Method for improving immunogenicity of protein/peptide antigen
Disclosed is a method for improving the immunogenicity of a protein/peptide antigen, the method comprising conjugating a protein/peptide antigen with a sugar to form a sugar-protein/peptide antigen conjugate, which has improved immunogenicity compared to an unconjugated protein/peptide antigen. In particular, the method involves conjugating a pathogen, such as a viral surface protein antigen or a fragment thereof, with a polysaccharide, in particular a capsular polysaccharide of Streptococcus pneumonia . The conjugate with improved immunogenicity can be used to prevent or treat diseases caused by pathogens, in particular diseases caused by coronaviruses.
1 . A method for improving the immunogenicity of a protein/peptide antigen, comprising forming a glyco-protein/peptide antigen conjugate by conjugating the protein/peptide antigen with a saccharide,
wherein the saccharide is capsular polysaccharide of Streptococcus pneumonia,
wherein the protein/peptide antigen is a virus pathogen-associated protein/peptide antigen selected from human respiratory syncytial virus glycoprotein G (RSV-gpG), hepatitis C virus envelope glycoprotein E1 protein (ECV-E1), hepatitis C virus envelope E2 protein (HCV-E2), influenza B hemagglutinin protein (FLU-B-HA1), influenza A H5N1 hemagglutinin (H5N1-HA), ebola virus glycoprotein (Ebola-GP), ebola virus glycoprotein GP1 (Ebola-GP1), zika virus envelope protein (ZIKV-E), and/or coronavirus spike protein.
2 . The method as claimed in claim 1 , wherein the saccharide is
capsular polysaccharide of Streptococcus pneumoniae serotype 14, 6B or 7F.
3 . The method as claimed in claim 1 , wherein the protein/peptide antigen is a protein/peptide comprising
viral antigen of
coronavirus spike protein receptor binding region RBD.
4 . The method as claimed in claim 3 , wherein the coronavirus is SARS-COV-2 or Middle East respiratory syndrome coronavirus.
5 . The method as claimed in claim 3 , wherein the protein/peptide antigen is a fusion protein
which is SARS-COV-2 RBD-mFc; SARS-COV-2 RBD-his; or MERS-COV RBD-his; and
the Fc fragment is a human or murine IgG Fc fragment.
6 . The method as claimed in claim 4 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:1, SEQ ID NO: 2 or SEQ ID NO:3.
7 . The method as claimed in claim 1 , wherein the molecular weight of the conjugate is 400-14000 KDa.
8 . The method as claimed in claim 1 , wherein the protein/peptide antigen is a fusion protein
which is RSV-gpG-his.
9 . The method as claimed in claim 1 , wherein the protein/peptide antigen comprises the sequence as set forth as SEQ ID NO:4 or SEQ ID NO:12.
10 . The method as claimed in claim 1 , wherein the protein/peptide antigen is a fusion protein
which is Flu-B-HA1-his or H5N1-HA-his.
11 . The method as claimed in claim 1 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:7, SEQ ID NO: 15, SEQ ID NO: 8 or SEQ ID NO: 16.
12 . The method as claimed in claim 1 , wherein the protein/peptide antigen is a fusion protein
which is Ebola-GP-Fc or Ebola-GP1-his;
wherein the Fc is a human or murine IgG Fc fragment.
13 . The method as claimed in claim 1 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO: 9, SEQ ID NO: 17, SEQ ID NO: 10 or SEQ ID NO: 18.
14 . The method as claimed in claim 1 , wherein the protein/peptide antigen is a fusion protein
which is ZIKV-E-Fc; wherein
Fc is a human or murine IgG Fc fragment; or
the fusion protein is HCV-E2-his and/or HCV-E1-his.
15 . The method as claimed in claim 1 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:11, SEQ ID NO: 19, SEQ ID NO: 6, SEQ ID NO:14, SEQ ID NO:5 or SEQ ID NO: 13.
16 . The method as claimed in claim 1 , wherein the glyco-protein/peptide antigen is further conjugated with a protein carrier.
17 . The method as claimed in claim 16 , wherein the protein carrier is a tetanus toxoid, a tetanus toxoid fragment C, a tetanus toxoid non-toxic mutant, a diphtheria toxoid, CRM197, or other non-toxic mutants of diphtheria toxoid.
