Caffeic acid phenethyl ester is protective in experimental ulcerative colitis via reduction in levels of pro-inflammatory mediators and enhancement of epithelial barrier function.

BACKGROUND: Inhibition of the nuclear factor kappa beta (NF-κß) pathway has been proposed as a therapeutic target due to its key role in the expression of pro-inflammatory genes, including pro-inflammatory cytokines, chemokines, and adhesion molecules. Caffeic acid phenethyl ester (CAPE) is a naturally occurring anti-inflammatory agent, found in propolis, and has been reported as a specific inhibitor of NF-κß. However, the impact of CAPE on levels of myeloperoxidases (MPO) and pro-inflammatory cytokines during inflammation is not clear. The aims of this study were to investigate the protective efficacy of CAPE in the mouse model of colitis and determine its effect on MPO activity, pro-inflammatory cytokines levels, and intestinal permeability. METHOD: Dextran sulphate sodium was administered in drinking water to induce colitis in C57/BL6 mice before treatment with intraperitoneal administration of CAPE (30 mg kg-1 day-1). Disease activity index (DAI) score, colon length and tissue histology levels of MPO, pro-inflammatory cytokines, and intestinal permeability were observed. RESULTS: CAPE-treated mice had lower DAI and tissue inflammation scores, with improved epithelial barrier protection and significant reduction in the level of MPO and pro-inflammatory cytokines. CONCLUSION: Our results show that CAPE is effective in suppressing inflammation-triggered MPO activity and pro-inflammatory cytokines production while enhancing epithelial barrier function in experimental colitis. Thus, we conclude that CAPE could be a potential therapeutic agent for further clinical investigations for treatment of inflammatory bowel diseases in humans.

Effect of poly(ethylene glycol) content and formulation parameters on particulate properties and intraperitoneal delivery of insulin from PLGA nanoparticles prepared using the double-emulsion evaporation procedure.

CONTEXT: Size, encapsulation efficiency and stability affect the sustained release from nanoparticles containing protein-type drugs. OBJECTIVES: Insulin was used to evaluate effects of formulation parameters on minimizing diameter, maximizing encapsulation efficiency and preserving blood glucose control following intraperitoneal (IP) administration. METHODS: Homogenization or sonication was used to incorporate insulin into poly(D,L-lactic-co-glycolic acid) (PLGA) nanoparticles with increasing poly(ethylene glycol) (PEG) content. Effects of polymer type, insulin/polymer loading ratio and stabilizer in the internal aqueous phase on physicochemical characteristics of NP, in vitro release and stability of encapsulated insulin were investigated. Entrapment efficiency and release were assessed by radioimmunoassay and bicinconnic acid protein assay, and stability was evaluated using SDS-PAGE. Bioactivity of insulin was assessed in streptozotocin-induced, insulin-deficient Type I diabetic mice. RESULTS: Increasing polymeric PEG increased encapsulation efficiency, while the absence of internal stabilizer improved encapsulation and minimized burst release kinetics. Homogenization was shown to be superior to sonication, with NP fabricated from 10% PEG-PLGA having higher insulin encapsulation, lower burst release and better stability. Insulin-loaded NP maintained normoglycaemia for 24 h in diabetic mice following a single bolus, with no evidence of hypoglycemia. CONCLUSIONS: Insulin-loaded NP prepared from 10% PEG-PLGA possessed therapeutically useful encapsulation and release kinetics when delivered by the IP route.

A non-inferiority randomized controlled trial comparing the clinical effectiveness of anesthesia obtained by application of a novel topical anesthetic putty with the infiltration of lidocaine for the treatment of lacerations in the emergency department.

STUDY OBJECTIVE: We test the hypothesis that anesthesia, measured as pain scores, induced by a novel topical anesthetic putty is non-inferior (margin=1.3) to that provided by conventional lidocaine infiltration for the repair of lacerations. METHODS: A randomized controlled trial was conducted in the emergency department (ED) of a local hospital. Participants were randomly allocated to receive either infiltration anesthesia or topical anesthetic putty as per the trial protocol. Pain scores were recorded 15 minutes after infiltration and 30 minutes after topical anesthetic putty application. Median pain scores were compared between groups. Wound evaluation scores were conducted after 7 to 10 days and adverse events were monitored for both groups of participants throughout the study. RESULTS: One hundred and ten participants were enrolled in the study, with 56 receiving infiltration and 54 receiving topical anesthetic putty. The median difference between the pain scores of the 2 groups was 0 (95% confidence interval -1 to 0). There were no substantial differences between the 2 groups in terms of either the wound evaluation scores or the incidence of adverse events. CONCLUSION: The novel topical anesthetic putty was not inferior to infiltration with lidocaine with respect to the pain experienced during suturing, and this putty is a feasible alternative to infiltration anesthesia of lacerations in the ED.

