI (SEQ ID NO: 252); and VLLSAGALTALMLIIFLMTCW (SEQ ID NO: 253).

[0213] In some embodiments, TMB contains the amino acid sequence of ILAYSTVASSLVLLVSLGAISF (SEQ ID NO:208).

[0214] In some embodiments, TMB is linked to the polypeptide described herein via a connector.

[0215] In some embodiments, TMB is located at the N-terminus of the polypeptide described herein.

[0216] In some embodiments, TMB is located at the C-terminus of the polypeptide described herein.

[0217] connector In some embodiments, the polypeptide of the present invention includes a linker. In some embodiments, the polypeptide of the present invention includes two or more antigens attached via the linker.

[0218] In some embodiments disclosed herein, a secretory signal peptide (SS) sequence or a transmembrane domain (TMB) is directly fused to the polypeptide described herein (i.e., there is no linker, such as an amino acid linker, connecting the SS sequence or TMB to the polypeptide described herein).

[0219] In other embodiments, the SS sequence and TMB disclosed herein are optionally attached to the polypeptide described herein via a linker. In some embodiments, the linker is an amino acid linker. In some embodiments, the length of the amino acid linker is from 1 to 10 amino acids (e.g., the amino acid linker has a length of 1, 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acids).

[0220] Exemplary examples of connectors include glycine polymer (Gly)n, where n is an integer of at least 1, 2, 3, 4, 5, 6, 7, or 8; glycine-serine polymer (GlySer)n, where n is an integer of at least one, two, three, four, five, six, seven, or eight; glycine-alanine polymer; alanine-serine polymer; and other flexible connectors known in the art.

[0221] Glycine and glycine-serine polymers are relatively unstructured and flexible, thus capable of serving as neutral linkers between the SS sequence and / or TMB and the peptides described herein. In some embodiments, the linker is SGS or GSG.

[0222] Other exemplary linkers include, but are not limited to, the following amino acid sequences: GGG; DGGGS (SEQ ID NO: 254); TGEKP (SEQ ID NO: 255) (Liu et al., Proc. Natl. Acad. Sci. [Proceedings of the National Academy of Sciences] 94:5525-5530. 1997); GGRR (SEQ ID NO: 256); (GGGGS)n (SEQ ID NO: 257), where n = 1, 2, 3, 4, or 5 (Kim et al., Proc. Natl. Acad. Sci. [Proceedings of the National Academy of Sciences] 93: 1156-1160. 1996); EGKSSGSGSESKVD (SEQ ID NO: 258) (Chaudhary et al., Proc. Natl. Acad. Sci. [Proceedings of the National Academy of Sciences] 87: 1066-1070. 1990); KESGSVSSEQLAQFRSLD (SEQ ID NO: 259) (Bird et al. Science. [Science] 242:423-426. 1988), GGRRGGGS (SEQ ID NO: 260); LRQRDGERP (SEQ ID NO: 261); LRQKDGGGSERP (SEQ ID NO: 262); and GSTGSGSGKPGSGEGSTKG (SEQ ID NO: 263) (Cooper et al. Blood. [Blood] 101(4): 1637-1644. 2003). Preferred linkers are shorter, for example, consisting of 3, 4, or 5 amino acids.

[0223] Additional examples of the connector are provided in Chen et al. (Adv Drug Deliv Rev. 65(10): 1357–1369. 2013), which are incorporated herein by reference.

[0224] Composition—nucleic acids and polypeptides This invention provides a composition comprising one or more nucleic acids disclosed herein. This invention also provides a composition comprising one or more polypeptides disclosed herein. The compositions of this invention can be pharmaceutical compositions, such as those comprising pharmaceutically acceptable carriers, excipients, or diluents. In some embodiments, the compositions of this invention are immunogenic compositions. "Immunogenic composition" means a composition comprising nucleic acids or proteins that, when administered to a subject, elicits an immune response, such as an antigen-specific immune response. The immune response can be a humoral (antibody) immune response or a cell-mediated immune response. The compositions of this invention can be vaccine compositions. Immunogenic compositions (e.g., vaccine compositions) can elicit immunity (e.g., an antibody response) against *Propionibacterium acnes* infection. Antibody responses can include binding to the surface of *Propionibacterium acnes* bacteria and recruiting immune effector cells (e.g., effector cells of the immune system, such as phagocytes). These antibodies can be able to elicit opsonization and phagocytosis in vitro. 。 Antibodies can be cross-reactive across a range of *Propionibacterium acnes* strains. These antibodies can reduce or neutralize cAMP2-mediated inflammation.

[0225] As used herein, “protective immunity” or “protective immune response” refers to the immunity or induction of an immune response against an infectious agent (e.g., *Propionibacterium acnes*) exhibited by a subject in order to prevent or reduce infection or at least one symptom thereof. Specifically, it is apparent that by administering the compositions of the present invention, protective immunity or a protective immune response can be induced in relation to the elimination or reduction of the presence of one or more symptoms of *Propionibacterium acnes* infection. As used herein, the term “immune response” refers to both humoral immune response and cell-mediated immune response. In some embodiments, treatment with the compositions of the present invention described herein provides protective immunity against *Propionibacterium acnes* infection.

[0226] Nucleic acid composition On one hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a polypeptide of *Dermatitis acnes* CAMP2 as described herein.

[0227] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a modified dermatitis acnes CAMP2 polypeptide as described herein.

[0228] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a polypeptide of *Dermatitis acnes* DsA1 as described herein.

[0229] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a polypeptide of *Dermatitis acnes* DsA2 as described herein.

[0230] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a PITP polypeptide of *Dermatophyte acnes* as described herein.

[0231] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a chimeric dermatitis acnes DsA1 / DsA2 polypeptide as described herein.

[0232] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a chimeric dermatophyte DsA1 / DsA2 / PITP polypeptide as described herein.

[0233] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a chimeric dermatophyte DsA1 / DsA2 / PITP / CAMP2 polypeptide as described herein.

[0234] Any composition described herein may comprise one or more nucleic acids encoding one or more antigens, as described in WO2021 / 165543 (incorporated herein by reference).

[0235] CAMP2 composition On one hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a polypeptide of *Dermatitis acnes* CAMP2 as described herein.

[0236] On the other hand, the present invention provides an immunogenic composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a polypeptide of *Dermatitis acnes* CAMP2 as described herein, and wherein the nucleic acid is mRNA.

[0237] On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a polypeptide of *Dermatitis acnes* CAMP2 as described herein, and wherein the composition further comprises an LNP.

[0238] Compositions containing the *Dermatitis acnes* CAMP2 peptide used in this invention, delivered, for example, as mRNA and / or in the form of compositions containing LNP, can induce antibodies in subjects. Such antibodies can neutralize the biological activities of the CAMP2 peptide, such as its inflammatory activity. Such antibodies can also neutralize the co-hemolytic activity of the CAMP2 peptide.

[0239] In some embodiments, any composition comprising (a) a nucleic acid as described herein, comprising a nucleotide sequence encoding the *Dermatitis acnes* CAMP2 polypeptide, further comprises one or more of the following: (b)(i) a nucleic acid as described herein, comprising a nucleotide sequence encoding the *Dermatitis acnes* DsA1 polypeptide; (b)(ii) a nucleic acid as described herein, comprising a nucleotide sequence encoding the *Dermatitis acnes* DsA2 polypeptide; (b)(iii) a nucleic acid as described herein, comprising a nucleotide sequence encoding the *Dermatitis acnes* PITP polypeptide; (b)(iv) a nucleic acid as described herein, comprising a nucleotide sequence encoding the *Dermatitis acnes* DsA1 / DsA2 polypeptide; and (b)(v) a nucleic acid as described herein, comprising a nucleotide sequence encoding the *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide. In some embodiments, the composition comprises the nucleic acid of (b)(i), the nucleic acid of (b)(ii), and the nucleic acid of (b)(iii). In some embodiments, the composition comprises the nucleic acid of (b)(v). In a preferred embodiment, the composition comprises the nucleic acid of (b)(iii) and the nucleic acid of (b)(iv).

[0240] In some embodiments, any two or more nucleic acids in (a) and (b)(i) through (v) are located on the same nucleic acid molecule or on different nucleic acid molecules (e.g., all nucleic acids in the composition are on the same nucleic acid molecule, or all nucleic acids in the composition are on separate nucleic acid molecules). In some embodiments, the nucleic acids in (a), (b)(iii), and (b)(iv) are located on the same nucleic acid molecule or on separate nucleic acid molecules. In some embodiments, the nucleic acids in (a) and (b)(v) are located on the same nucleic acid molecule or on separate nucleic acid molecules. In some embodiments, the composition comprises providing a nucleic acid that contains a nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein. Typically, the nucleic acids in (a), (b)(iii), and (b)(iv) are provided by separate nucleic acid molecules.

[0241] Modified CAMP2 composition On the other hand, the present invention provides a composition comprising a nucleic acid as described herein, the nucleic acid comprising a nucleotide sequence encoding a modified dermatitis acnes CAMP2 polypeptide as described herein.

[0242] In some embodiments, the composition further comprises one or more of the following: (i) a nucleic acid comprising a nucleotide sequence encoding a DsA1 polypeptide of *Propionibacterium acnes* as described herein, (ii) a nucleic acid comprising a nucleotide sequence encoding a DsA2 polypeptide of *Propionibacterium acnes* as described herein, (iii) a nucleic acid comprising a nucleotide sequence encoding a PITP polypeptide of *Propionibacterium acnes* as described herein, (iv) a nucleic acid comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 polypeptide of *Propionibacterium acnes* as described herein, and (v) a nucleic acid comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 / PITP polypeptide of *Propionibacterium acnes* as described herein.

[0243] In some embodiments, the composition comprises nucleic acid of (i), nucleic acid of (ii), and nucleic acid of (iii). In some embodiments, the composition comprises nucleic acid of (v). In a preferred embodiment, the composition comprises nucleic acid of (iii) and nucleic acid of (iv).

[0244] Composition of DsA1, DsA2 and PITP On the other hand, the present invention provides compositions comprising nucleic acids as described herein, the nucleic acids comprising one or more of the following: (i) nucleic acids comprising a nucleotide sequence encoding a DsA1 polypeptide of *Propionibacterium acnes* as described herein, (ii) nucleic acids comprising a nucleotide sequence encoding a DsA2 polypeptide of *Propionibacterium acnes* as described herein, (iii) nucleic acids comprising a nucleotide sequence encoding a PITP polypeptide of *Propionibacterium acnes* as described herein, (iv) nucleic acids comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 polypeptide of *Propionibacterium acnes* as described herein, (v) nucleic acids comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 / PITP polypeptide of *Propionibacterium acnes* as described herein, and (vi) nucleic acids comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 / PITP / CAMP2 polypeptide of *Propionibacterium acnes* as described herein.

[0245] Compared to using individual antigens, the combination of *Propionibacterium acnes* DsA1, DsA2, and PITP peptides can elicit an immune response against a wider number of *Propionibacterium acnes* strains. Compared to using individual antigens, using a combination of DsA1 and / or DsA2 with PITP may elicit an antibody pool that cross-reacts with a wider number of *Propionibacterium acnes* strains. Preferably, the *Propionibacterium acnes* DsA1 and DsA2 peptides are provided as a chimeric *Propionibacterium acnes* DsA1 / DsA2 peptide. The *Propionibacterium acnes* PITP peptide can be provided as a single peptide molecule, as part of a chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP peptide, or as part of a chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP / CAMP2 peptide. Typically, the *Propionibacterium acnes* PITP peptide is provided as a single peptide molecule.

[0246] In some embodiments, the composition comprises a nucleic acid containing a nucleotide sequence encoding a DsA1 polypeptide of *Propionibacterium acnes* as described herein, a nucleic acid containing a nucleotide sequence encoding a DsA2 polypeptide of *Propionibacterium acnes* as described herein, and a nucleic acid containing a nucleotide sequence encoding a PITP polypeptide of *Propionibacterium acnes* as described herein.

[0247] In some embodiments, the composition comprises a nucleic acid comprising, as described herein, a nucleotide sequence encoding a chimeric dermatitis acnes DsA1 / DsA2 polypeptide as described herein, and a nucleic acid comprising, as described herein, a nucleotide sequence encoding a dermatitis acnes PITP polypeptide as described herein. Typically, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises the amino acid sequence according to SEQ ID NO: 70, or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it, and the *Dermatitis acnes* PITP polypeptide comprises the sequence according to SEQ ID NO: 73, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 70 and the *Propionibacterium acnes* PITP polypeptide comprises the sequence of SEQ ID NO: 73. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide consists of the amino acid sequence according to SEQ ID NO: 70, and the *Propionibacterium acnes* PITP polypeptide consists of the amino acid sequence according to SEQ ID NO: 73.

[0248] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises the nucleotide sequence according to SEQ ID NO: 179 or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., 85%) identity with it, and the nucleic acid comprising the nucleotide sequence encoding the dermatitis acnes PITP polypeptide comprises the nucleotide sequence according to SEQ ID NO: 182 or a sequence having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., 75%) identity with it.

[0249] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide consists of the nucleotide sequence encoding the secretion signal peptide sequence (e.g., the viral secretion signal peptide sequence described herein) and the sequence according to SEQ ID NO: 179 or a sequence having at least 85% identity with it, and the nucleic acid comprising the nucleotide sequence encoding the *Dermatitis acnes* PITP polypeptide consists of the nucleotide sequence encoding the secretion signal peptide sequence (e.g., the viral secretion signal peptide sequence described herein) and the sequence according to SEQ ID NO: 182 or a sequence having at least 75% identity with it.

[0250] Any composition of the present invention may comprise a combination of nucleic acids described herein (e.g., they may be formulated in the same composition). Combinations of nucleic acids as described herein may alternatively be in the form of two or more separate compositions (e.g., as a combination of compositions for simultaneous, independent, or sequential administration (e.g., in therapeutic or prophylactic uses as described herein)).

[0251] The compositions of this disclosure, comprising one or more nucleic acids disclosed herein, may also include one or more additional components, such as small molecule immune enhancers (e.g., TLR agonists). The compositions of this disclosure may also include a delivery system for the nucleic acid (e.g., RNA) described herein, such as liposomes, oil-in-water emulsions, or microparticles. In some embodiments, the composition comprises lipid nanoparticles (LNPs). In some embodiments, the composition comprises nucleic acid molecules of the present invention encapsulated within LNPs.

[0252] In some embodiments, the compositions described herein are in the form of a frozen liquid. In some embodiments, the compositions described herein are in a lyophilized form (e.g., a freeze-dried form).

[0253] polypeptide composition On the one hand, the present invention provides a composition comprising the dermatitis acne-causing bacteria CAMP2 polypeptide described herein.

[0254] On the other hand, the present invention provides a composition comprising the modified dermatophyte cAMP2 polypeptide described herein.

[0255] On the other hand, the present invention provides a composition comprising the DsA1 polypeptide of *Dermatophyte acnes* described herein.

[0256] On the other hand, the present invention provides a composition comprising the DsA2 polypeptide of *Dermatophyte acnes* described herein.

[0257] On the other hand, the present invention provides a composition comprising the PITP polypeptide of *Dermatophyte acnes* as described herein.

[0258] On the other hand, the present invention provides a composition comprising the chimeric dermatophyte DsA1 / DsA2 polypeptide described herein.

[0259] On the other hand, the present invention provides a composition comprising the chimeric dermatophyte acnes DsA1 / DsA2 / PITP polypeptide described herein.

[0260] On the other hand, the present invention provides a composition comprising the chimeric dermatophyte DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein.

[0261] Any composition described herein may comprise any polypeptide antigen as described in WO 2021 / 165543.

[0262] CAMP2 composition On the one hand, the present invention provides a composition comprising the dermatitis acne-causing bacteria CAMP2 polypeptide described herein.

[0263] In some embodiments, a composition comprising (a) the *Dermatitis acnes* CAMP2 polypeptide described herein further comprises one or more of the following: (b)(i) the *Dermatitis acnes* DsA1 polypeptide described herein, (b)(ii) the *Dermatitis acnes* DsA2 polypeptide described herein, (b)(iii) the *Dermatitis acnes* PITP polypeptide described herein, (b)(iv) the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein, and (b)(v) the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide described herein. In some embodiments, the composition comprises the polypeptide of (b)(i), the polypeptide of (b)(ii), and the polypeptide of (b)(iii). In some embodiments, the composition comprises the polypeptide of (b)(v). In a preferred embodiment, the composition comprises the polypeptide of (b)(iii) and the polypeptide of (b)(iv).

[0264] In some embodiments, the polypeptides of (a), (b)(iii), and (b)(iv) are provided as chimeric polypeptides, such as the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein. In some embodiments, the polypeptides of (a) and (b)(v) are provided as chimeric polypeptides, such as the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein. Typically, the polypeptides of (a), (b)(iii), and (b)(iv) are provided as individual polypeptides.

[0265] Modified CAMP2 composition On the other hand, the present invention provides a composition comprising the modified dermatophyte cAMP2 polypeptide described herein.

[0266] In some embodiments, a composition comprising the modified *Propionibacterium acnes* CAMP2 polypeptide described herein further comprises one or more of the following: (i) the *Propionibacterium acnes* DsA1 polypeptide described herein, (ii) the *Propionibacterium acnes* DsA2 polypeptide described herein, (iii) the *Propionibacterium acnes* PITP polypeptide described herein, (iv) the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide described herein, and (v) the chimeric *Propionibacterium acnes* DsA1 / DsA2 / PITP polypeptide described herein.

[0267] In some embodiments, the composition comprises the polypeptide of (i), the polypeptide of (ii), and the polypeptide of (iii). In some embodiments, the composition comprises the polypeptide of (v). In a preferred embodiment, the composition comprises the polypeptide of (iii) and the polypeptide of (iv).

[0268] Composition of DsA1, DsA2 and PITP On the other hand, the present invention provides a composition comprising one or more of the following: (i) the *Dermatitis acnes* DsA1 polypeptide described herein, (ii) the *Dermatitis acnes* DsA2 polypeptide described herein, (iii) the *Dermatitis acnes* PITP polypeptide described herein, (iv) the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein, (v) the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide described herein, and (vi) the chimeric DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein.

