P17-09. Immunization with a single HIV-1 envelope sequence can generate CD8+ T lymphocytes capable of recognizing multiple variant forms of envelope

Background: The ability of CD8+ T lymphocytes to recognize a diversity of mutant forms of an HIV epitope is of central importance in the immune containment of this virus. The present studies were pursued to determine the mechanism employed by CD8+ T lymphocytes to recognize mutant viruses. In particular, we sought to determine whether mutant sequences are recognized by distinct CD8+ T lymphocyte populations or whether individual clonal populations of CD8+ T lymphocytes recognize a diversity of mutant sequences. Methods: We employed flow cytometry, Vβ repertoire analysis, and CDR3 sequencing methodologies to characterize the clonal diversity of CD8+ T lymphocytes that recognize variant forms of the HIV-1 envelope (Env) p41A epitope generated after infection by SHIV-89.6P or elicited by HIV-1 89.6P Env immunization of Mamu-A*01+ rhesus monkeys. To evaluate the capacity of the CD8+ T lymphocytes to recognize genetically diverse isolates of HIV-1, we employed a series of tetramers constructed with variants of the p41A epitope of HIV-1 Env. To define which T cell receptor mediated the recognition of each specific variant p41A, we isolated variant p41A-specific CD8+ T lymphocyte populations and analyzed the expression of 46 Vβ families and subfamilies genes. We then determined the precise clones employed for the recognition of each variant epitope peptide through CDR3 sequencing. Results: In both the infected and the vaccinated monkeys, we observed clonotypes capable of recognizing the majority of the variant epitope peptides. Conclusion: These data show that exposure to a single HIV-1 Env sequence can generate clonotypes capable of recognizing multiple variant forms of HIV-1 Env. Such Env-specific CD8+ T lymphocytes should be able to confer potent, effective protection against a diverse spectrum of circulating viruses

Contributions of Mamu-A*01 Status and TRIM5 Allele Expression, But Not CCL3L Copy Number Variation, to the Control of SIVmac251 Replication in Indian-Origin Rhesus Monkeys

CCL3 is a ligand for the HIV-1 co-receptor CCR5. There have recently been conflicting reports in the literature concerning whether CCL3-like gene (CCL3L) copy number variation (CNV) is associated with resistance to HIV-1 acquisition and with both viral load and disease progression following infection with HIV-1. An association has also been reported between CCL3L CNV and clinical sequelae of the simian immunodeficiency virus (SIV) infection in vivo in rhesus monkeys. The present study was initiated to explore the possibility of an association of CCL3L CNV with the control of virus replication and AIDS progression in a carefully defined cohort of SIVmac251-infected, Indian-origin rhesus monkeys. Although we demonstrated extensive variation in copy number of CCL3L in this cohort of monkeys, CCL3L CNV was not significantly associated with either peak or set-point plasma SIV RNA levels in these monkeys when MHC class I allele Mamu-A*01 was included in the models or progression to AIDS in these monkeys. With 66 monkeys in the study, there was adequate power for these tests if the correlation of CCL3L and either peak or set-point plasma SIV RNA levels was 0.34 or 0.36, respectively. These findings call into question the premise that CCL3L CNV is important in HIV/SIV pathogenesis

Variability in a dominant block to SIV early reverse transcription in rhesus monkey cells predicts in vivo viral replication and time to death