Fast-drying multi-laminate bioadhesive films for transdermal and topical drug delivery.

No bioadhesive patch-based system is currently marketed. This is despite an extensive number of literature reports on such systems detailing their advantages over conventional pressure sensitive adhesive-based patches in wet environments and describing successful delivery of a diverse array of drug substances. This lack of proprietary bioadhesive patches is largely due to the fact that such systems are exclusively water-based, meaning drying is difficult. In this paper we describe, for the first time, a novel multiple lamination method for production of bioadhesive patches. In contrast to patches produced using a conventional casting approach, which took 48 hours to dry, bioadhesive films prepared using the novel multiple lamination method were dried in 15 min and were folded into formed patches in a further 10 min. Patches prepared by both methods had comparable physicochemical properties. The multiple lamination method allowed supersaturation of 5-aminolevulinic acid to be achieved in formed patch matrices. However, drug release studies were unable to show an advantage for supersaturation with this particular drug, due to its water high solubility. The multiple lamination method allowed greater than 90% of incorporated nicotine to remain within formed patches, in contrast to the 48% achieved for patches prepared using a conventional casting approach. The procedure described here could readily be adapted for automation by industry. Due to the reduced time, energy and ensuing finance now required, this could lead to bioadhesive patch-based drug delivery systems becoming commercially viable. This would, in turn, mean that pathological conditions occurring in wet or moist areas of the body could now be routinely treated by prolonged site-specific drug delivery, as mediated by a commercially produced bioadhesive patch.

Furanone loaded aerogels are effective antibiofilm therapeutics in a model of chronic Pseudomonas aeruginosa wound infection.

Almost 80% of chronic wounds have a bacterial biofilm present. These wound biofilms are caused by a range of organisms and are often polymicrobial. Pseudomonas aeruginosa is one of the most common causative organisms in wound infections and readily forms biofilms in wounds. To coordinate this, P. aeruginosa uses a process known as quorum sensing. Structural homologues of the quorum sensing signalling molecules have been used to disrupt this communication and prevent biofilm formation by Pseudomonas. However, these compounds have not yet reached clinical use. Here, we report the production and characterisation of a lyophilised PVA aerogel for use in delivering furanones to wound biofilms. PVA aerogels successfully release a model antimicrobial and two naturally occurring furanones in an aqueous environment. Furanone loaded aerogels inhibited biofilm formation in P. aeruginosa by up to 98.80%. Further, furanone loaded aerogels successfully reduced total biomass of preformed biofilms. Treatment with a sotolon loaded aerogel yielded a 5.16 log reduction in viable biofilm bound cells in a novel model of chronic wound biofilm, equivalent to the current wound therapy Aquacel AG. These results highlight the potential utility of aerogels in drug delivery to infected wounds and supports the use of biofilm inhibitory compounds as wound therapeutics.

Gene Therapy for Neuropsychiatric Disorders: Potential Targets and Tools.

Neuropsychiatric disorders that affect the central nervous system cause considerable pressures on the health care system and have a substantial economic burden on modern societies. The present treatments based on available drugs are mostly ineffective and often costly. The molecular process of neuropsychiatric disorders is closely connected to modifying the genetic structures inherited or caused by damage, toxic chemicals, and some current diseases. Gene therapy is presently an experimental concept for neurological disorders. Clinical applications endeavor to alleviate the symptoms, reduce disease progression, and repair defective genes. Implementing gene therapy in inherited and acquired neurological illnesses entails the integration of several scientific disciplines, including virology, neurology, neurosurgery, molecular genetics, and immunology. Genetic manipulation has the power to minimize or cure illness by inducing genetic alterations at endogenous loci. Gene therapy that involves treating the disease by deleting, silencing, or editing defective genes and delivering genetic material to produce therapeutic molecules has excellent potential as a novel approach for treating neuropsychiatric disorders. With the recent advances in gene selection and vector design quality in targeted treatments, gene therapy could be an effective approach. This review article will investigate and report the newest and the most critical molecules and factors in neuropsychiatric disorder gene therapy. Different genome editing techniques available will be evaluated, and the review will highlight preclinical research of genome editing for neuropsychiatric disorders while also evaluating current limitations and potential strategies to overcome genome editing advancements.

