Xintao Hu

Neutralization sensitivity of HIV-1 subtype B'clinical isolates from former plasma donors in China

BackgroundHIV-1 subtype B’ isolates have been predominantly circulating in China. Their intra- and inter-subtype neutralization sensitivity to autologous and heterologous plasmas has not been well studied.ResultsTwelve HIV-1 B’ clinical isolates obtained from patients were tested for their intra- and inter-subtype neutralization sensitivity to the neutralization antibodies in the plasmas from patients infected by HIV-1 B’ and CRF07_BC subtypes, respectively. We found that the plasmas from the HIV-1 B’-infected patients could potently neutralize heterologous viruses of subtype B’ with mean ID50 titer (1/x) of about 67, but they were not effective in neutralizing autologous viruses of subtype B’ with mean ID50 titer (1/x) of about 8. The plasmas from HIV-1 CRF07_BC-infected patients exhibited weak inter-subtype neutralization activity against subtype B’ viruses with ID50 titer (1/x) is about 22. The neutralization sensitivity of HIV-1 B’ isolates was inversely correlated with the neutralizing activity of plasmas from HIV-1 B’-infected patients (Spearman’s r = −0.657, P = 0.020), and with the number of potential N-glycosylation site (PNGS) in V1-V5 region (Spearman’s r = −0.493, P = 0.034), but positively correlated with the viral load (Spearman’s r = 0.629, P = 0.028). It had no correlation with the length of V1-V5 regions or the CD4+ T cell count. Virus AH259V has low intra-subtype neutralization sensitivity, it can be neutralized by 17b (IC50: 10μg/ml) and 447-52D (IC50: 1.6μg/ml), and the neutralizing antibodies (nAbs) in plasma AH259P are effective in neutralizing infection by the primary HIV-1 isolates with different subtypes with ID50 titers (1/x) in the range of 32–396.ConclusionsThese findings suggest that the HIV-1 subtype B’ viruses may mutate under the immune pressure, thus becoming resistant to the autologous nAbs, possibly by changing the number of PNGS in the V1-V5 region of the viral gp120. Some of primary HIV-1 isolates are able to induce both intra- and inter-subtype cross-neutralizing antibody responses.

DNA Prime-Boost Vaccine Regimen To Increase Breadth, Magnitude, and Cytotoxicity of the Cellular Immune Responses to Subdominant Gag Epitopes of Simian Immunodeficiency Virus and HIV

HIV sequence diversity and the propensity of eliciting immunodominant responses targeting variable regions of the HIV proteome are hurdles in the development of an effective AIDS vaccine. An HIV-derived conserved element (CE) p24gag plasmid DNA (pDNA) vaccine is able to redirect immunodominant responses to otherwise subdominant and often more vulnerable viral targets. By homology to the HIV immunogen, seven CE were identified in SIV p27Gag. Analysis of 31 rhesus macaques vaccinated with full-length SIV gag pDNA showed inefficient induction (58% response rate) of cellular responses targeting these CE. In contrast, all 14 macaques immunized with SIV p27CE pDNA developed robust T cell responses recognizing CE. Vaccination with p27CE pDNA was also critical for the efficient induction and increased the frequency of Ag-specific T cells with cytotoxic potential (granzyme B+ CD107a+) targeting subdominant CE epitopes, compared with the responses elicited by the p57gag pDNA vaccine. Following p27CE pDNA priming, two booster regimens, gag pDNA or codelivery of p27CE+gag pDNA, significantly increased the levels of CE-specific T cells. However, the CE+gag pDNA booster vaccination elicited significantly broader CE epitope recognition, and thus, a more profound alteration of the immunodominance hierarchy. Vaccination with HIV molecules showed that CE+gag pDNA booster regimen further expanded the breadth of HIV CE responses. Hence, SIV/HIV vaccine regimens comprising CE pDNA prime and CE+gag pDNA booster vaccination significantly increased cytotoxic T cell responses to subdominant highly conserved Gag epitopes and maximized response breadth.