18 . A glyco-protein/peptide antigen conjugate with a saccharide increased immunogenicity compared to an unconjugated protein/peptide antigen;
wherein the saccharide is capsular polysaccharide of Streptococcus pneumonia,
wherein the protein/peptide antigen is a virus pathogen-associated protein/peptide antigen selected from human respiratory syncytial virus glycoprotein G (RSV-gpG), hepatitis C virus envelope glycoprotein E1 protein (ECV-E1), hepatitis C virus envelope E2 protein (HCV-E2), influenza B hemagglutinin protein (FLU-B-HA1), influenza A H5N1 hemagglutinin (H5N1-HA), ebola virus glycoprotein (Ebola-GP), ebola virus glycoprotein GP1 (Ebola-GP1), zika virus envelope protein (ZIKV-E), and/or coronavirus spike protein.
19 . The conjugate as claimed in claim 18 , wherein the saccharide is
capsular polysaccharide of Streptococcus pneumoniae serotype 14, 6B
or 7F.
20 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen is a protein/peptide comprising
viral antigens of
the coronavirus spike protein receptor binding region RBD.
21 . The conjugate as claimed in claim 20 , wherein the coronavirus is SARS-CoV-2 or Middle East respiratory syndrome coronavirus.
22 . The conjugate as claimed in claim 20 , wherein the protein/peptide antigen is a fusion protein
which is SARS-COV-2 RBD-mFc; SARS-COV-2 RBD-his; or MERS-COV RBD-his; and
the Fc fragment is a human or murine IgG Fc fragment.
23 . The conjugate as claimed in claim 21 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:1, SEQ ID NO: 2 or SEQ ID NO:3.
24 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen is a fusion protein
which is RSV-gpG-his.
25 . The conjugate as defined in claim 18 , wherein the protein/peptide antigen comprises the sequence as set forth as SEQ ID NO: 4 or SEQ ID NO:12.
26 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen is a fusion protein which is Flu-B-HA1-his or H5N1-HA-his.
27 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:7, SEQ ID NO:15, SEQ ID NO:8, or SEQ ID NO: 16.
28 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen is a fusion protein
which is Ebola-GP-Fc or Ebola-GP1-his; and
wherein the Fc is a human or murine IgG Fc fragment.
29 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:9, SEQ ID NO: 17, SEQ ID NO: 10, or SEQ ID NO: 18.
30 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen is a fusion protein
which is HCV-E2-his and/or HCV-E1-his.
31 . The conjugate as claimed in claim 18 , wherein the protein/peptide antigen comprises the sequence as set forth as any one of SEQ ID NO:11, SEQ ID NO: 19, SEQ ID NO: 6, SEQ ID NO:14, SEQ ID NO:5, or SEQ ID NO:13.
32 . The glyco-protein/peptide antigen conjugate as claimed in claim 18 , wherein the glyco-protein/peptide antigen is further conjugated to a protein carrier.
33 . The glyco-protein/peptide antigen conjugate as claimed in claim 32 , wherein the protein carrier is tetanus toxoid, tetanus toxoid fragment C, tetanus toxoid non-toxic mutant, diphtheria toxoid, CRM197, other non-toxic mutants of diphtheria toxoid.
34 . An immunogenic composition comprising the glyco-protein/peptide antigen conjugate of claim 18 , immune adjuvant and excipient.
35 . The immunogenic composition as claimed in claim 34 , the adjuvant selected from aluminium adjuvant, oil-in-water emulsion adjuvant, MF59, QS-21 and lipid monophosphate A.
36 . A method of preventing or treating disease caused by a pathogen-associated protein/peptide antigen, the method comprising administering the glyco-protein/peptide antigen conjugate of claim 18 to a subject in need thereof.
37 . A method of preventing or treating a disease caused by a pathogen-associated protein/peptide antigen, the method comprising preparing a vaccine or drug comprising the glyco-protein/peptide antigen conjugate of 18 and administering said vaccine or drug to a subject in need thereof.
38 . A method of preventing or treating a disease caused by a pathogen-associated protein/peptide antigen, the method comprising administering the immunogenic composition of claim 34 to a subject in need thereof.
39 . A method of preventing or treating a disease caused by a pathogen-associated protein/peptide antigen, the method comprising preparing a vaccine or drug comprising the immunogenic composition of claim 34 , and administering said vaccine or drug to a subject in need thereof.