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Biomedical subjects

Gert Storm

Publications and source records attributed to Gert Storm.

At least 73 records · Page 4Linked to original sources
PubMed2003

Liposome-mediated targeting of enzymes to cancer cells for site-specific activation of prodrugs: comparison with the corresponding antibody-enzyme conjugate.

PURPOSE: Immunoenzymosomes are tumor-targeted immunoliposomes bearing enzymes on their surface. These enzymes are capable of converting relatively nontoxic prodrugs into active cytostatic agents. The aims of this study were to compare the enzyme delivery capability of immunoenzymosomes with that of the corresponding antibody-enzyme conjugate and to evaluate whether immunoenzymosomes are able to mount a strong bystander effect. METHODS: Immunoenzymosomes exposing Fab' fragments of the monoclonal antibody 323/A3 and the bacterial enzyme beta-glucuronidase or the corresponding antibody-enzyme conjugate were incubated with OVCAR-3 cells (human ovarian carcinoma cells). Cell-associated enzymatic activity and the in vitro antiproliferative effect of a glucuronide prodrug of doxorubicin (DOX-GA3) were determined. RESULTS: At equal numbers of carrier units, the cell-associated enzymatic activity achieved by using immunoenzymosomes was 15-fold higher than that obtained after incubation with the corresponding antibody-enzyme conjugate. Increasing the amount of antibody-enzyme conjugate added to the cells could not compensate for their lower enzyme delivery capability. Immunoenzymosomes were able to induce inhibition of cell growth not only of tumor cells to which immunoenzymosomes were bound but also of a large number of neighboring cells. CONCLUSIONS: Immunoenzymosomes are able (a) to target prodrug-converting enzymes more efficiently to tumor cells than the corresponding antibody-enzyme conjugate and (b) to mount a strong bystander effect.

Animals↗
PubMed2002

Superoxide dismutase entrapped in long-circulating liposomes: formulation design and therapeutic activity in rat adjuvant arthritis.

The aim of this study was to investigate whether long-circulating liposomes can improve the anti-inflammatory activity of superoxide dismutase (SOD). Small-sized poly(ethyleneglycol) (PEG)-liposomes containing SOD were prepared via different preparation protocols and characterized in terms of encapsulation efficiency (EE), size, enzymatic activity and protein structure, to establish conditions where high EE can be combined with preservation of enzyme activity and structure. It was observed that structural information from circular dichroism analyses does not correlate with data on enzyme activity. SOD-containing PEG-liposomes prepared by the dehydration-rehydration method appeared to represent the most attractive formulation for in vivo evaluation. The therapeutic potential of selected SOD-containing PEG-liposomes was established and compared with SOD entrapped in stearylamine (SA)-liposomes and 'free' SOD upon intravenous (i.v.) injection in an arthritic rat model. Both small PEG-liposomes and SA-liposomes showed a superior therapeutic activity compared to 'free' SOD, with PEG-liposomes inducing stronger anti-inflammatory effects than SA-liposomes.

Amines↗
PubMed2002

Functional characterization of an endosome-disruptive peptide and its application in cytosolic delivery of immunoliposome-entrapped proteins.

Antibody-directed liposomes (immunoliposomes) are frequently used for targeted drug delivery. However, delivery of large biotherapeutic molecules (i.e. peptides, proteins, or nucleic acids) with immunoliposomes is often hampered by an inefficient cytosolic release of entrapped macromolecules after target cell binding and subsequent endocytosis of immunoliposomes. To enhance cytosolic drug delivery from immunoliposomes present inside endosomes, a pH-dependent fusogenic peptide (diINF-7) resembling the NH(2)-terminal domain of influenza virus hemagglutinin HA-2 subunit was used. Functional characterization of this dimeric peptide showed its ability to induce fusion between liposome membranes and leakage of liposome-entrapped compounds when exposed to low pH. In a second series of experiments, diINF-7 peptides were encapsulated in immunoliposomes to enhance the endosomal escape of diphtheria toxin A chain (DTA), which inhibits protein synthesis when delivered into the cytosol of target cells. Immunoliposomes targeted to the internalizing epidermal growth factor receptor on the surface of ovarian carcinoma cells (OVCAR-3) and containing encapsulated DTA did not show any cytotoxicity toward OVCAR-3 cells. Cytotoxicity was only observed when diINF-7 peptides and DTA were co-encapsulated in the immunoliposomes. Thus, diINF-7 peptides entrapped inside liposomes can greatly enhance cytosolic delivery of liposomal macromolecules by pH-dependent destabilization of endosomal membranes after cellular uptake of liposomes.

