Francisco Amat-Guerri

Intracellular Triggering of Fas Aggregation and Recruitment of Apoptotic Molecules into Fas-enriched Rafts in Selective Tumor Cell Apoptosis

We have discovered a new and specific cell-killing mechanism mediated by the selective uptake of the antitumor drug 1-O-octadecyl-2-O-methyl-rac-glycero-3-phosphocholine (ET-18-OCH3, Edelfosine) into lipid rafts of tumor cells, followed by its coaggregation with Fas death receptor (also known as APO-1 or CD95) and recruitment of apoptotic molecules into Fas-enriched rafts. Drug sensitivity was dependent on drug uptake and Fas expression, regardless of the presence of other major death receptors, such as tumor necrosis factor (TNF) receptor 1 or TNF-related apoptosis-inducing ligand R2/DR5 in the target cell. Drug microinjection experiments in Fas-deficient and Fas-transfected cells unable to incorporate exogenous ET-18-OCH3 demonstrated that Fas was intracellularly activated. Partial deletion of the Fas intracellular domain prevented apoptosis. Unlike normal lymphocytes, leukemic T cells incorporated ET-18-OCH3 into rafts coaggregating with Fas and underwent apoptosis. Fas-associated death domain protein, procaspase-8, procaspase-10, c-Jun amino-terminal kinase, and Bid were recruited into rafts, linking Fas and mitochondrial signaling routes. Clustering of rafts was necessary but not sufficient for ET-18-OCH3–mediated cell death, with Fas being required as the apoptosis trigger. ET-18-OCH3–mediated apoptosis did not require sphingomyelinase activation. Normal cells, including human and rat hepatocytes, did not incorporate ET-18-OCH3 and were spared. This mechanism represents the first selective activation of Fas in tumor cells. Our data set a framework for the development of more targeted therapies leading to intracellular Fas activation and recruitment of downstream signaling molecules into Fas-enriched rafts.

Correlations between photophysics and lasing properties of dipyrromethene–BF2 dyes in solution

Abstract UV–Vis absorption, fluorescence (steady-state and time-correlated) and lasing properties of three pyrromethene–BF 2 dyes (PM546, PM556 and PM567) have been determined in several solvents. Good correlations between photophysical properties and lasing characteristics of these dyes have been obtained not only for a given dye in several solvents but also for the three dyes in a common solvent. Both the observed spectral changes and the deactivation mechanism affecting the lasing gain are explained on the basis of the possible resonance structures of the chromophore.

Solid-state dye lasers based on copolymers of 2-hydroxyethyl methacrylate and methyl methacrylate doped with rhodamine 6G

Rhodamine 6G has been dissolved in copolymers of 2-HydroxyEthyl MethAcrylate (HEMA) and Methyl MethAcrylate (MMA) and the resulting solid-state solutions have been pumped at 337 nm and 532 nm. Lasing efficiencies similar to those found in ethanol solution have been obtained with a 1:1 vol/vol HEMA: MMA copolymer matrix, and lifetimes of ca. 10 000 (337 nm pumping) and ca. 75 000 (532 nm pumping) pulses at repetition rates up to 15 Hz and 10 Hz, respectively, have been demonstrated.

Solid-state dye laser based on Coumarin 540A-doped polymeric matrices

Abstract Coumarin 540-A has been dissolved in a copolymer of 2-hydroxyethyl methacrylate (HEMA) and methyl methacrylate (MMA) 1:1 v/v and in a pure poly(methyl methacrylate) homopolymer (PMMA). Laser action has been induced in the resulting solid-state solutions pumped with 1.2 mJ pulses at 337 nm from a nitrogen laser. The effects on the laser performance of different polymerization methods, dye concentration and polymeric matrix composition have been evaluated. Energy conversion efficiencies of 11% and lifetimes of about 2000 pulses at 2 Hz repetition rate have been demonstrated.

Proton-transfer lasing from solid organic matrices

Abstract Solid-state laser oscillation has been induced in 2-(2′-hydroxy-5′-fluorophenyl)benzimidazole dissolved in poly(methyl methacrylate), based on a four-level excited-state proton-transfer mechanism. The energy conversion efficiency at 493 nm was ≈ 1% for N2 laser pumping with pulses of 2 mJ at 337 nm.

Laser action from rhodamine 6G-doped poly (2-hydroxyethyl methacrylate) matrices with different crosslinking degrees

Abstract The lasing properties of rhodamine 6G dissolved in poly (2-hydroxyethyl methacrylate) matrices with different amounts of ethylene glycol dimethacrylate as crosslinking monomer are investigated. Increasing the rigidity of the matrix by decreasing the polymer free volume results in significant increases of both lasing efficiency and photostability. There exists an optimum degree of crosslinking for which efficiency and photostability reach the highest values.

