Noise-induced hearing loss - An examination of the methods of assessment in a cross-sectional study of 87 industrial workers.

OBJECTIVES: The surveillance of noise-induced hearing loss (NIHL) according to the Health and Safety Executive (HSE) differs from the medico-legal criteria used to assess NIHL. Our study compares the two systems and proposes a novel method of simplifying the medico-legal criteria and applying it to ascertain noise-induced hearing loss. DESIGN: The anonymised audiograms of a group of 87 industrial workers from a single site were analysed with both methods. RESULTS: The comparison showed approximately one-third of the workers assessed in this study had their noise-induced hearing loss underestimated by the HSE criteria. The majority of these individuals were over 40 years of age. CONCLUSIONS: The HSE criteria for noise-induced hearing loss need review and re-alignment with the medico-legal criteria to address the discrepancy between the two systems.

Site-specific, sustained release of drugs to the brain.

A dihydropyridine-pyridinium salt type of redox system is used in a general and flexible method for the site-specific or sustained delivery (or both) of drugs to the brain. A biologically active compound linked to a lipoidal dihydropyridine carrier easily penetrates the blood-brain barrier. Oxidation of the carrier part in vivo to the ionic pyridinium salt prevents its elimination from the brain, while elimination from the general circulation is accelerated. Subsequent cleavage of the quaternary carrier-drug species results in sustained delivery of the drug in the brain and facile elimination of the carrier part.

Usefulness of a novel Caco-2 cell perfusion system II. Characterization of monolayer properties and peptidase activity.

In this study, the enzymatic activity and the influence of support filters and extracellular matrix proteins on the differentiation of Caco-2 cells grown in a perfusion system (Minucells and MinutissueTM) were examined and compared to traditional culturing approaches. Differences were observed regarding the differentiation of Caco-2 cells using the traditional approach and perfusion system such that the cell monolayers grown in a perfusion system showed a significant increase in dipeptidase activities (18.20 +/- 0.43nmol x min(-1) x cm(-2)) compared to the cells cultivated using the 21-day protocol (9.45 +/- 0.50 nmol x min(-1) x cm(-2)). The peptidase activity of Caco-2 cells was strikingly inhibited when Matrigel extracellular protein was used for coating polycarbonate support filters. While the enzymatic activities of the cell monolayers differentiated in the perfusion system were up-regulated, the transepithelial electrical resistance values of the cell monolayers (171 +/- 52 and 251 +/- 62 omega x cm2 for polycarbonate and polyester, respectively) decreased compared to the traditional Snapwell inserts (644 +/- 119 omega x cm2). The results suggested that the perfusion systems were useful permeability models which reduce workload, resources and manpower needed to obtain useful Caco-2 monolayers. In addition, the approach offers an efficient tool for long-term culturing of highly differentiated Caco-2 cell monolayers.

Supersaturating drug delivery systems: effect of hydrophilic cyclodextrins and other excipients on the formation and stabilization of supersaturated drug solutions.

Supersaturating drug delivery systems (SDDS) utilize two important design elements in their preparation including converting the drug of interest into a high energy state or other rapidly dissolving form to facilitate the formation of supersaturated drug solutions and providing a means for stabilizing the formed supersaturated solution such that significant drug absorption is possible from the gastrointestinal tract. This has been referred to as a "spring" and "parachute" approach. The current effort is designed to assess materials which may affect properties in SDDS. To this end, a series of excipients was tested in a co-solvent/solvent quench method to assess their ability to attain and maintain supersaturation for a group of 14 drug development candidates. The approach focussed on hydrophilic cyclodextrins including hydroxypropyl-beta-cyclodextrin (HPbetaCD) and sulfobutyl-beta-cyclodextrin (SBEbetaCD). Various rheological polymers and surfactants were also included in the study. Consistent with previous investigations, the pharmaceutical polymers, as a class, had minimal effects on the extent of supersaturation but tended to be good stabilizers while the surfactants tended to provide for the greatest degree of supersaturation but the formed systems were poorly stable. This study found that hydrophilic cyclodextrins, especially SBEbetaCD, gave superior results in terms of attaining and maintaining supersaturation. A knowledge of the behavior and performance of excipients in this context can be useful in designing solid oral dosage forms for difficult-to-formulate drugs and drug candidates.

