Chlorhexidine Mouthwash Fails to Eradicate Oropharyngeal Gonorrhea in a Clinical Pilot Trial (MoNg).

ABSTRACT: This single-arm open-label pilot trial in Antwerp, Belgium, was ended early in accordance with the protocol because twice-daily gargling with chlorhexidine 0.2% for 6 days failed to eradicate Neisseria gonorrhoeae from the oropharynx of asymptomatic men who have sex with men (n = 3; efficacy of 0%; 95% confidence interval, 0%-56.1%).

Preexposure Intradermal Rabies Vaccination: A Noninferiority Trial in Healthy Adults on Shortening the Vaccination Schedule From 28 to 7 Days.

Background: The existing 4-week preexposure rabies vaccination schedule is costly and often not practicable. Shorter effective schedules would result in wider acceptance. Methods: We conducted a noninferiority trial in 500 healthy adults comparing the safety and immunogenicity of a 2-visit (days 0 and 7) intradermal (ID) primary vaccination (2 doses of 0.1 mL ID of the human diploid cell culture rabies vaccine [HDCV] at days 0 and 7) vs a standard 3-visit schedule (single dose of 0.1 mL ID at days 0, 7, and 28). One year to 3 years after primary vaccination, a single booster dose of 0.1 mL ID of HDCV was given to evaluate the anamnestic rabies antibody response. The primary endpoint for immunogenicity was the percentage of subjects with an adequate antibody level >0.5 IU/mL 7 days after the booster injection. The safety endpoint was the proportion of participants developing adverse reactions following the primary vaccination and/or booster dose. Results: All subjects in both study groups possessed a rabies antibody titer >0.5 IU/mL on day 7 following the booster dose. Following the booster dose, subjects exposed to the double-dose 2-visit ID schedule had a geometric mean titer of 37 IU/mL, compared with 25 IU/mL for the single-dose 3-visit schedule (P < .001). Local reactions at the injection site following primary vaccination were mild and transient. Conclusions: In healthy adults, ID administration of a double dose of 0.1 mL of HDCV over 2 visits (days 0 and 7) was safe and not inferior to the single-dose 3-visit schedule. Clinical Trials Registration: NCT01388985, EudraCT 2011-001612-62.

Comparative Immunogenicity and Safety Trial of 2 Different Schedules of Single-visit Intradermal Rabies Postexposure Vaccination.

BACKGROUND: Effective and safe single-visit rabies vaccination for pre- and postexposure prophylaxis (PrEP and PEP) could substantially simplify rabies prevention and therefore increase compliance. METHODS: In a comparative trial, 303 healthy adults received a primary vaccination that consisted of 2 intradermal (ID) doses of 0.1 mL of the purified chicken embryo cell vaccine (PCEV) during a single visit. One year later, participants were randomly assigned to receive either 4 or 2 ID PEP booster doses of 0.1 mL PCEV during a single visit. The primary endpoint for immunogenicity was the percentage of participants with an adequate antibody level (>0.5 IU/mL) 7 days after the booster doses. The safety endpoint was the proportion of participants who developed adverse events (AEs) following primary and/or booster vaccination. RESULTS: All participants, except 1 (99.3%) in each study group, had a rabies antibody titer >0.5 IU/mL on day 7 following the booster schedules. Participants exposed to the 4-dose PEP schedule had a geometric mean titer of 20 IU/mL vs 14 IU/mL for the 2-dose PEP schedule (P = .0228). Local reactions at the injection site following PrEP and PEP were mild and transient and only seen in 14.9% and 49.6%-53% of the participants, respectively. No serious AEs were reported. CONCLUSIONS: In healthy adults, a 2-dose (2 × 0.1 mL) single-visit ID PEP schedule was as immunologically adequate and safe as a 4-dose (4 × 0.1 mL) single-visit PEP schedule 7 to 28 months following a 2-dose (2 × 0.1 mL) single-visit ID PREP. CLINICAL TRIALS REGISTRATION: EudraCT 2014-00183612.

Antimalarial artesunate-mefloquine versus praziquantel in African children with schistosomiasis: an open-label, randomized controlled trial.

