Nanoparticle-mediated targeting of MAPK signaling predisposes tumor to chemotherapy

The MAPK signal transduction cascade is dysregulated in a majority of human tumors. Here we report that a nanoparticle-mediated targeting of this pathway can optimize cancer chemotherapy. We engineered nanoparticles from a unique hexadentate-polyD,L-lactic acid-co-glycolic acid polymer chemically conjugated to PD98059, a selective MAPK inhibitor. The nanoparticles are taken up by cancer cells through endocytosis and demonstrate sustained release of the active agent, resulting in the inhibition of phosphorylation of downstream extracellular signal regulated kinase. We demonstrate that nanoparticle-mediated targeting of MAPK inhibits the proliferation of melanoma and lung carcinoma cells and induces apoptosis in vitro. Administration of the PD98059-nanoparticles in melanoma-bearing mice inhibits tumor growth and enhances the antitumor efficacy of cisplatin chemotherapy. Our study shows the nanoparticle-mediated delivery of signal transduction inhibitors can emerge as a unique paradigm in cancer chemotherapy.Department of Defense Breast Cancer Research Program Era of Hope Award (W81XWH-07–1-0482)Mary Kay Ash Charitable Trus

Adhesive and conformational behaviour of mycolic acid monolayers

We have studied the pH-dependent interaction between mycolic acid (MA) monolayers and hydrophobic and hydrophilic surfaces using molecular (colloidal probe) force spectroscopy. In both cases, hydrophobic and hydrophilic monolayers (prepared by Langmuir-Blodgett and Langmuir-Schaefer deposition on silicon or hydrophobized silicon substrates, respectively) were studied. The force spectroscopy data, fitted with classical DLVO (Derjaguin, Landau, Verwey, and Overbeek) theory to examine the contribution of electrostatic and van der Waals forces, revealed that electrostatic forces are the dominant contribution to the repulsive force between the approaching colloidal probe and MA monolayers. The good agreement between data and the DLVO model suggest that beyond a few nm away from the surface, hydrophobic, hydration, and specific chemical bonding are unlikely to contribute to any significant extent to the interaction energy between the probe and the surface. The pH-dependent conformation of MA molecules in the monolayer at the solid-liquid interface was studied by ellipsometry, neutron reflectometry, and with a quartz crystal microbalance. Monolayers prepared by the Langmuir-Blodgett method demonstrated a distinct pH-responsive behaviour, while monolayers prepared by the Langmuir-Schaefer method were less sensitive to pH variation. It was found that the attachment of water molecules plays a vital role in determining the conformation of the MA monolayers. (C) 2010 Elsevier B.V. All rights reserved

Large sub-clonal variation in <i>Phytophthora infestans</i> from recent severe late blight epidemics in India

Abstract The population structure of the Phytophthora infestans populations that caused the recent 2013–14 late blight epidemic in eastern India (EI) and northeastern India (NEI) was examined. The data provide new baseline information for populations of P. infestans in India. A migrant European 13_A2 genotype was responsible for the 2013–14 epidemic, replacing the existing populations. Mutations have generated substantial sub-clonal variation with 24 multi-locus genotypes (MLGs) found, of which 19 were unique variants not yet reported elsewhere globally. Samples from West Bengal were the most diverse and grouped alongside MLGs found in Europe, the UK and from neighbouring Bangladesh but were not linked directly to most samples from south India. The pathogen population was broadly more aggressive on potato than on tomato and resistant to the fungicide metalaxyl. Pathogen population diversity was higher in regions around the international borders with Bangladesh and Nepal. Overall, the multiple shared MLGs suggested genetic contributions from UK and Europe in addition to a sub-structure based on the geographical location within India. Our data indicate the need for improved phytosanitary procedures and continuous surveillance to prevent the further introduction of aggressive lineages of P. infestans into the country

