Two-year carcinogenicity study of a novel plasticizer, bis(2-ethylhexyl) cyclohexane-1,4-dicarboxylate (Eco-DEHCH), by oral diet in Han Wistar rats.

Plasticizers are necessary for the usability of various products, including food contact materials. Exposure to plasticizers is most commonly made through the oral route. Several plasticizers have been reported to have adverse effects on humans and the environment. Thus, the present study aimed to determine the long-term toxicity and carcinogenicity of a novel plasticizer called bis(2-ethylhexyl) cyclohexane-1,4-dicarboxylate (Eco-DEHCH), which is an ecofriendly and biologically less harmful replacer. Groups of 50 male and 50 female Han Wistar rats were fed Eco-DEHCH at daily doses of 1,600, 5,000, or 16,000 ppm in their diet for at least 104 weeks. The rats were regularly monitored for mortality, clinical signs, body weight, food consumption, food efficiency, and perceivable mass. All animals were subjected to complete necropsy and histopathological examination. The results indicate that the rats well tolerated chronic exposure to Eco-DEHCH at highest daily doses of 16,000 ppm, with was equivalent to 805.1 mg/kg/day in males and 1060.6 mg/kg/day in females and did not show signs of toxicity or carcinogenicity. In conclusion, Eco-DEHCH could be a safe and promising alternative plasticizer.

Cyclohexane Inhalation Produces Long-Lasting Alterations in the Hippocampal Integrity and Reward-Seeking Behavior in the Adult Mouse.

Cyclohexane (CHX) is an organic solvent commonly used as a drug-of-abuse. This drug increases the oxidative stress and glial reactivity in the hippocampus, which suggests that this brain region is vulnerable to CHX effects. This study aimed to establish the behavioral changes and the pathological alterations that occur in the Cornu Ammonis 3 (CA3) and Dentate Gyrus (DG) after a long-lasting exposure to CHX. We exposed CD1 mice to a recreational-like dose of CHX (~ 30,000 ppm) for 30 days and explored its consequences in motor skills, reward-seeking behavior, and the CA3 and DG hippocampal subfields. Twenty-four hours after the last administration of CHX, we found a significant decrease in the number of c-Fos+ cells in the hippocampal CA3 and DG regions. This event coincided with an increased in NMDAR1 expression and apoptotic cells in the CA3 region. At day 13th without CHX, we found a persistent reduction in the number of c-Fos+ and TUNEL+ cells in DG. At both time points, the CHX-exposed mice showed a strong overexpression of neuropeptide Y (NPY) in the CA3 stratum lucidum and the hippocampal hilus. In parallel, we used an operant-based task to assess motor performance and operant conditioning learning. The behavioral analysis indicated that CHX did not modify the acquisition of operant conditioning tasks, but affected some motor skills and increased the reward-seeking behavior. Altogether, this evidence reveals that CHX exposure provokes long-lasting changes in the hippocampal subfields, induces motor impairments and increases the motivation-guided behavior. These findings can help understand the deleterious effect of CHX into the adult hippocampus and unveil its potential to trigger addiction-like behaviors.

Efficacy of fumagillin bicyclohexylamine on experimental corneal neovascularization in rat model.

