MONOCLONAL ANTIBODIES FOR HEPATITIS C VACCINATION ANDTHERAPY

Lead Research Organisation: MRC Virology Unit

Abstract

Abstracts are not currently available in GtR for all funded research. This is normally because the abstract was not required at the time of proposal submission, but may be because it included sensitive information such as personal details.

Technical Summary

There are two distinct parts to this project:

1. Development of an antibody-based vaccine
This proposal is based on a mouse monoclonal antibody (MAb) AP33, developed in-house, that is capable of neutralizing infection by all hepatitis C virus (HCV) major genotypes and subtypes. As AP33 is a broadly neutralizing antibody recognizing a highly conserved epitope in the HCV E2 glycoprotein, this region is a target for vaccine development. Attempts using appropriate antigen fragments were not successful in eliciting antibodies against this protective epitope, indicating that designing immunogens displaying the correct presentation of the epitope will be essential for generating the desired response. To this end, we will use innovative approaches to generate such antigens and immunize animals with them. We will test whether these immunogens are capable of eliciting appropriate immune responses in immunized animals, and whether the immune sera are able to neutralize virus infection in cell culture assays.

2. Identification and development of novel antibodies with therapeutic potential
We have generated a large repertoire (approximately 1200) of mouse hybridoma lines that secrete MAbs to plasma membrane proteins of human liver origin, with the objective of using them as a means to identify putative cellular receptor(s) for HCV entry into target cells. Recent advances in cell culture infection models provide an excellent opportunity to screen this bank of anti-liver membrane MAbs for the ability to inhibit virus infection. Any inhibitory MAbs identified thus will be humanized and evaluated for their therapeutic potential. Innovative approaches will be developed to overcome possible deleterious effects that antibodies directed against a cellular target may have on the host.

Both parts of this project are founded on extensive experience of MAb development. Arvind Patel's group has ample experience of raising mono- and polyclonal antibodies to numerous viral and cellular antigens. We have developed ELISA-based assays for screening and characterizing appropriate antibodies, and pilot immunizations have already yielded promising results. Crucially, we have also generated a novel reporter cell line that will allow high throughput screening for antibodies that inhibit infection by HCV bearing envelope glycoproteins of any genotype.
 
Title Mutant antibodies 
Description Derivatives of the broadly neutralsing antibody AP33 carying point mutations in residues located in the antigen-binding pocket have been generated. 
Type Of Material Technology assay or reagent 
Provided To Others? No  
Impact These mutant antibodies are currently proving very useful reagents in assays to assess the reactivity of anti-idiotypic antibodies to AP33. 
 
Title hybridomas producing anti-idiotype antibodies 
Description murine hybridoma cell lines producing anti-idiotype antibodies to MAb AP33 
Type Of Material Antibody 
Provided To Others? No  
Impact work in progress 
 
Title hybridomas producing antibodies to liver membrane proteins 
Description murine hybridoma cell lines producing novel antibodies to liver membrane proteins, as yet uncharacterised. 
Type Of Material Antibody 
Provided To Others? No  
Impact work in progress 
 