[0269] In some embodiments, the composition comprises the dermatitis acnes DsA1 polypeptide, the dermatitis acnes DsA2 polypeptide, and the dermatitis acnes PITP polypeptide described herein.

[0270] In some embodiments, the composition comprises the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide and the *Propionibacterium acnes* PITP polypeptide described herein. Typically, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises the amino acid sequence according to SEQ ID NO: 70, or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it, and the *Propionibacterium acnes* PITP polypeptide comprises the sequence according to SEQ ID NO: 73, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 70 and the *Propionibacterium acnes* PITP polypeptide comprises the sequence of SEQ ID NO: 73. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide consists of the amino acid sequence according to SEQ ID NO: 70, and the *Propionibacterium acnes* PITP polypeptide consists of the amino acid sequence according to SEQ ID NO: 73.

[0271] Any composition of the present invention may comprise a combination of peptides described herein (e.g., they may be formulated in the same composition). Combinations of peptides as described herein may alternatively be in two or more separate compositions (e.g., as a combination of compositions for simultaneous, individual or sequential administration (e.g., in therapeutic or preventative uses as described herein)).

[0272] Compositions of this disclosure comprising one or more polypeptides may include adjuvants as described herein. As used herein, "adjuvant" refers to a substance or carrier that enhances the immune response to an antigen. Adjuvants may include, but are not limited to, suspensions of minerals (e.g., alum, aluminum hydroxide, or phosphate) to which antigens are adsorbed; water-in-oil or oil-in-water emulsions of antigen solutions emulsified in mineral oil or water (e.g., Freund's incomplete adjuvant). Sometimes, killed mycobacteria (e.g., Freund's complete adjuvant) are included to further enhance antigenicity. The adjuvant may include squalene-based oil-in-water emulsion adjuvants (e.g., AF03, as described in WO 2007006939 and U.S. Patent No. 8,703,095; AS03, as described in WO 1995017209, WO 1995017210 and U.S. Patent Nos. 6,623,739, 7,029,678 and 7,510,698; and MF59, as described in WO 1990014837 and U.S. Patent Nos. 6,299,884 and 6,451,325). Immunostimulatory oligonucleotides (e.g., CpG motifs) can also be used as adjuvants (e.g., see U.S. Patent Nos. 6,194,388; 6,207,646; 6,214,806; 6,218,371; 6,239,116; 6,339,068; 6,406,705; and 6,429,199). Adjuvants may also include biomolecules such as Toll-like receptor (TLR) agonists (e.g., AS01, as described in WO 2007068907 and U.S. Patent Nos. 10,039,823 and 10,143,745; SPA14, as described in WO 2022090359; and LEQ, as described in WO 2023056089) and co-stimulatory molecules.

[0273] In some embodiments, the adjuvant is selected from the group consisting of: aluminum-based adjuvants (e.g., AlOOH), squalene-based oil-in-water emulsion adjuvants (e.g., AFO3, AS03, MF59), and liposome-based adjuvants containing saponins and TLR4 agonists (e.g., SPA14, LEQ, AS01).

[0274] In a preferred embodiment, the adjuvant is selected from the group consisting of AlOOH, AFO3 and SPA14.

[0275] In some embodiments, the compositions described herein are in the form of a cryoliposome. In some embodiments, the compositions described herein are in the form of a lyophilized liquid.

[0276] Combination—nucleic acids and peptides Nucleic acid combination This invention provides a combination comprising two or more nucleic acids of this invention.

[0277] CAMP2 group In some aspects, the present invention provides combinations of nucleic acids comprising a multinucleotide sequence encoding a *Dermatitis acnes* CAMP2 polypeptide; and one or more of the following: (i) a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* DsA1 polypeptide; (ii) a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* DsA2 polypeptide; (iii) a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* PITP polypeptide; (iv) a nucleic acid comprising a nucleotide sequence encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide; and (v) a nucleic acid comprising a nucleotide sequence encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide.

[0278] In some embodiments, the combination comprises nucleic acid of (i), nucleic acid of (ii), and nucleic acid of (iii). In some embodiments, the composition comprises nucleic acid of (v). In a preferred embodiment, the composition comprises nucleic acid of (iii); and nucleic acid of (iv). In some embodiments, the combination comprises nucleic acid of (v). In a preferred embodiment, the combination comprises nucleic acid of (iii) and nucleic acid of (iv).

[0279] In some embodiments, the combination comprises a nucleic acid containing a nucleotide sequence encoding the *Dermatitis acnes* CAMP2 polypeptide, the nucleic acid in (iii), and the nucleic acid in (iv). In some embodiments, the combination comprises a nucleic acid containing a nucleotide sequence encoding the *Dermatitis acnes* CAMP2 polypeptide and the nucleic acid in (v). In some embodiments, such combinations are provided as nucleic acids containing a nucleotide sequence encoding the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein. Typically, the nucleic acid containing the nucleotide sequence encoding the *Dermatitis acnes* CAMP2 polypeptide, the nucleic acid in (iii), and the nucleic acid in (iv) are provided as separate nucleic acid molecules.

[0280] Modified CAMP2 combination On the other hand, the present invention provides a combination comprising: a nucleic acid comprising a nucleotide sequence encoding a modified *Dermatitis acnes* CAMP2 polypeptide as described herein, and one or more of the following: (i) a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* DsA1 polypeptide as described herein, (ii) a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* DsA2 polypeptide as described herein, (iii) a nucleic acid comprising a nucleotide sequence encoding a *Dermatitis acnes* PITP polypeptide as described herein, (iv) a nucleic acid comprising a nucleotide sequence encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide as described herein, and (v) a nucleic acid comprising a nucleotide sequence encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide as described herein.

[0281] In some embodiments, the combination comprises nucleic acid (i), nucleic acid (ii), and nucleic acid (iii). In some embodiments, the combination comprises nucleic acid (v). In a preferred embodiment, the combination comprises nucleic acid (iii) and nucleic acid (iv).

[0282] DsA1, DsA2 and PITP combination On one hand, the present invention provides a nucleic acid comprising one or more of the following: (i) a nucleic acid comprising a nucleotide sequence encoding a DsA1 polypeptide of *Propionibacterium acnes*; (ii) a nucleic acid comprising a nucleotide sequence encoding a DsA2 polypeptide of *Propionibacterium acnes*; (iii) a nucleic acid comprising a nucleotide sequence encoding a PITP polypeptide of *Propionibacterium acnes*; (iv) a nucleic acid comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 polypeptide of *Propionibacterium acnes*; and (v) a nucleic acid comprising a nucleotide sequence encoding a chimeric DsA1 / DsA2 / PITP polypeptide of *Propionibacterium acnes*.

[0283] In some embodiments, a combination of nucleic acids comprising a nucleotide sequence encoding a DsA1 polypeptide of *Propionibacterium acnes* as described herein, a nucleic acid comprising a nucleotide sequence encoding a DsA2 polypeptide of *Propionibacterium acnes* as described herein, and a nucleic acid comprising a nucleotide sequence encoding a PITP polypeptide of *Propionibacterium acnes* as described herein.

[0284] In some embodiments, a nucleic acid comprising, as described herein, a nucleotide sequence encoding a chimeric dermatitis acnes DsA1 / DsA2 polypeptide as described herein, and a nucleic acid comprising, as described herein, a nucleotide sequence encoding a dermatitis acnes PITP polypeptide as described herein. Typically, the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide comprises the amino acid sequence according to SEQ ID NO: 70, or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it, and the *Dermatitis acnes* PITP polypeptide comprises the sequence according to SEQ ID NO: 73, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 70 and the *Propionibacterium acnes* PITP polypeptide comprises the sequence of SEQ ID NO: 73. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide consists of the amino acid sequence according to SEQ ID NO: 70, and the *Propionibacterium acnes* PITP polypeptide consists of the amino acid sequence according to SEQ ID NO: 73.

[0285] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric dermatitis acnes DsA1 / DsA2 polypeptide comprises the nucleotide sequence according to SEQ ID NO: 179 or a sequence having at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., 85%) identity with it, and the nucleic acid comprising the nucleotide sequence encoding the dermatitis acnes PITP polypeptide comprises the nucleotide sequence according to any one of SEQ ID NO: 182 or a sequence having at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., 75%) identity with it.

[0286] In some embodiments, the nucleic acid comprising the nucleotide sequence encoding the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide consists of the nucleotide sequence encoding the secretion signal peptide sequence (e.g., the viral secretion signal peptide sequence described herein) and the sequence according to SEQ ID NO: 179 or a sequence having at least 85% identity with it, and the nucleic acid comprising the nucleotide sequence encoding the *Dermatitis acnes* PITP polypeptide consists of the nucleotide sequence encoding the secretion signal peptide sequence (e.g., the viral secretion signal peptide sequence described herein) and the sequence according to SEQ ID NO: 182 or a sequence having at least 75% identity with it.

[0287] polypeptide combination This invention provides a combination comprising two or more polypeptides of this invention.

[0288] CAMP2 group In some aspects, the present invention provides a combination comprising the *Dermatitis acnes* CAMP2 polypeptide; and one or more of the following: (i) the *Dermatitis acnes* DsA1 polypeptide described herein, (ii) the *Dermatitis acnes* DsA2 polypeptide described herein, (iii) the *Dermatitis acnes* PITP polypeptide described herein, (iv) the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein, and (v) the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide described herein.

[0289] In some embodiments, the combination comprises the polypeptide of (i), the polypeptide of (ii), and the polypeptide of (iii). In some embodiments, the combination comprises the polypeptide of (v). In a preferred embodiment, the combination comprises the polypeptide of (iii) and the polypeptide of (iv). In some embodiments, such combinations are provided as chimeric polypeptides, such as the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP / CAMP2 polypeptide described herein. Typically, the *Dermatitis acnes* CAMP2 polypeptide, the polypeptide of (iii), and the polypeptide of (iv) are provided as separate polypeptides.

[0290] Modified CAMP2 combination On the other hand, the present invention provides a combination comprising the modified *Dermatitis acnes* CAMP2 polypeptide described herein and one or more of the following: (i) the *Dermatitis acnes* DsA1 polypeptide described herein, (ii) the *Dermatitis acnes* DsA2 polypeptide described herein, (iii) the *Dermatitis acnes* PITP polypeptide described herein, (iv) the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein, and (v) the chimeric *Dermatitis acnes* DsA1 / DsA2 / PITP polypeptide described herein.

[0291] In some embodiments, the combination comprises the polypeptide comprising (i), the polypeptide comprising (ii), and the polypeptide comprising (iii). In some embodiments, the combination comprises the polypeptide comprising (v). In a preferred embodiment, the combination comprises the polypeptide comprising (iii) and the polypeptide comprising (iv).

[0292] Composition of DsA1, DsA2 and PITP On one hand, the present invention provides a combination comprising one or more of the following: (i) the DsA1 polypeptide of *Propionibacterium acnes* described herein, (ii) the DsA2 polypeptide of *Propionibacterium acnes* described herein, (iii) the PITP polypeptide of *Propionibacterium acnes* described herein, (iv) the chimeric DsA1 / DsA2 polypeptide of *Propionibacterium acnes* described herein, and (v) the chimeric DsA1 / DsA2 / PITP polypeptide of *Propionibacterium acnes* described herein.

[0293] In some embodiments, the combination comprises the DsA1 polypeptide, the DsA2 polypeptide, and the PITP polypeptide of *Propionibacterium acnes* described herein.

[0294] In some embodiments, the composition comprises the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide and the *Propionibacterium acnes* PITP polypeptide described herein. Typically, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises the amino acid sequence according to SEQ ID NO: 70, or a sequence having at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it, and the *Propionibacterium acnes* PITP polypeptide comprises the sequence according to SEQ ID NO: 73, or a sequence having at least 75%, at least 80%, at least 85%, at least 86%, at least 87%, at least 88%, at least 89%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% (e.g., at least 90% or at least 95%) identity with it. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide comprises the sequence of SEQ ID NO: 70 and the *Propionibacterium acnes* PITP polypeptide comprises the sequence of SEQ ID NO: 73. In some embodiments, the chimeric *Propionibacterium acnes* DsA1 / DsA2 polypeptide consists of the amino acid sequence according to SEQ ID NO: 70, and the *Propionibacterium acnes* PITP polypeptide consists of the amino acid sequence according to SEQ ID NO: 73.

[0295] LNP In some embodiments, the compositions of the present invention (e.g., compositions comprising the nucleic acids of the present invention) further comprise lipid nanoparticles (LNPs). In some embodiments, the nucleic acids of the present invention are encapsulated in LNPs.

[0296] The LNP disclosed herein may include four types of lipids: (i) ionizable lipids (e.g., cationic lipids); (ii) PEGylated lipids; (iii) cholesterol-based lipids; and (iv) accessory lipids.

[0297] A. Ionizable lipids Ionizable lipids can promote mRNA encapsulation and can be cationic lipids. Cationic lipids provide a positively charged environment at low pH to promote the effective encapsulation of negatively charged mRNA drugs.

[0298] In some embodiments, the cationic lipid is OF-02:

[0299] Formula (I) OF-02 is a non-degradable structural analogue of OF-Deg-Lin. OF-Deg-Lin contains a degradable ester bond to connect the diketopiperazine core and the biunsaturated tail, while OF-02 contains a non-degradable 1,2-amino-ol bond to connect the same diketopiperazine core and biunsaturated tail (Fenton et al., Adv Mater. [Advanced Materials] (2016) 28:2939; U.S. Patent 10,201,618). The exemplary LNP formulation (lipid A) described herein contains OF-2.

[0300] In some embodiments, the cationic lipid is cKK-E10 (Dong et al., PNAS [Proceedings of the National Academy of Sciences] (2014) 111(11):3955-60; US Patent 9,512,073):

[0301] cKK-E10 Equation (II) The exemplary LNP formulation (lipid B) described herein contains cKK-E10.

[0302] In some embodiments, the cationic lipid is GL-HEPES-E3-E10-DS-3-E18-1(2-(4-(2-((3-(bis((Z)-2-hydroxyoctadecyl-9-en-1-yl)amino)propyl)dithioalkyl)ethyl)piperazin-1-yl)ethyl 4-(bis(2-hydroxydecyl)amino)butyrate), which is a HEPES-based disulfide cationic lipid with a piperazine core, having Formula III:

[0303] Equation (III) The exemplary LNP formulation (lipid C) described herein contains GL-HEPES-E3-E10-DS-3-E18-1. Lipid C has the same composition as lipid A or lipid B, except that it is a cationic lipid.

[0304] Typically, the cationic lipid is GL-HEPES-E3-E12-DS-4-E10(2-(4-(2-((3-(bis(2-hydroxydecyl)amino)butyl)dithioalkyl)ethyl)piperazin-1-yl)ethyl 4-(bis(2-hydroxydodecyl)amino)butyrate), which is a HEPES-based disulfide cationic lipid with a piperazine core, having Formula IV:

[0305] Formula (IV) The exemplary LNP formulation (lipid D) described herein contains GL-HEPES-E3-E12-DS-4-E10. Lipid D has the same composition as lipid A or lipid B, except that it is a different cationic lipid.

[0306] In some embodiments, the cationic lipid is GL-HEPES-E3-E12-DS-3-E14(2-(4-(2-((3-(bis(2-hydroxytetradecyl)amino)propyl)dithioalkyl)ethyl)piperazin-1-yl)ethyl 4-(bis(2-hydroxydodecyl)amino)butyrate), which is a HEPES-based disulfide cationic lipid with a piperazine core, having formula V:

[0307] Formula (V) The exemplary LNP formulation (lipid E) described herein contains GL-HEPES-E3-E12-DS-3-E14. Lipid E ​​has the same composition as lipid A or lipid B, except that it is a cationic lipid.

