The HIV-1 envelope glycoprotein, Env, facilitates the translocation of the viral capsid across the cellular membrane. Env is a trimer of hetero-dimers composed of a gp120 subunit and gp41 transmembrane protein. The gp120 subunit binds the primary receptor, CD4, leading to conformational changes of Env that then promote binding to the coreceptor, principally CCR5 or CXCR4. As the sole protein on the surface of the virion, Env is under continuous pressure from the host's antibody response. Two classes of antibodies target the highly conserved receptor-binding sites of gp120: CD4-binding site (CD4bs) and CD4-induced (CD4i) antibodies.
Here we focus on inhibiting HIV-1 entry by targeting both the CD4- and coreceptor-binding sites. In Chapter 2 we show that an improved CCR5-mimetic peptide, CCR5mim2-Ig, used in combination with CD4-Ig, neutralized HIV-1 isolates more potently than either inhibitor alone. Similarly, greater neutralization was achieved by combining a CD4i antibody, E51, with a CD4bs antibody, VRC01. We show that this increased potency derives from the ability of E51 to induce quaternary changes of the Env trimer, facilitating greater access of VRC01 to its epitope. Our studies also underscore key mechanistic differences between CD4-Ig and CD4bs antibodies.
Chapter 3 describes the usefulness of targeting both receptor-binding sites by characterizing a dual-acting inhibitor: eCD4-Ig. eCD4-Ig consists of CD4-Ig fused at its C-terminus with CCR5mim1. eCD4-Ig bound Env with higher avidity than CD4-Ig and its sulfopeptide-fusion inhibited the tendency of CD4-Ig to enhance infection. The potency of eCD4-Ig is similar to or better than the best HIV-1 broadly neutralizing antibodies (bNAbs). eCD4-Ig is also more broadly neutralizing than any current HIV-1 bNAb, and it efficiently neutralized a panel of 38 HIV-1 isolates resistant to the best CD4bs bNAbs as well as SIV and HIV-2 isolates. Lastly, eCD4-Ig retained its efficacy in vivo: it protected humanized mice from an HIV-1 challenge and was expressed by an adeno-associated virus (AAV) vector in rhesus macaques at levels greater than 100 µg/mL. Taken together, our studies highlight the advantages of targeting both receptor-binding sites simultaneously and describe a new HIV-1 entry inhibitor with potential clinical use.
|Advisor:||Farzan, Michael R.|
|Commitee:||Barouch, Dan, Gabuzda, Dana, Kuritzkes, Daniel, Sagar, Manish|
|Department:||Biology: Medical Sciences, Division of|
|School Location:||United States -- Massachusetts|
|Source:||DAI-B 75/10(E), Dissertation Abstracts International|
|Keywords:||Broadly neutralizing antibodies, Cd4i antibodies, HIV-1 entry, HIV-1 envelope glycoprotein, eCD4-Ig|
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