While it has long been appreciated that there is considerable variability in host containment of HIV/SIV replication, the determinants of that variability are not fully understood. Previous studies demonstrated that the degree of permissivity of a macaque's peripheral blood mononuclear cells (PBMC) for infection with simian immunodeficiency virus (SIV) in vitro predicted that animal's peak plasma virus RNA levels following SIV infection in vivo. The present study was conducted to define the mechanisms underlying the variable intrinsic susceptibility of rhesus monkey PBMC to SIVsmE660 infection. In a cohort of 15 unrelated Indian-origin rhesus monkeys, infectability of PBMC of individual animals with SIVsmE660, as defined by tissue culture infectious dose (TCID50), varied by more than 3 logs and was a stable phenotype over time. Susceptibility of a monkey's PBMC to wild type SIVsmE660 infection correlated with the susceptibility of that monkey's PBMC to infection with VSV-G pseudotyped SIVsm543-GFP. Moreover, the permissivity of an individual monkey's PBMC for infection with this construct correlated with the permissivity of a B-lymphoblastoid cell line (B-LCL) generated from PBMC of the same animal. We found that the degree of intrinsic resistance of monkey B-LCL correlated with the copy number of early reverse transcription (ERT) SIV DNA. The resistance of monkey B-LCL to SIVsmE660 replication could be abrogated by preincubation of cells with the SIV virus-like particles (VLPs) and SIV resistance phenotype could be transferred to a SIV susceptible B-LCL through cell fusion. Finally, we observed a positive correlation between susceptibility of monkey B-LCL to SIV infection with a VSV-G pseudotyped SIV-GFP construct in vitro and both the peak plasma virus RNA levels in vivo and time to death following wild type SIV infection. These findings suggest that a dominant early RT restricting factor that can be saturated by SIV capsid may contribute to the variable resistance to SIV infection in rhesus monkey B-LCL and that this differential intrinsic susceptibility contributes to the clinical outcome of an SIV infection

Association of Activating KIR Copy Number Variation of NK Cells with Containment of SIV Replication in Rhesus Monkeys

While the contribution of CD8+ cytotoxic T lymphocytes to early containment of HIV-1 spread is well established, a role for NK cells in controlling HIV-1 replication during primary infection has been uncertain. The highly polymorphic family of KIR molecules expressed on NK cells can inhibit or activate these effector cells and might therefore modulate their activity against HIV-1-infected cells. In the present study, we investigated copy number variation in KIR3DH loci encoding the only activating KIR receptor family in rhesus monkeys and its effect on simian immunodeficiency virus (SIV) replication during primary infection in rhesus monkeys. We observed an association between copy numbers of KIR3DH genes and control of SIV replication in Mamu-A*01– rhesus monkeys that express restrictive TRIM5 alleles. These findings provide further evidence for an association between NK cells and the early containment of SIV replication, and underscore the potential importance of activating KIRs in stimulating NK cell responses to control SIV spread

Early Divergent Host Responses in SHIVsf162P3 and SIVmac251 Infected Macaques Correlate with Control of Viremia

We previously showed intravaginal inoculation with SHIVsf162p3 results in transient viremia followed by undetectable viremia in most macaques, and some displayed subsequent immunity to superinfection with pathogenic SIVmac251. Here we compare early T cell activation, proliferation, and plasma cytokine/chemokine responses in macaques intravaginally infected with either SHIVsf162p3 or SIVmac251 to determine whether distinct differences in host responses may be associated with early viral containment. The data show SIVmac251 infection results in significantly higher levels of T cell activation, proliferation, and a mixed cytokine/chemokine “storm” in plasma in primary infection, whereas infection with SHIVsf162p3 resulted in significantly lower levels of T cell activation, proliferation, and better preservation of memory CD4+ T cells in early infection which immediately preceded control of viremia. These results support the hypothesis that early systemic immune activation, T cell proliferation, and a more prominent and broader array of cytokine/chemokine responses facilitate SIV replication, and may play a key role in persistence of infection, and the progression to AIDS. In contrast, immune unresponsiveness may be associated with eventual clearance of virus, a concept that may have key significance for therapy and vaccine design

Blocking TLR7- and TLR9-mediated IFN-α Production by Plasmacytoid Dendritic Cells Does Not Diminish Immune Activation in Early SIV Infection

