Date Published: June 18, 2018
Publisher: Public Library of Science
Author(s): Young C. Shin, Georg F. Bischof, William A. Lauer, Lucas Gonzalez-Nieto, Eva G. Rakasz, Gregory M. Hendricks, David I. Watkins, Mauricio A. Martins, Ronald C. Desrosiers, Daniel C. Douek.
The properties of the human immunodeficiency virus (HIV) pose serious difficulties for the development of an effective prophylactic vaccine. Here we describe the construction and characterization of recombinant (r), replication-competent forms of rhesus monkey rhadinovirus (RRV), a gamma-2 herpesvirus, containing a near-full-length (nfl) genome of the simian immunodeficiency virus (SIV). A 306-nucleotide deletion in the pol gene rendered this nfl genome replication-incompetent as a consequence of deletion of the active site of the essential reverse transcriptase enzyme. Three variations were constructed to drive expression of the SIV proteins: one with SIV’s own promoter region, one with a cytomegalovirus (cmv) immediate-early promoter/enhancer region, and one with an RRV dual promoter (p26 plus PAN). Following infection of rhesus fibroblasts in culture with these rRRV vectors, synthesis of the early protein Nef and the late structural proteins Gag and Env could be demonstrated. Expression levels of the SIV proteins were highest with the rRRV-SIVcmv-nfl construct. Electron microscopic examination of rhesus fibroblasts infected with rRRV-SIVcmv-nfl revealed numerous budding and mature SIV particles and these infected cells released impressive levels of p27 Gag protein (>150 ng/ml) into the cell-free supernatant. The released SIV particles were shown to be incompetent for replication. Monkeys inoculated with rRRV-SIVcmv-nfl became persistently infected, made readily-detectable antibodies against SIV, and developed T-cell responses against all nine SIV gene products. Thus, rRRV expressing a near-full-length SIV genome mimics live-attenuated strains of SIV in several important respects: the infection is persistent; >95% of the SIV proteome is naturally expressed; SIV particles are formed; and CD8+ T-cell responses are maintained indefinitely in an effector-differentiated state. Although the magnitude of anti-SIV immune responses in monkeys infected with rRRV-SIVcmv-nfl falls short of what is seen with live-attenuated SIV infection, further experimentation seems warranted.
There are good reasons for believing that development of an effective preventive vaccine against HIV-1 is going to be a very difficult task [1–3]. HIV is able to replicate continuously without relent despite apparently strong humoral and cellular immune responses to the virus. The HIV envelope glycoprotein is shielded with a large amount of carbohydrate and the trimer spike as it exists of the surface of virions is difficult for antibodies to access and difficult for antibodies to block infectivity. HIV-1 is highly variable from one individual to another and even within a single individual evolves to evade ongoing immune responses. The virus encodes a number of gene products that function at least in part to evade intrinsic, innate and adaptive immune responses. And during the course of an infection, HIV-1 gradually destroys CD4+ T lymphocytes, a key orchestrator of adaptive immune responses. The inability of infection by one HIV-1 strain to routinely provide protection against superinfection by a different HIV-1 strain supports this perception of great difficulty in development of a protective vaccine . Investigation of a variety of creative, non-standard approaches to a vaccine seem justified given this expected difficulty.
Herpesviruses have a number of potential advantages when being considered as vectors for vaccine delivery. Herpesviruses have large DNA genomes and can potentially accommodate significant amounts of inserted genetic information. That potential advantage has been borne out in our study described here in that 9,343 base pairs of genetic information have been successfully inserted. Being a DNA virus, inserts can be expected to be reasonably stable in the absence of direct repeats in the insert and with the absence of severe selective disadvantage. Furthermore, herpesviruses persist for the life of the infected individual and immune responses to their proteins persist in an up, on, active fashion for life. This is important when considering vaccine approaches for HIV/AIDS since immunological memory will probably never be enough to contain HIV-1; once a memory response kicks in for a previously-vaccinated, subsequently-infected individual, HIV-1 will employ its vast array of immune evasion and other strategies to allow continuous viral replication. Our results indeed indicate that rRRV-SIVnfl established persistent infection in RRV seronegative monkeys. Antibody responses increased to high levels and persisted through the 20 weeks of analysis. Similarly, cellular responses persisted to one extent or another in the 20 weeks of follow-up analyses. Also, the responding CD8+ T cells exhibited an effector memory phenotype consistent with recurrent antigen production. In contrast to the unconventionally MHC-restricted CD8+ TEM responses elicited by the 68.1 rhesus CMV vaccine developed by Hansen et al. [12–14], rRRV-SIVnfl-vaccinated macaques developed CD8+ T-cells capable of recognizing immunodominant SIV epitopes restricted by classical MHC class I molecules.