[0308] The cationic lipids GL-HEPES-E3-E10-DS-3-E18-1 (III), GL-HEPES-E3-E12-DS-4-E10 (IV), and GL-HEPES-E3-E12-DS-3-E14 (V) can be synthesized according to the general procedure listed in Scheme 1: Scheme 1: General synthetic schemes for lipids of formulas (III), (IV) and (V)

[0309] In some embodiments, the cationic lipid is MC3, which has formula VI:

[0310] Formula (VI) In some embodiments, the cationic lipid is SM-102 (9-heptadecyl 8-{(2-hydroxyethyl)[6-oxo-6-(undecyloxy)hexyl]amino}octanoate), which has formula VII:

[0311] Equation (VII) In some embodiments, the cationic lipid is ALC-0315 [(4-hydroxybutyl)azanidinediyl]bis(hexane-6,1-diyl)bis(2-hexyldecanoate), having formula VIII:

[0312] Formula (VIII) In some embodiments, the cationic lipid is cOrn-EE1, which has formula IX:

[0313] Formula (IX) In some embodiments, the cationic lipid may be selected from the group consisting of: cKK-E10; OF-02; [(6Z,9Z,28Z,31Z)-hexahepta-6,9,28,31-tetraen-19-yl] 4-(dimethylamino)butyrate (D-Lin-MC3-DMA); 2,2-dilinoleyl-4-dimethylaminoethyl-[1,3]-dioxolane (dLin-KC2-DMA); 1,2-dilinoleyloxy-N,N-dimethyl-3-aminopropane (dLin-DMA); 9-((4-(dimethylamino)butyryl)oxy)heptadecanedioic acid di((Z)-non-2-en-1-yl) ester (L319); 8-{(2-hydroxyethyl)[6-oxo-6-(undecyloxy)hexyl]amino}octanoic acid 9-heptadecanedioic acid (SM-102); [(4-hydroxybutyl)azonide di... [3-(dimethylamino)-2-[(Z)-octadecyl-9-enoyl]oxypropyl](Z)-octadecyl-9-enoyl ester (DODAP); 2,5-bis(3-aminopropylamino)-N-[2-[di(heptadecyl)amino]-2-oxoethyl]pentanamide (DOGS); [(3S,8S,9S,10R,13R,14S,17R)-10,13-dimethyl-17-[(2R)-6-methylheptane-2-yl]-2,3,4,7,8,9,11,12,14,1 5,16,17-Dodecylhydro-1H-cyclopentan[a]phenanthrene-3-yl]N-[2-(dimethylamino)ethyl]carbamate (DC-Chol); 3,3′,3″,3‴-(((methylazanediyl)bis(propane-3,1-diyl))bis(azanetriyl))tetra(8-methylnonyl)tetrapropionate (306Oi10); (2-(dioctylammonium)ethyl)decyl phosphate (9A1P9); 5,5-Di((Z)-heptadec-8-en-1-yl)-1-(3-(pyrrolidine-1-yl)propyl)-2,5-dihydro-1H-imidazol-2-carboxylate (A2-Iso5-2DC18) ); 3,3′-((3-methyl-9-oxo-10-oxa-13,14-dithia-3,6-diazahexadecyl)azanediyl)dipropionate bis(2-(dodecyldithioalkyl)ethyl) ester (BAME-O16B); 1,1′-((2-(4-(2-((2-(bis(2-hydroxydodecyl)amino)ethyl)(2-hydroxydodecyl)amino)ethyl)piperazine-1-yl)ethyl)azanediyl)bis(dodecane-2-ol) (C12-200); 3,6-bis(4-(bis(2-hydroxydodecyl)amino)butyl)piperazine-2,5-dione (cKK-E12);9,9′,9″,9‴,9″″,9‴″-((((benzene-1,3,5-tricarbonyl)tri(azanediyl))tri(propane-3,1-diyl))tri(azanetriyl))hexa(octyl-3-yl)hexyl nonanoate (FTT5); (((3,6-dioxopiperazine-2,5-diyl)bis(butane-4,1-diyl))bis(azanetriyl))tetra(ethane-2,1-diyl)(9Z,9′Z,9″Z,9‴Z,12Z,12′Z,12″Z,12‴Z)-tetra(octadec-9,12-dienoate) (OF-Deg-Lin); TT3; N1,N3,N5-tri(3-(dodecane) (aminopropyl)benzene-1,3,5-tricarboxamide; N1-[2-((1S)-1-[(3-aminopropyl)amino]-4-[di(3-aminopropyl)amino]butylcarbamoyl)ethyl]-3,4-di[oleylyloxy]-benzamide (MVL5); 8-((2-hydroxyethyl)(8-(nonoxy)-8-oxooctyl)amino)octanoic acid heptadecanyl-9-yl ester (lipid 5); GL-HEPES-E3-E10-DS-3-E18-1; GL-HEPES-E3-E12-DS-4-E10; GL-HEPES-E3-E12-DS-3-E14; and combinations thereof.

[0314] In some embodiments, the cationic lipid is IM-001, which has the formula X (EP23306049.0):

[0315] Formula (X) The exemplary LNP formulation (lipid F) described herein contains IM-001. Lipid F has the same composition as lipid A or lipid B, except that it is a cationic lipid.

[0316] The cationic lipid IM-001 (X) can be synthesized according to the general procedure described in Scheme 2: Scheme 2: General synthetic scheme for lipids of formula (X)

[0317] Option 2 can be carried out as described in Example 12.

[0318] In some embodiments, the cationic lipid is IS-001, having the formula XI (EP23306049.0):

[0319] Formula (XI) The exemplary LNP formulation (lipid G) described herein contains IS-001. Lipid G has the same composition as lipid A or lipid B, except that it is a cationic lipid.

[0320] The cationic lipid IS-001 (XI) can be synthesized according to the general procedure described in Scheme 3: Scheme 3: General synthetic scheme for lipids of formula (XI)

[0321] Option 3 can be carried out as described in Example 14.

[0322] In some embodiments, the cationic lipids are biodegradable.

[0323] In some embodiments, the cationic lipids are not biodegradable.

[0324] In some embodiments, the cationic lipid is cleavable.

[0325] In some embodiments, the cationic lipids are not cleavable.

[0326] Cationic lipids are described in further detail in Dong et al. (PNAS. 111(11):3955-60.2014); Fenton et al. (Adv. Mater. 28:2939.2016); U.S. Patent Nos. 9,512,073 and 10,201,618, which are each incorporated herein by reference.

[0327] B. PEGylated lipids PEGylated lipid components provide control over the particle size and stability of nanoparticles. The addition of such components can prevent complex aggregation and provides a means to increase cycle life and delivery of lipid-nucleic acid drug compositions to target tissues (Klibanov et al., FEBS Letters [European Federation of Biochemical Societies Letters] 268(1):235-7. 1990). These components can be selected for rapid exchange from the drug composition in vivo (see, for example, US Patent No. 5,885,613).

[0328] The PEGylated lipids under consideration include, but are not limited to, polyethylene glycol (PEG) chains up to 5 kDa in length, covalently attached to C6-C6 bonds. 20 (For example, C8, C) 10 C 12 C 14 C 16 Or C 18Lipids with one or more alkyl chains of length 1000, such as derivatized ceramides (e.g., N-octanoyl-sphingosine-1-[succinyl(methoxypolyethylene glycol)] (C8 PEG ceramide)). In some embodiments, the PEGylated lipid is 1,2-dimyristoyl-racemic-glycerol-3-methoxy polyethylene glycol (DMG-PEG); 1,2-distearyl-sn-glycerol-3-phosphoethanolamine-polyethylene glycol (DSPE-PEG); 1,2-dilauryl-sn-glycerol-3-phosphoethanolamine-polyethylene glycol (DLPE-PEG); or 1,2-distearyl-racemic-glycerol-polyethylene glycol (DSG-PEG); PEG-DAG; PEG-PE; PEG-S-DAG; PEG-S-DMG; PEG-cer; PEG-dialkoxypropylcarbamate; 2-[(polyethylene glycol)-2000]-N,N-tetracosylacetamide (ALC-0159); and combinations thereof.

[0329] In some embodiments, the PEG has a high molecular weight, for example, 2000-2400 g / mol. In some embodiments, the PEG is PEG2000 (or PEG-2K). In some embodiments, the PEGylated lipids described herein are DMG-PEG2000, DSPE-PEG2000, DLPE-PEG2000, DSG-PEG2000, C8 PEG2000, or ALC-0159 (2-[(polyethylene glycol)-2000]-N,N-bistetradecylacetamide). In some embodiments, the PEGylated lipids described herein are DMG-PEG2000.

[0330] C. Cholesterol-based lipids The cholesterol component provides stability to the lipid bilayer structure within the nanoparticles. In some embodiments, the LNP comprises one or more cholesterol-based lipids. Suitable cholesterol-based lipids include, for example: DC-Choi (N,N-dimethyl-N-ethylformamidocholesterol), 1,4-bis(3-N-oleenylaminopropyl)piperazine (Gao et al., BiochemBiophys Res Comm. (1991) 179:280; Wolf et al., BioTechniques (1997) 23:139; U.S. Patent 5,744,335), and imidazole cholesterol ester (“ICE”; WO 2011 / 068810), sitosterol (22,23-dihydrostigmasterol), β-stigmasterol, sitosterol, fucosterol, stigmasterol (stigmasterane-5,22-dien-3-ol), ergosterol; cyclosporine (3β-hydroxy-5,24-cholestadiene); lanosterol (8,24-lanosterdien-3β-ol); 7-dehydrocholesterol (Δ5,7-cholesterol); dihydrolanosterol (24,25-dihydrolanosterol); yeast sterol (5α-cholest-8,24-dien-3β-ol) ); cholesterol (5α-cholesterol-7-en-3β-ol); diosgenin ((3β,25R)-spirost-5-en-3-ol); campesterol (campesterol-5-en-3β-ol); campesteranol (5a-campesteranol-3b-ol); 24-methylene cholesterol (5,24(28)-cholestadien-24-methylene-3β-ol); cholesterol heptadecanate (cholesterol-5-en-3β-yl heptadecanate); cholesterol oleate; cholesterol stearate and other modified forms of cholesterol. In some embodiments, the cholesterol-based lipids used in the LNP are cholesterol.

[0331] D. Helper lipids The assisting lipid enhances the structural stability of the LNP and facilitates its escape from the endosome. It improves the uptake and release of the mRNA drug payload. In some embodiments, the assisting lipid is a zwitterionic lipid with fusion-promoting properties for enhancing the uptake and release of the drug payload. Examples of accessory lipids are 1,2-dioleoyl-SN-glycerol-3-phosphate ethanolamine (DOPE); 1,2-distearyl-sn-glycerol-3-phosphocholine (DSPC); 1,2-dioleoyl-sn-glycerol-3-phosphate-L-serine (DOPS); 1,2-ditransoleoyl-sn-glycerol-3-phosphate ethanolamine (DEPE); and 1,2-dioleoyl-sn-glycerol-3-phosphocholine (DPOC), dipalmitoylphosphatidylcholine (DPPC), DMPC, 1,2-dilauroyl-sn-glycerol-3-phosphocholine (DLPC), 1,2-distearylphosphatidylethanolamine (DSPE) and 1,2-dilauroyl-sn-glycerol-3-phosphocholine (DLPE).

[0332] Other exemplary cofactor lipids are dioleoylphosphatidylcholine (DOPC), dioleoylphosphatidylglycerol (DOPG), dipalmitoylphosphatidylglycerol (DPPG), palmitoyloleoylphosphatidylcholine (POPC), palmitoyloleoylphosphatidylethanolamine (POPE), dioleoylphosphatidylethanolamine 4-(N-maleiminomethyl)-cyclohexane-1-carboxylate (DOPE-mal), dipalmitoylphosphatidylethanolamine (DPPE), dimyristoylphosphatidylethanolamine (DMPE), phosphatidylserine, sphingolipids, sphingomyelin, ceramides, cerebrosides, gangliosides, 16-O-monomethylPE, 16-O-dimethylPE, 18-1-transPE, 1-stearoyl-2-oleoylphosphatidylethanolamine (SOPE), or combinations thereof. In some embodiments, the cofactor lipid is DOPE. In some embodiments, the cofactor lipid is DSPC.

[0333] In various embodiments, the LNP of the present invention comprises (i) a cationic lipid selected from the following: OF-02, cKK-E10, GL-HEPES-E3-E10-DS-3-E18-1, GL-HEPES-E3-E12-DS-4-E10, GL-HEPES-E3-E12-DS-3-E14, IM-001 or IS-001; (ii) DMG-PEG2000; (iii) cholesterol; and (iv) DOPE.

[0334] In other embodiments, the LNP of the present invention comprises (i) SM-102; (ii) DMG-PEG2000; (iii) cholesterol; and (iv) DSPC.

[0335] In some other embodiments, the LNP of the present invention comprises (i) ALC-0315; (ii) ALC-0159; (iii) cholesterol; and (iv) DSPC.

[0336] E. Molar ratio of lipid components The molar ratios of the aforementioned components are important for the effectiveness of LNP in delivering mRNA. The molar ratio of cationic lipids, PEGylated lipids, cholesterol-based lipids, and cofactor lipids is A:B:C:D, where A + B + C + D = 100%. In some embodiments, the molar ratio of cationic lipids to total lipids (i.e., A) in the LNP is 35%-55%, such as 35%-50% (e.g., 38%-42%, such as 40% or 45%-50%). In some embodiments, the molar ratio of the PEGylated lipid component to total lipids (i.e., B) is 0.25%-2.75% (e.g., 1%-2%, such as 1.5%). In some embodiments, the molar ratio of cholesterol-based lipids to total lipids (i.e., C) is 20%-50% (e.g., 27%-30%, such as 28.5% or 38%-43%). In some embodiments, the molar ratio of the auxiliary lipids to the total lipids (i.e., D) is 5%-35% (e.g., 28%-32%, such as 30% or 8%-12%, such as 10%). In some embodiments, the (PEGylated lipids + cholesterol) component has the same molar amount as the auxiliary lipids. In some embodiments, the molar ratio of cationic lipids to auxiliary lipids contained in the LNP is greater than 1.

[0337] In some embodiments, the LNP disclosed herein includes: Cationic lipids with a molar ratio of 35% to 55% or 40% to 50% (e.g., cationic lipids with molar ratios of 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, 50%, 51%, 52%, 53%, 54% or 55%). A polyethylene glycol (PEG) conjugated (PEGylated) lipid in a molar ratio of 0.25% to 2.75% or 1.00% to 2.00% (e.g., PEGylated lipids in molar ratios of 0.25%, 0.50%, 0.75%, 1.00%, 1.25%, 1.50%, 1.75%, 2.00%, 2.25%, 2.50%, or 2.75%). Cholesterol-based lipids with molar ratios of 20% to 50%, 25% to 45%, or 28.5% to 43% (e.g., cholesterol-based lipids with molar ratios of 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, 35%, 36%, 37%, 38%, 39%, 40%, 41%, 42%, 43%, 44%, 45%, 46%, 47%, 48%, 49%, or 50%); and The auxiliary lipids are in the molar ratio of 5% to 35%, 8% to 30%, or 10% to 30% (e.g., auxiliary lipids in the molar ratio of 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, 16%, 17%, 18%, 19%, 20%, 21%, 22%, 23%, 24%, 25%, 26%, 27%, 28%, 29%, 30%, 31%, 32%, 33%, 34%, or 35%). All molar ratios are relative to the total lipid content of the LNP.

[0338] In some embodiments, the LNP comprises: 40% cationic lipids; 1.5% PEGylated lipids; 28.5% cholesterol-based lipids; and 30% auxiliary lipids.

[0339] In some embodiments, the LNP disclosed herein comprises: cationic lipids in a molar ratio of 45% to 50%; PEGylated lipids in a molar ratio of 1.5% to 1.7%; cholesterol-based lipids in a molar ratio of 38% to 43%; and auxiliary lipids in a molar ratio of 9% to 10%.

[0340] In some embodiments, the PEGylated lipid is dimyristic PEG2000 (DMG-PEG2000).

[0341] In various embodiments, the cholesterol-based lipid is cholesterol.

[0342] In some embodiments, the auxiliary lipid is 1,2-dioleoyl-SN-glycerol-3-phosphoethanolamine (DOPE).

[0343] In some embodiments, the LNP comprises: OF-02 in a molar ratio of 35% to 55%; DMG-PEG2000 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DOPE in a molar ratio of 5% to 35%.

[0344] In some embodiments, the LNP comprises: cKK-E10 in a molar ratio of 35% to 55%; DMG-PEG2000 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DOPE in a molar ratio of 5% to 35%.

[0345] In some embodiments, the LNP comprises: GL-HEPES-E3-E10-DS-3-E18-1 in a molar ratio of 35% to 55%; DMG-PEG2000 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DOPE in a molar ratio of 5% to 35%.

[0346] In some embodiments, the LNP comprises: GL-HEPES-E3-E12-DS-4-E10 in a molar ratio of 35% to 55%; DMG-PEG2000 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DOPE in a molar ratio of 5% to 35%.

[0347] In some embodiments, the LNP comprises: GL-HEPES-E3-E12-DS-3-E14 in a molar ratio of 35% to 55%; DMG-PEG2000 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DOPE in a molar ratio of 5% to 35%.

[0348] In some embodiments, the LNP comprises: SM-102 in a molar ratio of 35% to 55%; DMG-PEG2000 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DSPC in a molar ratio of 5% to 35%.

[0349] In some embodiments, the LNP comprises: ALC-0315 in a molar ratio of 35% to 55%; ALC-0159 in a molar ratio of 0.25% to 2.75%; cholesterol in a molar ratio of 20% to 50%; and DSPC in a molar ratio of 5% to 35%.

[0350] In some embodiments, the LNP comprises: 40% OF-02 in a molar ratio; 1.5% DMG-PEG2000 in a molar ratio; 28.5% cholesterol in a molar ratio; and 30% DOPE in a molar ratio. This LNP formulation is designated herein as "Lipid A".

[0351] In some embodiments, the LNP comprises: 40% cKK-E10 in molar ratio; 1.5% DMG-PEG2000 in molar ratio; 28.5% cholesterol in molar ratio; and 30% DOPE in molar ratio. This LNP formulation is designated herein as "Lipid B".

[0352] In some embodiments, the LNP comprises: 40% GL-HEPES-E3-E10-DS-3-E18-1 in a molar ratio; 1.5% DMG-PEG2000 in a molar ratio; 28.5% cholesterol in a molar ratio; and 30% DOPE in a molar ratio. This LNP formulation is designated herein as "Lipid C".

[0353] Typically, an LNP contains: 40% GL-HEPES-E3-E12-DS-4-E10 in molar ratio; 1.5% DMG-PEG2000 in molar ratio; 28.5% cholesterol in molar ratio; and 30% DOPE in molar ratio. This LNP formulation is designated as "Lipid D" herein.

[0354] In some embodiments, the LNP comprises: 40% GL-HEPES-E3-E12-DS-3-E14 in a molar ratio; 1.5% DMG-PEG2000 in a molar ratio; 28.5% cholesterol in a molar ratio; and 30% DOPE in a molar ratio. This LNP formulation is designated herein as "Lipid E".

[0355] In some embodiments, the LNP comprises: 40% IM-001; 1.5% DMG-PEG2000; 28.5% cholesterol; and 30% DOPE. This LNP formulation is designated herein as "Lipid F".

[0356] In some embodiments, the LNP comprises: 40% IS-001; 1.5% DMG-PEG2000; 28.5% cholesterol; and 30% DOPE. This LNP formulation is designated herein as "Lipid G".

[0357] In some embodiments, the LNP comprises: 50% of 9-heptadecyl 8-{(2-hydroxyethyl)[6-oxo-6-(undecyloxy)hexyl]amino}octanoate (SM-102); 10% of 1,2-distearyl-sn-glycerol-3-phosphocholine (DSPC); 38.5% of cholesterol; and 1.5% of 1,2-dimyristoyl-racemic-glycerol-3-methoxy polyethylene glycol-2000 (DMG-PEG2000).

