Biomedical subjects

Katarina Edwards

Publications and source records attributed to Katarina Edwards.

30 records · Page 2Linked to original sources

PubMed2004

An evaluation of transmembrane ion gradient-mediated encapsulation of topotecan within liposomes.

Topotecan can be encapsulated in liposomes, however little is known about the role encapsulated counter ions play in drug loading efficiency and drug release. Using 1,2-distearoyl-sn-glycero-3 phosphatidylcholine and cholesterol liposomes (55:45 mole ratio), encapsulation was achieved using manganese ion gradients (MnSO(4) or MnCl(2)), with the addition of A23187, a divalent cation/proton exchanger, to maintain a pH gradient. This methodology was compared to procedures where the pH gradient was generated by use of encapsulated (NH(4))(2)SO(4) or citrate (300 mM, pH 3.5). All methods facilitated topotecan encapsulation. Liposomes prepared in the presence of the citrate and MnCl(2) (+A23187) exhibited reduced loading capacities. Liposomes prepared in the presence of (NH(4))(2)SO(4) and MnSO(4) (+A23187) could be used to generate liposomes exhibiting a drug-to-lipid ratio of 0.3 (wt/wt) with an encapsulation efficiency of >90%. In vitro drug release data suggested that the (NH(4))(2)SO(4) and MnSO(4) (+A23187) formulations released drug at a reduced rate. For these formulations, the drug release rates decreased as the drug-to-lipid ratio (wt/wt) increased from 0.1 to 0.2. Cryo-electron micrographs indicated that encapsulated topotecan precipitated as linear particles within liposomes. The stability of topotecan loaded liposomes appeared to be dependent on the presence of both a pH gradient and encapsulated sulfate.

Ammonium Sulfate↗

PubMed2004

pH gradient loading of anthracyclines into cholesterol-free liposomes: enhancing drug loading rates through use of ethanol.

Application of cholesterol-free liposomes as carriers for anticancer drugs is hampered, in part, because of standard pH gradient based loading methods that rely on incubation temperatures above the phase transition temperature (Tc) of the bulk phospholipid to promote drug loading. In the absence of cholesterol, liposome permeability is enhanced at these temperatures which, in turn, can result in the collapse of the pH gradient and/or unstable loading. Doxorubicin loading studies, for example, indicate that the drug could not be loaded efficiently into cholesterol-free DSPC liposomes. We demonstrated that this problem could be circumvented by the addition of ethanol as a permeability enhancer. Doxorubicin loading rates in cholesterol-free DSPC liposomes were 6.6-fold higher in the presence of ethanol. In addition, greater than 90% of the added doxorubicin was encapsulated within 2 h at 37 degrees C, an efficiency that was 2.3-fold greater than that observed in the absence of ethanol. Optimal ethanol concentrations ranged from 10% to 15% (v/v) and these concentrations did not significantly affect liposome size, retention of an aqueous trap marker (lactose) or, most importantly, the stability of the imposed pH gradient. Cryo-transmission electron micrographs of liposomes exposed to increasing concentrations of ethanol indicated that at 30% (v/v) perturbations to the lipid bilayer were present as evidenced by the appearance of open liposomes and bilayer sheets. Ethanol-induced increased drug loading was temperature-, lipid composition- and lipid concentration-dependent. Collectively, these results suggest that ethanol addition to preformed liposomes is an effective method to achieve efficient pH gradient-dependent loading of cholesterol-free liposomes at temperatures below the Tc of the bulk phospholipid.

Animals↗

PubMed2003

Disc formation in cholesterol-free liposomes during phase transition.

Cryogenic transmission electron microscopy (cryo-TEM) images of lysolipid-containing thermosensitive liposomes (LTSL) revealed that open liposomes and bilayer discs appeared when liposomes were cycled through the gel (Lbeta') to liquid-crystalline (Lalpha) phase transition. The amount of bilayer discs generated was dependent on the combined presence of PEG-lipid and lysolipid in the membrane. We hypothesize that micelle-forming membrane components stabilize the rim of bilayer openings and membrane discs that form when liposomes are cycled through TC.

