TY  - JOUR
AU  - Cavazzini, Davide
AU  - Spagnoli, Gloria
AU  - Mariz, Filipe Colaco
AU  - Reggiani, Filippo
AU  - Maggi, Stefano
AU  - Franceschi, Valentina
AU  - Donofrio, Gaetano
AU  - Müller, Martin
AU  - Bolchi, Angelo
AU  - Ottonello, Simone
TI  - Enhanced immunogenicity of a positively supercharged archaeon thioredoxin scaffold as a cell-penetrating antigen carrier for peptide vaccines.
JO  - Frontiers in immunology
VL  - 13
SN  - 1664-3224
CY  - Lausanne
PB  - Frontiers Media
M1  - DKFZ-2022-02059
SP  - 958123
PY  - 2022
AB  - Polycationic resurfaced proteins hold great promise as cell-penetrating bioreagents but their use as carriers for the intracellular delivery of peptide immuno-epitopes has not thus far been explored. Here, we report on the construction and functional characterization of a positively supercharged derivative of Pyrococcus furiosus thioredoxin (PfTrx), a thermally hyperstable protein we have previously validated as a peptide epitope display and immunogenicity enhancing scaffold. Genetic conversion of 13 selected amino acids to lysine residues conferred to PfTrx a net charge of +21 (starting from the -1 charge of the wild-type protein), along with the ability to bind nucleic acids. In its unfused form, +21 PfTrx was readily internalized by HeLa cells and displayed a predominantly cytosolic localization. A different intracellular distribution was observed for a +21 PfTrx-eGFP fusion protein, which although still capable of cell penetration was predominantly localized within endosomes. A mixed cytosolic/endosomal partitioning was observed for a +21 PfTrx derivative harboring three tandemly repeated copies of a previously validated HPV16-L2 (aa 20-38) B-cell epitope grafted to the display site of thioredoxin. Compared to its wild-type counterpart, the positively supercharged antigen induced a faster immune response and displayed an overall superior immunogenicity, including a substantial degree of self-adjuvancy. Altogether, the present data point to +21 PfTrx as a promising novel carrier for intracellular antigen delivery and the construction of potentiated recombinant subunit vaccines.
KW  - antigen carrier (Other)
KW  - epitope display (Other)
KW  - intracellular antigen delivery (Other)
KW  - peptide epitope (Other)
KW  - protein DNA interaction (Other)
KW  - protein scaffold engineering (Other)
KW  - protein scaffold vaccine (Other)
KW  - recombinant vaccine (Other)
LB  - PUB:(DE-HGF)16
C6  - pmid:36032169
C2  - pmc:PMC9405434
DO  - DOI:10.3389/fimmu.2022.958123
UR  - https://inrepo02.dkfz.de/record/181524
ER  -