Circular Dichroism↗
PubMed2002

Endothelial cells at inflammatory sites as target for therapeutic intervention.

In the course of an inflammation, vascular endothelial cells (VECs) are strongly involved in processes like leukocyte recruitment, cytokine production, and angiogenesis. Specific interference in these processes may yield great therapeutic benefit in the treatment of (chronic) inflammatory disorders. Drug targeting to VECs at inflamed sites may allow such intervention. VECs at inflamed sites represent a very well-accessible target cell population for circulating drug-targeting systems, which may also be selectively distinguished from normal VECs by the expression of several cell surface receptors involved in the inflammation. One group of specifically expressed molecules are the adhesion molecules (AMs), which have a major function in adhesion of cells to each other, to the extracellular matrix, or in the adhesion and subsequent recruitment of circulating immune cells. This review describes AMs with regard to their function in the inflammatory disease and their usefulness in functioning as a specific target receptor for drug-targeting approaches in general and with an emphasis on liposome-based drug delivery.

Animals↗
PubMedDate not supplied

A novel family of L-amino acid-based biodegradable polymer-lipid conjugates for the development of long-circulating liposomes with effective drug-targeting capacity.

The objective of this study was to develop biodegradable polypeptide-lipid conjugates for the design of polymer-coated long-circulating liposomes (LCL). Lipid conjugates of poly(hydroxyalkyl L-asparagine/L-glutamine) were synthesized and incorporated into 0.15 microm dipalmitoyl phosphatidylcholine (DPPC)-cholesterol liposomes. Circulation times and biodistribution were assessed in rats using a radioactive lipid marker. Evaluation of the therapeutic activity of prednisolone phosphate loaded in 0.1 microm PHEA-DPPC-cholesterol liposomes in a rat experimental arthritis model was performed to demonstrate the drug-targeting potential of the polymer-coated liposomes. Coating of liposomes with poly(hydroxyethyl L-asparagine) (PHEA) and poly(hydroxyethyl L-glutamine) (PHEG) extended the circulation half-life to a similar extent as poly(ethylene glycol) (PEG), which is normally used for the preparation of LCL. Glutamine polymers with a hydroxypropyl or a hydroxybutyl group instead of hydroxyethyl group also yield prolonged circulation, however, not to the same extent as PHEA/G. The pharmacokinetic properties of PHEA-liposomes were independent of the lipid dose even at very low lipid doses of around 50 nmol per rat. PLP was successfully entrapped in PHEA-liposomes. These liposomes were shown to be stable in the circulation and equally effective in rat experimental arthritis as PLP encapsulated in PEG-liposomes. PHEA and PHEG are attractive alternative polymers for the design of LCL: their performance is similar to that of PEG-liposomes but they have the advantage of being biodegradable.

1,2-Dipalmitoylphosphatidylcholine↗
PubMedDate not supplied

Enzymatic degradation of liposome-grafted poly(hydroxyethyl L-glutamine).

Liposomes coated with poly(hydroxyethyl L-glutamine) (PHEG) show prolonged circulation times and biodistribution patterns comparable to PEG-coated liposomes. While PEG is a nondegradable polymer, PHEG is expected to be hydrolyzed by proteases. In this study the enzymatic degradability of PHEG both in its free form and grafted onto liposomes was investigated, using the proteases papain, pronase E, and cathepsin B. Enzymatic action was monitored with a ninhydrin assay, which quantifies amine groups formed due to hydrolysis of amide bonds, and the degradation products were characterized by MALDI-ToF mass spectrometry. PHEG, both in its free form and when grafted onto liposomes, showed degradation into low molecular weight peptides by the enzymes. Thus, we present a polymer-coated long-circulating liposome with an enzymatically degradable coating polymer, avoiding the risk of cellular accumulation.

