Drug – bio-affecting and body treating compositions – Designated organic active ingredient containing – Carbohydrate doai
Reexamination Certificate
1997-02-11
2003-02-04
Riley, Jezia (Department: 1637)
Drug, bio-affecting and body treating compositions
Designated organic active ingredient containing
Carbohydrate doai
C424S472000, C424S497000
Reexamination Certificate
active
06514947
ABSTRACT:
BACKGROUND OF THE INVENTION
The following information is presented solely to assist the understanding of the reader, none of the information is admitted to describe prior art to the claims of the present invention.
This invention relates to compositions and methods for the introduction of a formulated nucleic acid into a cell for the expression of a peptide or polypeptide. It is useful for in vitro transfections and in vivo for gene therapy, for among other things administration of therapeutic proteins, polypeptides and peptides and for vaccination.
Non-viral administration of nucleic acid in vivo has been accomplished by a variety of methods. These include lipofectin/liposome fusion:
Proc. Natl. Acad. Sci
., Volume 84, pp. 7413-7417 (1993); polylysine condensation with and without adenovirus enhancement:
Human Gene Therapy
, Volume 3, pp. 147-154 (1992); and transferrin: transferrin receptor delivery of nucleic acid to cells:
Proc. Natl. Acad. Sci
., Volume 87, pp. 3410-3414 (1990). The use of a specific composition consisting of polyacrylic acid has been disclosed in WO 94/24983. Naked DNA has been administered as disclosed in WO 90/11092.
An important goal of gene therapy, as an initial step in the process of ultimately obtaining expression of a product encoded by a nucleic acid, is to effect the uptake of nucleic acid by cells. Uptake of nucleic acid by cells is dependent on a number of factors, one of which is the length of time during which a nucleic acid is in proximity to a cellular surface. For instance, after intramuscular (i.m.) administration of plasmid DNA in buffer, a marked reduction in gene expression is observed if the muscle is massaged, presumably due to DNA leakage out of the muscle either directly or via lymphatic vessels (
Human Gene Therapy
4:151-159; 1993). Accordingly, it would be desirable to formulate nucleic acids with compounds which would retard the rate at which nucleic acids diffuse or are carried away from a site at which cellular uptake of the nucleic acid is desired. Further, these compounds would be suitable for administration to an organism by means such as injection while maintaining or regaining the physical characteristics necessary to increase cellular uptake of nucleic acids.
SUMMARY OF THE INVENTION
This invention features compositions and methods for enhancing the administration to and uptake of nucleic acids by an organism. An efficient strategy for enhancing nucleic acid delivery in vivo is to protect the nucleic acid from degradation, thereby maintaining the administered nucleic acid at the target site in order to further increase its cellular uptake. Also, for in vitro administration, increasing the effective concentration of the nucleic acid at the cell surface should increase the efficiency of transfection. The compositions of the present invention which are used to administer nucleic acid comprise a compound which protects the nucleic acid and/or prolongs the localized bioavailability of the nucleic acid when administered to an organism in vivo, or in vitro in cell culture.
In connection with the compounds and compositions of this invention, the term “protects” or “protective” refers to an effect of the interaction between such a compound and a nucleic acid such that the rate of degradation of the nucleic acid is decreased in a particular environment. Such degradation may be due to a variety of different factors, which specifically include the enzymatic action of a nuclease. The protective action may be provided in different ways, for example, by exclusion of the nuclease molecules or by exclusion of water.
By “prolong the localized bioavailability of a nucleic acid” is meant that a nucleic acid when administered to an organism in a composition comprising such a compound will be available for uptake by cells for a longer period of time than if administered in a composition without such a compound, for example when administered in a formulation such as a saline solution. This increased availability of nucleic acid to cells could occur, for example, due to increased duration of contact between the composition containing the nucleic acid and a cell or due to protection of the nucleic acid from attack by nucleases. The compounds which prolong the localized bioavailability of a nucleic acid are suitable for internal administration.
By “suitable for internal administration” is meant that the compounds are suitable to be administered within the tissue of an organism, for example within a muscle or within a joint space, intradermally or subcutaneously. Other forms of administration which may be utilized are topical, oral, pulmonary, nasal and mucosal; for example, buccal, vaginal or rectal. Properties making a compound suitable for internal administration can include, for example, the absence of a high level of toxicity to the organism as a whole.
By “nucleic acid” is meant both RNA and DNA including: cDNA, genomic DNA, plasmid DNA or condensed nucleic acid, nucleic acid formulated with cationic lipids, nucleic acid formulated with peptides, cationic polymers, RNA or mRNA. In a preferred embodiment, the nucleic acid administered is plasmid DNA which comprises a “vector”.
A “vector” is a nucleic acid molecule incorporating sequences encoding therapeutic product(s) as well as, various regulatory elements for transcription, translation, transcript stability, replication, and other functions as are known in the art.
A “transcript stabilizer” is a sequence within the vector which contributes to prolonging the half life (slowing the elimination) of a transcript.
“Post-translational processing” means modifications made to the expressed gene product. These may include addition of side chains such as carbohydrates, lipids, inorganic or organic compounds, the cleavage of targeting signals or propeptide elements, as well as the positioning of the gene product in a particular compartment of the cell such as the mitochondria, nucleus, or membranes. The vector may comprise one or more genes in a linear or circularized configuration. The vector may also comprise a plasmid backbone or other elements involved in the production, manufacture, or analysis of a gene product.
An “expression vector” is a vector which allows for production of a product encoded for by a nucleic acid sequence contained in the vector. For example, expression of a particular growth factor protein encoded by a particular gene.
A “DNA vector” is a vector whose native form is a DNA molecule. A “viral vector” is a vector whose native form is as the genomic material of a viral particle.
A “gene product” means products encoded by the vector. Examples of gene products include mRNA templates for translation, ribozymes, antisense RNA, proteins, glycoproteins, lipoproteins and phosphoproteins. The nucleic acid may be associated with a targeting ligand to effect targeted delivery.
A “targeting ligand” is a component of the carrier or vehicle or a moiety (a molecule or group) attached to a protective, interactive, non-condensing compound which binds to receptors, with an affinity for the ligand, on the surface or within compartments of a cell for the purpose of enhancing uptake or intracellular trafficking of the vector. Galactosyl residues, e.g., Tris-galactosyl residues, carnitine derivatives, mannosyl residues, e.g., mannose-6-phosphate, monoclonal and polyclonal antibodies, peptide ligands, and DNA-binding proteins represent non-limiting examples of targeting ligands which can be used to enhance uptake.
“Targeted delivery” involves the use of targeting ligands which specifically enhance translocation of a nucleic acid to specific tissues or cells. Examples of cells which may be targeted include, but are not limited to, antigen-presenting cells, hepatocytes, myocytes, eptithelial cells, endothelial cells, and cancer cells.
A “target” is a specific organ, tissue, cell, or cellular region for which uptake of a vector and expression of a gene product is intended.
“Uptake” means the translocation of the vector from the extracellular to intracellular compartments. This can involve recept
Mumper Russell J.
Rolland Allain
Riley Jezia
Valentis, Inc.
Wise Michael J.
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