Heterodimeric IL15 treatment enhances tumor infiltration, persistence, and effector functions of adoptively transferred tumor-specific T cells in the absence of lymphodepletion

Purpose: Adoptive cell transfer (ACT) is a promising immunotherapeutic approach for cancer. Host lymphodepletion is associated with favorable ACT therapy outcomes, but it may cause detrimental effects in humans. We tested the hypothesis that IL15 administration enhances ACT in the absence of lymphodepletion. We previously showed that bioactive IL15 in vivo comprises a stable complex of the IL15 chain with the IL15 receptor alpha chain (IL15Rα), termed heterodimeric IL15 (hetIL15). Experimental Design: We evaluated the effects of the combination regimen ACT + hetIL15 in the absence of lymphodepletion by transferring melanoma-specific Pmel-1 T cells into B16 melanoma-bearing mice. Results: hetIL15 treatment delayed tumor growth by promoting infiltration and persistence of both adoptively transferred Pmel-1 cells and endogenous CD8+ T cells into the tumor. In contrast, persistence of Pmel-1 cells was severely reduced following irradiation in comparison with mice treated with hetIL15. Importantly, we found that hetIL15 treatment led to the preferential enrichment of Pmel-1 cells in B16 tumor sites in an antigen-dependent manner. Upon hetIL15 administration, tumor-infiltrating Pmel-1 cells showed a “nonexhausted” effector phenotype, characterized by increased IFNγ secretion, proliferation, and cytotoxic potential and low level of PD-1. hetIL15 treatment also resulted in an improved ratio of Pmel-1 to Treg in the tumor. Conclusions: hetIL15 administration improves the outcome of ACT in lymphoreplete hosts, a finding with significant implications for improving cell-based cancer immunotherapy strategies. Clin Cancer Res; 23(11); 2817–30. ©2016 AACR.

HIV Env conserved element DNA vaccine alters immunodominance in macaques

ABSTRACT Sequence diversity and immunodominance are major obstacles in the design of an effective vaccine against HIV. HIV Env is a highly-glycosylated protein composed of ‘conserved’ and ‘variable’ regions. The latter contains immunodominant epitopes that are frequently targeted by the immune system resulting in the generation of immune escape variants. This work describes 12 regions in HIV Env that are highly conserved throughout the known HIV M Group sequences (Env CE), and are poorly immunogenic in macaques vaccinated with full-length Env expressing DNA vaccines. Two versions of plasmids encoding the 12 Env CE were generated, differing by 0–5 AA per CE to maximize the inclusion of commonly detected variants. In contrast to the full-length env DNA vaccine, vaccination of macaques with a combination of these 2 Env CE DNA induced robust, durable cellular immune responses with a significant fraction of CD8+ T cells with cytotoxic phenotype (Granzyme B+ and CD107a+). Although inefficient in generating primary responses to the CE, boosting of the Env CE DNA primed macaques with the intact env DNA vaccine potently augmented pre-existing immunity, increasing magnitude, breadth and cytotoxicity of the cellular responses. Fine mapping showed that 7 of the 12 CE elicited T cell responses. Env CE DNA also induced humoral responses able to recognize the full-length Env. Env CE plasmids are therefore capable of inducing durable responses to highly conserved regions of Env that are frequently absent after Env vaccination or immunologically subdominant. These modified antigens are candidates for use as prophylactic and therapeutic HIV vaccines.

Therapeutic conserved elements (CE) DNA vaccine induces strong T-cell responses against highly conserved viral sequences during simian-human immunodeficiency virus infection

ABSTRACT HIV-specific T-cell responses play a key role in controlling HIV infection, and therapeutic vaccines for HIV that aim to improve viral control will likely need to improve on the T-cell responses induced by infection. However, in the setting of chronic infection, an effective therapeutic vaccine must overcome the enormous viral genetic diversity and the presence of pre-existing T-cell responses that are biased toward immunodominant T-cell epitopes that can readily mutate to evade host immunity and thus potentially provide inferior protection. To address these issues, we investigated a novel, epidermally administered DNA vaccine expressing SIV capsid (p27Gag) homologues of highly conserved elements (CE) of the HIV proteome in macaques experiencing chronic but controlled SHIV infection. We assessed the ability to boost or induce de novo T-cell responses against the conserved but immunologically subdominant CE epitopes. Two groups of animals were immunized with either the CE DNA vaccine or a full-length SIV p57gag DNA vaccine. Prior to vaccination, CE responses were similar in both groups. The full-length p57gag DNA vaccine, which contains the CE, increased overall Gag-specific responses but did not increase CE responses in any animals (0/4). In contrast, the CE DNA vaccine increased CE responses in all (4/4) vaccinated macaques. In SIV infected but unvaccinated macaques, those that developed stronger CE-specific responses during acute infection exhibited lower viral loads. We conclude that CE DNA vaccination can re-direct the immunodominance hierarchy towards CE in the setting of attenuated chronic infection and that induction of these responses by therapeutic vaccination may improve immune control of HIV.

