CLEC5A/TLR2 bispecific antibody suppresses dengue virus-induced pro-inflammatory cytokines production from macrophages

BACKGROUND: Dengue virus (DENV) infection drives pathological inflammation through coordinated activation of pattern recognition receptors (PRRs) in myeloid cells, yet no approved immunomodulatory therapy exists to interrupt this process. CLEC5A and Toll-like receptor 2 (TLR2) have each been implicated in DENV-induced cytokine production, but whether their co-engagement represents a convergent and therapeutically targetable axis remains unclear. METHODS: We define the cytokine landscape of human macrophages infected with all four DENV serotypes and reverse genetics DENV2 (rgDV2) strains carrying NS1 mutations derived from the severe 2015 Taiwan outbreak. We further demonstrate the potential therapeutic effect of bispecific antibodies simultaneously targeting CLEC5A and TLR2-engineered in both IgG1 and IgG4 formats. RESULTS: Human macrophages infected with all four DENV serotypes, as well as reverse genetics DENV2 (rgDV2), exhibit a consistently dominant pro-inflammatory program driven by CLEC5A-TLR2 co-signaling. Treatment with bispecific antibodies, in both IgG1 and IgG4 formats, potently suppress the production of TNF-α, IL-6, IL-8, and MCP-1 production across all viral strains tested. Notably, the IgG4 format preserves robust inhibitory efficacy while minimizing Fc receptor engagement, thereby offering a rational strategy to mitigate the risk of antibody-dependent enhancement (ADE). This dual-targeting approach also remains effective against NS1 mutant strains linked to enhanced virulence and cytokine storm. CONCLUSION: Together, these findings identify CLEC5A-TLR2 co-signaling as a convergent inflammatory axis in dengue pathogenesis and establish bispecific dual-receptor blockade as a mechanistically grounded, ADE-aware immunomodulatory strategy to mitigate cytokine-driven pathology.