The B Subunit of Shiga Toxin Fused to a Tumor Antigen Elicits CTL and Targets Dendritic Cells to Allow MHC Class I-Restricted Presentation of Peptides Derived from Exogenous Antigens

Abstract

Immunization with peptide or recombinant proteins generally fails to elicit CTL, which are thought to play a key role in the control of virus-infected cells and tumor growth. In this study we show that the nontoxic B subunit of Shiga toxin fused to a tumor peptide derived from the mouse mastocytoma P815 can induce specific CTL in mice without the use of adjuvant. The Shiga B subunit acts as a vector rather than as an adjuvant, because coinjection of the tumor peptide and the B subunit as separate entities does not lead to CTL induction. We also demonstrated that in vitro the B subunit mediates the delivery of various exogenous CD8 T cell epitopes into the conventional MHC class I-restricted pathway, as this process is inhibited by brefeldin A and lactacystin and requires a functional TAP system. In contrast to other nonviral methods for transport of exogenous Ags into the endogenous MHC class I pathway that involve macropinocytosis or phagocytosis, the Shiga B subunit targets this pathway in a receptor-dependent manner, namely via binding to the glycolipid Gb3. Because this receptor is highly expressed on various dendritic cells, it should allow preferential targeting of the Shiga B subunit to these professional APCs. Therefore, the Shiga B subunit appears to represent an attractive vector for vaccine development due to its ability to target dendritic cells and to induce specific CTL without the need for adjuvant.

Recombinant Cholera Toxin B Subunit Activates Dendritic Cells and Enhances Antitumor Immunity

“…Thus immunization with CTB by transcutaneous route may be suitable to stimulate immune response to cutaneous tumors as well as microbial infection. In regard to the other microbial toxins, it has been reported that Shiga toxin B subunit fused to tumor peptide elicits antitumor immune response to P815 murine mastocytoma (10). Molecular mechanism of adjuvant property of rCTB is not understood completely although binding to GM 1 gangliosides is required for rCTB to act as an adjuvant (21).…”

“…Therapeutic efficacy to the established Meth A tumor cells using DC immunization was also investigated. 10 6 Meth A tumor cells were injected subcutaneously into the shaved back of BALB/c mice on day 0. Splenic DC were pulsed with 1 µg/ml rCTB and Meth A tumor cell lysate on day 1 and day 2, respectively, as described above.…”

“…Another subunit of CT, a pentameric B subunit (CTB), is non-toxic and responsible for CT binding to GM 1 gangliosides, which are expressed on the cell membranes of most cells (6,17). Recently B subunit of Shiga toxin, homopentamer responsible for toxin binding to cell membrane, was proven to induce tumor-specific CTL in mice when it is fused with tumor peptide (10). In this study, we used recombinant CTB (rCTB) secreted by Bacillus brevis carrying expression vector pNU212-CTB (12) and investigated adjuvant capacity of rCTB in cancer vaccine against Meth A tumor using DC.…”

Efficient Delivery of Antennapedia Homeodomain Fused to CTL Epitope with Liposomes into Dendritic Cells Results in the Activation of CD8+ T Cells

“…Several vectors have been reported to have this ability, including toxins (43,44), the protein transduction domain from the HIV, TAT protein (45), and AntpHD (26). Their common attribute is their capacity to deliver peptides directly in the cytosol in a receptor-dependent manner for toxins (44) or following spontaneous crossing of cell membranes in the case of TAT (46) and AntpHD (47). They all differ from exogenous proteins in terms of their internalization mechanisms, which commonly use phagocytosis/endocytosis.…”

Correlation between Shiga toxin B‐subunit stability and antigen crosspresentation: A mutational analysis

“…It was shown that STxB delivers exogenous peptides into the MHC class I and II pathways of human and mouse dendritic cells (DCs) and induces humoral and cell-mediated immune responses that protect mice from tumor growth [18][19][20][21][22]. Functional TAP and proteasome activities are required for STxB-induced MHC class I-restricted antigen presentation, strongly suggesting that STxB delivers exogenous antigenic proteins and peptides across membranes into the cytosolic compartment [19]. The mechanism and exact intracellular site of this membrane translocation event have not yet been identified, but intracellular trafficking studies on human monocyte-derived DCs suggested that the retrograde route might not be involved [23].…”