Search

ObjectivesDolutegravir has replaced efavirenz as first-line treatment in universal HIV guidelines. We sought to ascertain the contributory effect of SNPs in four key genes linked to dolutegravir disposition (UGT1A1, ABCG2, CYP3A and NR1I2) on plasma dolutegravir pharmacokinetics.MethodsPaired pharmacogenetic/pharmacokinetic data from 93 subjects were analysed for association using multivariate linear regression.ResultsCo-occurring UGT1*28 and NR1I2 c.63396C>T homozygosity was associated with a 79% increase in AUC0–24 (P = 0.001; 27% if analysed individually), whilst combined ABCG2 c.421C>A and NR1I2 c.63396C>T variants were associated with a 43% increase in Cmax (P = 0.002) and a 39% increase in AUC0–24 (P = 0.002). When analysed individually, homozygosity for the NR1I2 c.63396C>T variant alleles was associated with a 28% increase in Cmax (P = 0.033) and homozygosity for the ABCG2 c.421C>A variant alleles was associated with a 28% increase in Cmax (P = 0.047). The UGT1A1*28 (rs8175347) poor metabolizer status (*28/*28; *28/*37; *37/*37) was individually associated with a 27% increase in AUC0–24 (P = 0.020). The combination of UGT1A1*28 poor metabolizer and UGT1A1*6 intermediate metabolizer statuses correlated with a 43% increase in AUC0–24 (P = 0.023).ConclusionsThis study showed a pharmacogenetic association between dolutegravir pharmacokinetics and variants in the ABCG2, UGT1A1 and NR1I2 genes, particularly when combined. Further research is warranted to confirm these associations in population-specific studies and to investigate their putative relationship with dolutegravir pharmacodynamics.

During the late phase of the HIV-1 replication cycle, the Gag polyproteins are transported to the plasma membrane (PM) for assembly. Gag targeting and assembly on the PM is dependent on interactions between its matrix (MA) domain and phosphatidylinositol-4,5-bisphosphate (PI(4,5)P2). Subsequent to Gag assembly, the envelope (Env) protein is recruited to the PM for incorporation into virus particles. Evidence suggests that the incorporation of the Env protein is mediated by interactions between the MA domain of Gag and the cytoplasmic tail of the gp41 subunit of Env (gp41CT), a mechanism that remains to be elucidated. Trimerization of the MA domain of Gag appears to be an obligatory step for this interaction. The interplay between gp41CT, the MA trimer, and the membrane has yet to be determined. Our lab has pioneered methods and approaches to investigate, at the molecular level, how the retroviral MA domains of Gag interact with membranes, a key requirement for understanding the Gag assembly and Env incorporation. Herein, we devised innovative approaches that will enable the structural characterization of the gp41CT–MA–membrane interactions. We employed structural biology (NMR and cryo-electron microscopy, biophysical methods, and biochemical tools to generate a macromolecular picture of how the MA domain of Gag binds to the membrane and how it interacts with gp41CT. To this end, we: (i) determined the three-dimensional structure of HIV-1 gp41CT and characterized its interaction with the membrane, (ii) engineered trimeric constructs of gp41CT and the MA to recapitulate the native and functional states of the proteins, and (iii) utilized membrane nanodisc technology to anchor the MA and gp41CT proteins. Our studies will allow for a detailed structural characterization of the gp41CT–MA–membrane interactions, which will advance our knowledge of HIV-1 Gag assembly and Env incorporation.

Neodymium-doped yttrium aluminum garnet (Nd:YAG) lasers are among the most commonly used lasers with a wide variety of applications from biomedicine to manufacturing. The ubiquity of these lasers increases the likelihood of accidental ocular injury resulting in permanent visual impairment. We performed dosimetry studies to determine retinal damage thresholds and hemorrhagic lesions in the porcine eye with Qswitched Nd:YAG lasers. The Yucatan miniature pig model exhibited similarities in ocular anatomy to human eyes. The Nd:YAG laser, tuned to 1064 nm with a pulse width of seven nanoseconds, delivered laser energy to the retina. Retinal imaging modalities including fundus photography, real-time video, confocal scanning laser ophthalmoscopy (cSLO), and spectral domain optical coherence tomography (SD-OCT) provided visualization of retinal morphology at multiple time points. Retinal damage thresholds were grouped into three categories: minimum visible lesion (MVL), contained hemorrhagic lesion (CHL), and vitreal hemorrhagic lesion (VHL). Probit analysis determined the effective dose for 50% probability of damage (ED50) for each lesion category. The threshold to produce a MVL was 0.193 mJ based on 24-hour assessments of the retina. The one-hour hemorrhagic lesion thresholds were 0.408 mJ and 1.52 mJ for CHL and VHL, respectively

