Isolation and characterization of Nanobodies against Junin virus as new and improved treatment option for Argentine hemorrhagic fever

LINERO, FLORENCIA; SCOLARO, LUIS; SAELENS, XAVIER

Workshop; Innovative Approaches for Identification of Antiviral Agents Summer School; 2014

European Society for Virology

Argentine Hemorrhagic Fever (AHF) is an endemic disease specific of the central part of Argentina with a case-fatality rate, without treatment, ranging from 15% to 30%. Although the use of a live attenuated vaccine Candid#1, has markedly reduced the incidence of AHF in the population at risk, supportive therapy remains very important in the management of patients with AHF. In this work we explored the development of a novel and improved therapeutic intervention against AHF, based on the generation and use of single domain antibodies (also known as Nanobodies or VHH). This technology combines unparalleled antigen-targeting specificities with ease of expression, purification and re-engineering opportunities. The generation of the nanobodies directed against JUNV was performed by the immunization of an alpaca with purified UV-inactivated Candid#1 JUNV. After immunization, a cDNA library was generated and subsequently used to isolate phages that express JUNV-binding nanobodies at their surface by biopanning. The JUNV-binding capacity of these VHHs was evaluated by ELISA from the periplasmic extracts and the neutralizing activity of these binders was then evaluated functionally by a virus neutralization assay against JUNV Candid#1 strain. Results showed that 4 of the 70 candidates were able to neutralize successfully Candid#1 JUNV strain replication at very low concentrations. These findings constitute the first demonstration that the nanobody technology could be used to avoid JUNV replication. Afterwards testing of the virus-neutralizing activity against pathogenic strains will allow demonstrating the potential use of this technology treat and potentially preventing arenavirus hemorrhagic fever in human.