Vaccines against klebsiella pneumoniae

Abstract

The present invention relates to novel immunogenic compounds comprising at least one antigen of formula (I), in particular of formula (II), and their use as medicaments, in particular as vaccines. The invention also relates to related aspects comprising intermediates and methods of making these immunogenic compounds. Furthermore, the present invention relates to pharmaceutical compositions comprising an immunogenic compound and to the use of the antigen of formula (I) in a biological assay. # imgabs0 #

Description

[0001] This invention was made with government support under IDSEP160030 - 01, IDSEP160030 - 02, and IDSEP160030 - 03 awarded by HHS / ASPR. The United States government has certain rights in this invention. Technical Field

[0002] The present invention relates to novel immunogenic compounds comprising at least one antigen of formula (I), in particular immunogenic compounds of formula (II), and their use as medicaments, in particular as vaccines. The present invention also relates to related aspects involving intermediates and methods for preparing immunogenic compounds. In addition, the present invention relates to pharmaceutical compositions comprising the immunogenic compounds and the use of the antigen of formula (I) in biological assays. Background Art

[0003] Klebsiella pneumoniae (K. pneumoniae) is a Gram - negative, facultatively anaerobic, rod - shaped bacterium that mainly colonizes the respiratory tract, intestine, urinary tract, and skin and causes Klebsiella pneumoniae infections (KPI). This bacterium mainly acts as an opportunistic pathogen. KPI is a major cause of nosocomial infections, which mainly affect immunocompromised patients. Due to the emergence of strains resistant to almost all available antimicrobials and their worldwide spread, infections caused by Klebsiella pneumoniae are an important challenge in the healthcare environment. Infections caused by Klebsiella pneumoniae can result in high morbidity and mortality. Therefore, it is highly desirable to prevent infections caused by Klebsiella pneumoniae, and vaccination is the most cost - effective and powerful means against KPI.

[0004] Klebsiella pneumoniae is a capsulated bacterium that expresses lipopolysaccharide (LPS) and capsular polysaccharide (CPS, K - antigen) on its outer membrane, which contribute to the virulence of the species.

[0005] LPS consists of three components, namely the lipid A part that serves as the membrane anchor, the core oligosaccharide covalently bound to lipid A, and the terminal antigen polysaccharide covalently bound to the core oligosaccharide and comprising repeating sugar units that form the O - antigen. It has been demonstrated that extracted LPS is pyrogenic, toxic, and capable of causing tissue damage. LPS can be masked by CPS and is usually less exposed on the surface than CPS.

[0006] CPS consists of repeating sugar units that form a layer on the outer surface of the bacterium. CPS is usually complex, linear or branched, and has a larger molecular weight than LPS. Its high immunogenicity and surface exposure make it an interesting target for vaccination strategies. For example, WO2016156338 discloses conjugates of synthetic oligosaccharides related to the CPS of carbapenem - resistant Klebsiella pneumoniae.

[0007] However, the variability of Klebsiella CPS is relatively high. Over 77 different CPS types (so-called K-types, K-serotypes or K-antigens) have been serologically identified, but there are at least 141 K-types. These additional K-types are identified based on the cps-locus or K-locus and are called the KL series.

[0008] On the other hand, the variability of LPS is lower and the currently known so-called O-types, O-serotypes or O-antigens are limited to the following 11 main groups: O1, O2a (previously called Gal-I), O2ac, O2afg (previously called Gal-III), O2aeh (previously O9), O3 (including sub-serotypes O3, O3a and O3b), O4, O5, O7, O8 and O12. In addition to the above types, 11 additional O-types have been reported based on the O-locus called the OL series. Although the immunogenicity of O-antigens is lower than that of K-antigens and they are less exposed on the membrane surface, they are also considered for use in vaccination strategies. In a recent survey, the relative prevalence of CPS and LPS serotypes was determined in a large number of clinical isolates, especially multi-drug resistant isolates (e.g., Lam et al., Microbial Genomics 2022; 8:000800; DOI 10.1099 / mgen.0.000800).

[0009] O-type O2afg is produced by many multi-drug resistant Klebsiella pneumoniae strains (e.g., ST258). These strains are spread globally and are highly drug resistant (e.g., carbapenem resistance). The repeat unit structure of O2afg is, for example, disclosed in Kelly et al., J. of Bacteriol., 178(17), 1996, 5205-5214:

[0010] O-type O2a is one of the five most common O-antigens in clinical isolates. Antibiotic resistance is also common in O2a. The repeat unit structure of O2a is, for example, disclosed in Kelly et al., J. of Bacteriol., 178(17), 1996, 5205-5214:

[0011] The O2afg serotype is not recognized by O2a-specific antibodies and vice versa (Szijarto et al., Int J Med Microbiol 2016; 306(2):89-98; PMID:26723873; http: / / dx.doi.org / 10.1016 / j.ijmm.2015.12.002).

[0012] It is well known that pure, isolated bacterial polysaccharides are thymus-independent antigens that activate B cells without T cell help. Thus, the immune response to carbohydrates is a primary immune response, in which antibodies consist mainly of low-affinity IgG, without affinity maturation / isotype switching and with IgG being less robust and short-lived. Typical thymus-independent antigens are, for example, CPS from Streptococcus pneumoniae, Haemophilus influenzae type b.

[0013] To overcome the limitations of thymus-independent antigens, the O-antigen of CPS or LPS can be covalently linked to a carrier protein. Immunization with these polysaccharide-protein conjugates (glycoconjugate vaccines) induces T cell-dependent B cell activation and can even induce long-lasting immunity in infants. For example, WO2019106201 discloses conjugates of synthetic oligosaccharides related to Klebsiella pneumoniae serotypes O1, O2, O2ac and O8 O-polysaccharides, as well as the O-polysaccharide of carbapenem-resistant Klebsiella pneumoniae ST258.

[0014] The underlying mechanism of how conjugate vaccines are presented to T cells is still under debate. The polysaccharide-protein conjugate can be recognized and internalized by the polysaccharide-specific B cell receptor (BCR) of follicular B cells. The protein part of the glycoconjugate is processed and presented on MHC-II molecules on the cell surface of B cells. The MHC-II-peptide complex is recognized by peptide-specific T cells, which then initiate cognate T cell / B cell interactions, in which the B cell receives activation signals from the T cell. Recent studies have shown that, after binding to the BCR and subsequent uptake into endosomes, the glycoconjugate can also be processed into glycopeptide fragments. The peptide part can then bind to MHC-II molecules, while the carbohydrate (glycan) is exposed to the T cell receptor, where it can interact with carbohydrate-specific CD4+ T cells.

[0015] In the case where two different synthetic glycan antigens (e.g., glycan "A" and glycan "B") are covalently linked to the same carrier protein molecule, the number of existing glycan "A" and glycan "B"-specific B cells and the affinity of the BCRs expressed on these B cells determine the generation of glycan-specific antibodies (Abs). If, for example, the number of B cells specific for glycan "A" is low and low-affinity BCRs are also expressed on the cell surface, the Ab response induced after vaccination will be dominated by glycan "B"-specific Abs. Currently, the emergence and strength of the immunodominance of glycoconjugates cannot be predicted.

[0016] Whether CPS or LPS, or which of them, is a suitable candidate or model sequence for a vaccine remains difficult and unpredictable, and in particular it is unpredictable whether or which shorter oligosaccharides are suitable for generating the desired immune response in vivo. In particular, the immunodominance of glycoconjugates remains an unpredictable issue.

[0017] To date, there is no approved vaccine available for Klebsiella pneumoniae, which clearly demonstrates the challenges associated with the development of these vaccines. Summary of the Invention

[0018] It has now surprisingly been found that the novel oligosaccharide-antigens have improved properties as potential vaccines against Klebsiella pneumoniae. Due to their specific non-natural hybrid structures that target multiple serotypes, they are able to reduce the amount of carrier protein per vaccination, thereby reducing undesired carrier-induced epitope suppression and at the same time reducing manufacturing costs. Brief Description of the Drawings

[0019] Hereinafter, the terms "Gal-I" and "O2a" are considered synonymous, and the terms "Gal-III" and "O2afg" are also considered synonymous. Figure 1 : Compared with CRM 197 vs D13-CRM 197 *, D17-CRM 197 * and D20-CRM 197 * HPLC-SEC characterization of glycoconjugates. Figure 2 : Compared with CRM 197 and marker (protein size marker is GelCode TM Blue Safe Protein Stain (ThermoScientific)) vs D13-CRM 197 *, D17-CRM 197 * and D20-CRM 197 * SDS-PAGE of glycoconjugates. Figure 3 : Showing immunogenicity tests in ZiKa rabbits (6 rabbits per group) using a 2 μg antigen dose / rabbit / immunization on days 0, 14 and 28 (D13-CRM 197 *) or on days 0, 14 and 34 (D17-CRM 197 * and D20-CRM 197 *); i.e. Figure 3 A shows ELISA against Gal-III LPS isolated from the PCM27 strain (Polish Collection of Microorganisms) using an LPS extraction kit (JH Science);Figure 3 Panel B shows an ELISA for O2a LPS isolated from strain NCTC 9148 using an LPS extraction kit (JH Science); sera were diluted 1:100. Figure 4 : showing immunogenicity testing in rabbits (6 rabbits per group) using a 2 μg antigen dose / rabbit / immunization at days 0, 14, and 28 (D13-CRM 197 *) or at days 0, 14, and 34 (D17-CRM 197 * and D20-CRM 197 *); i.e., Figure 4 Panel A shows an ELISA for inactivated PCM27 bacteria (Gal-III); Figure 4 Panel B shows an ELISA for inactivated NCTC 9148 (O2a) bacteria; sera were diluted 1:500 (Figure A) or 1:100 (Figure B). Figure 5 : showing survival data from a challenge experiment in mice. CD-1 mice received two intraperitoneal injections of rabbit anti-serum generated using D13-CRM 197 *(immunized by using a 2 μg D13-CRM197* antigen dose / rabbit / immunization at days 0, 14, and 28 and collected at day 35) ( Figure 5 A and Figure 5 B) or using D17-CRM 197 * or D20-CRM 197 *( Figure 5 C) (immunized by using a 2 μg D17-CRM197* or D20-CRM197* antigen dose / rabbit / immunization at days 0, 14, and 34 and collected at day 41) at -24 h and -1 h relative to untreated rabbit infection or control sera. At 0 h, mice were intraperitoneally infected with a lethal dose of Klebsiella pneumoniae O2a-expressing strain NCTC 9163 ( Figure 5 A) or O2afg-expressing strain ST258 ( Figure 5 B and Figure 5 C) along with galactosamine treatment (20 mg / mouse intraperitoneally). Mice were observed for 24 h survival. Indicated survival curves were significantly different statistically (log-rank (Mantel-Cox) test) with indicated P values. The number of mice per group was: 8 ( Figure 5 A and Figure 5 C) or 10 ( Figure 5 B). Detailed Description

[0020] 1) In a first aspect, the present invention relates to an immunogenic compound comprising at least one oligosaccharide hybrid antigen of formula (I) wherein R is OH or m is 3, 4, 5, 6, 7 or 8; and n is 1, 2, 3, 4, 5 or 6; or a pharmaceutically acceptable salt thereof.

[0021] "**" specified in the dotted line refers to the point of attachment. It means that at this position, the antigen is linked to the carrier protein via a linker and / or a spacer. The oligosaccharide hybrid antigen of formula (I) is responsible for immunogenic selectivity, i.e., responsible for the targeted (specific) antibody response against multiple O-serotypes of Klebsiella pneumoniae, particularly O2a and O2afg.

[0022] The phrase "at least one antigen" means that the immunogenic compound may comprise one or more antigens of formula (I). For example, the immunogenic compound comprises from 1 to 28 antigens of formula (I).

[0023] In certain embodiments, the immunogenic compound may comprise a mixture of different antigens of formula (I). Preferred immunogenic compounds are those having a uniform antigen of formula (I), i.e., those having only one specific type of antigen of formula (I).

[0024] The term "hybrid" means that the antigen comprises two different parts of the O-serotype (i.e., O2afg (Gal-III) and O2a (Gal-I)), wherein the O2afg part is distal from the point of attachment "**", and O2a is close to the point of attachment "**".

[0025] The oligosaccharide of the present invention is composed of galactan, i.e., -β-D-galactofuranose / β-D-Galf: The dotted line shows the points of attachment, i.e., C1 and C3 -α-D-galactopyranose / α-D-Galp: The dotted line shows the points of attachment, i.e., C1 and C3 Unless a definition that is clearly set forth otherwise provides a broader or narrower definition, the definitions provided herein are intended to apply uniformly to compounds of formula (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV) as defined in any one of embodiments 1) to 81) and (as appropriately modified) throughout the specification and claims. It is to be fully understood that a definition or preferred definition of a term is defined independently of (and in combination with) any definition or preferred definition of any one or all other terms as defined herein and may replace the corresponding term.

[0026] The oligosaccharide moieties (i.e., antigens or epitopes) of the compounds of formula (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV) are composed of D-galactopyranoside and D-galactofuranoside, respectively. The configuration at each anomeric center is α or β. The configuration at the anomeric center can give rise to an anomeric mixture, whereby the anomers are synthesized in the α or β form, preferably in the pure α or β anomeric form. The anomeric mixture can be separated in a manner known to those skilled in the art.

[0027] 2) Another embodiment relates to an immunogenic compound as in embodiment 1) or a pharmaceutically acceptable salt thereof, wherein m is 3, 4, 5 or 6, and n is 2, 3 or 4.

[0028] 3) Another embodiment relates to an immunogenic compound as in embodiment 1) or a pharmaceutically acceptable salt thereof, wherein m is 3, 4, 5 or 6, and n is 2 or 3, for example 2.

[0029] 4) Another embodiment relates to an immunogenic compound as in any one of embodiments 1), 2) or 3) or a pharmaceutically acceptable salt thereof, wherein R is OH.

[0030] 5) Another embodiment relates to an immunogenic compound as in embodiment 1) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is m is 4, n is 3 and R is OH.

[0031] 6) Another embodiment relates to an immunogenic compound as in embodiment 1) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH.

[0032] 7) Another embodiment relates to an immunogenic compound as in any one of embodiments 1), 2), 3), 4), 5) or 6) or a pharmaceutically acceptable salt thereof, wherein the immunogenic compound further comprises a carrier protein. The carrier protein is preferably non-toxic and suitable for inducing immunogenicity. Thus, the carrier protein is preferably a non-toxic carrier protein suitable for inducing immunogenicity.

[0033] 8) Another embodiment relates to an immunogenic compound as in any one of embodiments 1), 2), 3), 4), 5) or 6) or a pharmaceutically acceptable salt thereof, wherein the immunogenic compound further comprises a carrier protein selected from the group consisting of:

[0034] CRM 197 , diphtheria toxoid, tetanus toxoid, cholera toxin B subunit, outer membrane protein (OMP) of Neisseria meningitidis, capsid protein of bacteriophage Qβ, oligomers or virus-like particles prepared using the capsid protein of bacteriophage Qβ, detoxified exotoxin A (EPA) of Pseudomonas aeruginosa, maltose binding protein (MBP), Hc fragment of tetanus toxin (TetHc), detoxified hemolysin A of Staphylococcus aureus, clumping factor A (ClfA) and clumping factor B (ClfB) of Staphylococcus aureus, Escherichia coli FimH, Escherichia coli FimHC, detoxified variants of Escherichia coli heat-labile enterotoxin, detoxified variants of cholera toxin, Escherichia coli Sat protein, effector domain of Escherichia coli Sat protein, detoxified variants of pneumolysin of Streptococcus pneumoniae, Campylobacter jejuni AcrA, Pseudomonas PcrV protein, Campylobacter jejuni native glycoprotein, bovine serum albumin (BSA), fimbrial protein GBS80 from Streptococcus agalactiae, Escherichia coli heat-labile enterotoxin, tetanus toxin, cholera toxin and pneumolysin of Streptococcus pneumoniae.

[0035] The term "CRM 197 " refers to cross-reactive material 197, which is a non-toxic mutant form of diphtheria toxin, wherein a single amino acid exchange of glycine (Gly, G) for glutamic acid (Glu, E) at position 52 renders the protein non-toxic. It is described in more detail in embodiment 17).

[0036] The term "diphtheria toxoid" refers to the formalin-inactivated form of diphtheria toxin having SEQ ID NO:2 (Uniprot ID: P00587). The present invention encompasses proteins having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:2 (preferably having 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:2). Diphtheria toxoid can be prepared as described, for example, by Glenny et al. in Br J ExpPathol. 1923 Oct;4(5):283-8 (PMCID: PMC2047731).

[0037] The term "tetanus toxoid" refers to the formalin-inactivated form of tetanus toxin having SEQ ID NO:3 (Uniprot ID: P00587). The present invention encompasses proteins having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:3 (preferably having 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:3). Tetanus toxoid can be prepared as described, for example, by G. Ramon et al., CR SocBiol, 93(1925), pp. 508-509.

[0038] The term "cholera toxin B subunit" refers to the protein having SEQ ID NO:4 (Uniprot ID: P01556). The present invention encompasses proteins having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:4 (preferably having 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:4).

[0039] The term "Neisseria meningitidis outer membrane protein" (OMP) refers to the protein having SEQ ID NO:5 (Uniprot ID: Q51229). The present invention encompasses proteins having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:5 (preferably having 95%, 96%, 97%, 98%, 99% or 99.9% identity to the amino acid sequence SEQ ID NO:5).

[0040] The term "capsid protein of bacteriophage Qβ" refers to the protein having SEQ ID NO:6 (Uniprot ID: P01556). The present invention encompasses proteins having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.9% identity with the amino acid sequence SEQ ID NO:6 (preferably having 95%, 96%, 97%, 98%, 99% or 99.9% identity with the amino acid sequence SEQ ID NO:6).

[0041] 9) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 1), 2), 3), 4), 5) or 6), wherein the immunogenic compound further comprises a carrier protein selected from the group consisting of:

[0042] CRM 197 , diphtheria toxoid, tetanus toxoid, cholera toxin B subunit, Neisseria meningitidis outer membrane protein (OMP) and capsid protein of bacteriophage Qβ. The preferred carrier protein is CRM 197 .

[0043] 10) Another embodiment relates to an immunogenic compound as in any one of embodiments 7), 8) or 9), wherein the immunogenic compound further comprises a non-immunogenic linker and / or spacer that is covalently bonded to the antigen at the **-binding site on one side and covalently bonded to the carrier protein on the other side.

[0044] 11) In a second aspect, the present invention relates to an immunogenic compound of formula (Ia): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; and n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; i is at least 1, preferably a number from 1 to 90% of the count of lysine residues contained in the carrier protein CP; -L-T- represents a linker L and a spacer T that together form a bridge having a backbone of 5 to 25 atoms (covalently linked together), the length of which forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at the carrier protein CP, wherein the atoms of the backbone are selected from the group consisting of carbon, nitrogen, oxygen and sulfur; and CP is a carrier protein selected from the group consisting of: CRM 197, diphtheria toxoid, tetanus toxoid, cholera toxin B subunit, Neisseria meningitidis outer membrane protein (OMP), capsid protein of bacteriophage Qβ, oligomers or virus-like particles prepared using the capsid protein of bacteriophage Qβ, detoxified exotoxin A (EPA) of Pseudomonas aeruginosa, maltose-binding protein (MBP), Hc fragment of tetanus toxin (TetHc), detoxified hemolysin A of Staphylococcus aureus, Clumping factor A (ClfA) and Clumping factor B (ClfB) of Staphylococcus aureus, Escherichia coli FimH, Escherichia coli FimHC, detoxified variant of Escherichia coli heat-labile enterotoxin, detoxified variant of cholera toxin, Escherichia coli Sat protein, effector domain of Escherichia coli Sat protein, detoxified variant of pneumolysin of Streptococcus pneumoniae, Campylobacter jejuni AcrA, Pseudomonas aeruginosa PcrV protein, native glycoprotein of Campylobacter jejuni, bovine serum albumin (BSA), fimbrial protein GBS80 from group B streptococcus, Escherichia coli heat-labile enterotoxin, tetanus toxin, cholera toxin and pneumolysin of Streptococcus pneumoniae; or a pharmaceutically acceptable salt thereof.

[0045] Preferably, i is a number from 1 to 90% of the count of lysine residues contained in the carrier protein CP; more preferably, i is a number from 1 to 75% of the count of lysine residues contained in the carrier protein CP; even more preferably, i is a number from 1 to 40% of the count of lysine residues contained in the carrier protein CP. For example, if the carrier protein CP contains 39 lysine residues, the range "from 1 to a number corresponding to 40% of the count of lysine residues contained in the carrier protein CP" means that i is in the range of 1 to 16.