[0358] In some embodiments, the LNP comprises: 46.3% (4-hydroxybutyl)azanidinediyl]bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (ALC-0315); 9.4% 1,2-distearyl-sn-glycerol-3-phosphocholine (DSPC); 42.7% cholesterol; and 1.6% 2-[(polyethylene glycol)-2000]-N,N-bistetradecylacetamide (ALC-0159).

[0359] In some embodiments, the LNP comprises: 47.4% molar of [(4-hydroxybutyl)azanidinediyl]bis(hexane-6,1-diyl)bis(2-hexyldecanoate) (ALC-0315); 10% molar of 1,2-distearyl-sn-glycerol-3-phosphocholine (DSPC); 40.9% molar of cholesterol; and 1.7% molar of 2-[(polyethylene glycol)-2000]-N,N-bistetradecylacetamide (ALC-0159).

[0360] To calculate the actual amount of each lipid to be incorporated into the LNP formulation, the molar amount of the cationic lipid is first determined based on the desired N / P ratio, where N is the number of nitrogen atoms in the cationic lipid and P is the number of phosphate groups in the mRNA to be transported by LNP. Next, the molar amount of each other lipid is calculated based on the molar amount of the cationic lipid and the selected molar ratio. These molar amounts are then converted to weight using the molecular weight of each lipid.

[0361] F. Nucleic acid in LNP The LNP compositions described herein may contain the nucleic acids (e.g., mRNA) of the present invention.

[0362] If desired, the LNP can be multivalent. In some embodiments, the LNP may carry nucleic acids, such as mRNA, encoding more than one polypeptide (antigen), for example two, three, four, five, or six polypeptides (antigens). For example, the LNP may carry multiple nucleic acids (e.g., mRNAs) of the present invention, each encoding a different polypeptide described herein; or carry polycistronic mRNAs that can be translated into more than one polypeptide described herein (e.g., each antigen-coding sequence is separated by a nucleotide linker encoding a self-cleaving peptide such as a 2A peptide). An LNP carrying different nucleic acids (e.g., mRNA) typically contains (encapsulates) multiple copies of each nucleic acid. For example, an LNP carrying or encapsulating two different nucleic acids typically carries multiple copies of each of the two different nucleic acids.

[0363] Typically, two or more nucleic acids (e.g., two or three mRNAs) encoding different polypeptides as described herein are co-encapsulated in the same LNP. For example, the LNP described herein may co-encapsulate a nucleic acid (e.g., mRNA) encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide and a nucleic acid (e.g., mRNA) encoding a *Dermatitis acnes* PITP polypeptide. The LNP described herein may co-encapsulate a nucleic acid (e.g., mRNA) encoding a chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide and a nucleic acid (e.g., mRNA) encoding a modified *Dermatitis acnes* CAMP2 polypeptide (or *Dermatitis acnes* CAMP2 polypeptide). The LNP described herein may co-encapsulate a nucleic acid (e.g., mRNA) encoding a *Dermatitis acnes* PITP polypeptide and a nucleic acid (e.g., mRNA) encoding a modified *Dermatitis acnes* CAMP2 polypeptide (or *Dermatitis acnes* CAMP2 polypeptide). Any two nucleic acids (e.g., two mRNAs) described herein may be present in a 1:1 weight ratio.

[0364] The LNP described herein can co-encapsulate (i) nucleic acids (e.g., mRNA) encoding the chimeric *Dermatitis acnes* DsA1 / DsA2 polypeptide described herein, (ii) nucleic acids (e.g., mRNA) encoding the *Dermatitis acnes* PITP polypeptide described herein, and (iii) nucleic acids (e.g., mRNA) encoding the modified *Dermatitis acnes* CAMP2 polypeptide (or *Dermatitis acnes* CAMP2 polypeptide) described herein. The three nucleic acids (e.g., three mRNAs) described herein may be present in a 1:1:1 weight ratio.

[0365] LNPs are as described in this article (e.g., lipid D).

[0366] Alternatively, any two or more (e.g., three) nucleic acids (e.g., mRNA) encoding different polypeptides as described herein may be encapsulated in a single LNP. In some embodiments, a single LNP formulation may contain multiple (e.g., two, three, four, five, six, seven, eight, nine, ten or more) LNPs, each carrying a different nucleic acid (e.g., mRNA).

[0367] When the nucleic acid is mRNA, the mRNA can be unmodified (i.e., containing only natural ribonucleotides A, U, C, and / or G linked by phosphodiester bonds) or chemically modified (e.g., including nucleotide analogs such as pseudouridine (e.g., N-1-methylpseuuridine), 2'-fluororibonucleotides and 2'-methoxyribonucleotides and / or thiophosphate bonds). The mRNA molecule may contain a 5' cap and a poly-A tail.

[0368] G. Buffer and other components To stabilize nucleic acids and / or LNPs (e.g., to extend the shelf life of vaccine products), facilitate the administration of LNP pharmaceutical compositions, and / or enhance in vivo expression of nucleic acids, nucleic acids and / or LNPs can be formulated in combination with one or more carriers, targeting ligands, stabilizing agents (e.g., preservatives and antioxidants), and / or other pharmaceutically acceptable excipients. Examples of such excipients include parabens, thimerosal, sodium thiomersal, chlorobutanol, benzalkonium chloride, and chelating agents (e.g., EDTA).

[0369] The LNP compositions disclosed herein can be provided in either a cryo-liquid or lyophilized form. Various cryoprotectants can be used, including, but not limited to, sucrose, trehalose, glucose, mannitol, mannose, dextrose, etc. The cryoprotectant may comprise 5%-30% (w / v) of the LNP composition. In some embodiments, the LNP composition contains trehalose, for example, 5%-30% (e.g., 10%) (w / v). Once formulated with a cryoprotectant, the LNP composition can be frozen (or lyophilized and cryopreserved) at -20°C to -80°C.

[0370] The LNP composition can be provided to the patient in an aqueous buffer solution: if previously frozen, it is thawed, or if previously lyophilized, it is reconstituted at the bedside in an aqueous buffer solution. The buffer solution is preferably isotonic and suitable for, for example, intramuscular or intradermal injection. In some embodiments, the buffer solution is phosphate-buffered saline (PBS).

[0371] Nucleic acid The nucleic acid of the present invention can be RNA or DNA. The nucleic acid of the present invention can be single-stranded or double-stranded. In some embodiments, the nucleic acid is RNA, for example, mRNA.

[0372] mRNA In some embodiments, the nucleic acid of the present invention is messenger RNA (mRNA). The mRNA may be modified or unmodified. The mRNA may contain one or more coding and non-coding regions. The coding region may alternatively be referred to as an open reading frame (ORF). The non-coding region in the mRNA includes a 5' cap, a 5' untranslated region (UTR), a 3' UTR, and a poly-A tail. The mRNA may be purified from natural sources, generated and optionally purified using a recombinant expression system (e.g., in vitro transcription), or chemically synthesized.

[0373] In some embodiments, the mRNA contains an ORF encoding a target antigen. In some embodiments, the RNA (e.g., mRNA) further includes at least one 5' UTR, a 3' UTR, a poly(A) tail, and / or a 5' cap. In some embodiments, the mRNA contains (i) a 5' cap as defined herein; (ii) a 5' untranslated region (UTR) as defined herein; (iii) a protein-coding region; (iv) a 3' UTR as defined herein; and (v) a poly(A) tail. Typically, the 3' end of (i) is directly bonded to the 5' end of (ii) via a 3' to 5' phosphodiester linker; the 3' end of (ii) is directly bonded to the 5' end of (iii) via a 3' to 5' phosphodiester linker; the 3' end of (iii) is directly bonded to the 5' end of (iv) via a 3' to 5' phosphodiester linker; and the 3' end of (iv) is directly bonded to the 5' end of (v) via a 3' to 5' phosphodiester linker.

[0374] In some embodiments, the mRNA contains at least one, at least two, at least three, or more stop codons, wherein the one or more stop codons are selected from UAA, UGA, and UAG, and wherein the at least two, at least three, or more stop codons may be the same or different. Typically, the at least one stop codon includes UAA or UGA (e.g., UAA). Typically, the at least two stop codons include at least two identical stop codons, such as UAA or UGA (e.g., UAAUAA or UGAUGA) or at least two different stop codons, which may be particularly selected from UAA and UGA (e.g., UGAUAA). Typically, the at least three stop codons include UAA, UGA, and UAG (e.g., UGAUAAUAG).

[0375] Unless otherwise indicated, the mRNA sequence is presented in the 5' to 3' orientation.

[0376] 5' cap The 5' cap on mRNA can provide resistance to nucleases found in most eukaryotic cells and promote translation efficiency. Several types of 5' caps are known. The 7-methylguanosine cap (also known as "m7G" or "Cap-0") contains a guanosine nucleotide linked to the first transcribed nucleotide via a 5'–5'-triphosphate bond.

[0377] Typically, a 5' cap is added as follows: First, an RNA terminal phosphatase removes one terminal phosphate group from the 5' nucleotide, leaving two terminal phosphate groups; then, guanosine triphosphate (GTP) is added to the terminal phosphate groups via guanylate transferase, creating a 5'5'5 triphosphate bond; then, the 7-nitrogen of guanine is methylated via methyltransferase. Examples of cap structures include, but are not limited to, m7G(5')ppp, (5'(A,G(5')ppp(5')A, and G(5')ppp(5')G. Other cap structures are described in U.S. Publication Nos. US 2016 / 0032356 and US 2018 / 0125989, which are incorporated herein by reference.

[0378] According to the manufacturer’s protocol, the following chemical RNA cap analogues can be used to simultaneously cap polynucleotides during in vitro transcription to generate 5'-guanosine cap structures: 3'-O-Me-m7G(5')ppp(5')G (ARCA cap); G(5')ppp(5')A; G(5')ppp(5')G; m7G(5')ppp(5')A; m7G(5')ppp(5')G; m7G(5')ppp(5'))(2'oMeA)pG; m7G(5')ppp(5')(2'oMeA)pU; m7G(5')ppp(5')(2'oMeG)pG (New England BioLabs, Ipswich, MA; TriLink Biotechnologies). 5'-Capping of modified RNA can be performed post-transcriptionally using a vaccinia virus capping enzyme to produce a cap 0 structure: m7G(5')ppp(5')G. A cap 1 structure can be produced using both a vaccinia virus capping enzyme and a 2'-O methyltransferase, resulting in: m7G(5')ppp(5')G-2'-O-methyl. A cap 2 structure can be generated from the cap 1 structure, followed by 2'-O-methylation of the penultimate nucleotide at the 5' end using a 2'-O methyltransferase. A cap 3 structure can be generated from the cap 2 structure, followed by 2'-O-methylation of the penultimate nucleotide at the 5' end using a 2'-O methyltransferase.

[0379] In some embodiments, the mRNA disclosed herein includes a 5' cap selected from the group consisting of: 3'-O-Me-m7G(5')ppp(5')G (ARCA cap), G(5')ppp(5')A, G(5')ppp(5')G, m7G(5')ppp(5')A, m7G(5')ppp(5')G, m7G(5')ppp(5')(2'OMeA)pG, m7G(5')ppp(5')(2'OMeA)pU, and m7G(5')ppp(5')(2'OMeG)pG.

[0380] In some embodiments, the mRNA disclosed herein includes a 5' cap:

[0381] Untranslated Region (UTR) In some embodiments, the mRNA of the present invention comprises a 5' and / or a 3' untranslated region (UTR). In the mRNA, the 5' UTR begins at the transcription start site and continues to the start codon, but does not include the start codon. The 3' UTR begins immediately after the stop codon and continues until the transcription termination signal.

[0382] In some embodiments, the mRNA disclosed herein may include a 5' UTR, which includes one or more elements that affect the stability or translation of the mRNA. In some embodiments, the 5' UTR may have a length of about 10 to 5,000 nucleotides. In some embodiments, the 5' UTR may have a length of about 50 to 500 nucleotides. In some embodiments, the 5' UTR has a length of at least about 10 nucleotides, about 20 nucleotides, about 30 nucleotides, about 40 nucleotides, about 50 nucleotides, about 100 nucleotides, about 150 nucleotides, about 200 nucleotides, about 250 nucleotides, about 300 nucleotides, about 350 nucleotides, about 400 nucleotides, about 450 nucleotides, about 500 nucleotides, about 550 nucleotides, about 600 nucleotides, or about 650 nucleotides. The lengths are approximately 700 nucleotides, 750 nucleotides, 800 nucleotides, 850 nucleotides, 900 nucleotides, 950 nucleotides, 1,000 nucleotides, 1,500 nucleotides, 2,000 nucleotides, 2,500 nucleotides, 3,000 nucleotides, 3,500 nucleotides, 4,000 nucleotides, 4,500 nucleotides, or 5,000 nucleotides.

[0383] In some embodiments, the mRNA disclosed herein may include a 3' UTR, which includes one or more of the following: a polyadenylation signal, a protein binding site affecting the positional stability of the mRNA in the cell, or one or more binding sites of the miRNA. In some embodiments, the 3' UTR may have a length of 50 to 5,000 nucleotides or longer. In some embodiments, the 3' UTR may have a length of 50 to 1,000 nucleotides or longer. In some embodiments, the 3' UTRs have lengths of at least about 50 nucleotides, about 100 nucleotides, about 150 nucleotides, about 200 nucleotides, about 250 nucleotides, about 300 nucleotides, about 350 nucleotides, about 400 nucleotides, about 450 nucleotides, about 500 nucleotides, about 550 nucleotides, about 600 nucleotides, about 650 nucleotides, about 700 nucleotides, about 750 nucleotides, about 800 nucleotides, about 850 nucleotides, about 900 nucleotides, about 950 nucleotides, about 1,000 nucleotides, about 1,500 nucleotides, about 2,000 nucleotides, about 2,500 nucleotides, about 3,000 nucleotides, about 3,500 nucleotides, about 4,000 nucleotides, about 4,500 nucleotides, or about 5,000 nucleotides.

[0384] In some embodiments, the mRNA disclosed herein may include a 5' or 3' UTR derived from a gene different from the gene encoded by the mRNA transcript (i.e., the UTR is a heterologous UTR).

[0385] In some embodiments, the 5' and / or 3' UTR sequences may be derived from stable mRNAs (e.g., globin, actin, GAPDH, tubulin, histones, or citrate cycling enzymes) to increase mRNA stability. For example, the 5' UTR sequence may include a portion of the CMV, i.e., the early 1 (IE1) gene or a fragment thereof, to improve the mRNA's nuclease resistance and / or improve its half-life. Including a sequence encoding human growth hormone (hGH) or a fragment thereof at the 3' end or untranslated region of the mRNA is also considered. Generally, these modifications improve the stability and / or pharmacokinetic properties (e.g., half-life) of the mRNA relative to its unmodified counterpart, and include modifications, for example, to improve the resistance of such mRNAs to in vivo nuclease digestion.

[0386] The exemplary 5' UTR contains a sequence derived from the CMV, namely the early 1 (IE1) gene (US Publication Nos. 2014 / 0206753 and 2015 / 0157565, each of which is incorporated herein by reference) or the sequence GGGAUCCUACC (SEQ ID NO: 264) (US Publication No. 2016 / 0151409, which is incorporated herein by reference).

[0387] In various embodiments, the 5' UTR may be derived from the 5' UTR of a TOP gene. TOP genes are typically characterized by the presence of a 5'-terminal oligopyrimidine (TOP) bundle. Furthermore, most TOP genes are characterized by growth-related translational regulation. However, TOP genes with tissue-specific translational regulation are also known. In some embodiments, the 5' UTR derived from the 5' UTR of a TOP gene lacks the 5' TOP motif (oligopyrimidine bundle) (e.g., U.S. Publications 2017 / 0029847, 2016 / 0304883, 2016 / 0235864, and 2016 / 0166710, each of which is incorporated herein by reference).

[0388] In some embodiments, 5' UTR is derived from the ribosomal protein large 32 (L32) gene (US Publication No. 2017 / 0029847, ibid.).

[0389] In some embodiments, the 5' UTR is derived from the 5' UTR of the hydroxysteroid (17-b) dehydrogenase 4 gene (HSD17B4) (US Publication No. 2016 / 0166710, ibid.).

[0390] In some embodiments, the 5' UTR is derived from the 5' UTR of the ATP5A1 gene (US Publication No. 2016 / 0166710, ibid.).

[0391] In some embodiments, the internal ribosome entry site (IRES) is used instead of the 5' UTR.

[0392] In some embodiments, the 5' UTR contains the nucleic acid sequence shown in SEQ ID NO: 265, and is reproduced as follows: GGACAGAUCGCCUGGAGACGCCAUCCACGCUGUUUUGACCUCCAUAGAAGACACCGGGACCGAUCCAGCCUCCGCGGCCGGGAACGGUGCAUUGGAACGCGGAUUCCCCGUGCCAAGAGUGACUCACCGUCCUUGACACG (SEQ ID NO: 265).

[0393] In some embodiments, the 3' UTR contains the nucleic acid sequence shown in SEQ ID NO: 266, and is reproduced as follows: CGGGUGGCAUCCCUGUGACCCCUCCCCAGUGCCUCCUGGCCCUGGAAGUUGCCACACUCCAGUGCCCACCAGCCUUGUCCUAAUAAAAUUAAGUUGCAUC (SEQ ID NO: 266).

[0394] The 5' UTR and 3' UTR are described in further detail in WO 2012 / 075040 (incorporated hereby by reference).

[0395] polyadenylation tail As used herein, the terms “multiple (A) sequence,” “multiple (A) tail,” and “multiple (A) region” refer to the adenosine nucleotide sequence at the 3' end of an mRNA molecule. The multiple (A) tail can confer stability to mRNA and protect it from exonuclease degradation. The multiple (A) tail can enhance translation. In some embodiments, the multiple (A) tail is substantially homopolymeric. For example, a 100-adenosine nucleotide multiple (A) tail can substantially have a length of 100 nucleotides. In some embodiments, the multiple (A) tail can be interrupted by at least one nucleotide different from the adenosine nucleotide (e.g., a nucleotide that is not adenosine nucleotide). For example, a 100-adenosine nucleotide multiple (A) tail can have a length exceeding 100 nucleotides (including 100 adenosine nucleotides and at least one nucleotide different from the adenosine nucleotide or a segment of nucleotides). In some embodiments, the poly(A) tail contains the sequence AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAGCAU AUGACUAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA (SEQ ID NO: 267).

