The primary mechanism by which CD8 T cells contribute to controlling pathogens is by killing infected cells. The afflicted cells display pathogen epitopes on their surface via class I HLA molecules that constantly shuttle disused protein fragments to the cell surface in a manner akin to taking out the garbage. CD8 T cells that recognize the pathogen epitope in the context of an HLA molecule that matches the HLA molecule expressed by the CD8 T cell initiate a cascade of events that results in the infected cell being killed. Examples of the process have been captured on video (the Howard Hughes Medical Institute website has an excellent excerpt from a lecture by Bruce Walker showing such a video and explaining it). However, although this is the canonical mechanism of CD8 T cell activity (and the reason for their alternate moniker of cytotoxic T cell lymphocyte or CTL), CD8 T cells can also suppress pathogens by other means such as release of chemokines and cytokines and, in the case of HIV, an as-yet-unidentified antiviral factor that goes by the acronym of CAF. While there is a vast amount of evidence demonstrating the importance of CD8 T cells in suppressing replication of HIV (and other similar viruses such as SIV), the relative contributions of direct killing and indirect suppression have not been clearly delineated. Attempting to do so is a complex task, as the contribution of these activities may vary based on factors such as the extent of immunologic control and disease stage.
Two new studies in PLoS Pathogens make a valiant first attempt to shed light on this issue by evaluating whether artificial depletion of CD8 T cells impacts the speed of SIV suppression by antiretroviral therapy (ART) in macaques. The rationale of both studies is that the kinetics of ART-mediated viral suppression should be slower in the absence of CD8 T cells because infected cells will have a longer lifespan. In both experiments, no such effect is demonstrated, leading the researchers to argue that CD8 T cells do not impact the lifespan of productively infected cells and that therefore their primary mechanism of action in chronic SIV infection is indirect suppression as opposed to direct killing. In an accompanying commentary, Miles P. Davenport and Janka Petravic invoke the tortured language of former US Secretary of Defense Donald Rumsfeld and suggest that CD8 T cell activity against HIV may qualify as a “known unknown.”
But there are a number of issues that make interpreting these data complicated. Both studies involve chronically SIV-infected macaques, and CD8 T cell dysfunction and exhaustion in the setting of chronic infection is well documented. It could be that in a milieu in which viral control is only partial and both functional and dysfunctional SIV-specific CD8 T cells co-exist, indirect suppression plays a larger role than in the setting of robust immunological control (e.g. elite controllers). The selection pressure imposed upon SIV and HIV by HLA-restricted CD8 T cell responses is also extremely well documented, and although the authors of these new papers argue that indirect suppression could also exert selective pressure this scenario is somewhat difficult to envision. It is straightforward to grasp how a virus with an immune escape mutation could survive as a result of CD8 T cells being unable to kill the cell it is occupying. But in the case of indirect suppression, the escape mutation would have to abrogate localized release of suppressive substances by virus-specific CD8 T cells; it seems unlikely that the epitope specificities of CD8 T cells in a localized environment would be so uniform that a single virus carrying an escape mutation would switch off the suppression to a sufficient degree to obtain a survival advantage (although this question may be amenable to study).
Both papers note the possibility that CD8 T cells might kill virus-infected cells prior to the release of new virions, therefore making their activity essentially invisible in this particularly experimental system. The data cited in support of this possibility comes from Jonah Sacha, who has shown that epitopes from incoming virions can be processed and presented to CD8 T cells prior to virus integration, leading to killing of infected cells before the establishment of productive infection. Another possibility is that CD8 T cell-mediated killing reduces the average number of virions produced by each infected cell while having only a minor impact on the average lifespan (I think the mathematical model used in these papers assumes the same average virion production – or “burst size” – for every infected cell). Due to considerable variation from animal to animal, relatively subtle differences in infected cell lifespan may not be easy to capture.
Despite all the complexity and caveats, the papers are provocative and highlight the need to better understand the mechanism of action of CD8 T cells in SIV, and by extension, HIV infection.
