VHH antibodies against the B subunit of Shiga toxin 2 (Stx2B) as novel therapeutic tools against HUS

MARIA P. MEJÍAS; YANINA HIRIART; ROMINA J. FERNÁNDEZ-BRANDO; ROMINA PARDO; ANDREA BRUBALLA; FERNANDO A. GOLDBAUM; VANESA ZYLBERMAN; MARINA S. PALERMO

Ciudad de Bunos Aires

Congreso; IV LASID Meeting, II French-Argentinean Immunology Meeting and LXIII Reunión Científica Anual de la Sociedad Argentina de Inmunología; 2015

Sociedad Argentina de Inmunologia

VHH antibodies against the B subunit of Shiga toxin 2 (Stx2B) as novel therapeutic tools against HUS.  Maria P. Mejías1; YaninaHiriart2,3; Romina J. Fernández-Brando1; Romina Pardo2,3;  Andrea Bruballa1; Fernando A.Goldbaum2,3;  Vanesa Zylberman2,3*;Marina S. Palermo1.  1Institutode Medicina Experimental (IMEX-CONICET)- Academia Nacional de Medicina, división Inmunología, Ciudad Autónoma de Buenos Aires, Argentina. 2ConsejoNacional de Investigaciones Científicas y Técnicas (CONICET);3Inmunova S.A Ciudad Autónoma de Buenos Aires, Argentina. Ciudad Autónoma de Buenos Aires, Argentina. Keywords: VHH, HUS, Stx2 Background: Infection with Stx2-producingEscherichia coli (STEC) canprogress to Hemolytic Uremic Syndrome (HUS),for which no effective therapy is presently available. We generated achimeric BLS-Stx2B immunogen, which raised high affinity and protectiveanti-Stx2B antibodies. The aim of this work was to use this immunogen toproduce therapeuticVHH antibodies against Stx2.Methods: Two llamas were immunized with BLS-Stx2B.  Phagemid libraries were generated andanti-Stx2B VHH clones were selected. One highly neutralizing VHH (2vb27) wasselected. Three alternative formats were evaluated: monomeric 2vb27, a bivalent[(2vb27)2] and a heterotrimeric molecule [(2vb27)2-SA;SA: anti-human serum albumin VHH). Invitro neutralizing activity was measured by the Vero cell assay. For in vivo studies, Balb/c mice wereinjected i.v. with 1LD100 Stx2 or i.g. with 4x1011 CFU/kgof STEC and inoculated i.p. with the different VHH formats. The half-life of theVHHs was determined by persistence of antibody in plasma, assayed byneutralizing activity. Results: Although (2vb27)2and (2vb27)2-SA showed a similar in vitro neutralizing activity, only (2vb27)2-SA (0.1pmoles) was able to fully protect mice against mortality and Stx2-associatedrenal damage after i.v Stx2 or i.g. STEC challenge. The in vivo half-life studies showed that most of 2vb27 was removed within5 minutes from plasma, (2vb27)2 within 5 hours and (2vb27)2-SAwithin 10 days. Conclusions: (2vb27)2-SAshowed the highest in vivo protectionin both experimental models, probably due to the longer in vivo half-life as a result of the binding to serum albumin. TheseVHH are promising therapeutic agents against HUS.