Importance of STAT3 signalling in cancer, metastasis and therapeutic interventions.

The Signal Transducer and Activator of Transcription 3 (STAT3) protein is encoded on chromosome 17q21. The SH2 and the DNA binding domains are critical structural components of the protein, together with tyrosine and serine residues that initiate phosphorylation. STAT3 interacts with DNA directly and functions in cells as both a signal transducer and a transcription factor. Its cytoplasmic activation results in dimerisation and nuclear translocation, where it is involved in the transcription of a large number of target genes. STAT3 is hyperactive in cancer cells as a result of upstream STAT3 mutations or enhanced cytokine production in the tumour environment. The STAT3 signalling pathway promotes many hallmarks of carcinogenesis and metastasis, including enhanced cell proliferation and survival, as well as migration and invasion into the extracellular matrix. Recent investigations into novel STAT3-based therapies describe a range of innovative approaches, such as the use of novel oligonucleotide drugs. These limit STAT3 binding to its target genes by attaching to SH2 and DNA-binding domains. Yet, despite these significant steps in understanding the underpinning mechanisms, there are currently no therapeutic agents that addresses STAT3 signalling in a clinically relevant manner.

Crocin Inhibits Angiogenesis and Metastasis in Colon Cancer via TNF-α/NF-kB/VEGF Pathways.

Angiogenesis and metastasis play pivotal roles in the progression of cancer. We recently discovered that crocin, a dietary carotenoid derived from the Himalayan crocus, inhibited the growth of colon cancer cells. However, the exact role of crocin on the angiogenesis and metastasis in colorectal cancer remains unclear. In the present study, we demonstrated that crocin significantly reduces the viability of colon cancer cells (HT-29, Caco-2) and human umbilical vein endothelial cells (HUVEC), but was not toxic to human colon epithelial (HCEC) cells. Furthermore, pre-treatment of human carcinoma cells (HT-29 and Caco-2) with crocin inhibited cell migration, invasion, and angiogenesis in concentration -dependent manner. Further studies demonstrated that crocin inhibited TNF-α, NF-κB and VEGF pathways in colon carcinoma cell angiogenesis and metastasis. Crocin also inhibited cell migration, invasion, and tube formation in human umbilical vein endothelial cells (HUVEC) in a concentration -dependent manner. We also observed that crocin significantly reduced the secretion of VEGF and TNF-α induced activation of NF-kB by human colon carcinoma cells. In the absence of TNF-α, a concentration-dependent reduction in NF-kB was observed. Many of these observations were confirmed by in vivo angiogenesis models, which showed that crocin significantly reduced the progression of tumour growth. Collectively, these finding suggest that crocin inhibits angiogenesis and colorectal cancer cell metastasis by targeting NF-kB and blocking TNF-α/NF-κB/VEGF pathways.

Enhanced ROS production and cell death through combined photo- and sono-activation of conventional photosensitising drugs.

A combination of light and ultrasound activation of two conventional photosensitising drugs, methylene blue and rose bengal, was shown to generate higher levels of reactive oxygen species (ROS) and lower LD50 values than either light or ultrasound activation alone.

Furanone quorum-sensing inhibitors with potential as novel therapeutics against Pseudomonas aeruginosa.

Micro-organisms use quorum sensing (QS), a cell density-dependent process, to communicate. This QS mode of interchange leads to the production of a variety of virulence factors, co-ordination of complex bacterial behaviours, such as swarming motility, degradation of host tissue and biofilm formation. QS is implicated in numerous human infections and consequently researchers have sought ways of effectively inhibiting the process in pathogenic bacteria. Two decades ago, furanones were the first class of chemical compounds identified as Pseudomonas aeruginosa QS inhibitors (QSIs). P. aeruginosa is a ubiquitous organism, capable of causing a wide range of infections in humans, including eye and ear infections, wound infections and potentially fatal bacteraemia and thus novel treatments against this organism are greatly needed. This review provides a brief background on QS and the use of furanones as QSIs. Based on the effectiveness of action, both in vivo and in vitro, we will explore the use of furanones as potential antimicrobial therapeutics and conclude with open questions.

The secondary bile acids, ursodeoxycholic acid and lithocholic acid, protect against intestinal inflammation by inhibition of epithelial apoptosis.

