Dissecting antibody-dependent enhancement modulation by Fc-modified cross-neutralizing human monoclonal antibody

Abstract

Background. Dengue is a mosquito-borne disease caused by four dengue virus serotypes (DENV1 to DENV4). Secondary infections can generate flavivirus cross- reactive antibodies at sub-neutralizing levels. This phenomenon can significantly increase the severity of secondary infections through antibody-dependent enhancement (ADE). ADE is associated with a high risk of viral infection in immune effector cells, triggering cytokine cascades and activating the complement system, which can lead to severe symptoms. Despite extensive studies, therapeutic antibodies, particularly fully human monoclonal antibodies, which could serve as candidates for immune passive therapy, have not yet been discovered. Methodology. This study generated LALA-mutated human monoclonal antibody clone B3B9 (LALA-B3B9 HuMAb) which can neutralize all four DENV serotypes without enhancing viral activity. The number of infected cells in the ADE assay was compared among the wild-type antibody (B3B9), LALA-B3B9 HuMAb, and an Fc modified variant at position N297Q (N297Q-B3B9), with or without complement proteins. Moreover, the therapeutic efficacy of these HuMAbs against ADE infection by competing with natural antibodies in patients with acute dengue was evaluated using the in vitro suppression-of-enhancement assay in K562 cells. Result. Our novel Fc-modified antibody LALA-B3B9 (Leu234Ala/Leu235Ala mu- tations), exhibited neutralizing activity against all dengue virus serotypes without triggering ADE activity at any antibody concentration. This outcome was similar to that observed with the previously developed Fc-modified N297Q-B3B9 antibody (N297Q mutation).Wefurther evaluated the effect of complement protein on the enhancing and neutralizing activities of our Fc-modified antibodies. The results showed that LALA-B3B9 and N297Q-B3B9 HuMAbs were complement-independent, meaning that the reduced binding between complement protein (C1q) and the Fc portion of the antibody left the neutralizing and enhancing activities unchanged. Additionally, both LALA-B3B9 and N297Q-B3B9 HuMAbs demonstrated the suppression-of-enhancement activity in K562 cells induced by human anti-DENV serum antibodies. Overall, this study highlights the main advantages of our EDII-specific HuMAbs in inhibiting in vitro ADE, indicating that they are promising candidates for future dengue treatment.