Kupffer cells do not play a role in governing the efficacy of liposomal mitoxantrone used to treat a tumor model designed to assess drug delivery to liver.

A tumor model designed to assess liposome-mediated drug delivery to liver has been used in an attempt to better understand the mechanism of activity of liposomal mitoxantrone, a liposomal anticancer drug formulation that appears to be uniquely effective in treating this tumor model. Reductions in liposomal mitoxantrone accumulation in the liver were achieved either by use of poly(ethylene)glycol (PEG)-modified lipids or by methods designed to deplete liver phagocytes, a method referred to as hepatic mononuclear phagocytic system (MPS) blockade. A 2-fold reduction in mitoxantrone delivery to the liver was obtained using a mitoxantrone formulation with PEG-modified lipids, and a 3-fold reduction was obtained when liposomal mitoxantrone was given to animals pretreated to induce hepatic MPS blockade.

Liposomal encapsulation of topotecan enhances anticancer efficacy in murine and human xenograft models.

Topotecan was encapsulated in sphingomyelin/cholesterol liposomes using an ionophore-generated proton gradient. After i.v.

In vitro characterization of the anticancer activity of membrane-active cationic peptides. I. Peptide-mediated cytotoxicity and peptide-enhanced cytotoxic activity of doxorubicin against wild-type and p-glycoprotein over-expressing tumor cell lines.

Cationic amphipathic peptides, such as the defensins and cecropins, induce cell death in prokaryotic and eukaryotic cells by increasing membrane permeability. Increased permeability may lead to cell lysis or, alternatively, may produce subtle changes in the membrane's barrier function that promote cell death. The in vitro cytotoxic and lytic activity of short mammalian-derived extended-helical cationic peptides and insect-derived alpha-helical peptides was measured in this study with the objective of establishing the anticancer potential of these agents.

Eradication of human non-Hodgkin's lymphoma in SCID mice by BCL-2 antisense oligonucleotides combined with low-dose cyclophosphamide.

Cancers overexpressing Bcl-2 protein, which prevents programmed cell death (apoptosis), are less sensitive to stresses that produce cellular damage, including chemotherapy. If the level of Bcl-2 protein can be reduced sufficiently using antisense oligonucleotides (ASOs) targeting the gene message, then cytotoxic agents may be rendered more effective in eliminating disease and increasing cure rate. Preclinical studies in SCID mice bearing Bcl-2 overexpressing systemic human B-cell lymphoma (DoHH2) were undertaken to support development of a clinical trial.

Biological barriers to cellular delivery of lipid-based DNA carriers.

Although lipid-based DNA delivery systems are being assessed in gene therapy clinical trials, many investigators in this field are concerned about the inefficiency of lipid-based gene transfer technology, a criticism directed at all formulations used to enhance transfer of plasmid expression vectors. It is important to recognize that many approaches have been taken to improve transfection efficiency, however because of the complex nature of the formulation technology being developed, it has been extremely difficult to define specific carrier attributes that enhance transfection. We believe that these optimization processes are flawed for two reasons.

Clearance properties of liposomes involving conjugated proteins for targeting.

This review examines methods of protein conjugation onto liposomes and the effects of surface bound protein on the liposomes' biological behavior. It is evident that the presence of a conjugated protein significantly alters the attributes of targeted liposomes. Specifically, protein conjugation can result in dramatic increases in liposome size, enhanced immunogenicity, and increased plasma elimination.

Use of poly(ethylene glycol)-lipid conjugates to regulate the surface attributes and transfection activity of lipid-DNA particles.

We evaluated the use of poly(ethylene glycol) (PEG)-modified lipids to control the surface properties of a lipid-based gene transfer system. The lipid-DNA particles (LDPs) used form spontaneously when plasmid DNA is added to mixed detergent lipid micelles consisting of the non-ionic detergent n-octyl-D-glucopyranoside, the cationic lipid dioleyldimethylammonium chloride (DODAC), the zwitterionic lipid 1,2-dioleoyl-sn-glycero-3-phosphoethanolamine (DOPE), and selected PEG-modified phosphatidylethanolamines. The inclusion of DODAC is required to form the hydrophobic lipid-DNA complex.

15 targeted gene transfer : a practical guide based on experience with lipid-based plasmid delivery systems.

