Despite decades of effort, no current vaccine elicits neutralizing antibodies at concentrations blocking HIV infection. In addition to structural features of HIV's envelope spike that facilitate antibody evasion, we propose that the low density and limited lateral mobility of HIV spikes impedes bivalent binding by antibodies. The resulting predominantly monovalent binding minimizes avidity and thereby high affinity binding and potent neutralization, thus expanding the range of HIV mutations permitting antibody evasion. The HIV spike trimer geometry does not allow intra-spike cross-linking by naturally-occurring bivalent antibodies, but we can construct proteins capable of high-avidity trivalent binding to a spike for gene therapy and/or passive immunization. We will design, express, and test trimeric intra-spike cross-linking reagents that bind to two or three non-overlapping sites per spike monomer (6-9 sites per trimer). Choosing HIV-binding proteins and how to link them will be done combinatorially starting with a library of 15-30 HIV spike-binding proteins coupled to double-stranded DNA identifying tags. These will be linked using variable-length DNA to form bispecific reagents separated by different distances. Pooled bispecific reagents will separated by affinity chromatography to isolate the tightest binding pairs, which will be PCR amplified/sequenced to determine the two protein components and the linker length. Upon identifying the optimal length for the linker, the DNA is replaced with a comparable-length protein linker, and the two-binding-protein reagent is linked to a third binding protein from the HIV-binding library. Optimal two- or three-component HIV binding proteins will be trimerized by attaching a trimerization motif first via DNA linkers to determine the optimal length, then using protein-based linkers. The trimeric reagents intra-spike cross-linking would reduce the concentration required for sterilizing immunity, making HIVs low spike density irrelevant. Analogous to using ~3 drugs during HAART therapy, simultaneous binding to several different sites/monomer would reduce/abrogate sensitivity to spike mutations.
HIV/AIDS remains one of the most important current threats to global public health. We propose that HIV evolved an unusually low spike number and spike density to prevent antibodies from binding with both of their combining sites, thereby diminishing avidity effects that are important for antibody-mediated neutralization of other viruses for which humans mount an effective antibody response and/or a successful vaccine has been made. Here we describe a new methodology designed to produce novel anti-HIV binding proteins that can bind simultaneously to all three monomers in an HIV spike trimer, thus interacting with high avidity and rendering the low spike density of HIV and its high mutation rate irrelevant to effective neutralization.
|West Jr, Anthony P; Scharf, Louise; Scheid, Johannes F et al. (2014) Structural insights on the role of antibodies in HIV-1 vaccine and therapy. Cell 156:633-48|
|West Jr, Anthony P; Scharf, Louise; Horwitz, Joshua et al. (2013) Computational analysis of anti-HIV-1 antibody neutralization panel data to identify potential functional epitope residues. Proc Natl Acad Sci U S A 110:10598-603|