Optimization of Non-Viral Gene Therapeutics Using Bilamellar Invaginated Vesicles.

Bilamellar invaginated vesicles (BIVs) are unique liposomal nanoparticles (NPs) that are highly efficient vehicles for intravenous (iv) delivery of encapsulated therapeutics including plasmid DNA. Systemic administration of therapeutics is required to effectively treat or cure metastatic cancer, certain cardiovascular diseases, and other acquired or inherited diseases. In addition to having extended half-life and stability in circulation, BIVs are nontoxic, nonimmunogenic, biodegradable and can be repeatedly administered without losing potency.

Reversible masking using low-molecular-weight neutral lipids to achieve optimal-targeted delivery.

Intravenous injection of therapeutics is required to effectively treat or cure metastatic cancer, certain cardiovascular diseases, and other acquired or inherited diseases. Using this route of delivery allows potential uptake in all disease targets that are accessed by the bloodstream. However, normal tissues and organs also have the potential for uptake of therapeutic agents.

Oncolytic viruses for induction of anti-tumor immunity.

Oncolytic virotherapy is an evolving but, as yet, unrealized treatment option for cancer. This approach harnesses the cancer-restricted replicative activity of engineered viruses to achieve tumor cell kill. Tumors that are resistant to chemotherapy or radiotherapy can be susceptible to viral oncolysis because of distinct cell kill mechanisms.

Liposomes for gene transfer in cancer therapy.

We developed improved liposomes that produce efficacy for the treatment of cancer, cardiovascular diseases, and HIV-1-related diseases in small and large animal models. Because our processes are reproducible, we have standard operating procedures (SOPs) for the cGMP manufacture of these reagents that have been approved by the Food and Drug Administration for use in phase I/II clinical trials.

Nonviral delivery for genomic therapy of cancer.

We have developed improved liposomal nanoparticles that efficiently condense nucleic acids, proteins, viruses, drugs, and mixtures of these agents on the interior of bilamellar invaginated structures (BIVs) produced by a novel extrusion procedure. The liposomal complexes have extended half-life in the circulation, serum stability, and broad biodistribution; are targetable to specific organs and cell types; can penetrate through tight barriers in several organs; are fusogenic with cell membranes and avoid endosomes; are optimized for nucleic acid:lipid ratio and colloidal suspension in vivo; can be size fractionated to produce totally homogeneous populations of complexes prior to injection; are nontoxic, nonimmunogenic, and can be repeatedly administered; and they are stable in liquid suspensions and freeze-dried formulations. We can add specific ligands either by ionic interactions or by covalent attachments to the surface of these nucleic acid-liposome complexes to accomplish targeted delivery to specific cell surface receptors.

PDX-1 acts as a potential molecular target for treatment of human pancreatic cancer.

Objectives: The purpose of this study was to investigate whether pancreatic and duodenal homeobox factor 1 (PDX-1) could serve as a potential molecular target for the treatment of pancreatic cancer.

Methods: Cell proliferation, invasion capacity, and protein levels of cell cycle mediators were determined in human pancreatic cancer cells transfected with mouse PDX-1 (mPDX-1) alone or with mPDX-1 short hairpin RNA (shRNA) and/or human PDX-1 shRNA (huPDX-1 shRNA). Tumor cell growth and apoptosis were also evaluated in vivo in PANC-1 tumor-bearing severe combined immunodeficient mice receiving multiple treatments of intravenous liposomal huPDX-1 shRNA.

Oral immunization of rhesus macaques with adenoviral HIV vaccines using enteric-coated capsules.

Targeted delivery of vaccine candidates to the gastrointestinal (GI) tract holds potential for mucosal immunization, particularly against mucosal pathogens like the human immunodeficiency virus (HIV). Among the different strategies for achieving targeted release in the GI tract, namely the small intestine, pH sensitive enteric coating polymers have been shown to protect solid oral dosage forms from the harsh digestive environment of the stomach and dissolve relatively rapidly in the small intestine by taking advantage of the luminal pH gradient. We developed an enteric polymethacrylate formulation for coating hydroxy-propyl-methyl-cellulose (HPMC) capsules containing lyophilized Adenoviral type 5 (Ad5) vectors expressing HIV-1 gag and a string of six highly-conserved HIV-1 envelope peptides representing broadly cross-reactive CD4(+) and CD8(+) T cell epitopes.

