Synthetically modified methoxsalen for enhanced cytotoxicity in light and dark reactions.

Linear furocoumarins, also known as psoralens, are clinically useful photo-activated pharmaceuticals employed to address hyperproliferative skin diseases. Seven diverse cytotoxic pharmacophores have been synthetically attached to 8-methoxypsoralen via a 5-amino functionality. The resulting unique set of compounds was evaluated for dark and light toxicity against PAM212 keratinocytes in culture.

Intertidal mudflat and saltmarsh conservation and sustainable use in the UK: a review.

The adoption of the Convention on Wetlands of International Importance in Ramsar, Iran in 1971 committed the UK to conserve and sustainably use intertidal mudflats and saltmarshes for the benefit of present and future generations. Through consideration of their importance and value, current status, the characteristics, causes and consequences of their loss, and the associated responses to loss, this paper reviews the UK progress towards the conservation and sustainable use of intertidal mudflats and saltmarshes. Uncertainties in their current status and trends make it difficult to assess the overall net change in extent across the UK. However, it is apparent that losses due to erosion continue to exceed gains from intertidal mudflat and saltmarsh reparation (IMSR) schemes in south-east and southern England. IMSR schemes in the UK have been generally limited to relatively small-scale trials in comparison to elsewhere in Europe and in the USA. No research to date has unequivocally identified the causes of erosion. Regardless of the cause, the loss of intertidal mudflats and saltmarshes has adverse impacts on the provision of ecosystem services upon which humans and other species depend. The evidence presented in this paper suggests that alongside further science-based research, there is a need to develop a decision-making process capable of accommodating complexity, uncertainty and multiple diverse perspectives, through which more informed, timely decisions and more effective, concerted actions to conserve and sustainably use intertidal mudflats and saltmarshes can be taken.

Implementing a Learning Collaborative Framework for States Working to Improve Outcomes for Vulnerable Populations: The Opioid Use Disorder, Maternal Outcomes, and Neonatal Abstinence Syndrome Initiative Learning Community.

The opioid crisis has impacted vulnerable populations, specifically pregnant and postpartum women, and infants prenatally exposed to substances, including infants with Neonatal Abstinence Syndrome. Lack of access to clinical and social services; potential stigma or discrimination; and lack of resources for provision of services, including screening and treatment, have impacted the health of these populations. In 2018, using a systems change approach, the Association of State and Territorial Health Officials (ASTHO) and the Centers for Disease Control and Prevention (CDC) convened an Opioid use disorder, Maternal outcomes, Neonatal abstinence syndrome Initiative Learning Community (OMNI LC) that included other federal agencies, national clinical and nonclinical organizations, and 12 state leadership groups. The purpose of the OMNI LC was to determine areas of focus and identify strategies and best practices for implementing systems change to improve maternal and infant outcomes associated with opioid use disorder (OUD) during the perinatal period. Activities included in-person convenings with policy goal action plan development, virtual learning sessions, intensive technical assistance (TA), and temporary field placements. The OMNI LC partnering agencies and state teams met bimonthly for the first year of the initiative. At the in-person convening, state teams identified barriers to developing and implementing systems change in activity-specific action plans within five areas of focus: financing and coverage; access to and coordination of quality services; provider training and awareness; ethical, legal, and social considerations; and data, monitoring, and evaluation. State teams also identified stakeholder partnerships as a necessary component of strategy development in all areas of focus. Four virtual learning sessions were conducted on the areas of focus identified by state teams, and ASTHO conducted three intensive TA opportunities, and five states were identified for temporary field placement. To successfully address the impact of the opioid crisis on pregnant and postpartum women and infants, states developed innovative strategies focused on increasing support, services, and resources. Moving forward, state teams will participate in two additional in-person meetings, continue to identify barriers to the work, refine and customize action plans, and set new goals, to effect broad-ranging systems change for these vulnerable populations.

Examination of metformin hydrochloride in a continuous dissolution/HDM system.

A continuous dissolution/absorption system using a hexadecane membrane (HDM) as the permeation measurement has been examined for three distinct formulations of metformin hydrochloride. This system was used to correlate the absorption rate of metformin through the membrane after release from the dosage form to rate of appearance of metformin in the plasma from the same formulations. These correlations were then used to make predictions of the in vivo plasma profile for each formulation. Successful predictions of AUC were accomplished for both immediate release and extended release formulations of metformin hydrochloride.

SYNTHESIS AND EVALUATION OF WATER-SOLUBLE DIMETHYLAMINOETHYL ETHERS OF METHOXSALEN FOR PROLIFERATIVE SKIN DISORDERS.

