Platform ¨C recombinant overlapping peptides
Conventional method of designing vaccines is to use the whole (killed or attenuated) viruses/bacteria or whole viral/bacterial proteins to stimulate human body to produce mainly antibodies. A new technology proposed around 2003 by Dr Jiang and other scientists is to use a pool of short, serial and overlapped peptide fragments covering a viral, bacterial or tumour antigen as vaccines to stimulate mainly cellular immunity. The technology has been independently validated, demonstrating the applicability of the approach by two groups ¨C by Dr Jiang in an HIV study and by a Netherlands group in a Phase II trial of an HPV vaccine.
There is great potential for the use of this technology is the development of treatments for a broad range of infectious diseases and cancers. Compared with conventional vaccine technology (which only stimulates antibody and partially CD4+ T cells), there are major benefits from using a pool of synthetic, overlapping peptides (OPs)
Ø stronger and broader cellular immune responses are seen in vivo relating to a particular disease-associated protein (both CD8+ and CD4+ T cells).
Ø HLA restriction is overcome as multiple epitopes are covered.
Ø time-consuming epitope mapping is not required as overlapping peptides reduce the time taken to produce an effective vaccine.
Ø unrelated immunity is not stimulated as vaccines based on the overlapping peptides are also purely viral or bacterial elements.
Ø clearance of infections, as well as tumour cells is improved as therapeutic vaccines improve the body¡¯s cellular immunity, especially CD8+ T cells.
However, despite advances in solid-phase methods (which is the accepted method for creation of synthetic peptides in laboratories, particularly for peptides which are difficult to express in bacteria), use of synthetic peptides can be less than ideal in terms the large-scale manufacturing required for many infectious disease vaccines. They are both expensive and difficult to scale up and in addition require multiple regulatory submissions for approval.
Therefore, this technology has further been developed by Dr Jiang¡¯s research group at the University of Oxford. The research group proposed a recombinant method for producing overlapping peptides, where bacterial expression of an amino acid sequence corresponding to overlapping peptide sequences spanning the length of a protein of interest interspersed with enzymatic cleavage sites is used. The recombinant product can then be digested either in vitro, or, if cleavage sites for enzymes present in human cells are chosen, in vivo. The overlapping peptides made that way are called recombinant overlapping peptides (ROPs).
This novel patented methodology for making overlapping peptides gives many advantages over solid-phase synthesis. It has also shown to be effective in vaccine application The recombinant way of manufacturing overlapping peptides is at least a thousand times cheaper than the solid-phase synthesis and does not face the same regulatory hurdles. Proof of concept has been demonstrated - adjuvanted recombinant overlapping peptide vaccine to HIV-Nef protein can induce specific cellular immune responses (both CD8+ and CD4+ T cell) in 2 strains of mice and vaccination with the vaccine is also able to protect mice from high dose viral challenge of vaccinia virus expressing HIV-Nef.
This platform technology has maximum value in the development of therapeutic vaccines and in diagnostics. The advantages it provides include:
Ø significantly reduced manufacturing costs, leading to strong health economic arguments for the use of therapeutic vaccines
Ø simpler regulatory requirements in US, EU and Japan - each ROP requires one regulatory submission, not several as with synthetic overlapping peptides
Ø an effective alternative to antibody based vaccines, which are limited by their inability to be effective on intracellular microbial infections .
Ø effective stimulation of cellular immunity. The use of ROPs will overcome issues experienced with current methods used to stimulate cellular immunity
Ø an effective way of designing diagnostics for cellular immunity. A diagnostic method for cellular immunity against a viral infection can be developed in parallel with a vaccine development against the same viral infection so that the effectiveness of the vaccine can be monitored.
OVM-100 has been developed as a recombinant overlapping peptide to activate an immune response to the human papilloma virus (HPV). To date the following progress has been made:
Ø the ROP HPV cancer vaccine has been designed and manufactured ¨C OVM-100
Ø in vitro proof of concept has been established: OVM-100 has been shown to activate CD8+ T cells, with these activated T cells then attacking infected cells
Ø production scale up to 10L has already been established by Changzhou Vacmedix
ØFigure 7: OVM-200 proof of conceptpreclinical animal studies are ongoing in China ¨C in vivo proof of concept has been established in a mouse melanoma cell line B16 as shown below
Figure 5: OVM-100 proof of concept
Mice (10 per group) were immunized with either ROP- HPV E7 or the wild-type protein HPV E7 x3 with three weeks interval. Three weeks after the last immunization, mice were inoculated with mouse melanoma cell line B16 expressing either HPV E7. The percentage of survival rate was recorded for 20 days and showed a statistical difference strongly in favour of OVM-100.
Further preclinical work and manufacturing work on OVM-100 is ongoing in China. Requirements for a Phase 1 application for human use in the EU are given in Appendix 1.
The ROP OVM-200 has been developed to activate an immune response to Survivin. Survivin is one of a family of Inhibitor of Apoptosis Proteins (IAPs) that act to prevent the immune system from causing cell death in cancer cells. Survivin is highly expressed in many solid tumours and haematological malignancies and correlates with poor prognosis in several cancer types, including ovarian cancer, glioblastoma, breast cancer, lung cancer and colorectal cancer. Cancer vaccines using cytotoxic CD8+ T cells that target survivin epitopes have been developed and show tumour suppression in multiple animal models with minimal side effects. Further development of these vaccines in Phase 1 and Phase 2 trials are ongoing, mainly in academic centres
To date the following progress has been made with OVM-200:
Ø the ROP Survivin cancer vaccine has been designed and manufactured
Ø in vitro proof of concept has been established: OVM-200 has been shown to activate CD8+ T cells, with these activated T cells then attacking infected cells
Ø production scale up to 10L has already been established by Changzhou Vacmedix
Ø preclinical animal studies are ongoing in China ¨C in vivo proof of concept has been established in a mouse melanoma cell line B16 as shown below in Figure 7
Mice (10 per group) were immunized with either ROP- Survivin or the wild-type protein Survivin x3 with three weeks interval. Three weeks after the last immunization, mice were inoculated with mouse melanoma cell line B16 expressing Survivin. The percentage survival rate was recorded for 20 days and showed a statistical difference strongly in favour of OVM-200.
Further preclinical work and manufacturing work on OVM-200 is ongoing in China. Requirements for a Phase 1 application for human use in the EU are given in Appendix 1.
In addition to the two vaccines in preclinical development a therapeutic cancer vaccine to target the Epstein Barr virus (EBV) is being developed by OVM using the ROP technology platform. EBV is implicated in the onset of nasopharyngeal cancer. OVM also has the capability to develop further therapeutic cancer vaccines with the all the benefits of the ROP technology platform and with potential utility against a wide range of cancers. There are over 65 Phase 1 and Phase 2 trials ongoing using therapeutic cancer vaccines and targeting a wide range of solid tumours and haematological malignancies. Over a third of these trials are targeted at one of three indications ¨C glioblastoma, metastatic breast cancer and hormone refractory prostate cancer (HRPC).