Comparative Pharmacokinetics and Local Tolerance of Tenofovir Alafenamide (TAF) From Subcutaneous Implant in Rabbits, Dogs, and Macaques.

The administration of antiretrovirals (ARVs) for HIV pre-exposure prophylaxis (PrEP) is highly efficacious and may benefit from new long-acting (LA) drug delivery approaches. This paper describes a subcutaneous, reservoir-style implant for the LA delivery of tenofovir alafenamide (TAF) and documents the preclinical assessment of implant safety and pharmacokinetics (PK) in New Zealand White (NZW) rabbits (3 groups of n = 5), beagle dogs (2 groups of n = 6), and rhesus macaques (2 groups of n = 3). Placebo implants were placed in rabbits (n = 10) and dogs (n = 12). Implant parameters, including selection of the TAF form, choice of excipient, and PCL formulation were tuned to achieve targeted concentrations of the active anabolite of TAF, tenofovir diphosphate (TFV-DP), within peripheral blood mononuclear cells (PBMCs) and mucosal tissues relevant to HIV transmission. Sustained concentrations of TFV-DP in PBMCs over 100 fmol/106 cells were achieved in all animal species indicating that the implants effectively delivered TAF for 3-6 months. Unlike placebo implants without TAF, all active implants resulted in local adverse events (AEs) proximal to the implant ranging in severity from mild to moderate and included dermal inflammation and necrosis across all species. Despite these AEs, the implant performed as designed and achieved a constant drug release profile, supporting the continued development of this drug delivery platform.

Safety and efficacy of a biodegradable implant releasing tenofovir alafenamide for vaginal protection in a macaque model.

OBJECTIVES: To advance the initiative of ending the global epidemic, long-lasting HIV protection is needed through sustained release of antiretroviral drugs for months to years. We investigated in macaques the safety and efficacy of biodegradable polycaprolactone implants releasing tenofovir alafenamide for HIV pre-exposure prophylaxis (PrEP). METHODS: Implants were administered subcutaneously in the arm using a contraceptive trocar. Efficacy against vaginal simian-HIV (SHIV) infection was investigated in six pigtailed macaques that received two tenofovir alafenamide implants (0.35 mg/day), one in each arm, for a total release rate of tenofovir alafenamide at 0.7 mg/day. Macaques were exposed to SHIV twice weekly for 6 weeks. Statistical analyses were used to compare outcome with eight untreated controls. Histological assessments were performed on skin biopsies collected near implantation sites. RESULTS: Median (range) tenofovir diphosphate level in PBMCs was 1519 (1068-1898) fmol/106 cells. All macaques with tenofovir alafenamide implants were protected against vaginal SHIV infection. In contrast, 7/8 controls were infected after a median of 4 SHIV exposures (P = 0.0047). Histological assessment of tissues near tenofovir alafenamide implant sites showed inflammation and necrosis in 5/6 animals, which were not evident by visual inspection. CONCLUSIONS: We demonstrated complete protection against vaginal SHIV infection with two implants releasing a total of 0.7 mg of tenofovir alafenamide per day. We also identified tenofovir diphosphate concentrations in PBMCs associated with complete vaginal protection. Consistent with previous findings, we observed adverse local toxicity and necrosis near the tenofovir alafenamide implant site. Improved tenofovir alafenamide implants that are safe and maintain high efficacy have the potential to provide long-lasting protection against vaginal HIV infection.

Brillouin light scattering in niobium doped lead zirconate single crystal.

Brillouin light scattering experiments were performed for lead zirconate single crystals doped with niobium. Special attention was paid to the elastic mode softening near phase transition temperatures. The results are compared with data obtained by Raman light scattering experiments. We observed that the interaction between acoustic and optic modes is responsible for symmetry breaking far above TC, leading to polar regions' appearance. No changes in the acoustic mode frequency and its damping are observed at TC, where ε(T) exhibits a maximum value. The absence of these changes and the central peak observed in Raman experiments suggest that the phase transition at TC is mainly of the order-disorder type. The origin of other phase transitions is discussed as well.

Magnetic field driven phase transitions in EuTiO3.