[0396] As used herein, “multiple (A) tails” typically refers to RNA. However, in the context of this disclosure, the term also refers to the corresponding sequence in a DNA molecule (e.g., “multiple (T) sequences”).

[0397] The poly(A) tail may comprise about 10 to about 500 adenosine nucleotides, about 10 to about 200 adenosine nucleotides, about 40 to about 200 adenosine nucleotides, or about 40 to about 150 adenosine nucleotides. The length of the poly(A) tail may be at least about 10, 50, 75, 100, 150, 200, 250, 300, 350, 400, 450, or 500 adenosine nucleotides. In some embodiments, the poly(A) tail comprises at least 75 nucleotides (e.g., about 80 adenosine nucleotides). In some embodiments, the poly(A) tail comprises at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides). Typically, the poly(A) tail comprises at least 100 adenosine nucleotides.

[0398] In some embodiments where the nucleic acid is RNA, the poly(A) tail is obtained from a DNA template during in vitro transcription of RNA. In other embodiments, the poly(A) tail is obtained in vitro using common chemical synthesis methods, rather than transcribed from a DNA template. In various embodiments, the poly(A) tail is generated by enzymatic polyadenylation of RNA (after in vitro transcription of RNA) using commercially available polyadenylation kits and corresponding protocols, or alternatively by using immobilized poly(A) polymerases (e.g., using the methods and means described in WO 2016 / 174271).

[0399] Nucleic acids can include poly(A) tails obtained by enzymatic polyadenylation, wherein most nucleic acid molecules comprise about 100 (+ / -20) to about 500 (+ / -50) or about 250 (+ / -20) adenosine nucleotides.

[0400] In some embodiments, the nucleic acid may include a multi(A) tail derived from template DNA, and may additionally include at least one additional multi(A) tail generated by enzymatic polyadenylation, for example as described in WO 2016 / 091391.

[0401] In some embodiments, the nucleic acid includes at least one polyadenylation signal.

[0402] In various embodiments, the nucleic acid may include at least one multiple (C) sequence.

[0403] As used herein, the term "multiple (C) sequence" is intended to be a cytosine nucleotide sequence of up to about 200 cytosine nucleotides. In some embodiments, a multiple (C) sequence comprises about 10 to about 200 cytosine nucleotides, about 10 to about 100 cytosine nucleotides, about 20 to about 70 cytosine nucleotides, about 20 to about 60 cytosine nucleotides, or about 10 to about 40 cytosine nucleotides. In some embodiments, a multiple (C) sequence comprises about 30 cytosine nucleotides.

[0404] Chemical modification The mRNA disclosed herein may be modified or unmodified. In some embodiments, the mRNA may include at least one chemical modification. In some embodiments, the mRNA disclosed herein may contain one or more modifications that typically enhance RNA stability. Exemplary modifications may include backbone modifications, sugar modifications, or base modifications. In some embodiments, the disclosed mRNA may be synthesized from naturally occurring nucleotides and / or nucleotide analogs (modified nucleotides) (including, but not limited to, purines (adenine (A) and guanine (G))) or pyrimidines (thymine (T), cytosine (C), and uracil (U))). In some embodiments, the disclosed mRNA can be synthesized from modified nucleotide analogs or derivatives of purines and pyrimidines, such analogs or derivatives as 1-methyl-adenine, 2-methyl-adenine, 2-methylthio-N-6-isopentenyl-adenine, N6-methyl-adenine, N6-isopentenyl-adenine, 2-thio-cytosine, 3-methyl-cytosine, 4-acetyl-cytosine, 5-methyl-cytosine, 2,6-diaminopurine, 1-methyl-guanine, 2-methyl-guanine, 2,2-dimethyl-guanine, 7-methyl-guanine, inosine, 1-methyl-inosine, pseudouracil (5-uracil), dihydrouracil, 2-thio-uracil, 4-thio-uracil, 5-carboxyl-... Methylaminomethyl-2-thiouracil, 5-(carboxyhydroxymethyl)-uracil, 5-fluorouracil, 5-bromouracil, 5-carboxymethylaminomethyl-uracil, 5-methyl-2-thiouracil, 5-methyluracil, N-uracil-5-oxyacetic acid methyl ester, 5-methylaminomethyl-uracil, 5-methoxyaminomethyl-2-thiouracil, 5'-methoxycarbonylmethyl-uracil, 5-methoxyuracil, uracil-5-oxyacetic acid methyl ester, uracil-5-oxyacetic acid (v), 1-methyl-pseudouracil, piracetamidine, β-D-mannosyl-piracetamidine, aminophosphate, thiophosphate, peptide nucleotide, methylphosphonate, 7-deazoguanosine, 5-methylcytosine, and inosine.

[0405] In some embodiments, the disclosed mRNA may include at least one chemical modification, including but not limited to pseudouridine, N1-methylpseudouridine, 2-thiouridine, 4'-thiouridine, 5-methylcytosine, 2-thio-1-methyl-1-deazo-pseudouridine, 2-thio-1-methyl-pseudouridine, 2-thio-5-aza-uridine, 2-thio-dihydropseudouridine, 2-thio-dihydrouridine, 2-thio-pseudouridine, 4-methoxy-2-thio-pseudouridine, 4-methoxy-pseudouridine, 4-thio-1-methyl-pseudouridine, 4-thio-pseudouridine, 5-aza-uridine, dihydropseudouridine, 5-methyluridine, 5-methyluridine, 5-methoxyuridine, and 2'-O-methyluridine.

[0406] In some embodiments, the chemical modification is selected from the group consisting of: pseudouridine, N1-methylpseudouridine, 5-methylcytosine, 5-methoxyuridine, and combinations thereof.

[0407] In some embodiments, the chemical modification includes N1-methylpseuuridine. Typically, the chemical modification includes replacing each uridine with N1-methylpseuuridine, i.e., 100% of the U residues are N1-methylpseuuridine.

[0408] In some embodiments, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% of the uracil nucleotides in the mRNA are chemically modified.

[0409] In some embodiments, at least 20%, at least 30%, at least 40%, at least 50%, at least 60%, at least 70%, at least 80%, at least 85%, at least 90%, at least 95%, or 100% of the uracil nucleotides in the ORF are chemically modified.

[0410] Preparation of such analogues is described in, for example, U.S. Patent Nos. 4,373,071, 4,401,796, 4,415,732, 4,458,066, 4,500,707, 4,668,777, 4,973,679, 5,047,524, 5,132,418, 5,153,319, 5,262,530, and 5,700,642.

[0411] mRNA synthesis The mRNA disclosed herein can be synthesized according to any of a variety of methods. For example, the mRNA disclosed herein can be synthesized via in vitro transcription (IVT). Some methods for in vitro transcription are described, for example, in Geall et al. (2013) Semin. Immunol. [Journal of Immunology Symposium] 25(2): 152-159; Brunelle et al. (2013) Methods Enzymol. [Enzymological Methods] 530:101-14. In short, IVT is typically performed using: a linear or circular DNA template containing a promoter, a library of ribonucleoside triphosphates, a buffer system that may include DTT and magnesium ions, a suitable RNA polymerase (e.g., T3, T7, or SP6 RNA polymerase), DNase I, pyrophosphatase, and / or RNase inhibitors. The exact conditions may vary depending on the particular application. The presence of these reagents is generally undesirable in the final mRNA product, and these reagents can be considered impurities or contaminants that can be purified or removed to provide uncontaminated and / or homogeneous mRNA suitable for therapeutic use. While mRNA provided from an in vitro transcription reaction may be desirable in some embodiments, other mRNA sources, including wild-type mRNA derived from bacteria, fungi, plants, and / or animals, may be used in accordance with this disclosure.

[0412] Method for preparing the LNP vaccine of the present invention The LNPs of the present invention can be prepared using various techniques known in the art. For example, multilayer vesicles (MLVs) can be prepared using conventional techniques, such as by depositing selected lipids onto the inner wall of a suitable container or vessel (by dissolving the lipids in a suitable solvent and then evaporating the solvent to leave a thin film inside the vessel) or by spray drying. An aqueous phase can then be added to the vessel with vortex motion, which allows the MLV to form. Unilayer vesicles (ULVs) can then be formed by homogenizing, sonicating, or extruding the multilayer vesicles. Alternatively, ULVs can be formed using detergent removal techniques.

[0413] Various methods are described in US 2011 / 0244026, US 2016 / 0038432, US 2018 / 0153822, US 2018 / 0125989, and PCT / US2020 / 043223 (filed July 23, 2020), and can be used to practice this disclosure. One exemplary method requires encapsulating mRNA by mixing it with a mixture of lipids without first pre-forming lipid nanoparticles, as described in US 2016 / 0038432. Another exemplary method requires encapsulating mRNA by mixing pre-formed LNPs with mRNA, as described in US 2018 / 0153822.

[0414] In some embodiments, the process of preparing LNPs loaded with mRNA includes heating one or more solutions in a solution to a temperature above ambient temperature, wherein the one or more solutions are solutions containing pre-formed lipid nanoparticles, solutions containing mRNA, and mixed solutions containing mRNA encapsulated with LNPs. In some embodiments, the method includes heating one or both of the mRNA solution and the pre-formed LNP solution prior to a mixing step. In some embodiments, the method includes heating one or more of the following solutions during a mixing step: a solution containing pre-formed LNPs, a solution containing mRNA, and a solution containing mRNA encapsulated with LNPs. In some embodiments, the method includes heating the LNP-encapsulated mRNA after a mixing step. In some embodiments, the one or more solutions are heated to a temperature of or greater than about 30°C, 37°C, 40°C, 45°C, 50°C, 55°C, 60°C, 65°C, or 70°C. In some embodiments, one or more solutions are heated to a temperature range of about 25°C-70°C, about 30°C-70°C, about 35°C-70°C, about 40°C-70°C, about 45°C-70°C, about 50°C-70°C, or about 60°C-70°C. In some embodiments, the temperature is about 65°C.

[0415] Various methods can be used to prepare mRNA solutions suitable for this disclosure. In some embodiments, mRNA can be directly dissolved in the buffer solution described herein. In some embodiments, the mRNA solution can be generated by mixing the mRNA stock solution with the buffer solution prior to mixing with a lipid solution for encapsulation. In some embodiments, the mRNA solution can be generated by mixing the mRNA stock solution with the buffer solution immediately prior to mixing with a lipid solution for encapsulation. In some embodiments, a suitable mRNA stock solution may contain mRNA at a concentration in water or buffer of about 0.2 mg / ml, 0.4 mg / ml, 0.5 mg / ml, 0.6 mg / ml, 0.8 mg / ml, 1.0 mg / ml, 1.2 mg / ml, 1.4 mg / ml, 1.5 mg / ml, or 1.6 mg / ml, 2.0 mg / ml, 2.5 mg / ml, 3.0 mg / ml, 3.5 mg / ml, 4.0 mg / ml, 4.5 mg / ml, or 5.0 mg / ml.

[0416] In some embodiments, a pump is used to mix the mRNA stock solution with the buffer solution. Exemplary pumps include, but are not limited to, gear pumps, peristaltic pumps, and centrifugal pumps. Typically, the buffer solution is mixed at a rate greater than that of the mRNA stock solution. For example, the buffer solution may be mixed at a rate of at least 1x, 2x, 3x, 4x, 5x, 6x, 7x, 8x, 9x, 10x, 15x, or 20x that of the mRNA stock solution. In some embodiments, the buffer solution is mixed at a flow rate in the range of about 100-6000 ml / min (e.g., about 100-300 ml / min, 300-600 ml / min, 600-1200 ml / min, 1200-2400 ml / min, 2400-3600 ml / min, 3600-4800 ml / min, 4800-6000 ml / min, or 60-420 ml / min). In some embodiments, the buffer solution is mixed at a flow rate of about 60 ml / min, 100 ml / min, 140 ml / min, 180 ml / min, 220 ml / min, 260 ml / min, 300 ml / min, 340 ml / min, 380 ml / min, 420 ml / min, 480 ml / min, 540 ml / min, 600 ml / min, 1200 ml / min, 2400 ml / min, 3600 ml / min, 4800 ml / min, or 6000 ml / min.

[0417] In some embodiments, the mRNA stock solution is mixed at a flow rate ranging from about 10 to 600 ml / min (e.g., about 5 to 50 ml / min, about 10 to 30 ml / min, about 30 to 60 ml / min, about 60 to 120 ml / min, about 120 to 240 ml / min, about 240 to 360 ml / min, about 360 to 480 ml / min, or about 480 to 600 ml / min). In some embodiments, the mRNA stock solution is mixed at or above the following flow rates: approximately 5 ml / min, 10 ml / min, 15 ml / min, 20 ml / min, 25 ml / min, 30 ml / min, 35 ml / min, 40 ml / min, 45 ml / min, 50 ml / min, 60 ml / min, 80 ml / min, 100 ml / min, 200 ml / min, 300 ml / min, 400 ml / min, 500 ml / min, or 600 ml / min.

[0418] The process of incorporating desired mRNA into lipid nanoparticles is referred to as “addition”. An exemplary method is described in Lasic et al., FEBS Lett. [FEBS Communications] (1992) 312:255-8. The nucleic acid incorporated into the LNP can be entirely or partially located within the internal space of the lipid nanoparticle, within the bilayer of the lipid nanoparticle membrane, or associated with the outer surface of the lipid nanoparticle membrane. Incorporating mRNA into lipid nanoparticles is also referred to herein as “encapsulation,” in which the nucleic acid is completely or substantially contained within the internal space of the lipid nanoparticle.

[0419] Suitable LNPs can be prepared in various sizes. In some embodiments, reduced lipid nanoparticle size is associated with more efficient mRNA delivery. The selection of an appropriate LNP size can take into account the target cell or tissue site and, to some extent, the application for which the lipid nanoparticles will be used.

[0420] Several methods known in the art can be used to regulate the size of lipid nanoparticle clusters. The preferred method described herein utilizes the Zetasizer Nano ZS (Malvern Panalytical) to measure LNP particle size. In one protocol, 10 μl of an LNP sample is mixed with 990 μl of 10% trehalose. This solution is placed in a cuvette and then placed in the Zetasizer machine. The z-mean diameter (nm), or cumulative mean, is considered to be the average size of the LNPs in the sample. The Zetasizer machine can also be used to measure the polydispersity index (PDI) using dynamic light scattering (DLS) and cumulative analysis of autocorrelation functions. The average LNP diameter can be reduced by sonicating the formed LNPs. Intermittent sonication cycles can be alternated with quasi-elastic light scattering (QELS) assessments to guide efficient lipid nanoparticle synthesis.

[0421] In some embodiments, the LNP has an average diameter of 30 nm to 200 nm (e.g., an average diameter of 80 nm to 150 nm).

[0422] In some embodiments, the majority of purified LNPs (i.e., LNPs with a purity greater than about 50%, 55%, 60%, 65%, 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99%) have a size of about 70–150 nm (e.g., about 145 nm, about 140 nm, about 135 nm, about 130 nm, about 125 nm, about 120 nm, about 115 nm, about 110 nm, about 105 nm, about 100 nm, about 95 nm, about 90 nm, about 85 nm, or about 80 nm). In some embodiments, substantially all (e.g., more than 80% or 90%) of the purified lipid nanoparticles are about 70-150 nm in size (e.g., about 145 nm, about 140 nm, about 135 nm, about 130 nm, about 125 nm, about 120 nm, about 115 nm, about 110 nm, about 105 nm, about 100 nm, about 95 nm, about 90 nm, about 85 nm, or about 80 nm).

[0423] In some embodiments, the average size of the LNP in the composition of the present invention is less than 150 nm, less than 120 nm, less than 100 nm, less than 90 nm, less than 80 nm, less than 70 nm, less than 60 nm, less than 50 nm, less than 30 nm, or less than 20 nm.

[0424] In some embodiments, the size of more than 70%, 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the LNPs in the compositions of the present invention ranges from about 40-90 nm (e.g., about 45-85 nm, about 50-80 nm, about 55-75 nm, about 60-70 nm) or about 50-70 nm (e.g., 55-65 nm), which is particularly suitable for lung delivery via nebulization.

[0425] In some embodiments, the dispersibility or molecular size heterogeneity measure (PDI) of LNPs in the pharmaceutical compositions provided by this disclosure is less than about 0.5. In some embodiments, the PDI of LNPs is less than about 0.5, less than about 0.4, less than about 0.3, less than about 0.28, less than about 0.25, less than about 0.23, less than about 0.20, less than about 0.18, less than about 0.16, less than about 0.14, less than about 0.12, less than about 0.10, or less than about 0.08. The PDI can be measured using a Zetasizer machine as described above.

[0426] In some embodiments, greater than about 75%, 80%, 85%, 90%, 95%, 96%, 97%, 98%, or 99% of the purified LNPs in the pharmaceutical compositions provided herein encapsulate mRNA within each individual particle. In some embodiments, substantially all (e.g., greater than 80% or 90%) of the purified lipid nanoparticles in the pharmaceutical compositions encapsulate mRNA within each individual particle. In some embodiments, the encapsulation efficiency of the lipid nanoparticles is 50% to 99%; or greater than about 60%, 65%, 70%, 75%, 80%, 85%, 90%, 92%, 95%, 98%, or 99%. Typically, the lipid nanoparticles used herein have an encapsulation efficiency of at least 90% (e.g., at least 91%, 92%, 93%, 94%, or 95%).

[0427] In some embodiments, the N / P ratio of the LNP is between 1 and 10. In some embodiments, the N / P ratio of the lipid nanoparticles is greater than 1, about 1, about 2, about 3, about 4, about 5, about 6, about 7, or about 8. In a further embodiment, the typical N / P ratio of the LNP described herein is 4.