Widespread, exceptionally high levels of histone H3 lysine 4 trimethylation largely mediate "privileged" gene expression.

We examined the molecular determinants for sustained high-level expression of "privileged" genes, defined as the 0.03% most highly expressed genes within any specific cell. We identified histone modifications by chromatin immunoprecipitation analyses on Keratin 8, the most highly expressed gene in the human breast cancer cell line, MCF-7, based on serial analysis of gene expression.

Optimization of gene expression in nonactivated circulating lymphocytes.

Circulating lymphocytes are important target cells for the treatment of HIV-related and autoimmune diseases and for stimulating anti-tumor immunity. To date, gene transfection of these nonactivated cells after intravenous delivery of viral or nonviral vectors remains low although these circulating cells are highly accessible. Optimized lentiviral vectors currently can transduce less than 10% of nonactivated circulating lymphocytes.

Myths concerning the use of cationic liposomes in vivo.

Varied results have been obtained using cationic liposomes for in vivo delivery. Furthermore, optimisation of cationic liposomal complexes for in vivo applications is complicated, involving many diverse components. These components include nucleic acid purification, plasmid design, formulation of the delivery vehicle, administration route and schedule, dosing, detection of gene expression and others.

Cationic liposomes as in vivo delivery vehicles.

Optimization of cationic liposomal complexes for in vivo applications is complex involving many diverse components. These components include nucleic acid purification, plasmid design, formulation of the delivery vehicle, administration route and schedule, dosing, detection of gene expression, and others. This review will primarily focus on optimization of the delivery vehicle formulation.

Liposomal delivery of nucleic acids in vivo.

Optimization of cationic liposomal complexes for in vivo applications and therapeutics is complex involving many distinct components. These components include nucleic acid purification, plasmid design, formulation of the delivery vehicle, administration route and schedule, dosing, detection of gene expression, and others. This review will focus on optimization of these components for use in a variety of in vivo applications.

Enhanced gene expression in breast cancer cells in vitro and tumors in vivo.

Gene therapy clinical trials for cancer frequently produce inconsistent results. Some of this variability could result from differences in transcriptional regulation that limit expression of therapeutic genes in specific cancers. Systemic liposomal delivery of a nonviral plasmid DNA showed efficacy in animal models for several cancers.

Cationic liposome-mediated gene delivery in vivo.

Several improvements have been made in liposomal delivery, thus making this technology potentially useful for treatment of certain diseases in the clinic. Success in non-viral delivery is complicated and requires optimization of several components. These components include nucleic acid purification, plasmid design, formulation of the delivery vehicle, administration route and schedule, dosing, detection of gene expression, and others.

Identification of heat shock protein 60 as a molecular mediator of alpha 3 beta 1 integrin activation.

The alpha 3 beta 1 integrin is involved in the adhesion of metastatic breast cancer cells to the lymph nodes and to osteoblasts in the bone. Regulation of the affinity or avidity of integrins for their ligands may result from conformational changes induced by changes in the microenvironment of the integrin. Two surface proteins, 55 and 32 kDa, coimmunoprecipitated with the alpha 3 beta 1 integrin from breast carcinoma cells.

Bilamellar cationic liposomes protect adenovectors from preexisting humoral immune responses.

Adenoviral vectors have been widely used for gene therapy, but they are limited both by the presence of a humoral immune response that dramatically decreases the level of transduction after reinjection and by their requirement for target cells to express appropriate receptors such as Coxsackie adenovirus receptor (CAR). To overcome both limits, we encapsulated adenovectors using bilamellar DOTAP:chol liposomes. Electron micrography (EM) showed that these liposomes efficiently encapsulated the vectors, allowing CAR-independent adenovector transduction of otherwise resistant cells.