During the late phase of the HIV-1 replication cycle, the viral Gag polyproteins are targeted to the plasma membrane for assembly. The Gag-membrane interaction is mediated by binding of Gag's N-terminal myristoylated matrix (MA) domain to phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). The viral envelope (Env) glycoprotein is then recruited to the assembly sites and incorporated into budding particles. Evidence suggests that Env incorporation is mediated by interactions between Gag's MA domain and the cytoplasmic tail of the gp41 subunit of Env (gp41CT). MA trimerization appears to be an obligatory step for this interaction. Insufficient production of a recombinant MA trimer and unavailability of a biologically relevant membrane system have been barriers to detailed structural and biophysical characterization of the putative MA-gp41CT-membrane interactions. Here, we engineered a stable recombinant HIV-1 MA trimer construct by fusing a foldon domain (FD) of phage T4 fibritin to the MA C terminus. Results from NMR experiments confirmed that the FD attachment does not adversely alter the MA structure. Employing hydrogen-deuterium exchange MS, we identified an MA-MA interface in the MA trimer that is implicated in Gag assembly and Env incorporation. Utilizing lipid nanodiscs as a membrane mimetic, we show that the MA trimer binds to membranes 30-fold tighter than does the MA monomer and that incorporation of PI(4,5)P2 and phosphatidylserine enhances the binding of MA to nanodiscs. These findings advance our understanding of a fundamental mechanism in HIV-1 assembly and provide a template for investigating the interaction of MA with gp41CT. This work was supported by National Institutes of Health Grants 5R01GM117837 and 9R01AI150901 (to J.S.S.) Sí

Advancement in drug therapies and patient care have drastically improved the mortality rates of HIV-1 infected individuals. Many of these therapies were developed or improved upon by using structure-based techniques, which underscore the importance of understanding essential mechanisms in the replication cycle of HIV-1 at the structural level. One such process which remains poorly understood is the incorporation of the envelope glycoprotein (Env) into budding virus particles. Assembly of HIV particles is initiated by targeting of the Gag polyproteins to the inner leaflet of the plasma membrane (PM), a process mediated by the N-terminally myristoylated matrix (MA) domain and phosphatidylinositol 4,5-bisphosphate (PI(4,5)P2). There is strong evidence that formation of the Gag lattice on the PM is a prerequisite for the incorporation of Env into budding particles. It is also suggested that Env incorporation is mediated by an interaction between its cytoplasmic tail (gp41CT) and the MA domain of Gag. In this review, we highlight the latest developments and current efforts to understand the interplay between gp41CT, MA, and the membrane during assembly. Elucidation of the molecular determinants of Gag–Env–membrane interactions may help in the development of new antiviral therapeutic agents that inhibit particle assembly, Env incorporation and ultimately virus production.

BackgroundDemographic data show an increasingly aging human immunodeficiency virus (HIV) population worldwide. Recent concerns over dolutegravir-related neuropsychiatric toxicity have emerged, particularly amongst older people living with HIV (PLWH). We describe the pharmacokinetics (PK) of dolutegravir (DTG) 50 mg once daily in PLWH aged 60 and older. Additionally, to address calls for prospective neuropsychiatric toxicodynamic data, we evaluated changes in sleep quality and cognitive functioning in this population after switching to abacavir (ABC)/lamivudine (3TC)/DTG over 6 months.MethodsPLWH ≥60 years with HIV-viral load ResultsIn total, 43 participants enrolled, and 40 completed the PK phase. Overall, 5 discontinued (2 due to sleep-related adverse events, 4.6%). DTG maximum concentration (Cmax) was significantly higher in patients ≥60 years old versus controls (geometric mean 4246 ng/mL versus 3402 ng/mL, P = .005). In those who completed day 180 (n = 38), sleep impairment (Pittsburgh Sleep Quality Index) was marginally higher at day 28 (P = .02), but not at days 90 or 180. Insomnia, daytime functioning, and fatigue test scores did not change statistically over time.ConclusionsDTG Cmax was significantly higher in older PLWH. Our data provides clinicians with key information on the safety of prescribing DTG in older PLWH.