Schistosomiasis treatment entirely relies on a single drug, praziquantel, prompting research into alternative therapeutics. Here we evaluated the efficacy and safety of the antimalarial combination artesunate-mefloquine for the treatment of schistosomiasis in a proof-of-concept, pragmatic, open-label, randomized controlled trial in primary schools of six villages endemic for schistosomiasis in northern Senegal. Children (6-14 years) were eligible if Schistosoma eggs were detected by microscopy in urine and/or stool. In total, 726 children were randomized 1:1 to praziquantel (standard care: 40 mg kg-1 single dose; n = 364) or to artesunate-mefloquine (antimalarial dosage: artesunate 4 mg kg-1 and mefloquine 8 mg kg-1 daily for three consecutive days; n = 362). Eight children not meeting the inclusion criteria were excluded from efficacy analysis. Median age of the remaining 718 participants was 9 years; 399 (55.6%) were male, and 319 (44.4%) female; 99.3% were infected with Schistosoma haematobium and 15.2% with S. mansoni. Primary outcomes were cure rate, assessed by microscopy, and frequency of drug-related adverse effects of artesunate-mefloquine versus praziquantel at 4 weeks after treatment. Cure rate was 59.6% (208/349) in the artesunate-mefloquine arm versus 62.1% (211/340) in the praziquantel arm. The difference of -2.5% (95% confidence interval (CI) -9.8 to 4.8) met the predefined criteria of noninferiority (margin set at 10%). All drug-related adverse events were mild or moderate, and reported in 28/361 children receiving artesunate-mefloquine (7.8%; 95% CI 5.4 to 11.0) versus 8/363 (2.2%; 95% CI 1.1 to 4.3) receiving praziquantel (P < 0.001). Artesunate-mefloquine at antimalarial dosage was moderately safe and noninferior to standard-care praziquantel for the treatment of schistosomiasis, predominantly due to S. haematobium. Multicentric trials in different populations and epidemiological settings are needed to confirm these findings. ClinicalTrials.gov identifier: NCT03893097 .

Effect of screening for Neisseria gonorrhoeae and Chlamydia trachomatis on incidence of these infections in men who have sex with men and transgender women taking HIV pre-exposure prophylaxis (the Gonoscreen study): results from a randomised, multicentre, controlled trial.

BACKGROUND: Guidelines recommend screening for Neisseria gonorrhoeae and Chlamydia trachomatis at three anatomical sites (urethra, anus, and pharynx) every 3 months (3 × 3) in men who have sex with men (MSM) and transgender women taking HIV pre-exposure prophylaxis (PrEP). We present the first randomised controlled trial to compare the effect of screening versus non-screening for N gonorrhoeae and C trachomatis on the incidence of these infections in MSM and transgender women taking PrEP. METHODS: A multicentre, randomised, controlled trial of 3 × 3 screening for N gonorrhoeae and C trachomatis versus non-screening was done among MSM and transgender women taking PrEP in five HIV reference centers in Belgium. Participants attended the PrEP clinics quarterly for 12 months. N gonorrhoeae and C trachomatis was tested at each visit in both arms, but results were not provided to the non-screening arm, if asymptomatic. The primary outcome was incidence rate of N gonorrhoeae and C trachomatis infections in each arm, assessed in the per-protocol population. Non-inferiority of the non-screening arm was proven if the upper limit of the 95% CI of the incidence rate ratio (IRR) was lower than 1·25. This trial is registered with ClinicalTrials.gov, NCT04269434, and is completed. FINDINGS: Between Sept 21, 2020, and June 4, 2021, 506 participants were randomly assigned to the 3 × 3 screening arm and 508 to the non-screening arm. The overall incidence rate of N gonorrhoeae and C trachomatis was 0·155 cases per 100 person-days (95% CI 0·128-0·186) in the 3 × 3 screening arm and 0·205 (95% CI 0·171-0·246) in the non-screening arm. The incidence rate was significantly higher in the non-screening arm (IRR 1·318, 95% CI 1·068-1·627). Participants in the non-screening arm had a higher incidence of C trachomatis infections and symptomatic C trachomatis infections. There were no significant differences in N gonorrhoeae infections. Participants in the non-screening arm consumed significantly fewer antimicrobial drugs. No serious adverse events were reported. INTERPRETATION: We failed to show that non-screening for N gonorrhoeae and C trachomatis is non-inferior to 3 × 3 screening in MSM and transgender women taking PrEP in Belgium. However, screening was associated with higher antibiotic consumption and had no effect on the incidence of N gonorrhoeae. Further research is needed to assess the benefits and harms of N gonorrhoeae and C trachomatis screening in this population. FUNDING: Belgian Health Care Knowledge Centre.

