Studies of cationic amphiphilic drug catalysed membrane degradation

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Studies of cationic amphiphilic drug catalysed membrane degradation

Although drug molecules are designed to bind specifically to targets such as receptors that are embedded within biological membranes, it is becoming increasingly evident that a large fraction of these compounds interact non-specifically with the membrane and I or other proteins. In particular, the non-specific binding of positron emission tomography (PET) radioligands to tissue, both in-vivo and in-vitro, is poorly understood. This phenomenon is a major confounding factor in the development of new radioligands for receptor imaging in-vivo. To address this issue, studies on the interaction of central nervous system (eNS) drugs, belonging to the cationic amphiphilic drug (CAD) family, with model membrane assemblies were conducted. Experiments were performed using three CADs: Haloperidol (HPD), Spiperone .(SPIP) and WAY, on condensed fluid lamellar phases and membrane vesicles. CADmembranes interactions were studied by small angle X-ray scattering (SAXS), soIidstate nuclear magnetic resonance (SS-NMR), fluorescence assays, and fluorescence microscopy. CADs were found to partition rapidly to the polar I apolar region of the membrane; this was demonstrated by SAXS where CADs affected the bilayer spacing. At physiological pH, the protonated groups on the CAD catalyze the acid-hydrolysis of the ester linkage present in the phospholipid chains, producing a fatty acid and singlechain lipid. 'The single-chain lipids destabilize the membrane, causing membranous . fragments and small vesicles to separate and diffuse away from the host. These membrane fragments carry the drug molecules with them. The entire process, from drug adsorption to drug release within mi~ell~ fragments, occurs on a timescale of seconds I minutes. Given the rate at which this occurs it is probable that this process is a significant mechanism for drug transport. Kinetic studies were conducted to determine the rate of the lipid hydrolysis in the condensed fluid lamellar phase, by varying CAD's counterions, the lipid composition and the stored curvature elastic stress in the bilayer. The lipid hydrolysis kinetics was fitted to a pseudo-first order exponential decay, and hydrolysis rates were determined. Hydrolysis rates are specific to the CAD molecules, with WAY hydrolysing the bilayer as twice as fast as SPIP. In addition, evidence is presented that the stored curvature elastic stress in the membrane modulates the hydrolysis kinetics. Interestingly, the rate of membrane hydrolysis appears to correlate with in-vivo nonspecific binding of the PET radioligands. The measured rate of membrane hydrolysis may provide useful insight into the mechanism of non-specific binding on a molecular level and possibly in the design ofnew radiotracers.EThOS - Electronic Theses Online ServiceGBUnited Kingdo

Photoresponse of Complexes between Surfactants and Azobenzene-Modified Polymers Accounting for the Random Distribution of Hydrophobic Side Groups

The design of photoresponsive macromolecules has opened the route to many applications, in particular to trigger macroscopic responses induced by light irradiation in complex fluids and polymer−surfactant formulations. In this report, we studied the association of three sets of azobenzene modified polymer (AMPs) derived from poly(acrylic)acid with varying integration levels of azobenzene and various azobenzene hydrophobic moieties, with the neutral surfactant Triton X 100 (TX 100). Binding isotherms in dilute aqueous solutions were determined by spectrophotometry (to measure the fraction of bound azobenzene) and capillary electrophoresis (to measure the amount of bound TX 100). The degree of binding of TX 100 to AMPs increases markedly with increasing azobenzene hydrophobicity and density in AMPs. A noticeable and reversible photoresponse of the associates was observed upon exposure to UV/visible lights, although the magnitude of the UV-triggered photodissociation and blue-triggered association depends on the chemical structure of both the azobenzene and AMPs. We introduce a critical distance <i>l_c</i> that accounts as single parameter for the balance between energy gain of hydrophobic binding and energy loss due to chain conformational constraints. Only segments of chains flanked with two azobenzene groups at their ends and shorter than <i>l_c</i> are assumed to bind tightly. <i>l_c</i> is used to fit both the maximum fraction of azobenzene transferred into TX 100 micelles (with saturation well below 100% despite the presence of excess free TX 100) and the amount of bound TX 100 as a function of the density of azobenzene in the chains. The model includes the effect of random distribution of azobenzene moieties along the chains. From this analysis, we find criterions for optimization of the photoresponse as a function of the azobenzene hydrophobicity and density in the chain, and the chain length

Permeabilization of lipid membranes and cells by a light-responsive copolymer.

International audienceMembrane permeabilization is achieved via numerous techniques involving the use of molecular agents such as peptides used in antimicrobial therapy. Although high efficiency is reached, the permeabilization mechanism remains global with a noticeable lack of control. To achieve localized control and more gradual increase in membrane perturbation, we have developed hydrophobically modified poly(acrylic acid) amphiphilic copolymers with light-responsive azobenzene hydrophobic moieties. We present evidence for light triggered membrane permeabilization in the presence azobenzene-modified polymers (AMPs). Exposure to UV or blue light reversibly switches the polarity of the azobenzene (cis-trans isomerization) in AMPs, hence controlling AMP-loaded lipid vesicles permeabilization via in situ activation. Release of encapsulated probes was studied by microscopy on isolated AMP-loaded giant unilamellar vesicles (pol-GUVs). We show that in pH and ionic strength conditions that are biologically relevant pol-GUVs are kept impermeable when they contain predominantly cis-AMPs but become leaky with no membrane breakage upon exposure to blue light due to AMPs switch to a trans-apolar state. In addition, we show that AMPs induce destabilization of plasma membranes when added to mammal cells in their trans-apolar state, with no loss of cell viability. These features make AMPs promising tools for remote control of cell membrane permeabilization in mild conditions