Purpose of reviewThis review of recent published literature and data presented at scientific meetings on integrase stand transfer inhibitors (InSTIs) examines how these findings may impact on their future clinical use.Recent findingsElvitegravir (EVG), raltegravir (RAL) and dolutegravir (DTG) are InSTIs recommended as first-line options for treatment naive patients by the European AIDS Clinical Society, British HIV Association, International AIDS Society-USA and DHHS. InSTIs have gained a leading role in the management of HIV-1 because of increased viral suppression and maintaining undetectability with fewer side-effects.RAL 1200 mg once-daily (QD) has been shown to be noninferior to 400 mg BD, and the European Medicines Agency has approved QD RAL for review. RAL and DTG are not metabolized via cytochrome P450 (CYP) resulting in fewer drug interactions and less toxicity risk in patients receiving direct-acting antivirals and other coadministered medications.EVG is currently available as a single tablet regimen and requires cobisistat, a pharmacokinetic booster and CYP3A inhibitor to allow QD dosing. EVG will soon be available in combination with tenofovir alfenamide, which is as efficacious as tenofovir disoproxil fumarate, but offers better renal and bone outcomes.DTG has a high genetic barrier to resistance and has been the subject of a number of simplification and treatment failure trials and shown promise. There are some emerging reports of neuropsychiatric and gastrointestinal side-effects associated with DTG, which were not reported in clinical trials emphasizing the importance of real-life data.Carbotegravir, a long-acting InSTI, is currently in the pipeline of development.SummaryAll three InSTIs have impressive data on efficacy, tolerability and safety. The unique differences of each InSTI's pharmacokinetics and pharmacodynamics lend themselves to various clinical scenarios, enabling us as clinicians to provide better patient-centred care.

Background:Dolutegravir combined with darunavir/cobicistat is a promising NRTI-sparing and/or salvage strategy for the treatment of HIV-1 infection. Methods:This Phase I, open-label, 57 day, crossover, pharmacokinetic (PK) study, enrolled healthy volunteers aged 18-65 years, who were randomized to one of two groups. Group 1 received dolutegravir (50 mg) once daily for 14 days followed by a 7 day washout, then a 14 day dolutegravir/darunavir/cobicistat (DTG/DRV/COBI) once-daily co-administration period followed by a 7 day washout and finally a 14 day period of darunavir/cobicistat (800/150 mg) once daily. Group 2 followed the same sequence starting with darunavir/cobicistat and concluding with dolutegravir. Each group underwent intensive PK sampling over 24 h on day 14 of each drug period and DTG/DRV/COBI concentrations were measured using validated LC-MS/MS methods. Results:Twenty participants completed all PK phases. Thirteen were female and median age and BMI were 33.5 years and 27 kg/m2. Dolutegravir geometric mean ratios (GMR, DTG/DRV/COBI versus dolutegravir alone) and 90% CI for Cmax, AUC0-24 and C24 were 1.01 (0.92-1.11), 0.95 (0.87-1.04) and 0.9 (0.8-1.0), respectively. Darunavir GMR (DRV/COBI/DTG versus darunavir/cobicistat alone) and 90% CI for Cmax, AUC0-24 and C24 were 0.90 (0.83-0.98), 0.93 (0.86-1.00) and 0.93 (0.78-1.11), respectively. No grade 3 or 4 adverse events or laboratory abnormalities were observed. Conclusions:Concentrations of dolutegravir and darunavir, when boosted with cobicistat, decreased by

Computed tomography (CT) imaging was applied to Mormon Mesa petrocalcic and related materials to observe the location, arrangement, and interconnectedness of void spaces and to visualize density differences that may define pedogenic mineral relationships. Density variations visualized by these scans reveal features that may result from long developed pedogenic processes within these unique soil materials. Voids are present as cracks (shown to interconnect three-dimensionally), elongated spaces filled with unconsolidated materials, and oblate-spheroidal shapes that appear to be associated with detrital carbonate clasts. Density variations viewed through CT imaging may show the products of mineral neoformation, mineral packing differences during crystal growth, and/or older, denser petrocalcic materials eroded and incorporated via subsequent recementation (pisoliths). Computed tomography scanning can be used to understand unique forms and processes that are vital to our understanding of dissolution/precipitation and other pedogenic processes at work in these mature petrocalcic soils. We encourage the application of CT imaging to similar soil materials.