The natural product 8-methoxypsoralen (methoxsalen or 8-MOP) in combination with long wavelength ultraviolet light (UVA, 320-400 nm), also referred to as PUVA therapy, is used for the treatment of cutaneous proliferative disorders including psoriasis, vitiligo and mycosis fungoides. The use of 8-MOP (3) is limited by its poor water solubility and there remains a need to develop more water-soluble psoralens to enhance bioavailability following oral administration of the drug. In the present studies a water-soluble dimethylaminoethyl ether analog of 8-MOP was synthesized and analyzed for biological activity. This analog, (8-[2-(N,N-dimethylamino)ethoxy]-psoralen hydrochloride (1) [or CAS name: 9-[2-(dimethylamino)ethoxy]-7H-furo[3,2-g][1]benzopyran-7-one, hydrochloride], was found to be significantly more active than 3 in keratinocyte growth inhibition assays (IC50 = 12 nM and 130 nM for 1 and 3, respectively). The partially reduced dihydro derivative of 1, 8-[2-(N,N-dimethylamino)ethoxy]-4',5'-dihydropsoralen hydrochloride (2) [or CAS name: 9-[2-(dimethylamino)ethoxy]-2,3-dihydro-7H-furo[3,2-g][1]benzopyran-7-one, hydrochloride] and the partially reduced 4',5'-dihydro-8-methoxypsoralen (4) lacking the water-solubilizing side-chain were significantly less active. As inhibitors of keratinocyte growth they ranked as IC50 = 13,000 nM and 70,000 nM for 2 and 4, respectively, indicating that an unsaturated furan ring in the psoralen was required for maximal activity. Compound (1) was found to readily intercalate and damage DNA following UVA light treatment as determined by plasmid DNA nicking and unwinding experiments in neutral and alkaline agarose gels. Taken together, these data demonstrate that a water-soluble dimethylaminoethyl ether psoralen targets DNA, is highly active as a photosensitizer, and may be useful in the treatment of skin diseases involving abnormal keratinocyte proliferation.

Predicting human drug pharmacokinetics from in vitro permeability using an absorption-disposition model.

The purpose of this research is to simulate the in vivo performance of drugs with a wide range of solubility and permeability characteristics formulated as oral dosage forms. The absorption-disposition model was developed using a number of physiological parameters as well as in vitro permeability data generated with Caco-2 cells, 2/4/A1 cells, and hexadecane membranes. A total of 13 drugs with varying solubility and permeability properties were examined using the absorption-disposition model to predict their pharmacokinetic profile. The correlation of predicted and experimentally determined AUC and Cmax, as measures of the pharmacokinetic profile, were >0.96 for all permeation techniques examined. The predictive ability of the model is influenced by the type of permeation method employed; 2/4/A1 cell data yielded the highest degree of accuracy in predicting Cmax and AUC values. The absorption-disposition model developed in this work accurately predicts the in vivo performance of a wide range of orally administered drugs with 8 of 9 drugs examined falling within 80-125% of the experimental value of AUC when using 2/4/A1 cells.

Development of a molecularly imprinted polymer for the analysis of avermectin.

Five molecularly imprinted polymers (MIPs) were synthesized for a large molecule, avermectin, using different preparation techniques, monomers, and polymerization solvents. Selectivities (α) of each were compared using HPLC and different mobile phases containing various levels of acetic acid. Selectivity (α) for avermectin was greatest (α estimated ≥18) when the polymer was prepared non-covalently (utilizing only non-covalent interactions between avermectin and monomer) in chloroform using methacrylic acid (MAA) monomer and evaluated in chloroform. When evaluated in acetonitrile, an MIP prepared in acetonitrile provided better selectivity (α=8.4) than the polymer prepared in chloroform. Optimizing mobile phase conditions by adding acetic acid was much more important when MIPs were evaluated in chloroform than in acetonitrile. MIPs prepared with MAA provided better selectivity than a polymer prepared with acrylamide monomer. Covalent preparation of two MIPs utilizing a covalent bond between avermectin and monomer before polymerization did not improve selectivity but did improve peak shape in chromatograms. Specificity was demonstrated by comparing the selectivity of avermectin with eprinomectin (α=3.0), a compound with a very similar structure. Results indicate that an MIP can be prepared for the large avermectin molecule, and has the potential to simplify sample preparation and to reduce the time needed for analysis.

Inflammatory cytokines induce protein tyrosine nitration in rat astrocytes.

