Julian Sutton

Characterization of CD4+ CTLs Ex Vivo

The cytotoxic potential of CD8+ T cells and NK cells plays a crucial role in the immune response to pathogens. Although in vitro studies have reported that CD4+ T cells are also able to mediate perforin-mediated killing, the in vivo existence and relevance of cytotoxic CD4+ T cells have been the subject of debate. Here we show that a population of CD4+ perforin+ T cells is present in the circulation at low numbers in healthy donors and is markedly expanded in donors with chronic viral infections, in particular HIV infection, at all stages of the disease, including early primary infection. Ex vivo analysis shows that these cells have cytotoxic potential mediated through the release of perforin. In comparison with more classical CD4+ T cells, this subset displays a distinct surface phenotype and functional profile most consistent with end-stage differentiated T cells and include Ag experienced CD4+ T cells. The existence of CD4+ cytotoxic T cells in vivo at relatively high levels in chronic viral infection suggests a role in the immune response.

Development of a DNA-MVA/HIVA vaccine for Kenya.

Without going into the details of the devastation that human immunodeficiency virus (HIV) infection causes especially in the developing world, the best hope for changing the course of this epidemic is development of a safe, effective, accessible prophylactic HIV vaccine. While the inaccessibility of potentially neutralising epitopes on primary HIV isolates has hampered the development of envelope-based vaccines, there is a number of new potent technologies capable of inducing high levels of circulating virus-specific CD8(+) cytotoxic T lymphocytes (CTL). Our original finding that a successive immunisation with DNA and modified vaccinia virus Ankara (MVA) vaccines expressing a common immunogen is a potent way of inducing CD8(+) CTL, which has been since reinforced by us and others, prompted us to test this approach in humans. With the view of proceeding into a high-risk cohort in Kenya for the efficacy trial, we designed the immunogen, termed HIVA, to match the HIV strain responsible locally for over 70% infections. It consists of a consensus clade A gag p24/p17 and a string of clade A-derived CTL epitopes. Pre-clinical studies demonstrated high immunogenicities of both the pTHr.HIVA and MVA.HIVA vaccines. In mice, these induced strong T cells-mediated immune responses which lasted at least 155 days. In rhesus macaques, the prime-boost immunisation elicited T cell responses specific for multiple HIV-derived epitopes. Phase I trials in healthy low-risk volunteers have commenced in Oxford and Nairobi, and the preliminary immunogenicity analysis from the Oxford site indicated that both vaccine components alone induced T cell responses in a majority of volunteers. These results have boosted expectations for the prime-boost vaccinations.

Comparison between HIV‐ and CMV‐specific T cell responses in long‐term HIV infected donors

The mechanisms underlying non‐progression in HIV‐1 infection are not well understood; however, this state has been associated previously with strong HIV‐1‐specific CD8+ T cell responses and the preservation of proliferative CD4+ T cell responses to HIV‐1 antigens. Using a combination of interferon‐gamma (IFN‐γ) ELISpot assays and tetramer staining, the HIV‐1‐specific CD8+ T cell populations were quantified and characterized in untreated long‐term HIV‐1‐infected non‐progressors and individuals with slowly progressive disease, both in relation to CD4+ T cell responses, and in comparison with responses to cytomegalovirus (CMV) antigens. High levels of CD8+ T cell responses specific for HIV‐1 or CMV were observed, but neither their frequency nor their phenotype seemed to differ between the two patient groups. Moreover, while CMV‐specific CD4+ T cell responses were preserved in these donors, IFN‐γ release by HIV‐1‐specific CD4+ T cells was generally low. These data raise questions with regard to the role played by CD8+ T cells in the establishment and maintenance of long‐term non‐progression.

Correlates of Delayed Disease Progression in HIV-1-Infected Kenyan Children

Without treatment most HIV-1-infected children in Africa die before their third birthday (>89%) and long-term nonprogressors are rare. The mechanisms underlying nonprogression in HIV-1-infected children are not well understood. In the present study, we examined potential correlates of delayed HIV disease progression in 51 HIV-1-infected African children. Children were assigned to progression subgroups based on clinical characterization. HIV-1-specific immune responses were studied using a combination of ELISPOT assays, tetramer staining, and FACS analysis to characterize the magnitude, specificity, and functional phenotype of HIV-1-specific CD8+ and CD4+ T cells. Host genetic factors were examined by genotyping with sequence-specific primers. HIV-1 nef gene sequences from infecting isolates from the children were examined for potential attenuating deletions. Thymic output was measured by T cell rearrangement excision circle assays. HIV-1-specific CD8+ T cell responses were detected in all progression groups. The most striking attribute of long-term survivor nonprogressors was the detection of HIV-1-specific CD4+ Th responses in this group at a magnitude substantially greater than previously observed in adult long-term nonprogressors. Although long-term survivor nonprogressors had a significantly higher percentage of CD45RA+CD4+ T cells, nonprogression was not associated with higher thymic output. No protective genotypes for known coreceptor polymorphisms or large sequence deletions in the nef gene associated with delayed disease progression were identified. In the absence of host genotypes and attenuating mutations in HIV-1 nef, long-term surviving children generated strong CD4+ T cell responses to HIV-1. As HIV-1-specific helper cells support anti-HIV-1 effector responses in active disease, their presence may be important in delaying disease progression.

Persistent HIV-1-specific cellular responses despite prolonged therapeutic viral suppression

Design Antiretroviral therapy (ART) currently represents the best way to avert the lethal consequences of chronic persistent HIV-1 infection. It leads to significant reductions of plasma viremia, often to undetectable levels, but it can also be linked with the reduction and disappearance of detectable HIV-specific CD8 T-cell responses. Results Here we describe a group of patients in whom ongoing replication of HIV, particularly transcription of Nef mRNA species, was detected despite prolonged and clinically successful antiretroviral treatment. Modest, but significant, numbers of HIV-specific CD8 T cells and CD4 T-cell responses were found in these subjects, with the strongest responses directed towards Nef epitopes. Detailed phenotypic analysis of the HIV-specific CD8 cells demonstrated low perforin levels and persistent expression of CD27, a phenotype associated with incomplete differentiation of cytotoxic T lymphocytes (CTL). Conclusion This immature CTL phenotype has been described previously in association with chronic HIV disease, but its continued persistence is surprising in the setting of prolonged viral suppression on therapy and the presence of HIV-specific CD4 cell activity.

Transition from long-term nonprogression to HIV-1 disease associated with escape from cellular immune control

Transition from long-term nonprogressive infection to progressive HIV-1 disease presents an opportunity to investigate pathogenesis in a defined immunogenetic background. We studied a male long-term nonprogressor (LTNP) who remained asymptomatic and viremic and had normal CD4 T-cell counts without antiretroviral therapy for >18 years and then experienced a transition to disease progression. We analyzed the complete HIV-1 genomic RNA sequence from plasma and cellular immune responses to predefined human leukocyte antigen-matched autologous viral peptides spanning the viral genome, before and after progression. Serial viral sequences did not seem attenuated and consistently utilized coreceptor CCR5. LTNP status was associated with elongated V2 domains and broad low-level T-cell immune responses targeting several regions of the viral genome. The transition to progressive disease was associated with the acquisition of viral mutations conferring escape from CD8 T-cell responses. Multiple changes in HIV-1 sequence and loss of immune response over time most likely contributed to the transition from LTNP status to progressive disease. These data are relevant to vaccine design and identification of the correlates of protection from disease progression.