Purinergic signalling and immune cells

This review article provides a historical perspective on the role of purinergic signalling in the regulation of various subsets of immune cells from early discoveries to current understanding. It is now recognised that adenosine 5'-triphosphate (ATP) and other nucleotides are released from cells following stress or injury. They can act on virtually all subsets of immune cells through a spectrum of P2X ligand-gated ion channels and G protein-coupled P2Y receptors. Furthermore, ATP is rapidly degraded into adenosine by ectonucleotidases such as CD39 and CD73, and adenosine exerts additional regulatory effects through its own receptors. The resulting effect ranges from stimulation to tolerance depending on the amount and time courses of nucleotides released, and the balance between ATP and adenosine. This review identifies the various receptors involved in the different subsets of immune cells and their effects on the function of these cells

Effect of mycobacterial lipids on surface properties of Curosurf (TM): Implications for lung surfactant dysfunction in tuberculosis

In this study, the effect of mycobacterial lipids on the surface activity of lung surfactant was evaluated. Mycolic acid and cord factor, the most abundant surface active lipids of the mycobacterial cell wall when combined with Curosurf (TM) led to alteration of its surface properties. Addition of graded amounts of mycolic acid increased the minimum surface tension of Curosurf (TM) monolayers from <10 mN/m to similar to 20-27 mN/m. Presence of mycolic acid also slowed the rate of Curosurf (TM) adsorption. Similarly, presence of cord factor increased the minimum surface tension achieved by Curosurf (TM) to similar to 16-27 mN/m. AFM imaging revealed presence of aggregates on addition of mycobacterial lipids to Curosurf (TM) monolayers. Results of this study show that mycolic acid and cord factor can biophysically inactivate porcine lung surfactant extract (Curosurf (TM)). This suggests that biophysical inhibition of lung surfactant is possible in vivo in pulmonary tuberculosis which could aggravate areas of alveolar atelectasis. (c) 200

Effect of mycolic acid on surface activity of binary surfactant lipid monolayers

In pulmonary tuberculosis, Mycobacterium tuberculosis lies in close physical proximity to alveolar surfactant. Cell walls of the mycobacteria contain loosely bound, detachable surface-active lipids. In this study, the effect of mycolic acid (MA), the most abundant mycobacterial cell wall lipid, on the surface activity of phospholipid mixtures from lung surfactant was investigated using Langmuir monolayers and atomic force microscopy (AFM). In the presence of mycolic acid, all the surfactant lipid mixtures attained high minimum surface tensions (between 20 and 40 mN/m) and decreased surface compressibility moduli < 50 mN/m. AFM images showed that the smooth surface topography of surfactant lipid monolayers was altered with addition of MA. Aggregates with diverse heights of at least two layer thicknesses were found in the presence of mycolic acid. Mycolic acids could aggregate within surfactant lipid monolayers and result in disturbed monolayer surface activity. The extent of the effect of mycolic acid depended on the initial state of the monolayer, with fluid films of DPPC-POPC and DPPC-CHOL being least affected. The results imply inhibitory effects of mycolic acid toward lung surfactant lipids and could be a mechanism of lung surfactant dysfunction in pulmonary tuberculosis. (C) 200

Evaluation of antitubercular drug-loaded surfactants as inhalable drug-delivery systems for pulmonary tuberculosis

Pulmonary tuberculosis is associated with a year-long chemotherapy, poor alveolar drug levels, drug-related systemic toxicity, and patient noncompliance. In this study, exogenous pulmonary surfactant is proposed as a drug carrier for antitubercular drugs. Dipalmitoylphosphatidylcholine (DPPC), the major lung-surfactant lipid, has been combined with antitubercular drugs isoniazid (INH), rifampicin (RFM), and ethambutol (ETH) in 1:1 ratio by weight, in which drugs had a ratio of 1:23 by weight. At 37 degrees C, the formulation had better surfactant function with quicker reduction of surface tension on adsorption (32.71 +/- 0.65 mN/m) than DPPC liposomes (44.67 +/- 0.57 mN/m) and maintained 100% airway patency in a capillary surfactometer. Drug-loacted surfactant liposomes were about 2 mu m and had entrapment efficiency of 30.04%+/- 2.05%, 18.85%+/- 2.92%, and 61.47%+/- 3.32% for INH, RFM, and ETH, respectively. Sustained release of the drugs from surfactants was observed over 24 h. In vitro alveolar deposition efficiency using the twin impinger showed 12.06%+/- 1.87% of INH, 43.30%+/- 0.87% of RFM, and 22.07%+/- 2.02%, of ETH deposited in the alveolar chamber upon nebulization for a minute using a jet nebulizer. The formulation was biocompatible and stable with physicochemical properties being retained even after storage for a month at 4 degrees C. Antitubercular drug-loaded surfactants developed Could serve dual purposes of alveolar stabilization due to surfactant action and better reach of these drugs to the alveoli due to antiatelectatic effect of the surfactant. (C) 2008 . J Biomed Mater Res 89A: 281-292, 200

