A Novel Modular Antigen Delivery System for Immuno Targeting of Human 6-sulfo LacNAc-Positive Blood Dendritic Cells (SlanDCs)

Claudia C Bippes,Adrian Schwarzer,Achim Temme,Knut Schäkel,Michael Bachmann,Marc Cartellieri,Marc Schmitz,Holger Bartsch,Biji T Kurien,R Hal Scofield,Rebekka Wehner,Senming Zhao,Slava Stamova,Anja Feldmann,E Peter Rieber

doi:10.1371/journal.pone.0016315

Abstract

BackgroundPreviously, we identified a major myeloid-derived proinflammatory subpopulation of human blood dendritic cells which we termed slanDCs (e.g. Schäkel et al. (2006) Immunity 24, 767–777). The slan epitope is an O-linked sugar modification (6-sulfo LacNAc, slan) of P-selectin glycoprotein ligand-1 (PSGL-1). As slanDCs can induce neoantigen-specific CD4+ T cells and tumor-reactive CD8+ cytotoxic T cells, they appear as promising targets for an in vivo delivery of antigens for vaccination. However, tools for delivery of antigens to slanDCs were not available until now. Moreover, it is unknown whether or not antigens delivered via the slan epitope can be taken up, properly processed and presented by slanDCs to T cells.Methodology/Principal FindingsSingle chain fragment variables were prepared from presently available decavalent monoclonal anti-slan IgM antibodies but failed to bind to slanDCs. Therefore, a novel multivalent anti-slanDC scaffold was developed which consists of two components: (i) a single chain bispecific recombinant diabody (scBsDb) that is directed on the one hand to the slan epitope and on the other hand to a novel peptide epitope tag, and (ii) modular (antigen-containing) linker peptides that are flanked at both their termini with at least one peptide epitope tag. Delivery of a Tetanus Toxin-derived antigen to slanDCs via such a scBsDb/antigen scaffold allowed us to recall autologous Tetanus-specific memory T cells.Conclusions/SignificanceIn summary our data show that (i) the slan epitope can be used for delivery of antigens to this class of human-specific DCs, and (ii) antigens bound to the slan epitope can be taken up by slanDCs, processed and presented to T cells. Consequently, our novel modular scaffold system may be useful for the development of human vaccines.

Highlights

B cells assemble a highly diverse repertoire of immunoglobulin heavy and light chains by recombination of variable (V), diversity (D), and joining (J) gene fragments, a process known as VDJ or VJ recombination e. g. [1]
In contrast to the polyvalent maternal IgM antibody, the recombinant monovalent single chain fragment variables (scFvs) failed to bind to slanDCs (Fig. 1C, data not shown)
We hoped that the formation of bi- (Fig. 1H), tri(Fig. 1I), or other multivalent anti-slan single chain bispecific recombinant diabody (scBsDb)/linker peptide scaffolds would allow us to sufficiently restore a binding to slanDCs

Summary

Introduction

B cells assemble a highly diverse repertoire of immunoglobulin heavy and light chains by recombination of variable (V), diversity (D), and joining (J) gene fragments, a process known as VDJ (heavy chain) or VJ (light chain) recombination e. g. [1]. Antibody-encoding genes can undergo further modifications including somatic hypermutations (SHM) and class switch recombinations (CSR). After CSR and SHM, a positive selection favors the survival of those B cells which express high affinity antibodies (‘‘affinity maturation’’). SHM and positive selection improve the affinity of the paratope(s) of an antibody towards its epitope. Monovalently binding recombinant antibody fragments including for example single chain fragment variables (scFvs) prepared from IgM type antibodies usually retain only very low if any binding affinity e.g. Tools for delivery of antigens to slanDCs were not available until now. It is unknown whether or not antigens delivered via the slan epitope can be taken up, properly processed and presented by slanDCs to T cells

Methods

Results

Conclusion