Cryoelectron Microscopy↗

PubMed2003

Evidence of surfactant-induced formation of transient pores in lipid bilayers by using magnetic-fluid-loaded liposomes.

It is often assumed that surfactant-induced permeability of lipid membranes obeys a pore-formation mechanism, but, to date, this has not been totally proven. A novel approach is developed using a magnetic fluid composed of calibrated nanocrystals of maghemite (gamma-Fe2O3) as a permeability marker. It is shown that low amounts of surfactant molecules catalyze the transient opening of unilamellar phospholipid vesicles which permit the passage of 8 nm maghemite nanospheres before closing up.

Chromatography, Gel↗

PubMed2003

Interactions between pH-sensitive liposomes and model membranes.

The structure and dynamics of two different pH-sensitive liposome systems were investigated by means of cryo-transmission electron microscopy and different photophysical techniques. Both systems consisted of dioleoylphosphatidylethanolamine (DOPE) and contained either oleic acid (OA) or a novel acid-labile polyethylene glycol-conjugated lipid (DHCho-MPEG5000) as stabiliser. Proton induced leakage, lipid mixing and structural changes were studied in the absence and presence of EPC liposomes, as well as in the presence of liposomes designed to model the endosome membrane. Neither DHCho-MPEG5000- nor OA-stabilised liposomes showed any tendency for fusion with pure EPC liposomes or endosome-like liposomes composed of EPC/DOPE/SM/Cho (40/20/6/34 mol.%). Our investigations showed, however, that incorporation of lipids from the pH-sensitive liposomes into the endosome membrane may lead to increased permeability and formation of non-lamellar structures. Taken together the results suggest that the observed ability of DOPE-containing liposomes to mediate cytoplasmic delivery of hydrophilic molecules cannot be explained by a mechanism based on a direct, and non-leaky, fusion between the liposome and endosome membranes. A mechanism involving destabilisation of the endosome membrane due to incorporation of DOPE, seems more plausible.

Cell Membrane↗

PubMed2003

Liposomes, disks, and spherical micelles: aggregate structure in mixtures of gel phase phosphatidylcholines and poly(ethylene glycol)-phospholipids.

Poly(ethylene glycol) (PEG) decorated lipid bilayers are widely used in biomembrane and pharmaceutical research. The success of PEG-lipid stabilized liposomes in drug delivery is one of the key factors for the interest in these polymer/lipid systems. From a more fundamental point of view, it is essential to understand the effect of the surface grafted polymers on the physical-chemical properties of the lipid bilayer. Herein we have used cryo-transmission electron microscopy and dynamic light scattering to characterize the aggregate structure and phase behavior of mixtures of PEG-lipids and distearoylphosphatidylcholine or dipalmitoylphosphatidylcholine. The PEG-lipids contain PEG of molecular weight 2000 or 5000. We show that the transition from a dispersed lamellar phase (liposomes) to a micellar phase consisting of small spherical micelles occurs via the formation of small discoidal micelles. The onset of disk formation already takes place at low PEG-lipid concentrations (<5 mol %) and the size of the disks decreases as more PEG-lipid is added to the lipid mixture. We show that the results from cryo-transmission electron microscopy correlate well with those obtained from dynamic light scattering and that the disks are well described by an ideal disk model. Increasing the temperature, from 25 degrees C to above the gel-to-liquid crystalline phase transition temperature for the respective lipid mixtures, has a relatively small effect on the aggregate structure.

Biophysical Phenomena↗

PubMed2003

Linkage identity is a major factor in determining the effect of PEG-ylated surfactants on permeability of phosphatidylcholine liposomes.

