In vitro phototoxicity of 5-aminolevulinic acid and its methyl ester and the influence of barrier properties on their release from a bioadhesive patch.

opical administration of excess exogenous 5-aminolevulinic acid (ALA) leads to selective accumulation of the potent photosensitiser protoporphyrin IX (PpIX) in neoplastic cells, which can then be destroyed by irradiation with visible light. Due to its hydrophilicity, ALA penetrates deep lesions, such as nodular basal cell carcinomas (BCCs) poorly. As a result, more lipophilic esters of ALA have been employed to improve tissue penetration. In this study, the in vitro release of ALA and M-ALA from proprietary creams and novel patch-based systems across normal stratum corneum and a model membrane designed to mimic the abnormal stratum corneum overlying neoplastic skin lesions were investigated. Receiver compartment drug concentrations were compared with the concentrations of each drug producing high levels of PpIX production and subsequent light-induced kill in a model neoplastic cell line (LOX). LOX cells were found to be quite resistant to ALA- and M-ALA-induced phototoxicity. However, drug concentrations achieved in receiver compartments were comparable to those required to induce high levels of cell death upon irradiation in cell lines reported in the literature. Patches released significantly less drug across normal stratum corneum and significantly more across the model membrane. This is of major significance since the selectivity of PDT for neoplastic lesions will be further enhanced by the delivery system. ALA/M-ALA will only be delivered in significant amounts to the abnormal tissue. PpIX will only then accumulate in the neoplastic cells and the normal surrounding tissue will be unharmed upon irradiation.

A two compartment in vitro release model for the testing of HIV microbicide formulations

BACKGROUND: In vitro release testing of vaginal formulations is usually performed in a one-compartment model (OCM) where the release medium, usually comprising pH-adjusted water, an aqueous surfactant solution or a solvent-water solution, provides sink conditions throughout the release experiment. Although this model is useful in evaluating the effect of formulation parameters upon release, it rarely reflects in vivo conditions. Here we report use of a two-compartment diffusion cell model (TCM, comprising a small volume donor, a large volume receptor, and separated by a model epithelial membrane) to more closely mimic in vivo vaginal release and tissue absorption following administration of a UC781 vaginal ring.

METHODS: Macaque-sized matrix silicone elastomer vaginal rings containing 100mg UC781 were prepared by injection molding, and in vitro release testing performed using both OCM (20mL simulated vaginal fluid, SVF) and TCM (5mL SVF in donor cell and variable quantities of Tween 80; silicone elastomer membrane; 100mL 3:2 ethanol/water in receptor cell). In the TCM, drug levels were measured by HPLC in both donor and receptor cells, representing fluid and tissue levels respectively. Rings containing 100mg UC781 and 10% w/w Tween 80 were also manufactured and tested.

RESULTS: The amount of UC781 released from rings was significantly influenced by the choice of release model. Greatest release (56mg/14 days) was measured in the ethanol/water OCM, compared with no measurable release into SVF only. Increasing the concentration of Tween 80 in the SVF medium (1, 3 and 5% w/w) led to increased UC781 release (11, 16 and 18mg, respectively), demonstrating that vaginal fluid solubility of UC781 may be rate-determining in vivo. In the TCM, UC781 accumulates in the receptor cell medium over time, despite not being measured in the donor medium containing the ring device. Incorporation of Tween 80 directly into the ring provided enhanced release in both donor and receptor cells.

CONCLUSIONS: Release of UC781 was influenced by the choice of release medium and the inclusion of Tween 80 in the ring. Although use of SVF-only in the OCM indicated no measurable UC781 release from rings, data from the TCM confirms that UC781 is not only released but is also capable of penetrating across the model epithelial membrane. The TCM may therefore provide a more representative in vitro release model for mimicking in vivo absorption.

Rheological, mechanical and membrane penetration properties of novel dual drug systems for percutaneous delivery

In this study it has been demonstrated that mixtures of two solid drugs, ibuprofen and methyl nicotinate, with different but complementary pharmacological activities and which exist as a single liquid phase over a wide composition range at skin temperature, can be formulated as o/w emulsions without the use of an additional hydrophobic carrier. These novel dual drug systems provided significantly enhanced in vitro penetration rates through a model lipophilic barrier membrane compared to conventional individual formulations of each active. Thus, for ibuprofen, drug penetration flux enhancements of three- and 10-fold were observed when compared to an aqueous ibuprofen suspension and a commercial alcohol-based ibuprofen formulation, respectively. Methyl nicotinate penetration rates were shown to be similar for aqueous gels and emulsified systems. Mechanisms explaining these observations are proposed. Novel dual drug formulations of ibuprofen and methyl nicotinate, formulated within the liquid range at skin temperature, were investigated by oscillatory rheology and texture profile analysis. demonstrating the effects of drug and viscosity enhancer concentrations, and disperse phase type upon the rheological, mechanical and drug penetration properties of these systems. (C) 2000 Elsevier Science B.V. All rights reserved.