[0428] In some embodiments, the pharmaceutical composition according to this disclosure contains at least about 0.5 μg, 1 μg, 5 μg, 10 μg, 100 μg, 500 μg, or 1000 μg of encapsulated mRNA. In some embodiments, the pharmaceutical composition contains about 0.1 μg to 1000 μg, at least about 0.5 μg, at least about 0.8 μg, at least about 1 μg, at least about 5 μg, at least about 8 μg, at least about 10 μg, at least about 50 μg, at least about 100 μg, at least about 500 μg, or at least about 1000 μg of encapsulated mRNA.

[0429] In some embodiments, mRNA can be prepared by chemical synthesis or by in vitro transcription (IVT) of a DNA template. In this process, during IVT, a cDNA template is used to generate mRNA transcripts, and the DNA template is degraded by a DNase. The transcripts are purified by deep filtration and tangential flow filtration (TFF). The purified transcripts are further modified by adding caps and tails, and the modified RNA is purified again by deep filtration and TFF.

[0430] The mRNA is then prepared in an aqueous buffer and mixed with an amphiphilic solution containing the lipid components of the LNP. The amphiphilic solution for dissolving the four lipid components of the LNP can be an alcohol solution. In some embodiments, the alcohol is ethanol. The aqueous buffer can be, for example, a citrate, phosphate, acetate, or succinate buffer and can have a pH of about 3.0–7.0 (e.g., about 3.5, about 4.0, about 4.5, about 5.0, about 5.5, about 6.0, or about 6.5). The buffer may contain other components, such as salts (e.g., sodium, potassium, and / or calcium salts). In a particular embodiment, the aqueous buffer has 1 mM citrate, 150 mM NaCl, and a pH of 4.5.

[0431] The process for preparing a composition comprising LNPs and one or more mRNAs involves mixing a buffered mRNA solution with a solution of lipids in ethanol in a controlled homogenizing manner, wherein the lipid:mRNA ratio is maintained throughout the mixing process. In this illustrative example, the mRNA is presented in an aqueous buffer containing citrate monohydrate, trisodium citrate dihydrate, and sodium chloride. The mRNA solution is added to a solution (1 mM citrate buffer, 150 mM NaCl, pH 4.5). A lipid mixture of four lipids (e.g., cationic lipids, PEGylated lipids, cholesterol-based lipids, and accessory lipids) is dissolved in ethanol. The aqueous mRNA solution and the ethanol lipid solution are mixed at a 4:1 volume ratio in a T-shaped mixer with a near-pulseless pump system. The resulting mixture is then subjected to downstream purification and buffer exchange. Buffer exchange can be performed using a dialysis cassette or a TFF system. A TFF can be used to concentrate and buffer exchange the resulting nascent LNPs immediately after formation via the T-mixing process. The percolation process is a continuous operation, and the volume is kept constant by adding an appropriate buffer at the same rate as the percolation stream.

[0432] carrier On the one hand, this document discloses vectors including the mRNA compositions disclosed herein. RNA sequences encoding target proteins (e.g., mRNA encoding the polypeptides disclosed herein) can be cloned into various types of vectors. For example, the nucleic acid can be cloned into vectors including, but not limited to, plasmids, phage particles, phage derivatives, animal viruses, and granules. Specific target vectors may include expression vectors, replication vectors, probe generation vectors, sequencing vectors, and vectors optimized for in vitro transcription.

[0433] In some embodiments, the vector can be used to express mRNA in host cells. In various embodiments, the vector can be used as a template for IVT. The construction of the most preferably translated IVT mRNA for therapeutic use is disclosed in detail in Sahin et al. (2014). Nat. Rev. Drug Discov. 13, 759–780; Weissman (2015). Expert Rev. Vaccines 14, 265–281.

[0434] In some embodiments, the vectors disclosed herein may include at least the following from 5' to 3': an RNA polymerase promoter; a polynucleotide sequence encoding a 5' UTR; a polynucleotide sequence encoding an ORF; a polynucleotide sequence encoding a 3' UTR; and a polynucleotide sequence encoding at least one RNA aptamer. In some embodiments, the vectors disclosed herein may include polynucleotide sequences encoding multiple (A) sequences and / or multiple adenosine esterification signals.

[0435] Various RNA polymerase promoters are known. In some embodiments, the promoter may be the T7 RNA polymerase promoter. Other useful promoters may include, but are not limited to, the T3 and SP6 RNA polymerase promoters. Common nucleotide sequences for the T7, T3, and SP6 promoters are known.

[0436] This document also discloses host cells (e.g., mammalian cells, such as human cells) including the vectors or RNA compositions disclosed herein. “Host cell” includes individual cells or cell cultures that may be or are already recipients of exogenous nucleic acids. Host cells include the offspring of a single host cell, and may not necessarily be identical to the original parent cell (morphologically or in terms of total DNA complement) due to natural, accidental, or intentional mutations and / or alterations. Host cells include cells transfected or infected in vivo or in vitro with the nucleic acids or vectors disclosed herein.

[0437] The vector can be introduced into target cells using any of a variety of different methods, such as commercially available methods including, but not limited to, electroporation (Amaxa Nucleofector-II (Amaxa Biosystems, Cologne, Germany)), (ECM 830 (BTX) (Harvard Instruments, Boston, MA) or Gene Pulser II (BioRad, Denver, Colorado), Multiporator (Eppendorf, Hamburg, Germany), cationic liposome-mediated transfection using lipid transfection, polymer encapsulation, peptide-mediated transfection, bioballistic particle delivery systems such as “gene guns” (see, for example, Nishikawa, et al. (2001). Hum Gene Ther. [Human Gene Therapy] 12(8):861-70) or TransIT-RNA transfection kit (Mirus, Madison, WI).

[0438] Chemical means for introducing a carrier into a host cell include colloidal dispersion systems such as macromolecular complexes, nanocapsules, microspheres, beads, and lipid-based systems (including oil-in-water emulsions, micelles, mixed micelles, and liposomes). An exemplary colloidal system used as a carrier for in vitro and in vivo delivery is a liposome (e.g., an artificial membrane vesicle).

[0439] Regardless of the method used to introduce exogenous nucleic acids into host cells or otherwise expose cells to the inhibitors disclosed herein, various measurements can be performed to confirm the presence of mRNA sequences in host cells.

[0440] Self-replicating RNA, trans-replicating RNA, and non-replicating RNA Typically, the nucleic acid molecules described herein are non-replicating RNAs. However, the nucleic acid molecules described herein may alternatively be self-replicating RNAs or trans-replicating RNAs.

[0441] Self-replicating RNA Self-replicating (or self-amplifying) RNA can be produced by using replication elements derived from, for example, alphaviruses and replacing structural viral proteins with nucleotide sequences encoding target proteins (such as the polypeptides described herein). Self-replicating RNA is typically a positive-strand molecule that can be directly translated upon delivery to the cell, and this translation provides an RNA-dependent RNA polymerase that then produces both antisense and sense transcripts from the delivered RNA. Thus, the delivered RNA results in the production of multiple daughter RNAs. These daughter RNAs, along with the collinear subgenomic transcripts themselves, can be translated to provide in situ expression of the encoded antigen, or can be transcribed to provide additional transcripts with the same meaning as the delivered RNA, which are then translated to provide in situ expression of the antigen. The overall result of this transcriptional sequence is a massive amplification of the number of introduced replicon RNAs, and thus the encoded antigen becomes the dominant polypeptide product of the cell.

[0442] A suitable system for self-replication in this manner is the use of alphavirus-based replicons. These replicons are positive-strand (sense-strand) RNAs that, upon delivery to the cell, induce translation by a replicase (or replicase-transcriptionase). The replicase is translated into a polyprotein, which is automatically cleaved to provide a replication complex that forms a genomic strand copy of the positive-strand delivered RNA. These negative-strand transcripts can themselves be transcribed to produce further copies of the positive-strand parental RNA and also to produce subgenomic transcripts encoding the antigen. Translation of the subgenomic transcripts then induces in situ expression of the antigen by the infected cell. Suitable alphavirus replicons can utilize replicases from Sindbis virus, Semliki Forest virus, Eastern Equine encephalitis virus, Venezuelan Equine encephalitis virus, etc. Mutated or wild-type viral sequences can be used; for example, the attenuated TC83 mutant of VEEV has been used in replicons, see reference WO 2005 / 113782, which is incorporated herein by reference.

[0443] In one embodiment, each self-replicating RNA described herein encodes (i) an RNA-dependent RNA polymerase, which can transcribe RNA from the self-replicating RNA molecule, and (ii) an influenza protein antigen. The polymerase may be an alphavirus replicase, for example, including one or more of the alphavirus proteins nsP1, nsP2, nsP3, and nsP4. Although the native alphavirus genome encodes structural virion proteins in addition to non-structural replicase polyproteins, in some embodiments, the self-replicating RNA molecule does not encode alphavirus structural proteins. Therefore, the self-replicating RNA may result in the production of copies of its own genomic RNA within the cell, but not in the production of RNA-containing virions. The inability to produce these virions means that, unlike wild-type alphavirus, the self-replicating RNA molecule cannot permanently persist itself in an infectious form. The alphavirus structural proteins necessary for permanent persistence in wild-type virus are not present in the self-replicating RNA disclosed herein, and their positions are replaced by one or more genes encoding a target immunogen, such that the subgenomic transcript encodes an immunogen, rather than alphavirus structural virion proteins. A further detailed description of the self-replicating RNA is found in WO 2011005799, which is incorporated herein by reference.

[0444] Trans-replicating RNA Trans-replicating (or trans-amplified) RNA has elements similar to those of self-replicating RNA described above. However, for trans-replicating RNA, two separate RNA molecules are used. The first RNA molecule encodes the aforementioned RNA replicase (e.g., alphavirus replicase) and the second RNA molecule encodes the target protein (e.g., the polypeptide described herein). The RNA replicase can replicate one or both of the first and second RNA molecules, thereby greatly increasing the copy number of the RNA molecule encoding the target protein. Trans-replicating RNA is described in further detail in WO 2017162265, which is incorporated herein by reference.

[0445] Non-replicating RNA Non-replicating (or non-amplifying) RNA is RNA that does not have the ability to replicate itself.

[0446] Therapeutic uses On the other hand, the present invention provides polypeptides, nucleic acids, combinations, or compositions of the present invention for use as medicaments. The present invention also provides the use of polypeptides, nucleic acids, combinations, or compositions of the present invention in the manufacture of medicaments. These medicaments can be used to treat or prevent the diseases described herein. The present invention further provides a method of treating or preventing a disease, the method comprising administering the polypeptides, nucleic acids, combinations, or compositions of the present invention to a subject in need. The polypeptides, nucleic acids, combinations, or compositions of the present invention can be administered, for example, in an amount effective in treating or preventing the disease in the subject. Therefore, polypeptides, nucleic acids, combinations, or compositions can be administered in an effective amount.

[0447] On the other hand, the present invention provides polypeptides, nucleic acids, combinations, or compositions of the present invention for use in treating or preventing *Propionibacterium acnes* infection in a subject (e.g., a human). The present invention also provides the use of polypeptides, nucleic acids, combinations, or compositions of the present invention in the manufacture of medicaments for treating or preventing *Propionibacterium acnes* infection in a subject (e.g., a human). The present invention further provides a method for treating or preventing *Propionibacterium acnes* infection in a subject (e.g., a human), the method comprising administering to the subject the polypeptides, nucleic acids, combinations, or compositions of the present invention. The polypeptides, nucleic acids, combinations, or compositions of the present invention may be administered, for example, in an amount effective in treating or preventing *Propionibacterium acnes* infection in the subject (i.e., in an effective amount). *Propionibacterium acnes* infection may be mild, moderate, or severe (e.g., moderate or severe).

[0448] The peptides, nucleic acids, combinations or compositions of the present invention can be used to induce an immune response in subjects (e.g., an immune response against Propionibacterium acnes infection).

[0449] On the other hand, the present invention provides polypeptides, nucleic acids, combinations, or compositions of the present invention for treating or preventing acne (also known as acne vulgaris) in a subject (e.g., a human). The present invention also provides the use of polypeptides, nucleic acids, combinations, or compositions of the present invention in the preparation of medicaments for treating or preventing acne in a subject (e.g., a human). The present invention further provides a method for treating or preventing acne in a subject (e.g., a human), the method comprising administering the polypeptides, nucleic acids, combinations, or compositions of the present invention to the subject. The polypeptides, nucleic acids, combinations, or compositions of the present invention may be administered, for example, in an amount effective in treating or preventing acne in the subject (i.e., in an effective amount). Acne may be caused by *Propionibacterium acnes*. Acne may be mild, moderate, or severe (e.g., moderate or severe).

[0450] In some embodiments, the treatment as described herein achieves a reduction in the number of inflammatory acne lesions by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% (e.g., at least 50%) of the number of inflammatory acne lesions relative to the number of inflammatory acne lesions before treatment, for example, at 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after treatment (e.g., 3 or 6 months) following treatment (e.g., after the last dose). In some embodiments, the inflammatory acne lesions are located on the subject's face. In some embodiments, the inflammatory acne lesions are located on the subject's face and another body area (e.g., the chest or back).

[0451] In some embodiments, the treatment as described herein achieves a reduction in the number of non-inflammatory acne lesions by at least 5%, at least 10%, at least 15%, at least 20%, at least 25%, at least 30%, at least 35%, at least 40%, at least 45%, at least 50%, at least 55%, at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, or at least 90% (e.g., at least 50%) of the number of non-inflammatory acne lesions relative to the number of non-inflammatory acne lesions before treatment, for example, at 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after treatment (e.g., 3 or 6 months) following treatment (e.g., after the last dose).

[0452] In some embodiments, the treatment as described herein achieves a reduction in the Investigator Global Assessment (IGA) score for the subject relative to the pre-treatment score at 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months (e.g., 3 or 6 months) after treatment administration (e.g., after the last dose). In some embodiments, the treatment as described herein achieves at least a two-level improvement in the Investigator Global Assessment (IGA) score for the subject relative to the pre-treatment score at 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months (e.g., 3 or 6 months) after treatment administration (e.g., after the last dose). In some embodiments, for example at 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or 12 months after treatment administration (e.g., 3 or 6 months) (e.g., after the last dose of treatment), the improvement in the IGA score results in an IGA score of 0, 1, or 2 (e.g., 0 or 1).

[0453] The polypeptides, nucleic acids, combinations or compositions of the present invention can be used to treat or prevent chronic blepharitis and / or endophthalmitis associated with Propionibacterium acnes infection.

[0454] This invention provides polypeptides, nucleic acids, combinations, or compositions of the invention for use in methods of providing protective immunity against *Propionibacterium acnes* infection in a subject. The invention also provides the use of the polypeptides, nucleic acids, combinations, or compositions of the invention in the manufacture of a medicament for use in methods of providing protective immunity against *Propionibacterium acnes* infection in a subject. The invention further provides a method of providing protective immunity against *Propionibacterium acnes* infection in a subject, the method comprising administering the polypeptides, nucleic acids, combinations, or compositions of the invention to the subject. The polypeptides, nucleic acids, combinations, or compositions of the invention may be administered, for example, in an amount that effectively provides protective immunity against *Propionibacterium acnes* infection in a subject (i.e., in an effective amount). Protective immunity can prevent the development of pathological conditions caused by *Propionibacterium acnes* infection (e.g., acne). Protective immunity can prevent or reduce the development of *Propionibacterium acnes*-related indications.

[0455] The peptides, nucleic acids, combinations, or compositions of the present invention can elicit antibodies (e.g., antigen-specific antibodies), such as IgG. Antibodies (DsA1, DsA2, and / or PITP-specific antigens) can bind to the surface of *Propionibacterium acnes* bacteria. Antibodies (DsA1, DsA2, and / or PITP-specific antigens) can opsonize *Propionibacterium acnes* bacteria. This may allow immune effector cells (e.g., phagocytes) to be recruited to the bacteria. *Propionibacterium acnes* bacteria can be phagocytosed by the recruited immune effector cells. Within the IgG isotype, the antibody can be an IgG1, IgG2, IgG3, or IgG4 subclass. The antibody may have a κ or λ light chain. In some embodiments, the peptides, nucleic acids, combinations, or compositions of the present invention can elicit an antibody response that is cross-reactive against two or more strains or phylogenetic types of *Propionibacterium acnes*, for example, a cross-reactive antibody response against two or more phylogenetic types (e.g., cross-reactive against phylogenetic types IA1, IA2, IB, IC, II, and III).

[0456] The peptides, nucleic acids, combinations, or compositions of the present invention can reduce inflammation associated with (e.g., caused by) *Propionibacterium acnes* infection. The peptides, nucleic acids, combinations, or compositions of the present invention can reduce *Propionibacterium acnes*-mediated tissue inflammation.

[0457] The polypeptides, nucleic acids, combinations, or compositions of the present invention can inhibit biofilm formation of *Propionibacterium acnes*. In some embodiments, biofilm formation can be prevented. In some embodiments, biofilm formation can be reduced.

[0458] Administration of the peptides, nucleic acids, combinations, or compositions of the present invention to subjects enables them to generate a population of *Propionibacterium acnes* antigen-responsive memory B cells upon exposure to *Propionibacterium acnes* bacteria or *Propionibacterium acnes* antigens. The peptides, nucleic acids, combinations, or compositions of the present invention can be used to induce primary and / or enhance immune responses.

[0459] The polypeptides, nucleic acids, combinations, or compositions of the present invention can be used in primary-boost vaccination regimens. Immunization against *Propionibacterium acnes* infection according to the present invention can be provided by administering a primary vaccine comprising the polypeptides, nucleic acids, combinations, or compositions of the present invention, followed by a booster vaccine. The booster vaccine can be the same as the initial vaccine.

[0460] In some embodiments, the subject is a vertebrate, such as a mammal, such as a human or a veterinary mammal (e.g., cat, dog, horse, cow, sheep, cattle, deer, goat, pig, rodent (e.g., mouse)). In a preferred embodiment, the subject is a human. The subject (e.g., a human subject) can be male or female. The subject can be a child (0-10 years old), an adolescent (10-18 years old), or an adult (18 years and older). In some embodiments, the human subject can be 0-30 or 0-45 years old (e.g., 5-18, 12-45, 18-45, or 9-45). For example, the subject can be 9-45, 12-45, or 18-45 (e.g., 9-45 years old).