Parallel artificial membrane permeability assay (PAMPA) combined with a 10-day multiscreen Caco-2 cell culture as a tool for assessing new drug candidates.

The parallel artificial membrane permeability assay (PAMPA) is extensively used for the evaluation of early drug candidates. It is high throughput, low cost and is amenable to automation. This method has been shown useful in assessing transmembrane, non-energy dependent, diffusion of drugs such that reasonable predictability with in vivo (passive) absorption is possible. Cell cultures mimicking the gastrointestinal tract such as the CACO-2 cultures have the advantage of taking into account other transport mechanism including paracellular and carrier-mediated uptake but are lower throughput and labor-intensive. In this study, the applicability of two high throughput permeability assays namely PAMPA (PSR4p, pION Inc.) and 96-well Caco-2 cell assay (MultiScreen, Millipore) were used to rank drug permeability as well as to predict passive and active drug absorption/secretion for a series of marketed drugs as well as a collection of structurally diverse drug candidates. CACO-2 cells were cultured using MultiScreen hardware over a period of 10 days with the integrity of the cells assessed using transepithelial electrical resistance (TEER) and by the ability of the monolayer to the transport a paracellular marker, sodium fluorescence. Effective permeability (Peff) data were calculated using spectrophotometric data and were binned based on a pre-defined cut-off values as either highly and poorly permeable. A comparison of a well characterized drug training set indicate at least 85% concordance between the data generated from PAMPA and Caco-2 MultiScreen. The values obtained using the MultiScreen approach were also similar to data obtained from the literature using the conventional 21-day Caco-2 cell assay. Differences between PAMPA and CACO-2 ranking were useful indicators of either drug efflux (PAMPA (Peff) > CACO-2 (Peff)) or absorptive transport (CACO-2 (Peff) > PAMPA (Peff)). These results indicate that PAMPA combined with the MultiScreen Caco-2 cell culture may be a useful high throughput screening for predicting passive diffusion and active transport of new drugs.

Spontaneously self-assembled micelles from poly(ethylene glycol)-b-poly(epsilon-caprolactone-co-trimethylene carbonate) for drug solubilization.

Di-block copolymers composed of polyethylene glycol (PEG) and a second block of (co)polyesters of epsilon-caprolactone (CL) and/or trimethylene carbonate (TMC) were synthesized and characterized. Tin octoate was used as catalyst and polymerization were completed over a period of 24 h with high conversion (> 95%). Self-assembling properties in water were evaluated. All di-block copolymers behave similarly except when PCL served as the second block. Stable crew-cut micelles of about 20 nm were obtained by direct dissolution of the liquid di-block copolymers in water at room temperature. When PCL was present as the second block, no solubilization occurred. Drug encapsulation of poorly water-soluble drugs belonging to biopharmaceutics classification system (BCS) class II (ketoprofen and furosemide) was evaluated. Experimental solubility for these two drugs shows a significant enhancement such that a maximum value of 23.4 mg/ml was obtained for ketoprofen in a 10% w/v micellar solution as compared to 0.14 mg in water. In the case of furosemide, the solubility increased from 0.04 mg/ml in water to about 3.2 mg/ml in a 10% w/v micellar solution. Enzymatic degradation of diblock copolymers was also studied in the presence of Pseudomonas lipase in a phosphate buffer solution (pH 7.4). Results indicated rapid degradation of copolymers containing relatively higher amounts of CL compared to TMC suggesting the potential in vivo degradation.

Delivery of a DNAzyme targeting c-myc to HT29 colon carcinoma cells using a gold nanoparticulate approach.