The influence of an external static magnetic field (up to 480 mT) on the structural properties of EuTiO3(ETO) polycrystalline samples was examined by powder XRD at the Elettra synchrotron facilities in the temperature range 100-300 K. While the cubic to tetragonal structural phase transition temperature in this magnetic field range remains almost unaffected, significant lattice effects appear at two characteristic temperatures (∼200 K and ∼250 K), which become more pronounced at a critical threshold field. At ∼200  K a change in the sign of magnetostriction is detected attributed to a modification of the local magnetic properties from intrinsic ferromagnetism to intrinsic antiferromagnetism. These data are a clear indication that strong spin-lattice interactions govern also the high temperature phase of ETO and trigger the appearance of magnetic domain formation and phase transitions.

Wildland Fire Emission Sampling at Fishlake National Forest, Utah Using an Unmanned Aircraft System.

Emissions from a stand replacement prescribed burn were sampled using an unmanned aircraft system (UAS, or "drone") in Fishlake National Forest, Utah, U.S.A. Sixteen flights over three days in June 2019 provided emission factors for a broad range of compounds including carbon monoxide (CO), carbon dioxide (CO2), nitric oxide (NO), nitrogen oxide (NO2), particulate matter < 2.5 microns in diameter (PM2.5), volatile organic compounds (VOCs) including carbonyls, black carbon, and elemental/organic carbon. To our knowledge, this is the first UAS-based emission sampling for a fire of this magnitude, including both slash pile and crown fires resulting in wildfire-like conditions. The burns consisted of drip torch ignitions as well as ground-mobile and aerial helicopter ignitions of large stands comprising over 1,000 ha, allowing for comparison of same-species emission factors burned under different conditions. The use of a UAS for emission sampling minimizes risk to personnel and equipment, allowing flexibility in sampling location and ensuring capture of representative, fresh smoke constituents. PM2.5 emission factors varied 5-fold and, like most pollutants, varied inversely with combustion efficiency resulting in lower emission factors from the slash piles than the crown fires.

Methodology for characterizing emissions from small (0.5-2 MTD) batch-fed gasification systems using multiple waste compositions.

A compact, containerized gasification system was characterized for air emissions while burning four waste types. A methodology is presented for developing a standardized test waste composition and demonstrated using three military and one civilian waste types. Batch charges of waste were processed through a gasification chamber, afterburner, and wet scrubber. The 0.5-2 metric ton per day (MTD) system was designed for mobile deployment by the military in forward operations but would be applicable to small scale civilian applications. Emissions data from these types of small capacity, cyclically operated systems are lacking, limiting efforts to compare technologies and their environmental performance. Eight tests were conducted in a 7-day period at the Kilauea Military Camp (KMC) in Hawaii. The pollutants characterized were chosen based on their regulatory and health relevance: particulate matter (PM), mercury (Hg), elemental composition, volatile organic compounds (VOCs), polyaromatic hydrocarbons (PAHs), and polychlorinated dibenzo-p-dioxins (PCDDs) and polychlorinated dibenzofurans (PCDFs). Averaged data from 4-hour runs, including startups and shutdowns, indicated that five of the nine EPA-regulated compounds (lead, cadmium, Hg, sulfur dioxide, and hydrogen chloride) were under the emission limits set for Other Solid Waste Incineration Units (OSWI) while four, PCDD/PCDF, PM, nitrogen oxides, and carbon monoxide, were higher. The procedures through which waste compositions were created and emissions were characterized provide a methodology by which differing waste to energy technologies can be compared on an equivalent basis. This system's emissions compare favorably with alternative disposal methods. PM and PCDD/PCDF emission factors were, respectively, over 39 and 9 times lower from this unit than from published data on burning simulated military waste in an air curtain incinerator and in open burn piles ("burn pits").

Progression-dependent transport heterogeneity of breast cancer liver metastases as a factor in therapeutic resistance.

