Delivery of therapeutic gene products by intestinal cell expression

Abstract

Intestinal epithelial cells of a mammalian subject are genetically altered to operatively incorporate a gene which expresses a protein which has a desired therapeutic effect. Intestinal cell transformation is accomplished by administration of a formulation containing DNA and is preferably administered orally. Oral or other intragastrointestinal routes of administration provide a simple method of administration. The expressed protein is secreted directly into the gastrointestinal tract and / or bloodstream to obtain therapeutic blood levels of the protein, thereby treating the patient in need of the protein. The delivery system of the invention provides short or long term delivery of therapeutic gene products.

Description

PRIOR APPLICATIONS [0001] This application is a continuation of U.S. patent application Ser. No. 09 / 811,323, filed on Mar. 16, 2001, which is a continuation of U.S. patent application Ser. No. 09 / 254,988, filed on Jun. 11, 1999 (now U.S. Pat. No. 6,258,789), which is a national phase application under 35 U.S.C. § 371 of International Application No. PCT / US97 / 16523, filed on Sep. 18, 1997, which is a continuation-in-part of U.S. application Ser. No. 08 / 717,084, filed Sep. 20, 1996 (now U.S. Pat. No. 6,225,290), each of which is incorporated by reference herein in its entirety for all purposes.FIELD OF THE INVENTION [0002] This invention relates generally to the field of drug delivery, more particularly to the delivery of therapeutic gene products by transformation of cells of the intestine. BACKGROUND OF THE INVENTION [0003] Proteins are essential to all biological functions, from metabolism, to growth, to reproduction, to immunity. As such, they have an important potential role as p...

AI Technical Summary

Benefits of technology

[0016] An important advantage of the present invention is that it allows administration of protein drugs by mouth. Rather than attempting to deliver the protein itself, the delivery system of the invention does so indirectly by administering the gene encoding the therapeutic proteins. Despite the conventional wisdom that any DNA in the gastrointestinal tract would be destroyed rapidly by the digestive process (either by stomach acid or intestinal DNAse), oral delivery of DNA encoding a desired therapeutic protein is successful in the present invention. The DNA is taken up by intestinal cells, which synthesize the encoded protein and secrete it into the bloodstream or the gastrointestinal tract to achieve therapeutic results. The flexibility of this technology allows for the delivery of a wide variety of protein pharmaceuticals, systemically, into the gastrointestinal tract, as well as locally, making it well suited for a broad spectrum of therapeutic applications.

[0017] Another advantage of the present invention is that the short term expression of the therapeutic gene in the individual allows for regulation of administration of the therapeutic gene product to the patient. Because intestinal cells turn over rapidly, expression can be easily modified or altered by varying the dose and / or formulation of the oral preparation. Short-term expression is thus a consequence of the rapid turnover of transformed cells that are normally lost (or “turned over”) within about two or three days. This aspect of the invention is both advantageous for dose control and reduction of risk of long-term complications from DNA integration (mutagenesis).

[0018] Another advantage of the invention is that the method completely avoids invasive procedures, and allows the vector to be administered in the simplest possible fashion—by the oral administration of a pill or other material. The lumen of the gastrointestinal tract is actually “outside” the body, and is separated from it by a single continuous layer of cells. As such, anything that passes into the gastrointestinal tract through the mouth remains in the exterior space, and cannot enter the body proper and its bloodstream, unless it first crosses the cells that line the gastrointestinal tract. However, once the gene is expressed within the intestinal cells and the protein product released into the bloodstream via natural secretory pathways, the therapeutic protein acts in the same manner as current, injectable forms of the drug.

[0019] The present invention is also advantageous over gene-based therapies that administer the gene vector into the bloodstream to other tissues and organs in that it involves administration of the DNA of interest directly to the target cells of the subject without first being distributed broadly via the bloodstream. Thus delivery of the DNA using the invention is more efficient and avoids the need for additional mechanisms to target the DNA of interest to a particular tissue.

[0020] Still another advantage is that the present invention minimizes the exposure of the transforming DNA to the bloodstream, the major source of adverse reactions to treatment. Reaction by the immune system to the delivery vector (particularly viral vectors) is a major obstacle for conventional gene-based therapies. Delivery of vectors by other routes (e.g., intravenous, intramuscular injection, or pulmonary administration) exposes them to blood and extracellular fluid. This exposure commonly results in inflammation and an immune response. These adverse reactions often worsen with reapplication, to the point where treatment cannot continue and becomes completely ineffective. The present invention presents the vector directly to the intestines without having to pass first through the blood or tissue of the subject. This shields the DNA delivery process as much as possible from the systemic circulation where immunological and inflammatory responses are initiated, and in this way minimizes their interference with therapy.

[0021] Another advantage of the invention is that naked DNA can be used as the vector, rather than viral vectors. Although viral vectors have been popular for gene therapy due to ease of administration and the incorporation of DNA into the genome, viral vectors have been found to produce substantial antigenic reactions that prevent their multiple administration. Use of naked DNA avoids this problem.

Problems solved by technology

Although protein drugs have enormous therapeutic potential, their more widespread use has been limited by several restrictive technical factors.

First, proteins remain difficult and expensive to manufacture compared to other pharmaceuticals.

Large scale purification of proteins in bioactive form can be a limiting step in the commercialization of these drugs.

This results in the need for frequent re administration.

This increases the complexity and expense of the treatment, and the disagreeable nature of administration also limits potential clinical applications.

However, past efforts to accomplish in vivo transformation of intestinal cells have met with severe obstacles.

Efforts to achieve in vivo transformation may be further complicated by the mucus layer of the intestine, which is thought to block access of the gene therapy transforming formulation to the target cells (Sandberg et al., supra).

The presence of high concentrations of DNAses in the intestinal tract is also thought to be a formidable barrier to the effective introduction of DNA into intestinal tract cells.

Many of the vectors and delivery systems developed for in vivo cellular transformation either have their own inherent drawbacks or are not entirely suitable for in vivo intestinal cell transformation.

For example, recombinant viruses, particularly retroviruses, may be slow in gaining FDA approval due to concerns generally associated with the administration of live viruses to humans.

In addition, it has become clear that viral vectors present problems with the possibility of multiple administrations of the gene construct due to immune responses, and may greatly limit their utility.

Mechanical means, such as the gene gun, are designed for use in transformation of skeletal muscle cells and are not particularly useful in intestinal cell transformation due to problems of access and to the delicate nature of organ.

However ex vivo techniques require complex procedures to accomplish transformation, put the subject at risk of rejection of the transplant, require at least minor invasive procedures, and limit implantation to modest numbers of cells.

In vivo methods (e.g., direct administration through blood or to muscle) also frequently require invasive procedures and meet with difficulties in delivery of the transforming material to the target cell.

Moreover, delivery of the transforming material via the bloodstream of the individual results in exposure of the DNA and any carrier associated with it to the immune system, which can result in adverse reactions (e.g., inflammatory reactions to the DNA administered and / or to components of the formulation containing the DNA).

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