Enzymes↗
PubMedDate not supplied

A liposomal system for contrast-enhanced magnetic resonance imaging of molecular targets.

Pegylated paramagnetic and fluorescent immunoliposomes were designed to enable the parallel detection of the induced expression of molecular markers on endothelial cells with magnetic resonance imaging (MRI) and fluorescence microscopy. MRI is capable of three-dimensional noninvasive imaging of opaque tissues at near cellular resolution, while fluorescence microscopy can be used to investigate processes at the subcellular level. As a model for the expression of a molecular marker, human umbilical vein endothelial cells (HUVEC) were treated with the pro-inflammatory cytokine tumor necrosis factor alpha (TNFalpha) to upregulate the expression of the adhesion molecule E-selectin/CD62E. E-selectin-expressing HUVEC were incubated with pegylated paramagnetic fluorescently labeled liposomes carrying anti-E-selectin monoclonal antibody as a targeting ligand. Both MRI and fluorescence microscopy revealed the specific association of the liposomal MR contrast agent with stimulated HUVEC. This study suggests that this newly developed system may serve as a useful diagnostic tool to investigate pathological processes in vivo with MRI.

Antibodies↗
PubMedDate not supplied

Annexin A5-functionalized bimodal lipid-based contrast agents for the detection of apoptosis.

Apoptosis, or programmed cell death, plays an important role in the etiology of a variety of diseases, including cancer and myocardial infarction. Visualization of apoptosis would allow both early detection of therapy efficiency and evaluation of disease progression. To that aim, we synthesized two types of lipid-based bimodal contrast agents that enable the detection of apoptotic cells with both MRI and optical techniques. MR contrast was provided either by entrapment of iron oxide particles within pegylated micelles or by incorporation of Gd-DTPA-bis(stearylamide) (Gd-DTPA-BSA) lipids within the lipid bilayer of pegylated liposomes. The resulting contrast agents were approximately 10 and 100 nm in diameter, respectively. Additional fluorescent lipids were incorporated in the lipid (bi)layer of the contrast agents to allow parallel detection with optical methods. Multiple human recombinant annexin A5 molecules were covalently coupled to introduce specificity for apoptotic cells. Both annexin A5-conjugated contrast agents were shown to significantly increase the relaxation rates of apoptotic cell pellets compared to untreated control cells and apoptotic cells that were treated with nonfunctionalized nanoparticles. Increased relaxation rates were confirmed to originate from association of the contrast agents to apoptotic cells by confocal microscopy. The targeted nanoparticles presented in this study, which differ both in size and in magnetic properties, may have applications for the in vivo detection of apoptosis.

Annexin A5↗
PubMedDate not supplied

1H NMR spectroscopy as a tool for determining the composition of poly(hydroxyethyl-L-asparagine)-coated liposomes.

Liposomes coated with the poly(amino acid) poly(hydroxyethyl-L-asparagine) (PHEA) show long-circulation properties comparable to the frequently used PEG-liposomes. The pharmacokinetic characteristics of long-circulating liposomes are dependent on the density of the shielding polymer on the liposome surface. Therefore, it is necessary to know the exact composition of the liposomes including the amount of coating polymer present on the liposome surface. In this study, a 1H NMR method to establish the composition of liposomes coated with PHEA was developed and validated.

Asparagine↗
PubMedDate not supplied

Annexin A5-conjugated quantum dots with a paramagnetic lipidic coating for the multimodal detection of apoptotic cells.

Apoptosis, or programmed cell death, plays an important role in the etiology of a variety of diseases, including cancer. Visualization of apoptosis would allow both early detection of therapy efficiency and evaluation of disease progression. To that aim we developed a novel annexin A5-conjugated bimodal nanoparticle. The nanoparticle is composed of a quantum dot that is encapsulated in a paramagnetic micelle to enable its use both for optical imaging and MRI. Multiple recombinant human annexin A5 protein molecules were covalently coupled to the nanoparticle for targeting. In this study the specificity of the annexin A5-conjugated nanoparticles for apoptotic cells was demonstrated both with fluorescence microscopy and MRI, which confirms its potential for the detection of apoptosis with both imaging modalities in vivo.