The permeability effects induced by single-chained and double-chained poly(ethylene glycol)-surfactants were investigated by measuring the leakage of the fluorescent dye 5(6)-carboxy fluorescein from EPC liposomes. The standard incorporated amount of the surfactants was 5 mol%. Depending on the size of the poly(ethylene glycol) chain and especially on the type of linkage between the polymer and the hydrophobic moiety different leakage profiles were obtained. The presence of a long PEG-polymer resulted in a slower leakage compared with a short analogue. More importantly, the linkage identity was decisive for whether an overall reduction or increase in permeability was obtained. When the hydrocarbon chains were attached to the PEG chain via an ether or an ester the leakage increased compared to pure EPC liposomes. In contrast, if the link was an amide, the leakage was significantly reduced. This effect is assumed to originate from headgroup-headgroup interactions, and most probably hydrogen bonding, between amide and phosphate groups of the PEG-surfactant and the EPC, respectively.

Hydrogen Bonding↗

PubMed2003

Catanionic drug-surfactant mixtures: phase behavior and sustained release from gels.

PURPOSE: To study mixtures of SDS and the drugs diphenhydramine, tetracaine, and amitriptyline to compile phase diagrams and to investigate the use of interesting phases for sustained release from gels. METHODS: Phase diagrams were composed by studying large numbers of different compositions of negatively charged SDS and positively charged drug compounds visually, rheologically, and by cryotransmission electron microscopy. Drug release from Carbopol 940 and agar gels containing interesting phases, e.g., vesicle and branched micelle phases, was measured in vitro by the USP paddle method. RESULTS: Vesicles and elongated and branched micelles were formed on the SDS-rich side in all three systems examined. The tetracaine system differed from the other two in that it showed a vesicle area in the drug-rich side. Release of diphenhydramine from Carbopol 940 gels was slowed by at least a factor of 10 when in the form of vesicles or branched micelles. The same delay was found for both drug-rich and SDS-rich tetracaine vesicles. CONCLUSIONS: Mixtures of SDS and positively charged drugs form the same interesting phases as traditional catanionic mixtures. This may prove useful in obtaining functional controlled-release systems when using gels as drug carriers.

Amitriptyline↗

PubMed2003

Introductory experiments on ligand liposomes as delivery agents for boron neutron capture therapy.

Liposomes are, when coupled to receptor ligands, candidates for receptor mediated delivery of boron for tumour therapy since they have capacity to deliver large amounts of boron per receptor interaction. With EGF-liposomes we present a pegylated ligand liposome delivery vehicle, containing water soluble boronated phenanthridine, WSP1, or water soluble boronated acridine, WSA1, for EGFR targeting. In the case of WSA1 a ligand dependent uptake was obtained and the boron uptake was as good as if free WSA1 was given. No ligand dependent boron uptake was seen for WSP1 containing liposomes. Thus, WSA1 is a candidate for further studies. Approximately 10(5) boron atoms were in each liposome. A critical assessment indicates that after optimization up to 10(6) boron atoms can be loaded. Since it is known that, for therapeutic effect, approximately 10(8)-10(9) boron atoms are needed in a single tumour cell it is realized that 10(2)-10(3) receptor interactions are needed to meet the demand. Tests applying cultured glioma cells indicate, without optimization of the delivery conditions, a boron uptake in the ppm range, which is necessary for successful BNCT. Thus, it seems possible to kill micro-invasive tumour cells with targeted liposomes if the delivery conditions are optimal.

Acridines↗

PubMed2002

Formation of transition metal-doxorubicin complexes inside liposomes.