PURPOSE: Fumagillin has been previously used to treat corneal microsporidial keratitis and also identified as an angiogenesis inhibitor. This study aimed to evaluate efficacy of fumagillin bicyclohexylamine on the rat model of corneal neovascularization induced by silver nitrate cauterization. METHODS: Twenty-four Albino Wistar rats (n = 24) were divided into three groups. Following silver nitrate-induced corneal injury, eyes in Group 1 received one drop of 5 mg/mL topical fumagillin bicyclohexylamine four times daily for 10 days. Group 2 received subconjunctival injection of 0.1 mL fumagillin bicyclohexylamine (2.5 mg/mL) on day 1 and day 5. Group 3 received artificial tears and lubricants four times daily for 10 days as control. On day 10, animals were sacrificed. Corneal specimens were obtained and prepared to assess vascular endothelial growth factor (VEGF-C) levels and corneal angiogenic microvessel density. RESULTS: There was no significant difference in VEGF-C levels between the groups (P = 0.994). Assessment of angiogenic microvessel density for peripheral corneal zone also did not reveal significant difference between the groups (P = 0.113). However, mean vascular density in Group 1 and Group 2 was significantly higher for both midperipheral and central corneal zones in comparison with Group 3 (P = 0.003, P = 0.015). CONCLUSIONS: Previously proved to be effective for treatment of microsporidial keratitis in humans, topical and subconjunctival concentration or dosing of fumagillin bicyclohexylamine failed to reduce corneal neovascularization induced by silver nitrate in this study. Further studies comparing different concentrations and dosing may detect inhibitory effects of fumagillin on corneal neovascularization without inducing toxicity.

Evaluation of development, locomotor behavior, oxidative stress, immune responses and apoptosis in developing zebrafish (Danio rerio) exposed to TBECH (tetrabromoethylcyclohexane).

Tetrabromoethylcyclohexane (TBECH), as one emerging brominated flame retardants, is ubiquitous in the environment, including water and aquatic organisms. TBECH was found to exhibit endocrine-disrupting effects in different models, whereas a survey of comprehensive toxic effects of TBECH on zebrafish is limited. In the present study, zebrafish (Danio rerio) were waterborne exposed continuously to TBECH from embryonic stage (3 h post-fertilization (hpf)) to the time when the respective parameters were evaluated. Exposure to TBECH reduced hatchability of zebrafish embryos at 72 and 96 hpf, diminished heart rate of zebrafish larvae at 48 hpf, and increased malformation in zebrafish larvae at 96 hpf. In addition, exposure to TBECH diminished free swimming distance both in the light and under a photoperiod of 10 min light/10 min dark cycles in zebrafish larvae at 6 days post-fertilization (dpf). Moreover, exposure to TBECH elevated activities of superoxide dismutase (SOD) and catalase (CAT), malondialdehyde (MDA) content, whereas it reduced glutathione (GSH) content, in zebrafish larvae at 6 dpf. Accordingly, RT-qPCR analysis demonstrated that TBECH exposure increased the mRNA levels of sod1, sod2, cat, and gpx1 in zebrafish larvae at 6 dpf. With respect to the immune aspect, the mRNA levels of pro-inflammatory genes, including il-1b, il-6, il-8, and tnfa, in larval zebrafish at 6 dpf were increased by exposure to TBECH, while pretreatment with TBECH inhibited 24 h of exposure to LPS-stimulated elevation in the mRNA levels of the abovementioned four pro-inflammatory genes in zebrafish larvae at 6 dpf. Furthermore, TBECH treatment increased caspase-3 enzyme activities and regulated apoptosis-related genes in larval zebrafish at 6 dpf. Taken together, the data obtained in this study demonstrated that TBECH caused developmental and locomotor behavioral toxicity, immunotoxicity, oxidative stress and proapoptotic effects in early life zebrafish. The present study will help to understand the comprehensive toxicity of TBECH in zebrafish.

Anti-angiogenic drug loaded liposomes: Nanotherapy for early atherosclerotic lesions in mice.

Fumagillin-loaded liposomes were injected into ApoE-KO mice. The animals were divided into several groups to test the efficacy of this anti-angiogenic drug for early treatment of atherosclerotic lesions. Statistical analysis of the lesions revealed a decrease in the lesion size after 5 weeks of treatment.

Enterocytozoon bieneusi Microsporidiosis in Stem Cell Transplant Recipients Treated with Fumagillin

Enterocytozoon bieneusi microsporidiosis is an emerging disease in immunocompromised patients. We report 2 cases of this disease in allogeneic hematopoietic stem cell transplant patients successfully treated with fumagillin. Thrombocytopenia occurred but without major adverse events. Modifications of immunosuppression could be avoided when E. bieneusi is rapidly identified and fumagillin therapy is started promptly.

