Leland Stanford Junior University

The Brigham and Women's Hospital, Inc.

Gary H Gibbons

Victor J Dzau

John P Cooke

Michael J Mann

Ryuichi Morishita

Frank P Diet

Matthew J Pollman

Heiko Von Der Leyen

The invention provides for the use of oligodeoxynucleotide decoys for the prophylactic or therapeutic treatment of diseases associated with the binding of endogenous transcription factors to genes involved in cell growth, differentiation and signalling or to viral genes. By inhibiting endogenous trans-activating factors from binding transcription regulatory regions, the decoys modulate gene expression and thereby regulating pathological processes including inflammation, intimal hyperplasia, angiogenesis, neoplasia, immune responses and viral infection. The decoys are administered in amounts and under conditions whereby binding of the endogenous transcription factor to the endogenous gene is effectively competitively inhibited without significant host toxicity. The subject compositions comprise the decoy molecules in a context which provides for pharmacokinetics sufficient for effective therapeutic use.

Therapeutic use of cis-element decoys in vivo

Therapeutic use of CIS-element decoys in vivo

Vascular function and structure is maintained or improved by long term administration of physiologically acceptable compounds which enhance the level of endogenous nitric oxide or other intermediates in the NO induced relaxation pathway in the host. Alternatively, or in combination, other compounds may be administered which provide for short term enhancement of nitric oxide, either directly or by physiological processes.

Enhancement of vascular function by modulation of endogenous nitric oxide production or activity

Vascular function and structure is maintained or improved by long term administration of physiologically acceptable compounds which enhance the level of endogenous nitric oxide or other intermediates in the NO induced relaxation pathway in the host. Alternatively, or in combination, other compounds may be administered which provide for short term enhancement of nitric oxide, either directly or by physiological processes

Atherogenesis and restenosis are treated by long term administration of physiologically acceptable compounds which enhance the level of endogenous nitric oxide or other intermediates in the NO induced relaxation pathway in the host. Alternatively, or in combination, other compounds may be administered which provide for short term enhancement of nitric oxide, either directly or by physiological processes. In addition, cells may be genetically engineered to provide a component in the synthetic pathway to nitric oxide, so as to drive the process to enhance nitric oxide concentration, particularly in conjunction with the administration of a nitric oxide precursor.

Treatment of vascular degenerative diseases by modulation of endogenous

Naked nucleic acids (DNA, RNA, and/or analogs), drugs, and/or other molecules in an extracellular environment enter cells in living intact tissue upon application of pressure to the cells and extracellular environment. Nucleic acids localize to the cell nuclei. Transfection efficiencies greater than 90% are achievable for naked DNA and RNA. A sealed enclosure, defined by an enclosing means and/or tissue, contains the cells and their extracellular environment. The enclosure is pressurized to an incubation pressure on the order of atmospheres. A protective inelastic sheath may be used to prevent distension and trauma in tissue that is part of the enclosure boundary. Suitable enclosures include pressurization chambers and organs such as blood vessels or the heart. Parts of organs, entire organs, and/or entire organisms are pressurized. Suitable target tissue types include blood vessel (in particular vein) tissue, heart, kidney, liver, and bone marrow tissue. Gene therapy applications include ex-vivo treatment of grafts prior to transplantation, and in-vivo treatment of tissue. Useful therapy targets include cell cycle regulatory genes for blocking cell proliferation, and interleukin (IL) and cell adhesion molecule (CAM) genes for reducing immune responses to grafts.

Intracellular delivery of nucleic acids using pressure

Vascular function and structure is maintained or improved by long term administration of physiologically acceptable compounds, namely L-arginine, L-lysine, physiologically acceptable salts thereof, and polypeptide precursors thereof, which enhance the level of endogenous nitric oxide or other intermediates in the NO induced relaxation pathway in the host. In or in combination, other compounds, such as B.sub.6, folate, B.sub.12, or an antioxidant, which provide for short term enhancement of nitric oxide, either directly or by physiological processes may be employed.

Enhancement of vascular function by modulation of endogenous nitric

Neointimal cells are shown to express high levels of an anti-apoptotic gene, bcl-x, while the medial cells of the vessel itself express only low levels. The difference in gene expression exploited to provide for selective deletion of the neointimal cells. Apoptosis is induced by administering anti-sense bcl-x oligonucleotides to the affected vessel. Apoptosis is desirable as a treatment because it does not induce inflammation, further tissue injury or reactive hyperplasia. A significant reduction in lesion size is seen after treatment.

Apoptotic regression of intimal vascular lesions

The invention presents a system for inducing cells in living intact tissue, in vivo or ex vivo, to accept nucleotides from their extracellular environment and to localize those nucleotides into the cells' nuclei. This system relies on the fact that, when subjected to high pressure, cells take in nucleotides and localize those nucleotides into their nuclei with a transfection rate of greater than 90% in some cases. This invention employs various techniques for placing under high pressure either cells in isolated tissue cultures, or cells in tissues still connected to a living body. Such pressurization techniques include enclosing the tissue in an impermeable and inelastic sheath and injecting fluid into the tissue until a sufficiently high pressure is reached; placing a tissue culture in a dish and placing the dish in a pressurized chamber; mechanical pressurization techniques such as compression of a space containing fluids; occluding the ingress and egress of a vessel or organ to create a watertight compartment within the vessel or organ; and pressurization of an entire organism after delivery of nucleotide to the extracellular environment. This invention also employs various techniques for delivering a solution or suspension containing the nucleotide into the tissue; such techniques include the use of a syringe-like mechanism and a catheter.

Apparatus and method for delivering a nucleotide into cell nuclei

Atherogenesis and restenosis are treated by long term administration of physiologically acceptable compounds which enhance the level of endogenous nitric oxide in the host. Alternatively, or in combination, other compounds may be administered which provide for short term enhancement of nitric oxide, either directly or by physiological processes. In addition, cells may be genetically engineered to provide a component in the synthetic pathway to nitric oxide, so as drive the process to enhance nitric oxide concentration, particularly in conjunction with the administration of a nitric oxide precursor.

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Palo Alto, CA, United States of America

Gary H Gibbons