The objective of the current study was to develop cellular delivery approaches for catalytic DNA enzymes (DNAzymes) which cleave targeted messenger RNA, using vectors based on colloidal gold. The model DNAzyme was a 32mer oligonucleotide designed to specifically interact with and cleave c-myc mRNA. Colloidal gold particles were prepared by reduction of tetrachlororauric [III] acid with sodium citrate. Particles could be produced in the 1-90 nm range. A cationic substrate linked to transferrin was electrostatically/hydrophobically bound to the gold particle. These vectors were then treated with the DNAzyme to yield the condensed DNA-cationic polymer-particulate product. The pH (4-11.5), the quantity of the DNAzymes (0.079-0.567 microg/probe), the cationic polymer (polylysine (PL) or polyethylenimine (PEI)) as well as the surfactant (PVP) concentration (0-0.5%) were varied to give stable constructs which decomplexed under the desired conditions (i.e., in lysosomes and at lower pH values). Cellular uptake of the FITC-labelled c-myc DNAzyme incorporated in this vector was measured using FACS analysis in human HT29 colon carcinoma cells. Data suggested that PEI gave better delivery efficiencies than PL. The use of PVP to stabilize the formed dispersions was detrimental to DNAzyme delivery when PL was used but had little effect in the PEI systems. In the best cases, delivery to 77% of the cells was possible using PEI with the PVP stabilizer and completing the DNA condensation at pH 5.5 with 0.118 microg of DNAzyme/probe. In contrast, the best conditions for PL gave only transfection to 43% of the cells (no PVP, condensed at pH 5.7 and with a loading of 0.079 microg DNAzyme/probe). The PL probe tended to be more toxic than the PEI-based systems (65% cell death in PL transfected cells compared to 22% for PEI). These results suggest that cellular targeting using colloidal gold appears feasible for DNAzyme delivery.

Delayed diagnosis of synovial sarcoma of the foot.

Soft tissue sarcomas are relatively rare in the population, rarely encountered by the average orthopedic surgeon, and may only be seen once in a lifetime by a general practitioner. Although rare, it is a serious condition that, if diagnosed early, can result in improved prognosis. This article presents a case of a delayed diagnosis of a soft tissue sarcoma and highlights possible pitfalls and causes for such delays in hope of educating and reducing such incidences in the future.

Characterization of self-assembling copolymers in aqueous solutions using Electron Paramagnetic Resonance and Fluorescence spectroscopy.

Electron Paramagnetic Resonance and fluorescence spectroscopy have been used to determine the micropolarity and microviscosity of self-assembling systems based on mmePEG-p(CL-co-TMC) having different PEG chain lengths and different CL/TMC ratios and PEG/MOG/SA (45/5/50) polymers with different PEG chain lengths. Four reporter probes have been used: two spin probes, 16-doxyl stearic acid and 5-doxylstearic acid, and two fluorescent probes, pyrene and 1,3-bis(1-pyrenyl) propane (P3P). We found that the micelles based on mmePEG-p(CL-co-TMC) polymers are of a biphasic nature. The micelles are made of a hydrophilic corona with low viscosity while the core of the micelle is more hydrophobic and more viscous. The outer shell is made up of PEG chains, the hydrophobic part of the chains making the core. The partial hydration of the shell seems to lead to a looser chain network than that associated with deeper domains in the micelles. By contrast, in micelles composed of PEG/MOG/SA, there is no clear domain separation. This is consistent with a spatial configuration of random polymeric chains forming a loose network. In these micelles, the microviscosity is low and the hydrophobicity is high.

Characterization of physico-chemical properties and pharmaceutical performance of sucrose co-freeze-dried solid nanoparticulate powders of the anti-HIV agent loviride prepared by media milling.

In order to improve the dissolution and absorption properties of loviride, a poorly soluble antiviral agent, sucrose co-freeze-dried nanopowders were prepared, characterized and evaluated. Tween 80/poloxamer 188-stabilized nanosuspensions were produced on a laboratory scale using media milling. The milling process was monitored by dynamic light scattering (DLS) and resulted in particles with a mean size of 264+/-14nm and a distribution width of 59+/-6nm after 4h of milling. Co-freeze-drying of the nanosuspensions with sucrose had an inhibiting effect on nanoparticle agglomeration and yielded solid "nanopowders" that were resuspendable and homogeneous with respect to loviride content. X-ray powder diffraction (XRPD) confirmed the presence of small loviride crystallites and indicated that sucrose and poloxamer 188 were crystalline. Differential scanning calorimetry (DSC) showed melting peaks of poloxamer 188, sucrose and loviride. Time-resolved XRPD indicated that sucrose crystallization was complete within 24h of storage. Scanning electron microscopy (SEM) suggested the formation of sheet-like matrix structures. The dissolution rate of loviride from the nanopowders was excellent. A Caco-2 experiment on the nanopowder showed a significantly higher cumulative amount transported after 120min (1.59+/-0.02microg) compared to the physical mixture (0.93+/-0.01microg) and the untreated loviride (0.74+/-0.03mcirog).