Five accelerated schedules for the tick-borne encephalitis vaccine FSME-Immun® in last-minute travellers: an open-label, single-Centre, randomized controlled pilot trial.

BACKGROUND: The purpose of this exploratory study was to evaluate different accelerated tick-borne encephalitis (TBE) vaccine schedules for last-minute travellers. METHODS: In a single-centre, open-label pilot study, 77 TBE-naïve Belgian soldiers were randomized to one of the following five schedules with FSME-Immun®: group 1 ('classical accelerated' schedule) received one intramuscular (IM) dose at day 0 and day 14, group 2 two IM doses at day 0, group 3 two intradermal (ID) doses at day 0, group 4 two ID doses at day 0 and day 7, group 5 two ID doses at day 0 and day 14. The last dose(s) of the primary vaccination scheme were given after one year: IM (1 dose) or ID (2 doses). TBE virus neutralizing antibodies were measured in a plaque reduction neutralization test (PRNT90 and 50) at day 0, 14, 21, 28, month 3, 6, 12, and 12 + 21 days. Seropositivity was defined as neutralizing antibody titres ≥10. RESULTS: The median age was 19-19.5 years in each group. Median time-to-seropositivity up to day 28 was shortest for PRNT90 in ID-group 4 and for PRNT50 in all ID groups. Seroconversion until day 28 peaked highest for PRNT90 in ID-group 4 (79%) and for PRNT50 in ID-groups 4 and 5 (both 100%). Seropositivity after the last vaccination after 12 months was high in all groups. Previous yellow fever vaccination was reported in 16% and associated with lower GMTs of TBE-specific antibodies at all time points. The vaccine was generally well tolerated. However, mild to moderate local reactions occurred in 73-100% of ID compared to 0-38% of IM vaccinations, persistent discolouration was observed in nine ID vaccinated individuals. CONCLUSION: The accelerated two-visit ID schedules might offer a better immunological alternative to the recommended classical accelerated IM schedule but an aluminium-free vaccine would preferable.

Evaluation of Artesunate-mefloquine as a Novel Alternative Treatment for Schistosomiasis in African Children (SchistoSAM): protocol of a proof-of-concept, open-label, two-arm, individually-randomised controlled trial.

INTRODUCTION: Alternative drugs and diagnostics are needed for the treatment and control of schistosomiasis. The exclusive use of praziquantel (PZQ) in mass drug administration programmes may result in the emergence of drug resistance. PZQ has little activity against Schistosoma larvae, thus reinfection remains a problem in high-risk communities. Furthermore, the insufficient sensitivity of conventional microscopy hinders therapeutic response assessment. Evaluation of artesunate-mefloquine (AM) as a Novel Alternative Treatment for Schistosomiasis in African Children (SchistoSAM) aims to evaluate the safety and efficacy of the antimalarial combination artesunate-mefloquine, re-purposed for the treatment of schistosomiasis, and to assess the performance of highly sensitive novel antigen-based and DNA-based assays as tools for monitoring treatment response. METHODS AND ANALYSIS: The SchistoSAM study is an open-label, two-arm, individually randomised controlled non-inferiority trial, with a follow-up of 48 weeks. Primary school-aged children from the Richard Toll district in northern Senegal, an area endemic for Schistosoma mansoni and Schistosoma haematobium, are allocated to the AM intervention arm (3-day courses at 6-week intervals) or the PZQ control arm (single dose of 40 mg/kg). The trial's primary endpoints are the efficacy (cure rate (CR), assessed by microscopy) and safety (frequency and pattern of drug-related adverse events) of one AM course versus PZQ at 4 weeks after treatment. Secondary endpoints include (1) cumulative CR, egg reduction rate and safety after each additional course of AM, and at weeks 24 and 48, (2) prevalence and severity of schistosomiasis-related morbidity and (3) malaria prevalence, incidence and morbidity, both after 24 and 48 weeks. CRs and intensity reduction rates are also assessed by antigen-based and DNA-based diagnostic assays, for which performance for treatment monitoring is evaluated. ETHICS AND DISSEMINATION: Ethics approval was obtained both in Belgium and Senegal. Oral assent from the children and signed informed consent from their legal representatives was obtained, prior to enrolment. The results will be disseminated in peer-reviewed journals and at international conferences. TRIAL REGISTRATION NUMBER: NCT03893097; pre-results.