[0461] In some embodiments, the subject has at least one of the following: (i) an IGA scale score of 3 or 4; (ii) at least 25 non-inflammatory lesions (e.g., open and / or closed comedones), optionally located on the subject's face; (iii) optionally at least 20 inflammatory lesions (e.g., papules and / or pustules) on the subject's face; (iv) optionally two or fewer nodular cystic lesions (e.g., nodules and / or cysts) on the subject's face. In some embodiments, the subject has at least two of (i)-(iv). In some embodiments, the subject has at least three of (i)-(iv). Typically, the subject has all four of (i)-(iv). A subject having all four of (i)-(iv) may have moderate / severe acne.

[0462] Acne vulgaris (also referred to herein as acne) exhibits varying degrees of severity: mild, moderate, and severe. Moderate and severe acne account for more than one-third of all cases and require medication. In some embodiments, subjects may have a mild, moderate, or severe (e.g., moderate or severe acne) *Propionibacterium acnes* infection. Subjects may have mild, moderate, or severe acne (e.g., moderate or severe acne). In some embodiments, subjects may have mild, moderate, or severe acne (e.g., moderate or severe acne).

[0463] Application mode The polypeptides, nucleic acids, combinations, or compositions of the present invention can be administered parenterally (e.g., intramuscular, intradermal, subcutaneous, intraperitoneal, intravenous, or into the interstitial space of tissues) or via rectal, oral, vaginal, topical, transdermal, intranasal, sublingual, ocular, ocular, pulmonary, or other mucosal administration. In some embodiments, delivery is via mucosal administration. Typically, administration is intramuscular.

[0464] In some embodiments, the nucleic acids, peptides, compositions, or combinations thereof (e.g., compositions) of the present invention are provided for intramuscular (IM) injection. The nucleic acids, peptides, compositions, or combinations thereof (e.g., compositions) may be applied to the thigh or upper arm of a subject, such as the deltoid muscle of the upper arm. In some embodiments, the nucleic acids, peptides, compositions, or combinations thereof (e.g., compositions) are provided in a pre-filled syringe or injector (e.g., single-chamber or multi-chamber). Injection may be performed with a needle (e.g., a hypodermic needle), but needle-free injection may be used alternatively. A typical intramuscular dose is 0.5 ml. In some embodiments, the nucleic acids, peptides, compositions, or combinations thereof (e.g., compositions) are provided for inhalation and are provided in a pre-filled pump, aerosol, or inhaler.

[0465] In some embodiments, the nucleic acids, peptides, compositions, or combinations of the present invention (e.g., compositions) are provided for skin injection, such as in the epidermis, dermis, or subcutaneous tissue of the skin. In some embodiments, the nucleic acids, peptides, compositions, or combinations of the present invention (e.g., compositions) are provided in devices suitable for skin injection (e.g., needles (e.g., epidermal, dermal, or hypodermal needles)), needle-free devices, microneedle devices, or microprotrusion array devices. Examples of microneedle or microprotrusion array devices suitable for skin injection according to the present invention are described in US 20230270842 A1, US 20220339416 A1, US 20210085598 A1, US 20200246450 A1, US 20220143376 A1, US 20180264244 A1, US 20180263641 A1, and US20110245776 A1.

[0466] The nucleic acids, polypeptides, compositions, or combinations thereof of the present invention (e.g., compositions) can be used to induce systemic, skin, and / or mucosal immunity.

[0467] Dosage therapy can be administered on a single-dose schedule or a multiple-dose schedule. Multiple (e.g., two or three) doses may be used in the initial immunization schedule and / or in the booster immunization schedule. A booster schedule may follow the initial dose schedule. Multiple doses (e.g., two or three) are typically administered to subjects in need at intervals of at least one week (e.g., about 2 weeks, about 3 weeks, about 4 weeks, about 6 weeks, about 8 weeks, about 10 weeks, about 12 weeks, about 16 weeks, etc.) to achieve the desired therapeutic or preventative effect (e.g., two months). Doses (e.g., initial and booster doses) may be spaced apart by intervals such as 1 week, 2 weeks, 3 weeks, 4 weeks, 1 month, 2 months, 3 months, 4 months, 5 months, 6 months, 1 year, 2 years, 5 years, or 10 years (e.g., two months). In some embodiments, a single dose is administered intramuscularly to the subject. In some embodiments, two doses are administered intramuscularly to the subject (e.g., two months apart).

[0468] The compositions of the present invention may be in the form of temporary preparations, for example, the compositions of the present invention may be lyophilized. Such compositions may be reconstituted with physiological buffer (e.g., PBS) immediately before use. The compositions of the present invention may be provided in the form of aqueous solutions or frozen aqueous solutions and may be administered directly to subjects without reconstitution (or after thawing if previously frozen).

[0469] In some embodiments of the compositions comprising one or more nucleic acids (e.g., mRNA) described herein, a single dose of the composition comprises 1-300 (or 1-50) μg of the mRNA described herein (e.g., monovalent or polyvalent). For example, a single dose may contain about 2.5 μg, about 5 μg, about 7.5 μg, about 10 μg, about 12.5 μg, about 15 μg, about 30 μg, about 45 μg, about 60 μg, about 75 μg, about 90 μg, about 105 μg, about 120 μg, about 135 μg, about 150 μg, about 165 μg, about 180 μg, about 195 μg, about 210 μg, about 225 μg, about 240 μg, about 250 μg, about 260 μg, about 275 μg, or about 300 μg of one or more nucleic acids (e.g., one or more mRNAs) described herein, for example for intramuscular (IM) injection. In some embodiments, the composition comprises 40-50 μg (e.g., about 45 μg) of one or more nucleic acids (e.g., one or more mRNAs). In some embodiments, the composition comprises 110-140 µg (e.g., about 120 µg) of one or more nucleic acids (e.g., one or more mRNAs). In some embodiments, the composition comprises 200-250 µg (e.g., about 225 µg) of one or more nucleic acids (e.g., one or more mRNAs).

[0470] The composition may comprise the three nucleic acids described herein (e.g., three mRNAs) encoding the different polypeptides described herein, and these nucleic acids may be present in a 1:1:1 weight ratio. In some embodiments, the composition comprises a total of about 45 μg, about 120 μg, or about 225 μg of nucleic acids. Thus, the composition may comprise about 15 μg, about 40 μg, or about 75 μg of each of the three nucleic acids.

[0471] The composition may contain two nucleic acids (e.g., two mRNAs) encoding different polypeptides described herein, and these nucleic acids may be present in a 1:1 weight ratio.

[0472] In further embodiments, the compositions of the present invention can be provided as multivalent single doses containing a plurality of (e.g., 2, 3 or 4) LNPs, each LNP for a different antigen, and each LNP having an mRNA amount of, for example, 2.5 μg, about 5 μg, about 7.5 μg, about 10 μg, about 12.5 μg, about 15 μg, about 30 μg, about 45 μg, about 60 μg, about 75 μg, about 90 μg, about 105 μg, about 120 μg, about 135 μg, about 150 μg, about 165 μg, about 180 μg, about 195 μg, about 210 μg, about 225 μg, about 250 μg, about 275 μg or about 300 μg.

[0473] In some embodiments, one or more nucleic acid compositions of the present invention are administered to a subject. The nucleic acid composition may comprise a nucleic acid containing a nucleotide sequence encoding a polypeptide antigen as described herein. The nucleic acid compositions may be administered simultaneously, separately, or sequentially. In some embodiments, a nucleic acid combination of the present invention is administered to a subject. The nucleic acid combination includes a combination of two or more (e.g., three) nucleic acids as described herein. The nucleic acids within the combination may be administered simultaneously, separately, or sequentially.

[0474] In some embodiments, one or more polypeptide compositions of the present invention are administered to a subject. The polypeptide composition may contain a polypeptide antigen as described herein. The polypeptide compositions may be administered simultaneously, separately, or sequentially. In some embodiments, a polypeptide combination of the present invention is administered to a subject. The polypeptide combination includes a combination of two or more (e.g., three) polypeptides as described herein. Nucleic acids within the combination may be administered simultaneously, separately, or sequentially.

[0475] In some embodiments, one or more nucleic acid compositions and one or more polypeptide compositions of the present invention are administered to a subject. The nucleic acid compositions and one or more polypeptide compositions may be administered simultaneously, separately, or sequentially.

[0476] Compositions administered alone or sequentially may be administered within 12 months, within 6 months, or within 1 month or less (e.g., within 10 days). Compositions may be administered within 7 days, 3 days, 2 days, or 24 hours of each other. Simultaneous administration may involve the simultaneous administration of the compositions of the invention. Simultaneous administration may include administering the compositions of the invention to a patient within 12 hours, 6 hours, 3 hours, 2 hours, or 1 hour of each other (typically within the same visit to a clinical center).

[0477] The present invention also provides a kit comprising one or more compositions described herein in one or more containers, or provides one or more compositions as described herein in one or more containers and physiological buffer for reconstitution in another container. The one or more containers may contain a single-use dose or a multiple-use dose. The containers may be pretreated glass vials or ampoules. The kit may include instructions for use.

[0478] definition The term “comprising” encompasses both “including” and “consisting of”, for example, a composition that “comprising” X may consist of only X or may contain other substances, such as X + Y.

[0479] With numerical values x The relevant term "about" is optional and means, for example... x ± 10%.

[0480] As used herein, the term "effective amount" means an amount sufficient to achieve a beneficial or desired outcome (e.g., an amount of nucleic acid, peptide, combination, or composition as described herein). An effective amount may be administered by one or more applications, uses, or doses and is not intended to be limited to a particular formulation or route of administration. The term "effective amount" includes, for example, therapeutically effective amounts and / or preventively effective amounts. As used herein, the term "effective amount" means an amount (e.g., a ...

Claims

1. A nucleic acid comprising a nucleotide sequence encoding a modified C. acnes CAMP2 polypeptide, wherein the modified C. acnes CAMP2 polypeptide comprises an amino acid sequence comprising a C. acnes CAMP2 polypeptide sequence and a transmembrane domain sequence.

2. The nucleic acid of claim 1, wherein: (a) the transmembrane domain sequence is located at the C-terminal end of the modified C. acnes CAMP2 polypeptide; and / or (b) the transmembrane domain sequence comprises a viral transmembrane domain sequence, optionally selected from the group consisting of an influenza hemagglutinin (HA) transmembrane domain sequence, a SARS CoV-2 spike transmembrane domain sequence, a VZV gB transmembrane domain sequence, a VZV gE transmembrane domain sequence, a VZV gI transmembrane domain sequence, a VZV gK transmembrane domain sequence, a measles F protein transmembrane domain sequence, a rubella E1 protein transmembrane domain sequence, a rubella E2 protein transmembrane domain sequence, a mumps F protein transmembrane domain sequence, an Ebola GP protein transmembrane domain sequence, and a rabies transmembrane domain sequence, e.g., wherein the transmembrane domain comprises an amino acid sequence according to any one of the sequences in Table 4.

3. The nucleic acid of claim 1 or 2, wherein the modified C. acnes CAMP2 polypeptide comprises a non-native secretion signal peptide sequence (e.g., a viral secretion signal peptide sequence), optionally wherein: (a) the secretion signal peptide sequence is located at the N-terminal end of the modified C. acnes CAMP2 polypeptide; and / or (b) the secretion signal peptide sequence is a viral secretion signal peptide sequence, optionally selected from the group consisting of an influenza hemagglutinin (HA) secretion signal peptide sequence, a SARS CoV-2 spike secretion signal peptide sequence, a VZV gB secretion signal peptide sequence, a VZV gE secretion signal peptide sequence, a VZV gI secretion signal peptide sequence, a VZV gK secretion signal peptide sequence, a measles F protein secretion signal peptide sequence, a rubella E1 protein secretion signal peptide sequence, a rubella E2 protein secretion signal peptide sequence, a mumps F protein secretion signal peptide sequence, an Ebola GP protein secretion signal peptide sequence, a smallpox 6 kDa IC protein secretion signal peptide sequence, and a rabies G protein secretion signal peptide sequence, e.g., wherein the secretion signal peptide sequence comprises a sequence according to any one of the sequences in Table 3.

4. The nucleic acid of any one of claims 1-3, wherein the nucleic acid is a messenger RNA (mRNA), optionally wherein: (a) the mRNA comprises a 5’ cap, at least one 5’ untranslated region (5’ UTR), at least one 3’ untranslated region (3’ UTR), and / or at least one polyadenylation (poly(A)) sequence; (b) the mRNA is unmodified or comprises at least one chemical modification, optionally wherein the mRNA comprises at least one chemical modification, e.g., wherein the chemical modification comprises N1-methyl pseudouridine; and / or (c) the mRNA comprises at least one chemical modification, e.g., wherein the chemical modification comprises N1-methyl pseudouridine. (c) the mRNA is a self-replicating mRNA or a non-replicating mRNA, e.g., a non-replicating mRNA.

5. The nucleic acid of any one of claims 1-4, wherein: (a) the modified C. acnes CAMP2 polypeptide comprises a sequence according to any one of SEQ ID NO: 207 or SEQ ID NOs: 5-9 (e.g., SEQ ID NO: 207) or a sequence having at least 60% (e.g., at least 85%, 90%, or 95%) identity thereto; and / or (b) the nucleic acid comprises a nucleotide sequence according to any one of SEQ ID NOs: 90-94 or SEQ ID NOs: 391-392 (e.g., SEQ ID NO: 91) or a sequence having at least 50% (e.g., at least 75%, 80%, or 85%, e.g., 75% or 80%) identity thereto.

6. The nucleic acid of any one of claims 1-5, wherein the nucleic acid is an mRNA comprising or consisting of (e.g., consisting of) the following structural elements: (i) a 5’ cap having the structure: (ii) a 5’ untranslated region (5’ UTR) having a nucleic acid sequence according to SEQ ID NO: 265; (iii) a protein coding region having a nucleic acid sequence according to any one of SEQ ID NOs: 90-91 or SEQ ID NOs: 391-392 (e.g., SEQ ID NO: 91); (iv) a 3’ untranslated region (3’ UTR) having a nucleic acid sequence according to SEQ ID NO: 266; and (v) a poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), e.g., wherein the poly-A tail comprises at least 100 adenosine nucleotides.

7. A modified C. acnes CAMP2 polypeptide having an amino acid sequence comprising a C. acnes CAMP2 polypeptide sequence and a transmembrane domain sequence.

8. The modified C. acnes CAMP2 polypeptide of claim 7, wherein: (A) the transmembrane domain sequence is located at the C-terminus of the modified C. acnes CAMP2 polypeptide; and / or (B) the transmembrane domain comprises a viral transmembrane domain sequence, optionally selected from the group consisting of an influenza hemagglutinin (HA) transmembrane domain sequence, a SARS CoV-2 spike transmembrane domain sequence, a VZV gB transmembrane domain sequence, a VZV gE transmembrane domain sequence, a VZV gI transmembrane domain sequence, a VZV gK transmembrane domain sequence, a measles F protein transmembrane domain sequence, a rubella E1 protein transmembrane domain sequence, a rubella E2 protein transmembrane domain sequence, a mumps F protein transmembrane domain sequence, an Ebola GP protein transmembrane domain sequence, and a rabies transmembrane domain sequence, e.g., wherein the transmembrane domain comprises an amino acid sequence according to any one of the sequences in Table 4.

9. The modified C. acnes CAMP2 polypeptide of any of claims 7-8, comprising a sequence according to SEQ ID NO: 207 or any of SEQ ID NOs: 5-9 (e.g., SEQ ID NO: 207) or a sequence having at least 60% (e.g., at least 85%, 90%, or 95%) identity thereto.

10. A composition comprising: (a) the nucleic acid (e.g., mRNA) of any of claims 1-6, optionally wherein the composition is an immunogenic composition; (b) a nucleic acid comprising a nucleotide sequence encoding a C. acnes CAMP2 polypeptide, wherein the nucleic acid is mRNA, and wherein the composition is an immunogenic composition; or (c) a nucleic acid (e.g., mRNA) comprising a nucleotide sequence encoding a C. acnes CAMP2 polypeptide and one or more of the following: (i) a nucleic acid comprising a nucleotide sequence encoding a C. acnes DsAl polypeptide; (ii) a nucleic acid comprising a nucleotide sequence encoding a C. acnes DsA2 polypeptide; (iii) a nucleic acid comprising a nucleotide sequence encoding a C. acnes PITP polypeptide; (iv) a nucleic acid comprising a nucleotide sequence encoding a chimeric C. acnes DsAl / DsA2 polypeptide; and (v) a nucleic acid comprising a nucleotide sequence encoding a chimeric C. acnes DsAl / DsA2 / PITP polypeptide; optionally wherein the composition according to (c) is an immunogenic composition, further optionally wherein the composition according to any of (a)-(c) is in a frozen liquid form or in a lyophilized form (e.g., in a lyophilized form).

11. The composition of claim 10(c), wherein the composition comprises the nucleic acid according to (c)(iii) and / or the nucleic acid according to (c)(iv), optionally wherein: (A) the C. acnes CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 203 or a sequence having at least 60% (e.g., at least 85%) identity thereto; (B) the chimeric C. acnes DsAl / DsA2 polypeptide comprises a sequence according to SEQ ID NO: 70 or a sequence having at least 90% identity thereto; and (C) the C. acnes PITP polypeptide comprises a sequence according to SEQ ID NO: 73 or a sequence having at least 75% identity thereto.

12. The composition of claim 10, wherein the composition is as defined in claim 10(a) or (b), and wherein the composition further comprises one or more of the following: (i) a nucleic acid comprising a nucleotide sequence encoding a C. acnes DsAl polypeptide; (ii) a nucleic acid comprising a nucleotide sequence encoding a C. acnes DsA2 polypeptide; (iii) a nucleic acid comprising a nucleotide sequence encoding a C. acnes PITP polypeptide; (iv) a nucleic acid comprising a nucleotide sequence encoding a chimeric P. acnes DsAl / DsA2 polypeptide; and (v) a nucleic acid comprising a nucleotide sequence encoding a chimeric P. acnes DsAl / DsA2 / PITP polypeptide.

