Precision glycoconjugates as therapeutic tools

Abstract

The present description relates to glycoconjugates, glycoconjugate immunogens and glycoconjugate vaccines comprising carbohydrate antigens coupled to immunogenic carrier proteins, or materials used for detection and screening of resulting antibodies. Improved methods of more directly and precisely conjugating carbohydrate antigens to free thiol groups of immunogenic carrier proteins are described, including “click-chemistry” approaches based on photocatalytic thiol-ene reactions.

1 - 71 . (canceled) 
     
     
         72 . A synthetic glycoconjugate comprising a carbohydrate antigen coupled to a carrier material via a linker, the carbohydrate antigen being an unprotected carbohydrate antigen that is water-soluble, wherein:
 (a) the synthetic glycoconjugate has the structure:   
       
         
           
           
               
               
           
         
         
           wherein: CA is the carbohydrate antigen; S-CM is the carrier material having z sulfur atoms available for conjugation, wherein z is at least 1; X is O, S, NR 1 , or CH 2 ; R 1  is —H, —COH, —COCH 3 , or —COEt; n is 1, 2, 3, 4, or 5; and R 2  is H or Me; or a stereoisomer thereof; or 
         
         (b) the synthetic glycoconjugate has the structure: 
       
       
         
           
           
               
               
           
         
         
           wherein: CA is the carbohydrate antigen; S-CM is the carrier material having z sulfur atoms available for conjugation, wherein z is at least 1; X is S, NR 1 , CH 2  or O; R 1  is —H, —COH, —COMe, or —COEt; n is 1, 2, 3, 4, or 5; R 2  is H or Me; q is 1, 2, 3, 4, or 5; R 3  and R 4  are each a hydrogen atom and m is 1, 2, 3, 4 or 5, or R 3  and R 4  form together a radical —CO—CH 2 — or a radical —CO—CH 2 —CH 2 — with the carbonyl linked to the nitrogen atom, and m is 1; or a stereoisomer thereof; or 
         
         (c) the synthetic glycoconjugate has the structure: 
       
       
         
           
           
               
               
           
         
         
           wherein: CA is the carbohydrate antigen; y is at least 1; and when y is more than 1, CA are identical or different; [S] z -CM is the carrier material having z sulfur atoms available for conjugation, wherein z is at least equal to y; and L is a linker selected from the group consisting of linkers having the structure: 
         
       
       
         
           
           
               
               
           
         
         
           wherein: X is O, S, NR 1 , or CH 2 ; R 1  is —H, —COH, —COCH 3 , or —COEt; n is 1, 2, 3, 4, or 5; and R 2  is H or Me; and when y is more than 1, L are identical or different; or a stereoisomer thereof; or 
         
         (d) the synthetic glycoconjugate has the structure: 
       
       
         
           
           
               
               
           
         
         
           wherein: CA is the carbohydrate antigen; y is at least 1; and when y is more than 1, CA are identical or different; S-CM is the carrier material having z sulfur atoms available for conjugation, wherein z is at least 1 and is at least equal to y; and L is a linker selected from the group consisting of linkers having the structure: 
         
       
       
         
           
           
               
               
           
         
         
           wherein: X is S, NR 1 , CH 2  or O; R 1  is —H, —COH, —COMe, or —COEt; n is 1, 2, 3, 4, or 5; R 2  is H or Me; q is 1, 2, 3, 4, or 5; R 3  and R 4  are each a hydrogen atom and m is 1, 2, 3, 4 or 5, or R 3  and R 4  form together a radical —CO—CH 2 — or a radical —CO—CH 2 —CH 2 — with the carbonyl linked to the nitrogen atom, and m is 1; and when y is more than 1, L are identical or different; or a stereoisomer thereof; or 
         
         (e) the synthetic glycoconjugate has the structure: 
       
       
         
           
           
               
               
           
         
         
           wherein: CA is the carbohydrate antigen; y is at least 1; and when y is more than 1, CA are identical or different; [S] z -CM is the carrier material having z sulfur atoms available for conjugation, wherein z is at least equal to y; and L is a linker selected from the group consisting of linkers having the structure: 
         
       
       
         
           
           
               
               
           
         
         
           wherein: X is S, NR 1 , CH 2  or O; R 1  is —H, —COH, —COMe, or —COEt; n is 1, 2, 3, 4, or 5; R 2  is H or Me; q is 1, 2, 3, 4, or 5; r is 1, 2, 3, 4 or 5; R 5  is S-CM, a covalent bond, or a radical of structure: 
         
       
       
         
           
           
               
               
           
         
         
           wherein R 3  and R 4  are each a hydrogen atom and m is 1, 2, 3, 4 or 5, or R 3  and R 4  form together a radical —CO—CH 2 — or a radical —CO—CH 2 —CH 2 — with the carbonyl linked to the nitrogen atom, and m is 1; and when y is more than 1, L are identical or different; or a stereoisomer thereof. 
         
       
     
     
         73 . The synthetic glycoconjugate of  claim 72  having the structure as defined in (e), wherein the linker has the structure: 
       
         
           
           
               
               
           
         
       
       wherein: X is S, NR 1 , CH 2  or O; R 1  is —H, —COH, —COMe, or —COEt; n is 1, 2, 3, 4, or 5; R 2  is H or Me; q is 1, 2, 3, 4, or 5; and r is 1, 2, 3, 4 or 5. 
     
