Fusion Protein Vaccine by Domains of Bacterial Exotoxin Linked with a Tumor Antigen Generates Potent Immunologic Responses and Antitumor Effects

Abstract

Antigen-specific immunotherapy represents an attractive approach for cancer treatment because of the capacity to eradicate systemic tumors at multiple sites in the body while retaining the requisite specificity to discriminate between neoplastic and nonneoplastic cells. It has been shown that certain domains of bacterial exotoxins facilitate translocation from extracellular and vesicular compartments into the cytoplasm. This feature provides an opportunity to enhance class I and/or II presentation of exogenous antigen to T lymphocytes. We investigated previously whether the translocation domain (domain II) of Pseudomonas aeruginosa exotoxin A with a model tumor antigen, human papillomavirus type 16 E7, in the context of a DNA vaccine could enhance vaccine potency. We then attempted to determine whether this chimeric molecule could also generate strong antigen-specific immunologic responses and enhance the potency of cancer vaccine in the protein format. Our results show that vaccination with the PE(#III)-E7-KDEL3 fusion protein enhances MHC class I and II presentation of E7, leading to dramatic increases in the number of E7-specific CD8 + and CD4 + T-cell precursors and markedly raised titers of E7-specific antibodies. Furthermore, the PE(#III)-E7-KDEL3 protein generates potent antitumor effects against s.c. E7-expressing tumors and preestablished E7-expressing metastatic lung tumors. Further, mice immunized with PE(#III)-E7-KDEL3 protein vaccine also retained long-term immunologic responses and antitumor effects. Our results indicate that retrograde-fusion protein via the delivery domains of exotoxins with an antigen greatly enhances in vivo antigen-specific immunologic responses and represents a novel strategy to improve cancer vaccine potency.

HER2-targeting recombinant protein with truncated Pseudomonas Exotoxin: A translocation domain efficiently kills breast cancer cells

“…[4][5][6][7] Previous studies have demonstrated that the PEA translocation domain can promote heterologous protein entry into cells. 8,9 Although earlier studies used PEA fragment from amino acids 253-412 or 253-364 for protein translocation, these data suggested that amino acids 253-358 is sufficient for PEA translocation. [10][11][12][13] In order to determine the minimum domain length required for translocation, a set of deletions in the PEA translocation domain were introduced into the previously constructed ImmunoGrB, 13 which is composed of a single-strand antibody against HER2, translocation domain from PEA, and active granzyme B.…”

Connective tissue growth factor linked to the E7 tumor antigen generates potent antitumor immune responses mediated by an antiapoptotic mechanism

“…We further assessed the therapeutic potential of each vaccine by performing an in vivo tumor treatment experiment using the lung hematogenous spread model described by Liao et al 23 The representative pulmonary tumor nodules are shown in Figure 3b. As shown in These data indicate that CTGF when linked to the E7 antigen could generate more potent preventive and therapeutic effects than the wild-type E7 DNA constructs in both subcutaneous and lung hematogenous spread models.…”

“…The amplified product was then cloned into the XhoI/EcoRI sites of pcDNA3 vector (Invitrogen Corp., Carlsbad, CA, USA). To generate pcDNA3-CTGF/ E7, E7 was generated as described in a previous study 23 and cloned into the HindIII sites of pcDNA3-CTGF to generate pcDNA3-CTGF/E7. To generate pcDNA3-CTGF/E7/GFP, pcDNA3-CTGF/E7 was first digested by the restriction enzyme HindIII.…”

Morphine-Sparing Effect by COX-1 Inhibitor Sustains Analgesic Function without Compromising Antigen-Specific Immunity and Antitumor Effect of Naked DNA Vaccine

“…Before intracellular cytokine staining, 3.5x10 5 pooled splenocytes from each vaccinated group were incubated for 16 h with I ug/ml ofE7 peptide (aa 49-57) containing an MHC class I epitope (20) for detecting E7-specific CD8+ T cell precursors. Cell surface marker staining ofCD8 and intracellular cytokine staining for IFNy, as well as flowcytometric analysis, were performed using conditions described previously (21)(22)(23)(24).…”