13. The composition of claim 12, wherein the composition comprises a nucleic acid according to (iii) and / or a nucleic acid according to (iv), optionally wherein: (A) the P. acnes CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 207 or a sequence having at least 60% (e.g., at least 85%) identity thereto; the chimeric P. acnes DsAl / DsA2 polypeptide comprises a sequence according to SEQ ID NO: 70 or a sequence having at least 90% identity thereto; and the P. acnes PITP polypeptide comprises a sequence according to SEQ ID NO: 73 or a sequence having at least 75% identity thereto; or (B) the nucleotide sequence encoding the P. acnes CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 91 or a sequence having at least 50% identity thereto; the nucleotide sequence encoding the chimeric P. acnes DsAl / DsA2 polypeptide comprises a sequence according to SEQ ID NO: 113 or a sequence having at least 75% identity thereto; and the nucleotide sequence encoding the P. acnes PITP polypeptide comprises a sequence according to SEQ ID NO: 115 or a sequence having at least 50% identity thereto.

14. The composition of claim 10(c) or any one of claims 11-13, wherein any two or more nucleic acids are on the same nucleic acid molecule or on different nucleic acid molecules (e.g., wherein all nucleic acids in the composition are on the same nucleic acid molecule, or wherein all nucleic acids in the composition are on separate nucleic acid molecules).

15. The composition of any one of claims 10-14, wherein: (a) the mRNA comprises a 5’ cap, at least one 5’ untranslated region (5’ UTR), at least one 3’ untranslated region (3’ UTR), and / or at least one polyadenylation (poly(A)) sequence; (b) the mRNA is unmodified or comprises at least one chemical modification, optionally wherein the mRNA comprises at least one chemical modification, e.g., wherein the chemical modification comprises Nl-methyl pseudouridine; and / or (c) the mRNA is a self-replicating mRNA or a non-replicating mRNA, e.g., a non-replicating mRNA.

16. The composition of any one of claims 10-15, wherein the composition comprises: (A) a first mRNA comprising or consisting of (e.g., consisting of) the following structural elements: (i) a 5’ cap; (ii) a 5’ UTR having a nucleic acid sequence according to SEQ ID NO: 265; (iii) a protein-coding region having a nucleic acid sequence according to SEQ ID NO: 91; (iv) a 3’ UTR having a nucleic acid sequence according to SEQ ID NO: 266; and (v) a poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), e.g., wherein the poly-A tail comprises at least 100 adenosine nucleotides; (B) a second mRNA comprising or consisting of (e.g., consisting entirely of) the following structural elements: (i) a 5’ cap; (ii) a 5’ UTR having a nucleic acid sequence according to SEQ ID NO: 265; (iii) a protein coding region having a nucleic acid sequence according to SEQ ID NO: 113; (iv) a 3’ UTR having a nucleic acid sequence according to SEQ ID NO: 266; and (v) a poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), e.g., wherein the poly-A tail comprises at least 100 adenosine nucleotides; and (C) a third mRNA comprising or consisting of (e.g., consisting entirely of) the following structural elements: (i) a 5’ cap; (ii) a 5’ UTR having a nucleic acid sequence according to SEQ ID NO: 265; (iii) a protein coding region having a nucleic acid sequence according to SEQ ID NO: 115; (iv) a 3’ UTR having a nucleic acid sequence according to SEQ ID NO: 266; and (v) a poly-A tail, optionally wherein the poly-A tail comprises at least 75 adenosine nucleotides (e.g., about 80 adenosine nucleotides) or at least 100 adenosine nucleotides (e.g., about 115 adenosine nucleotides), e.g., wherein the poly-A tail comprises at least 100 adenosine nucleotides; wherein the 5’ cap has the following structure: ， and optionally wherein one or more (e.g., all three) of the first, second, and third mRNAs comprise at least one chemical modification, e.g., wherein the chemical modification comprises N1-methyl pseudouridine, preferably wherein all uridines are replaced with N1-methyl pseudouridine.

17. The composition of any of claims 10-16, wherein the composition comprises a total of about 45 µg, about 120 µg, or about 225 µg of the one or more nucleic acids (e.g., one or more mRNAs).

18. The composition of any of claims 10-17, wherein the composition further comprises a lipid nanoparticle (LNP), optionally wherein: (a) any one or more nucleic acids are encapsulated in the LNP; (b) any two or more nucleic acids are co-encapsulated in a single LNP; and / or (c) any two or more nucleic acids are encapsulated in separate LNPs.

19. The composition of claim 18, wherein the LNP comprises at least one cationic lipid, and wherein the cationic lipid is selected from the group consisting of OF-02, cKK-E10, IM-001, IS-001, and GL-HEPES-E3-E12-DS-4-E10, preferably wherein the cationic lipid is GL-HEPES-E3-E12-DS-4-E10.

20. The composition of claim 18 or 19, wherein the LNP further comprises a polyethylene glycol (PEG)-conjugated (PEGylated) lipid, a cholesterol-based lipid, and a helper lipid.

21. The composition of any one of claims 18-20, wherein the LNP comprises GL-HEPES-E3-E12-DS-4-E10 at a molar ratio of 35% to 55%; DMG-PEG2000 at a molar ratio of 0.25% to 2.75%; cholesterol at a molar ratio of 20% to 50%; and DOPE at a molar ratio of 5% to 35%.

22. The composition of any one of claims 18-21, wherein the LNP comprises GL-HEPES-E3-E12-DS-4-E10 at a molar ratio of 40%, DMG-PEG2000 at a molar ratio of 1.5%, cholesterol at a molar ratio of 28.5%, and DOPE at a molar ratio of 30%.

23. A composition, optionally an immunogenic composition, comprising: (a) a modified C. acnes CAMP2 polypeptide as defined in any one of claims 7-9; or (b) a C. acnes CAMP2 polypeptide and one or more of the following: (i) a C. acnes DsAl polypeptide, (ii) a C. acnes DsA2 polypeptide, (iii) a C. acnes PITP polypeptide, (iv) a chimeric C. acnes DsAl / DsA2 polypeptide, and (v) a chimeric C. acnes DsAl / DsA2 / PITP polypeptide.

24. The composition of claim 23, wherein the composition is as defined in claim 23(a), and wherein the composition further comprises one or more of the following: (A) a C. acnes DsAl polypeptide, (B) a C. acnes DsA2 polypeptide, (C) a C. acnes PITP polypeptide, (D) a chimeric C. acnes DsAl / DsA2 polypeptide, and (E) a chimeric C. acnes DsAl / DsA2 / PITP polypeptide.

25. The composition of any one of claims 10-24, wherein the C. acnes CAMP2 polypeptide, the C. acnes DsAl polypeptide, the C. acnes DsA2 polypeptide, the C. acnes PITP polypeptide, the chimeric C. acnes DsAl / DsA2 polypeptide, and the chimeric C. acnes DsAl / DsA2 / PITP polypeptide further comprise a non-native transmembrane domain sequence (e.g., a viral transmembrane domain sequence) and / or a non-native secretion signal peptide sequence.

26. The composition of any one of claims 10-25, wherein: (a) one or more of the C. acnes CAMP2 polypeptide, the C. acnes DsAl polypeptide, the C. acnes DsA2 polypeptide, the C. acnes PITP polypeptide, the chimeric C. acnes DsAl / DsA2 polypeptide, and the chimeric C. acnes DsAl / DsA2 / PITP polypeptide comprises a mutation at one or more (e.g., all) positions corresponding to an N-glycosylation site and / or one or more (e.g., all) positions corresponding to an O-glycosylation site in the corresponding native C. acnes polypeptide, optionally wherein the mutation is a single amino acid substitution; and / or (b) one or more of the C. acnes DsAl polypeptide, the C. acnes DsA2 polypeptide, the C. acnes PITP polypeptide, the chimeric C. acnes DsAl / DsA2 polypeptide, and the chimeric C. acnes DsAl / DsA2 / PITP polypeptide comprises a single amino acid substitution at one or more (e.g., all) positions corresponding to a cysteine residue in the corresponding native C. acnes polypeptide, optionally wherein the single amino acid substitution is a substitution of the cysteine with a serine.

27. The composition of any one of claims 10-26, wherein: (a) the C. acnes CAMP2 polypeptide comprises a sequence according to any one of SEQ ID NO: 203, SEQ ID NO: 43-58, SEQ ID NO: 1-4, SEQ ID NO: 10-16, or SEQ ID NO: 339-363 (e.g., SEQ ID NO: 203) or a sequence having at least 60% (e.g., at least 85%, at least 90%, at least 95%) identity thereto; (b) the C. acnes DsAl polypeptide comprises a sequence according to any one of SEQ ID NO: 204 or SEQ ID NO: 17-19 or SEQ ID NO: 59-61 or a sequence having at least 75% identity thereto; (c) the C. acnes DsA2 polypeptide comprises a sequence according to any one of SEQ ID NO: 205 or SEQ ID NO: 20-27 or SEQ ID NO: 62-69 or a sequence having at least 75% identity thereto; (d) the C. acnes PITP polypeptide comprises a sequence according to any one of SEQ ID NO: 206, SEQ ID NO: 31-37, SEQ ID NO: 73-79 or a sequence having at least 75% (e.g., 90% or 95%) identity thereto; (e) the chimeric C. acnes DsAl / DsA2 polypeptide comprises a sequence according to any one of SEQ ID NOs: 28-30, SEQ ID NO: 39, or SEQ ID NOs: 70-72, or SEQ ID NO: 81, or a sequence with at least 90% identity thereto; and / or (f) the chimeric C. acnes DsAl / DsA2 / PITP polypeptide comprises (i) a sequence according to any one of SEQ ID NOs: 28-30, SEQ ID NO: 39, or SEQ ID NOs: 70-72, or SEQ ID NO: 81, or a sequence with at least 90% identity thereto; and (ii) a PITP polypeptide sequence, optionally wherein the chimeric C. acnes DsAl / DsA2 / PITP polypeptide comprises a sequence according to any one of SEQ ID NO: 38, SEQ ID NO: 40-41, SEQ ID NO: 80, SEQ ID NO: 82-83, SEQ ID NO: 367-368, or a sequence with at least 90% identity thereto.

28. The composition of any one of claims 10-27, wherein the composition comprises: (a) a C. acnes CAMP2 polypeptide comprising or consisting of (e.g., consisting of) a sequence according to SEQ ID NO: 203; (b) a chimeric C. acnes DsAl / DsA2 polypeptide comprising or consisting of (e.g., consisting of) a sequence according to SEQ ID NO: 70; and (c) a C. acnes PITP polypeptide comprising or consisting of (e.g., consisting of) a sequence according to SEQ ID NO:

73.

29. The composition of any one of claims 23-28, wherein the composition further comprises an adjuvant, optionally wherein the adjuvant is selected from the group consisting of an aluminum-based adjuvant (e.g., AIOOH), a squalene-based oil-in-water emulsion adjuvant (e.g., AF03, AS03, MF59), and a liposome-based adjuvant comprising a saponin and a TLR4 agonist (e.g., SPA14, AS01, LEQ), further optionally wherein the adjuvant is selected from the group consisting of AIOOH, AF03, and SPA14.

30. A nucleic acid comprising a nucleotide sequence encoding a chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide, wherein the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises: (a) a chimeric C. acnes DsAl / DsA2 polypeptide; (b) an immunogenic fragment of a C. acnes PITP polypeptide, optionally wherein the immunogenic fragment comprises an ENFD of a C. acnes PITP polypeptide; and (c) a C. acnes CAMP2 polypeptide or an immunogenic fragment thereof.

31. A chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprising: (a) a chimeric C. acnes DsAl / DsA2 polypeptide; (b) an immunogenic fragment of a C. acnes PITP polypeptide, optionally wherein the immunogenic fragment comprises an ENFD of a C. acnes PITP polypeptide; and (c) a C. acnes CAMP2 polypeptide or an immunogenic fragment thereof.

32. The nucleic acid of claim 30 or the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide of claim 31, wherein i. the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a CSD1 of a C. acnes DsAl polypeptide, a CSD2 of a C. acnes DsA2 polypeptide, and a CSD3 of a C. acnes DsAl polypeptide, optionally wherein the chimeric C. acnes DsAl / DsA2 polypeptide of (a) comprises a sequence according to SEQ ID NO: 70 or a sequence having at least 90% (e.g., at least 95%) identity thereto; ii. the immunogenic fragment of a C. acnes PITP polypeptide in (b) comprises a sequence corresponding to amino acid residues 1-133 or residues 1-146 (e.g., residues 1-146) of SEQ ID NO: 73 or a sequence having at least 90% identity thereto; and / or iii. (c) is (1) a C. acnes CAMP2 polypeptide, optionally wherein (c) comprises SEQ ID NO: 203 or a sequence having at least 90% identity thereto; or (2) an immunogenic fragment of a C. acnes CAMP2 polypeptide comprising an N-terminal domain of a C. acnes CAMP2 polypeptide, e.g., wherein the immunogenic fragment comprises amino acid residues 29-176 of SEQ ID NO: 202 or a sequence having at least 90% identity to the sequence of amino acid residues 29-176 of SEQ ID NO:

202.

33. The nucleic acid of claim 30 or 32 or the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide of claim 31 or 32, wherein the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a transmembrane domain sequence, optionally wherein the transmembrane domain sequence comprises an amino acid sequence according to any one of the sequences in Table 4 or SEQ ID NO: 84 (e.g., SEQ ID NO: 208 or 84).

34. The nucleic acid of claim 30, 32, or 33 or the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide of claims 31-33, wherein: (i) the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 374-375; or (ii) the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 373 and a transmembrane domain sequence (e.g., a transmembrane sequence according to SEQ ID NO: 208 or 84), for example, wherein the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 374, or wherein the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 373 and a TMB sequence according to SEQ ID NO:

84.

35. The nucleic acid of claim 30 or 32-34, wherein: (i) the nucleotide sequence encoding the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to any one of SEQ ID NO: 377-382 or 384-389 (e.g., SEQ ID NO: 384-389); or (ii) the nucleotide sequence encoding the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 384 and a sequence according to SEQ ID NO: 395; or a sequence according to SEQ ID NO: 385 and a sequence according to SEQ ID NO: 396, for example, wherein the nucleotide sequence encoding the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 387, or the nucleotide sequence encoding the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide comprises a sequence according to SEQ ID NO: 385 and a sequence according to SEQ ID NO:

396.

36. A composition, optionally an immunogenic composition, comprising the nucleic acid (e.g., mRNA) of any one of claims 30 or 32-35, optionally wherein the composition further comprises a lipid nanoparticle (LNP), further optionally wherein the nucleic acid (e.g., mRNA) is encapsulated in the LNP.

37. The composition of claim 36, wherein: (a) the mRNA comprises a 5’ cap, at least one 5’ untranslated region (5’ UTR), at least one 3’ untranslated region (3’ UTR), and / or at least one polyadenylation (poly(A)) sequence; (b) the mRNA is unmodified or comprises at least one chemical modification, optionally wherein the mRNA comprises at least one chemical modification, e.g., wherein the chemical modification comprises Nl-methyl pseudouridine; and / or (c) the mRNA is a non-replicating mRNA.

38. The composition of claim 36 or 37, wherein: (i) the LNP comprises at least one cationic lipid, and wherein the cationic lipid is selected from the group consisting of OF-02, cKK-E10, IM-001, IS-001, and GL-HEPES-E3-E12-DS-4-E10 (e.g., wherein the cationic lipid is GL-HEPES-E3-E12-DS-4-E10); and / or (ii) the LNP comprises a polyethylene glycol (PEG)-conjugated (PEGylated) lipid, a cholesterol-based lipid, and a helper lipid, e.g., wherein the LNP comprises GL-HEPES-E3-E12-DS-4-E10 at a molar ratio of 35% to 55%; DMG-PEG2000 at a molar ratio of 0.25% to 2.75%; cholesterol at a molar ratio of 20% to 50%; and DOPE at a molar ratio of 5% to 35% (e.g., wherein the LNP comprises GL-HEPES-E3-E12-DS-4-E10 at a molar ratio of 40%, DMG-PEG2000 at a molar ratio of 1.5%, cholesterol at a molar ratio of 28.5%, and DOPE at a molar ratio of 30%).

39. A composition, optionally an immunogenic composition, comprising the chimeric C. acnes DsAl / DsA2 / PITP / CAMP2 polypeptide of any one of claims 31-34.

40. A nucleic acid of any one of claims 1-6, 30, or 32-35, a polypeptide of any one of claims 7-9 or 31-34, or a composition of any one of claims 10-29 or 36-39, for use as a medicament.

41. A nucleic acid of any one of claims 1-6, 30, or 32-35, a polypeptide of any one of claims 7-9 or 31-34, or a composition of any one of claims 10-29 or 36-39, for use in a method of treating or preventing a C. acnes infection in a subject.

42. A nucleic acid of any one of claims 1-6, 30, or 32-35, a polypeptide of any one of claims 7-9 or 31-34, or a composition of any one of claims 10-29 or 36-39, for use in a method of treating or preventing acne in a subject, wherein the acne is mild, moderate, or severe (e.g., moderate or severe).

43. The nucleic acid, polypeptide, or composition for use of any one of claims 40-42, wherein the method comprises: (i) administering one dose of the nucleic acid, the polypeptide, or the composition; or (ii) administering two doses of the nucleic acid, the polypeptide, or the composition, optionally wherein the two doses are administered two months apart.

44. The nucleic acid, polypeptide, or composition for use of any one of claims 41-43, wherein the subject is a human subject, and wherein the subject is between 9 and 45 years old or between 12 and 45 years old (e.g., between 9 and 18 years old or between 18 and 45 years old).

45. The nucleic acid, polypeptide, or composition for use of any one of claims 40-44, wherein the method comprises intramuscular or injection through the skin, e.g., intradermal, administration of the nucleic acid, the polypeptide, or the composition, e.g., wherein the composition is administered intramuscularly.

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