Effects of MetAP2 inhibition on hyperphagia and body weight in Prader-Willi syndrome: A randomized, double-blind, placebo-controlled trial.

AIMS: There are no treatments for the extreme hyperphagia and obesity in Prader-Willi syndrome (PWS). The bestPWS clinical trial assessed the efficacy, safety and tolerability of the methionine aminopeptidase 2 (MetAP2) inhibitor, beloranib. MATERIALS AND METHODS: Participants with PWS (12-65 years old) were randomly assigned (1:1:1) to biweekly placebo, 1.8 mg beloranib or 2.4 mg beloranib injection for 26 weeks at 15 US sites. Co-primary endpoints were the changes in hyperphagia [measured by Hyperphagia Questionnaire for Clinical Trials (HQ-CT); possible score 0-36] and weight by intention-to-treat. ClinicalTrials.gov registration: NCT02179151. RESULTS: One-hundred and seven participants were included in the intention-to-treat analysis: placebo (n = 34); 1.8 mg beloranib (n = 36); or 2.4 mg beloranib (n = 37). Improvement (reduction) in HQ-CT total score was greater in the 1.8 mg (mean difference -6.3, 95% CI -9.6 to -3.0; P = .0003) and 2.4 mg beloranib groups (-7.0, 95% CI -10.5 to -3.6; P = .0001) vs placebo. Compared with placebo, weight change was greater with 1.8 mg (mean difference - 8.2%, 95% CI -10.8 to -5.6; P < .0001) and 2.4 mg beloranib (-9.5%, 95% CI -12.1 to -6.8; P < .0001). Injection site bruising was the most frequent adverse event with beloranib. Dosing was stopped early due to an imbalance in venous thrombotic events in beloranib-treated participants (2 fatal events of pulmonary embolism and 2 events of deep vein thrombosis) compared with placebo. CONCLUSIONS: MetAP2 inhibition with beloranib produced statistically significant and clinically meaningful improvements in hyperphagia-related behaviours and weight loss in participants with PWS. Although investigation of beloranib has ceased, inhibition of MetAP2 is a novel mechanism for treating hyperphagia and obesity.

A randomized, placebo-controlled trial of beloranib for the treatment of hypothalamic injury-associated obesity.

AIMS: Hypothalamic injury-associated obesity (HIAO) results from damage to the hypothalamus that often occurs with surgical removal/radiation therapy of tumours in the hypothalamic region, such as craniopharyngioma. There is currently no rigorously studied pharmaceutical treatment for the intractable weight gain and cardiometabolic consequences that occur in patients with HIAO. We aimed to assess efficacy, safety and tolerability of beloranib treatment for 4 to 8 weeks in patients with HIAO. MATERIALS AND METHODS: This Phase 2a, double-blind, placebo-controlled study included 14 patients with HIAO, randomized to receive beloranib 1.8 mg or placebo subcutaneously twice weekly for 4 weeks with an optional 4-week open-label extension in which all patients received beloranib. The primary endpoint was change in weight from baseline to Week 4. RESULTS: Participants were 64% female, with a mean (SD) age of 32 (9) years, BMI of 43 (7) kg/m2 and weight of 126 (22) kg. Compared with placebo (N = 4), beloranib 1.8 mg (N = 8) resulted in a mean (95% CI) difference in weight of -3.2 (-5.4, -0.9) kg after 4 weeks. Weight loss continued through the 8 weeks in patients randomized to beloranib (mean -6.2 [-8.2, -4.1] kg). Beloranib treatment was associated with improvements in high-sensitivity CRP. Adverse events were mild to moderate. No patients who received beloranib discontinued treatment. CONCLUSION: Beloranib treatment resulted in progressive weight loss in patients with HIAO that was comparable to that observed with beloranib in patients with exogenous obesity. These findings indicate a novel mechanism for treating obesity in patients with HIAO.

