DNA vaccine against Hemolytic Uremic Syndrome (HUS) induces neutralizing antibodies to Stx2

BENTANCOR, L; RAMOS, MV; FERNANDEZ-BRANDO, R; FERNANDEZ, G; CAMERANO, G; GHIRINGHELLI, D; PALERMO, M

Cordoba, Argentina

Congreso; VII Congreso Latinoamericano de Inmunología; 2005

ALAI

HUS is caused by the infection with Shiga toxin (Stx)-producing E.coli. There is neither a vaccine nor an effective treatment. Vaccine against HUS was not effective because the B subunit has low immunoprophylactic potential and the A subunit is toxic, so we developed a DNA vaccine expressing the toxin Stx2 without its active site. This plasmid construction conserved the last 31 aas. from A subunit and the complete B subunit. The correct eukaryotic expression and the absence of cytotoxic effects were confirmed previously. Adults BALB/C mice were immunized with pStx2DAB, pStx2B, and both injected with pGM-CSF three days before; controls with pCDNA3.1 and pGM-CSF were evaluated. Sera 1/20 were assayed for neutralizing activity in vitro on Vero cells using 0.8CD50 of Stx2. The maximal neutralizing activity was observed in pGM-CSF+pStx2DAB group (100%) at 15d.p.v., this capacity was maintained for at least 30 days. This group also showed the higher neutralizing activity ex vivo (50% survival). Sera collected recognized the A and B subunits of Stx2 assayed by Western Blot (32 and 7.7Kda). The recombinant protein was purified using pET22b system in order to study its stability and conformational properties, and to be used in combined immunization protocols (DNA followed by protein). These results showed that sera from immunized mice could be used as a therapy to diminish the toxic activity of Stx2.