Generation and Characterization of a Bivalent HIV-1 Subtype C gp120 Protein Boost for Proof-of-Concept HIV Vaccine Efficacy Trials in Southern Africa.

Carlo Zambonelli,Susan W Barnett,Eden P Go,Celia Labranche,Antu K Dey,Mark Wininger,Hua-Xin Liao,Heather Desaire,Ronald I Swanstrom,Samuel Stephenson,David Montefiori,Barton F Haynes,Susan Hilt,Andrea Carfi,Daniel F Clark,Cynthia Ann Derdeyn

doi:10.1371/journal.pone.0157391

Abstract

The viral envelope glycoprotein (Env) is the major target for antibody (Ab)-mediated vaccine development against the Human Immunodeficiency Virus type 1 (HIV-1). Although several recombinant Env antigens have been evaluated in clinical trials, only the surface glycoprotein, gp120, (from HIV-1 subtype B, MN, and subtype CRF_01AE, A244) used in the ALVAC prime-AIDSVAX gp120 boost RV144 Phase III HIV vaccine trial was shown to contribute to protective efficacy, although modest and short-lived. Hence, for clinical trials in southern Africa, a bivalent protein boost of HIV-1 subtype C gp120 antigens composed of two complementary gp120s, from the TV1.C (chronic) and 1086.C (transmitted founder) HIV-1 strains, was selected. Stable Chinese Hamster Cell (CHO) cell lines expressing these gp120s were generated, scalable purification methods were developed, and a detailed analytical analysis of the purified proteins was conducted that showed differences and complementarity in the antigenicity, glycan occupancy, and glycan content of the two gp120 molecules. Moreover, mass spectrometry revealed some disulfide heterogeneity in the expressed proteins, particularly in V1V2-C1 region and most prominently in the TV1 gp120 dimers. These dimers not only lacked binding to certain key CD4 binding site (CD4bs) and V1V2 epitope-directed ligands but also elicited reduced Ab responses directed to those epitopes, in contrast to monomeric gp120, following immunization of rabbits. Both monomeric and dimeric gp120s elicited similarly high titer Tier 1 neutralizing Abs as measured in standard virus neutralization assays. These results provide support for clinical evaluations of bivalent preparations of purified monomeric TV1.C and 1086.C gp120 proteins.

Highlights

Human Immunodeficiency Virus type 1 (HIV-1) infection and acquired immunodeficiency syndrome (AIDS) represent a major public health concern
The presence of a dimeric gp120 fraction was detected by Western blot analysis of the culture supernatants and clones with the highest percentage of monomeric gp120 were selected for further evaluations
It was critical that envelope glycoprotein (Env) subunit protein vaccine candidates from HIV-1 subtype C strains, relevant to that region, be produced for the boost component of the vaccine

Summary

Introduction

HIV-1 infection and acquired immunodeficiency syndrome (AIDS) represent a major public health concern. Subsequent approaches adopted vaccines designed to preferentially stimulate cytolytic CD8+ T cell (CTL) immunity These trials failed to show protection, and a potential enhancement of disease was reported in some individuals [14, 15]. The first evidence of HIV vaccine efficacy came from the RV144 Phase 3 trial in Thailand [17] This trial tested a recombinant canarypox prime followed by a bivalent gp120 boost. We report the generation of stable CHO cell lines expressing these two gp120s, development of a scalable purification process, a comprehensive analytical characterization of the purified gp120s, and confirmation of the immunogenicity of the candidate proteins These studies serve as a foundation for cGMP manufacture of these candidates for post-RV144 clinical evaluations

Materials & Methods

Results

Discussion