[0046] The definition of the carrier protein as disclosed in embodiments 8) and 17) is expressly referred to.

[0047] To avoid any doubt, throughout this application, the terms "lysine residue" and "lysine site" are used synonymously.

[0048] The immunogenic compound according to the invention is an oligosaccharide-carrier protein conjugate, and the term "immunogenic compound" is used synonymously with "oligosaccharide-carrier protein conjugate".

[0049] The immunogenic compound of embodiment 11) may alternatively be referred to as an oligosaccharide-carrier protein conjugate of formula (Ia).

[0050] The term "heterozygous" means that the antigen comprises two different parts of the O-serotype, namely O2afg (Gal-III) and O2a (Gal-I), where the O2afg part is far from the carrier protein and O2a is close to the carrier protein.

[0051] 12) Another embodiment relates to an immunogenic compound as in embodiment 11) or a pharmaceutically acceptable salt thereof, wherein R is OH.

[0052] 13) Another embodiment relates to an immunogenic compound as in embodiment 11) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is m is 4, n is 3 and R is OH.

[0053] 14) Another embodiment relates to an immunogenic compound as in embodiment 11) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH.

[0054] 15) Another embodiment relates to an immunogenic compound as in embodiment 11), 12), 13) or 14) or a pharmaceutically acceptable salt thereof, wherein CP is a carrier protein selected from the group consisting of:

[0055] CRM 197 , diphtheria toxoid, tetanus toxoid, cholera toxin B subunit, Neisseria meningitidis outer membrane protein (OMP), and capsid protein of bacteriophage Qβ (especially CRM 197 ).

[0056] 16) The immunogenic compound as in embodiment 11), 12), 13), 14) or 15) may have a linker-spacer-L-T- as disclosed in any one of embodiments 17) to 47). This means that the linker-spacer-L-T- of formula (Ia) is the same as the linker spacer-L-T- described in binding CRM 197 . Accordingly, the same descriptions and definitions (with appropriate modifications) apply to the carrier protein CP.

[0057] 17) In another aspect, the present invention relates to an immunogenic compound of formula (II) wherein R is OH or m is 3, 4, 5, 6, 7 or 8; n is 1, 2, 3, 4, 5 or 6; i is from 1 to 28; and -L-T- represents a linker L and a spacer T, which together form a bridge having a backbone of 5 to 25 atoms (covalently linked together), and this length forms the oxygen at the reducing end C1 of the oligosaccharide and the carrier protein CRM 197The shortest distance between the amino nitrogens of lysine residues, where the atoms of the backbone are selected from the group consisting of carbon, nitrogen, oxygen, and sulfur; or a pharmaceutically acceptable salt thereof.

[0058] It should be understood that the term "substantially" as used in terms such as "substantially pure" in the context of the present invention particularly means that at least 90% by weight, particularly at least 95% by weight, and especially at least 99% by weight of the corresponding immunogenic compound / oligosaccharide / oligosaccharide-conjugate compound / oligosaccharide-conjugate-spacer compound / glycoconjugate is composed of the corresponding pure immunogenic compound / oligosaccharide / oligosaccharide-conjugate compound / oligosaccharide-conjugate-spacer compound / glycoconjugate.

[0059] When a substituent is indicated as optional, it should be understood that the substituent may be absent (i.e., with respect to the optional substituent, the corresponding residue is unsubstituted), in which case all positions with free valences (to which the optional substituent may be attached; e.g., the ring carbon atoms and / or ring nitrogen atoms with free valences in an aromatic ring) are hydrogen-substituted as needed. Similarly, if the term "optionally" is used in the context of (ring) heteroatoms, the term means that the corresponding optional heteroatom or the like is absent (i.e., some moieties are heteroatom-free / carbocyclic / or the like), or the corresponding optional heteroatom or the like is present as clearly defined.

[0060] “CRM 197 ” refers to cross-reacting material 197, which is a non-toxic mutant form of diphtheria toxin, where a single amino acid exchange of glycine (Gly, G) with glutamic acid (Glu, E) at position 52 renders the protein non-toxic.

[0061] CRM 197 is produced by Corynebacterium diphtheriae infected with the non-toxigenic phage β197tox, which is generated by nitrosoguanidine mutagenesis of the toxigenic phage β (Uchida et al., J. Biol. Chem., 1973, Vol. 245, No. 11, pp. 3838 - 3844). CRM 197 protein is a safe and effective T cell-dependent carrier for sugars. CRM 197 For example, it is described by Giannini et al. in Nucleic Acids Research, Vol 12, No. 10, 1984, pp. 4063 - 4069. Regarding CRM 197 and further details of its production can be found, for example, in US5,614,382, which is incorporated herein by reference. CRM 197It can be produced in various expression systems (e.g., in Corynebacterium diphtheriae, Escherichia coli or Pseudomonas fluorescens) (Hickey et al., J. Pharm. Sci., 2018, 107, 1806 - 1819).

[0062] In the present invention, the term "CRM" 197 " encompasses proteins having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99%, 99.8% or 99.9% identity with the amino acid sequence SEQ ID NO:1 (preferably having at least 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98%, 99% or 99.9% identity with the amino acid sequence SEQ ID NO:1; and particularly having at least 95%, 96%, 97%, 98%, 99% or 99.9% identity with the amino acid sequence SEQ ID NO:1), which optionally includes an additional methionine (Met, M) at the N - terminus, and / or optionally contains residues generated by functionalizing CRM at lysine sites 197 which can be in a capped (i.e., inactivated) form.

[0063] The phrase "residues generated by functionalizing CRM at lysine sites" means that CRM is functionalized at lysine sites with functional groups suitable for forming covalent bonds with a linker and / or spacer moiety (i.e., the oligosaccharide - linker moiety of the conjugate) attached to an antigen 197 . Such lysine - functionalized CRM 197 is well - known to those skilled in the art. These functional groups are particularly suitable for attaching thiols or performing click chemistry. For example, these functional groups are groups containing bromoacetamide, iodoacetamide, maleimide, azide or alkyne groups. This means that CRM 197 optionally contains lysine residues functionalized with bromoacetamide, iodoacetamide, maleimide, azide or alkyne groups (preferably bromoacetamide, iodoacetamide, maleimide groups), which can be in a capped form. 197

[0064] Preferred functionalized CRM 197 contains groups carrying bromoacetamide, iodoacetamide or maleimide groups, all of which are suitable for reacting with thiol groups provided by the oligosaccharide / linker moiety. Unreacted functional groups in CRM 197 can subsequently be quenched using any pharmaceutically acceptable thiol (e.g., L - cysteine or cysteamine (2 - aminoethane - 1 - thiol)) to provide a "capped form".

[0065] ​Preferred lysine-functionalized CRM 197 is selected from the group consisting of: where Z is Br or I, q is 2 or 3, and t is from 1 to 28; where r is 2 or 3, and t' is from 1 to 28; and where Z is Br or I, and t'' is from 1 to 28.

[0066] In a preferred embodiment, CRM is not functionalized in the above manner 197 . This means that there is no "prefunctionalization", but rather the oligosaccharide / linker / spacer moiety is directly attached thereto using "natural" lysine residues (i.e., the unmodified amino groups of lysine residues).

[0067] CRM 197 The amino acid sequence of is known to those skilled in the art and is outlined in SEQ ID NO:1 below.

[0068] The use of CRM for the synthesis of conjugates and preferred binding sites on CRM 197 have been reported (e.g., 197 et al., Sci.Rep.6,20488; doi:10.1038 / srep20488(2016), which is incorporated herein by reference).

[0069] The phrase "-L-T- represents a linker L and a spacer T, which together form a bridge having a backbone of 5 to 25 atoms (covalently linked together), and this length forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at the carrier protein CRM 197 (or CP, if applicable), where the atoms of the backbone are selected from the group consisting of carbon, nitrogen, oxygen, and sulfur" means that the backbone can be saturated, unsaturated, unsubstituted, or substituted with one or more (especially 1, 2, 3, or 4) substituents independently selected from oxo, (C 1-4 )alkyl, fluorine, and (C 1-2 )alkoxy (especially oxo), and optionally a part of the ring structure can be part of the backbone. The ring structure can be a saturated, unsaturated, or aromatic 3- to 8-membered ring of a fused ring system containing 2 to 4 rings, where the ring atoms are selected from carbon, nitrogen, oxygen, and sulfur (especially selected from carbon and nitrogen), and the ring is unsubstituted or substituted with one or more (especially 1, 2, 3, or 4) substituents independently selected from oxo, (C 1-4 )alkyl, halogen, and (C 1-2 )alkoxy (especially oxo). ​

[0070] To avoid any doubt, the count of 5 to 25 atoms refers to the count of atoms in the main chain (not the bridge).

[0071] Examples of optional ring structures that can be part of the main chain are pyrrolidine-2,5-dione, cyclobut-3-ene-1,2-dione, triazole, isoindolin-1-one, 8,9-dihydro-1H-dibenzo[b,f][1,2,3]triazolo[4,5-d]azepine, cyclohexane, and benzene as follows: The introduction of these rings or ring systems is known to those skilled in the art of linker chemistry.

[0072] The phrase "the main chain may be unsaturated" means that the main chain may contain one or more double bonds that may or may not be part of a ring system.

[0073] For example, in a bridge having a saturated main chain containing 3 oxo-substituents and the main chain being part of a ring system, the atom count is as follows: Thus, the count of atoms forming the main chain starts with the first atom after the C1 oxygen and ends with the last atom of the lysine nitrogen attached to the CRM 197 of the lysine nitrogen.

[0074] Oxygen atoms in a saturated chain are preferably separated from another oxygen atom by one or more (especially 2, 3, 4, or 5, and especially 2) carbon atoms.

[0075] Sulfur atoms in a saturated chain are preferably separated from another sulfur atom by one or more (especially 1, 2, 3, 4, or 5) carbon atoms.

[0076] The term "halogen" means fluorine, chlorine, or bromine, preferably fluorine or chlorine, more preferably fluorine.

[0077] The term "oxo" refers to the functional group =O, i.e., a substituent oxygen atom linked to another atom (preferably a carbon atom) by a double bond.

[0078] The term "alkyl", when used alone or in combination, means a straight-chain or branched saturated hydrocarbon chain containing 1 to 4 carbon atoms. The term "(C x-y )alkyl" (where x and y are each integers) refers to an alkyl containing x to y carbon atoms as defined above. For example, (C 1-4 )alkyl contains 1 to 4 carbon atoms. Examples of (C 1-4 )alkyl are methyl, ethyl, n-propyl, isopropyl, n-butyl, isobutyl, sec-butyl, and tert-butyl. Examples of (C 1-2 )alkyl are methyl and ethyl.

[0079] The term "alkoxy", when used alone or in combination, refers to an alkyl-O-group, wherein the alkyl is as defined above. The term "(C x-y )alkoxy" (wherein x and y are each integers) refers to an alkoxy containing from x to y carbon atoms as defined previously. For example, (C 1-2 )alkoxy means a group of the formula (C 1-2 )alkyl-O-, wherein the term "(C 1-2 )alkyl" has the meaning given previously. Examples of (C 1-2 )alkoxy are methoxy and ethoxy.

[0080] As used herein, the term "oligosaccharide-carrier protein conjugate" is considered synonymous with the term "glycoconjugate". In addition, the "immunogenic compound" as described herein is an "oligosaccharide-carrier protein conjugate".

[0081] The term "hybrid" means that the antigen comprises two different portions of the O-serotype (i.e., O2afg (Gal-III) and O2a (Gal-I)), wherein the O2afg portion is remote from the carrier protein and the O2a is close to the carrier protein.

[0082] 18) Another embodiment relates to the immunogenic compound or a pharmaceutically acceptable salt thereof as in embodiment 17), wherein m is 3, 4, 5 or 6, and n is 2, 3 or 4.

[0083] 19) Another embodiment relates to the immunogenic compound or a pharmaceutically acceptable salt thereof as in embodiment 17), wherein m is 3, 4, 5 or 6, and n is 2 or 3, for example 2.

[0084] 20) Another embodiment relates to the immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 17), 18) or 19), wherein R is OH.

[0085] 21) Another embodiment relates to the immunogenic compound or a pharmaceutically acceptable salt thereof as in embodiment 17), wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is m is 4, n is 3 and R is OH.

[0086] 22) Another embodiment relates to the immunogenic compound or a pharmaceutically acceptable salt thereof as in embodiment 17), wherein m is 4, n is 2 and R is OH.

[0087] 23) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 11) to 15) and as in any one of embodiments 17), 18), 19), 20), 21) or 22), wherein the bridge does not contain an aromatic or heteroaromatic ring.

[0088] 24) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 11) to 15) and as in any one of embodiments 17), 18), 19), 20), 21) or 22), wherein

[0089] -L-T- represents a linker L and a spacer T which together form a bridge having a backbone of 5 to 25 atoms (covalently linked together), the length of which forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at CRM 197 (or CP, if applicable), and having at most one double bond,

[0090] wherein the atoms of the backbone are selected from the group consisting of carbon, nitrogen, oxygen and sulfur, and

[0091] wherein the backbone may be substituted with one or more (especially 1, 2, 3 or 4) substituents independently selected from oxo, (C 1-4 ) alkyl, fluorine and (C 1-2 ) alkoxy (especially oxo), and

[0092] wherein a part of the backbone may optionally be part of a 4-, 5- or 6-membered ring selected from the following: whereby, "at most one double bond" is preferably the double bond of a cyclobut - 3 - ene - 1,2 - dione ring.

[0093] 25) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 11) to 15) and as in any one of embodiments 17), 18), 19), 20), 21) or 22), wherein

[0094] -L-T- represents a linker L and a spacer T which together form a bridge consisting of a backbone which is a saturated chain counted as 5 to 25 atoms (selected from the group consisting of carbon, nitrogen, oxygen and sulfur (especially carbon, nitrogen and oxygen)), the chain may be unsubstituted or substituted with one or more (especially 1, 2, 3 or 4) substituents independently selected from oxo, (C 1-4 ) alkyl, fluorine and (C 1-2) substituted by substituents of alkoxy groups (especially oxo). This means that the bridge consists of a saturated chain counting from 5 to 25 atoms (selected from the group consisting of carbon, nitrogen, oxygen and sulfur (especially carbon, nitrogen and oxygen)), and the chain can be unsubstituted or substituted by one or more (especially 1, 2, 3 or 4) substituents independently selected from oxo, (C 1-4 ) alkyl, fluorine and (C 1-2 ) alkoxy (especially oxo). To avoid any doubt, in this embodiment, the bridge does not contain a ring structure.

[0095] 26) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as described in any one of embodiments 11) to 15) and any one of embodiments 17), 18), 19), 20), 21) or 22), wherein

[0096] -L-T- represents a linker L and a spacer T, which together form a bridge composed of a main chain, which is a saturated chain counting from 5 to 25 atoms (selected from the group consisting of carbon, nitrogen and oxygen (especially carbon and nitrogen)), and the chain can be unsubstituted or substituted by one or more (especially 1, 2, 3 or 4) substituents independently selected from oxo, (C 1-4 ) alkyl, fluorine and (C 1-2 ) alkoxy (especially oxo). This means that the bridge consists of a saturated chain counting from 5 to 25 atoms (selected from the group consisting of carbon, nitrogen and oxygen (especially carbon and nitrogen)), and the chain can be unsubstituted or substituted by one or more (especially 1, 2, 3 or 4) substituents independently selected from oxo, (C 1-4 ) alkyl, fluorine and (C 1-2 ) alkoxy (especially oxo). To avoid any doubt, in this embodiment, the bridge does not contain a ring structure.

[0097] 27) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as described in any one of embodiments 11) to 15) and any one of embodiments 17), 18), 19), 20), 21), 22), 23), 24), 25) or 26), wherein the main chain of the bridge has a length of 8 to 20, preferably 8 to 16 atoms (covalently linked together), which forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at the carrier protein CRM 197 (or CP, if applicable).

[0098] 28) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as described in any one of embodiments 11) to 15) and any one of embodiments 17), 18), 19), 20), 21), 22) or 27), wherein L represents *-(C2-10 ) alkylene-NH-; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 ) fluoroalkylene-NH-; *-(CH2) c NHC(O)(CH2) c’ -NH-, where c and c' are independently 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d' are independently 2 to 6; *-(C 1-10 ) alkylene-C(O)-NH-(C 2-10 ) alkylene-NH-; or *-(C 2-10 ) alkylene-O-NH-; T represents -C(O)-(C 0-10 ) alkylene-C(O)-; -C(O)-CH2CH2-(OCH2CH2) j -C(O)-, where j is 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)-, where k is 0 or 1, k' is 0 or 1, and k" is 1, 2 or 3; where l is 1 or 2; or where p is 1 to 4, and p' is 1 or 2; or L-T represents *-(C 2-10 ) alkylene-S-R 1 ; and R 1 represents where q is 2 or 3; where r is 2 or 3; or

[0099] The designated "*" in the linker L means that at this position the linker is attached to the oligosaccharide.

[0100] The designated "*" in the spacer T means that at this position the spacer is attached to the linker L.

[0101] R 1 The designated "#" in R 1 means that at this position R

[0102] The term "-(C x-y )alkylene-alk (where x and y are each integers) when used alone or in combination refers to a saturated straight-chain or branched hydrocarbon chain with a double bond containing from x to y carbon atoms. For example, (C 2-10 )alkylene contains from two to ten carbon atoms, and (C 0-10 )alkylene is a bond (i.e., absent, C is zero) or an alkylene of one to ten carbon atoms. The straight-chain -(C x-y )alkylene-, i.e., -(CH2) x-y - is preferred. Representative examples of (C 2-10 )alkylene are ethylene, propylene, butylene, pentylene, hexylene, heptylene, octylene, nonylene, and decylene (especially 1,2-ethylene, 1,3-propylene, 1,4-butylene, 1,5-pentylene, 1,6-hexylene, 1,7-heptylene, 1,8-octylene, 1,9-nonylene, and 1,10-decylene).

[0103] The term "(C x-y )fluoroalkylene" (where x and y are each integers) when used alone or in combination refers to a saturated straight-chain or branched hydrocarbon group with a double bond containing from x to y carbon atoms, where one or more (and possibly all) of the hydrogen atoms have been replaced by fluorine. The straight-chain -(C x-y )fluoroalkylene- is preferred.

[0104] To avoid any doubt, in this embodiment, the length of the backbone of -L-T- is 5 to 25 atoms, 8 to 20 atoms, or 8 to 16 atoms. This means that the linker L and the spacer T (including R 1 and where applicable) together form a bridge with a backbone of length 5 to 25 (8 to 20 or 8 to 16) atoms (covalently linked together), which length forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at the carrier protein CRM 197 (or CP, if applicable).

[0105] 29) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in embodiment 28), wherein L represents *-(CH2) a-NH-; wherein a is from 2 to 10; *-(CH2CH2O) b -CH2CH2NH-, wherein b is 1, 2 or 3; * -CH2CH2S-CH2CH2NH-; *-(C 2-10 ) fluoroalkylene-NH-, wherein the fluoroalkylene is a saturated straight chain; *-(CH2) c NHC(O)(CH2) c’ -NH-, wherein c and c' are independently of each other from 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, wherein d and d' are independently of each other from 2 to 6; *-(CH2) e -C(O)-NH-(CH2) e’ -NH-; wherein e is from 1 to 10 and e' is from 2 to 10; or *-(CH2) f -O-NH-, wherein f is from 2 to 10; or L-T represents *-(CH2) g -S-R 1 , wherein g is from 2 to 10.

[0106] 30) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2) a -NH-; wherein a is from 2 to 10; *-(CH2CH2O) b -CH2CH2NH-, wherein b is 1, 2 or 3; *-(C 2-10 ) fluoroalkylene-NH-, wherein the fluoroalkylene is a saturated straight chain; *-(CH2) e -C(O)-NH-(CH2) e’ -NH-; wherein e is from 1 to 10 and e' is from 2 to 10; or *-(CH2) f -O-NH-, wherein f is from 2 to 10; or L-T represents *-(CH2) g -S-R 1 , wherein g is from 2 to 10.