The overall goal of gene therapy is to cure or stabilize a disease process that results from the production of a mutant protein (for example, the chloride channel protein important in cystic fibrosis) or overproduction of a normal protein (such as the products of certain oncogenes). We can achieve this goal by replacing the defective gene or by reducing the overexpression of the target gene using an antisense strategy, thus reducing the production of the diseasepromoting protein (1,2). For either method, it is critical to transfer DNA into target cells in a concentration high enough to be effective in modifying the disease.

Antibody conjugation methods for active targeting of liposomes.

Liposomes are useful drug delivery vehicles since they may protect encapsulated drugs from enzymatic degradation and rapid clearance in vivo, or alter biodistribution, potentially leading to reduced toxicities (1,2). A major limitation to the development of many specialized applications is the problem of directing liposomes to tissues where they would not normally accumulate. Consequently, a great deal of effort has been made over the years to develop liposomes that have targeting vectors attached to the bilayer surface.

Role of drug release and liposome-mediated drug delivery in governing the therapeutic activity of liposomal mitoxantrone used to treat human A431 and LS180 solid tumors.

A previous study suggested that drug release is the dominating factor controlling biological activity of liposomal mitoxantrone in tissues where the rate of liposome accumulation is rapid. The studies described here attempted to address the question: under conditions where the rate of liposome accumulation is slow, does drug release or liposome-mediated drug delivery become the dominant factor controlling therapeutic activity? Liposomal mitoxantrone formulations exhibiting different drug-release characteristics were injected i.v.

A multi-step lipid mixing assay to model structural changes in cationic lipoplexes used for in vitro transfection.

Formation of liposome/polynucleotide complexes (lipoplexes) involves electrostatic interactions, which induce changes in liposome structure. The ability of these complexes to transfer DNA into cells is dependent on the physicochemical attributes of the complexes, therefore characterization of binding-induced changes in liposomes is critical for the development of lipid-based DNA delivery systems. To clarify the apparent lack of correlation between membrane fusion and in vitro transfection previously observed, we performed a multi-step lipid mixing assay to model the sequential steps involved in transfection.

Controlled destabilization of a liposomal drug delivery system enhances mitoxantrone antitumor activity.

Programmable fusogenic vesicles (PFVs) are lipid-based drug-delivery systems that exhibit time-dependent destabilization. The rate at which this destabilization occurs is determined by the exchange rate of a bilayer-stabilizing component, polyethylene glycol-phosphatidylethanolamine (PEG-PE) from the vesicle surface. This exchange rate is controlled, in turn, by the acyl chain composition of the PEG-PE.

Electrostatically mediated interactions between cationic lipid-DNA particles and an anionic surface.

In an effort to model the interaction of lipid-based DNA delivery systems with anionic surfaces, such as a cell membrane, we have utilized microelectrophoresis to characterize how electrokinetic measurements can provide information on surface charge and binding characteristics. We have established that cationic lipids, specifically N-N-dioleoyl-N,N-dimethylammonium chloride (DODAC), incorporated into liposomes prepared with 1, 2-dioleoyl-i-glycero-3-phosphoethanolamine (DOPE) or 1, 2-dioleoyl-sn-glycero-3-phosphocholine (DOPC) at 50 mol%, change the inherent electrophoretic mobility of anionic latex polystyrene beads. Self-assembling lipid-DNA particles (LDPs), prepared at various cationic lipid to negative DNA phosphate charge ratios, effected no changes in bead mobility when the LDP charge ratio (+/-) was equal to or less than 1.

ThrH, a homoserine kinase isozyme with in vivo phosphoserine phosphatase activity in Pseudomonas aeruginosa.

Homoserine kinase, the product of the thrB gene, catalyses an obligatory step of threonine biosynthesis. In Pseudomonas aeruginosa, unlike Escherichia coli, inactivation of the previously identified thrB gene does not result in threonine auxotrophy. A new gene, named thrH, was isolated that, when expressed in E.

RpoS-dependent stress tolerance in Pseudomonas aeruginosa.

Pseudomonas aeruginosa is able to persist during feast and famine in many different environments including soil, water, plants, animals and humans. The alternative sigma factor encoded by the rpoS gene is known to be important for survival under stressful conditions in several other bacterial species. To determine if the P.

Liposomal lipid and plasmid DNA delivery to B16/BL6 tumors after intraperitoneal administration of cationic liposome DNA aggregates.

The transfer of plasmid expression vectors to cells is essential for transfection after administration of lipid-based DNA formulations (lipoplexes). A murine i.p.

Phase I study of liposomal vincristine.