Physicochemical and drug diffusion analysis of rifampicin containing polyethylene glycol-poly(epsilon-caprolactone) networks designed for medical device applications

This study reports the physicochemical and drug diffusion properties of rifampicin containing poly(epsilon-caprolactone) (PCL)/polyethylene glycol (PEG) networks, designed as bioactive biomaterials. Uniquely, the effects of the states of both rifampicin and PEG and the interplay between these components on these properties are described. PCL matrices containing rifampicin (1-5%, w/w) and PEG 200 (0-15%, w/w) were prepared by casting from an organic solvent (dichloromethane). The films were subsequently characterized in terms of their thermal/thermorheological, surface and tensile properties, biodegradation and drug diffusion/release properties. Incorporation of PEG and/or rifampicin significantly affected the tensile and surface properties of PCL, lowering the ultimate tensile strength, % elongation at break, Young modulus and storage and loss moduli. Both in the absence and presence of PEG, solubilisation of rifampicin within the crystalline domains of PCL was observed. PEG was present as a dispersed liquid phase. The release of rifampicin (3% loading) was unaffected by the presence of PEG. Similarly the release of rifampicin (5%) was unaffected by low concentrations of PEG (5-10%) however, at higher loadings, the release rate of rifampicin was enhanced by the presence of PEG. Rifampicin release (10% loading) was enhanced by the presence of PEG in a concentration dependent fashion. These observations were accredited to enhanced porosity of the matrix. In all cases, diffusion-controlled release of rifampicin occurred which was unaffected by polymer degradation. This study has uniquely illustrated the effect of hydrophilic pore formers on the physicochemical properties of PCL. Interestingly, enhanced diffusion controlled release was only observed from biomaterials containing high loadings of PEG and rifampicin (5, 10%), concentrations that were shown to affect the mechanical properties of the biomaterials. Care should therefore be shown when adopting this strategy to enhance release of bioactive agents from biomaterials. (C) 2011 Elsevier B.V. All rights reserved.

Chlorhexidine release from poly(epsilon-caprolactone) films prepared by solvent evaporation

The effect of selected formulation variables on the release of chlorhexidine from poly(epsilon-caprolactone) films was evaluated in vitro using a complete factorial experimental design. Repeated measures analysis of variance showed chlorhexidine type (diacetate or base), drug load (10, 20 or 30% w/w), chlorhexidine particle size (

A modified SILCS contraceptive diaphragm for long-term controlled release of the HIV microbicide dapivirine

Background: There is considerable interest in developing new multipurpose prevention technologies to address women's reproductive health needs. This study describes an innovative barrier contraceptive device--based on the SILCS diaphragm--that also provides long-term controlled release of the lead candidate anti-HIV microbicide dapivirine.

Study design: Diaphragm devices comprising various dapivirine-loaded polymer spring cores overmolded with a nonmedicated silicone elastomer sheath were fabricated by injection molding processes. In vitro release testing, thermal analysis and mechanical characterization were performed on the devices.

Results: A diaphragm device containing a polyoxymethylene spring core loaded with 10% w/w dapivirine provided continuous and controlled release of dapivirine over a 6-month period, with a mean in vitro daily release rate of 174 mcg/day. The mechanical properties of the new diaphragm were closely matched to the SILCS diaphragm.

Conclusions: The study demonstrates proof of concept for a dapivirine-releasing diaphragm with daily release quantities potentially capable of preventing HIV transmission. In discontinuous clinical use, release of dapivirine may be readily extended over 1 or more years. © 2013 Elsevier Inc. All rights reserved.