[0107] 31) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2) a -NH-; where a is from 2 to 10, preferably from 2 to 6; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3, preferably 1 or 2; *-(CH2) e -C(O)-NH-(CH2) e’ -NH-; where e is from 1 to 10, preferably from 1 to 6 and e' is from 2 to 10, preferably from 2 to 6; or *-(CH2) f -O-NH-, where f is from 2 to 10, preferably from 2 to 6; or L-T represents *-(CH2) g -S-R 1 , where g is from 2 to 10, preferably from 2 to 6.

[0108] 32) Another embodiment relates to an oligosaccharide-carrier protein conjugate as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2) a -NH-; where a is from 2 to 10, preferably from 2 to 6; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3, preferably 1 or 2; or *-(CH2) f -O-NH-, where f is from 2 to 10, preferably from 2 to 6.

[0109] 33) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2) a -NH-; where a is from 2 to 10, preferably from 2 to 6; or *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3, preferably 1 or 2;

[0110] 34) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2) a -NH-; where a is from 2 to 10, preferably from 2 to 6.

[0111] 35) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH-.

[0112] 36) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2)5-NH-.

[0113] 37) Another embodiment relates to an immunogenic compound as in embodiment 28) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 1; preferably 1.

[0114] 38) Another embodiment relates to an immunogenic compound as in any one of embodiments 28) to 37) or a pharmaceutically acceptable salt thereof, wherein T represents -C(O)-(CH2) h -C(O)-, where h is from 0 to 10; -C(O)-CH2CH2-(OCH2CH2) j -C(O)-, where j is from 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)-, where k is 0 or 1, k' is 0 or 1, and k'' is 1, 2 or 3; where l is 1 or 2; or where p is from 1 to 4, and p' is 1 or 2.

[0115] 39) Another embodiment relates to an immunogenic compound as in any one of embodiments 28) to 37) or a pharmaceutically acceptable salt thereof, wherein T represents -C(O)-(CH2) h -C(O)-, where h is from 0 to 10, preferably from 0 to 6; -C(O)-CH2CH2-(OCH2CH2) j -C(O)-, where j is from 1 to 5, preferably from 1 to 3, more preferably 1; -C(O)-CH2(CH2) k-(SCH2(CH2) k’ ) k” -C(O)-, where k is 0 or 1, k' is 0 or 1, and k'' is 1, 2, or 3, preferably 1; or

[0116] 40) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28) to 37), wherein T represents -C(O)-(CH2) h -C(O)-, where h is from 0 to 6; -C(O)-CH2CH2-(OCH2CH2) j -C(O)-, where j is 1 or 2; or -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)-, where k is 0 or 1, k' is 0 or 1, and k'' is 1, preferably where k and k' are 0 and k'' is 1.

[0117] 41) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28) to 37), wherein T represents -C(O)-(CH2) h -C(O)-, where h is 0, 1, 2, 3, 4, 5, or 6, preferably 4; or -C(O)-CH2CH2-(OCH2CH2) j -C(O)-, where j is 1 or 2.

[0118] 42) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28) to 37), wherein T represents -C(O)-(CH2) h -C(O)-, where h is 0, 1, 2, 3, 4, 5, or 6, preferably 4.

[0119] 43) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28) to 37), wherein T represents where l is 1 or 2; or where p is from 1 to 4, preferably 1, and p' is 1 or 2.

[0120] 44) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28) to 37), wherein T represents wherein l is 1 or 2.

[0121] 45) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28), 29), 30), 38), 43) or 44), wherein R 1 represents wherein q is 2 or 3; wherein r is 2 or 3; or Preferably, R 1 represents: wherein q is 2 or 3; or

[0122] 46) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 11), 12), 13), 14), 15), 17), 18), 19), 20), 21) or 22), wherein L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH-; and T represents -C(O)-C(O)-, -C(O)-CH2-C(O)-, -C(O)-(CH2)2-C(O)-, -C(O)-(CH2)3-C(O)-, -C(O)-(CH2)4-C(O)-, -C(O)-(CH2)5-C(O)- or -C(O)-(CH2)6-C(O)-.

[0123] 47) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 11), 12), 13), 14), 15), 17), 18), 19), 20), 21) or 22), wherein L represents *-(CH2)5-NH- and T represents -C(O)-(CH2)4-C(O)-.

[0124] 48) A preferred embodiment is an immunogenic compound having the following formula (II): wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is m is 4, n is 3 and R is OH. i is from 1 to 28; L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH-; and T represents -C(O)-C(O)-, -C(O)-CH2-C(O)-, -C(O)-(CH2)2-C(O)-, -C(O)-(CH2)3-C(O)-, -C(O)-(CH2)4-C(O)-, -C(O)-(CH2)5-C(O)- or -C(O)-(CH2)6-C(O)-; or a pharmaceutically acceptable salt thereof.

[0125] 49) Another embodiment relates to an immunogenic compound as in embodiment 48) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH; L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH- (preferably *-(CH2)5-NH-); T represents -C(O)-C(O)-, -C(O)-CH2-C(O)-, -C(O)-(CH2)2-C(O)-, -C(O)-(CH2)3-C(O)-, -C(O)-(CH2)4-C(O)-, -C(O)-(CH2)5-C(O)- or -C(O)-(CH2)6-C(O)- (preferably -C(O)-(CH2)4-C(O)-); and i is from 1 to 28; preferably from 6 to 15.

[0126] 50) Another embodiment relates to an immunogenic compound as in any one of embodiments 11) to 49) or a pharmaceutically acceptable salt thereof, wherein i is from 1 to 28, from 1 to 25, from 1 to 23, from 1 to 20, from 1 to 18, from 3 to 25, from 3 to 23, from 3 to 20, from 3 to 18, from 5 to 23, from 5 to 20, from 5 to 18, from 6 to 23, from 6 to 20, from 6 to 18, from 6 to 15. The variable i describes the loading of the antigen (i.e., the oligosaccharide conjugate) on the CRM 197 protein carrier and relates to an integer of a single molecule. However, when considering the glycoconjugate as a product of more than one single molecule, it should be noted that the loading can be described as a statistical distribution (i.e., a substantially Gaussian distribution). The chemical process that produces the product yields a molecular mixture having such a statistical distribution of the loading, and then the loading is provided in the form of the mean value of the statistical distribution, particularly the Gaussian distribution. It should be understood that for i ≥ 2, the m and / or n of two or more oligosaccharides respectively linked to the carrier protein CP or CRM via -L-T- 197 can be the same or different. Preferably, all i oligosaccharides are represented by the same combination of m and n (i.e., having the same structure) or all i oligosaccharides are represented by a first combination of m and n or a second combination of m and n (i.e., having one or the other structure); most preferably, all i oligosaccharides are represented by the same combination of m and n. The linker-spacer unit -L-T- is the same for the i oligosaccharides of a particular oligosaccharide-carrier protein conjugate. In other words, preferred compounds are those having uniform oligosaccharide / linker / spacer residues, i.e., those having only one particular type of oligosaccharide / linker / spacer residue linked to the carrier protein, preferably linked to CRM 197 .

[0127] 51) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 11) to 49), wherein i is from 6 to 15.

[0128] 52) A preferred embodiment relates to an immunogenic compound selected from the group consisting of: wherein i is from 1 to 28, or a pharmaceutically acceptable salt thereof. To avoid any doubt, the immunogenic compounds of formula (IIa), (IIb), and (IIc) of this embodiment can also be schematically drawn as follows: wherein i is from 1 to 28, or a pharmaceutically acceptable salt thereof. CRM 197 ’ means CRM as defined herein 197 , with the only difference being that in formulae (IIa’), (IIb’), and (IIc’), the amino group of the lysine residue is specifically shown as the attachment position of the linker / spacer moiety -L-T-. The preferred values of i are those disclosed in embodiment 50) or especially 51).

[0129] 53) Another preferred embodiment relates to an immunogenic compound of formula (IIa): wherein i is from 1 to 28, or a pharmaceutically acceptable salt thereof. Preferred values of i are those disclosed in embodiment 50) or especially 51).

[0130] 54) Another preferred embodiment relates to an immunogenic compound selected from the group consisting of: wherein i is from 6 to 15, or a pharmaceutically acceptable salt thereof.

[0131] 55) Another preferred embodiment relates to an immunogenic compound of formula (IIa): wherein i is from 6 to 15, or a pharmaceutically acceptable salt thereof.

[0132] Accordingly, the present invention relates to compounds of formula (I) as defined in embodiment 1) and these compounds further restricted by the characteristics of any one of embodiments 2) to 10), compounds of formula (Ia) as defined in embodiment 11) and these compounds further restricted by the characteristics of any one of embodiments 12) to 16), compounds of formula (II) as defined in embodiment 17) and these compounds further restricted by the characteristics of any one of embodiments 18) to 55) (taking into account the corresponding dependencies), their pharmaceutically acceptable salts; and the use of these compounds as further described below. In particular, the compounds of formula (Ia) and (II) are sub-formulas of formula (I), and the compounds of formula (III) and (IV) are intermediates for preparing the compounds of formula (I), (Ia) and (II), respectively. It should be understood that the embodiments relating to the L-T definition of the compounds of formula (II) as specified in embodiments 23) to 55) also apply to the L-T definition of the compounds of formula (Ia) in any one of embodiments 11) to 15), and vice versa.

[0133] To avoid any doubt, the following embodiments, especially those related to the compounds of formula (I) and (II), are thus possible and intended and hereby specifically disclosed in individualized form: 1、2+1、3+1、4+1、4+2+1、4+3+1、5+1、6+1、11、12+11、13+11、14+11、15+11、15+12+11、15+13+11、15+14+11、17、18+17、19+17、20+17、20+18+17、20+19+17、21+17、22+17、23+11、23+12+11、23+13+11、23+14+11、23+15+11、24+11、24+12+11、24+13+11、24+14+11、24+15+11、25+11、25+12+11、25+13+11、25+14+11、25+15+11、26+11、26+12+11、26+13+11、26+14+11、26+15+11、27+11、27+12+11、27+13+11、27+14+11、27+15+11、28+11、28+12+11、28+13+11、28+14+11、28+15+1129+28+17、29+28+18+17、29+28+19+17、29+28+20+17、29+28+21+17、29+28+22+17、29+28+27+17、30+28+17、30+28+18+17、30+28+19+17、30+28+20+17、30+28+21+17、30+28+22+17、30+28+27+17、31+28+17、31+28+18+17、31+28+19+17、31+28+20+17、31+28+21+17、31+28+22+17、31+28+27+17、32+28+17、32+28+18+17、32+28+19+17、32+28+20+17、32+28+21+17、32+28+22+17、32+28+27+17、33+28+17、33+28+18+17、33+28+19+17、33+28+20+17、33+28+21+17、33+28+22+17、33+28+27+17、34+28+17、34+28+18+17、34+28+19+17、34+28+20+17、34+28+21+17、34+28+22+17、34+28+27+17、35+28+17、35+28+18+17、35+28+19+17、35+28+20+17、35+28+21+17、35+28+22+17、35+28+27+17、36+28+17、36+28+18+17、36+28+19+17、36+28+20+17、36+28+21+17、36+28+22+17、36+28+27+17、37+28+17、37+28+18+17、37+28+19+17、37+28+20+17、37+28+21+17、37+28+22+17、37+28+27+17、38+28+17、38+28+18+17、38+28+19+17、38+28+20+17、38+28+21+17、38+28+22+17、38+28+27+17、38+29+28+17、38+29+28+18+17、38+29+28+19+17、38+29+28+20+17、38+29+28+21+17、38+29+28+22+17、38+29+28+27+17、38+30+28+17、38+30+28+18+17、38+30+28+19+17、38+30+28+20+17、38+30+28+21+17、38+30+28+22+17、38+30+28+27+17、38+31+28+17、38+31+28+18+17、38+31+28+19+17、38+31+28+20+17、38+31+28+21+17、38+31+28+22+17、38+31+28+27+17、38+32+28+17、38+32+28+18+17、38+32+28+19+17、38+32+28+20+17、38+32+28+21+17、38+32+28+22+17、38+32+28+27+17、38+33+28+17、38+33+28+18+17、38+33+28+19+17、38+33+28+20+17、38+33+28+21+17、38+33+28+22+17、38+33+28+27+17、38+34+28+17、38+34+28+18+17、38+34+28+19+17、38+34+28+20+17、38+34+28+21+17、38+34+28+22+17、38+34+28+27+17、38+35+28+17、38+35+28+18+17、38+35+28+19+17、38+35+28+20+17、38+35+28+21+17、38+35+28+22+17、38+35+28+27+17、38+36+28+17、38+36+28+18+17、38+36+28+19+17、38+36+28+20+17、38+36+28+21+17、38+36+28+22+17、38+36+28+27+17、38+37+28+17、38+37+28+18+17、38+37+28+19+17、38+37+28+20+17、38+37+28+21+17、38+37+28+22+17、38+37+28+27+17、39+28+17、39+28+18+17、39+28+19+17、39+28+20+17、39+28+21+17、39+28+22+17、39+28+27+17、39+29+28+17、39+29+28+18+17、39+29+28+19+17、39+29+28+20+17、39+29+28+21+17、39+29+28+22+17、39+29+28+27+17、39+30+28+17、39+30+28+18+17、39+30+28+19+17、39+30+28+20+17、39+30+28+21+17、39+30+28+22+17、39+30+28+27+17、39+31+28+17、39+31+28+18+17、39+31+28+19+17、39+31+28+20+17、39+31+28+21+17、39+31+28+22+17、39+31+28+27+17、39+32+28+17、39+32+28+18+17、39+32+28+19+17、39+32+28+20+17、39+32+28+21+17、39+32+28+22+17、39+32+28+27+17、39+33+28+17、39+33+28+18+17、39+33+28+19+17、39+33+28+20+17、39+33+28+21+17、39+33+28+22+17、39+33+28+27+17、39+34+28+17、39+34+28+18+17、39+34+28+19+17、39+34+28+20+17、39+34+28+21+17、39+34+28+22+17、39+34+28+27+17、39+35+28+17、39+35+28+18+17、39+35+28+19+17、39+35+28+20+17、39+35+28+21+17、39+35+28+22+17、39+35+28+27+17、39+36+28+17、39+36+28+18+17、39+36+28+19+17、39+36+28+20+17、39+36+28+21+17、39+36+28+22+17、39+36+28+27+17、39+37+28+17、39+37+28+18+17、39+37+28+19+17、39+37+28+20+17、39+37+28+21+17、39+37+28+22+17、39+37+28+27+17、40+28+17、40+28+18+17、40+28+19+17、40+28+20+17、40+28+21+17、40+28+22+17、40+28+27+17、40+29+28+17、40+29+28+18+17、40+29+28+19+17、40+29+28+20+17、40+29+28+21+17、40+29+28+22+17、40+29+28+27+17、40+30+28+17、40+30+28+18+17、40+30+28+19+17、40+30+28+20+17、40+30+28+21+17、40+30+28+22+17、40+30+28+27+17、40+31+28+17、40+31+28+18+17、40+31+28+19+17、40+31+28+20+17、40+31+28+21+17、40+31+28+22+17、40+31+28+27+17、40+32+28+17、40+32+28+18+17、40+32+28+19+17、40+32+28+20+17、40+32+28+21+17、40+32+28+22+17、40+32+28+27+17、40+33+28+17、40+33+28+18+17、40+33+28+19+17、40+33+28+20+17、40+33+28+21+17、40+33+28+22+17、40+33+28+27+17、40+34+28+17、40+34+28+18+17、40+34+28+19+17、40+34+28+20+17、40+34+28+21+17、40+34+28+22+17、40+34+28+27+17、40+35+28+17、40+35+28+18+17、40+35+28+19+17、40+35+28+20+17、40+35+28+21+17、40+35+28+22+17、40+35+28+27+17、40+36+28+17、40+36+28+18+17、40+36+28+19+17、40+36+28+20+17、40+36+28+21+17、40+36+28+22+17、40+36+28+27+17、40+37+28+17、40+37+28+18+17、40+37+28+19+17、40+37+28+20+17、40+37+28+21+17、40+37+28+22+17、40+37+28+27+17、41+28+17、41+28+18+17、41+28+19+17、41+28+20+17、41+28+21+17、41+28+22+17、41+28+27+17、41+29+28+17、41+29+28+18+17、41+29+28+19+17、41+29+28+20+17、41+29+28+21+17、41+29+28+22+17、41+29+28+27+17、41+30+28+17、41+30+28+18+17、41+30+28+19+17、41+30+28+20+17、41+30+28+21+17、41+30+28+22+17、41+30+28+27+17、41+31+28+17、41+31+28+18+17、41+31+28+19+17、41+31+28+20+17、41+31+28+21+17、41+31+28+22+17、41+31+28+27+17、41+32+28+17、41+32+28+18+17、41+32+28+19+17、41+32+28+20+17、41+32+28+21+17、41+32+28+22+17、41+32+28+27+17、41+33+28+17、41+33+28+18+17、41+33+28+19+17、41+33+28+20+17、41+33+28+21+17、41+33+28+22+17、41+33+28+27+17、41+34+28+17、41+34+28+18+17、41+34+28+19+17、41+34+28+20+17、41+34+28+21+17、41+34+28+22+17、41+34+28+27+17、41+35+28+17、41+35+28+18+17、41+35+28+19+17、41+35+28+20+17、41+35+28+21+17、41+35+28+22+17、41+35+28+27+17、41+36+28+17、41+36+28+18+17、41+36+28+19+17、41+36+28+20+17、41+36+28+21+17、41+36+28+22+17、41+36+28+27+17、41+37+28+17、41+37+28+18+17、41+37+28+19+17、41+37+28+20+17、41+37+28+21+17、41+37+28+22+17、41+37+28+27+17、42+28+17、42+28+18+17、42+28+19+17、42+28+20+17、42+28+21+17、42+28+22+17、42+28+27+17、42+29+28+17、42+29+28+18+17、42+29+28+19+17、42+29+28+20+17、42+29+28+21+17、42+29+28+22+17、42+29+28+27+17、42+30+28+17、42+30+28+18+17、42+30+28+19+17、42+30+28+20+17、42+30+28+21+17、42+30+28+22+17、42+30+28+27+17、42+31+28+17、42+31+28+18+17、42+31+28+19+17、42+31+28+20+17、42+31+28+21+17、42+31+28+22+17、42+31+28+27+17、42+32+28+17、42+32+28+18+17、42+32+28+19+17、42+32+28+20+17、42+32+28+21+17、42+32+28+22+17、42+32+28+27+17、42+33+28+17、42+33+28+18+17、42+33+28+19+17、42+33+28+20+17、42+33+28+21+17、42+33+28+22+17、42+33+28+27+17、42+34+28+17、42+34+28+18+17、42+34+28+19+17、42+34+28+20+17、42+34+28+21+17、42+34+28+22+17、42+34+28+27+17、42+35+28+17、42+35+28+18+17、42+35+28+19+17、42+35+28+20+17、42+35+28+21+17、42+35+28+22+17、42+35+28+27+17、42+36+28+17、42+36+28+18+17、42+36+28+19+17、42+36+28+20+17、42+36+28+21+17、42+36+28+22+17、42+36+28+27+17、42+37+28+17、42+37+28+18+17、42+37+28+19+17、42+37+28+20+17、42+37+28+21+17、42+37+28+22+17、42+37+28+27+17、43+28+17、43+28+18+17、43+28+19+17、43+28+20+17、43+28+21+17、43+28+22+17、43+28+27+17、43+29+28+17、43+29+28+18+17、43+29+28+19+17、43+29+28+20+17、43+29+28+21+17、43+29+28+22+17、43+29+28+27+17、43+30+28+17、43+30+28+18+17、43+30+28+19+17、43+30+28+20+17、43+30+28+21+17、43+30+28+22+17、43+30+28+27+17、43+31+28+17、43+31+28+18+17、43+31+28+19+17、43+31+28+20+17、43+31+28+21+17、43+31+28+22+17、43+31+28+27+17、43+32+28+17、43+32+28+18+17、43+32+28+19+17、43+32+28+20+17、43+32+28+21+17、43+32+28+22+17、43+32+28+27+17、43+33+28+17、43+33+28+18+17、43+33+28+19+17、43+33+28+20+17、43+33+28+21+17、43+33+28+22+17、43+33+28+27+17、43+34+28+17、43+34+28+18+17、43+34+28+19+17、43+34+28+20+17、43+34+28+21+17、43+34+28+22+17、43+34+28+27+17、43+35+28+17、43+35+28+18+17、43+35+28+19+17、43+35+28+20+17、43+35+28+21+17、43+35+28+22+17、43+35+28+27+17、43+36+28+17、43+36+28+18+17、43+36+28+19+17、43+36+28+20+17、43+36+28+21+17、43+36+28+22+17、43+36+28+27+17、43+37+28+17、43+37+28+18+17、43+37+28+19+17、43+37+28+20+17、43+37+28+21+17、43+37+28+22+17、43+37+28+27+17、44+28+17、44+28+18+17、44+28+19+17、44+28+20+17、44+28+21+17、44+28+22+17、44+28+27+17、44+29+28+17、44+29+28+18+17、44+29+28+19+17、44+29+28+20+17、44+29+28+21+17、44+29+28+22+17、44+29+28+27+17、44+30+28+17、44+30+28+18+17、44+30+28+19+17、44+30+28+20+17、44+30+28+21+17、44+30+28+22+17、44+30+28+27+17、44+31+28+17、44+31+28+18+17、44+31+28+19+17、44+31+28+20+17、44+31+28+21+17、44+31+28+22+17、44+31+28+27+17、44+32+28+17、44+32+28+18+17、44+32+28+19+17、44+32+28+20+17、44+32+28+21+17、44+32+28+22+17、44+32+28+27+17、44+33+28+17、44+33+28+18+17、44+33+28+19+17、44+33+28+20+17、44+33+28+21+17、44+33+28+22+17、44+33+28+27+17、44+34+28+17、44+34+28+18+17、44+34+28+19+17、44+34+28+20+17、44+34+28+21+17、44+34+28+22+17、44+34+28+27+17、44+35+28+17、44+35+28+18+17、44+35+28+19+17、44+35+28+20+17、44+35+28+21+17、44+35+28+22+17、44+35+28+27+17、44+36+28+17、44+36+28+18+17、44+36+28+19+17、44+36+28+20+17、44+36+28+21+17、44+36+28+22+17、44+36+28+27+17、44+37+28+17、44+37+28+18+17、44+37+28+19+17、44+37+28+20+17、44+37+28+21+17、44+37+28+22+17、44+37+28+27+17、45+28+17、45+28+18+17、45+28+19+17、45+28+20+17、45+28+21+17、45+28+22+17、45+28+27+17、45+29+28+17、45+29+28+18+17、45+29+28+19+17、45+29+28+20+17、45+29+28+21+17、45+29+28+22+17、45+29+28+27+17、45+30+28+17、45+30+28+18+17、45+30+28+19+17、45+30+28+20+17、45+30+28+21+17、45+30+28+22+17、45+30+28+27+17、45+38+28+17、45+38+28+18+17、45+38+28+19+17、45+38+28+20+17、45+38+28+21+17、45+38+28+22+17、45+38+28+27+17、45+38+29+28+17、45+38+29+28+18+17、45+38+29+28+19+17、45+38+29+28+20+17、45+38+29+28+21+17、45+38+29+28+22+17、45+38+29+28+27+17、45+38+30+28+17、45+38+30+28+18+17、45+38+30+28+19+17、45+38+30+28+20+17、45+38+30+28+21+17、45+38+30+28+22+17、45+38+30+28+27+17、45+38+31+28+17、45+38+31+28+18+17、45+38+31+28+19+17、45+38+31+28+20+17、45+38+31+28+21+17、45+38+31+28+22+17、45+38+31+28+27+17、45+38+32+28+17、45+38+32+28+18+17、45+38+32+28+19+17、45+38+32+28+20+17、45+38+32+28+21+17、45+38+32+28+22+17、45+38+32+28+27+17、45+38+33+28+17、45+38+33+28+18+17、45+38+33+28+19+17、45+38+33+28+20+17、45+38+33+28+21+17、45+38+33+28+22+17、45+38+33+28+27+17、45+38+34+28+17、45+38+34+28+18+17、45+38+34+28+19+17、45+38+34+28+20+17、45+38+34+28+21+17、45+38+34+28+22+17、45+38+34+28+27+17、45+38+35+28+17、45+38+35+28+18+17、45+38+35+28+19+17、45+38+35+28+20+17、45+38+35+28+21+17、45+38+35+28+22+17、45+38+35+28+27+17、45+38+36+28+17、45+38+36+28+18+17、45+38+36+28+19+17、45+38+36+28+20+17、45+38+36+28+21+17、45+38+36+28+22+17、45+38+36+28+27+17、45+38+37+28+17、45+38+37+28+18+17、45+38+37+28+19+17、45+38+37+28+20+17、45+38+37+28+21+17、45+38+37+28+22+17、45+38+37+28+27+17、45+43+28+17、45+43+28+18+17、45+43+28+19+17、45+43+28+20+17、45+43+28+21+17、45+43+28+22+17、45+43+28+27+17、45+43+29+28+17、45+43+29+28+18+17、45+43+29+28+19+17、45+43+29+28+20+17、45+43+29+28+21+17、45+43+29+28+22+17、45+43+29+28+27+17、45+43+30+28+17、45+43+30+28+18+17、45+43+30+28+19+17、45+43+30+28+20+17、45+43+30+28+21+17、45+43+30+28+22+17、45+43+30+28+27+17、45+43+31+28+17、45+43+31+28+18+17、45+43+31+28+19+17、45+43+31+28+20+17、45+43+31+28+21+17、45+43+31+28+22+17、45+43+31+28+27+17、45+43+32+28+17、45+43+32+28+18+17、45+43+32+28+19+17、45+43+32+28+20+17、45+43+32+28+21+17、45+43+32+28+22+17、45+43+32+28+27+17、45+43+33+28+17、45+43+33+28+18+17、45+43+33+28+19+17、45+43+33+28+20+17、45+43+33+28+21+17、45+43+33+28+22+17、45+43+33+28+27+17、45+43+34+28+17、45+43+34+28+18+17、45+43+34+28+19+17、45+43+34+28+20+17、45+43+34+28+21+17、45+43+34+28+22+17、45+43+34+28+27+17、45+43+35+28+17、45+43+35+28+18+17、45+43+35+28+19+17、45+43+35+28+20+17、45+43+35+28+21+17、45+43+35+28+22+17、45+43+35+28+27+17、45+43+36+28+17、45+43+36+28+18+17、45+43+36+28+19+17、45+43+36+28+20+17、45+43+36+28+21+17、45+43+36+28+22+17、45+43+36+28+27+17、45+43+37+28+17、45+43+37+28+18+17、45+43+37+28+19+17、45+43+37+28+20+17、45+43+37+28+21+17、45+43+37+28+22+17、45+43+37+28+27+17、45+44+28+17、45+44+28+18+17、45+44+28+19+17、45+44+28+20+17、45+44+28+21+17、45+44+28+22+17、45+44+28+27+17、45+44+29+28+17、45+44+29+28+18+17、45+44+29+28+19+17、45+44+29+28+20+17、45+44+29+28+21+17、45+44+29+28+22+17、45+44+29+28+27+17、45+44+30+28+17、45+44+30+28+18+17、45+44+30+28+19+17、45+44+30+28+20+17、45+44+30+28+21+17、45+44+30+28+22+17、45+44+30+28+27+17、45+44+31+28+17、45+44+31+28+18+17、45+44+31+28+19+17、45+44+31+28+20+17、45+44+31+28+21+17、45+44+31+28+22+17、45+44+31+28+27+17、45+44+32+28+17、45+44+32+28+18+17、45+44+32+28+19+17、45+44+32+28+20+17、45+44+32+28+21+17、45+44+32+28+22+17、45+44+32+28+27+17、45+44+33+28+17、45+44+33+28+18+17、45+44+33+28+19+17、45+44+33+28+20+17、45+44+33+28+21+17、45+44+33+28+22+17、45+44+33+28+27+17、45+44+34+28+17、45+44+34+28+18+17、45+44+34+28+19+17、45+44+34+28+20+17、45+44+34+28+21+17、45+44+34+28+22+17、45+44+34+28+27+17、45+44+35+28+17、45+44+35+28+18+17、45+44+35+28+19+17、45+44+35+28+20+17、45+44+35+28+21+17、45+44+35+28+22+17、45+44+35+28+27+17、45+44+36+28+17、45+44+36+28+18+17、45+44+36+28+19+17、45+44+36+28+20+17、45+44+36+28+21+17、45+44+36+28+22+17、45+44+36+28+27+17、45+44+37+28+17、45+44+37+28+18+17、45+44+37+28+19+17、45 + 44 + 37 + 28 + 20 + 17, 45 + 44 + 37 + 28 + 21 + 17, 45 + 44 + 37 + 28 + 22 + 17, 45 + 44 + 37 + 28 + 27 + 17, 46 + 11, 46 + 12 + 11, 46 + 13 + 11, 46 + 14 + 11, 46 + 15 + 11, 46 + 15 + 12 + 11, 46 + 15 + 13 + 11, 46 + 15 + 14 + 11, 46 + 17, 46 + 18 + 17, 46 + 19 + 17, 46 + 20 + 17, 46 + 20 + 18 + 17, 46 + 20 + 19 + 17, 46 + 21 + 17, 46 + 22 + 17, 47 + 11, 47 + 12 + 11, 47 + 13 + 11, 47 + 14 + 11, 47 + 15 + 11, 47 + 15 + 12 + 11, 47 + 15 + 13 + 11, 47 + 15 + 14 + 11, 47 + 17, 47 + 18 + 17, 47 + 19 + 17, 47 + 20 + 17, 47 + 20 + 18 + 17, 47 + 20 + 19 + 17, 47 + 21 + 17 and 47 + 22 + 17; in the above list, the numbers refer to the numbers of the embodiments provided above, and the "+" indicates subordination from another embodiment. The different individualized embodiments are separated by commas. In other words, for example, "4 + 2 + 1" refers to embodiment 4) which is subordinate to embodiment 2) and subordinate to embodiment 1), i.e., embodiment "4 + 2 + 1" corresponds to the compound of embodiment 1) further restricted by the features of embodiments 2) and 4).