Metastatic disease is a major cause of mortality in cancer patients. While many drug delivery strategies for anticancer therapeutics have been developed in preclinical studies of primary tumors, the drug delivery properties of metastatic tumors have not been sufficiently investigated. Therapeutic efficacy hinges on efficient drug permeation into the tumor microenvironment, which is known to be heterogeneous thus potentially making drug permeation heterogeneous, also. In this study, we have identified that 4 T1 liver metastases, treated with pegylated liposomal doxorubicin, have unfavorable and heterogeneous transport of doxorubicin. Our drug extravasation results differ greatly from analogous studies with 4 T1 tumors growing in the primary site. A probabilistic tumor population model was developed to estimate drug permeation efficiency and drug kinetics of liver metastases by integrating the transport and structural properties of tumors and delivered drugs. The results demonstrate significant heterogeneity in metastases with regard to transport properties of doxorubicin within the same animal model, and even within the same organ. These results also suggest that the degree of heterogeneity depends on the stage of tumor progression and that differences in transport properties can define transport-based tumor phenotypes. These findings may have valuable clinical implications by illustrating that therapeutic agents can permeate and eliminate metastases of "less resistant" transport phenotypes, while sparing tumors with more "resistant" transport properties. We anticipate that these results could challenge the current paradigm of drug delivery into metastases, highlight potential caveats for therapies that may alter tumor perfusion, and deepen our understanding of the emergence of drug transport-based therapeutic resistance.

Control of collagen gel mechanical properties through manipulation of gelation conditions near the sol-gel transition.

The ability to control the mechanical properties of cell culture environments is known to influence cell morphology, motility, invasion and differentiation. The present work shows that it is possible to control the mechanical properties of collagen gels by manipulating gelation conditions near the sol gel transition. This manipulation is accomplished by performing gelation in two stages at different temperatures. The mechanical properties of the gel are found to be strongly dependent on the duration and temperature of the first stage. In the second stage the system is quickly depleted of free collagen which self assembles into a highly branched network characteristic of gelation at the higher temperature (37 °C). An important aspect of the present work is the use of advanced rheometric techniques to assess the transition point between viscoelastic liquid and viscoelastic solid behaviour which occurs upon establishment of a sample spanning network at the gel point. The gel time at the stage I temperature is found to indicate the minimum time that the gelling collagen sample must spend under stage I conditions before the two stage gelation procedure generates an enhancement of mechanical properties. Further, the Fractional Maxwell Model is found to provide an excellent description of the time-dependent mechanical properties of the mature collagen gels.

The Canine POMC Gene, Obesity in Labrador Retrievers and Susceptibility to Diabetes Mellitus.

BACKGROUND: Diabetes mellitus (DM) in dogs is a common endocrinopathy with a complex genetic architecture. Disease susceptibility in several breeds is associated with polymorphisms in immune response genes, but in the Labrador retriever breed, no genetic associations with DM have been identified. A deletion in the pro-opiomelanocortin (POMC) gene in Labrador retrievers is associated with increased appetite and risk of obesity. HYPOTHESIS/OBJECTIVES: To characterize the POMC deletion in Labrador retrievers, to develop a simple genetic test for this mutation, and to test the hypothesis that the POMC gene deletion is associated with an increased risk of DM in this breed. ANIMALS: Sixty-one non-diabetic Labrador retrievers aged >6 years and 57 Labrador retrievers with DM. METHODS: Case-control genotyping study to compare the frequency of the POMC deletion in dogs with and without DM. After polymerase chain reaction (PCR) and sequencing to characterize the mutation, a PCR-based test was developed and validated using 2 different restriction fragment length polymorphism assays. RESULTS: A 14-base-pair deletion was confirmed and localized to exon 3 of the canine POMC gene. A PCR-based test for the deletion was successfully developed. There was no association between the presence of the POMC deletion mutation and DM in this population of Labrador retriever dogs (P = .31). CONCLUSIONS AND CLINICAL IMPORTANCE: This study adds to the existing scientific literature indicating that there is little evidence for a direct link between obesity and DM in dogs.

Emissions from prescribed burning of agricultural fields in the Pacific Northwest.

