eCD4-Ig Limits HIV-1 Escape More Effectively than CD4-Ig or a Broadly Neutralizing Antibody

Abstract

The engineered antibody-like entry inhibitor eCD4-Ig neutralizes every human immunodeficiency virus type 1 (HIV-1), HIV-2, and simian immunodeficiency virus isolate it has been tested against. The exceptional breadth of eCD4-Ig derives from its ability to closely and simultaneously emulate the HIV-1 receptor CD4 and coreceptors, either CCR5 or CXCR4. Here we investigated whether viral escape from eCD4-Ig is more difficult than that from CD4-Ig or the CD4-binding site antibody NIH45-46. We observed that a viral swarm selected with high concentrations of eCD4-Ig was increasingly resistant to but did not fully escape from eCD4-Ig. In contrast, viruses selected under the same conditions with CD4-Ig or NIH45-46 fully escaped from those inhibitors. eCD4-Ig-resistant viruses acquired unique changes in the V2 apex, V3, V4, and CD4-binding regions of the HIV-1 envelope glycoprotein (Env). Most of the alterations did not directly affect neutralization by eCD4-Ig or neutralizing antibodies. However, alteration of Q428 to an arginine or lysine resulted in markedly greater resistance to eCD4-Ig and CD4-Ig, with correspondingly dramatic losses in infectivity and greater sensitivity to a V3 antibody and to serum from an infected individual. Compensatory mutations in the V3 loop (N301D) and in the V2 apex (K171E) partially restored viral fitness without affecting serum or eCD4-Ig sensitivity. Collectively, these data suggest that multiple mutations will be necessary to fully escape eCD4-Ig without loss of viral fitness. IMPORTANCE HIV-1 broadly neutralizing antibodies (bNAbs) and engineered antibodylike inhibitors have been compared for their breadths, potencies, and in vivo half-lives. However, a key limitation in the use of antibodies to treat an established HIV-1 infection is the rapid emergence of fully resistant viruses. Entry inhibitors of similar breadths and potencies can differ in the ease with which viral escape variants arise. Here we show that HIV-1 escape from the potent and exceptionally broad entry inhibitor eCD4-Ig is more difficult than that from CD4-Ig or the bNAb NIH45-46. Indeed, full escape was not observed under conditions under which escape from CD4-Ig or NIH45-46 was readily detected. Moreover, viruses that were partially resistant to eCD4-Ig were markedly less infective and more sensitive to antibodies in the serum of an infected person. These data suggest that eCD4-Ig will be more difficult to escape and that even partial escape will likely extract a high fitness cost.

Nanoparticles presenting clusters of CD4 expose a universal vulnerability of HIV-1 by mimicking target cells

“…A modified version of a recently published in vitro evolution protocol ( 36 ) was used to evaluate whether HIV-1 can escape when intermittently exposed to increasing CD4-VLP concentrations. As above, the diversified HIV-1 YU2 swarm was exposed to CD4-Ig and CD4-VLPs at starting concentrations of 17.5 μg/mL and 5 × 10 8 VLPs/mL, respectively, for 1 h, followed by the addition of fresh target cells.…”

“…Our in vitro evolution experiments show that CD4-VLPs effectively suppressed viral replication at neutralizing concentrations, and that intermittent exposure to increasing CD4-VLP concentrations failed to select resistant viral swarms. This evolution strategy has been shown to generate viral populations completely resistant to CD4-Ig and the CD4bs bNAb NIH45-46 ( 36 ). A modest loss in viral sensitivity was also observed for the potent antibody-like inhibitor eCD4-Ig, a fusion of CD4-Ig and a CCR5-mimetic sulfopeptide that protected rhesus macaques from simian-HIV challenge following delivery using an adeno-associated virus vector ( 44 ).…”

Induction of Neutralizing Responses against Autologous Virus in Maternal HIV Vaccine Trials

“…However, these data do not preclude administration of a cocktail of bNAbs targeting multiple specificities to mitigate the potential introduction of mutations that enable viral escape from any single bNAb. Similar in intent and yet different in approach to bNAbs, engineered antibody-like entry inhibitor molecules (eCD4-Ig) that simultaneously emulate both the target CD4 receptor and a coreceptor, CXCR4 or CCR5, have demonstrated a capacity for high neutralization breadth and a lower potential for viral escape than a single bNAb (35). In contrast to the concerns associated with the administration of single bNAb as a maternal intervention, a recent phase I study has demonstrated the safety, tolerability, and pharmacokinetics of passive administration of VRC01-LS bNAb in HIV-exposed infants as adjunctive therapy to ART to further reduce postnatal transmission via breastfeeding (36).…”

Promise and Progress of an HIV-1 Cure by Adeno-Associated Virus Vector Delivery of Anti-HIV-1 Biologics

“…Recently we have shown that three mutations in CD4 domain 1 increase its potency by an average of 9-fold (Fetzer et al, 2018). Furthermore, in vitro studies showed that continuous passaging of virus in the presence of eCD4-Ig only yielded partial resistance to the inhibitor (Fellinger et al, 2019). These resistance mutations also conferred a high fitness cost to the virus in entry assays.…”