Characterization of the biological activity of a panel of TLR5 agonists

BIEDMA, MARINA; MORENO, GRISELDA; ROSSI, ANDRÉS; TABAREAU, JULIEN; BERGUER, PAULA; SIRARD, JEAN CLAUDE; RUMBO, MARTÍN

Ciudad de Buenos Aires

Congreso; First International Joint Meeting between the Latin American Society for Immunodeficiencies (LASID), the Argentinean Society for Immunology (SAI), and the French Society for Immunology (SFI); 2015

Flagellin (FliC) activates innate immune responses via its recognition by Toll-like receptor 5 (TLR5) and adaptive immunity in the host. TLR5 is a receptor widely distributed in mucosal epithelia and FliC, has been proposed as mucosal adjuvant. The structure of FliC includes four domains, D0 and D1 are involved in TLR5 activation, whereas D2 and D3 contain the main antigenic sites of the molecule. However, the contribution of D2 and D3 to TLR5 activation has not been fully elucidated. In this study, we evaluated 11 different deletion variants of FliC affecting D2 and D3 domains using wild type FliC as reference. We found that the deletions affect differentially the capacity to trigger TLR5 in vitro. Circular dichroism (CD) analysis indicates that FliC mutants conserved mostly the secondary structure present in the wt molecule and underwent a reversible thermal transition between 31 and 43 °C. The deletions in D2-D3 domains resulted in low capacity to elicit anti-flagellin antibodies, however, all variants were able to trigger in vivo innate response activation at dose of 1 ug. Furthermore, to characterize the adjuvant properties, we assessed antibodies responses to OVA following i.n. co-immunizations. Recombinant FliCs present mucosal adjuvant activity that correlates with TLR5 activating capacity. The highest adjuvant capacity was shown by variants which have a potency to trigger TLR5 activation comparable to flagellin wt. Our results indicate that flagellin deletion variants showing maximal TLR5 activating capacity have the most promising features to be used as mucosal adjuvants