Prescribed burns of winter wheat stubble and Kentucky bluegrass fields in northern Idaho and eastern Washington states (U.S.A.) were sampled using ground-, aerostat-, airplane-, and laboratory-based measurement platforms to determine emission factors, compare methods, and provide a current and comprehensive set of emissions data for air quality models, climate models, and emission inventories. Batch measurements of PM2.5, volatile organic compounds (VOCs), polycyclic aromatic hydrocarbons (PAHs), and polychlorinated dibenzodioxins/dibenzofurans (PCDDs/PCDFs), and continuous measurements of black carbon (BC), particle mass by size, CO, CO2, CH4, and aerosol characteristics were taken at ground level, on an aerostat-lofted instrument package, and from an airplane. Biomass samples gathered from the field were burned in a laboratory combustion facility for comparison with these ground and aerial field measurements. Emission factors for PM2.5, organic carbon (OC), CH4, and CO measured in the field study platforms were typically higher than those measured in the laboratory combustion facility. Field data for Kentucky bluegrass suggest that biomass residue loading is directly proportional to the PM2.5 emission factor; no such relationship was found with the limited wheat data. CO2 and BC emissions were higher in laboratory burn tests than in the field, reflecting greater carbon oxidation and flaming combustion conditions. These distinctions between field and laboratory results can be explained by measurements of the modified combustion efficiency (MCE). Higher MCEs were recorded in the laboratory burns than from the airplane platform. These MCE/emission factor trends are supported by 1-2 min grab samples from the ground and aerostat platforms. Emission factors measured here are similar to other studies measuring comparable fuels, pollutants, and combustion conditions. The size distribution of refractory BC (rBC) was single modal with a log-normal shape, which was consistent among fuel types when normalized by total rBC mass. The field and laboratory measurements of the Angstrom exponent (α) and single scattering albedo (ω) exhibit a strong decreasing trend with increasing MCEs in the range of 0.9-0.99. Field measurements of α and ω were consistently higher than laboratory burns, which is likely due to less complete combustion. When VOC emissions are compared with MCE, the results are consistent for both fuel types: emission factors increase as MCE decreases.

Hidden magnetism in the paramagnetic phase of EuTiO3.

EuTiO3 is investigated experimentally by temperature dependent conductivity and dielectric constant measurements. Both data sets evidence a crossover behavior in their temperature dependence around T * ~ 200 K indicating a change of the electrical and magnetic behavior of EuTiO3 as already observed by muon spin rotation (µSR) measurements. Around T ' = 80 K an additional anomaly appears which is consistent with previous resonant ultrasound spectroscopy (RUS) data. By applying a magnetic field (1.2 T at room temperature) the bulk conductivity is anomalously enhanced by more than one order of magnitude. The bulk dielectric constant ε(r) decreases with increasing temperature and is enhanced by the magnetic field by up to 22%. All data reveal a substantial hysteresis which is diminished in a magnetic field. The unusual magnetic field dependence of all quantities is suggested to stem from magnetically active domain walls which are mobile between the structural phase transition temperature (T(S) = 282 K) and T * and are pinned below it.

Evaluation of the Sherlock 3CG Tip Confirmation System on peripherally inserted central catheter malposition rates.

Peripherally inserted central catheters are often positioned blindly in the central circulation, and this may result in high malposition rates, especially in critically ill patients. Recently, a new technology has been introduced (Sherlock 3CG Tip Positioning System) that uses an electro-magnetic system to guide positioning in the superior vena cava, and then intra-cavity ECG to guide positioning at the cavo-atrial junction. In this observational study, we investigated how the Sherlock 3CG Tip Positioning System would affect peripherally inserted central catheter malposition rates, defined using a post-insertion chest radiograph, in critically ill patients. A total of 239 catheters positioned using the Sherlock 3CG Tip Positioning System were analysed. When an adequate position was defined as low superior vena cava or cavo-atrial junction, 134 catheters (56.1%; 95% CI 50-62%) were malpositioned. When an adequate position was defined as mid/low superior vena cava, cavo-atrial junction or high right atrium (≤ 2 cm from cavo-atrial junction), 49 (20.5%; 95% CI 16-26%) catheters were malpositioned. These malposition rates are significantly lower than our own historical data, which used a 'blind' anthropometric technique to guide peripherally inserted central catheter insertion.