Protein tyrosine nitration may be relevant for the pathogenesis of hepatic encephalopathy (HE). Infections, sepsis, and trauma precipitate HE episodes. Recently, serum levels of tumor necrosis factor (TNF)-alpha were shown to correlate with severity of HE in chronic liver failure. Here the effects of inflammatory cytokines on protein tyrosine nitration in cultured rat astrocytes and rat brain in vivo were studied. In cultured rat astrocytes TNF-alpha (50 pg/ml-10 ng/ml) within 6h increased protein tyrosine nitration. TNF-alpha-induced tyrosine nitration was related to an increased formation of reactive oxygen and nitrogen intermediates, which was downstream from a NMDA-receptor-dependent increase of intracellular [Ca(2+)](i) and nNOS-catalyzed NO production. Astroglial tyrosine nitration was also elevated in brains of rats receiving a non-lethal injection of lipopolysaccharide, as indicated by colocalization of nitrotyrosine immunoreactivity with glial fibrillary acidic protein and glutamine synthetase, and by identification of the glutamine synthetase among the tyrosine-nitrated proteins. It is concluded that reactive oxygen and nitrogen intermediates as well as protein tyrosine nitration by inflammatory cytokines may alter astrocyte function in an NMDA-receptor-, Ca(2+)-, and NOS-dependent fashion. This may be relevant for the pathogenesis of HE and other conditions involving cytokine exposure the brain.

Hypoosmotic swelling increases protein tyrosine nitration in cultured rat astrocytes.

Astrocyte swelling is observed in different types of brain injury. We studied a potential contribution of swelling to protein tyrosine nitration (PTN) by using cultured rat astrocytes exposed to hypoosmotic (205 mosmol/L) medium. Hypoosmolarity (2 h) increases total PTN by about 2-fold in 2 h. The hypoosmotic PTN is significantly inhibited by the NMDA receptor antagonist MK-801, the nitric oxide synthase (NOS) inhibitor L-NMMA, the extracellular Ca2+ chelator EGTA and the calmodulin antagonist W13, suggesting the involvement of NMDA receptor activation, influx of extracellular Ca2+ and Ca2+/calmodulin-dependent NO synthesis. Further, superoxide dismutase plus catalase and uric acid strongly inhibit hypoosmotic PTN, suggesting the involvement of the toxic metabolite peroxynitrite (ONOO-) as a nitrating agent. Hypoosmotic astrocyte swelling rapidly stimulates generation of reactive oxygen intermediates; this process is prevented by MK-801 and EGTA. In addition, MK-801 inhibits the hypoosmotic elevation of [Ca2+]i. The findings support the view that astrocyte swelling as induced, for example, by toxins relevant for hepatic encephalopathy is sufficient to produce oxidative stress and PTN and thus contributes to altered astroglial and neuronal function.

Benzodiazepine-induced protein tyrosine nitration in rat astrocytes.

Recent studies indicate that ammonia and hypoosmotic astrocyte swelling can induce protein tyrosine nitration (PTN) in astrocytes with potential pathogenetic relevance for hepatic encephalopathy (HE). Because HE episodes are known to be precipitated also by sedatives, the effects of benzodiazepines on PTN in cultured rat astrocytes and rat brain in vivo were studied. In cultured rat astrocytes, diazepam, PK11195, Ro5-4864, and the benzodiazepine binding inhibitor (DBI), which acts on peripheral-type benzodiazepine receptors, induced PTN. Clonazepam, a specific ligand of the central benzodiazepine receptor, failed to induce PTN. Nanomolar concentrations of DBI and PK11195 were sufficient to increase PTN, and diazepam effects were already observed at concentrations of 1 micromol/L. Diazepam-induced PTN was insensitive to NOS inhibition and uric acid but was blunted by MK-801, BAPTA-AM, W13, and catalase, suggesting an involvement of NMDA-receptor activation, elevation of the cytosolic Ca(2+) concentration [Ca(2+)](i), and hydrogen peroxide. Diazepam induced a plateau-like increase in [Ca(2+)](i) and the generation of reactive oxygen intermediates (ROIs), which are both blunted by MK-801 and BAPTA-AM. The expression of functional N-methyl-D-aspartate (NMDA) receptors on cultured rat astrocytes was confirmed by reverse transcriptase polymerase chain reaction, Western blot analysis, immunhistochemistry, and receptor autoradiography. Astroglial PTN is also found in brains from rats challenged with diazepam, indicating the in vivo relevance of the present findings. In conclusion, production of ROIs and increased PTN by benzodiazepines may alter astrocyte function and thereby contribute to the precipitation of HE episodes.