α-Adrenergic agonism enhances the growth hormone (GH) response to GH-releasing hormone through an inhibition of hypothalamic somatostatin release in normal men

The purpose of this study was to investigate the precise mechanism by which central a-adrenergic pathways modulate GH secretion in humans. In 10 normal subjects we compared the pattern of clonidine-induced GH release to that elicited by GH-releasing hormone (GHRH) given at a time of presumably similar responsiveness of the somatotrope. We also evaluated the effect of stimulation by GHRH (either endogenous, by administration of clonidine, or exogenous) on the GH response to a further exogenous GHRH stimulation. In 2 experiments the administration of clonidine (0.150 mg, orally) at 0 or 60 min was followed by a GHRH [GRF-(1-29); 1 µg/kg, iv] challenge at 180 min. In other experiments subjects received on separate occasions placebo or clonidine at 0 min, followed by GHRH at 60 min and again at 180 min. In a further experiment the administration of clonidine at 0 min was followed by 2 GHRH challenges (60 and 180 min later). The administration of clonidine 60 or 120 min, but not 180 min, before the GHRH bolus significantly (P <0.01) increased the GH responses to this challenge compared to those elicited by GHRH when given after placebo in a period of a similar somatotrope responsiveness. These, in turn, were significantly (P <0.05) higher than those elicited by clonidine alone. The close relationship between pre-GHRH plasma GH values and GHRH-elicited GH peaks, not observed for clonidine, was lost after pretreatment with this drug. These data indicate that clonidine was able to disrupt the intrinsic hypothalamic-somatotroph rhythm, suggesting that a-adrenergic pathways have a major inhibitory effect on somatostatin release. Our data also indicate that GH responses to a GHRH bolus administered 120 min after a prior GHRH challenge are dependent on two parameters: the intrinsic hypothalamic-somatotroph rhythm at the time of the second GHRH bolus, and the magnitude of GH secretion elicited by the previous somatotroph stimulation. In summary, a-adrenergic agonism appears to act primarily in GH control by inhibiting the hypothalamic release of somatostatin, rather than by stimulating GHRH secretion.

Modified Silicone Elastomer Vaginal Gels for Sustained Release of Antiretroviral HIV Microbicides

We previously reported nonaqueous silicone elastomer gels (SEGs) for sustained vaginal administration of the CCR5-targeted entry inhibitor maraviroc (MVC). Here, we describe chemically modified SEGs (h-SEGs) in which the hydrophobic cyclomethicone component was partially replaced with relatively hydrophilic silanol-terminated polydimethylsiloxanes (st-PDMS). MVC and emtricitabine (a nucleoside reverse transcriptase inhibitor), both currently under evaluation as topical microbicides to counter sexual transmission of human immunodeficiency virus type 1 (HIV-1), were used as model antiretroviral (ARV) drugs. Gel viscosity and in vitro ARV release were significantly influenced by st-PDMS molecular weight and concentration in the h-SEGs. Unexpectedly, gels prepared with lower molecular weight grades of st-PDMS showed higher viscosities. h-SEGs provided enhanced release over 24 h compared with aqueous hydroxyethylcellulose (HEC) gels, did not modify the pH of simulated vaginal fluid (SVF), and were shown to less cytotoxic than standard HEC vaginal gel. ARV solubility increased as st-PDMS molecular weight decreased (i.e., as percentage hydroxyl content increased), helping to explain the in vitro release trends. Dye ingression and SVF dilution studies confirmed the increased hydrophilicity of the h-SEGs. h-SEGs have potential for use in vaginal drug delivery, particularly for ARV-based HIV-1 microbicides.

Respiratory Infections Cause the Release of Extracellular Vesicles: Implications in Exacerbation of Asthma/COPD

Background: Infection-related exacerbations of respiratory diseases are a major health concern; thus understanding the mechanisms driving them is of paramount importance. Despite distinct inflammatory profiles and pathological differences, asthma and COPD share a common clinical facet: raised airway ATP levels. Furthermore, evidence is growing to suggest that infective agents can cause the release of extracellular vesicle (EVs) in vitro and in bodily fluids. ATP can evoke the P2X7/caspase 1 dependent release of IL-1β/IL-18 from EVs; these cytokines are associated with neutrophilia and are increased during exacerbations. Thus we hypothesized that respiratory infections causes the release of EVs in the airway and that the raised ATP levels, present in respiratory disease, triggers the release of IL-1β/IL-18, neutrophilia and subsequent disease exacerbations.

Methods: To begin to test this hypothesis we utilised human cell-based assays, ex vivo murine BALF, in vivo pre-clinical models and human samples to test this hypothesis.

Results: Data showed that in a murine model of COPD, known to have increased airway ATP levels, infective challenge causes exacerbated inflammation. Using cell-based systems, murine models and samples collected from challenged healthy subjects, we showed that infection can trigger the release of EVs. When exposed to ATP the EVs release IL-1b/IL-18 via a P2X₇/caspase-dependent mechanism. Furthermore ATP challenge can cause a P2X₇ dependent increase in LPS-driven neutrophilia.

Conclusions: This preliminary data suggests a possible mechanism for how infections could exacerbate respiratory diseases and may highlight a possible signalling pathway for drug discovery efforts in this area.