Subcutaneous delivery of a bacterial protease inhibitor in a vaccine formulation increases the half life of Ag inside DCs and enhances immune responses.

CORIA, LORENA; IBAÑEZ ANDRES; PASQUEVICH KARINA; CARAVAJAL MARIANELA; BERGUER PAULA; BRUNO LAURA; RISSO GABRIELA; PAULA L. GONZALEZ COBIELLO; FERNANDA M. FRANK; CASSATARO JULIANA

Congreso; Congreso Argentino de Inmunologia; 2014

Previous results demonstrated that a Brucella spp. protein (Bp) is a broad spectrum protease inhibitor as it can inhibit proteases present in the stomach and gut (mostly serine proteases) and also lysosomal proteases (cysteine proteases). Moreover, subcutaneous co-administration of Bp as adjuvant and ovalbumin (OVA) as model antigen (Ag) induces OVA specific IFN-γ producing CD8+ T cells and cytotoxic T cells in mice. In this work we characterized the mechanism of inhibition of Bp on cysteine proteases using a specific fluorogenic substrate for cathepsin L. We have found that Bp has a competitive mechanism of inhibition of cathepsin L with an inhibition constant in the μM range. We also found that Bp increases the amount of the co-administered Ag inside Lamp2+ compartments in dendritic cells (DCs) by confocal microscopy (P<0.05). This result indicates that Bp is able to increase the Ag half life inside lysosomes promoting Ag storage inside DCs that enhance its presentation to T cells. We also studied Bp?s inflammatory properties when used as an adjuvant. Bp increased the recruitment of antigen presenting cells to draining lymph nodes after its subcutaneous administration (P<0.01). Furthermore, Bp induced the up-regulation of co-stimulatory molecules and secretion of pro-inflammatory cytokines from BMDCs (P<0.001) indicating it can activate DCs. Finally, the adjuvant capacity of Bp in vaccine formulations using real microbial derived Ags was studied using a whole homogenate of Trypanozoma cruzi parasites (WH). Subcutaneous co-administration of Bp with WH reduced parasite loads and increased survival after T. cruzi infection in mice (P<0.05). Altogether, these results indicate that Bp behaves as an ideal component in vaccine formulations against infectious diseases.