Doxorubicin complexation with the transition metal manganese (Mn(2+)) has been characterized, differentiating between the formation of a doxorubicin-metal complex and doxorubicin fibrous-bundle aggregates typically generated following ion gradient-based loading procedures that rely on liposome encapsulated citrate or sulfate salts. The physical and chemical characteristics of the encapsulated drug were assessed using cryo-electron microscopy, circular dichroism (CD) and absorbance spectrophotometric analysis. In addition, in vitro and in vivo drug loading and release characteristics of the liposomal formulations were investigated. Finally, the internal pH after drug loading was measured with the aim of linking formation of the Mn(2+) complex to the presence or absence of a transmembrane pH gradient. Doxorubicin was encapsulated into either 1,2-dimyristoyl-sn-glycero-3-phosphocholine (DMPC)/cholesterol (Chol) or 1,2-distearoyl-sn-glycero-3-phosphocholine (DSPC)/Chol liposomes, where the entrapped salts were citrate, MnSO(4) or MnCl(2). In response to a pH gradient or a Mn(2+) ion gradient, doxorubicin accumulated inside to achieve a drug-to-lipid ratio of approximately 0.2:1 (wt/wt). Absorbance and CD spectra of doxorubicin in the presence of Mn(2+) suggested that there are two distinct structures captured within the liposomes. In the absence of added ionophore A23187, drug loading is initiated on the basis of an established pH gradient; however, efficient drug uptake is not dependent on maintenance of the pH gradient. Drug release from DMPC/Chol is comparable regardless of whether doxorubicin is entrapped as a citrate-based aggregate or a Mn(2+) complex. However, in vivo drug release from DSPC/Chol liposomes indicate less than 5% or greater than 50% drug loss over a 24-h time course when the drug was encapsulated as an aggregate or a Mn(2+) complex, respectively. These studies define a method for entrapping drugs possessing coordination sites capable of complexing transition metals and suggest that drug release is dependent on lipid composition, internal pH, as well as the nature of the crystalline precipitate, which forms following encapsulation.

Animals↗

PubMedDate not supplied

Ligand liposomes and boron neutron capture therapy.

Boron neutron capture therapy (BNCT) has been used both experimentally and clinically for the treatment of gliomas and melanomas, with varying results. However, the therapeutic effects on micro-invasive tumor cells are not clear. The two drugs that have been used clinically, p-boronophenylalanine, (BPA), and the sulfhydryl borane, (BSH), seem to be taken up preferentially in solid tumor areas but it is uncertain whether enough boron is taken up by micro-invasive tumor cells. To increase the selective uptake of boron by such cells, would be to exploit tumor transformation related cellular changes such as over-expression of growth factor receptors. However, the number of receptors varies from small to large and the uptake of large amounts of boron for each receptor interaction is necessary in order to deliver sufficient amounts of boron. Therefore, each targeting moiety must deliver large number of boron atoms. One possible way to meet these requirements would be to use receptor-targeting ligand liposomes, containing large number of boron atoms. This will be the subject of this review and studies of boron containing liposomes, with or without ligand, will be discussed. Two recent examples from the literature are ligand liposomes targeting either folate or epidermal growth factor (EGF) receptors on tumor cells. Other potential receptors on gliomas include PDGFR and EGFRvIII. Besides the appropriate choice of target receptor, it is also important to consider delivery of the ligand liposomes, their pharmacodynamics and pharmacokinetics and cellular processing, subjects that also will be discussed in this review.

Animals↗

PubMedDate not supplied

Development of EGF-conjugated liposomes for targeted delivery of boronated DNA-binding agents.

Liposomes are of interest as drug delivery tools for therapy of cancer and infectious diseases. We investigated conjugation of epidermal growth factor, EGF, to liposomes using the micelle-transfer method. EGF was conjugated to the distal end of PEG-DSPE lipid molecules in a micellar solution and the EGF-PEG-DSPE lipids were then transferred to preformed liposomes, either empty or containing the DNA-binding compound, water soluble acridine, WSA. We found that the optimal transfer conditions were a 1-h incubation at 60 degrees C. The final conjugate, (125)I-EGF-liposome-WSA, contained approximately 5 mol % PEG, 10-15 EGF molecules at the liposome surface, and 10(4) to 10(5) encapsulated WSA molecules could be loaded. The conjugate was shown to have EGF-receptor-specific cellular binding in cultured human glioma cells.

Acridines↗