Time of administration of rabies immunoglobulins and adequacy of antibody response upon post-exposure prophylaxis: a descriptive retrospective study in Belgium.

Background: Data on rabies post-exposure prophylaxis (PEP) and the use of human rabies immunoglobulins (HRIG) in Belgium are scarce. The main objective of this study was to evaluate the timely administration of HRIG after rabies exposure. The secondary objective was to evaluate the adequate antibody response following PEP.Methods: We reviewed all medical records from July 2017 to June 2018 of patients seeking care at, or referred to, the Institute of Tropical Medicine and the University Hospital, Antwerp for the administration of human rabies immunoglobulins following potential rabies exposure abroad or in Belgium.A timely response was defined as starting HRIG with a delay of ≤48 h and rabies vaccination in the first 7 days after exposure.Adequate antibody response was defined as a titer of >5.0 IU/mL in case of bat-related exposure and >3.0 IU/mL in case of exposure to other animals. Titers were measured 10 days after the last PEP vaccine dose, using the rapid fluorescent focus inhibition test (RFFIT).Results: Of the 92 cases treated with HRIG, 75 were evaluated.The majority of injuries were acquired in Asia (n = 26,34%) and in Western Europe (n = 18, 24%), of which 17 in Belgium. The five most frequently recorded countries overseas were Indonesia (n = 13), Thailand (n = 7), Morocco (n = 4), Peru (n = 3) and Costa Rica (n = 3). Administration of immunoglobulins was related to injuries by dogs (36%), monkeys (25%) or bats (22%).A timely response was observed in 16 (21,33%) and in 55 (73,33%) of subjects receiving HRIG (≤48 h) or rabies vaccine (<7days) respectively. The mean time between exposure and the first administered dose of rabies vaccine and HRIG was 7.7 and 8.7 days, respectively. The mean delay for HRIG administration was 9.6 days and 6 days for abroad and inland risks, respectively.In 15 of 16 (94%) bat-related cases the antibody titer after full PEP was >5.0 IU/ml. In 38 of 47 (81%) cases related to other animals the RFFIT titer was >3.0 IU/ml. All low-responders received additional rabies injections.Conclusion: This study showed a substantial time delay between the animal-related risk and the administration of HRIG, in particular when the injury occurred abroad. More targeted communication about the risks of rabies and preventable measures may reduce this delay.Furthermore, the antibody response was inadequate in some cases following full PEP administration according to the Belgian recommendation.

Antibacterial mouthwash to prevent sexually transmitted infections in men who have sex with men taking HIV pre-exposure prophylaxis (PReGo): a randomised, placebo-controlled, crossover trial.