Evaluation of antitubercular drug insertion into preformed dipalmitoylphosphatidylcholine monolayers

Insertion profiles of antitubercular drugs isoniazid (INH), rifampicin (RFM) and ethambutol (ETH) into dipalmitoylphosphatidylcholine (DPPC) membrane models were evaluated by Langmuir monolayer technique. Maximum drug insertion into DPPC monolayer was observed with rifampicin with a surface pressure increase (Δπmax) in the range of 21-33 mN/m depending upon rifampicin concentration. Isoniazid had minimal insertion resulting in a lower Δπmax of about 2-3 mN/m, suggestive of minimal interactions between INH and DPPC. Ethambutol surface pressure increment on insertion resulted in an intermediate rise in the Δπmax (6-10 mN/m). Antitubercular drug combination in the ratio of 2 mM:0.7 mM:4.5 mM for INH:RFM:ETH, attained Δπmax between 25 and 33 mN/m. Insertion profiles similar to rifampicin were exhibited by the antitubercular drug mixture suggestive of predominant rifampicin insertion into the DPPC monolayer. The extent of drug insertion into the DPPC monolayer is suggestive of the drug penetration potential into biological membranes in vivo. Higher RFM Δπmax is suggestive of excellent cell membrane penetration, which explains broad reach of the drug to all the organs including the cerebrospinal fluid while lower Δπmax of INH suggests poor membrane penetration restricting the entry of the drug in different biological membranes. DPPC membrane destabilization was observed at higher antitubercular drug concentrations indicated by the negative slopes of the surface pressure-time curves. This may correlate with the dose related toxic effects observed in tuberculosis affected patients. Drug insertion studies offer a potential tool in understanding the pharmacotoxicological behavior of the various pharmacological agents.© Elsevie

Effect of antitubercular drugs on dipalmitoylphosphatidylcholine monolayers: implications for drug loaded surfactants

The year long chemotherapy in pulmonary tuberculosis results in dose related side effects and may not reach atelectatic areas. On account of its spreading properties and the ability to re-expand atelectatic areas, exogenous surfactant may act as a pulmonary drug delivery agent. We investigated the interactions between antitubercular drugs and the main surfactant component, dipalmitoylphosphatidylcholine (DPPC) with the aim of developing more effective antitubercular drug loaded surfactants. The surface properties were evaluated using a Langmuir-Blodgett trough and Wilhelmy balance at 37degreesC. Lung surfactant was modeled as DPPC monolayers. The isoniazid (INH)-DPPC combination in 1:1 ratio by weight significantly improved the adsorption of DPPC, reached a minimum surface tension of zero, formed a low compressibility film and required 32.7% area change to decrease surface tension from 30 to 10 mN/m. The triple drug (INH-rifampicin-ethambutol in 1:23 ratio by weight) DPPC combination when used in 1: 1 or 1:2 ratios by weight also achieved surface properties superior to those of DPPC alone. A significant improvement in the adsorption was observed (surface tensions of 34.7 mN/m for 1: 1 and 32.0 mN/m for 1:2 triple drug: DPPC combinations in the first second), and the films had low compressibility reaching a minimum surface tension of zero on compression. Thus, we observed statistically significant improvements in all the surface parameters and we feel encouraged to continue developing a tuberculosis therapy consisting of surfactant liposomes carrying antitubercular drugs. (C) 2004