Purpose: A phase I study of vincristine encapsulated inside 120-nm-diameter distearoylphosphatidylcholine-cholesterol liposomes was performed. The primary objectives were to determine the maximum-tolerated dose (MTD), recommended phase II dose, toxicity, and pharmacokinetics of liposomal vincristine (ONCO-TCS).

Patients And Methods: Twenty-five patients with histologically confirmed malignancies were enrolled and assessable.

Assembly of XcpR in the cytoplasmic membrane is required for extracellular protein secretion in Pseudomonas aeruginosa.

A broad range of extracellular proteins secreted by Pseudomonas aeruginosa use the type II or general secretory pathway (GSP) to reach the medium. This pathway requires the expression of at least 12 xcp gene products. XcpR, a putative nucleotide-binding protein, is essential for the secretion process across the outer membrane even though the protein contains no hydrophobic sequence that could target or anchor it to the bacterial envelope.

Intravenous pretreatment with empty pH gradient liposomes alters the pharmacokinetics and toxicity of doxorubicin through in vivo active drug encapsulation.

Liposomes have been used widely to improve the therapeutic activity of pharmaceutical agents. The traditional approach for such applications has been to formulate the pharmaceutical agent in liposomes prior to administration in vivo. In this report we demonstrate that liposomes exhibiting a transmembrane pH gradient injected intravenously (iv) can actively encapsulate doxorubicin in the circulation after iv administration of free drug.

GSP-dependent protein secretion in gram-negative bacteria: the Xcp system of Pseudomonas aeruginosa.

Bacteria have evolved several secretory pathways to release proteins into the extracellular medium. In Gram-negative bacteria, the exoproteins cross a cell envelope composed of two successive hydrophobic barriers, the cytoplasmic and outer membranes. In some cases, the protein is translocated in a single step across the cell envelope, directly from the cytoplasm to the extracellular medium.

Preclinical pharmacology, toxicology and efficacy of sphingomyelin/cholesterol liposomal vincristine for therapeutic treatment of cancer.

Purpose: To establish the pharmacodynamic relationships between drug biodistribution and drug toxicity/efficacy, a comprehensive preclinical evaluation of sphingomyelin/cholesterol (SM/chol) liposomal vincristine and unencapsulated vincristine in mice was undertaken.

Methods: Pharmaceutically acceptable formulations of unencapsulated vincristine and liposomal vincristine at drug/lipid ratios of 0.05 or 0.

Comparison of different hydrophobic anchors conjugated to poly(ethylene glycol): effects on the pharmacokinetics of liposomal vincristine.

Poly(ethylene glycol) (PEG) conjugated lipids have been used to increase the circulation longevity of liposomal carriers encapsulating therapeutic compounds. PEG is typically conjugated to distearoylphosphatidylethanolamine (DSPE) via a carbamate linkage that results in a net negative charge on the phosphate moiety at physiological pH. It was anticipated that the presence of this negative charge could have deleterious effects on liposome pharmacokinetic characteristics.

Characterization of lipid DNA interactions. I. Destabilization of bound lipids and DNA dissociation.

We have recently described a method for preparing lipid-based DNA particles (LDPs) that form spontaneously when detergent-solubilized cationic lipids are mixed with DNA. LDPs have the potential to be developed as carriers for use in gene therapy. More importantly, the lipid-DNA interactions that give rise to particle formation can be studied to gain a better understanding of factors that govern lipid binding and lipid dissociation.

Pharmacokinetic behavior of vincristine sulfate following administration of vincristine sulfate liposome injection.

The pharmacokinetic behavior of vincristine sulfate (VINC) following administration of vincristine sulfate liposome injection (VSLI), 0.16 mg/ml, as an intravenous infusion over 60 min in 24 of 25 patients enrolled in a phase I clinical study of this drug is described. Plasma samples for determination of the pharmacokinetic behavior of VINC were collected during the infusion at 15, 30 and 60 min as well as at 2, 4, 8, 12, 48 and 72 h postinfusion.

An immune response to ovalbumin covalently coupled to liposomes is prevented when the liposomes used contain doxorubicin.

It is now well established that liposomes with surface associated proteins are immunogenic. Repeated administration of protein coated liposomes elicits the generation of antibodies and the elimination of proteoliposome increases markedly in animals 'immunized' with such liposomes. This immune response compromises the therapeutic potential of liposomal formulations that rely on the use of protein- or peptide-based targeting ligands to enhance cell specificity.