BACKGROUND: Bacterial sexually transmitted infections (STIs) are highly prevalent among men who have sex with men who use HIV pre-exposure prophylaxis (PrEP), which leads to antimicrobial consumption linked to the emergence of antimicrobial resistance. We aimed to assess use of an antiseptic mouthwash as an antibiotic sparing approach to prevent STIs. METHODS: We invited people using PrEP who had an STI in the past 24 months to participate in this single-centre, randomised, double-blind, placebo-controlled, AB/BA crossover superiority trial at the Institute of Tropical Medicine in Antwerp, Belgium. Using block randomisation (block size eight), participants were assigned (1:1) to first receive Listerine Cool Mint or a placebo mouthwash. They were required to use the study mouthwashes daily and before and after sex for 3 months each and to ask their sexual partners to use the mouthwash before and after sex. Participants were screened every 3 months for syphilis, chlamydia, and gonorrhoea at the oropharynx, anorectum, and urethra. The primary outcome was combined incidence of these STIs during each 3-month period, assessed in the intention-to-treat population, which included all participants who completed at least the first 3-month period. Safety was assessed as a secondary outcome. This trial is registered with Clinicaltrials.gov, NCT03881007. FINDINGS: Between April 2, 2019, and March 13, 2020, 343 participants were enrolled: 172 in the Listerine followed by placebo (Listerine-placebo) group and 171 in the placebo followed by Listerine (placebo-Listerine) group. The trial was terminated prematurely because of the COVID-19 pandemic. 151 participants completed the entire study, and 89 completed only the first 3-month period. 31 participants withdrew consent, ten were lost to follow-up, and one acquired HIV. In the Listerine-placebo group, the STI incidence rate was 140·4 per 100 person-years during the Listerine period, and 102·6 per 100 person-years during the placebo period. In the placebo-Listerine arm, the STI incidence rate was 133·9 per 100 person-years during the placebo period, and 147·5 per 100 person-years during the Listerine period. We did not find that Listerine significantly reduced STI incidence (IRR 1·17, 95% CI 0·84-1·64). Numbers of adverse events were not significantly higher than at baseline and were similar while using Listerine and placebo. Four serious adverse events (one HIV-infection, one severe depression, one Ludwig's angina, and one testicular carcinoma) were not considered to be related to use of mouthwash. INTERPRETATION: Our findings do not support the use of Listerine Cool Mint as a way to prevent STI acquisition among high-risk populations. FUNDING: Belgian Research Foundation - Flanders (FWO 121·00).

In vitro synergistic activity against CCR5-tropic HIV-1 with combinations of potential candidate microbicide molecules HHA, KRV2110 and enfuvirtide (T20).

OBJECTIVES: To block the different mechanisms of HIV mucosal transmission, it is likely that use of several microbicide molecules will lead to the best protection against HIV transmission. Indeed, the combination of microbicides with complementary mechanisms of action is expected to increase the antiviral potency of the formulation. METHODS: The gp120-interacting plant lectin HHA ('Hippeastrum hybrid agglutinin'), the non-nucleoside reverse transcriptase inhibitor KRV2110 and the fusion inhibitor enfuvirtide (T20) were combined in 12 drug associations by using the Ray combination design method. Their activity against HIV-1(BaL) was assessed by the lymphocyte infectivity reduction assay and by the single-cycle BaL pseudovirus (PV) assay. In addition, their cell tolerance was evaluated for HEC-1 and HeLa epithelial cell lines, both originating from genital tissue. RESULTS: All evaluated combinations showed synergistic activity in both lymphocyte infectivity reduction and single-cycle BaL PV assays. The combination HHA + KRV2110 resulted in the highest cell viability, whereas the combinations including T20 exhibited a dose-dependent decrease in cell viability, demonstrating the differential tolerance of epithelial cell lines to the combinations. CONCLUSIONS: These observations provide a rational basis for in vitro testing of microbicide candidate molecule combinations, including anti-HIV-1 and cytotoxic cellular assays.

Human immunodeficiency virus type 1 (HIV-1) integration: a potential target for microbicides to prevent cell-free or cell-associated HIV-1 infection.

Conceptually, blocking human immunodeficiency virus type 1 (HIV-1) integration is the last possibility for preventing irreversible cellular infection. Using cocultures of monocyte-derived dendritic cells and CD4(+) T cells, which represent primary targets in sexual transmission, we demonstrated that blocking integration with integrase strand transfer inhibitors (InSTIs), particularly L-870812, could consistently block cell-free and cell-associated HIV-1 infection. In a pretreatment setting in which the compound was present before and during infection and was afterwards gradually diluted during the culture period, the naphthyridine carboxamide L-870812 blocked infection with the cell-free and cell-associated HIV-1 Ba-L strain at concentrations of, respectively, 1,000 and 10,000 nM. The potency of L-870812 was similar to that of the nucleotide reverse transcriptase inhibitor R-9-(2-phosphonylmethoxypropyl) adenine (PMPA) but one or two orders of magnitude lower than those of the nonnucleoside reverse transcriptase inhibitors UC781 and TMC120. In contrast, the diketo acid RDS derivative InSTIs showed clear-cut but weaker antiviral activity than L-870812. Moreover, L-870812 completely blocked subtype C and CRFO2_AG primary isolates, which are prevalent in the African heterosexual epidemic. Furthermore, the addition of micromolar concentrations of L-870812 even 24 h after infection could still block both cell-free and cell-associated Ba-L, opening the prospect of postexposure prophylaxis. Finally, an evaluation of the combined activity of L-870812 with either T20, zidovudine, PMPA, UC781, or TMC120 against replication-deficient HIV-1 Ba-L (env) pseudovirus suggested synergistic activity for all combinations. Importantly, compounds selected for the study by using the coculture model were devoid of acute or delayed cytotoxic effects at HIV-blocking concentrations. Therefore, these findings provide evidence supporting consideration of HIV-1 integration as a target for microbicide development.