Crosstalk between angiotensin and the nonamyloidogenic pathway of Alzheimer's amyloid precursor protein.

The association between hypertension and an increased risk for Alzheimer's disease (AD) and dementia is well established. Many data suggest that modulation of the renin-angiotensin system may be meaningful for the prevention and therapy of neurodegenerative disorders, in particular AD. Proteolytic cleavage of the amyloid precursor protein (APP) by α-secretase precludes formation of neurotoxic Aß peptides and is expected to counteract the development of AD. An established approach for the up-regulation of α-secretase cleavage is the activation of G protein-coupled receptors (GPCRs). Therefore, our study aimed to analyze whether stimulation of angiotensin AT1 or AT2 receptors stably expressed in HEK cells influence the nonamyloidogenic pathway of APP processing. Treatment of both receptors with angiotensin II clearly showed that only activation of the AT1 receptor increased several fold the α-secretase-mediated shedding of APP. This effect was completely abolished by treatment with the AT1 receptor-specific antagonist telmisartan. Using the BIM-46187 inhibitor, we demonstrate that the Gαq protein-mediated pathway is involved in this stimulation process. Stimulation of AT1 receptors with the ß-arrestin-biased agonist SII was ineffective regarding α-secretase-mediated APP shedding. This result discloses that only the G protein-dependent pathway is involved in the Ang II-induced APP shedding. Blocking of Gßγ subunits by the inhibitor gallein completely prevented constitutive and Ang II-induced APP shedding. Our findings provide evidence that induction of APP shedding via Ang II/AT1 receptor stimulation is effected by G protein activation with Gßγ subunits playing important roles.

Anti-angiogenic Nanotherapy Inhibits Airway Remodeling and Hyper-responsiveness of Dust Mite Triggered Asthma in the Brown Norway Rat.

Although angiogenesis is a hallmark feature of asthmatic inflammatory responses, therapeutic anti-angiogenesis interventions have received little attention. Objective: Assess the effectiveness of anti-angiogenic Sn2 lipase-labile prodrugs delivered via αvß3-micellar nanotherapy to suppress microvascular expansion, bronchial remodeling, and airway hyper-responsiveness in Brown Norway rats exposed to serial house dust mite (HDM) inhalation challenges. Results: Anti-neovascular effectiveness of αvß3-mixed micelles incorporating docetaxel-prodrug (Dxtl-PD) or fumagillin-prodrug (Fum-PD) were shown to robustly suppress neovascular expansion (p<0.01) in the upper airways/bronchi of HDM rats using simultaneous 19F/1H MR neovascular imaging, which was corroborated by adjunctive fluorescent microscopy. Micelles without a drug payload (αvß3-No-Drug) served as a carrier-only control. Morphometric measurements of HDM rat airway size (perimeter) and vessel number at 21d revealed classic vascular expansion in control rats but less vascularity (p<0.001) after the anti-angiogenic nanotherapies. CD31 RNA expression independently corroborated the decrease in airway microvasculature. Methacholine (MCh) induced respiratory system resistance (Rrs) was high in the HDM rats receiving αvß3-No-Drug micelles while αvß3-Dxtl-PD or αvß3-Fum-PD micelles markedly and equivalently attenuated airway hyper-responsiveness and improved airway compliance. Total inflammatory BAL cells among HDM challenged rats did not differ with treatment, but αvß3+ macrophages/monocytes were significantly reduced by both nanotherapies (p<0.001), most notably by the αvß3-Dxtl-PD micelles. Additionally, αvß3-Dxtl-PD decreased BAL eosinophil and αvß3+ CD45+ leukocytes relative to αvß3-No-Drug micelles, whereas αvß3-Fum-PD micelles did not. Conclusion: These results demonstrate the potential of targeted anti-angiogenesis nanotherapy to ameliorate the inflammatory hallmarks of asthma in a clinically relevant rodent model.