In a related development, a group of researchers headed by members of the Human Immunology Laboratory at the International AIDS Vaccine Initiative (IAVI) have just published details of an assay that measures HIV inhibition by CD8 T cells in vitro. Although the numbers of people studied is small, the paper reports that inhibition measured by the assay correlated with viral load control, i.e. CD8 T cell-mediated inhibition was strong among untreated individuals with viral loads <10,000 copies/mL but weak in those above that threshold. The researchers also use the assay to measure the inhibitory capacity of CD8 T cells from seven uninfected individuals immunized with a DNA/Ad5 HIV vaccine regimen, reporting that significant inhibition could be documented but only after receipt of the Ad5 boost. Importantly, there was no correlation between the degree of in vitro virus inhibition and the numbers of vaccine-induced CD8 T cell responses measured by ELISpot, which up until now has been the standard way to measure the immunogenicity of T cell-based vaccine candidates. The new assay takes three weeks to run and involves less than 2 million cells, making it more practical than those developed previously. The study authors write: “We believe the viral inhibition assay will be a useful tool in the study of HIV‐1 pathogenesis and vaccine development, complementing existing methods used to prioritize candidates for further trials.”
PLoS Pathog 6(1): e1000747. doi:10.1371/journal.ppat.1000747
CD8+ Lymphocytes Control Viral Replication in SIVmac239-Infected Rhesus Macaques without Decreasing the Lifespan of Productively Infected Cells
Nichole R. Klatt1,2, Emi Shudo3, Alex M. Ortiz1, Jessica C. Engram1, Mirko Paiardini1, Benton Lawson2, Michael D. Miller4, James Else2, Ivona Pandrea5, Jacob D. Estes6, Cristian Apetrei5, Joern E. Schmitz7, Ruy M. Ribeiro3, Alan S. Perelson3, Guido Silvestri1,2
1 Department of Pathology and Laboratory Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America, 2 Yerkes National Primate Research Center, Emory University, Atlanta, Georgia, United States of America, 3 Theoretical Biology and Biophysics, Los Alamos National Laboratory, Los Alamos, New Mexico, United States of America, 4 Gilead Sciences, Inc., Foster City, California, United States of America, 5 Tulane National Primate Research Center and Tulane Health Sciences Center, Tulane University, New Orleans, Louisiana, United States of America, 6 AIDS and Cancer Virus Program, Science Applications International Corporation-Frederick, Inc., National Cancer Institute, Frederick, Maryland, United States of America, 7 Beth Israel Deaconess Medical Center, Harvard Medical School, Boston, Massachusetts, United States of America
While CD8+ T cells are clearly important in controlling virus replication during HIV and SIV infections, the mechanisms underlying this antiviral effect remain poorly understood. In this study, we assessed the in vivo effect of CD8+ lymphocyte depletion on the lifespan of productively infected cells during chronic SIVmac239 infection of rhesus macaques. We treated two groups of animals that were either CD8+ lymphocyte-depleted or controls with antiretroviral therapy, and used mathematical modeling to assess the lifespan of infected cells either in the presence or absence of CD8+ lymphocytes. We found that, in both early (day 57 post-SIV) and late (day 177 post-SIV) chronic SIV infection, depletion of CD8+ lymphocytes did not result in a measurable increase in the lifespan of either short- or long-lived productively infected cells in vivo. This result indicates that the presence of CD8+ lymphocytes does not result in a noticeably shorter lifespan of productively SIV-infected cells, and thus that direct cell killing is unlikely to be the main mechanism underlying the antiviral effect of CD8+ T cells in SIV-infected macaques with high virus replication.