Prediction of drug solubility in amphiphilic di-block copolymer micelles: the role of polymer-drug compatibility.

The goal of the current study was to assess the value of predictive computational approaches for estimating drug solubility in hydrated micelles formed from di-block copolymers of polyethylene glycol (PEG) and random copolyesters of epsilon-caprolactone (CL) and trimethylene carbonate (TMC) using drug-polymer compatibility as assessed through the Flory-Huggins interaction parameter (chi). In order to accomplish this, the compatibility of several well-known model drugs (associated with the four biopharmaceutics classification system (BCS) classes) was assessed with both segments of the amphiphilic di-block copolymer PEG-b-P(CL-co-TMC). Compatibilities were estimated based on the Hansen modification of the Hildebrand approach using Molecular Modeling Pro software. Experimental solubilities for model drugs were determined using a shake-flask technique at various polymer concentrations. The solubilities of 8 compounds in 10% w/v micelle solutions were in relatively good agreement with the predicted drug-polymer compatibility. In addition, the approach allows for the selection of a suitable di-block copolymer for optimal solubilization of a specific drug. Furosemide was assessed as a model with results suggesting that it can be best entrapped in a di-block copolyester containing a relatively high CL content. The data suggests that prediction of drug solubilization of block copolymer-based micelles may be facilitated by assessing the compatibility of the drug for the component polymeric domains.

A survey on IVIVC/IVIVR development in the pharmaceutical industry - Past experience and current perspectives.

The present work aimed to describe the current status of IVIVC/IVIVR development in the pharmaceutical industry, focusing on the use and perception of specific approaches as well as successful and failed case studies. Two questionnaires have been distributed to 13 EFPIA partners of the Oral Biopharmaceutics Tools Initiative and to the Pharmacokinetics Working Party of the European Medicines Agency in order to capture the perspectives and experiences of industry scientists and agency members, respectively. Responses from ten companies and three European Agencies were received between May 21st 2014 and January 19th 2016. The majority of the companies acknowledged the importance of IVIVC/IVIVR throughout the drug development stages and a well-balanced rate of return on investment. However, the IVIVC/IVIVR approach seemed to be underutilized in regulatory submissions. Four of the ten companies stated to have an internal guidance related to IVIVC/IVIVR modelling, whereas three felt that an overall strategy is not necessary. Successful models mainly served to support formulation development and to provide a better mechanistic understanding. There was not yet much experience with safe-space IVIVRs as well as the use of physiologically based modelling in the field of IVIVC. At the same time, the responses from both industry and agencies indicated that there might be a need for a regulatory framework to guide the application of these novel approaches. The relevance of IVIVC/IVIVR for oral IR drug products was recognized by most of the companies. For IR formulations, relationships other than Level A correlation were more common outcomes among the provided case studies, such as multiple Level C correlation or safe-space IVIVR, which could be successfully used for requesting regulatory flexibility. Compared to the responses from industry scientists, there was a trend towards a higher appreciation of the BCS among the regulators, but a less positive attitude towards the utility of non-compendial dissolution methods for establishing a successful IVIVC/IVIVR. The lack of appropriate in vivo data and regulatory uncertainty were considered the major difficulties in IVIVC/IVIVR development. The results of this survey provide unique insights into current IVIVC/IVIVR practices in the pharmaceutical industry. Pursuing an IVIVC/IVIVR should be generally encouraged, considering its high value from both industry and regulators' perspective.

Intestinal uptake and biodistribution of novel polymeric micelles after oral administration.

To determine the fate of polymeric micelles after oral administration, we investigated the possible transport of polymeric micelles across Caco-2 monolayers and their biodistribution in rats after per os administration of [14C]-labelled mmePEG750P(CL-co-TMC) micelles containing risperidone (BCS Class II drug). mmePEG750P(CL-co-TMC) was able to cross Caco-2 monolayer via a saturable transport mechanism. The oral bioavailability of the polymer was 40%. Polymeric micelles based on mmePEG750P(CL-co-TMC) showed very low clearance by the reticuloendothelial system (RES) and a renal excretion. A sustained release of risperidone was observed.