Bedside to bench to bedside research: Estrogen receptor beta ligand as a candidate neuroprotective treatment for multiple sclerosis.

Protective effects of pregnancy during MS have led to clinical trials of estriol, the pregnancy estrogen, in MS. Since estriol binds to estrogen receptor (ER) beta, ER beta ligand could represent a "next generation estriol" treatment. Here, ER beta ligand treatment was protective in EAE in both sexes and across genetic backgrounds. Neuroprotection was shown in spinal cord, sparing myelin and axons, and in brain, sparing neurons and synapses. Longitudinal in vivo MRIs showed decreased brain atrophy in cerebral cortex gray matter and cerebellum during EAE. Investigation of ER beta ligand as a neuroprotective treatment for MS is warranted.

A Pharmacokinetic and Pharmacodynamic Study of Maraviroc as Acute Graft-versus-Host Disease Prophylaxis in Pediatric Allogeneic Stem Cell Transplant Recipients with Nonmalignant Diagnoses.

Maraviroc is an allosteric small molecule antagonist of chemokine receptor type 5 (CCR5) and has been used in adult allogeneic hematopoietic stem cell transplant (HSCT) recipients to prevent acute graft-versus-host disease (GVHD) of the gastrointestinal (GI) tract and liver. The goal of this study was to establish feasibility and pharmacokinetic and pharmacodynamic profiles of maraviroc in pediatric HSCT recipients. Children ages 2 to 12 years were enrolled and maraviroc was added to standard GVHD prophylaxis, which included a calcineurin inhibitor and either steroids or mycophenolate mofetil. Maraviroc was started on day -3 and administered at a dose of approximately 300 mg/m(2) orally twice daily until day +30 after stem cell infusion. On days 0 and day +10, samples for pharmacokinetic analysis were collected before the dose and 1, 2, 4, 6, 8, and 12 hours after maraviroc administration. Additional trough concentrations were collected on days +7, 14, and 21. Patients were followed until day +100 for acute GVHD. Functional blockade of CCR5 was assessed in a pharmacodynamic assay by flow cytometry. Thirteen patients, median age of 4 years (range, 2 to 11 years), were prospectively enrolled. Underlying diagnoses included a primary immune deficiency (n = 6), hemoglobinopathy (n = 4), metabolic disorder (n = 1), and bone marrow failure syndrome (n = 2). Patients received either a myeloablative preparative regimen (n = 7) or a reduced-intensity conditioning regimen (n = 6). Cyclosporine and methylprednisolone (n = 7) was the predominant GVHD prophylactic regimen, followed by tacrolimus and mycophenolate mofetil (n = 4) and tacrolimus and steroids (n = 2). Two formulations of maraviroc (150-mg tablets and 20-mg/mL solution) were used on study. Mean (± SD) area under the concentration-time curve from 0 to 12 hours was 4805 ± 3265 hour * ng/mL on day 0 and 5917 ± 4048 hour * ng/mL on day +10. Four patients developed grade 1 or 2 acute skin GVHD before day +100 and were successfully treated. Two patients developed grade 3 acute GI GVHD on days +23 and +24 after HSCT and both had discontinued maraviroc before development of GI GVHD. No adverse effects attributable to maraviroc were observed and administration by enteral tubes was well tolerated by children and accepted by parents. All evaluable patients demonstrated functional CCR5 blockade on day 0. Administration of maraviroc is feasible in most pediatric HSCT recipients with good safety and tolerability profile.

Biophysical characterization of small molecule antiviral-loaded nanolipogels for HIV-1 chemoprophylaxis and topical mucosal application.

