Abstract
P15.25 Background: The human monoclonal antibody, 10E8, targets the membrane-proximal external region of gp41 on HIV-1 Env and neutralizes ∼98% of HIV-1 isolates. However, it is prone to aggregation at neutral pH and this less-than-ideal solubility poses a challenge to the development of 10E8 as a prophylactic agent for passive protection from HIV-1 infections in humans. Methods: We used somatic variants and employed structural and computational approaches to enhance the solubility of 10E8, which we assessed by measuring the turbidity of variants in phosphate buffered saline. Neutralization potency was measured via luciferase/TZM-bl cell-based assay on a 9-virus panel. Results: We identified a somatic variant of the heavy chain 10E8 (named HC6), which slightly improved neutralization potency relative to 10E8, while displaying similar breadth and solubility. We altered four hydrophobic residues in the framework 3 region of HC6 and found that this alteration (L72D, I75K, F77T, and M84T) improved solubility by ∼10-fold. When we introduced these mutations into another heavy chain variant of 10E8, H6dN152, solubility increased even further (>20-fold relative to 10E8); however, this variant neutralized the selected viruses over 10-fold more weakly than 10E8. Mapping sequence differences between HC6 and H6dN152 suggested that this reduced potency might relate to changes in four residues in close proximity to the gp41 epitope: D28, N31, T52 and Y98. Incorporation of changes at these residues into H6dN152 (L72D, S74T, I75K, and F77T) increased potency to a level (geometric mean IC50=0.168 ug/ml), which was comparable to that of 10E8, while retaining a>20-fold increase in solubility. Conclusions: By combining a structure-based approach and the natural variation in potency and solubility for somatic variants of 10E8, we successfully engineered a variant of 10E8 with solubility improved by more than 20-fold, while retaining its impressive neutralization breadth and potency.
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