Increased epithelial permeability is a key feature of IBD pathogenesis and it has been proposed that agents which promote barrier function may be of therapeutic benefit. We have previously reported the secondary bile acid, ursodeoxycholic acid (UDCA), to be protective in a mouse model of colonic inflammation and that its bacterial metabolism is required for its beneficial effects. The current study aimed to compare the effects of UDCA, LCA, and a non-metabolizable analog of UDCA, 6-methyl-UDCA (6-MUDCA), on colonic barrier function and mucosal inflammation in a mouse model of colonic inflammation. Bile acids were administered daily to C57Bl6 mice by intraperitoneal injection. Colonic inflammation, induced by addition of DSS (2.5%) to the drinking water, was measured as disease activity index (DAI) and histological score. Epithelial permeability and apoptosis were assessed by measuring FITC-dextran uptake and caspase-3 cleavage, respectively. Cecal bile acids were measured by HPLC-MS/MS. UDCA and LCA, but not 6-MUDCA, were protective against DSS-induced increases in epithelial permeability and colonic inflammation. Furthermore, UDCA and LCA inhibited colonic epithelial caspase-3 cleavage both in DSS-treated mice and in an in vitro model of cytokine-induced epithelial injury. HPLC-MS/MS analysis revealed UDCA administration to increase colonic LCA levels, whereas LCA administration did not alter UDCA levels. UDCA, and its primary metabolite, LCA, protect against intestinal inflammation in vivo, at least in part, by inhibition of epithelial apoptosis and promotion of barrier function. These data suggest that clinical trials of UDCA in IBD patients are warranted.

Correction: Frankincense essential oil suppresses melanoma cancer through down regulation of Bcl-2/Bax cascade signaling and ameliorates heptotoxicity via phase I and II drug metabolizing enzymes.

[This corrects the article DOI: 10.18632/oncotarget.26930.].

Anti-bacterial activity of inorganic nanomaterials and their antimicrobial peptide conjugates against resistant and non-resistant pathogens.

This review details the antimicrobial applications of inorganic nanomaterials of mostly metallic form, and the augmentation of activity by surface conjugation of peptide ligands. The review is subdivided into three main sections, of which the first describes the antimicrobial activity of inorganic nanomaterials against gram-positive, gram-negative and multidrug-resistant bacterial strains. The second section highlights the range of antimicrobial peptides and the drug resistance strategies employed by bacterial species to counter lethality. The final part discusses the role of antimicrobial peptide-decorated inorganic nanomaterials in the fight against bacterial strains that show resistance. General strategies for the preparation of antimicrobial peptides and their conjugation to nanomaterials are discussed, emphasizing the use of elemental and metallic oxide nanomaterials. Importantly, the permeation of antimicrobial peptides through the bacterial membrane is shown to aid the delivery of nanomaterials into bacterial cells. By judicious use of targeting ligands, the nanomaterial becomes able to differentiate between bacterial and mammalian cells and, thus, reduce side effects. Moreover, peptide conjugation to the surface of a nanomaterial will alter surface chemistry in ways that lead to reduction in toxicity and improvements in biocompatibility.

Dietary Crocin is Protective in Pancreatic Cancer while Reducing Radiation-Induced Hepatic Oxidative Damage.

Pancreatic cancer is one of the fatal causes of global cancer-related deaths. Although surgery and chemotherapy are standard treatment options, post-treatment outcomes often end in a poor prognosis. In the present study, we investigated anti-pancreatic cancer and amelioration of radiation-induced oxidative damage by crocin. Crocin is a carotenoid isolated from the dietary herb saffron, a prospect for novel leads as an anti-cancer agent. Crocin significantly reduced cell viability of BXPC3 and Capan-2 by triggering caspase signaling via the downregulation of Bcl-2. It modulated the expression of cell cycle signaling proteins P53, P21, P27, CDK2, c-MYC, Cyt-c and P38. Concomitantly, crocin treatment-induced apoptosis by inducing the release of cytochrome c from mitochondria to cytosol. Microarray analysis of the expression signature of genes induced by crocin showed a substantial number of genes involved in cell signaling pathways and checkpoints (723) are significantly affected by crocin. In mice bearing pancreatic tumors, crocin significantly reduced tumor burden without a change in body weight. Additionally, it showed significant protection against radiation-induced hepatic oxidative damage, reduced the levels of hepatic toxicity and preserved liver morphology. These findings indicate that crocin has a potential role in the treatment, prevention and management of pancreatic cancer.

Microneedle arrays allow lower microbial penetration than hypodermic needles in vitro.