UNLABELLED: Nanocarriers are versatile vehicles for drug delivery, and emerging as platforms to formulate and deliver multiple classes of antiretroviral (ARV) drugs in a single system. Here we describe the fabrication of hydrogel-core and lipid-shell nanoparticles (nanolipogels) for the controlled loading and topical, vaginal delivery of maraviroc (MVC) and tenofovir disoproxil fumarate (TDF), two ARV drugs with different mechanisms of action that are used in the treatment of HIV. The nanolipogel platform was used to successfully formulate MVC and TDF, which produced ARV drug-loaded nanolipogels that were characterized for their physical properties and antiviral activity against HIV-1 BaL in cell culture. We also show that administration of these drug carriers topically to the vaginal mucosa in a murine model leads to antiviral activity against HIV-1 BaL in cervicovaginal lavages. Our results suggest that nanolipogel carriers are promising for the encapsulation and delivery of hydrophilic small molecule ARV drugs, and may expand the nanocarrier systems being investigated for HIV prevention or treatment. STATEMENT OF SIGNIFICANCE: Topical, mucosal intervention of HIV is a leading strategy in the efforts to curb the spread of viral infection. A significant research thrust in the field has been to characterize different dosage forms for formulation of physicochemically diverse antiretroviral drugs. Nanocarriers have been used to formulate and deliver small molecule and protein drugs for a range of applications, including ARV drugs for HIV treatment. The broad significance of our work includes evaluation of lipid-shell, hydrogel-core nanoparticles for formulation and topical, vaginal delivery of two water-soluble antiretroviral drugs. We have characterized these nanocarriers for their physical properties and their biological activity against HIV-1 infection in vitro, and demonstrated the ability to deliver drug-loaded nanocarriers in vivo.

Maraviroc and reverse transcriptase inhibitors combinations as potential preexposure prophylaxis candidates.

OBJECTIVE: Receptive anal intercourse in both men and women is associated with the highest probability for sexual acquisition of HIV infection. As part of a program to develop an effective prevention strategy, we performed an ex-vivo preclinical evaluation to determine the efficacy of multiple double combinations of maraviroc (MVC) and reverse transcriptase inhibitors (RTIs). DESIGN: The entry inhibitor, MVC, a nucleotide RTI, tenofovir and two non-nucleoside RTIs, UC781 and TMC120 (dapivirine, DPV), were used in double, combinations against a panel of CCR5-using clade B and clade C HIV-1 isolates and against MVC-escape variants. A gel-formulated version of MVC-DPV combination was also tested. METHODS: Indicator cells, cocultures of immature dendritic cells with CD4T cells, and colorectal tissue explants were used to assess antiviral activity of drug combinations. RESULTS: All dual MVC-RTI combinations tested inhibited MVC-sensitive and resistant isolates in cellular and colorectal explants models. All the combinations were positive with no reduction in the activity of MVC. In tissue explants, the combinations against all viral isolates tested produced an increase in the activity of MVC. An initial gel-formulation of MVC-DPV combination showed greater and prolonged antiviral activity of MVC in mucosal tissue explants. CONCLUSION: This study demonstrates that combinations based on antiretroviral drugs inhibiting HIV transmission at viral entry and reverse transcription have potential as prevention strategies against colorectal transmission of HIV-1 including MVC-resistant isolates. Preclinical evaluation with colorectal tissue explants indicates that a gel-formulation of MVC-DPV is an effective candidate colorectal microbicide.

No selection of CXCR4-using variants in cell reservoirs of dual-mixed HIV-infected patients on suppressive maraviroc therapy.

We used ultradeep sequencing to investigate the evolution of the frequency of CXCR4-using viruses in the peripheral blood mononuclear cells of 22 patients infected with both CCR5 and CXCR4-using viruses treated with the CCR5 antagonist maraviroc for 24 weeks and a stable antiviral therapy. The mean CXCR4-using virus frequency in peripheral blood mononuclear cells was 59% before maraviroc intensification and 52% after 24 weeks of effective treatment, indicating no selection by maraviroc.

A Rapid Screening Assay Identifies Monotherapy with Interferon-ß and Combination Therapies with Nucleoside Analogs as Effective Inhibitors of Ebola Virus.

