Abstract
Priming of naive CD8+ and CD4+ T cells by dendritic cells (DCs) requires effective antigen presentation on both MHC class I and II molecules. We have developed a novel technology to use recombinant overlapping peptides (ROP) that stimulate both CD8+ and CD4+ T cell immune responses. The single chain protein of a ROP is made up of overlapping peptides linked by the target sequence (LRMK) for cathepsin S, a protease found in the endosomes of DCs. We designed synthetic genes encoding ROPs derived from ovalbumin (OVA), tuberculosis protein (CFP10-ESAT6), human papilloma virus (HPV) protein (E7) and survivin, a protein commonly over-expressed in tumour cells. An epitope from ROP-OVA was cross-presented and detected by a CD8+ T cell receptor-like antibody (TCR like Ab). Human DCs pulsed with ROP-survivin activated CD8+ T cells. CD4-low PBMCs from HIV and TB co-infected patients recognized ROP-CFP10-ESAT6 compared to a soluble form of the antigen. Immunization of mice with ROP-survivin or ROP-HPV-E7 generated specific cellular immune responses and protected mice from inoculation with melanoma B16 cells expressing survivin or HPV-E7 proteins. Together these data provide evidence to support ROP as a central component of a new platform for therapeutic vaccines and diagnostics.
Highlights
Priming naive antigen-specific CD8+ and CD4+ T cells requires effective antigen presentation by Major histocompatibility complex (MHC) class I and II molecules on the surface of antigen presenting cells (APC), such as dendritic cells (DC) [1, 2]
We examined antigen processing and presentation of recombinant overlapping peptides (ROP)-OVA in DC2.4 cells (H-2b) with native ovalbumin protein (OVA) as the comparator (Figure 1A)
The H-2bSIINFEKL complex was readily detected after incubation of ROP-OVA with DC2.4 cells for 13 hours (Figure 1B, red peak in the right panel), indicating that ROP-OVA was efficiently cross-presented on MHC class I
Summary
Priming naive antigen-specific CD8+ and CD4+ T cells requires effective antigen presentation by MHC class I and II molecules on the surface of antigen presenting cells (APC), such as dendritic cells (DC) [1, 2]. For CD8+ T cells, an antigen must be endogenous, i.e. expressed in or delivered to the cytoplasmic compartment of an APC, before being processed to produce peptide epitopes that are bound to MHC class I in the endoplasmic reticulum (ER) [3]. The antigens are degraded and processed, resulting in peptide epitopes bound to MHC class II. The MHC-peptide complex moves to the surface of the APC where it can stimulate CD4+ T cells [4]. Crosspresentation, a process in which exogenous antigen is presented by MHC class I on APCs to activate CD8+ T cells occurs. The mechanisms by which this cross-presentation occurs remain unclear [5, 6]
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