Solid‐state dye lasers based on modified rhodamine 6G dyes copolymerized with methacrylic monomers

Modified rhodamine 6G molecules with polymerizable double bonds have been copolymerized with methacrylic monomers and the resulting polymers have been pumped at 337 nm with a N2 laser in a transversal configuration. The preparation of these new materials is described in detail and their lasing properties are evaluated. Important increases in photostability, with lasing efficiencies similar to those found for the parent dye rhodamine 6G in ethanol solution, have been obtained for some of these materials. Lifetimes (measured as an 80% efficiency drop) in excess of 20 000 shots at repetition rates of 2 Hz have been demonstrated. Strong dependence on pump repetition rate was observed. Possible mechanisms and processes responsible for the behavior of these materials are discussed. By using a rotating system where the sample is scanned in a continuous way, the laser output remained stable, with no sign of degradation, after 500 000 shots.

The Solid State, Solution and Tubulin-Bound Conformations of Agents that Promote Microtubule Stabilization

Taxol (paclitaxel), a complex diterpene obtained from Taxus brevifolia and its semisynthetic analogue Taxotere are two of the most important new drugs for cancer chemotherapy. Their mechanism of cytotoxic action involves stabilization of microtubules leading to mitotic arrest. A similar mechanism has been proposed for an expanding set of other natural products, for instance, the epothilones, eleutherobin, the sarcodictyins, discodermolide, laulimalide, Rhazinilam, WS9885B, certain steroids and a group of polyisoprenyl benzophenones. In this review, we focus on the conformations of small molecule microtubule (MT) stabilizing compounds which have been isolated or synthesized and subjected to structural analysis. NMR and fluorescense spectroscopies, X-ray crystallography, high resolution microscopy (electron crystallography) and theoretical calculations comprise the most common methods used in this context. In particular, we describe how the structures were determined and with what accuracy. We also discuss the conformational diversity apparent from the three dimensional structures and compare the various proposals for bioactive conformations at the target MT binding sites. Of critical importance are the recently disclosed models for Taxol and its biomimetics binding to beta-tubulin. Several different conformational schemes derived from both pharmacophore construction and modeled protein ligand complexes are compared and critically evaluated. Although full consensus has yet to be reached, emphasis is placed on pharmacophore models for the various anti-MT agents that are internally consistent and encompass more than one structural class.

Singlet oxygen photogeneration by ionized and un-ionized derivatives of Rose Bengal and Eosin Y in diluted solutions

Abstract Quantum yields of singlet oxygen photogeneration (relative to commercial Rose Bengal) were determined for the pure dye, Eosin Y, their corresponding methyl, benzyl, and p-isopropylbenzyl esters, and the product of Rose Bengal decarboxylation, all in solution in basic methanol (in order to assure total ionization). The similar values found for each series of salts indicate a low influence, if any, of the different C-2′ groups on their behaviour as sensitizers. As expected, the monodehalogenated parent dyes showed in the same medium somewhat lower yields. Slightly lower values were also found for the un-ionized forms of the methyl esters and for the non-ionizable methyl ether of Rose Bengal methyl ester in acidic dioxane as medium. In order to avoid the formation of dye aggregates and the quenching of the triplets of the sensitizer by the acceptor, all the yields were determined using solutions of 10−5 M sensitizer and 10−4 M or 10−5 M acceptor. The methodology, including the spectrophotometric procedure used to determine relative β values, is also reported.

Solvent influence on the kinetics of the photodynamic degradation of trolox, a water-soluble model compound for vitamin E

The kinetics of the aerobic dye-sensitized photo-oxidation of 6-hydroxy-2,5,7,8-tetramethylchroman-2-carboxylic acid (Trolox), a water-soluble model compound for vitamin E, has been studied in aqueous solutions of pH values 6 and 12 and in organic media of different polarity. Experimental evidence indicates that the process largely occurs through a Trolox-singlet molecular oxygen O2(1Δg) interaction, the photo-oxidation rates increasing with the pH value and the polarity of the medium. Low polarity solvents favour the physical quenching of O2(1Δg) by Trolox, as already observed in other tocopherols. High polarity solvents drive the interaction predominantly towards a reactive pathway that yields the same substituted p-benzoquinone that can be obtained by thermal oxidation of Trolox. Overall rate constants for the quenching of O2(1Δg) by Trolox (determined by time-resolved phosphorescence spectroscopy) and reactive rate constants (obtained by relative methods) are both in the range 107−108 M−1s−1. On the basis of these and their results and their comparison with previous results on the photo-oxidation of tocopherol derivatives, the role of the solvent in the behaviour of vitamin E is discussed.