METHODS: In this study we determined, for the first time, the ability of microorganisms to traverse microneedle-induced holes using two different in vitro models. RESULTS: When employing Silescol membranes, the numbers of Candida albicans, Pseudomonas aeruginosa and Staphylococcus epidermidis crossing the membranes were an order of magnitude lower when the membranes were punctured by microneedles rather than a 21G hypodermic needle. Apart from the movement of C. albicans across hypodermic needle-punctured membranes, where 40.2% of the microbial load on control membranes permeated the barrier over 24 h, the numbers of permeating microorganisms was less than 5% of the original microbial load on control membranes. Experiments employing excised porcine skin and radiolabelled microorganisms showed that the numbers of microorganisms penetrating skin beyond the stratum corneum were approximately an order of magnitude greater than the numbers crossing Silescol membranes in the corresponding experiments. Approximately 10(3) cfu of each microorganism adhered to hypodermic needles during insertion. The numbers of microorganisms adhering to MN arrays were an order of magnitude higher in each case. CONCLUSION: We have shown here that microneedle puncture resulted in significantly less microbial penetration than did hypodermic needle puncture and that no microorganisms crossed the viable epidermis in microneedle-punctured skin, in contrast to needle-punctured skin. Given the antimicrobial properties of skin, it is, therefore, likely that application of microneedle arrays to skin in an appropriate manner would not cause either local or systemic infection in normal circumstances in immune-competent patients. In supporting widespread clinical use of microneedle-based delivery systems, appropriate animal studies are now needed to conclusively demonstrate this in vivo. Safety in patients will be enhanced by aseptic or sterile manufacture and by fabricating microneedles from self-disabling materials (e.g. dissolving or biodegradable polymers) to prevent inappropriate or accidental reuse.

Microneedle arrays permit enhanced intradermal delivery of a preformed photosensitizer.

Silicon microneedle (MN) arrays were used to puncture excised murine and porcine skin in vitro and transdermal and intradermal delivery of meso-tetra (N-methyl-4-pyridyl) porphine tetra tosylate (TMP) investigated using topical application of a bioadhesive patch containing 19 mg TMP cm(-2). Animal studies, using nude mice, were then conducted to investigate the in vivo performance of the bioadhesive patch following MN puncture of skin. MN puncture significantly enhanced both intradermal and transdermal delivery of TMP in vitro, though the total amounts of drug delivered (25.22% into porcine skin and 0.07% across murine skin) were still quite small in each case. Notwithstanding this, in vivo experiments showed that MN puncture was capable of permitting a prolonged increase in TMP fluorescence at the site of application. Importantly, fluorescence was negligible at distant sites, meaning systemic delivery of the drug was not sufficient to induce TMP accumulation other than at the application site. In this study we have conclusively demonstrated proof of principle; MN puncture allows true intradermal delivery of a preformed photosensitizer in animal skin models in vitro and in vivo. Importantly, transdermal delivery was much reduced in each case. Increasing MN density would allow increased amounts of photosensitizer to be delivered. However, as MNs create aqueous pores in the stratum corneum, a preformed photosensitizer must possess at least some degree of water solubility in order to permit enhanced intradermal delivery in this way. We believe that use of MN array technology in this way has the potential to significantly improve topical photodynamic therapy of skin tumors.

Response of vulval lichen sclerosus and squamous hyperplasia to photodynamic treatment using sustained topical delivery of aminolevulinic acid from a novel bioadhesive patch system.

This study evaluated the clinical and histopathological responses of vulval lichen sclerosus (LS) and squamous hyperplasia (SH) to photodynamic therapy (PDT). A novel bioadhesive patch containing aminolevulinic acid (ALA) at a dose of (38 mg/cm(2)) was used to treat 10 patients before irradiation with light of 630 nm. Clinical, histopathological and pathological responses to treatment were assessed at 6 weeks post-treatment. After 17 cycles of PDT, six patients reported significant symptomatic relief and no cutaneous photosensitivity. Histopathological differences were not demonstrated, but statistically significant induction of apoptosis was seen. It can be concluded that ALA-PDT patch-based formulation is pragmatic and primarily offers symptomatic management of vulval LS and SH.

Aminolevulinic acid-loaded Witepsol microparticles manufactured using a spray congealing procedure: implications for topical photodynamic therapy.