PLoS Pathog 6(1): e1000748. doi:10.1371/journal.ppat.1000748
In Vivo CD8+ T-Cell Suppression of SIV Viremia Is Not Mediated by CTL Clearance of Productively Infected Cells
Joseph K. Wong1,2#*, Matthew C. Strain2, Rodin Porrata3, Elizabeth Reay4, Sumathi Sankaran-Walters4, Caroline C. Ignacio2, Theresa Russell2, Satish K. Pillai1,2, David J. Looney2, Satya Dandekar4#
1 Department of Medicine, University of California San Francisco, San Francisco, California, United States of America, 2 Department of Medicine, VA San Diego Healthcare System, University of California San Diego, La Jolla, California, United States of America, 3 Department of Physics, University of California Berkeley, Berkeley, California, United States of America, 4 Department of Medical Microbiology and Immunology, University of California Davis, Davis, California, United States of America
The CD8+ T-cell is a key mediator of antiviral immunity, potentially contributing to control of pathogenic lentiviral infection through both innate and adaptive mechanisms. We studied viral dynamics during antiretroviral treatment of simian immunodeficiency virus (SIV) infected rhesus macaques following CD8+ T-cell depletion to test the importance of adaptive cytotoxic effects in clearance of cells productively infected with SIV. As previously described, plasma viral load (VL) increased following CD8+ T-cell depletion and was proportional to the magnitude of CD8+ T-cell depletion in the GALT, confirming a direct relationship between CD8+ T-cell loss and viral replication. Surprisingly, first phase plasma virus decay following administration of antiretroviral drugs was not slower in CD8+ T-cell depleted animals compared with controls indicating that the short lifespan of the average productively infected cell is not a reflection of cytotoxic T-lymphocyte (CTL) killing. Our findings support a dominant role for non-cytotoxic effects of CD8+ T-cells on control of pathogenic lentiviral infection and suggest that cytotoxic effects, if present, are limited to early, pre-productive stages of the viral life cycle. These observations have important implications for future strategies to augment immune control of HIV.
PLoS Pathog 6(1): e1000728. doi:10.1371/journal.ppat.1000728
CD8+ T Cell Control of HIV—A Known Unknown
Miles P. Davenport*, Janka Petravic
Complex Systems in Biology Group, Centre for Vascular Research, University of New South Wales, Sydney, New South Wales, Australia
The Journal of Infectious Diseases 2010;201:720–729
DOI: 10.1086/650492
MAJOR ARTICLE
Viral Inhibition Assay: A CD8 T Cell Neutralization Assay for Use in Clinical Trials of HIV‐1 Vaccine Candidates
Aggeliki Spentzou,1,a Philip Bergin,1,a Dilbinder Gill,1 Hannah Cheeseman,1 Ambreen Ashraf,1 Harry Kaltsidis,1 Michelle Cashin‐Cox,1 Insiyah Anjarwalla,1 Alan Steel,2 Christopher Higgs,2 Anton Pozniak,2 Alicja Piechocka‐Trocha,3 Johnson Wong,4 Omu Anzala,6 Etienne Karita,7 Len Dally,5 Frances Gotch,1 Bruce Walker,3 Jill Gilmour,1 and Peter Hayes1
1Human Immunology Laboratory, International AIDS Vaccine Initiative, Imperial College London, and 2Chelsea and Westminster Hospital, London, United Kingdom; 3Ragon Institute of MGH, MIT and Harvard and 4Massachusetts General Hospital, Boston; 5Emmes Corporation, Rockville, Maryland; 6Kenya AIDS Vaccine Initiative, University of Nairobi, Nairobi, Kenya; 7Projet San Francisco, Kigali, Rwanda
We have characterized an assay measuring CD8 T cell–mediated inhibition of human immunodeficiency virus (HIV) type 1 replication, demonstrating specificity and reproducibility and employing a panel of primary HIV‐1 isolates. The assay uses relatively simple autologous cell culture and enzyme‐linked immunosorbent assay, avoids generation of T cell clones, and can be performed with <2 million peripheral blood mononuclear cells. Efficient CD8 T cell–mediated cross‐clade inhibition of HIV‐1 replication in vitro was demonstrated in antiretroviral therapy–naive HIV‐1–infected subjects with controlled viral replication in vivo but not in viremic subjects. An HIV‐1 vaccine candidate, consisting of DNA and recombinant adenovirus 5 vectors tested in a phase I clinical trial, induced CD8 T cells that efficiently inhibited HIV‐1 in a HLA‐I–dependent manner. Assessment of direct antiviral T cell function by this assay provides additional information to guide vaccine design and the prioritizing of candidates for further clinical trials.