CD4 mimetic miniproteins: potent anti-HIV compounds with promising activity as microbicides.

OBJECTIVES: The antiviral activity of CD4 miniproteins was evaluated as potential HIV microbicides, using relevant in vitro models. METHODS: Compounds were tested in a single-cycle HIV-1 pseudovirus assay and against replication competent HIV-1 in co-cultures of monocyte-derived dendritic cells (MO-DC) and CD4+ T cells. Cytotoxic activity was evaluated in an MTT assay. RESULTS: Monomeric miniproteins (M47 and M48) showed 50% effective concentration (EC(50)) values of 79-105 nM against a subtype B, CCR5 co-receptor-using Ba-L pseudovirus. Higher activity was found for the dimeric miniproteins M48D30, M48D50 and M48D100 (EC(50) between 15 and 30 nM), in contrast to the tetrameric miniproteins M48T30, M48T50 and M48T100 (EC(50) between 107 and 377 nM). The hetero-bivalent miniprotein M48-Hep and miniproteins that targeted the Phe-43 cavity on gp120 (M48-U1, M48-U2 and M48-U3) were highly active, with EC(50) values as low as 2 nM for M48-U1. All miniproteins showed high activity against CCR5 or CXCR4 co-receptor-using subtype B and CRF-01_A/E pseudoviruses. Many early M48-based compounds were much less active against subtype C pseudoviruses, whereas M48-U compounds that targeted the Phe-43 cavity were very active against all pseudoviruses, including subtype C. In MO-DC/CD4+ T cell co-cultures with replication-competent HIV-1 Ba-L, EC(50) values ranged between 13 and 1719 nM depending on the miniprotein, with M48-U1, M48-U2 and M48-U3 again being the most potent. Importantly, the latter compounds completely prevented viral replication by treating the cultures from 2 h before until 24 h after infection, at non-toxic concentrations of 66-6564 nM. CONCLUSIONS: These novel CD4 miniproteins might constitute a promising class of HIV microbicides.

Antiviral compounds show enhanced activity in HIV-1 single cycle pseudovirus assays as compared to classical PBMC assays.

HIV-1 Env pseudotyped viruses (PV) are an attractive tool for studying the antiviral activities of compounds interfering with virus entry into a target cell. To investigate whether results obtained in PV assays are relevant biologically, the antiviral activity of 6 reference compounds was compared on 5 virus isolates of different clades using three assays: (1) replicating virus in peripheral blood mononuclear cells (PBMCs), (2) PV in CD4 and CCR5- or CXCR4 co-receptor expressing Ghost cells, and (3) PV in PBMCs. A significant linear relationship was found between both single-cycle PV assays (P<0.0001, R2=0.75). Moreover, both assays showed enhanced sensitivity to the antiretrovirals tested (P=0.013 and 0.015, respectively) as compared to the PBMC assay with replication-competent virus. Most importantly, results from the latter assay could be predicted significantly from both PV assays, in which either Ghost target cells (P<0.0001, R2=0.61) or PBMCs (P<0.0001, R2=0.55) were used. The usefulness of the PV assay was demonstrated further by investigating the impact of the HIV-1 Env subtype on the antiviral activity of five new compounds derived from the entry inhibitor BMS806.

A dual chamber model of female cervical mucosa for the study of HIV transmission and for the evaluation of candidate HIV microbicides.

