Generation and characterization of an anti ITGB4 monoclonal antibody that blocks Zika virus entry

Zika virus (ZIKV) causes congenital disease and neurological complications, yet no approved vaccines or antivirals are available. Integrin β4 (ITGB4) has been identified as an entry receptor for ZIKV.

Recombinant human ITGB4 ectodomain was expressed using a baculovirus-insect cell system and used to immunize BALB/c mice. Hybridoma screening identified monoclonal antibody 7C3. Binding affinity to ITGB4 was determined by biolayer interferometry. Epitope relationships between 7C3 and the previously reported anti-ITGB4 antibody 13H10 were assessed by competitive binding assay. Antiviral activity was evaluated in four cell lines by RT-qPCR quantification of cell-associated ZIKV RNA after antibody pretreatment and viral challenge.

7C3 bound ITGB4 with picomolar affinity (K_D=30.9 pM) and high specificity. Competitive binding showed that 7C3 and 13H10 (K_D=7.98 pM) recognize non-overlapping epitopes on ITGB4. Pretreatment with 7C3 significantly reduced cell-associated ZIKV RNA in all four cell lines compared with PBS and isotype controls.

A second, independently generated anti-ITGB4 antibody with picomolar affinity can robustly block ZIKV entry across multiple cell types, including SY5Y neuroblastoma cells relevant to congenital neuropathology. The non-overlapping epitopes of 7C3 and 13H10 suggest potential for combination strategies to enhance ITGB4 blockade. High germline identity of the 7C3 variable regions (V_H=94.46%, V_L=95.82%) supports future humanization. These findings establish 7C3 as a host-directed antiviral candidate and provide a basis for ITGB4-targeting approaches against congenital ZIKV disease.