[0134] It should be understood that the ranges of i as set forth in embodiments 50) and 51) should be considered as being explicitly disclosed for each of the above-listed combinations.

[0135] If the plural forms of compounds, conjugates, salts, pharmaceutical compositions, diseases or the like are used, this is also intended to refer to a single compound, conjugate, salt, pharmaceutical composition, disease or the like.

[0136] If appropriate and convenient, any compound of (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV) as defined in any one of embodiments 1) to 81) mentioned should be understood to also refer to the salts of these compounds (and especially their pharmaceutically acceptable salts).

[0137] The term "pharmaceutically acceptable salts" refers to salts that retain the desired biological activity of the target compound and exhibit minimal undesired toxicological effects. These salts include inorganic or organic acid and / or base addition salts, depending on the presence of basic and / or acidic groups in the target compound. They may also be in the form of buffers or lyophilized products containing buffers for stabilization. For references, see, for example, "Handbook of Pharmaceutical Salts. Properties, Selection and Use.", P. Heinrich Stahl, Camille G. Wermuth (eds.), Wiley-VCH, 2008 and "Pharmaceutical Salts and Co-crystals", Johan Wouters and Luc Quéré (eds.), RSC Publishing, 2012.

[0138] This embodiment also includes isotopically labeled, in particular 2 compounds of formula (I), (Ia), (II), (IIa), (IIb), and (IIc) labeled with 2 H (deuterium), which are identical to the compounds of formula (I), (Ia), (II), (IIa), (IIb), and (IIc), but in which one or more atoms have each been replaced by an atom having the same atomic number but an atomic weight different from that usually found in nature. Isotopically labeled, in particular 2 compounds of formula (I), (Ia), (II), (IIa), (IIb), and (IIc) labeled with

[0139] The compounds of formula (I), (Ia), (II), (IIa), (IIb) and (IIc) as defined in any one of embodiments 1) to 55) and their pharmaceutically acceptable salts can be used as medicaments, for example in the form of pharmaceutical compositions for parenteral, enteral (e.g., oral) or nasal administration, especially parenteral administration (e.g., intramuscular, subcutaneous and intradermal injection).

[0140] 56) Accordingly, one aspect of the invention relates to a pharmaceutical composition comprising, as an active ingredient, an immunogenic compound as defined in any one of embodiments 1) to 55), especially embodiments 52), 53), 54) and 55), or a pharmaceutically acceptable salt thereof, and at least one therapeutically inert excipient.

[0141] The pharmaceutical composition can be produced in a manner familiar to any person skilled in the art (for example, see Remington, The Science and Practice of Pharmacy, 23rd Edition (2021), published by Elsevier Inc., ISBN: 978-0-12-820007-0; Vaccine Development and Manufacturing, 1st Edition (2014), published by John Wiley & Sons, ISBN: 9780470261941) by formulating the compounds of formula (I), (Ia), (II), (IIa), (IIb) and (IIc) or their pharmaceutically acceptable salts (optionally in combination with other therapeutically valuable substances) together with a suitable, non-toxic, inert, therapeutically compatible solid or liquid carrier material and (optionally) conventional pharmaceutical adjuvants into a galenical administration form.

[0142] In addition to the immunogenic compound as defined in any one of embodiments 1) to 55), the pharmaceutical composition may also comprise one or more (preferably 1, 2, 3 or 4; more preferably 2, 3 or 4; and most preferably 2 or 3) other immunogenic compounds (oligosaccharide - carrier protein conjugates) that are immunogenic against one or more other Klebsiella pneumoniae serotypes (especially against O1, O2ac, O2aeh, O3, O3a, O3b, O4, O5, O7, O8 and / or O12; and especially against O1, O3, O3b and / or O5).

[0143] The pharmaceutical composition is suitable for eliciting a protective immune response in a human and / or animal (especially mammalian (including human)) host, and can thus be used for the prevention and / or treatment of diseases associated with Klebsiella pneumoniae bacteria. Preferably, the pharmaceutical composition is suitable for humans.

[0144] The terms “prevention (prevention, preventing and / or prophylaxis)” are used synonymously and refer to inhibiting the initial onset of a pathological process such that the pathological process that could ultimately lead to the development of symptoms never occurs or those symptoms occur at a lower, non - dangerous intensity (i.e., preventing the occurrence of a disease, disorder or condition in a prophylactic manner).

[0145] The present pharmaceutical composition is suitable for administration to animal (and especially, human) patients and thus encompasses both human and veterinary uses. It can be used in a method of enhancing an immune response in a patient, the method comprising the step of administering the composition to the patient.

[0146] The pharmaceutical composition of the present invention can be administered before and / or after a subject is exposed to Klebsiella pneumoniae. Preferably, it is used before the subject is exposed to Klebsiella pneumoniae.

[0147] The pharmaceutical composition is preferably in an aqueous form (especially upon administration), but it can also be presented in a non - aqueous liquid form or in a dry form (such as in gelatin capsules or as a lyophilizate, etc.). Solid powders obtained, for example, by spray drying, spray freeze - drying, vacuum drying or air drying or lyophilization can be reconstituted before use. Lyophilization is preferred if a solid powder is required.

[0148] The pharmaceutical composition can include one or more pharmaceutically inert excipients. The excipient can be selected from the group consisting of: citric acid monohydrate, sodium citrate, sodium citrate dihydrate, acetic acid, sodium hydroxide, tromethamine, tromethamine hydrochloride (for adjusting pH), cholesterol, sorbitan trioleate, DSPC (l,2 - distearoyl - sn - glycero - 3 - phosphocholine) and bis(hexane - 6,1 - diyl)bis(2 - hexyl decanoate)(4 - hydroxybutyl)azanediyl ester), polydimethylsiloxane (antifoaming agent), ascorbic acid (antioxidant).

[0149] An excipient (such as sodium chloride (NaCl)) can be used to adjust the tonicity and it can be present at 1 to 20 mg / ml. Other salts that can be present include potassium chloride, potassium dihydrogen phosphate, disodium phosphate anhydrous, magnesium chloride, calcium chloride, etc.

[0150] The pharmaceutical composition can contain one or more excipients used as preservatives, which can be selected from the group consisting of: 2 - phenoxyethanol, benzethonium chloride, EDTA (ethylenediaminetetraacetic acid), formaldehyde, phenol and thimerosal (sodium ethylmercurithiosalicylate). Compositions free of mercury are preferred and a vaccine free of preservatives can be prepared.

[0151] The pharmaceutical composition may comprise one or more excipients used as surfactants, which may be selected from the group consisting of: polysorbate 20 (polyoxyethylene (20) sorbitan monolaurate), polysorbate 80 (polyoxyethylene (80) sorbitan monooleate), nonylphenol ethoxylate, octoxynol-10, and sodium deoxycholate.

[0152] The pharmaceutical composition may comprise a compound (with or without an insoluble metal salt) in purified water (such as water for injection, w.f.i.), but typically comprises one or more buffers. Typical buffers include: phosphate buffer, Tris buffer, borate buffer, succinate buffer, histidine buffer (especially with aluminum hydroxide adjuvant), or citrate buffer. The buffer salts are typically included in the range of 5 to 20 mM.

[0153] The pharmaceutical composition typically has a pH between 5.0 and 9.5 (such as between 6.0 and 8.0).

[0154] The pharmaceutical composition may further comprise one or more stabilizers.

[0155] The pharmaceutical composition is preferably sterile and gluten-free.

[0156] 57) Another embodiment of the present invention relates to a pharmaceutical composition as in embodiment 56), which further comprises an adjuvant.

[0157] As used herein, the term "adjuvant" refers to an immunological adjuvant (i.e., a material for use in a vaccine composition) that modifies or increases the effect of a vaccine by enhancing the immune response to a given antigen contained therein, without being antigenically related thereto. For those skilled in the art, classical recognized examples of immunological adjuvants include (but are not limited to) aluminum- or calcium-based salts, saponins or saponin-based adjuvants (e.g., Matrix-M), CpG oligodeoxynucleotide-based adjuvants (e.g., CpG1018), oil-in-water emulsions (e.g., Freund's adjuvant, MF59), activators of natural killer T cells (NKT cells) or invariant NKT cells (e.g., glycosphingolipids (e.g., KRN7000)), Toll-like receptor 1 / 2 (TLR-1 / 2) agonists (e.g., Pam3CSK4), TLR-3 agonists (e.g., poly(I:C)), TLR-4 agonists (e.g., lipopolysaccharide), TLR-5 agonists (e.g., flagellin), TLR-7 / 8 agonists (e.g., resiquimod), immunomodulatory proteins (e.g., detoxified heat-labile enterotoxin from Escherichia coli (dmLT)), the TLR-4 agonist glucopyranosyl lipid adjuvant-stable emulsion (GLA-SE) and monophosphoryl lipid A (MPL), nonionic block polymers, cytokines (e.g., type I interferon (IFN), granulocyte-macrophage colony-stimulating factor (GM-CSF), interleukins), the papain-like cysteine protease family, and many others (e.g., AS04, AS03, AS01 B ) and formulations of the above-mentioned adjuvants in the form of liposomes or nanoparticles prepared using lipids (e.g., DOPC (1,2-dioleoyl-sn-glycero-3-phosphocholine), DSPC (1,2-distearoyl-sn-glycero-3-phosphocholine), cholesterol, and / or ALC-0315), formulations in the form of virus-like particles, and composite formulations of the above-mentioned adjuvants, particularly composite formulations containing aluminum- or calcium-based salts.

[0158] The adjuvant "aluminum", "aluminum-based adjuvant", or "aluminum-salt-based adjuvant" is one or more of the following: amorphous aluminum hydroxyphosphate sulfate (AAHS), aluminum hydroxide, aluminum phosphate, and potassium alum.

[0159] An example of a calcium- or calcium-salt-based adjuvant is calcium phosphate.

[0160] Matrix-M is a saponin-based adjuvant composed of nanoparticles of saponins extracted from Quillaja saponaria (soap bark), cholesterol, and phospholipids.

[0161] CpG-based adjuvants are immunostimulatory oligodeoxynucleotides having one or more CpG motifs (CpG ODNs), which are unmethylated cytosine-guanine dinucleotides. The methylation status of the CpG immunostimulatory motif generally refers to the cytosine residue in the dinucleotide. An immunostimulatory oligonucleotide containing at least one unmethylated CpG dinucleotide is an oligonucleotide containing 5'-unmethylated cytosine linked via a phosphodiester bond to 3'-guanine and activating the immune system via ligation to Toll-like receptor 9 (TLR-9).

[0162] Freund's adjuvant is an oil-in-water adjuvant based on mineral oil.

[0163] MF59 is an oil-in-water emulsion that includes 4.3% w / v squalene, 0.5% w / v polysorbate 80 (Tween 80), and 0.5% w / v sorbitan trioleate (Span 85).