Objectives To evaluate dolutegravir and elvitegravir/cobicistat pharmacokinetics in HIV-negative volunteers up to 10 days after drug cessation. Methods Healthy volunteers received 50 mg of dolutegravir once-daily for 10 days, then underwent a 9 day wash-out period, and then received elvitegravir/cobicistat as part of Stribild(®) (245 mg of tenofovir, 200 mg of emtricitabine, 150 mg of elvitegravir and 150 mg of cobicistat) for 10 days. Serial pharmacokinetic (PK) sampling occurred prior to the final dose of each course and at regular intervals for up to 216 h (10 days) after drug cessation. Concentrations were determined by LC-MS/MS, and PK parameters were illustrated as geometric mean and 90% CI. Results Seventeen volunteers completed the study. For dolutegravir, plasma terminal elimination t1/2 to the last measurable concentration (within 216 h) was longer than its t1/2 within the dosing interval (0-24 h): 14.3 h (12.9-15.7 h) versus 23.1 h (19.7-26.6 h); conversely, the terminal elimination t1/2 for elvitegravir was lower than its t1/2 within the dosing interval (0-24 h): 10.8 h (9.7-13.0 h) versus 5.2 h (4.7-6.1 h). Dolutegravir concentrations were above the protein-adjusted (PA) IC90 (64 ng/mL) in 100% of subjects after 36 and 48 h and in 94% after 60 and 72 h. All subjects had detectable dolutegravir concentrations at 96 h, a mean of 23.5% above the IC90. Elvitegravir concentrations were above the PA IC95 (45 ng/mL) in 100% of subjects at 24 h, 65% at 36 h but 0% after 48 h. Conclusions Our data show marked differences in the elimination rates of dolutegravir and elvitegravir following treatment interruption, which is likely to impact the extent to which drug doses can be delayed or missed. They suggest that clinical differences may emerge in patients who have suboptimal adherence.

Dolutegravir and Elvitegravir belongs to a class of integrase inhibitors which has recently been approved by the FDA for the treatment of HIV-infection. Elvitegravir and its co-administered booster drug, Cobicistat, has shown the potential to be a candidate for a one pill once a day regimen and is currently a component of many clinical trials. A sensitive LC-MS/MS method has been developed and validated for the simultaneous determination of these three drugs in human plasma. A liquid- liquid extraction was used as a sample preparation technique using 100μL of plasma. The method was validated from 10 to 4000ng/mL for Dolutegravir, Elvitegravir and Cobicistat. Chromatography was performed on XBridge C18 2.1mm×50mm column, using an 80:20 methanol/water mobile phase containing 0.1% formic acid on a gradient program. This method was successfully applied for ongoing clinical trials.

We performed single-breath wash-in tests for He and SF6 in four subjects exposed to 14 days of microgravity (microG) during the Spacelab flight Spacelab Life Sciences-2. Subjects inspired a vital capacity breath of 5% He-1.25% SF6-balance O2 and then exhaled to residual volume at 0.5l/s. The tests were also performed with a 10-s breath hold at the end of inspiration. Measurements were also made with the subjects standing and supine in 1 G. Phase III slope was measured after the dead-space washout and before the onset of airway closure. In all subjects in 1 G, whether standing or supine, phase III slope for SF6 was significantly steeper than that for He. However, in microG, the slopes became the same. Furthermore, after breath holding in microG, the SF6 slopes were significantly flatter than those for He. On return to 1 G, the changes were reversed, and there was no difference between preflight and postflight values. Because most of the phase III slope reflects events occurring in the acinar regions of the lung, the results suggest that microG causes conformational changes in the acini or changes in cardiogenic mixing in the lung periphery, but in either case the mechanism is unclear.

Conductivity measurements of ultrahigh vacuum-sublimed (MPcF)n thin films (M=Al, Ga; Pc = phthalocyanine) were performed under ultrahigh vacuum and well-defined gas conditions. The intrinsic dark conductivity of as-prepared (GaPcF)n thin films is remarkably high (4 × 10−6 S/cm at 300 K) if compared to pressed pellets (3 × 10−6 S/cm at 300 K). After heating to 485 K, the conductivity decreases to 2.4 × 10−6 S/cm. This is the result of changes in the geometric structure observed in scanning electron microscopy. In contrast to (GaPcF)nm, a very low dark conductivity (