Description Structure of AP33 and related vaccine studies 
Organisation Imperial College London
Country United Kingdom 
Sector Academic/University 
PI Contribution Generated and purified large quantities of the hepatitis C virus (HCV) monoclonal antibody (MAb) AP33 Fab fragment (a pan-genotypic virus neutralizing antibody) and supplied appropriate peptides to University of St Andrews for co-crystallisation studies. We used MAb AP33 as a template to reverse engineer an immunogen that induces similar antibodies upon vaccination. To our knowledge, this is the first time in the HCV vaccine field, the success of such a focused, structure-based approach has been demonstrated. We have supplied this candidate vaccine to our University of St Andrews collaborator for structural studies. We recently established collaoartion with ICL to validate this vaccine in vivo in an HCV infection small animal model In collaboration with the group at University of Naples Federico II, Naples we characterised monoclonal antibodies generated in animals following immunisation with a cyclic peptide representing the AP33 epitope for binding to the HCV glycoprotein E2 and for virus neutralising activity. In collaboration with the Intercollegiate Faculty of Biotechnology , Gdansk, analysed sera from animals immunised with the hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope.
Collaborator Contribution Determined the crystal structure of the antibody AP33 bound to its epitope peptide and just recently recently, that of our vaccine candidate in complex with AP33. Further structural analysis of the latter is currently in progress. The University of Naples Federico II group generated antibodies to the cyclic variant of the peptide corresponding to the AP33 epitope and performed their structural analyses when bound to the peptide. Intercollegiate Faculty of Biotechnology , Gdansk, developed a hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope for vaccine research Our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine.
Impact We have recently crystallized MAb AP33 bound to its cognate epitope sequence which identified antibody residues essential for this interaction. Based on this we are generated mutated derivatives of this antibody which are proving very useful in our on-going studies on understanding the antigen-antibody interactions at both structural and biochemical levels (Paper published: PMID 22993159). As an extension to this work, we have a novel vaccine candidate capable of eliciting MAb AP33-like anitbodies in immunized animals. Furthermore, a crystal structure of this antibody-based vaccine candidate in complex with its ligand has just been generated. Further work (including in vivo evaluation of the vaccine) is in progress. Indeed, more recently our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine. We explored the potential of cyclic peptides mimicking the AP33 epitope structure to elicit anti-HCV antibodies. Antibodiess that specifically recognize a cyclic variant of the epitope bound to soluble E2 with a lower affinity than other blocking antibodies and do not neutralize virus. The structure of the complex between one such antibody and the cyclic epitope, together with new structural data showing the linear peptide bound to neutralizing antibodies in extended conformations, suggests that the epitope displays a conformational flexibility that contributes to neutralization escape. Such features can be of major importance for the design of epitope-based anti-HCV vaccines (published PMID:26819303). Collaboration is multidisciplinary involving Protein biochemistry, molecular biology, structural biology and virology.
Start Year 2009
 
Description Structure of AP33 and related vaccine studies 
Organisation Medical University of Gdansk
Department Intercollegiate Faculty of Biotechnology
Country Poland 
Sector Academic/University 
PI Contribution Generated and purified large quantities of the hepatitis C virus (HCV) monoclonal antibody (MAb) AP33 Fab fragment (a pan-genotypic virus neutralizing antibody) and supplied appropriate peptides to University of St Andrews for co-crystallisation studies. We used MAb AP33 as a template to reverse engineer an immunogen that induces similar antibodies upon vaccination. To our knowledge, this is the first time in the HCV vaccine field, the success of such a focused, structure-based approach has been demonstrated. We have supplied this candidate vaccine to our University of St Andrews collaborator for structural studies. We recently established collaoartion with ICL to validate this vaccine in vivo in an HCV infection small animal model In collaboration with the group at University of Naples Federico II, Naples we characterised monoclonal antibodies generated in animals following immunisation with a cyclic peptide representing the AP33 epitope for binding to the HCV glycoprotein E2 and for virus neutralising activity. In collaboration with the Intercollegiate Faculty of Biotechnology , Gdansk, analysed sera from animals immunised with the hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope.
Collaborator Contribution Determined the crystal structure of the antibody AP33 bound to its epitope peptide and just recently recently, that of our vaccine candidate in complex with AP33. Further structural analysis of the latter is currently in progress. The University of Naples Federico II group generated antibodies to the cyclic variant of the peptide corresponding to the AP33 epitope and performed their structural analyses when bound to the peptide. Intercollegiate Faculty of Biotechnology , Gdansk, developed a hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope for vaccine research Our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine.
Impact We have recently crystallized MAb AP33 bound to its cognate epitope sequence which identified antibody residues essential for this interaction. Based on this we are generated mutated derivatives of this antibody which are proving very useful in our on-going studies on understanding the antigen-antibody interactions at both structural and biochemical levels (Paper published: PMID 22993159). As an extension to this work, we have a novel vaccine candidate capable of eliciting MAb AP33-like anitbodies in immunized animals. Furthermore, a crystal structure of this antibody-based vaccine candidate in complex with its ligand has just been generated. Further work (including in vivo evaluation of the vaccine) is in progress. Indeed, more recently our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine. We explored the potential of cyclic peptides mimicking the AP33 epitope structure to elicit anti-HCV antibodies. Antibodiess that specifically recognize a cyclic variant of the epitope bound to soluble E2 with a lower affinity than other blocking antibodies and do not neutralize virus. The structure of the complex between one such antibody and the cyclic epitope, together with new structural data showing the linear peptide bound to neutralizing antibodies in extended conformations, suggests that the epitope displays a conformational flexibility that contributes to neutralization escape. Such features can be of major importance for the design of epitope-based anti-HCV vaccines (published PMID:26819303). Collaboration is multidisciplinary involving Protein biochemistry, molecular biology, structural biology and virology.
Start Year 2009
 