[0164] When glycolipids present MHC class I-associated molecules (e.g., CD1d), glycolipids are a class of lipids that stimulate unconventional invariant T-cell receptors on NKT cells or iNKT cells.

[0165] Pam3CSK4 (Pam3CysSerLys4) is a synthetic triacylated lipopeptide that is a ligand for TLR-1 and TLR-2. It mimics the acylated amino terminus of bacterial lipopeptides.

[0166] Poly(I:C) is a polymer and analog of double-stranded RNA that consists of a polymer chain of inosinic acid and a polymer chain of cytidylic acid. It stimulates TLR-3 and mimics viral infection.

[0167] Lipopolysaccharide (LPS) is a membrane component of Gram-negative bacteria and a stimulator of TLR-4.

[0168] Flagellin is a globular protein that forms the filaments of bacterial flagella. Flagellin activates TLR-5 and TLR-11.

[0169] Resiquimod (R848; 1-[4-amino-2-(ethoxymethyl)-1H-imidazo[4,5-c]quinolin-1-yl]-2-methylpropan-2-ol) is an immunomodulator and a small molecule that activates TLR-7 and TLR-8.

[0170] dmLT is a double mutant (and thus detoxified) of the heat-labile enterotoxin from Escherichia coli. It is an effective mucosal and systemic adjuvant.

[0171] GLA-SE is an oil-in-water emulsion adjuvant that is prepared by combining an aqueous solution of glucopyranosyl lipid A (GLA), a TLR-4 agonist, with squalene.

[0172] MPL (monophosphoryl lipid A, truncated LPS) is a TLR-4 agonist used clinically.

[0173] Suitable nonionic block polymers (NBP) as adjuvants are simple copolymers of polyoxyethylene (POE) and hydrophobic polyoxypropylene (POP) and differ in molecular weight, POE percentage, and the linkage pattern of the POE and POP groups.

[0174] Cytokines are small proteins secreted by cells that affect the interaction and communication between cells. Generally, cytokines activate target cells, thereby causing the secretion of additional cytokines and a signal transduction cascade. Cytokines are involved in the induction of innate and adaptive immunity. As adjuvants, cytokines can be used as recombinant proteins or can be encoded on DNA molecules (such as plasmids).

[0175] Papain-like cysteine protease families are derived from viruses, bacteria, yeast, protozoa, plants, or animals and contain a cysteine thiol at the active site. Such proteases can stimulate a Th2-type immune response.

[0176] AS04 (adjuvant system 04) is a complex of MPL (3-O-deacylated-4'-monophosphoryl lipid A) and aluminum hydroxide or aluminum phosphate.

[0177] AS03 (adjuvant system 03) is an oil-in-water squalene emulsion with DL-α-tocopherol (vitamin E) and polysorbate 80.

[0178] AS01 B is a mixture of 3-O-deacylated-4'-monophosphoryl lipid A (MPL) and saponin QS-21.

[0179] Preferred adjuvants are aluminum-based adjuvants, especially aluminum hydroxide.

[0180] 58) Another aspect of the present invention relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as described in any one of embodiments 1) to 55), especially embodiments 52), 53), 54), and 55), for use as a medicament, especially as a vaccine. In other words, the present invention relates to a vaccine comprising an immunogenic compound or a pharmaceutically acceptable salt thereof as described in any one of embodiments 1) to 55), especially embodiments 52), 53), 54), and 55). Preferably, the vaccine is for active vaccination.

[0181] 59) Another aspect of the present invention relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as described in any one of embodiments 1) to 55), especially embodiments 52), 53), 54), and 55), for the prevention and / or treatment of Klebsiella pneumoniae infection.

[0182] 60) Another embodiment of the present invention relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 1) to 55), particularly embodiments 52), 53), 54) and 55) for the prevention and / or treatment of Klebsiella pneumoniae infections in individuals 50 years of age or older, hospital-acquired (i.e., nosocomial) Klebsiella pneumoniae infections (e.g., nosocomial pneumonia, nosocomial bloodstream infections and nosocomial urinary tract infections), community-acquired Klebsiella pneumoniae infections, and pneumonia, bronchitis, meningitis, urinary tract infections, intra-abdominal infections, wound infections, blood infections, osteomyelitis, bacteremia, sepsis, liver abscesses and inflammatory bowel disease (IBD) all caused by Klebsiella pneumoniae infections.

[0183] Population-based strategies are desired for vaccinating individuals 50 years of age or older against Klebsiella pneumoniae infections, as this population is particularly vulnerable to Klebsiella pneumoniae infections, especially individuals 60 years of age or older and at risk of exposure to Klebsiella pneumoniae and / or with a predicted weakened immune system.

[0184] Klebsiella pneumoniae is a notorious pathogen that commonly causes hospital-acquired (i.e., nosocomial) respiratory and urinary tract infections. It is the second most common cause of Gram-negative bacteremia. Drug-resistant isolates are associated with high mortality (greater than 50% according to some studies), significantly increase the length of hospital stay, and are particularly problematic in the ICU.

[0185] Therefore, it is desirable to prevent hospital-acquired (i.e., nosocomial) Klebsiella pneumoniae infections, especially in populations at high risk of exposure, including patients undergoing elective surgery (e.g., joint replacement) with a hospital stay longer than 72 hours, patients with a weakened immune system, and patients predicted to have a weakened immune system (e.g., those on the waiting list for solid organ transplantation, undergoing non-emergency solid tumor surgery and subsequent chemotherapy). In these populations, vaccination 2 - 8 weeks before surgery, optionally followed by a booster, is applicable.

[0186] In addition, it is desirable to prevent community-acquired infections in specific target groups in long-term care facilities or nursing homes (e.g., healthcare workers or the elderly (60 years of age or older)). The term "community-acquired Klebsiella pneumoniae infection" refers to any Klebsiella pneumoniae infection acquired in the community. It is different from nosocomial (hospital-acquired) infections.

[0187] In addition, the immunogenic compound of the present invention or a pharmaceutically acceptable salt thereof according to any one of embodiments 1) to 55), particularly embodiments 52), 53), 54) and 55), can be used for the prevention and / or treatment of pneumonia, bronchitis, meningitis, urinary tract infection, intra-abdominal infection, wound infection, blood infection, osteomyelitis, bacteremia, sepsis, liver abscess and inflammatory bowel disease (IBD), all of which are caused by Klebsiella pneumoniae infection.

[0188] 61) Another embodiment of the present invention relates to an immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of embodiments 1) to 55), particularly embodiments 52), 53), 54) and 55), for the prevention and / or treatment of Klebsiella pneumoniae infection as listed in embodiments 59) and 60) above, wherein Klebsiella pneumoniae is selected from O-serotypes (including O2a and O2afg).

[0189] 62) To avoid any doubt, the immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of embodiments 1) to 55), particularly embodiments 52), 53), 54) and 55), the pharmaceutical composition according to embodiment 56) or 57), and the vaccine according to embodiment 58) are also suitable for the prevention and / or treatment of Klebsiella pneumoniae infection as listed in any one of embodiments 59), 60) and 61).

[0190] 63) Preferably, the immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of embodiments 1) to 55), particularly embodiments 52), 53), 54) and 55), the pharmaceutical composition according to embodiment 56) or 57), and the vaccine according to embodiment 58) are suitable for the prevention (prevention and / or prophylaxis) of Klebsiella pneumoniae infection as listed in any one of embodiments 59), 60) and 61).

[0191] 64) Another aspect of the present invention relates to a method for eliciting an immune response against Klebsiella pneumoniae in a human and / or animal (especially mammalian (including human)) host, which comprises administering to the human and / or animal an effective amount of an immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of embodiments 1) to 55), particularly embodiments 52), 53), 54) and 55). The dosage administered is preferably 0.05 μg to 30 μg of glycan / immunization of a human patient. The term "glycan" refers to an antigen, i.e., an oligosaccharide that does not contain the linker L and the spacer T. Possibly, more than one immunization is required.

[0192] 65) Similarly, embodiments of the present invention relate to methods of eliciting an immune response against Klebsiella pneumoniae in a human and / or animal (especially mammalian (including human)) host, which comprise administering to the human and / or animal an effective amount of a composition as in embodiment 56) or 57) and a vaccine as in embodiment 58).

[0193] 66) To avoid any doubt, if an immunogenic compound as in any one of embodiments 1) to 55), especially embodiments 52), 53), 54) and 55), or a pharmaceutically acceptable salt thereof is described as being useful for preventing and / or treating a Klebsiella pneumoniae infection as in any one of embodiments 59), 60) and 61), then these immunogenic compounds are equally suitable for use in the preparation of a medicament for preventing and / or treating such Klebsiella pneumoniae infection as in any one of embodiments 59), 60) and 61).

[0194] 67) Another aspect of the present invention relates to a multivalent vaccine, which comprises an immunogenic compound as in any one of embodiments 1) to 55), preferably an immunogenic compound as in embodiments 52), 53), 54) and 55) or a pharmaceutically acceptable salt thereof.

[0195] The term "multivalent vaccine" in this context relates to a vaccine comprising antigens against two or more different Klebsiella pneumoniae strains, especially against two or more pathogenic Klebsiella pneumoniae strains.

[0196] 68) Another aspect of the present invention relates to an intermediate compound for the preparation of an immunogenic compound as in any one of embodiments 28) to 55), which has the formula (III) wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; L 1 represents *-(C 2-10 ) alkylene - NH2; *-(CH2CH2O) b -CH2CH2NH2, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH2; *-(C 2-10 ) fluoroalkylene - NH2; *-(CH2) c NHC(O)(CH2) c’ -NH2, where c and c' are independently of each other 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH2, where d and d' are each independently 2 to 6; *-(C 1-10 )alkylene-C(O)-NH-(C 2-10 )alkylene-NH2; *-(C 2-10 )alkylene-O-NH2; or *-(C 2-10 )alkylene-SH; or a pharmaceutically acceptable salt thereof.

[0197] 69) Another embodiment relates to an intermediate compound as in embodiment 68) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is m is 4, n is 3 and R is OH.

[0198] 70) Another embodiment relates to an intermediate compound as in embodiment 68) or 69) or a pharmaceutically acceptable salt thereof, wherein L 1 represents *-(CH2) a -NH2; where a is 2 to 10; *-(CH2CH2O) b -CH2CH2NH2, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH2; *-(C 2-10 )fluoroalkylene-NH2, where the fluoroalkylene is a saturated straight chain; *-(CH2) c NHC(O)(CH2) c’ -NH2, where c and c' are each independently 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH2, where d and d' are each independently 2 to 6; *-(CH2) e -C(O)-NH-(CH2) e’ -NH2; where e is 1 to 10 and e' is 2 to 10; *-(CH2) f -O-NH2, where f is 2 to 10; or *-(CH2) g -SH, where g is from 2 to 10.

[0199] It should be understood that, in a similar manner, Embodiments 30) to 37) disclose other preferred Ls having terminal amino- or SH-groups as shown in Embodiment 68). 1 .

[0200] 71) Another embodiment relates to an intermediate compound or a pharmaceutically acceptable salt thereof as in Embodiment 68) or 69), wherein L 1 represents *-(CH2)2-NH2, *-(CH2)3-NH2, *-(CH2)4-NH2, *-(CH2)5-NH2 or *-(CH2)6-NH2; preferably *-(CH2)5-NH2.

[0201] 72) Another embodiment relates to an immunogenic compound or a pharmaceutically acceptable salt thereof as in Embodiment 69), wherein m is 4, n is 2 and R is OH; and L 1 represents *-(CH2)2-NH2, *-(CH2)3-NH2, *-(CH2)4-NH2, *-(CH2)5-NH2 or *-(CH2)6-NH2; preferably *-(CH2)5-NH2.

[0202] 73) Another aspect of the present invention relates to an intermediate compound for preparing an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of Embodiments 28) to 55), which has the formula (IV): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; L represents *-(C 2-10 )alkylene-NH-; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 )fluoroalkylene-NH-; *-(CH2) cNHC(O)(CH2) c’ -NH-, where c and c' are each independently 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d' are each independently 2 to 6; *-(C 1-10 ) alkylene - C(O) - NH - (C 2-10 ) alkylene - NH -; or *-(C 2-10 ) alkylene - O - NH -; and T 1 represents -C(O)-(C 0-10 ) alkylene - C(O)X; -C(O)-CH2CH2-(OCH2CH2) j -C(O)X, where j is 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)X, where k is 0 or 1, k' is 0 or 1, and k" is 1, 2, or 3; where l is 1 or 2; or where p is 1 to 4, preferably 1, and p' is 1 or 2; -C(O)X represents -C(O)OH or an activated ester; and Y represents Me, Et, Bu, or -(CH2CH2O)3CH3 (especially Me, Et, n-Bu, or -(CH2CH2O)3CH3). The term "activated ester" refers to a functionalized carboxylic acid that has enhanced reactivity towards an amine (compared to a carboxylic acid) for reaction with the amino group of a lysine residue of CRM 197 .

[0203] 74) Another embodiment relates to an intermediate compound as in embodiment 73) or a pharmaceutically acceptable salt thereof, where m is 4, n is 2, and R is OH; m is 4, n is 2, and R is m is 4, n is 3, and R is OH.

[0204] 75) Another embodiment relates to an intermediate compound as in embodiment 73) or 74) or a pharmaceutically acceptable salt thereof, where L represents *-(CH2) a -NH-; where a is from 2 to 10; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 ) fluoroalkylene-NH-, where the fluoroalkylene is a saturated straight chain; *-(CH2) c NHC(O)(CH2) c’ -NH-, where c and c’ are independently of each other from 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d’ are independently of each other from 2 to 6; *-(CH2) e -C(O)-NH-(CH2) e’ -NH-; where e is from 1 to 10 and e’ is from 2 to 10; or *-(CH2) f -O-NH-, where f is from 2 to 10; and T 1 represents -C(O)-(CH2) h -C(O)-, where h is from 0 to 10; -C(O)-CH2CH2-(OCH2CH2) j -C(O)X, where j is from 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)X, where k is 0 or 1, k’ is 0 or 1, and k” is 1, 2 or 3; where l is 1 or 2; or where p is from 1 to 4, preferably 1, and p’ is 1 or 2. -C(O)X represents -C(O)OH or an activated ester; and Y represents Me, Et, Bu or -(CH2CH2O)3CH3 (especially Me, Et, n-Bu or -(CH2CH2O)3CH3).

[0205] 76) Another embodiment relates to an intermediate compound as in embodiment 73) or 74) or a pharmaceutically acceptable salt thereof, wherein L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH-; preferably *-(CH2)5-NH-; T 1 represents -C(O)-C(O)X, -C(O)-CH2-C(O)X, -C(O)-(CH2)2-C(O)X, -C(O)-(CH2)3-C(O)X, -C(O)-(CH2)4-C(O)X, -C(O)-(CH2)5-C(O)X or -C(O)-(CH2)6-C(O)X; preferably -C(O)-(CH2)4-C(O)X; and -C(O)X represents -C(O)OH or an activated ester.

[0206] 77) Another embodiment relates to an immunogenic compound as in embodiment 76) or a pharmaceutically acceptable salt thereof, wherein m is 4, n is 2 and R is OH; L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH-; preferably *-(CH2)5-NH-; T 1 represents -C(O)-C(O)X, -C(O)-CH2-C(O)X, -C(O)-(CH2)2-C(O)X, -C(O)-(CH2)3-C(O)X, -C(O)-(CH2)4-C(O)X, -C(O)-(CH2)5-C(O)X or -C(O)-(CH2)6-C(O)X; preferably -C(O)-(CH2)4-C(O)X; and -C(O)X represents -C(O)OH or an activated ester. In embodiments 73), 74), 75), 76) and 77), preferably X represents It should be understood that embodiments 30) to 37) disclose other preferred Ls covered by this embodiment. In addition, it should be understood that in a similar manner, embodiments 39) to 44) disclose the conversion to T 1Other preferred Ts, wherein the terminal "C(O)-" is replaced by "C(O)X", and in the case of squaric acid, the point of attachment to the carrier protein (e.g., to CRM 197 ) is represented as "O-Y", and R 1 is H. These preferred Ts 1 should be regarded as expressly disclosed.

[0207] 78) Yet another aspect of the present invention relates to an immunoassay comprising an oligosaccharide hybrid antigen of formula (I) wherein R is OH or m is 3, 4, 5, 6, 7 or 8; and n is 1, 2, 3, 4, 5 or 6. In this embodiment, the "**" in the dashed line refers to the point of attachment to the array surface preferably via a linker and / or spacer. The oligosaccharide hybrid antigen of formula (I) can be attached to any carrier surface suitable for an array or microarray with or without a linker and / or spacer.

[0208] 79) A preferred embodiment of the present invention relates to an immunoassay as in embodiment 78), which comprises a compound of formula (Ia): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; and n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; i is at least 1, preferably a number from 1 to 90% of the count corresponding to the number of lysine residues contained in the carrier protein CP; -L-T- represents a linker L and a spacer T as disclosed in any one of embodiments 17) or 23) to 47); and CP is a carrier protein suitable for immunological assays, particularly ELISA. Preferred CPs are described in embodiments 8) and 9). A particularly preferred carrier protein is BSA. The synthesis of the compound of formula (Ia) antigen bound to BSA is described and illustrated in the experimental section. It should be understood that this synthesis applies analogously to all antigens of formula (Ia) so that those skilled in the art can prepare them. The assay method of this embodiment is suitable for detecting antibodies against Klebsiella pneumoniae O2a and O2afg strains.

[0209] 80) Another aspect of the present invention relates to an immunogenic compound as in any one of embodiments 28) to 55) or a pharmaceutically acceptable salt thereof, wherein the immunogenic compound can be obtained or prepared by binding a compound of formula (IV): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; L represents *-(C 2-10 )alkylene-NH-; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 )fluoroalkylene-NH-; *-(CH2) c NHC(O)(CH2) c’ -NH-, where c and c' are independently 2 to 6 from each other; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d' are independently 2 to 6 from each other; *-(C 1-10 )alkylene-C(O)-NH-(C 2-10 )alkylene-NH-; or *-(C 2-10 )alkylene-O-NH-; and a) T 1 represents -C(O)-(C 0-10 )alkylene-C(O)X; -C(O)-CH2CH2-(OCH2CH2) j -C(O)X, where j is 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)X, where k is 0 or 1, k' is 0 or 1, and k'' is 1, 2 or 3; -C(O)X represents -C(O)OH or an activated ester; and Y represents to CRM197 Me, Et, Bu or -(CH2CH2O)3CH3 of the lysine residue; or b) binding a compound of formula (IV) having the following characteristics: wherein T 1 represents wherein l is 1 or 2; or wherein p is 1 to 4, preferably 1, and p' is 1 or 2; or wherein L-T 1 represents *-(C 2-10 ) alkylene - SH; to a modified CRM selected from the group consisting of 197 : wherein Z is Br or I, q is 2 or 3, and t is 1 to 28; wherein r is 2 or 3, and t' is 1 to 28; and wherein Z is Br or I, and t'' is 1 to 28. Preferably, X represents: It should be understood that embodiments 29) to 37) disclose other preferred Ls, and embodiments 73) and 75) to 77) disclose other preferred Ts 1 , and all these groups are covered in this embodiment. In addition, it should be understood that in a similar manner, embodiments 39) to 44) disclose other preferred Ts converted to T 1 wherein the terminal "C(O)-" is replaced by "C(O)X", and in the case of squaric acid, the attachment point to the carrier protein (e.g., to CRM 197 ) is represented as "O - Y", and R 1 is H. These preferred Ts 1 should be regarded as clearly disclosed.

[0210] 81) Another aspect of the present invention relates to a process for preparing an immunogenic compound or a pharmaceutically acceptable salt thereof as in any one of embodiments 28) to 55), wherein the process comprises binding a compound of formula (IV): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; L represents *-(C2-10 ) alkylene-NH-; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 ) fluoroalkylene-NH-; *-(CH2) c NHC(O)(CH2) c’ -NH-, where c and c’ are independently 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d’ are independently 2 to 6; *-(C 1-10 ) alkylene-C(O)-NH-(C 2-10 ) alkylene-NH-; or *-(C 2-10 ) alkylene-O-NH-; and a) T 1 represents -C(O)-(C 0-10 ) alkylene-C(O)X; -C(O)-CH2CH2-(OCH2CH2) j -C(O)X, where j is 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)X, where k is 0 or 1, k’ is 0 or 1, and k” is 1, 2 or 3; -C(O)X represents an activated ester; Y represents Me, Et, Bu or -(CH2CH2O)3CH3 of the lysine residue to CRM 197 ; or b) binding a compound of formula (IV) having the following characteristics: where T 1 represents where l is 1 or 2; or where p is 1 to 4, preferably 1, and p’ is 1 or 2; or where L-T 1 represents *-(C 2-10 ) alkylene-SH; to a modified CRM selected from the group consisting of the following 197 : where Z is Br or I, q is 2 or 3, and t is from 1 to 28; where r is 2 or 3, and t' is from 1 to 28; and where Z is Br or I, and t'' is from 1 to 28.