To date there are no approved antiviral drugs for the treatment of Ebola virus disease (EVD). While a number of candidate drugs have shown limited efficacy in vitro and/or in non-human primate studies, differences in experimental methodologies make it difficult to compare their therapeutic effectiveness. Using an in vitro model of Ebola Zaire replication with transcription-competent virus like particles (trVLPs), requiring only level 2 biosafety containment, we compared the activities of the type I interferons (IFNs) IFN-α and IFN-ß, a panel of viral polymerase inhibitors (lamivudine (3TC), zidovudine (AZT) tenofovir (TFV), favipiravir (FPV), the active metabolite of brincidofovir, cidofovir (CDF)), and the estrogen receptor modulator, toremifene (TOR), in inhibiting viral replication in dose-response and time course studies. We also tested 28 two- and 56 three-drug combinations against Ebola replication. IFN-α and IFN-ß inhibited viral replication 24 hours post-infection (IC50 0.038µM and 0.016µM, respectively). 3TC, AZT and TFV inhibited Ebola replication when used alone (50-62%) or in combination (87%). They exhibited lower IC50 (0.98-6.2µM) compared with FPV (36.8µM), when administered 24 hours post-infection. Unexpectedly, CDF had a narrow therapeutic window (6.25-25µM). When dosed >50µM, CDF treatment enhanced viral infection. IFN-ß exhibited strong synergy with 3TC (97.3% inhibition) or in triple combination with 3TC and AZT (95.8% inhibition). This study demonstrates that IFNs and viral polymerase inhibitors may have utility in EVD. We identified several 2 and 3 drug combinations with strong anti-Ebola activity, confirmed in studies using fully infectious ZEBOV, providing a rationale for testing combination therapies in animal models of lethal Ebola challenge. These studies open up new possibilities for novel therapeutic options, in particular combination therapies, which could prevent and treat Ebola infection and potentially reduce drug resistance.

Mucosal Topical Microbicide Candidates Exert Influence on the Subsequent SIV Infection and Survival by Regulating SIV-Specific T-Cell Immune Responses.

OBJECTIVE: To determine whether mucosal topical microbicides have any influence on disease progression during subsequent simian immunodeficiency virus (SIV) infection. DESIGN: A 2-phase study was performed in primate monkeys. The first phase mimicked microbicide efficacy studies; the second phase served to determine the disease progression in a productive infection model. METHODS: During the first phase, monkeys were intrarectally pretreated with tenofovir, sifuvirtide (SFT), or maraviroc-formulated microbicides and then challenged with low-dose SHIV-1157ipd3N4. Second, all monkeys were rechallenged with a single high dose of SIVmac239 to generate productive infections. The survival rate, viral loads, CD4(+) T-cell counts, and SIV-specific T-cell responses were determined during the 104-week following up. RESULTS: Repeated rectal challenges did not result in productive infection in all groups, evidenced by undetectable viral loads with occasional viral blips during the first phase of this study. All monkeys were productively infected after the high-dose rechallenge with SIVmac239. Two groups, including maraviroc-treated and tenofovir-treated groups, experienced 100% mortality during the 104-week following up. In contrast, the SFT-treated group showed significantly higher survival, and only 25% died at week 95. Interestingly, SIV-specific T-cell responses were also significantly higher in the SFT group. Transcriptomic analyses evidenced immune imprint in immune system among different microbicide-treated groups. CONCLUSIONS: This study provides preliminary but important evidence for the influence of prophylactically applied microbicides on disease progression of subsequent SIV infection and suggests that the long-term immune safety concern for microbicides should be also considered in the effort to develop effective microbicides.

Single oral dose of maraviroc does not prevent ex-vivo HIV infection of rectal mucosa in HIV-1 negative human volunteers.