OBJECTIVES: The aim was to enhance aminolevulinic acid (ALA) stability by incorporation into low-melting microparticles prepared using a spray congealing procedure and to evaluate temperature-triggered release, allowing topical bioavailability following melting at skin temperature. METHODS: ALA-loaded Witepsol microparticles were prepared using a novel spray congealing technique. Entrapment efficiency was compared with conventional emulsion-based methods and modelled drug release profiles determined using a membrane separation technique. Raised receiver medium temperature was used to determine triggered release. Bioavailability and lipid-mediated enhancement of ALA penetration were determined in excised murine skin. KEY FINDINGS: ALA-loaded Witepsol microparticles were spherical, with a mean diameter of 20 mum. Loading and stability studies demonstrated effective encapsulation, ranging from 91% to 100%, with no evidence of degradation to pyrazine derivatives. ALA release correlated with dissolution medium temperature, triggered at temperatures close to that of skin. Results suggested that molten Witepsol enhanced cutaneous permeation, whereas incorporation of microparticles in a semi-solid vehicle attenuated ALA penetration. Optimal use was direct application under occlusion. CONCLUSIONS: Spray congealing is superior to the emulsion-based procedures with respect to encapsulation efficiency of ALA in Witepsol matrices, providing temperature-triggered release, enhanced stability and improved penetration of ALA through keratinised skin. These features could improve ALA delivery to superficial lesions as part of photodynamic therapy.

Influence of formulation factors on PpIX production and photodynamic action of novel ALA-loaded microparticles.

A novel 5-aminolevulinic acid (ALA)-containing microparticulate system was produced recently, based on incorporation of ALA into particles prepared from a suppository base that maintains drug stability during storage and melts at skin temperature to release its drug payload. The novel particulate system was applied to the skin of living animals, followed by study of protoporphyrin IX (PpIX) production. The effect of formulating the microparticles in different vehicles was investigated and also the phototoxicity of the PpIX produced using a model tumour.Particles formulated in propylene glycol gels (10% w/w ALA loading) generated the highest peak PpIX fluorescence levels in normal mouse skin. Peak PpIX levels induced in skin overlying subcutaneously implanted WiDr tumours were significantly lower than in normal skin for both the 10% w/w ALA microparticles alone and the 10% w/w ALA microparticles in propylene glycol gels during continuous 12 h applications. Tumours not treated with photodynamic therapy continued to grow over the 17 days of the anti-tumour study. However, those treated with 12 h applications of either the 10% w/w ALA microparticles alone or the 10% w/w ALA microparticles in propylene glycol gel followed by a single laser irradiation showed no growth. The gel formulation performed slightly better once again, reducing the tumour growth rate by approximately 105%, compared with the 89% reduction achieved using particles alone. Following the promising results obtained in this study, work is now going on to prepare particle-loaded gels under GMP conditions with the aim of initiating an exploratory clinical trial.

Processing difficulties and instability of carbohydrate microneedle arrays.

BACKGROUND: A number of reports have suggested that many of the problems currently associated with the use of microneedle (MN) arrays for transdermal drug delivery could be addressed by using drug-loaded MN arrays prepared by moulding hot melts of carbohydrate materials. METHODS: In this study, we explored the processing, handling, and storage of MN arrays prepared from galactose with a view to clinical application. RESULTS: Galactose required a high processing temperature (160 degrees C), and molten galactose was difficult to work with. Substantial losses of the model drugs 5-aminolevulinic acid (ALA) and bovine serum albumin were incurred during processing. While relatively small forces caused significant reductions in MN height when applied to an aluminium block, this was not observed during their relatively facile insertion into heat-stripped epidermis. Drug release experiments using ALA-loaded MN arrays revealed that less than 0.05% of the total drug loading was released across a model silicone membrane. Similarly, only low amounts of ALA (approximately 0.13%) and undetectable amounts of bovine serum albumin were delivered when galactose arrays were combined with aqueous vehicles. Microscopic inspection of the membrane following release studies revealed that no holes could be observed in the membrane, indicating that the partially dissolved galactose sealed the MN-induced holes, thus limiting drug delivery. Indeed, depth penetration studies into excised porcine skin revealed that there was no significant increase in ALA delivery using galactose MN arrays, compared to control (P value < 0.05). Galactose MNs were unstable at ambient relative humidities and became adhesive. CONCLUSION: The processing difficulties and instability encountered in this study are likely to preclude successful clinical application of carbohydrate MNs. The findings of this study are of particular importance to those in the pharmaceutical industry involved in the design and formulation of transdermal drug delivery systems based on dissolving MN arrays. It is hoped that we have illustrated conclusively the difficulties inherent in the processing and storage of carbohydrate-based dissolving MNs and that those in the industry will now follow alternative approaches.