Transdermal Delivery of a Hydrogen Sulphide Donor, ADT-OH Using Aqueous Gel Formulations for the Treatment of Impaired Vascular Function: an Ex Vivo Study.

PURPOSE: Hydrogen sulphide (H2S) is an important signalling molecule involved in the regulation of several physiological and pathophysiological processes. The objective of this study was to investigate the feasibility of transdermal delivery of ADT-OH, a H2S donor, by investigating the transdermal flux of aqueous gels loaded with penetration enhancers or liposomes. Furthermore, we explored the ability of permeated ADT-OH to promote angiogenesis and mitochondrial bioenergetics in HUVEC cells. METHODS: Aqueous hypromellose gels (5% w/v) were prepared with up to 10% v/v propylene glycol (PG) or deformable liposomes with 0.025% w/w ADT-OH. ADT-OH permeation from formulations across excised murine skin into PBS was quantified over 24 h using HPLC-UV detection. Media was collected and applied to HUVEC cells to evidence ADT-OH functionality following permeation. Tube formation assays were performed as indicative of angiogenesis and mitochondrial oxygen consumption was evaluated using a Seahorse XF24. RESULTS: Increasing the loading of PG caused an increase in ADT-OH permeation rate across skin and a decrease in dermal drug retention whereas liposomal gels produced a slow-release profile. Treatment of HUVEC's using conditioned media collected from the ADT-OH loaded permeation studies enhanced tube formation and the basal oxygen consumption rates after 30 min of treatment. CONCLUSIONS: These findings demonstrate that transdermal delivery of ADT-OH may provide a promising approach in the treatment of impaired vascular function. Gels prepared with 10% v/v PG have the potential for use in conditions requiring rapid H2S release whereas liposomal loaded gels for treatment requiring sustained H2S release.

Phytoestrogens Modulate Breast Cancer Resistance Protein Expression and Function at the Blood-Cerebrospinal Fluid Barrier.

PURPOSE: Breast cancer resistance protein (BCRP/ABCG2) is a drug efflux transporter expressed at the blood cerebrospinal fluid barrier (BCSFB), and influences distribution of drugs into the central nervous systems (CNS). Current inhibitors have failed clinically due to neurotoxicity. Novel approaches are needed to identify new modulators to enhance CNS delivery. This study examines 18 compounds (mainly phytoestrogens) as modulators of the expression/function of BCRP in an in vitro rat choroid plexus BCSFB model. METHODS: Modulators were initially subject to cytotoxicity (MTT) assessment to determine optimal non-toxic concentrations. Reverse-transcriptase PCR and confocal microscopy were used to identify the presence of BCRP in Z310 cells. Thereafter modulation of the intracellular accumulation of the fluorescent BCRP probe substrate Hoechst 33342 (H33342), changes in protein expression of BCRP (western blotting) and the functional activity of BCRP (membrane insert model) were assessed under modulator exposure. RESULTS: A 24 hour cytotoxicity assay (0.001 µM-1000 µM) demonstrated the majority of modulators possessed a cellular viability IC50 > 148 µM. Intracellular accumulation of H33342 was significantly increased in the presence of the known BCRP inhibitor Ko143 and, following a 24 hour pre-incubation, all modulators demonstrated statistically significant increases in H33342 accumulation (P < 0.001), when compared to control and Ko143. After a 24 hour pre-incubation with modulators alone, a 0.16-2.5 -fold change in BCRP expression was observed for test compounds. The functional consequences of this were confirmed in a permeable insert model of the BCSFB which demonstrated that 17-ß-estradiol, naringin and silymarin (down-regulators) and baicalin (up-regulator) can modulate BCRP-mediated transport function at the BCSFB. CONCLUSION: We have successfully confirmed the gene and protein expression of BCRP in Z310 cells and demonstrated the potential for phytoestrogen modulators to influence the functionality of BCRP at the BCSFB and thereby potentially allowing manipulation of CNS drug disposition.