[0211] Preferably, X represents It should be understood that embodiments 29) to 37) disclose other preferred Ls, and embodiments 73) and 75) to 77) disclose other preferred Ts 1 , and all of these groups are encompassed in this embodiment. In addition, it should be understood that, in a similar manner, embodiments 39) to 44) disclose other preferred Ts converted to T 1 where the terminal "C(O)-" is replaced by "C(O)X", and in the case of squaric acid, the point of attachment to the carrier protein (e.g., to CRM 197 ) is represented as "O-Y", and R 1 is H. These preferred Ts 1 should be regarded as being explicitly disclosed.

[0212] Whenever the words "between... and" or "to" are used to describe a numerical range, it should be understood that the endpoints of the indicated range are explicitly disclosed and included within that range. For example: if the temperature range is described as being between 40°C and 80°C (or 40°C to 80°C), this means that the endpoints 40°C and 80°C are included in the range; or if a variable is defined as an integer between 1 and 4 (or 1 to 4), this means that the variable is the integer 1, 2, 3, or 4.

[0213] However, to avoid any doubt, the definition "a bridge having a backbone of 5 to 25 atoms (covalently linked together) that forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at CRM 197 " means that the oxygen at C1 and the amino nitrogen of lysine at CRM 197 are not counted in the numbering of the defined backbone.

[0214] Unless otherwise indicated with respect to temperature, the term "about" (or alternatively "around") placed before a numerical value "X" in this application means the range extending from X - 10%X to X + 10%X, and preferably means the range extending from X - 5%X to X + 5%X. In the specific case of temperature, the term "about" (or alternatively "around") placed before a temperature "Y" in this application means the range extending from temperature Y - 10 °C to Y + 10 °C, and preferably means the range extending from Y - 5 °C to Y + 5 °C. In addition, as used herein, the term "room temperature" means a temperature of about 25 °C.

[0215] Preparation of the compounds of formulas (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV)

[0216] Another aspect of the present invention is a process for preparing the compounds of formulas (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV). The compounds of formulas (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV) of the present invention can be prepared from commercially available or well-known starting materials according to the methods described in the experimental section, by similar methods or according to the general sequence of reactions outlined below, where L, T, L 1 , T 1 , X and Y are as defined in formulas (I), (Ia), (II), (IIa), (IIb), (IIc), (III) and (IV). Other abbreviations used herein are either explicitly defined or as defined in the experimental section.

[0217] The synthesis of the compounds of the present invention requires a protecting group strategy. Although this protecting group strategy can be complex, the use of protecting groups is well known in the art (for example, see "Protective Groups in Organic Synthesis", T.W. Greene, P.G.M. Wuts, Wiley-Interscience, 1999). The compounds obtained can also be converted into salts, especially pharmaceutically acceptable salts, in a manner known per se.

[0218] General preparation route: Antigen illustration

[0219] Reaction Figure 1 : Synthesis of AG-CRM using the NHS-ester method 197 Conjugate Means all linkers L as set forth in embodiments 28) and 29) to 47) having a terminal amino group 1 Treat antigen AG-L in a suitable solvent (e.g., DMSO) in a vial at room temperature with the activated Bis-NHS ester of diacid 2’ (e.g., adipic acid bis-NHS ester, which is commercially available or can be prepared by those skilled in the art using the corresponding diacid and N-hydroxysuccinimide) (Odom, O.W., Biochemistry, Vol. 29, No. 48, 1990) (5 - 20 equivalents) in DMSO in the presence of triethylamine 1 1’ and stir at room temperature for 3 h. Precipitate antigen-NHS ester 3’ by adding EtOAc, and centrifuge, then wash the precipitate with EtOAc, dry in vacuo, and then use for the next step. Stir a buffer solution containing antigen-NHS ester 3’ (25 - 100 equivalents) and CRM 197 for 20 - 24 h at room temperature. Wash, purify, and store the resulting antigen-CRM 197 conjugate 4’. In the synthetic pathway above, CRM 197 can be replaced by any of the carrier proteins as set forth in embodiments 8) or 9).

[0220] Reaction Figure 2 : Synthesize AG-CRM conjugate using the squarate method 197 conjugate Treat antigen AG-L in a suitable solvent (e.g., H2O-EtOH buffer) in a vial at room temperature with the desired squarate alkyl ester 5’ (e.g., 3,4-dibutoxy-3-cyclobutene-1,2-dione, 3,4-(bis(2-(2-(2-methoxyethoxy)ethoxy)ethoxy)-3-cyclobutene-1,2-dione) (Ganesh et al., JACS, 2014, 136, 16260 - 16269 and Xu et al., Carbhydr.Res, 2018, 456, 24 - 29)) 1 1’ and stir at room temperature in a solvent having a suitable pH (7 - 8). Neutralize the reaction mixture with acetic acid and then concentrate in vacuo (or lyophilize). Purify the crude product using a C18 (or SEC) column with water-acetonitrile as the eluent. Freeze and lyophilize the fraction containing the product to provide 6’. Stir a buffer solution containing antigen-squarate 6’ (25 - 100 equivalents) and CRM 197Stir for 24 - 72 h in 0.5 M pH 9 borate buffer solution (S. Hou et al., Carbhydr. Res, 2008, 343, 196 - 210). Wash, purify and store the resulting antigen - CRM 197 conjugate 7' using an appropriate buffer solution. In the above synthetic pathway, CRM 197 can be replaced by any one of the carrier proteins described in embodiments 8) or 9).

[0221] Reaction Figure 3 : Synthesis of antigen - thiol Treat antigen AG - L in a suitable solvent (e.g., DMSO) in a vial at room temperature with 8' (e.g., DSP (dithiobis(succinimidyl propionate)) or DTSSP (3,3' - dithiobis(sulfosuccinimidyl propionate))) to obtain the corresponding disulfide, which is then reduced by DTT (dithiothreitol) or TCEP (tris(2 - carboxyethyl)phosphine) to provide antigen - thiol 9'. 1 1'

[0222] Reaction Figure 4 : Synthesis of AG - CRM 197 conjugate using antigen - thiol and functionalized CRM 197 conjugate a) Synthesize AG - CRM 197 conjugate using the antigen - thiol - maleimide method Stir a buffer solution containing antigen - thiol 9' (25 - 100 equivalents) and CRM functionalized with maleimide 10' 197 (e.g., which can be prepared by those skilled in the art by treating CRM with 3 - maleimidopropionic acid succinimidyl ester or any other suitable NHS ester with maleimide)(Robert M.F. van der Put et al., ACS Cent. Sci. 2022, 8, 4, 449 - 460) at room temperature for 20 - 24 h. Then quench the excess maleimide moiety by adding L - cysteine in buffer to RM and stir for one hour at room temperature. Wash, purify and store the resulting antigen - CRM 197 -thiol - maleimide conjugate 11' using an appropriate buffer solution. 197 In the above synthetic pathway, CRM 197 can be replaced by any one of the carrier proteins described in embodiments 8) or 9). b) Synthesis of AG-CRM using the antigen-thiol-ether method 197 Conjugate At room temperature, a buffer solution containing antigen-thiol 9’ (25 - 100 equivalents) and protein functionalized with α-bromoacetate 10’ (e.g., CRM synthesized using CRM 197 and SBAP (3-(2-bromoacetamido)propionic acid N-maleimidyl ester) or any other suitable NHS ester with α-bromoacetate) (Schumann, B. et al., Chem. Sci., 2014, 5, 1992 - 2002) was stirred for 24 h. Then, L-cysteine in the buffer was added to the RM to quench the excess α-bromoacetate moiety and stirred for one hour at room temperature. The resulting antigen-CRM 197 -thiol-ether conjugate 11’ was washed, purified, and stored using an appropriate buffer solution. 197 -thiol-ether conjugate 11’. In the synthetic pathway above, CRM 197 can be replaced by any of the carrier proteins described in embodiments 8) or 9). General retrosynthetic approach of AG-heterologous-antigen RS-1 Antigen RS-1 can be synthesized using functionalized building blocks as shown in reaction Figure 5 . The fully deprotected antigen RS-1 has a linker L1 at its reducing end, which is necessary for binding to the protein carrier. Linker L1 is disclosed in embodiments 28) and 29) to 47). RS-1 can be obtained from the deprotection of fully protected RS-2. The deprotection strategy may involve removing esters, amides, imides, carbamates via (acidic or basic) hydrolysis, hydrogenolysis, birch reduction, or reduction of azide to amine. The deprotection sequence depends on the protecting groups and their compatibility with the reaction conditions. Those skilled in the art can successfully achieve this. RS-2 can be obtained from the glycosylation of RS-3 as the donor and RS-4 as the receptor. RS-4 can be obtained from intermediates RS-5 and RS-6, which are equipped with appropriate linkers (Lx). RS-9 donor can be treated with various linkers selected from those listed in (Table A) to obtain RS-6. RS-5 and RS-9 synthesized using RS-7 and RS-8 can also be obtained from the repeating unit RS-8. Thus, the common intermediate RS-8 can be obtained from the monosaccharide building blocks RS-10 and RS-11.

[0223] Reaction Figure 5 : Retrosynthetic approach of antigen RS-1 Lx = linker with protected functional groups LG1, LG2, LG3, LG4, LG5 = leaving group = imidoester, phosphate, STol, 5-butyl-o-thiocresol, SPh or SEt TPG = temporary protecting group = OLev or ONap

[0224] Reaction Scheme 6: Introduction of the linker Lx TPG = OLev or ONap LG4 = imidoester, phosphate, STol, 5-tert-butyl-o-thiocresol, SPh or SEtLx = linker with a protected functional group Add the linker nucleophile Ln (e.g., 5-azidopentan-1-ol) and the RS-9 donor to an RBF and dry azeotropically with anhydrous toluene under vacuum. Add the mixture to a suitable solvent (e.g., DCM) at room temperature, add 4A molecular sieves to it, and stir under an N2 atmosphere for 30 - 45 min. Cool the RM to a suitable temperature (e.g., 0 °C to -20 °C) and add an activator (e.g., TMSOTf, TfOH) to the RM and stir the RM for 20 min. Then allow the RM to warm slowly to room temperature over 1 hr. Monitor the completion of the reaction by TLC. (e.g., using sat. NaHCO3, Na2S2O3 solution) Quench the RM and extract with a solvent (e.g., DCM, EtOAc). Wash the combined organics with water, brine, dry, evaporate in vacuo to obtain the crude product. Purify the crude product by silica column chromatography using EA / cyclohexane as the eluent. Evaporate the fraction containing the product and dry in vacuo to obtain the product RS-6. Table A: List of nucleophilic linkers Ln

[0225] Experimental Section: Abbreviations (as used herein and in the above description): AcOH acetic acid aq. aqueous Bn benzyl BSA bovine serum albumin CDCl3 deuterochloroform Cs2CO3 cesium carbonate Cy cyclohexane D2O deuterium oxide DCM dichloromethane DDQ 2,3-dichloro-5,6-dicyano-1,4-benzoquinone DMAP 4-(dimethylamino)pyridine DMF N,N-dimethylformamide DMSO dimethyl sulfoxide ELISA enzyme-linked immunosorbent assay equiv equivalent ESI electrospray ionization Et3N(TEA) triethylamine EtOAc(EA) ethyl acetate EtOH ethanol EtSH ethanethiol Fr fraction h hour H2 hydrogen H2O water H2SO4 sulfuric acid HCl hydrochloric acid HPLC high performance liquid chromatography HPLC-SEC high performance liquid chromatography-size exclusion chromatography I2 iodine ICU intensive care unit IPA isopropanol LPS lipopolysaccharide M molarity MeOH methanol Min minute MS molecular sieve N2 nitrogen Na sodium Na2S2O3 sodium thiosulfate Na2SO4 sodium sulfate NaCl sodium chloride NaHCO3 sodium bicarbonate NaOMe sodium methoxide NaPi buffer sodium phosphate buffer NH2NH2 hydrazine NIS N-iodosuccinimide NMR nuclear magnetic resonance spectroscopy PBS phosphate buffered saline PBS-T phosphate buffered saline with 0.1% (v / v) Tween-20 Pd(OH)2 palladium hydroxide Pd / C palladium on carbon py pyridine RBF round bottom flask RM reaction mixture rt room temperature sat. saturated SDS-PAGE Sodium dodecyl sulfate polyacrylamide gel electrophoresis SM Starting material sol. Solution TBAF Tetrabutylammonium fluoride TBS Tris-buffered saline TDS Dimethyl-tert-hexylchlorosilane TLC Thin layer chromatography TMB 3,3’,5,5’-Tetramethylbenzidine TMSOTf Trimethylsilyl trifluoromethanesulfonate UV Ultraviolet light

[0226] I. Chemistry The following examples illustrate the preparation of the bioactive compounds of the invention, but in no way limit its scope. General information: All reagents and solvents were used as purchased and the solvents used in the reactions were anhydrous. All reactions were carried out in dry glassware (purchased from VWR and ROTH) under a N2 atmosphere, except for reactions containing water as a solvent. Before glycosylation, it is strongly recommended to dehydrate the acceptor and donor by azeotroping twice with anhydrous toluene. Experiments were carried out using a Heidolph magnetic stirrer. Thin layer chromatography (TLC) was carried out on silica gel 60F254 glass plates (Merck) or aluminum plates (VWR). Visualization of the developed TLC plates was carried out under short-wave UV light and by heating the plates soaked in the sugar stain solution (3-methoxyphenol (0.225 mL), H2SO4 (6 mL) and EtOH (200 mL)). All automated flash chromatography purifications were carried out on silica gel (FlashPureSilica 40μm irregular: BUCHI column) using Biotage Isolera and Biotage Select. Solvents were evaporated using a BUCHI rotary evaporator. The reaction mixture was cooled using dry ice and acetone and an ice / water combination to obtain the desired temperature. All NMR experiments were carried out on a BRUKER 400 MHz instrument. Temperatures are indicated in degrees Celsius (°C). In mixtures, the parts of solvents or eluents or reagent mixtures in liquid form are given in volume relationship (v / v) unless otherwise stated.

[0227] Characterization methods used: HPLC-SEC: The glycoconjugates for immunization were analyzed by HPLC-SEC to observe the conjugated and unbound CRM 197Mass difference between proteins. The sample was diluted in 50 mM Tris, 20 mM NaCl (pH 7.2) and run on an Agilent 1100 HPLC system equipped with a Tosoh TSK G2000 column (SWxl, 7.8 mm x 30 cm, 5 μm) and a Tosoh TSK Gel Guard column (SWxl 6.0 mm x 4 cm, 7 μm). The flow rate was maintained at 1 mL / min. SDS-PAGE: The sample was diluted in Laemmli loading buffer and heated at 95 °C for 5 min. After cooling at room temperature for 5 min, approximately 2 - 2.5 μg of the sample was loaded into the wells of a 10% polyacrylamide gel together with approximately 5 μL of protein size markers. The sample was run at a constant voltage of 120 V for approximately 30 - 45 min. Staining was performed using Gel Code TM Blue Safe Protein Stain according to the manufacturer's instructions. The gel was washed with deionized water overnight and scanned.

[0228] Synthesis of thiol disaccharide D1: To a solution of disaccharide A3 (see WO2019106201, page 164), 68 g, 61 mmol) in HPLC-grade DCM (305 mL) was added EtSH (27.1 mL, 366 mmol) and TsOH·H2O (3.15 g, 18.29 mmol). After stirring at room temperature for 40 min, TLC analysis (EtOAC / hexane, 1 / 1) showed disappearance of the starting material and the presence of new spots. The reaction mixture was then quenched with triethylamine (2.55 mL, 18.29 mmol) and concentrated under reduced pressure. The residue was purified by flash silica column chromatography (gradient DCM / MeOH, 0 to 10%). The fractions containing the product were concentrated in vacuo and dried under high vacuum to afford D1 as a colorless oil (55 g, 88%). HRMS C 59 H 70 NO 14 Si + [M+NH4] + Calculated value: 1044.4560, experimental value: 1044.460.

[0229] Synthesis of disaccharide acceptor D2: Benzoic anhydride (12.51 g, 55.3 mmol) and triethylamine (38.9 g, 53.6 mL) were added to a solution of thiol D1 (49.4 g, 48.1 mmol) in anhydrous DCM (385 mL), and the reaction mixture was stirred overnight at room temperature. The reaction mixture was transferred to a separatory funnel and washed with sat. aq. sol. NaHCO3 (150 mL). The layers were separated and the aqueous layer was extracted with DCM (150 mL). The combined organic layers were dried over Na2SO4 and the solvent was concentrated in a rotary evaporator. The residue was purified using an automated purification system (Cy / EtOAc, gradient 0 to 100%). The tubes containing the product were combined and the solvent was evaporated to give the product D2 as a white foam (48.5 g, 89%). HRMS C 66 H 74 NO 15 Si + [M+NH4] + Calculated: 1148.4822, Found: 1148.487.

[0230] Synthesis of protected trisaccharide D3: To a solution of phenyl 4,6-di-O-benzoyl-2,3-di-O-benzyl-1-thio-β-D-galactopyranoside donor (25.5 g, 38.6 mmol) and acceptor D2 (32 g, 28.3 mmol) in toluene:dioxane (3:1, 515 mL) was added freshly activated MS, and the mixture was stirred at room temperature for 45 min. Then NIS (10.18 g, 45.3 mmol) was added and the reaction mixture was cooled to 0 °C. TMSTOf (0.51 mL, 2.83 mmol) was added and the reaction mixture was stirred at 0 °C for 1.5 h. The reaction mixture was filtered, quenched with NaHCO3 (150 mL), diluted with ethyl acetate (150 mL) and extracted with 0.1 M aq. sol. Na2S2O3, sat. aq. sol. NaHCO3 and brine. The organic layer was dried over Na2SO4 and the solvent was concentrated in a rotary evaporator. Purification by an automated purification system (Cy / EtOAc, gradient 0 to 100%) provided the product D3 as a white foam (43.8 g, 92%) after evaporation of the solvent. HRMS C 100 H 104 NO 22 Si + [M+NH4] + Calculated: 1698.6814, Found: 1699.695.

[0231] Synthesis of tritol D4: At 0 °C, DDQ (10.12 g, 44.6 mmol) was added to a solution of the NAP-protected trisaccharide D3 (60 g, 35.7 mmol) in DCM:MeOH (9:1, 350 mL) in a 500 mL RBF. The reaction mixture was warmed to room temperature and stirred for 2.5 h. The reaction was monitored by TLC (EtOAc in Cy, 3:1). The reaction solution was diluted with DCM (100 mL) and quenched with sat. aq. sol. NaHCO3 (100 mL). The organic layer was washed with saturated aqueous NaHCO3 (2 x 100 mL) and brine (100 mL). The organic layer was dried over Na2SO4, filtered and the filtrate was concentrated in vacuo to obtain the crude product. The crude product was purified by automated flash chromatography (Cy / EtOAc, gradient 0 to 100%). The solvent was concentrated to give the product D4 as a white foam (43.5 g, 79%). HRMS C 89 H 96 NO 22 Si + [M+NH4] + Calcd for: 1558.6188, found: 1559.633.