Recombinant canine IgE Fc and an IgE Fc-TRAIL fusion protein bind to neoplastic canine mast cells.

Screening for expression of the high affinity receptor for IgE by reverse transcriptase PCR, revealed that almost all canine mast cell tumors expressed FcɛRIα mRNA, supporting the rationale for developing anti-neoplastic treatments based on molecules that could target this receptor. Use of cytotoxic cytokines to trigger an apoptotic signal is one strategy for inducing cell death in malignant mast cells. The coding sequences for canine IgE and tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) were identified through genome analyses. Selected regions of the coding sequences for these genes were cloned and compared to the predicted genome sequences. The Fc region of canine IgE, death domain of canine TRAIL and an IgE Fc: TRAIL fusion construct were generated and epitope-tagged proteins expressed, using a eukaryotic expression system. Specific binding of recombinant canine IgE Fc-containing proteins to recombinant human FcɛRIα and to a canine mast cell tumor line expressing FcɛRIα (C2), but not one failing to express FcɛRIα (MCLA), was demonstrated. Specific binding of the IgE: TRAIL fusion protein was not abrogated by the TRAIL moiety. These results are proof of principle that canine IgE targeting to FcɛRIα can be used as a platform for selective delivery of therapies to FcɛRIα-expressing cells, potentially enhancing their therapeutic index and efficacy.

Lattice and polarizability mediated spin activity in EuTiO3.

EuTiO3 is shown to exhibit novel strong spin-charge-lattice coupling deep in the paramagnetic phase. Its existence is evidenced by an, until now, unknown response of the paramagnetic susceptibility at temperatures exceeding the structural phase transition temperature T(S) = 282 K. The 'extra' features in the susceptibility follow the rotational soft zone boundary mode temperature dependence above and below TS. The theoretical modeling consistently reproduces this behavior and provides reasoning for the stabilization of the soft optic mode other than quantum fluctuations.

A polymorphism in the melanocortin 4 receptor gene (MC4R:c.92C>T) is associated with diabetes mellitus in overweight domestic shorthaired cats.

BACKGROUND: Feline diabetes mellitus (DM) shares many pathophysiologic features with human type 2 DM. Human genome-wide association studies have identified genes associated with obesity and DM, including melanocortin 4 receptor (MC4R), which plays an important role in energy balance and appetite regulation. HYPOTHESIS/OBJECTIVES: To identify single nucleotide polymorphisms (SNPs) in the feline MC4R gene and to determine whether any SNPs are associated with DM or overweight body condition in cats. ANIMALS: Two-hundred forty domestic shorthaired (DSH) cats were recruited for the study. Of these, 120 diabetics were selected (60 overweight, 60 lean), along with 120 nondiabetic controls (60 overweight and 60 lean). Males and females were equally represented. METHODS: A prospective case-control study was performed. Genomic DNA was extracted from blood samples and used as template for PCR amplification of the feline MC4R gene. The coding region of the gene was sequenced in 10 cats to identify polymorphisms. Subsequently, genotyping by restriction fragment length polymorphism (RFLP) analysis assessed MC4R:c.92C > T allele and genotype frequencies in each group of cats. RESULTS: No significant differences in MC4R:c.92C>T allele or genotype frequencies were identified between nondiabetic overweight and lean cats. In the overweight diabetic group, 55% were homozygous for the MC4R:c.92C allele, compared to 33% of the lean diabetics and 30% of the nondiabetics. The differences between the overweight diabetic and the nondiabetics were significant (P < .01). CONCLUSIONS AND CLINICAL IMPORTANCE: We identified a polymorphism in the coding sequence of feline MC4R that is associated with DM in overweight DSH cats, similar to the situation in humans.

Nonlinear pressure dependence of TN in almost multiferroic EuTiO3.

