Role of Occult and Post-acute Phase Replication in Protective Immunity Induced with a Novel Live Attenuated SIV Vaccine

In order to evaluate the role of persisting virus replication during occult phase immunisation in the live attenuated SIV vaccine model, a novel SIVmac239Δnef variant (SIVrtTA) genetically engineered to replicate in the presence of doxycycline was evaluated for its ability to protect against wild-type SIVmac239. Indian rhesus macaques were vaccinated either with SIVrtTA or with SIVmac239Δnef. Doxycycline was withdrawn from 4 of 8 SIVrtTA vaccinates before challenge with wild-type virus. Unvaccinated challenge controls exhibited ~107 peak plasma viral RNA copies/ml persisting beyond the acute phase. Six vaccinates, four SIVmac239Δnef and two SIVrtTA vaccinates exhibited complete protection, defined by lack of wild-type viraemia post-challenge and virus-specific PCR analysis of tissues recovered post-mortem, whereas six SIVrtTA vaccinates were protected from high levels of viraemia. Critically, the complete protection in two SIVrtTA vaccinates was associated with enhanced SIVrtTA replication in the immediate post-acute vaccination period but was independent of doxycycline status at the time of challenge. Mutations were identified in the LTR promoter region and rtTA gene that do not affect doxycycline-control but were associated with enhanced post-acute phase replication in protected vaccinates. High frequencies of total circulating CD8+T effector memory cells and a higher total frequency of SIV-specific CD8+ mono and polyfunctional T cells on the day of wild-type challenge were associated with complete protection but these parameters were not predictive of outcome when assessed 130 days after challenge. Moreover, challenge virus-specific Nef CD8+ polyfunctional T cell responses and antigen were detected in tissues post mortem in completely-protected macaques indicating post-challenge control of infection. Within the parameters of the study design, on-going occult-phase replication may not be absolutely required for protective immunity

Phase I Randomised Clinical Trial of an HIV-1CN54, Clade C, Trimeric Envelope Vaccine Candidate Delivered Vaginally

We conducted a phase 1 double-blind randomised controlled trial (RCT) of a HIV-1 envelope protein (CN54 gp140) candidate vaccine delivered vaginally to assess immunogenicity and safety. It was hypothesised that repeated delivery of gp140 may facilitate antigen uptake and presentation at this mucosal surface. Twenty two healthy female volunteers aged 18–45 years were entered into the trial, the first receiving open-label active product. Subsequently, 16 women were randomised to receive 9 doses of 100 µg of gp140 in 3 ml of a Carbopol 974P based gel, 5 were randomised to placebo solution in the same gel, delivered vaginally via an applicator. Participants delivered the vaccine three times a week over three weeks during one menstrual cycle, and were followed up for two further months. There were no serious adverse events, and the vaccine was well tolerated. No sustained systemic or local IgG, IgA, or T cell responses to the gp140 were detected following vaginal immunisations. Repeated vaginal immunisation with a HIV-1 envelope protein alone formulated in Carbopol gel was safe, but did not induce local or systemic immune responses in healthy women

Plasma vRNA profiles in completely and partially protected macaques and challenge controls.

<p>Complete time-course of total SIV RNA <i>gag</i> levels determined by real-time RT-PCR detailing kinetics of the vaccine and challenge viruses (red arrows signify start of wild-type (WT) SIVmac239 challenge period) showing completely protected, partially protected and unvaccinated naïve control macaques. Administration of challenge virus is indicated by red arrows.</p

Comparison of the frequency of Nef-specific CD8⁺ T cells in mesenteric lymph nodes 130 days after superinfection challenge with respect to protection status.

<p>Cell populations were determined by multi-parametric flow cytometry following stimulation of MNC with SIV-Nef peptide pools. Background responses detected in medium alone control samples were subtracted for every combination of cytokines and a cut-off of >0.01% after background subtraction was used as the threshold for positive reactivity (dashed line). Box plots show the 25^th and 75^th percentiles and median (solid line) and mean (dotted line) for each cytokine combination.</p

Frequency and distribution of SIV-specific CD8⁺ peripheral blood T cells on the day of superinfection challenge with respect to protection status.

<p>Cell populations were determined by multi-parametric flow cytometry following stimulation of PBMC separately with SIV-Gag, Rev and Tat peptide pools. Background responses detected in medium alone control samples were subtracted for every combination of cytokines and a cut-off of >0.01% after background subtraction was used as the threshold for positive reactivity (dashed line). Frequencies were derived by addition of results for Gag, Rev and Tat. Box plots show the 25^th and 75^th percentiles and median (solid line) and mean (dotted line) for each cytokine combination (a & c). Statistically significant differences are highlighted by blue boxes. Proportionate functionality for each macaque (b & d) is shown as a pie chart, with quadruple positivity shown in black and triple to mono positivity shown as shades of grey. Arcs show the combination of cytokine reactivities.</p

Evidence of wild-type infection the spleen of protected and partially protected macaques.

<p>Foci of SIV positive wild-type SIVmac239 infection in all SIVrtTA vaccinates compared with Group C macaques vaccinated with SIVmac239Δ<i>nef</i> (E76 and E77) stained with KK77 monoclonal antibody that detects wild-type Nef only. Distribution of wild-type SIV positive cells in Group D (E80, E81) and a naïve, uninfected macaque (M1464) are shown for comparison. Staining for protected macaques E65 and E70 is more diffuse and sporadic compared to naïve wild-type SIVmac239 challenge control staining. Magnification factor X40.</p

Frequency and distribution of SIV-specific CD8⁺ peripheral blood T cells 130 days after superinfection challenge with respect to protection status.

<p>Cell populations were determined by multiparametric flow cytometry following stimulation of PBMC separately with SIV-Gag, Rev and Tat peptide pools. Background responses detected in medium alone control samples were subtracted for every combination of cytokines and a cut-off of >0.01% after background subtraction was used as the threshold for positive reactivity (dashed line). Frequencies were derived by addition of results for Gag, Rev and Tat. Box plots show the 25^th and 75^th percentiles and median (solid line) and mean (dotted line) for each cytokine combination (a & c). Proportionate functionality for each macaque (b & d) is shown as a pie chart, with quadruple positivity shown in black and triple to mono positivity shown as shades of grey. Arcs show the combination of cytokine reactivities.</p