[0232] Synthesis of tert-hexyldimethylsilyl 4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→4)-6-O-benzoyl-2-O-benzyl-3-O-propionyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranoside D5: The trisaccharide starting material D4 (30 g, 19.46 mmol) was dissolved in anhydrous DCM (195 mL) and LevOH (9.04 g, 78 mmol), followed by the addition of EDCI (14.92 g, 78 mmol) and DMAP (3.82 g, 68.1 mmol). The reaction mixture was stirred at room temperature and monitored by TLC. After 72 h, the mixture was partitioned between DCM and brine. The organic layer was dried over Na2SO4, filtered and concentrated to obtain the crude product. The crude product was loaded onto isolute and purified using an automated purification system and Cy / EtOAc (0-100%) to give the product D5 as a white foam (28.4 g, 89%). HRMS C 94 H 102 NO 24 Si + [M+NH4] + Calcd for: 1656.6556, found: 1657.664. 11H NMR (400 MHz, CDCl3) δ 7.92 - 7.86 (m, 4H), 7.86 - 7.76 (m, 6H), 7.67 - 7.62 (m, 2H), 7.45 - 7.14 (m, 20H), 7.11 - 6.90 (m, 13H), 5.80 (d, J = 2.0 Hz, 1H), 5.74 - 5.67 (m, 1H), 5.33 (s, 1H), 5.21 (dd, J = 10.8, 2.6 Hz, 1H), 5.18 (s, 1H), 5.12 (d, J = 3.7 Hz, 1H), 4.78 - 4.71 (m, 2H), 4.57 - 4.29 (m, 13H), 4.14 (d, J = 2.4 Hz, 1H), 4.10 (d, J = 5.5 Hz, 1H), 4.07 - 4.00 (m, 1H), 3.99 - 3.94 (m, 1H), 3.82 (dd, J = 10.7, 3.5 Hz, 1H), 3.74 (dd, J = 10.1, 3.2 Hz, 1H), 2.63 - 2.53 (m, 2H), 2.53 - 2.39 (m, 2H), 2.00 (s, 3H), 1.51 - 1.45 (m, 1H), 0.71 (dd, J = 6.8, 0.9 Hz, 6H), 0.68 (s, 6H), 0.03 (s, 3H), 0.00 (s, 3H). 13 13C NMR (101 MHz, CDCl3) δ 206.3, 172.5, 166.3, 166.1, 166.1, 165.8, 165.8, 165.3, 138.2, 138.1, 138.0, 133.3, 133.2, 133.1, 133.0, 130.1, 130.0, 129.9, 129.9, 129.8, 129.7, 129.6, 129.4, 128.8, 128.6, 128.5, 128.5, 128.5, 128.5, 128.4, 128.4, 128.4, 128.3, 128.1, 128.0, 127.9, 127.7, 127.5, 101.1, 101.0, 99.7, 85.1, 83.5, 81.8, 77.4, 76.5, 74.5, 73.8, 73.3, 72.8, 72.1, 71.7, 70.7, 69.4, 68.4, 67.8, 63.9, 62.7, 62.1, 38.0, 34.2, 29.9, 28.3, 24.9, 20.2, 20.0, 18.7, 18.6, -2.2, -3.3.

[0233] Synthesis of trisaccharide hemiacetal D6: The TDS-protected trisaccharide starting material D5 (28.4 g, 17.32 mmol) was dissolved in anhydrous DCM (139 mL). AcOH (20.5 mL, 358 mmol) was added. The solution was stirred for 5 min and TBAF (350 mL, 1 M in THF) was added. The reaction mixture was stirred at room temperature and monitored by TLC. After 16 h, the mixture was diluted with water (50 mL) and DCM (100 mL). The reaction mixture was quenched with sat. aq. sol. NaHCO3 (150 mL). The organic layer was separated and washed with brine (150 mL). The organic layer was dried over anhydrous Na2SO4, filtered and evaporated to give the crude product. The crude product was loaded onto isolute and purified using an automated purification system and Cy / EtOAc (0 - 100%) to give the white foam product D6 (25 g, 96%). HRMS C 86 H 84 NO 24 + [M+NH4] + calcd for: 1514.5378, found: 1514.542.

[0234] Synthesis of the trisaccharide imidate ester donor D7: The trisaccharide hemiacetal starting material D6 (14.5 g, 9.68 mmol) was dissolved in anhydrous DCM (97 mL). Cs2CO3 (9.46 g, 29.0 mmol) and 2,2,2-trifluoro-N-phenylimidoyl acetyl chloride (4.02 g, 19.36 mmol) were added. The reaction mixture was stirred at room temperature and monitored by TLC. After 4.5 h, the reaction mixture was filtered through celite. The solvent was evaporated to give the crude product. The crude product was loaded onto isolute and purified using an automated purification system and Cy / EtOAc (0 - 100%, with 0.1% triethylamine) to give the white foam product D7 (13.5 g, 84%). HRMS C 94 H 84 F3NNaO 24 + [M+NH4] + calcd for: 1690.5228, found: 1691.536.

[0235] Synthesis of the undecasaccharide D8: To a solution of the donor D7 (0.44 g, 0.26 mmol) and the acceptor A17 ((see WO2019106201, page 248), 0.83 g, 0.22 mmol) in anhydrous DCM (9 mL) was added MS was used to stir the mixture for 30 min. The reaction mixture was cooled to 0 °C. TMSOTf (0.008 mL, 0.044 mmol) was added and the reaction mixture was stirred at the same temperature for 30 min. The reaction solution was diluted with DCM (10 mL), filtered and quenched by adding sat. aq. sol. NaHCO3 (5 mL). The organic layer was separated, dried over Na2SO4 and filtered. The solvent was evaporated to obtain an oily residue. The crude reaction mixture was purified by an automated purification system using Cy / EtOAc (0 - 100%) to give the product D8 (0.88 g, 78%). MALDI-TOF C 307 H 281 N3NaO 80 + [M+Na] + Calculated value: 5311.7904, experimental value: 5315.83.

[0236] Synthesis of undecasaccharide receptor D9: To a solution of Lev-protected undecasaccharide D8 (1.95 g, 0.375 mmol) in DCM (10 mL) was added a hydrazine hydrate solution (0.12 mL, 3.75 mmol) dissolved in AcOH (0.8 mL) and py (1.2 mL). The resulting reaction mixture was stirred at room temperature for 2 h. The reaction solution was quenched by adding acetone (1 mL) and the solvent was removed in vacuo to obtain a crude product, which was purified by automated flash chromatography using Cy / EtOAc (0 - 100%) as the eluent. The solvent in the tube containing the product D9 (based on TLC) was concentrated in vacuo to give a white foam (1.88 g, 98%). MALDI-TOF C 302 H 275 NNaO 78 + [M+Na-N2] + Calculated value: 5185.7475, experimental value: 5187.08.

[0237] Synthesis of 5-azido-pentyl 4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→4)-6-O-benzoyl-2-O-benzyl-3-O-acetyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-4-[4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→)]-6-O-benzoyl-2-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)]-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranoside D10: The acceptor D9 (1.45 g, 0.28 mmol) was co-evaporated with toluene twice and dissolved in anhydrous toluene (8 mL). Freshly activated MS was added and the mixture was stirred for 30 min. The reaction mixture was cooled to 0 °C and TMSOTf (5 μL, 0.028 mmol) was added dropwise. The donor D7 (0.56 g, 0.33 mmol) was co-evaporated with toluene twice and dissolved in toluene (3 mL). The donor was added dropwise to the reaction mixture over 10 min. Toluene (1 mL) was added to wash the flask containing the donor and the solution was added dropwise to the reaction mixture. The reaction mixture was slowly warmed to 10 °C and maintained for 1.5 h. TLC showed complete consumption of the acceptor. The reaction solution was filtered, diluted with ethyl acetate (10 mL) and quenched by adding sat. aq. sol. NaHCO3 (10 mL). The organic layer was separated, dried over Na2SO4 and filtered. The solvent was evaporated to give an oily residue. The crude reaction mixture was purified by an automated purification system using Cy / EtOAc (0 - 65%) to give the product D10 (1.4 g, 75%). MALDI-TOFC 388 H 354 N3O 101 +[M+H] + Calculated value: 6670.2651, experimental value: 6670.32. 1 H NMR (400 MHz, CDCl3) δ 8.14 - 7.57 (m, 63H), 7.51 - 7.38 (m, 24H), 7.36 - 7.27 (m, 30H), 7.24 - 7.02 (m, 61H), 6.99 - 6.61 (m, 32H), 6.05 (s, 1H), 5.89 - 5.58 (m, 12H), 5.55 - 5.44 (m, 4H), 5.22 - 5.11 (m, 4H), 5.11 - 3.65 (m, 140H), 3.62 - 3.53 (m, 1H), 3.33 - 3.25 (m, 1H), 3.10 (t, J = 6.9 Hz, 2H), 2.24 - 1.84 (m, 4H), 1.71 (s, 3H), 1.52 - 1.39 (m, 4H), 1.33 - 1.23 (m, 2H).

[0238] Synthesis of tetradecasaccharide D11: To a solution of Lev - protected tetradecasaccharide D10 (1.3 g, 0.195 mmol) in DCM (10 mL) was added hydrazine hydrate solution (0.12 mL, 1.95 mmol) and pyridine (1.2 mL) dissolved in acetic acid (0.8 mL). The resulting reaction mixture was stirred at room temperature for 2 h. The reaction solution was quenched by adding acetone (1 mL) and the solvent was removed in vacuo to obtain a crude product, which was purified by automated flash chromatography using Cy / EtOAc (0 - 100%) as the eluent. The solvent from the tube containing product D11 (based on TLC) was concentrated in vacuo to give a white foam (1.08 g, 84%). MALDI - TOFC 383 H 347 NNaO 99 + [M+Na] + Calculated value: 6566.2041, experimental value: 6568.51.

[0239] Synthesis of partially protected tetradecasaccharide D12: At room temperature, an excess of 0.5 M NaOMe solution in methanol (2.93 mL, 1.464 mmol) was added to a solution of tetradecasaccharide D11 (275 mg, 0.042 mmol) in THF (5 mL). The reaction mixture was heated to 55 °C and stirred for 18 h. Then, the solvent was evaporated to dryness in vacuo. Water was added to the reaction mixture and neutralized with acetic acid. The aqueous layer was extracted with EtOAc (3 × 10 mL). The combined organic layers were washed with sat. NaHCO3 (2 × 10 mL), brine (10 mL), dried (Na2SO4), and evaporated in vacuo to give a crude product. SEC purification on LH-20 was carried out using 50% CHCl3 / MeOH as the eluent. The fractions containing the sugar stain-active spots were collected and evaporated, and dried in vacuo (125 mg). 1 1H NMR and MALDI-TOF analysis showed that fewer benzoyl groups remained in the molecule. At room temperature, the substrate (125 mg, 0.031 mmol) was added to THF (5 mL), and 0.5 M NaOMe solution in methanol (1.53 mL, 0.766 mmol) was added to the resulting solution. The reaction mixture was stirred at 60 °C for 18 h. The reaction solvent was evaporated to dryness in vacuo. Water was added to the residue and neutralized with acetic acid. The aqueous layer was extracted with EtOAc (3 × 10 mL). The combined organic layers were washed with sat. NaHCO3 (2 × 10 mL), brine (10 mL), dried (Na2SO4), and evaporated in vacuo to give a crude product. SEC purification on LH-20 was carried out using 30% CHCl3 / MeOH as the eluent. The fractions containing the sugar stain-active spots were collected, evaporated, and dried in vacuo to afford a light yellow fluffy solid D12 (90 mg, 59%). MALDI-TOF C 187 1H 239 4NO 71 + [M+NH4] + Calculated: 3676.5209, Found: 3677.00.

[0240] Synthesis of 5-Amino-pentyl α-D-galactopyranosyl-(1→4)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-4-[α-D-galactopyranosyl-(1→)]-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranoside D13: Deacylated tetrasaccharide D12 (50 mg) was added to a mixture of tBuOH:DCM:PBS (3:0.75:0.38) mL. Pd / C (100 mg) was added and the mixture was hydrogenated under a H2 atmosphere of ~5 bar for 20 h. The reaction mixture was filtered through a PTFE filter using 50% methanol in water (3 × 6 mL). The filtrate was concentrated in vacuo to give a crude product as a white solid. The crude product was purified using a Sep-Pak C18 (0.5 g) column with a water-acetonitrile gradient as the eluent to give the desired product as a fluffy white solid after lyophilization. The product was further purified by SEC column on LH-20 resin using water as the eluent and the fractions containing product D13 were combined, frozen and lyophilized to afford a fluffy white solid (19 mg, 59%). 1 H NMR (400 MHz, D2O) δ 5.20 (s, 5H), 5.11 (d, J = 3.8 Hz, 1H), 5.09 - 5.05 (m, 6H), 5.03 (d, J = 1.5 Hz, 1H), 4.95 (d, J = 3.8 Hz, 1H), 4.43 - 4.37 (m, 4H), 4.35 - 3.53 (m, 82H), 3.03 - 2.96 (m, 2H), 1.74 - 1.60 (m, 4H), 1.50 - 1.38 (m, 2H). MALDI-TOF C 89 H 153 NNaO 71 + [M+Na] + Calculated: 2394.8285, Found: 2394.83.

[0241] Synthesis of 5-azido-pentyl 2,3,5,6-tetra-O-benzoyl-β-D-galactofuranosyl-(1→3)-4-[4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→)]-6-O-benzoyl-2-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-4-[4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→)]-6-O-benzoyl-2-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)]-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranoside D15: The acceptor D11 (0.25 g, 0.038 mmol) was co-evaporated twice with toluene and dissolved in anhydrous toluene (3 mL). Freshly activated MS was added and the mixture was stirred for 30 min. The reaction mixture was cooled to 0 °C and TMSOTf (1.34 μL, 0.038 mmol, in 0.1 mL solution in toluene) was added dropwise. The donor D14 (see WO2019106201, page 198) (0.088 g, 0.114 mmol) was co-evaporated twice with toluene and dissolved in DCM (1.5 mL). The donor was added dropwise to the reaction mixture over 10 min. The reaction mixture was slowly warmed to 10 °C and maintained for 1.5 h. TLC showed complete consumption of the acceptor. The reaction solution was filtered, diluted with ethyl acetate (10 mL) and quenched by adding sat. aq. sol. NaHCO3 (10 mL). The organic layer was separated, dried over Na2SO4 and filtered. The solvent was evaporated to give the crude product. The crude reaction mixture was purified by an automated purification system using Cy / EtOAc (0 - 100%) to give the product D15 as a white foam (0.17 g, 62%). MALDI-TOF C 417H 374 N3O 108 + [M+H] + Calculated value: 7150.3860, experimental value: 7150.75. 1 H NMR (400 MHz, CDCl3) δ 8.16 - 7.27 (m, 128H), 7.26 - 6.71 (m, 102H), 6.04 (s, 1H), 5.91 (s, 1H), 5.87 - 5.73 (m, 8H), 5.71 (s, 1H), 5.66 - 5.57 (m, 2H), 5.57 - 5.41 (m, 5H), 5.17 - 5.10 (m, 4H), 5.10 - 4.91 (m, 13H), 4.89 - 4.78 (m, 2H), 4.75 - 4.17 (m, 68H), 4.11 - 3.94 (m, 20H), 3.88 - 3.74 (dd, J = 23.9, 10.4 Hz, 8H), 3.67 - 3.53 (m, 1H), 3.39 - 3.23 (m, 1H), 3.13 (t, J = 6.9 Hz, 2H), 1.58 - 1.39 (m, 4H), 1.28 (q, J = 7.7, 7.2 Hz, 2H).

[0242] Synthesis of partially protected pentadecasaccharide D16: The sodium methoxide solution (3 mL, 1.5 mmol) in 0.5 M MeOH was added to the solution of pentadecasaccharide D15 (160 mg, 0.022 mmol) in THF (3 mL). The reaction mixture was stirred overnight at 60 °C. The reaction solvent was evaporated to obtain the crude product. The crude product was washed with H2O (2 × 2 mL) and AcOH (0.1 mL in 1 mL H2O). The solid was dissolved in MeOH and evaporated in a rotary evaporator. The residue was washed with cyclohexane and then purified by SEC using LH-20 CHCl3:MeOH (1:2) to obtain the product D16 as a white solid (70 mg, 82%). MALDI-TOF C 193 H 246 N3O 76 + [M+H] + Calculated value: 3821.5471, experimental value: 3821.09.

[0243] Synthesis of 5-Amino-pentyl β-D-galactofuranosyl-(1→3)-4-[α-D-galactopyranosyl-(1→)]-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-4-[α-D-galactopyranosyl-(1→)]-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranoside D17: The debenzoylated pentadecose D16 (40 mg, 0.010 mmol) was dissolved in a mixture of DCM:tBuOH:H2O (2:8:1, 3.5 mL). Pd / C (70 mg) was added and the reaction mixture was purged with hydrogen (5 times) and stirred under a hydrogen pressure of 5 bar for 20 h. Then, the reaction mixture was filtered through a PTFE filter using H2O:ACN (1:1), the organic solvents were evaporated in a rotary evaporator and the crude material was lyophilized. The crude product was purified by Sep-Pak C18 and then by SEC LH-20 using miliQ H2O and lyophilized to give the product D17 as a white solid (12.9 mg, 49%). MALDI-TOF C 95 H 163 NNaO 76 + [M+Na] + Calculated: 2556.8813, Found: 2558.40. 1 1H NMR (400 MHz, D2O) δ 5.20 (s, 6H), 5.14 - 4.96 (m, 9H), 4.43 - 4.36 (m, 4H), 4.35 - 3.54 (m, 88H), 2.99 (t, J = 7.5, 6.4 Hz, 2H), 1.75 - 1.56 (m, 4H), 1.50 - 1.36 (m, 2H).

[0244] Synthesis of 5-azido-pentyl 4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→4)-6-O-benzoyl-2-O-benzyl-3-O-acetyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-4-[4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→)]-6-O-benzoyl-2-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-4-[4,6-di-O-benzoyl-2,3-di-O-benzyl-α-D-galactopyranosyl-(1→)]-6-O-benzoyl-2-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranosyl-(1→3)]-6-O-benzoyl-2,4-di-O-benzyl-α-D-galactopyranosyl-(1→3)-2,5,6-tri-O-benzoyl-β-D-galactofuranoside D18: The acceptor D11 (0.55 g, 0.084 mmol) was co-evaporated twice with toluene and dissolved in anhydrous toluene (4 mL). Freshly activated Add MS and stir for 30 min. Cool the reaction mixture to 0 °C and add TMSOTf (3.02 μL, 0.017 mmol, 0.1 mL solution in toluene) dropwise. Co-evaporate the donor D7 (0.28 g, 0.167 mmol) with toluene twice and dissolve it in DCM (2 mL). Add the donor dropwise to the reaction mixture over 10 min. Slowly warm the reaction mixture to 10 °C and hold for 1.5 h. Filter the reaction solution, dilute with ethyl acetate (10 mL) and quench by adding sat. aq. sol. NaHCO3 (10 mL). Separate the organic layer, dry over Na2SO4 and filter. Evaporate the solvent in vacuo to obtain the crude product. Purify the crude reaction mixture by an automated purification system using Cy / EtOAc (0 - 100%) to give the product D18 as a white foam (0.46 g, 68%). MALDI-TOF C 469 H 426 N3O 122 + [M+H] + Calculated: 8050.7217, Found: 8050.19. 1 1H NMR (400 MHz, CDCl3) δ 8.15 - 7.62 (m, 76H), 7.55 - 7.27 (m, 86H), 7.22 - 6.75 (m, 93H), 6.12 (s, 1H), 6.07 - 5.98 (d, J = 12.2 Hz, 2H), 5.95 - 5.62 (m, 17H), 5.58 - 5.48 (d, J = 11.3 Hz, 5H), 5.26 - 5.14 (m, 6H), 5.14 - 4.91 (m, 18H), 4.90 - 3.72 (m, 148H), 3.66 - 3.54 (m, 1H), 3.33 (d, J = 9.7 Hz, 1H), 3.14 (t, J = 6.9 Hz, 2H), 2.23 - 1.81 (m, 4H), 1.71 (s, 3H), 1.57 - 1.45 (m, 4H), 1.37 - 1.29 (m, 2H).

[0245] Synthesis of the partially protected heptadecasaccharide D19: A solution of sodium methoxide (4 mL, 2 mmol) in 0.5 M MeOH was added to a solution of heptadecasaccharide D18 (160 mg, 0.022 mmol) in THF (3 mL). The reaction mixture was stirred overnight at 60 °C. The reaction solvent was evaporated to give a crude product. The crude product was washed with H2O (2 × 2 mL) and AcOH (0.1 mL in 1 mL H2O). The solid was dissolved in MeOH and the solvent was evaporated to dryness in vacuo. The residue was washed with cyclohexane and then purified by SEC using LH-20 CHCl3:MeOH (1:2) to give the product D19 as a white solid (220 mg, 87%). MALDI-TOF C 226 H 284 N3O 86 + [M+H] + Calculated: 4415.7936, found: 4415.43.

