Sulfated ß-glucan from Agaricus subrufescens inhibits flavivirus infection and nonstructural protein 1-mediated pathogenesis.

Despite substantial morbidity and mortality, no therapeutic agents exist for treatment of dengue or Zika, and the currently available dengue vaccine is only recommended for dengue virus (DENV)-immune individuals. Thus, development of therapeutic and/or preventive drugs is urgently needed. DENV and Zika virus (ZIKV) nonstructural protein 1 (NS1) can directly trigger endothelial barrier dysfunction and induce inflammatory responses, contributing to vascular leak in vivo. Here we evaluated the efficacy of the (1-6,1-3)-ß-D-glucan isolated from Agaricus subrufescens fruiting bodies (FR) and its sulfated derivative (FR-S) against DENV-2 and ZIKV infection and NS1-mediated pathogenesis. FR-S, but not FR, significantly inhibited DENV-2 and ZIKV replication in human monocytic cells (EC50 = 36.5 and 188.7 µg/mL, respectively) when added simultaneously with viral infection. No inhibitory effect was observed when FR or FR-S were added post-infection, suggesting inhibition of viral entry as a mechanism of action. In an in vitro model of endothelial permeability using human pulmonary microvascular endothelial cells (HPMECs), FR and FR-S (0.12 µg/mL) inhibited DENV-2 NS1- and ZIKV NS1-induced hyperpermeability by 50% and 100%, respectively, as measured by Trans-Endothelial Electrical Resistance. Treatment with 0.25 µg/mL of FR and FR-S inhibited DENV-2 NS1 binding to HPMECs. Further, FR-S significantly reduced intradermal hyperpermeability induced by DENV-2 NS1 in C57BL/6 mice and protected against DENV-induced morbidity and mortality in a murine model of dengue vascular leak syndrome. Thus, we demonstrate efficacy of FR-S against DENV and ZIKV infection and NS1-induced endothelial permeability in vitro and in vivo. These findings encourage further exploration of FR-S and other glycan candidates for flavivirus treatment alone or in combination with compounds with different mechanisms of action.

Intradermal Delivery of Dendritic Cell-Targeting Chimeric mAbs Genetically Fused to Type 2 Dengue Virus Nonstructural Protein 1.

Targeting dendritic cells (DCs) by means of monoclonal antibodies (mAbs) capable of binding their surface receptors (DEC205 and DCIR2) has previously been shown to enhance the immunogenicity of genetically fused antigens. This approach has been repeatedly demonstrated to enhance the induced immune responses to passenger antigens and thus represents a promising therapeutic and/or prophylactic strategy against different infectious diseases. Additionally, under experimental conditions, chimeric αDEC205 or αDCIR2 mAbs are usually administered via an intraperitoneal (i.p.) route, which is not reproducible in clinical settings. In this study, we characterized the delivery of chimeric αDEC205 or αDCIR2 mAbs via an intradermal (i.d.) route, compared the elicited humoral immune responses, and evaluated the safety of this potential immunization strategy under preclinical conditions. As a model antigen, we used type 2 dengue virus (DENV2) nonstructural protein 1 (NS1). The results show that the administration of chimeric DC-targeting mAbs via the i.d. route induced humoral immune responses to the passenger antigen equivalent or superior to those elicited by i.p. immunization with no toxic effects to the animals. Collectively, these results clearly indicate that i.d. administration of DC-targeting chimeric mAbs presents promising approaches for the development of subunit vaccines, particularly against DENV and other flaviviruses.