Profiling the neutralizing antibody response in chronically HIV-1 CRF07_BC-infected intravenous drug users naïve to antiretroviral therapy

Characterizing neutralizing antibody (NAb) responses in individuals infected with diverse HIV-1 strains is necessary to reveal the novel targets for regional preventive and therapeutic strategies development. We evaluated the prevalence, breadth, and potency of NAb responses in 98 CRF07_BC-infected individuals using a large, multi-subtype panel of 30 tier 2-3 Env-pseudotyped viruses. Furthermore, we compared the neutralization pattern of CRF07_BC-infected people with that of subtype B’-infected individuals in China. Of the 98 plasma samples tested, 18% neutralized more than 80% of viruses in the panel, and 53% neutralized more than 50%, suggesting the presence of broadly NAbs in these individuals. A preferential intra-subtype neutralization of CRF07_BC was found. Notably, CRF07_BC-infected individuals generated higher neutralization titers against intra-subtype viruses than subtype B’-infected individuals with longer infection length. However, subtype B’-infected individuals mounted broader neutralization responses against inter-subtype viruses than CRF07_BC infection with shorter infection time, indicating the transition from narrow autologous to broad heterologous neutralization over time. Neutralization activity of the top six plasmas from each cohort was attributable to IgG fraction, and half of them developed CD4 binding site antibody reactivity. Heatmap analysis identified three statistically robust clusters of plasmas that offer valuable resources for further in-depth virological and immunological study.

Gag and env conserved element CE DNA vaccines elicit broad cytotoxic T cell responses targeting subdominant epitopes of HIV and SIV Able to recognize virus-infected cells in macaques

ABSTRACT HIV sequence diversity and the propensity of eliciting immunodominant responses targeting inessential variable regions are hurdles in the development of an effective AIDS vaccine. We developed a DNA vaccine comprising conserved elements (CE) of SIV p27Gag and HIV-1 Env and found that priming vaccination with CE DNA is critical to efficiently overcome the dominance imposed by Gag and Env variable regions. Here, we show that DNA vaccinated macaques receiving the CE prime/CE+full-length DNA co-delivery booster vaccine regimens developed broad, potent and durable cytotoxic T cell responses targeting conserved protein segments of SIV Gag and HIV Env. Gag CE-specific T cells showed robust anamnestic responses upon infection with SIVmac239 which led to the identification of CE-specific cytotoxic lymphocytes able to recognize epitopes covering distinct CE on the surface of SIV infected cells in vivo. Though not controlling infection overall, we found an inverse correlation between Gag CE-specific CD8+ T cell responses and peak viremia. The T cell responses induced by the HIV Env CE immunogen were recalled in some animals upon SIV infection, leading to the identification of two cross-reactive epitopes between HIV and SIV Env based in sequence homology. These data demonstrate that a vaccine combining Gag and Env CE DNA subverted the normal immunodominance patterns, eliciting immune responses that included subdominant, highly conserved epitopes. These vaccine regimens augment cytotoxic T cell responses to highly conserved epitopes in the viral proteome and maximize response breadth. The vaccine-induced CE-specific T cells were expanded upon SIV infection, indicating that the predicted CE epitopes incorporated in the DNA vaccine are processed and exposed by infected cells in their natural context within the viral proteome.

G-109 Reverse vaccinology identified promising vaccine candidates focusing responses to relatively conserved regions associated with control of viremia

Background:HIV sequence diversity and potential “decoy” epitopes are hurdles in the development of an effective AIDS vaccine. To target immune responses towards invariable viral regions, we engineered DNA-based immunogens encoding conserved elements (CE) of HIV-1 selected on the basis of stringent conservation, functional importance, HLA-independence, and association with immune control. Methods:Macaques were immunized by IM/EP with DNA plasmids expressing the novel immunogens alone or in prime-boost regimens using DNA expressing the complete p55gag or rMVA. Results:Two molecules were tested expressing regions from (1) p24gag (p24CE) or (2) gag, pol, vif and nef (HTI). These DNA vaccines induced robust central and effector memory responses with a significant fraction of cytotoxic T cells. Repeated vaccination with p55gag DNA was inefficient in inducing primary responses against the CE epitopes but it was able to boost the preexisting responses induced by p24CE, strongly increasing both magnitude and breadth. Although designed as CTL vaccine, these vectors elicited Ab responses, which could be boosted, thereby inducing highest and broadest immunity. Conclusions:The novel HIV-1 T cell vaccines induce broadly balanced responses to vulnerable sites of HIV-1 while avoiding responses to decoy targets that may divert effective T-cell responses towards variable and less protective viral determinants. Combination of CE and full-length immunogen provides a novel strategy to increase the magnitude and breadth of cellular and humoral immunity while targeting efficiently the conserved regions of the virus. This strategy allows for the development and expansion of subdominant responses and greater breadth of immune response.