Human endogenous retroviruses (HERVs) are thought to be inactive elements of our genome, normally held in check by host cellular restriction proteins such as APOBEC3G (11
). We hypothesized that the impairment of these controls in HIV-1 infection could expose the immune system to HERV antigens that act as immutable targets for cytotoxic lysis, resulting in the containment of HIV-1 viremia. Previously, our lab has shown that HERV expression occurs in HIV-1-infected cells and that HERV-specific CD8+
T cell responses are stimulated in primary HIV-1 infection (12
). These responses are negatively correlated with HIV-1 viral load (12
). Here, we extend these observations to include individuals who have long-term chronic HIV-1 infection, focusing on a rare subset of individuals who are able to suppress HIV-1 indefinitely in the absence of combination therapy. The mechanism of control in these HIV-1 controllers remains to be fully defined. Although there is evidence that the HIV-specific T cell response contributes to virologic containment (1
), it is clear from a number of studies that other factors are likely to be involved (10
). A recent genome-wide association study in a multiethnic cohort of HIV-1 controllers and progressors showed that differences in HLA alleles explain 19% of the variance of host control (27
). In order to determine what effect, if any, HERV-specific T cell responses have on viral control, we studied a cohort of HIV-1 controllers and compared their responses to those of untreated virologic noncontrollers and immunologic progressors, patients on HAART, and HIV-1-uninfected controls. While it is likely that robust CD4+
T cell help contributes to strong CD8+
T cell responses, we sought to rule out that it was the primary cause of any observed differential responses to HERV antigens. Therefore, to avoid a confounding effect of generalized immune dysfunction due to a lack of CD4+
T cell help, we performed additional analyses that excluded the immunologic progressor group. Our finding that the magnitude of CMV pp65-specific responses was similar in the controller, HAART-suppressed, and noncontroller groups supports the fulfillment of this criterion.
We found that in chronic HIV-1 infection, controllers have HERV responses with higher magnitude and greater breadth than patients with viral suppression on HAART, virologic noncontrollers, immunologic progressors, and HIV-1-negative subjects. Interestingly, controllers who lack HLA alleles that are associated with protection from HIV-1 disease progression (HLA-B27 and -B57) constituted a large proportion of the subjects with the strongest HERV responses, suggesting that there may be an alternative mechanism of HIV control (such as HERV-specific cytotoxic T cells) in these controllers. There was no difference in the magnitude of the HIV-1-specific responses between the controllers, noncontrollers, and progressors (as defined by IFN-γ production) and no correlation between this measure and HIV-1 plasma viremia. These data are consistent with other studies reporting that neither the magnitude nor the breadth of the HIV-1-specific CD8+
T cell response is associated with a difference in viral load (2
). Of note, there was also no significant correlation between the strength of HERV-specific responses and HIV-1-specific responses in untreated patients, suggesting that these are independent variables. In contrast, we discovered that the HERV-specific T cell response magnitude was inversely correlated with HIV-1 viral load (even when the immunologic progressors were excluded), suggesting a potential role for these responses in the control of HIV-1. There was also a positive correlation between HERV response magnitude and CD4+
T cell count in untreated individuals. The group with viral suppression on HAART responded to fewer HERV epitopes and had weaker responses, lending support to the hypothesis that these responses are a cause rather than a consequence of viral control.
Longitudinal analyses of HERV responses in two subjects revealed interesting differences. One individual (P2) had a strong, titratable response to the HERV-K FAFTIPAI epitope. This person was able to contain HIV-1 replication to low levels and maintain normal CD4+ T cell counts for almost a decade. Interestingly, the patient eventually initiated HAART, and we observed a decline in the magnitude of this person's HERV response in the few years preceding this juncture. In contrast, another subject (elite controller P1) with more than 15 years of undetectable HIV-1 viral load and normal CD4+ T cell count without HAART had an increase in his response to a different HERV-K epitope over 5 years. These data are generally consistent with our primary hypothesis that the generation and maintenance of HERV-specific responses may contribute to virus control.