A dual chamber system was established to model heterosexual HIV transmission. Cell-associated, but not cell-free HIV, added to a confluent layer of cervical epithelial cells in the apical chamber, reproducibly infected monocyte-derived dendritic cells (MO-DC) and CD4(+) T cells in the basal compartment. Only minimal epithelial transmigration of HIV-infected mononuclear cells (HIV-PBMCs) was observed. Most evidence points to transepithelial migration of virus, released from HIV-PBMCs after their activation by epithelial cells. We used this model for evaluation of the therapeutic index of various potentially preventive antiviral compounds, including non-nucleoside reverse transcriptase inhibitors (NNRTIs, including UC781 and various diaryltriazines and diarylpyrimidines), poly-anionic entry inhibitors (including PRO2000, cellulose sulphate, dextrane sulphate 5000 and polystyrene sulphonate) and the fusion inhibitor T-20. The epithelium was pre-treated with compound and incubated with HIV-PBMCs for 24 h. Afterwards the apical chamber was removed and MO-DC/CD4(+) T cell co-cultures were further cultured without compound. NNRTIs, including a TMC120 gel, blocked infection of the sub-epithelial targets at sub-micromolar concentrations. Polyanionic entry inhibitors (up to 100 microg/ml) and T-20 (up to 449 microg/ml) failed to inhibit transmission. Moreover, whereas the NNRTIs used interfered with epithelial integrity with cervical epithelium only at very high concentrations, the evaluated entry inhibitors showed toxicity at concentrations that did not prevent infection.

In vitro pre- and post-exposure prophylaxis using HIV inhibitors as microbicides against cell-free or cell-associated HIV-1 infection.

Several classes of microbicides are being evaluated for the prevention of sexual HIV transmission. In vivo, the infectious dose and viral source involved in transmission remain uncertain and it is likely that women will use microbicides both before and after high-risk HIV exposure. Therefore, we evaluated HIV entry inhibitors (EIs) and reverse transcriptase inhibitors (RTIs) for their ability to block cell-free and cell-associated HIV-1 infection in co-cultures of monocyte-derived dendritic cells (MO-DC) and CD4+ T-cells using settings of pre- and post-exposure prophylaxis. In the pre-exposure assay, where compound was present before, during and 24 h after infection, all tested EIs (BMS806, TAK779 and T20) and RTIs (PMPA, TMC120 and UC781) blocked infection with 10(-4) multiplicity of infection (MOI) of cell-free virus at a dose between 100 and 10,000 nM, dependent on the compound used. At 10(-3) MOI, however, only T20 and the RTIs completely blocked infection. Furthermore, in experiments with cell-associated virus, EIs were ineffective, whereas RTIs actively blocked infection with similar potency as in the experiments with cell-free virus. In the post-exposure assay, where compound was added 2 h after infection and remained present for 24 h, EIs were inactive whereas RTIs blocked cell-free and cell-associated viral infections equally efficiently. Moreover, post-exposure prophylaxis initiated 24 h after infection with cell-free or cell-associated HIV-1 was still effective with 1,000 nM of TMC120. Both EIs and RTIs were non-cytotoxic at any tested concentration for MO-DC and CD4+ T-cells in co-culture. Our study shows that RTIs are potent inhibitors of cell-free and cell-associated virus used either in pre- or post-exposure settings. It highlights that parameters such as viral input, viral source, the time of compound addition and the target cells should be considered in microbicides evaluation.

Mid-trimester maternal ADAM12 levels differ according to fetal gender in pregnancies complicated by preeclampsia.

An overrepresentation of adverse pregnancy outcomes has been observed in pregnancies associated with a male fetus. We investigated the association between fetal gender and candidate biomarkers for preeclampsia. Proteins were quantified in samples taken at 20 weeks from women recruited to the SCreening fOr Pregnancy Endpoints (SCOPE) study (preeclampsia n = 150; no preeclampsia n = 450). In contrast to placental growth factor, soluble endoglin, and insulin-like growth factor acid labile subunit, levels of metallopeptidase domain 12 (ADAM12) at 20 weeks were dependent on fetal gender in pregnancies complicated by preeclampsia, for male (n = 73) fetuses the multiples of the median (MoM; interquartile range [IQR] 1.1-1.5) was 1.3, whereas for female fetuses (n = 75) MoM was 1.1 (1.0-1.3); P < .01. Prediction of preeclampsia using ADAM12 levels was improved for pregnancies associated with a male fetus (area under receiver-operator curve [AUC] 0.73 [95% confidence interval [CI] 0.67-0.80]) than that of a female fetus (AUC 0.62 [0.55-0.70]); P = .03. The data presented here fit a contemporary hypothesis that there is a difference between the genders in response to an adverse maternal environment and suggest that an alteration in ADAM12 may reflect an altered placental response in pregnancies subsequently complicated by preeclampsia.