Description Structure of AP33 and related vaccine studies 
Organisation University of Naples
Department Institute of Biostructures and Bioimaging
Country Italy 
Sector Academic/University 
PI Contribution Generated and purified large quantities of the hepatitis C virus (HCV) monoclonal antibody (MAb) AP33 Fab fragment (a pan-genotypic virus neutralizing antibody) and supplied appropriate peptides to University of St Andrews for co-crystallisation studies. We used MAb AP33 as a template to reverse engineer an immunogen that induces similar antibodies upon vaccination. To our knowledge, this is the first time in the HCV vaccine field, the success of such a focused, structure-based approach has been demonstrated. We have supplied this candidate vaccine to our University of St Andrews collaborator for structural studies. We recently established collaoartion with ICL to validate this vaccine in vivo in an HCV infection small animal model In collaboration with the group at University of Naples Federico II, Naples we characterised monoclonal antibodies generated in animals following immunisation with a cyclic peptide representing the AP33 epitope for binding to the HCV glycoprotein E2 and for virus neutralising activity. In collaboration with the Intercollegiate Faculty of Biotechnology , Gdansk, analysed sera from animals immunised with the hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope.
Collaborator Contribution Determined the crystal structure of the antibody AP33 bound to its epitope peptide and just recently recently, that of our vaccine candidate in complex with AP33. Further structural analysis of the latter is currently in progress. The University of Naples Federico II group generated antibodies to the cyclic variant of the peptide corresponding to the AP33 epitope and performed their structural analyses when bound to the peptide. Intercollegiate Faculty of Biotechnology , Gdansk, developed a hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope for vaccine research Our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine.
Impact We have recently crystallized MAb AP33 bound to its cognate epitope sequence which identified antibody residues essential for this interaction. Based on this we are generated mutated derivatives of this antibody which are proving very useful in our on-going studies on understanding the antigen-antibody interactions at both structural and biochemical levels (Paper published: PMID 22993159). As an extension to this work, we have a novel vaccine candidate capable of eliciting MAb AP33-like anitbodies in immunized animals. Furthermore, a crystal structure of this antibody-based vaccine candidate in complex with its ligand has just been generated. Further work (including in vivo evaluation of the vaccine) is in progress. Indeed, more recently our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine. We explored the potential of cyclic peptides mimicking the AP33 epitope structure to elicit anti-HCV antibodies. Antibodiess that specifically recognize a cyclic variant of the epitope bound to soluble E2 with a lower affinity than other blocking antibodies and do not neutralize virus. The structure of the complex between one such antibody and the cyclic epitope, together with new structural data showing the linear peptide bound to neutralizing antibodies in extended conformations, suggests that the epitope displays a conformational flexibility that contributes to neutralization escape. Such features can be of major importance for the design of epitope-based anti-HCV vaccines (published PMID:26819303). Collaboration is multidisciplinary involving Protein biochemistry, molecular biology, structural biology and virology.
Start Year 2009
 