To investigate possible mechanisms of protection of the HERV-specific response, we took advantage of P2's comparably strong ELISPOT responses to two very similar HERV and HIV-1 peptides (HERV-K FI8 and HIV-1 TI8). After defining the HLA restriction (B51) of the HERV response, we compared its functionality and phenotype to those of the HIV-1 epitope and a CMV pp65 peptide pool. We found that within the HERV-K FI8-specific population, there were as many cells producing two cytokines as there were producing just one. In contrast, the HIV response was dominated by monofunctional (IFN-γ-secreting) cells. IFN-γ secretion is a first-line antiviral defense mechanism and promotes the expression of TNF-α receptors on the cell surface, among other functions (28
). TNF-α, in turn, has broad, beneficial effects in protective immunity and can kill virally infected target cells by binding to the cell surface receptor and triggering an apoptosis signaling cascade (reviewed in reference 21
). The combination of the two cytokines is therefore critical for an effective immune response and has been shown to clear hepatitis B from hepatocytes and lymphocytic choriomeningitis virus (LCMV) from acutely infected mice (14
). HIV-1-specific CD8+
T cells that can generate a multifunctional response have been linked to slower disease progression, and polyfunctional T cell responses are associated with chronic control of other viral infections such as CMV, Epstein-Barr virus (EBV), and influenza virus (6
). We were surprised to find that even a HERV epitope that shares close homology with an HIV-1 sequence (having six out of eight amino acids in common) could trigger a distinct CD8+
T cell response. Given the inverse correlation between HIV-1 viral load and the HERV response in our cohort, we might expect this pattern to hold true for the total HERV-specific and HIV-1-specific CD8+
T cell population in this individual and others, though this remains to be tested.
The observed differences in functionality of the HERV and HIV-1 responses in our subject hinted that there could also be a divergence in their phenotypic parameters, leading us to examine their differentiation and activation profiles. Dual staining with the HLA-B51 tetramers confirmed that two distinct and specific T cell populations that correspond to each of these antigens exist and also ruled out the potential of a cross-reactive effect. In our previous report (12
), we compared the phenotype of CD8+
T cells responding to a CMV pool and to two unique HERV and HIV-1 peptides without a high degree of sequence homology. Unlike the maturationally deficient HIV-1-specific population, the HERV-specific CD8+
T cells were similar to the CMV-specific CD8+
T cells in having a terminally differentiated phenotype that is associated with improved viral control (25
). Our present study confirms this even though the two HERV and HIV-1 epitopes were similar and also demonstrates that the HERV response was comparable to the CMV response in its level of activation. The HERV-specific population had a late-differentiation (CD27−
) and low-activation (as measured by CD38 MFI) phenotype, which has previously been associated with greater cytotoxic activity (3
). In contrast, the HIV-1-specific population was more immature and had a higher level of activation. A disparity in activation level between HIV-1-specific cells and CMV-specific cells within the same individual was also recently reported by Barbour, et al. (5
). A lack of CD38 on the surface of virus-specific T cells has been correlated with control of infections, including HIV-1 and EBV, and has been linked to polyfunctional cytokine production, MIP-1β chemokine secretion, proliferative capacity, and lack of exhaustion as measured by PD-1 expression (6
). Conversely, HIV-specific CD8+
T cells from viremic patients tend to be CD38+
and monofunctional (secreting only IFN-γ) (6
). While the combination of these measures was not examined in the current study, the lack of CD38 expression on the HERV-specific T cell population suggests that this epitope stimulates cytotoxic T lymphocytes (CTLs) with these favorable characteristics and warrants additional exploration.
Another factor contributing to the ability of HERV-specific CTLs to proliferate and kill target cells may be linked to their level of avidity. As HERVs are encoded in the human genome and therefore represent self-antigens, T cells directed against them may bind to their cognate epitopes with low avidity. This is consistent with the increased peptide concentrations required to generate a response in many of our subjects. Low-avidity T cells (which are more prone to tolerance induction and are stimulated only with high antigen load) have been shown to proliferate better than high-avidity T cells and to lack their replicative defects (16
). Therefore, HERV-specific CTLs may play a role in the containment of HIV-1 due to a superior ability to proliferate. Of note, we previously showed that despite being directed against self-antigens, HERV-specific CTLs are not impaired in their ability to kill cells pulsed with their cognate peptide (12
). Overall, our present study has identified phenotypic and functional traits of the HERV-specific response that may confer an enhanced ability beyond that of most HIV-specific CTLs to effectively target and lyse infected cells. Our finding that the HIV-1 viral load was lower in patients with stronger and broader HERV responses provides empirical evidence for this.
Our results constitute the discovery of a novel correlate of immune protection from disease progression in chronic HIV-1 infection and have implications for both a better understanding of HIV-1 pathogenesis and a potential new approach to HIV-1 vaccines. While the primary advantage of a therapeutic agent derived from HERVs is their conserved nature, features associated with a superior cytotoxic ability offer an additional benefit to the induction of HERV-specific responses. Our findings show that individuals who can control HIV-1 in the absence of HAART have the greatest HERV responses. This HERV-specific immunity is correlated with the containment of HIV-1 replication and preservation of CD4+ T cell count in untreated subjects and provides strong support for the further investigation of an HERV-based HIV-1 vaccine.