Title:
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Studies of cationic amphiphilic drug catalysed membrane degradation
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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.
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