Metabolic activation of nucleoside and nucleotide reverse transcriptase inhibitors in dendritic and Langerhans cells.

BACKGROUND: Langerhans cells and interstitial dendritic cells are the earliest targets for HIV infection through sexual transmission of HIV. Metabolism of nucleoside analogues markedly differs in proliferating T lymphocytes and resting monocyte/macrophages, and thus their antiviral efficacy can substantially differ between both cell types. METHODS: The metabolism of radio-labelled zidovudine (ZDV), lamivudine (3TC) and tenofovir (PMPA) to their antivirally active metabolites was studied in primary cells, representative of early in vivo targets of HIV [i.e. monocyte-derived dendritic cells (MO-DC), MO-derived Langerhans cells (MO-LC), PHA/IL-2-activated T-blast cells] as well as in a laboratory T-lymphocyte (CEM) cell line. RESULTS: Whereas lamivudine metabolism to its active triphosphate derivative (3TC-TP) did not markedly differ between T-cells and MO-derived LC and DC, zidovudine was much better converted to ZDV-TP in T-cells than in MO-LC and MO-DC. In contrast, tenofovir was markedly more abundantly converted to its antivirally active diphosphate metabolite PMPApp in MO-DC and MO-LC than zidovudine and lamivudine. CONCLUSION: Our metabolic data suggest that tenofovir may be superior to zidovudine and lamivudine for inhibition of HIV replication in dendritic/Langerhans cells, the first-line cell types targeted by a primary HIV infection.

Activity of reverse transcriptase inhibitors in monocyte-derived dendritic cells: a possible in vitro model for postexposure prophylaxis of sexual HIV transmission.

Because prevention of heterosexual HIV transmission is not always possible, it is important to develop effective strategies of postexposure prophylaxis (PEP). Since in vivo comparison of drug potency is difficult, we developed an in vitro model with cells resembling primary targets during sexual transmission: monocyte-derived dendritic cells (MO-DCs), Langerhans cells (MO-LCs), and resting autologous CD4(+) T cells. Nucleoside and nonnucleoside reverse transcriptase inhibitors (NRTIs and NNRTIs, respectively) were evaluated for their antiviral activity, when added immediately after infection or at a later time point. In parallel, their immune-suppressive effect was examined by measuring inhibition of mixed MO-DC/allogeneic CD4(+) T cell cultures. Most RTIs potently inhibited HIV replication, even if added 24 hr after infection (representing PEP). The sensitivity to antiretroviral drugs was similar in HIV-infected MO-DCs and MO-LCs, but decreased in cocultures with resting autologous CD4(+) T cells. The NNRTIs efavirenz and UC-781 as well as the NRTIs AZT, 3TC, and d4T showed a similar high potency in MO-DC plus autologous CD4(+) T cell cocultures as compared with CEM T cells, whereas their activity in phytohemagglutinin/interleukin 2 (PHA/IL-2)-activated CD4(+) T cells was lower. The dideoxynucleoside RTI abacavir as well as the phosphonates (R)-PMPA and PMEA were more active in infected MO-DCs as compared with either CEM T cells or PHA/IL-2 activated CD4(+) T cells. Infection in cocultures of MO-DCs and autologous CD4(+) T cells could be aborted in a proportion of the cultures, with high concentrations of PMEA and/or efavirenz, but not with AZT. Suppressive activity in mixed leukocyte cultures was observed only at very high concentrations of RTI. Our data suggest that cocultures of MO-DCs and autologous CD4(+) T cells can be used as a possible in vitro model to explore protocols for PEP after sexual HIV transmission.