Persistent production of type I interferon (IFN) by activated plasmacytoid dendritic cells (pDC) is a leading model to explain chronic immune activation in human immunodeficiency virus (HIV) infection but direct evidence for this is lacking. We used a dual antagonist of Toll-like receptor (TLR) 7 and TLR9 to selectively inhibit responses of pDC but not other mononuclear phagocytes to viral RNA prior to and for 8 weeks following pathogenic simian immunodeficiency virus (SIV) infection of rhesus macaques. We show that pDC are major but not exclusive producers of IFN-α that rapidly become unresponsive to virus stimulation following SIV infection, whereas myeloid DC gain the capacity to produce IFN-α, albeit at low levels. pDC mediate a marked but transient IFN-α response in lymph nodes during the acute phase that is blocked by administration of TLR7 and TLR9 antagonist without impacting pDC recruitment. TLR7 and TLR9 blockade did not impact virus load or the acute IFN-α response in plasma and had minimal effect on expression of IFN-stimulated genes in both blood and lymph node. TLR7 and TLR9 blockade did not prevent activation of memory CD4+ and CD8+ T cells in blood or lymph node but led to significant increases in proliferation of both subsets in blood following SIV infection. Our findings reveal that virus-mediated activation of pDC through TLR7 and TLR9 contributes to substantial but transient IFN-α production following pathogenic SIV infection. However, the data indicate that pDC activation and IFN-α production are unlikely to be major factors in driving immune activation in early infection. Based on these findings therapeutic strategies aimed at blocking pDC function and IFN-α production may not reduce HIV-associated immunopathology. © 2013 Kader et al

Simian-Human Immunodeficiency Infection – Is the Course Set in the Acute Phase?

Identifying early predictors of infection outcome is important for the clinical management of HIV infection, and both viral load and CD4+ T cell level have been found to be useful predictors of subsequent disease progression. Very high viral load or extensively depleted CD4+ T cells in the acute phase often result in failure of immune control, and a fast progression to AIDS. It is usually assumed that extensive loss of CD4+ T cells in the acute phase of HIV infection prevents the establishment of robust T cell help required for virus control in the chronic phase. We tested this hypothesis using viral load and CD4+ T cell number of SHIV-infected rhesus macaques. In acute infection, the lowest level of CD4+ T cells was a good predictor of later survival; animals having less than 3.3% of baseline CD4+ T cells progressed to severe disease, while animals with more than 3.3% of baseline CD4+ T cells experienced CD4+ T cell recovery. However, it is unclear if the disease progression was caused by early depletion, or was simply a result of a higher susceptibility of an animal to infection. We derived a simple relationship between the expected number of CD4+ T cells in the acute and chronic phases for a constant level of host susceptibility or resistance. We found that in most cases, the depletion of CD4+ T cells in chronic infection was consistent with the prediction from the acute CD4+ T cell loss. However, the animals with less than 3.3% of baseline CD4 T cells in the acute phase were approximately 20% more depleted late in the infection than expected based on constant level of virus control. This suggests that severe acute CD4 depletion indeed impairs the immune response

TRIM5α Modulates Immunodeficiency Virus Control in Rhesus Monkeys

The cytoplasmic TRIM5α proteins of certain mammalian lineages efficiently recognize the incoming capsids of particular retroviruses and potently restrict infection in a species-specific manner. Successful retroviruses have evolved capsids that are less efficiently recognized by the TRIM5α proteins of the natural hosts. To address whether TRIM5α contributes to the outcome of retroviral infection in a susceptible host species, we investigated the impact of TRIM5 polymorphisms in rhesus monkeys on the course of a simian immunodeficiency virus (SIV) infection. Full-length TRIM5α cDNAs were derived from each of 79 outbred monkeys and sequenced. Associations were explored between the expression of particular TRIM5 alleles and both the permissiveness of cells to SIV infection in vitro and clinical sequelae of SIV infection in vivo. Natural variation in the TRIM5α B30.2(SPRY) domain influenced the efficiency of SIVmac capsid binding and the in vitro susceptibility of cells from the monkeys to SIVmac infection. We also show the importance in vivo of the interaction of SIVmac with different allelic forms of TRIM5, demonstrating that particular alleles are associated with as much as 1.3 median log difference in set-point viral loads in SIVmac-infected rhesus monkeys. Moreover, these allelic forms of TRIM5 were associated with the extent of loss of central memory (CM) CD4+ T cells and the rate of progression to AIDS in the infected monkeys. These findings demonstrate a central role for TRIM5α in limiting the replication of an immunodeficiency virus infection in a primate host