Spurious readings from a mass spectrometer have been reported following the administration of aerosol bronchodilators. We quantified the response of various respiratory gas analyzers to the aerosol propellant of albuterol inhalant (Proventil). The mass spectrometer systems tested, two Advantage systems, a SARA system, and a Model 6000 Ohmeda system, all displayed artifactual readings in response to the albuterol propellant. Each metered dose of the Proventil brand of albuterol contains 4 ml of Freon 11 (trichloromonofluoromethane) and 11 ml of Freon 12 (dichlorodifluoromethane). The concentration of propellant was expressed in doses/L, where each liter of gas contains 0.4 vol % of Freon 11 and 1.1 vol % of Freon 12 per dose. In proportion to the concentration of albuterol propellant, the two Advantage systems showed substantial readings of isoflurane (%) when no isoflurane was present (13% and 16% per dose/L) and reduced readings of enflurane (-8% and -10% per dose/L) and carbon dioxide (CO2) (-3 and +5 mm Hg per dose/L). The SARA system showed substantial CO2 readings when no CO2 was present (5 mm Hg per dose/L) and displayed small enflurane readings (0.1% per dose/L) when no enflurane was present. The Model 6000 unit showed CO2 readings when no CO2 was present (5 mm Hg per dose/L). Neither the Raman spectrometer, the infrared spectrometers, nor the piezoadsorptive analyzer we tested showed an artifactual effect of albuterol propellant on any of its readings. Simulation and clinical tests demonstrated that a single dose of albuterol propellant into a breathing circuit at the onset of inspiration resulted in concentrations of 0.8 and 0.3 dose/L, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Beta-endorphin, morphine, and saline were given intravenously to a single schizophrenic subject on separate occasions in a double-blind design. EEG spectral analyses performed on data collected before and after drug injection demonstrated that β-endorphin and morphine produced similar increases in alpha power within 5 to 15 minutes after injection. This effect could be distinguished from two placebo (saline) injections. These data suggest that intravenous β-endorphin can produce changes in the central nervous system in humans.

Compartmental Schiff-base ligands have been prepared from 2,3-dihydro-2-hydroxy-4H-1-benzopyran-4-one and alkane-α,ω-diamines. Mononuclear copper(II), nickel(II), cobalt(II), and dioxouranium(VI) complexes have been prepared and the site occupancy established: the nickel(II) and cobalt(II) are N2O2, the dioxouranium(VI) is O2O2, and positional isomers are available for copper(II). Homobinuclear copper(II), nickel(II), and oxovanadium(IV) complexes were prepared, as were heterobinuclear complexes containing copper(II) and dioxouranium(VI), and nickel(II) and dioxouranium(VI). Three-dimensional X-ray crystal structure analyses have been carried out on {3,3′-(ethane-1,2-diyl-di-imino)bis[1-(o-hydroxyphenyl)-prop-2-enonato]-(N,N′,O1O1′)}nickel(II), (8a; M = Ni), {3,3′-(propane-1,3-diyldi-imino)bis[1-(o-hydroxyphenyl)prop-2-enonato](N,N′,O1,O1′)}copper(II), (8b; M = Cu), and the related complex {3,3′-(ethane-1,2-diyldi-imino)bis[1-(o-hydroxyphenyl)but-2-enonato](N,N′,O1,O1′)}nickel(II), (10). Two crystallographically independent molecules were found for (10). Complex (8a; M = Ni) has Z= 8 in a monoclinic cell with space group C2/c(no. 15, C62h), and dimensions a= 23.373(13), b= 7.800(1), c= 20.819(6)A, β= 110.14(4)°; R= 0.1341 for 603 reflections. Complex (8b; M = Cu) has Z= 8 in a monoclinic cell with space group C2/c(no. 15, C62h) and dimensions a= 23.098(18), b= 9.525(5), c= 18.046(8)A, β= 104.15(5)°; R= 0.1063 for 891 reflections. Complex (10) has Z= 16 in a monoclinic cell with space group B21/c(a non-standard setting of P21/c, no. 14, C52h) and dimensions a= 26.11(4), b= 7.797(4), c= 38.42(6)A, β= 93.656(8)°; R= 0.0376 for 4 344 reflections.

sons frequenting swimming pools. Of principal importance are the standards related to the bacterial quality of the water and the procedures for purification. Because of its simplicity of application, its economy of use, and its continuing, bactericidal action through the maintenance of residual levels, chlorine has been and is being used extensively to control the quality of swimming pool water. Many studies have been conducted to measure the effectiveness of chlorination of swimming pool water under varying conditions and to develop improved methods of application, and much consideration has been given to the amount and type of chlorine residual of the swimming pool water that should be maintained and the choice of organism or group of organisms that should be used as an indication of pollution.'"7 The Joint Committee on Bathing Places of the American Public Health

Large-scale geologic CO2 storage (GCS) can be limited by overpressure, while geothermal energy production is often limited by pressure depletion. We investigate how synergistic integration of these complementary systems may enhance the viability of GCS by relieving overpressure, which reduces pore-space competition, the Area of Review, and the risks of CO2 leakage and induced seismicity, and by producing geothermal energy and water, which can defray parasitic energy and water costs of CO2 capture.