BACKGROUND: Although broadly neutralizing monoclonal antibodies (bNAbs) have 
always been considered to be a potential therapeutic option for the prophylaxis 
and treatment of HIV infection, their lack of breadth against all HIV variants 
has been one of the limiting factors. To provide sufficient neutralization 
breadth and potency against diverse viruses, including neutralization escape 
mutants, strategies to combine different bNAbs have been explored recently.
METHODS: We rationally designed and engineered a novel bispecific 
HIV-1-neutralizing antibody (bibNAb), iMabm36. The potency and breadth of 
iMabm36 against HIV were extensively characterized in vitro.
RESULTS: iMabm36 comprises the anti-CD4 Ab ibalizumab (iMab) linked to 2 copies 
of the single-domain Ab m36, which targets a highly conserved CD4-induced 
epitope. iMabm36 neutralizes a majority of a large, multiclade panel of 
pseudoviruses (96%, n = 118) at an IC50 concentration of less than 10 µg/mL, 
with 83% neutralized at an IC50 concentration of less than 0.1 µg/mL. In 
addition, iMabm36 neutralizes a small panel of replication-competent 
transmitted-founder viruses to 100% inhibition at a concentration of less than 
0.1 µg/mL in a peripheral blood mononuclear cell-based neutralizing assay. 
Mechanistically, the improved antiviral activity of iMabm36 is dependent on both 
the CD4-binding activity of the iMab component and the CD4i-binding activity of 
the m36 component. After characterizing that viral resistance to iMabm36 
neutralization was due to mutations residing in the bridging sheet of gp120, an 
optimized m36 variant was engineered that, when fused to iMab, improved 
antiviral activity significantly.
CONCLUSIONS: The interdependency of this dual mechanism of action enables 
iMabm36 to potently inhibit HIV-1 entry. These results demonstrate that 
mechanistic-based design of bibNAbs can generate potential preventive and 
therapeutic candidates for HIV/AIDS.