The antiferromagnetic (AFM) phase transition temperature TN of EuTiO3 has been studied as a function of pressure p. The data reveal a nonlinear dependence of TN on p with TN increasing with increasing pressure. The exchange interactions exhibit an analogous dependence on p as TN (if the absolute value of the nearest neighbor interaction is considered) and there is evidence that the AFM transition is robust with increasing pressure. The corresponding Weiss temperature ΘW remains anomalous since it always exhibits positive values. The data are analyzed within the Bloch power law model and provide excellent agreement with experiment.

Magnetic field enhanced structural instability in EuTiO3.

EuTiO(3) undergoes a structural phase transition from cubic to tetragonal at T(S) = 282 K which is not accompanied by any long range magnetic order. However, it is related to the oxygen octahedral rotation driven by a zone boundary acoustic mode softening. Here, we show that this displacive second order structural phase transition can be shifted to higher temperatures by the application of an external magnetic field (ΔT(S) âˆ¼ 4 K for µ(0)H = 9 T). This observed field dependence is in agreement with theoretical predictions based on a coupled spin-anharmonic-phonon interaction model.

Novel diabetes mellitus treatment: mature canine insulin production by canine striated muscle through gene therapy.

Muscle-targeted gene therapy using insulin genes has the potential to provide an inexpensive, low maintenance alternative or adjunctive treatment method for canine diabetes mellitus. A canine skeletal muscle cell line was established through primary culture, as well as through transdifferentiation of canine fibroblasts after infection with a myo-differentiation gene containing adenovirus vector. A novel mutant furin-cleavable canine preproinsulin gene insert (cppI4) was designed and created through de novo gene synthesis. Various cell lines, including the generated canine muscle cell line, were transfected with nonviral plasmids containing cppI4. Insulin and desmin immunostaining were used to prove insulin production by muscle cells and specific canine insulin ELISA to prove mature insulin secretion into the medium. The canine myoblast cultures proved positive on desmin immunostaining. All cells tolerated transfection with cppI4-containing plasmid, and double immunostaining for insulin and desmin proved present in the canine cells. Canine insulin ELISA assessment of medium of cppI4-transfected murine myoblasts and canine myoblast and fibroblast mixture proved presence of mature fully processed canine insulin, 24 and 48 h after transfection. The present study provides proof of principle that canine muscle cells can be induced to produce and secrete canine insulin on transfection with nonviral plasmid DNA containing a novel mutant canine preproinsulin gene that produces furin-cleavable canine preproinsulin. This technology could be developed to provide an alternative canine diabetes mellitus treatment option or to provide a constant source for background insulin, as well as C-peptide, alongside current treatment options.

An engineered Plasmodium falciparum C-terminal 19-kilodalton merozoite surface protein 1 vaccine candidate induces high levels of interferon-gamma production associated with cellular immune responses to specific peptide sequences in Gambian adults naturally exposed to malaria.

The 19-kDa C-terminal region of merozoite surface protein 1 (MSP1(19)), a major blood stage malaria vaccine candidate, is the target of cellular and humoral immune responses in humans naturally infected with Plasmodium falciparum. We have previously described engineered variants of this protein, designed to be better vaccine candidates, but the human immune response to these proteins has not been characterized fully. Here we have investigated the antigenicity of one such variant compared to wild-type MSP1(19)-derived protein and peptides. Gambian adults produced both high T helper type 1 (Th1) [interferon (IFN)-γ] and Th0/Th2 [interleukin (IL)-13 and sCD30] responses to the wild-type MSP1(19) and the modified protein as wells as to peptides derived from both forms. Response to the modified MSP1(19) (with three amino acid substitutions: Glu27Tyr, Leu31Arg and Glu43Leu) relative to the wild-type, included higher IFN-γ production. Interestingly, some peptides evoked different patterns of cytokine responses. Modified peptides induced higher IL-13 production than the wild-type, while the conserved peptides P16 and P19 induced the highest IFN-γ and IL-13 and/or sCD30 release, respectively. We identified P16 as the immunodominant peptide that was recognized by cells from 63% of the study population, and not restricted to any particular human leucocyte antigen D-related (HLA-DR) type. These findings provide new and very useful information for future vaccine development and formulation as well as potential Th1/Th2 immunmodulation using either wild-type or modified protein in combination with their peptides.