OBJECTIVE: Maraviroc (MVC) is a potential candidate for 'on demand' preexposure prophylaxis. In the present study, we evaluated the efficacy of a single oral dose of MVC to prevent ex-vivo HIV-1 infection of rectal tissue in humans. DESIGN AND METHODS: Eight HIV-1-negative healthy volunteers received a single oral dose of MVC (300 or 600 mg), and two additional volunteers received tenofovir disoproxil fumarate/emtricitabine (TDF/FTC, 300/200 mg) for 10 days. Rectal biopsies were performed prior to the ex-vivo challenge (day 0), at day 7 (4 h after MVC) or after 10 days with TDF/FTC. Rectal biopsies were infected ex-vivo, and viral inhibition and CCR5 occupancy was analyzed. MVC concentration in plasma and rectal tissue was measured just after biopsy and after viral incubation. RESULTS: Ex-vivo rectal tissue protection with MVC was incomplete in all but two participants, whereas TDF/FTC avoided ex-vivo infection in the two controls. Median dose-normalized concentration of MVC was significantly higher in rectal tissue than in plasma (561.1 and 155.1 ng/ml, respectively). A significant loss of MVC during the virus incubation (about 60%) and a low CCR5 occupancy (approximately 45%) were detected in rectal cells. CONCLUSIONS: An ex-vivo challenge with a single oral dose of MVC does not prevent ex-vivo infection of human rectal mucosa. The lack of prophylactic efficacy observed suggests that 'on demand' MVC preexposure prophylaxis would not prevent rectal HIV-1 transmission.

Maraviroc Clinical Test (MCT) as an alternative tool to decide CCR5-antagonists prescription in naïve HIV-infected patients.

Our aim was to analyze the virological response to a combined antiretroviral therapy started after Maraviroc Clinical Test (MCT) in naïve HIV-infected patients. Forty-one patients were exposed to MCT, based on an 8-day MVC monotherapy. If undetectability or a viral load reduction >1 log10 HIV-RNA copies/ml was achieved, a MVC-containing cART was prescribed. Forty patients showed a positive MCT; undetectability after 48weeks on cART was achieved in 34/41 (82.9%) patients. The result of MCT was compared with a genotypic tropism method and with Trofile®, showing 10.7% and 18.75% discordance rates, respectively. MCT is a reliable tool to decide CCR5-antagonists prescription, also in the naïve scenario where most patients show a virological response to MVC independently the tropism result reported by genotypic or phenotypic methods.

Protection Against Rectal Chimeric Simian/Human Immunodeficiency Virus Transmission in Macaques by Rectal-Specific Gel Formulations of Maraviroc and Tenofovir.

BACKGROUND: Rectal human immunodeficiency virus (HIV) transmission is an important driver of the HIV epidemic. Optimally formulated gels of antiretroviral drugs are under development for preventing rectally acquired HIV. We investigated in a macaque model the pharmacokinetics and efficacy of 3 rectal gel formulations METHODS: Single-dose pharmacokinetics of low-osmolar 1% maraviroc (MVC), 1% tenofovir (TFV), or 1% MVC/1% TFV combination gel were evaluated in blood, rectal fluids, colorectal biopsy specimens, and rectal lymphocytes. Efficacy was evaluated over 10 twice-weekly rectal SHIV162p3 challenges in rhesus macaques that received either placebo (n = 7), MVC (n = 6), TFV (n = 6), or MVC/TFV (n = 6) gel 30 minutes before each challenge. RESULTS: MVC and TFV were detected in plasma 30 minutes after gel application and remained above 95% inhibitory concentrations in rectal fluids at 24 hours. MVC, TFV, and TFV diphosphate (TFV-DP) concentrations in colorectal tissues collected up to 30 cm from the anal margin were all high at 2 hours, demonstrating rapid and extended tissue dosing. TFV-DP concentrations in tissue homogenates and rectal lymphocytes were highly correlated (r(2) = 0.82). All 3 gel formulations were highly protective (82% efficacy; P ≤ .02 by the log-rank test). CONCLUSIONS: Desirable pharmacokinetic profiles and high efficacy in this macaque model support the clinical development of these gel formulations for preventing rectal HIV infection.