Encapsulation of amphotericin B in poly(ethylene glycol)-block-poly(epsilon-caprolactone-co-trimethylenecarbonate) polymeric micelles.

The aim of this work was to evaluate the potential of self-assembling poly(ethyleneglycol)(750)-block-poly(epsilon-caprolactone-co-trimethylenecarbonate)(4500) 50/50 copolymers (PEG-p(CL-co-TMC)) to solubilize amphotericin B in polymeric micelles and to disaggregate the drug to the less toxic monomeric form. Amphotericin B was encapsulated in the micelles upon dilution of a mixture of the liquid polymer and the drug in water. Its solubility was increased by two orders of magnitude depending on polymer concentration. The aggregation state of amphotericin B was decreased by PEG-p(CL-co-TMC). The preparation method and the loading of the polymeric micelles influenced it. The antifungal activity of the drug was reduced by encapsulation in the polymeric micelles whereas the onset of amphotericin B-induced hemolysis was delayed. PEG-p(CL-co-TMC) micelles could be an easy method for amphotericin B encapsulation.

A permanently charged tamoxifen derivative displays anticancer activity and improved tissue selectivity in rodents.

A quaternized form of tamoxifen (TAM), tamoxifen methiodide (TMI), was shown to demonstrate very low brain uptake compared to TAM and, unexpectedly, was considerably less estrogenic than TAM in the uterus. The agonist activity of TMI in the bone was similar to that of TAM. TMI manifested significant dose-dependent tumoricidal activity with a rapid onset of action against MCF-7 human breast cancer implants in nude mice and a mean reduction in tumor size of 60% over six weeks.

Transfer of phosphatidylcholine facilitated by a component of human plasma.

A constituent of lipoprotein-free (p greater than 1.21) human plasma from normolipemic donors facilitates the transfer of diacyl phosphatidylcholine from unilamellar egg yolk phosphatidylcholine liposomes to liver mitochondria. The active component is heat labile, has a hydrated density greater than 1.25 and an apparent molecular weight of more than 100 000. The presence of this protein in plasma may facilitate movement of diacylphospholipids between the surfaces of lipid-containing particles such as lipoproteins and erythrocytes. Knowledge of the properties and behavior of this protein are important in designing methods of drug therapy based on encapsulation in biodegradable lipid vesicles.

The use of human nasal in vitro cell systems during drug discovery and development.

The nasal route is widely used for the administration of drugs for both topical and systemic action. At an early stage in drug discovery and during the development process, it is essential to gain a thorough insight of the nasal absorption potential, metabolism and toxicity of the active compound and the components of the drug formulation. Human nasal epithelial cell cultures may provide a reliable screening tool for pharmaco-toxicological assessment of potential nasal drug formulations. The aim of this review is to give an overview of the information relevant for the development of a human nasal epithelial cell culture model useful during drug discovery and development. A primary goal in the development of in vitro cell culture systems is to maintain differentiated morphology and biochemical features, resembling the original tissue as closely as possible. The potential and limitations of the existing in vitro human nasal models are summarized. The following topics related to cell culture methodology are discussed: (i) primary cultures versus cell lines; (ii) cell-support substrate; (iii) medium and medium supplements; and (iv) the air-liquid interface model versus liquid-liquid. Several considerations with respect to the use of in vitro systems for pharmaceutical applications (transport, metabolism, assessment of ciliary toxicity) are also discussed.

Stable ciliary activity in human nasal epithelial cells grown in a perfusion system.