[0246] Synthesis of 5-amino-pentyl α-D-galactopyranosyl-(1→4)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-4-[α-D-galactopyranosyl-(1→)]-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-4-[α-D-galactopyranosyl-(1→)]-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranosyl-(1→3)-α-D-galactopyranosyl-(1→3)-β-D-galactofuranoside D20: The debenzoylated heptadecasaccharide D19 (37 mg, 0.084 mmol) was dissolved in a mixture of DCM:tBuOH:H2O (2:8:1, 2.75 mL). Pd / C (40 mg) was added and the reaction mixture was purged with hydrogen (5 times) and stirred under a hydrogen pressure of 5 bar for 20 h. Then, the reaction mixture was filtered through a PTFE filter using H2O:ACN (1:1), the solvent was evaporated in a rotary evaporator and the crude material was lyophilized. The crude product was purified by Sep-Pak C18 and then by SEC LH-20 using miliQ H2O and lyophilized to give the product D20 as a white solid (5.3 mg, 22%). MALDI-TOF C 107 H 183 NNaO 86+ [M+Na] + Calculated value: 2880.9869, experimental value: 2882.40. 1 H NMR (400 MHz, D2O) δ 5.25 - 5.17 (m, 6H), 5.13 - 4.98 (m, 10H), 4.94 (d, J=3.5 Hz, 1H), 4.41 (s, 4H), 4.33 - 4.03 (m, 40H), 3.95 - 3.80 (m, 32H), 3.76 - 3.56 (m, 28H), 2.98 (t, J=7.2 Hz, 2H), 1.75 - 1.58 (m, 4H), 1.51 - 1.36 (m, 2H).

[0247] Synthesis of the conjugate: Synthesis of the NHS ester of compound D13 (D13 - adipic acid - NHS ester) Dissolve compound D13 (4 mg, 1.686 μmol) in DMSO - H2O (100 μL - 10 μL) in a 15 mL falcon tube at room temperature. Add triethylamine (8 μL, 0.059 mmol) thereto. Add adipic acid - NHS ester (bis(2,5 - dioxopyrrolidin - 1 - yl) adipate) (11.5 mg, 0.034 mmol) in DMSO (100 μL) and stir at room temperature for 2 h. Precipitate D13 - adipic acid - NHS ester by adding EtOAc (5 mL) and centrifuge. Wash the precipitate with EtOAc (3 mL × 2) and dry in vacuo to obtain a white solid (4 mg, 91%) and use it for the next step.

[0248] D13 and CRM 197 conjugate of (D13 - adipic acid - CRM 197 ester or D13 - CRM 197 *) Synthesis: Dissolve compound D13 - adipic acid - NHS ester (4 mg, 1.54 μmol) in 0.1 M NaPi buffer (pH 7.0, 150 μL) in a 15 mL falcon tube. Add freshly washed CRM 197 (obtained from EirGenix, Inc., Taiwan Province, China; expression system: Escherichia coli) (2 mg, 0.034 μmol) drop - by - drop to it. Rinse the vial with 0.1 M NaPi buffer (pH 7.0, 50 μL) and transfer to the reaction mixture in the falcon tube, and stir at room temperature for 20 h. The obtained D13 - adipic acid - CRM 197The ester solution was transferred to an Amicon Ultra vial (10 kDa, MWCO) and centrifuged for 5 minutes at 2 - 8 °C. 300 μL of 0.1 M NaPi was added to the reaction falcon tube, rinsed and transferred to the filter and centrifuged again. Additional washes were performed more than 5 times using a TBS buffer (pH 7.4) solution. After the final wash, the conjugate was sterile filtered and stored at 2 - 8 °C in TBS (1.0 mL) (pH 7.4). The loading experimental value using MALDI - TOF MS was 12.3. The conjugate was analyzed using SDS - PAGE, BCA, SEC - HPLC and endotoxin testing.

[0249] Synthesis of the conjugate with BSA (D13 - adipic acid - BSA ester or D13 - BSA*) In a 15 mL falcon tube, the compound D13 - adipic acid - NHS ester (1.7 mg, 0.654 μmol) was dissolved in 0.1 M NaPi buffer (pH 7.0, 150 μL). Freshly washed BSA (obtained from Sigma Aldrich, heat shock fraction, pH 7, ≥98%; product number: A7906) (3 mg, 0.045 μmol) in 0.1 M NaPi buffer (pH 7.0, 150 μL) in a vial was added dropwise thereto. The vial was rinsed with 0.1 M NaPi buffer (pH 7.0, 50 μL) and transferred to the reaction mixture in the falcon tube, and stirred at room temperature for 20 h. The obtained D13 - adipic acid - BSA ester solution was transferred to an Amicon Ultra vial (10 kDa, MWCO) and centrifuged for 5 minutes at 2 - 8 °C. 300 μL of 0.1 M NaPi was added to the reaction falcon tube, rinsed and transferred to the filter and centrifuged again. Additional washes were performed more than 5 times using a 1×PBS buffer (pH 7.4) solution. After the final wash, the conjugate was sterile filtered and stored at 2 - 8 °C in TBS (0.5 mL) (pH 7.4). The loading experimental value using MALDI - TOF MS was 6.96. The conjugate was analyzed using SDS - PAGE and SEC - HPLC.

[0250] Synthesis of the NHS ester of compound D17 (D17 - adipic acid - NHS ester) The NHS ester of compound D17 was synthesized using a similar procedure as described above for D13 - adipic acid - NHS ester and dried in vacuo to obtain a white solid (7 mg, 92%) and used in the next step.

[0251] Conjugate of D17 with CRM 197 (D17 - adipic acid - CRM197 Ester or D17-CRM 197 *) was synthesized using D17-adipic acid-NHS ester and CRM 197 Using the similar procedure described above for D13-adipic acid-CRM 197 ester, the conjugate of D17 and CRM was synthesized 197 (3 mg), and the experimental value of loading using MALDI-TOF MS was 9.69

[0252] The conjugate with BSA (D17-adipic acid-BSA ester or D17-BSA*) was synthesized using D17-adipic acid-NHS ester and BSA. Using the similar procedure described above for D13-adipic acid-BSA ester, the conjugate of D17 and BSA (1 mg) was synthesized, and the experimental value of loading using MALDI-TOF MS was 12.78

[0253] Synthesis of the NHS ester of compound D20 (D20-adipic acid-NHS ester) Using the similar procedure described above for D13-adipic acid-NHS ester, the NHS ester of compound D20 was synthesized and dried in vacuo to obtain a white solid (7 mg, 93%), and was used for the next step

[0254] The conjugate of D20 and CRM 197 (D20-adipic acid-CRM 197 ester or D20-CRM 197 *) was synthesized using D20-adipic acid-NHS ester and CRM 197 Using the similar procedure described above for D13-adipic acid-CRM 197 ester, the conjugate of D20 and CRM was synthesized 197 (3 mg), and the experimental value of loading using MALDI-TOF MS was 7.45

[0255] The conjugate with BSA (D20-adipic acid-BSA ester or D20-BSA*) was synthesized using D17-adipic acid-NHS ester and BSA. Using the similar procedure described above for D13-adipic acid-BSA ester, the conjugate of D17 and BSA (1 mg) was synthesized, and the experimental value of loading using MALDI-TOF MS was 9.41

[0256] II. Biology Materials · ELISA plates (high binding, EIA / RIA plates, 96-well, flat bottom with low evaporation lid, company: 3361) · Detection antibody: Goat anti-rabbit IgG peroxidase conjugate (Sigma, #A4914) · Blocking solution: Commercial blocking reagent (Roche, cat.no.11112589001) · Antibody diluent: PBS + 1% BSA (w / v) · Wash buffer: PBS + 0.1% Tween 20 (PBS-T) · Developing solution: 1Step TM Ultra TMB-ELISA developer. (ThermoScientific, Cat#: 34028) · Stop solution - 2M sulfuric acid (H2SO4) · Microplate reader: FLUOstar Omega (BMG LABTECH) · Software: GraphPad Prism version 7 or higher for data plotting and analysis · Alum: Aluminum hydroxide adjuvant ( HPA), Chemtrade · QuantiPro TM BCA assay kit (SIGMA) Product: QPBCA-1KT; Lot number: SLBR7451V; P code: 1002296464 · Mini- TGX TM Gel - 10%, 10 wells (30 μL / well) Control number: 64175708 · GelCode TM Blue Safe protein stain; ThermoScientific; Ref: 1860957; Lot number: TA260266

[0257] Method: Bacterial strains and LPS. Klebsiella pneumoniae strains with different LPS (O-antigen) were used to isolate and purify the corresponding LPS. The purified LPS was used as the coating antigen in the enzyme-linked immunosorbent assay (ELISA). LPS was isolated using a commercial LPS extraction kit (JH Science) according to the manufacturer's protocol. Table 1. Klebsiella pneumoniae strains used for LPS isolation. # LPS / O-antigen 1NCTC 9148 O2a 2PCM27 Galactan-III (O2afg)

[0258] Formulation of candidate vaccines for immunization. All preparations are made under aseptic conditions. The drug substance (DS) and buffer (10 mM TRIS-HCl, pH 7.4) are mixed with an appropriately pre-calculated dilution factor (see below) to obtain the desired glycan dose, excluding the required volume of aluminum hydroxide adjuvant (0.25 mg / mL). The DS-buffer mixture is gently mixed and the aluminum hydroxide adjuvant (“aluminum”) stock is added until the final aluminum concentration is 0.250 mg / mL of aluminum. The mixture is immediately mixed by gentle pipetting and then mixed at 250 rpm on a horizontal shaker at room temperature for 2 h. Aliquots are stored in type 1 glass vials at 4 °C until further use.

[0259] Prepare the vaccine described above to contain the following expected glycan doses (e.g., 2 μg glycan / injection). Determine the average loading factor (in antigen moles / carrier protein) of the glycan antigen by subtracting the measured molecular weight (m / z = 1) of CRM 197 from the measured molecular weight (m / z = 1) of the DS by MALDI-TOF MS, and then divide this mass difference by the theoretical molecular weight of the glycan antigen containing the linker (here: alkyl) and spacer (here: adipoyl) moieties. Multiply the resulting loading factor by the theoretical molecular weight of the glycan antigen (excluding the linker and spacer moieties) to provide the total mass of glycan attached to an average of each DS molecule. Divide the total mass of this glycan by the measured molecular weight of the CRM 197 protein to produce the mass ratio of glycan to protein of the DS. Multiply this ratio by the measured protein concentration of the DS (as determined by a BCA assay kit (Sigma) according to the manufacturer's protocol) to produce the glycan concentration of the DS. To obtain the dilution factor required to dilute the DS to obtain the desired glycan dose / immunization, divide the glycan concentration of the DS by the desired glycan concentration (e.g., 4 μg / mL glycan concentration for a 2 μg glycan dose for a rabbit with an injection volume of 500 μL). Then use this dilution factor to dilute the DS for the final volume of the vaccine formulation.

[0260] Immunization: Female Zika rabbits are immunized via the intramuscular (i.m.) route using an injection volume of 500 μL / dose. The animals are maintained under specific pathogen-free conditions and provided with water and food ad libitum.

[0261] ELISA: Use antigen-coated plates: Coat with isolated LPS. Dissolve LPS in isopropanol to a concentration of 10 μg / mL and use 100 μL for coating so that each well is coated with 1 μg LPS. Allow the LPS solution to evaporate overnight inside a biosafety cabinet at room temperature. Blocking: Block the plates with 100 μL of commercial blocking solution and incubate for 1 h at room temperature. After blocking, wash the plates 3 times with PBS with 0.1% (v / v) Tween-20 (PBS-T). Incubate with diluted serum: Dilute pooled or individual sera from different time points to their respective dilutions using 1% BSA (w / v) in PBS. Add 50 - 100 μL of diluted serum in duplicate to ELISA wells and incubate for 1 h at room temperature. 100 μL / well of 1% BSA (w / v) in PBS is used as a blank. After incubation with serum, wash the plates 3 times with PBS-T. Incubate with detection antibody: Dilute anti-rabbit IgG HRP conjugate 1:10,000 in 1% BSA (w / v) in PBS and add 100 μL / well, and incubate for 30 minutes at room temperature. After incubation with the detection antibody, wash the plates 3 times with PBS-T. Substrate addition: Add 100 μL of TMB substrate to each well and incubate for approximately 15 min. Terminate the reaction by adding 50 μL / well of 2 M H2SO4. Measure the absorbance at 450 nm using a plate reader. Analyze the absorbance values using GraphPad Prism software.

[0262] Challenge experiment: Twenty-four hours and 1 hour before bacterial challenge, female CD-1 mice received pooled post-immune antiserum from rabbits (30 - 250 μL) via the intraperitoneal route. Before bacterial challenge, mice were pretreated with galactosamine at 20 mg / animal. Infect the mice with 2 × 10 7 colony-forming units (CFU) of Klebsiella pneumoniae strain NCTC 9163 (O2a) or 1 × 10 8 CFU strain ST258 (Gal-III) via the intraperitoneal route. Observe the mice for 24 h to obtain clinical scores and euthanize at the humane endpoint.

[0263] SEQ ID NO:1 (CRM 197 ) SEQ ID NO:2 (Diphtheria toxin (Uniprot ID:P00587)) SEQ ID NO:3 (Tetanus toxin (Uniprot ID:P04958)) SEQ ID NO:4 (Cholera toxin B subunit (Uniprot ID:P01556)) SEQ ID NO:5 (Neisseria meningitidis outer membrane protein (OMP) (Uniprot ID:Q51229)) SEQ ID NO:6 (Capsid protein of bacteriophage Qβ (Uniprot ID:P03615))

Claims

1. An immunogenic compound comprising at least one oligosaccharide heterozygous antigen having the structure of formula (I) wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; and n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; or a pharmaceutically acceptable salt thereof.

2. The immunogenic compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is or; m is 4, n is 3 and R is OH.

3. The immunogenic compound or a pharmaceutically acceptable salt thereof according to claim 1, wherein m is 4, n is 2 and R is OH.

4. An immunogenic compound of formula (II) wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; i is from 1 to 28; and -L-T- represents a linker L and a spacer T which together form a bridge having a backbone of 5 to 25 atoms covalently linked together, the length of which forms the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of a lysine residue at the carrier protein CRM 197 wherein these atoms of the backbone are selected from the group consisting of carbon, nitrogen, oxygen and sulfur; or a pharmaceutically acceptable salt thereof.

5. The immunogenic compound or a pharmaceutically acceptable salt thereof according to claim 4, wherein m is 4, n is 2 and R is OH; m is 4, n is 2 and R is or; m is 4, n is 3 and R is OH.

6. The immunogenic compound or a pharmaceutically acceptable salt thereof according to claim 4, wherein m is 4, n is 2 and R is OH.

7. The immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of claims 4 to 6, wherein -L-T- represents a linker L and a spacer T which together form a bridge having a backbone of 5 to 25 atoms covalently linked together, the length forming the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at 197 the carrier protein CRM, and having at least one double bond, wherein these atoms of the main chain are selected from the group consisting of carbon, nitrogen, oxygen and sulfur, and wherein the main chain may be substituted by one or more (especially 1, 2, 3 or 4) substituents independently selected from oxo, (C 1-4 )alkyl, fluorine and (C 1-2 )alkoxy (especially oxo), and wherein a part of the main chain may optionally be part of a 4-, 5- or 6-membered ring selected from the following:

8. The immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of claims 4 to 7, wherein the backbone of the bridge has a length of 8 to 20, preferably 8 to 16 atoms covalently linked together, the length forming the shortest distance between the oxygen at the reducing end C1 of the oligosaccharide and the amino nitrogen of the lysine residue at 197 the CRM of the carrier protein.

9. The immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of claims 4, 5, 6 or 8, wherein L represents *-(CH2) a -NH-; wherein a is from 2 to 10; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 ) fluoroalkylene-NH-, wherein the fluoroalkylene is a saturated straight chain; *-(CH2) c NHC(O)(CH2) c’ -NH-, where c and c' are each independently 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d' are each independently from 2 to 6; *-(CH2) e -C(O)-NH-(CH2) e’ -NH-; wherein e is from 1 to 10 and e' is from 2 to 10; or *-(CH2) f -O-NH- wherein f is from 2 to 10; or L-T represents *-(CH2) g -S-R 1 , where g is from 2 to 10; and T represents -C(O)-(CH2) h -C(O)-, where h is from 0 to 10; -C(O)-CH2CH2-(OCH2CH2) j -C(O)-, where j is from 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)-, where k is 0 or 1, k' is 0 or 1, and k'' is 1, 2 or 3; where l is 1 or 2; or where p is from 1 to 4, and p' is 1 or 2; and R 1 represent where q is 2 or 3; where r is 2 or 3; or 10. The immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of claims 4, 5 or 6, wherein L represents *-(CH2)2-NH-, *-(CH2)3-NH-, *-(CH2)4-NH-, *-(CH2)5-NH- or *-(CH2)6-NH-; preferably *-(CH2)5-NH-; and T represents -C(O)-C(O)-, -C(O)-CH2-C(O)-, -C(O)-(CH2)2-C(O)-, -C(O)-(CH2)3-C(O)-, -C(O)-(CH2)4-C(O)-, -C(O)-(CH2)5-C(O)- or -C(O)-(CH2)6-C(O)-; preferably -C(O)-(CH2)4-C(O)-.

11. The immunogenic compound or a pharmaceutically acceptable salt thereof according to any one of claims 4 to 10, wherein i is from 6 to 15, or a pharmaceutically acceptable salt thereof.

12. The immunogenic compound or a pharmaceutically acceptable salt thereof according to claim 4, which is selected from the group consisting of the structures of formula (IIa), (IIb) and (IIc): wherein i is from 1 to 28; preferably from 6 to 15.

13. A pharmaceutical composition comprising, as an active ingredient, an immunogenic compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt thereof, and at least one therapeutically inert excipient.

14. The pharmaceutical composition according to claim 13, which further comprises an adjuvant.

15. The immunogenic compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt thereof, for use in pharmacy, in particular as a vaccine.

16. The immunogenic compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt thereof, for the prevention and / or treatment of Klebsiella pneumoniae infection.

17. A multivalent vaccine comprising an immunogenic compound according to any one of claims 1 to 12 or a pharmaceutically acceptable salt thereof.

18. An intermediate compound for preparing an immunogenic compound according to any one of claims 4 to 12, which has the structure of formula (III): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; L 1 represent *-(C 2-10 )alkylene-NH2, preferably *-(CH2) a -NH2; wherein a is from 2 to 10, more preferably l is 5; *-(CH2CH2O) b -CH2CH2NH2, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH2; *-(C 2-10 ) fluoroalkylene-NH2; *-(CH2) c NHC(O)(CH2) c’ -NH2, where c and c' are each independently from 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH2, where d and d' are each independently from 2 to 6; *-(C 1-10 )alkylene - C(O)-NH-(C 2-10 )alkylene - NH2; *-(C 2-10 )alkylene-O-NH2; or *-(C 2-10 )alkylene - SH; or a pharmaceutically acceptable salt thereof.

19. An intermediate compound for preparing an immunogenic compound according to any one of claims 4 to 12 or a pharmaceutically acceptable salt thereof, which has the structure of formula (IV): wherein R is OH or m is 3, 4, 5, 6, 7 or 8; preferably 3, 4, 5 or 6; n is 1, 2, 3, 4, 5 or 6; preferably 2, 3 or 4; L represents *-(C 2-10 )alkylene-NH-, preferably *-(CH2) a -NH-, where a is from 2 to 10, more preferably 5; *-(CH2CH2O) b -CH2CH2NH-, where b is 1, 2 or 3; *-CH2CH2S-CH2CH2NH-; *-(C 2-10 ) fluoroalkylene-NH-; *-(CH2) c NHC(O)(CH2) c’ -NH-, wherein c and c' are each independently from 2 to 6; *-(CH2) d NHC(O)NH(CH2) d’ -NH-, where d and d' are each independently from 2 to 6; *-(C 1-10 )alkylene - C(O) - NH - (C 2-10 )alkylene - NH -; or *-(C 2-10 )alkylene - O - NH-; T 1 represent -C(O)-(C 0-10 )alkylene - C(O)X; -C(O)-CH2CH2-(OCH2CH2) j -C(O)X, where j is from 1 to 5; -C(O)-CH2(CH2) k -(SCH2(CH2) k’ ) k” -C(O)X, where k is 0 or 1, k' is 0 or 1, and k" is 1, 2, or 3; where l is 1 or 2; or wherein p is from 1 to 4, preferably 1, and p' is 1 or 2; -C(O)X represents -C(O)OH or an activated ester, wherein preferably X represents and Y represents Me, Et, Bu or -(CH2CH2O)3CH3.

Images