Selection of unadapted, pathogenic SHIVs encoding newly transmitted HIV-1 envelope proteins

Abstract

Infection of macaques with chimeric viruses based on SIVMAC but expressing the HIV-1 envelope (Env) glycoproteins (SHIVs) remains the most powerful model for evaluating prevention and therapeutic strategies against AIDS. Unfortunately, only a few SHIVs are currently available. Furthermore, their generation has required extensive adaptation of the HIV-1 Env sequences in macaques so they may not accurately represent HIV-1 Env proteins circulating in humans, potentially limiting their translational utility. We developed a strategy for generating large numbers of SHIV constructs expressing Env proteins from newly transmitted HIV-1 strains. By inoculating macaques with cocktails of multiple SHIV variants, we selected SHIVs that can replicate and cause AIDS-like disease in immunologically intact rhesus macaques without requiring animal-to-animal passage. One of these SHIVs could be transmitted mucosally. We demonstrate the utility of the SHIVs generated by this method for evaluating neutralizing antibody administration as a protection against mucosal SHIV challenge.

Figure 1. Cloning Strategy and In Vitro Replication of R5-SHIVs

(A) Flowchart of the strategy used for cloning and screening R5-SHIVs. Restriction sites used to introduce the Env PCR products and ligate the two genome halves are noted. (B) Replication of SHIVs encoding the indicated Env proteins in MT2 cells engineered to express CCR5. Infectious virus in supernatant samples collected from MT2-R5 cells at the indicated times p.i. was measured using TZM-bl cells. Results from a representative experiment (of two performed) are shown. See also Figure S1 and Table S1.

Figure 2. In Vivo Selection of R5-SHIVs

(A and B) Plasma viral load in macaques inoculated with cocktails of SHIVs from Group 1 (A) or Group 2 (B). Animals were depleted of CD8⁺ cells at the time of and 1 week post inoculation. Red x indicates euthanasia due to AIDS. (C and D) Phylogenetic analysis of Env sequences found in the plasma of Group 1 (C) or Group 2 (D) SHIV-infected macaques. Black lines represent unmodified Env sequences from Group 1 or Group 2 viruses inoculated into animals and colored lines Env sequences obtained from infected animals at the indicated times p.i. Predominant Env sequences are labeled. See also Figure S2.

Figure 3. Replication of Individual R5-SHIVs in Macaques

(A) Plasma viremia, CD4⁺ T cell counts in blood, and CD4⁺ T cell frequency in the GALT for 2 macaques (one in red and one in black) inoculated intravenously with 2.9 × 10⁵ i.u. of SHIV_AD081. Asterisks in bar graphs indicate unavailable samples. (B) As in (A), except that two macaques were inoculated intravenously with 4.8 × 10⁵ i.u. of SHIV₁₀₅₄. (C) Plasma viremia for six macaques inoculated intrarectally with varying doses of SHIV₁₀₅₄. Arrows indicate time and colors the dose of viral challenge; white arrow is 3 × 10³ i.u., blue arrow is 3 × 10⁴ i.u., black arrow is 7.4 × 10⁴ i.u., and red arrow is 2.1x10⁵ i.u. See also Figure S3.

Figure 4. HIV-1 Monoclonal Antibody Protects against T/F Env Virus Challenge

(A) PGT121 antibody concentration in serum of animals infused with the antibody on day −1. Day 0 is the day of challenge with SHIV₁₀₅₄. Dotted line indicates antibody concentration that achieves 90% neutralization of SHIV₁₀₅₄ in vitro. (B) Plasma viremia in macaques receiving pre-exposure transfusion of PGT121 or control monoclonal antibodies at 10 mg/kg, followed by an intrarectal SHIV₁₀₅₄ challenge using 2.1 × 10⁵ i.u. of virus. Animals in both groups are shown in #1 blue, #2 green, #3 purple, #4 red, #5 black, and #6 orange. See also Figure S4 and Table S3.