PURPOSE: Explore the usefulness of a perfusion system in order to establish human nasal epithelial cell cultures suitable for long-term in vitro ciliary beat frequency (CBF) and cilio-toxicity studies. METHODS: The cells were obtained by protease digestion of nasal biopsy material. The cells were plated at a density of 0.8-1 x 10(6)/cm2 on Vitrogen-coated polyethylene terephthalate membranes, and cultured under submerged conditions in a CO2 incubator or in a perfusion system (initiated on days 8-9 after plating). The CBF was determined at 24.1 +/- 0.8 degrees C by a computerized microscope photometry system. The morphology of the cultured cells was characterized by transmission electron microscopy (TEM). RESULTS: Under CO2 incubator culture conditions, stable ciliary activity was expressed and maintained from day 2 to day 24. Under perfusion system culture conditions, the CBF (mean+/-S.D., n = 4) amounted to 8.4 +/- 0.9 and 8.8 +/- 0.4 Hz on days 7 and 14, respectively. These values were lower as compared to the corresponding CBF obtained in the CO2 incubator cultures (9.5 +/- 0.6 and 9.9 +/- 1.0 Hz, respectively). Reference cilio-stimulatory (glycocholate) and cilio-inhibitory (chlorocresol) compounds were used to assess CBF reactivity. In the CO2 incubator and 7- and 14-days perfusion system cultures, glycocholate (0.5%) showed a reversible cilio-stimulatory effect of 23, 26 and 21%, respectively, while chlorocresol (0.005%) exerted a reversible cilio-inhibitory effect of 36, 40 and 36%, respectively. TEM revealed polarized cuboidal to columnar epithelial morphology, with well-differentiated ciliated cells under CO2 and perfusion system conditions (up to day 23). CONCLUSION: Culturing human nasal epithelial cells on Vitrogen-coated polyethylene terephthalate membranes in submerged conditions in a CO2 incubator and in a perfusion system offers the possibility for long-term preservation (up to 22-24 days) of stable and reactive CBF in vitro.

Comparison of distortion product otoacoustic emissions and pure tone audiometry in occupational screening for auditory deficit due to noise exposure.

OBJECTIVE: To examine whether distortion product otoacoustic emissions can serve as a replacement for pure tone audiometry in longitudinal screening for occupational noise exposure related auditory deficit. METHODS: A retrospective review was conducted of pure tone audiometry and distortion product otoacoustic emission data obtained sequentially during mandatory screening of brickyard workers (n = 16). Individual pure tone audiometry thresholds were compared with distortion product otoacoustic emission amplitudes, and a correlation of these measurements was conducted. RESULTS: Pure tone audiometry threshold elevation was identified in 13 out of 16 workers. When distortion product otoacoustic emission amplitudes were compared with pure tone audiometry thresholds at matched frequencies, no evidence of a robust relationship was apparent. Seven out of 16 workers had substantial distortion product otoacoustic emissions with elevated pure tone audiometry thresholds. CONCLUSION: No clinically relevant predictive relationship between distortion product otoacoustic emission amplitude and pure tone audiometry threshold was apparent. These results do not support the replacement of pure tone audiometry with distortion product otoacoustic emissions in screening. Distortion product otoacoustic emissions at frequencies associated with elevated pure tone audiometry thresholds are evidence of intact outer hair cell function, suggesting that sites distinct from these contribute to auditory deficit following ototrauma.

Downstream processing of polymer-based amorphous solid dispersions to generate tablet formulations.

Application of amorphous solid dispersions (ASDs) is considered one of the most promising approaches to increase the dissolution rate and extent of bioavailability of poorly water soluble drugs. Such intervention is often required for new drug candidates in that enablement, bioavailability is not sufficient to generate a useful product. Importantly, tableting of ASDs is often complicated by a number of pharmaceutical and technological challenges including poor flowability and compressibility of the powders, compression-induced phase changes or phase separation and slow disintegration due to the formation of a gelling polymer network (GPN). The design principles of an ASD-based system include its ability to generate supersaturated systems of the drug of interest during dissolution. These metastable solutions can be prone to precipitation and crystallization reducing the biopharmaceutical performance of the dosage form. The main aim of the research in this area is to maintain the supersaturated state and optimally enhance bioavailability, meaning that crystallization should be delayed or inhibited during dissolution, as well as in solid phase (e.g., during manufacturing and storage). Based on the expanding use of ASD technology as well as their downstream processing, there is an acute need to summarize the results achieved to this point to better understand progress and future risks. The aim of this review is to focus on the conversion of ASDs into tablets highlighting results from various viewpoints.