Annexin A5↗
PubMedDate not supplied

Targeted liposomes for delivery of protein-based drugs into the cytoplasm of tumor cells.

Our goal was to deliver therapeutically active macromolecules into the cytosol of target cells. First, attempts were made to prepare virosomes that specifically interact with OVCAR-3 cells (human ovarian cancer cells). Detergent solubilized influenza virus envelopes were reconstituted forming virosomes. Cell specificity was introduced by incorporating PEG-derivatized lipids with mAB 323/A3 (Fab' fragments) connected to their distal PEG end. These cell-specific, modified virosomes maintained their fusogenic activity when lowering the pH. Most importantly, antibody-mediated binding was a prerequisite for low-pH induced membrane fusion. However, basically, there are two problems with this approach: (1) these virosomes are quite leaky and (2) virosomes can be expected to be immunogenic. A solution to tackle leakage and potential immunogenicity of these site-specific liposomal structures is to use immuno-PEG-liposomes with a pH-dependent fusogen inside the liposome. The system that we designed to test this concept consisted of (1) the fusogenic di-peptide dINF-7, (2) the monoclonal antibody 425 connected to the distal end of PEG-PE (for site specific binding and endosomal uptake), (3) diphtheria toxin chain A (DTA, as carrier-dependent active compound) and phosphatidylcholine/cholesterol as 'bilayer backbone'. A series of tests were performed to show that selective binding and pH-dependent destabilization of (endosomal) membranes indeed occurred. To test the cytotoxic activity of these DTA loaded liposomes, OVCAR-3 cells were used for testing. OVCAR-3 cells express the epidermal growth factor receptor, which is the ligand for antibody 425. In vitro, these site specific and fusogenic liposomes showed a remarkable, cell specific cytotoxic effect.

Cytoplasm↗
PubMedDate not supplied

Liposome-enabled synergistic interaction of antimicrobial agents.

Antimicrobial agents may interact synergistically when both drugs are present at the infected site for an adequate period of time at sufficient concentrations. Generally speaking, the agents in the combination show different tissue distributions and pharmacokinetics. By co-encapsulation of the drugs in a drug carrier, like liposomes, parallel tissue distributions of both drugs may be ensured and drug concentrations at the site of infection may be increased. In this presentation therapeutic efficacy of liposome-co-encapsulated gentamicin (GN) and ceftazidime (CZ) will be shown in a GN-CZ-susceptible and GN-CZ-resistant Klebsiella pneumoniae-pneumonia in rats.

Animals↗
PubMedDate not supplied

Ligand-targeted liposomes directed against pathological vasculature.

The development of liposomes targeted to angiogenic endothelial cells offers exciting prospects for intervention in cancer and inflammation. Several proteins are (strongly) over-expressed on angiogenic endothelial cells as compared to the quiescent endothelium, and could potentially serve as targets for site-specific drug delivery. In this contribution particular attention is given to the design of targeted long-circulating liposomes directed against the alpha v beta 3-integrin protein.

Cells, Cultured↗
PubMedDate not supplied

Administration of prednisolone phosphate-liposomes reduces wound contraction in a rat partial-thickness wound model.

Macrophages play an important role in the inflammatory phase of wound healing and their activity regulates fibroblasts and keratinocytes. Modulation of macrophage function may result in improvement of the wound healing process. Prednisolone phosphate (PLP) encapsulated into liposomes was administered to partial-thickness wounds in rats. A single dose of 75 microg/kg, applied directly after wounding, resulted in up to a 30% reduction of wound contraction at 28 days after wounding. This effect could not be achieved in the group that was administered free PLP or liposomes containing phosphate-buffered saline to the wound. The number of myofibroblasts was up to 50% lower in wounds treated with the liposomal PLP at 4 days after wounding. The number of macrophages present in the wounds was not statistically different between groups. Most probably, the production of cytokines and growth factors by macrophages is altered after phagocytosing the liposomes, resulting in reduced wound contraction.

Administration, Topical↗