CONSORT flowchart.

<p>CONSORT flowchart.</p

Study flow diagram.

<p>Paired measures from compartment concentrations (CC) and both CC and biopsy samples (*) taken at the bolded visits: V3 (Visit 3: ∼30 mins post single oral dose), V5 (1–6 days post V3 dose), V6 (7–9 days post V3 dose), V7 (∼30 mins post single topical dose), V9 (1–3 days post V7 dose), V10 (7–12 days post V7 does) and V12 (∼30 mins post 7^th daily dose) used in the dose-response analysis.</p

Rectal Administration of Tenofovir Gel Protected a High Proportion of Macaques against Subsequent Acquisition of SIV by Rectal Transmission

<p>The results of VI from PBMC are shown as + or − for each animal. The temporal profiles of plasma vRNA concentration (red dot) and frequency of PMBC-associated proviral DNA (blue triangle) are shown for each animal in the study.</p

SIV-Specific IFN-γ Secreting T Cells Were Detected in SIV-Challenged Macaques in the Absence of Serum Antibody Responses and Evidence of Overt Infection

<div><p>(A) IFN-γ secreting T cell frequencies in PBMC from protected animals (D68–D14) compared to those in an SIV-infected animal (E81) measured 20 wk after virus exposure measured by ex vivo ELISpot. The mean frequencies of three replicate determinations plus one standard deviation are shown for each peptide pool used.</p> <p>(B) SIV-specific IFN-γ secreting T cell frequencies in MNC isolated from ileum–jejunum tissue of four protected animals measured post mortem at 21 wk after virus challenge by ex vivo ELISpot.</p> <p>(C) The group mean +/− standard deviation profile of anti-SIV Gag p27 binding antibody titres (measured by ELISA) from animals infected with SIV (○) and the individual profile for an SIV-infected macaque E81 (○), in which T cell ELISpot was analysed was compared with animals from which no virus was detected following challenge (▴). </p></div

Colorectal Explants from Macaques Supported Replication of SIV That Was Inhibited by Pretreatment with Tenofovir In Vitro and In Vivo

<div><p>(A) Replication dynamics of SIVmac_251/32H in explants from two untreated animals (group F: M3, M6) in the presence or absence of exogenously added tenofovir at 100 μg/ml. A total of 10⁴ TCID₅₀ of virus was added to each well containing three explants in a total volume of 200 μl of medium. Virus replication was assayed by SIV Gag p27 production and mean values +/− standard deviations are shown for four replicates of each tissue.</p> <p>(B) Colorectal explants from four animals (group G: M1, M38, M5, M32) dosed in vivo with tenofovir per rectum 3 h before tissue removal were exposed to virus in vitro (as described above) and culture supernatants assayed for Gag p27. Mean percent inhibition of SIV Gag p27 production plus standard deviations are shown.</p></div

Compartment TFV and TFVdp efficacy concentrations (EC_50,90,95) predicted by logistic regression to suppress HIV infection following single oral TDF, single topical TFV 1% gel and 7-day topical TFV 1% gel <i>in vivo</i> product use.

§<p>Predicted compartment dose concentrations to suppress 50, 90 and 95% of HIV infection interpolated from the logistic regression probability curve where infection was defined as cumulative p24 pg/mL ≥500.</p><p>Actual drug concentrations in the delivered product were 300 mg tenofovir disoproxil fumarate in the oral pill (equivalent to 245 mg of tenofovir disoproxil) and 40 mg/4 mL tenofovir in each topical gel application.</p><p>Compartment TFV and TFVdp efficacy concentrations (EC_50,90,95) predicted by logistic regression to suppress HIV infection following single oral TDF, single topical TFV 1% gel and 7-day topical TFV 1% gel <i>in vivo</i> product use.</p

Solution and Gel Formulated Tenofovir Inhibited SIV_mac251/32H Infectivity In Vitro at Similar Doses and in the Same Range as for Representative HIV-1 Isolates

<p>Infection of TZM-bl indicator cells in the presence and absence of tenofovir was compared by luminescence analysis of cell lysates and the results expressed as percent inhibition. The graph shows the full titration of drug formulations on infection with SIV_mac251/32H; the virus stock used in subsequent challenge experiments in vivo. Each point represents the mean of three independent experiments performed in triplicate +/− standard deviation. The results for a panel of HIV-1 strains in comparison to SIV_mac251/32H are shown as IC₅₀ values in the inset.</p

Graphic summary of the ability of each compound to activate HIV within each cell model: (A) primary CD4 T cell models and patient cell outgrowth assay (QVOA), and (B) J-Lat T cell line clones.

<p>Each compound and concentration tested is listed on the X-axis. In the primary CD4 cell models, each compound was tested using cells from 2, 3 or 4 different donors and in duplicate or triplicate with cells from each donor (See <a href="http://www.plospathogens.org/article/info:doi/10.1371/journal.ppat.1003834#s4" target="_blank">Methods</a> Section for details). For the QVOA, results from the limiting dilution cultures from 3 patients were pooled to calculate one common IUPM (infectious units per million cells) value which was then normalized to that obtained with PHA. With the J-Lat clones, experiments were performed in triplicate. Asterisks represent “not done”.</p

List of stimuli used in this study and their corresponding signaling pathways.

<p>List of stimuli used in this study and their corresponding signaling pathways.</p

Heatmap visualization of the ability of each compound to activate HIV within each model when excluding (A) and including (B) data from the QVOA model.

<p>A reduced set of compounds was analyzed in (<b>B</b>) since not every compound was run at every concentration in the QVOA. The clustergram at the left of each heatmap reflects the relationships between compounds based on their ability to activate HIV across compounds. Since cells in all models responded to PHA with high strength, ranking was normalized within each model to the response to PHA at 10 µg/mL and, therefore, all models display in the heatmap the same relative responsiveness to this treatment. The clustergram at the top of each heatmap reflects the relationship between each model based on their response to compounds. Clustergrams were created by calculating Euclidean distances and then clustering distances using the average linkage method. The numbers at the nodes of clusters are AU p-values where 95% represents a <i>p</i>-value cut-off of 0.05 and only values 95% or greater are depicted. Red cells in the heatmaps reflect HIV activation whereas blue or blank cells indicate that the compound did not effectively activate HIV.</p