     
         74 . The synthetic glycoconjugate of  claim 72  having the structure as defined in (e), wherein the linker has the structure: 
       
         
           
           
               
               
           
         
       
       wherein: X is S, NR 1 , CH 2  or O; R 1  is —H, —COH, —COMe, or —COEt; n is 1, 2, 3, 4, or 5; R 2  is H or Me; q is 1, 2, 3, 4, or 5; and r is 1 or 2. 
     
     
         75 . The synthetic glycoconjugate of  claim 72 , wherein the carbohydrate antigen is or comprises a viral saccharide antigen, a tumor associated carbohydrate antigen (TACA), or a bacterial capsular polysaccharide (CPS). 
     
     
         76 . The synthetic glycoconjugate of  claim 72 , wherein the carbohydrate antigen is, is from, or comprises Tn antigen, Thomsen-Friedenreich (TF) antigen, a sialylated analog of Tn or TF, or any combination thereof. 
     
     
         77 . The synthetic glycoconjugate of  claim 76 , wherein the carbohydrate antigen is, is from, or comprises: S-Tn, (2,3)-S-TF, (2,6)-S-TF, or any combination thereof. 
     
     
         78 . The synthetic glycoconjugate of  claim 72 , wherein the carbohydrate antigen is, is from, or comprises: Globo H, GD2, GD3, GM2, GM3, N-glycolyl-GM3, Lea, sLea, Lex, sLex, or any combination thereof. 
     
     
         79 . The synthetic glycoconjugate of  claim 72 , wherein the carbohydrate antigen is, is from, or comprises a Pneumococcal and/or Streptococcal polysaccharide serotype, meningococcal CPS, or influenza CPS saccharide antigen. 
     
     
         80 . The synthetic glycoconjugate of  claim 72 , wherein the carrier material is or comprises a polymer, a polypeptide, a carrier protein, a solid support, a particle, or any other material having at least one or more a free thiol group suitable for conjugation to the carbohydrate antigen via a photocatalytic thiol-ene reaction. 
     
     
         81 . The synthetic glycoconjugate of  claim 72 , wherein the conjugate material is coupled to at least two of the same carbohydrate antigen or to more than one type of carbohydrate antigen, thereby producing a multi-valent synthetic glycoconjugate. 
     
     
         82 . The synthetic glycoconjugate of  claim 72 , wherein the conjugate material is a carrier protein that is immunogenic when administered to a subject, and wherein conjugation of the carbohydrate antigen to the carrier protein increases the immunogenicity of the carbohydrate antigen upon administration to the subject, as compared to administration of the unconjugated carbohydrate antigen. 
     
     
         83 . The synthetic glycoconjugate of  claim 82 , wherein the carrier protein is not denatured or retains its native conformation, antigenicity, and/or structure when conjugated to the carbohydrate antigen. 
     
     
         84 . The synthetic glycoconjugate of  claim 82 , wherein the carbohydrate antigen is not cleavable from the carrier protein by an endogenous enzyme of the subject. 
     
     
         85 . The synthetic glycoconjugate of  claim 82 , wherein the carbohydrate antigen comprises a B cell epitope, and/or induces a humoral immune response in the subject; and/or comprises T cell epitope, and/or induces a cell-mediated immune response in the subject. 
     
     
         86 . The synthetic glycoconjugate of  claim 82 , wherein the carrier protein comprises a human T cell epitope, and/or induces a cell-mediated immune response in the subject. 
     
     
         87 . The synthetic glycoconjugate of  claim 82 , wherein the carrier protein is an immunogenic peptide. 
     
     
         88 . The synthetic glycoconjugate of  claim 82 , wherein the carrier protein is, is from, or comprises: Tetanus Toxoid (TT), Diphtheria Toxoid (DT), cross-reacting material 197 (CRM197), Meningococcal Outer Membrane Protein Complex (OMPC),  H. Influenzae  Protein D (HiD), a cytokine, Tetanus Toxin 831-844 (SEQ ID NO:01 or SEQ ID NO:02), an MHC-restricted peptide, albumin, or an immunogenic fragment thereof. 
     
     
         89 . The synthetic glycoconjugate of  claim 72 , wherein the carbohydrate antigen is coupled to the carrier material by providing an unprotected carbohydrate antigen covalently linked via a linker to a terminal alkene (alkenyl carbohydrate antigen), the terminal alkene being directly conjugatable to a thiol group via a thiol-ene reaction; providing a carrier material having one or more free thiol groups; and performing a photocatalytic thiol-ene reaction to directly conjugate the unprotected carbohydrate antigen to the carrier material at the one or more free thiol groups, thereby producing the synthetic glycoconjugate. 
     
     
         90 . An immunogenic composition comprising the synthetic glycoconjugate of  claim 82  formulated with a pharmaceutically acceptable excipient, and/or an adjuvant. 
     
     
         91 . A method for detecting or screening a biological sample for the presence of an antibody against a carbohydrate antigen of interest, the method comprising: contacting a biological sample from a subject having or suspected of having the antibody with the synthetic glycoconjugate as defined in  claim 72 , wherein the carbohydrate antigen is the carbohydrate antigen of interest; and detecting the presence of antibody bound to the carbohydrate antigen.