Description Structure of AP33 and related vaccine studies 
Organisation University of St Andrews
Department Centre for Biomolecular Sciences
Country United Kingdom 
Sector Academic/University 
PI Contribution Generated and purified large quantities of the hepatitis C virus (HCV) monoclonal antibody (MAb) AP33 Fab fragment (a pan-genotypic virus neutralizing antibody) and supplied appropriate peptides to University of St Andrews for co-crystallisation studies. We used MAb AP33 as a template to reverse engineer an immunogen that induces similar antibodies upon vaccination. To our knowledge, this is the first time in the HCV vaccine field, the success of such a focused, structure-based approach has been demonstrated. We have supplied this candidate vaccine to our University of St Andrews collaborator for structural studies. We recently established collaoartion with ICL to validate this vaccine in vivo in an HCV infection small animal model In collaboration with the group at University of Naples Federico II, Naples we characterised monoclonal antibodies generated in animals following immunisation with a cyclic peptide representing the AP33 epitope for binding to the HCV glycoprotein E2 and for virus neutralising activity. In collaboration with the Intercollegiate Faculty of Biotechnology , Gdansk, analysed sera from animals immunised with the hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope.
Collaborator Contribution Determined the crystal structure of the antibody AP33 bound to its epitope peptide and just recently recently, that of our vaccine candidate in complex with AP33. Further structural analysis of the latter is currently in progress. The University of Naples Federico II group generated antibodies to the cyclic variant of the peptide corresponding to the AP33 epitope and performed their structural analyses when bound to the peptide. Intercollegiate Faculty of Biotechnology , Gdansk, developed a hepatitis B virus surface antigen-based virus-like particles displaying the AP33 epitope for vaccine research Our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine.
Impact We have recently crystallized MAb AP33 bound to its cognate epitope sequence which identified antibody residues essential for this interaction. Based on this we are generated mutated derivatives of this antibody which are proving very useful in our on-going studies on understanding the antigen-antibody interactions at both structural and biochemical levels (Paper published: PMID 22993159). As an extension to this work, we have a novel vaccine candidate capable of eliciting MAb AP33-like anitbodies in immunized animals. Furthermore, a crystal structure of this antibody-based vaccine candidate in complex with its ligand has just been generated. Further work (including in vivo evaluation of the vaccine) is in progress. Indeed, more recently our collaborators at the ICL have shown protection from HCV infection in a chimeric human live mouse model following passive immunization with serum IgGs derived from mice vaccinated with our anti-idiotype antibody vaccine. We explored the potential of cyclic peptides mimicking the AP33 epitope structure to elicit anti-HCV antibodies. Antibodiess that specifically recognize a cyclic variant of the epitope bound to soluble E2 with a lower affinity than other blocking antibodies and do not neutralize virus. The structure of the complex between one such antibody and the cyclic epitope, together with new structural data showing the linear peptide bound to neutralizing antibodies in extended conformations, suggests that the epitope displays a conformational flexibility that contributes to neutralization escape. Such features can be of major importance for the design of epitope-based anti-HCV vaccines (published PMID:26819303). Collaboration is multidisciplinary involving Protein biochemistry, molecular biology, structural biology and virology.
Start Year 2009
 
Title Potential prophylactic vaccine, B2.1A 
Description B2.1A is an anti-idiotype antibody-based vaccine that is capable of eliciting antibodies target an epitope recognised by our broadly neutralizing antibody AP33 in vaccinated animals. A GB patent application (reference 1415714.3) was filed in 5th September 2014. Further data were added in the priority year. The efficacy of B2.1A is currently being evaluated in an immunocompetent mouse model of HCV via a collaborative study with Marcus Dorner at Imperial College. There is a collaborative agreement in place between the institutions (i.e. MRCT, GU, and Imperial), which addresses matters such as IP management and commercialisation. The intension is to market the technology once further data has been added to the patent application, as a licence and / or collaborative opportunity. Funding: MRC 
Type Therapeutic Intervention - Vaccines
Current Stage Of Development Initial development
Year Development Stage Completed 2015
Development Status Under active development/distribution
Impact Not yet, as the product is till under assessment