Contents of This Section

1. What is the relevance of the fields of gene therapy and genomics to brain aneurysms?

As you read the section on Gene Therapy, below, and the section on Genomics elsewhere in this Website ( take me to the Genomics section now), you will learn that understanding the genetic basis for why aneurysms form and rupture is extremely important. Why? Because this knowledge may lead to the identification of a precise molecular target for gene-based aneurysm screening and treatment and, hopefully, will pave the way to preventing the formation and rupture of these lesions altogether. This may seem like a long shot, but my colleagues and I have already made great strides in the fields of cerebrovascular gene therapy and genomics, and we're dedicated to making this dream a reality. Here are some studies published in premier cerebrovascular, gene therapy and neurosurgical journals, reflecting my personal involvement in this field, and some important scientific advances we've made to date:

2. What is gene therapy and how does it relate to the Human Genome Project?

In order to understand what the term "gene therapy" means, the two key words "gene" and "therapy" must first be defined:

Human Gene Therapy Model:

Figure 1 shows key components of a human gene therapy model. In the ideal scenario, a patient with a disease whose precise molecular defect is known undergoes safe and specific gene therapy, resulting in an objective clinical benefit (ideally, a "cure").

The promise of gene therapy is that this therapeutic technology will one day be safe, widely available, and highly effective in curing human diseases. However, two important things can be said about this statement:

3. What is gene transfer?

Gene transfer is the science behind gene therapy (see Figure 2, below). That is, gene therapy relies on gene transfer technology. What comprises gene transfer technology? There are four key components:

General Mechanism of Gene Transfer:

Figure 2 illustrates the mechanism of gene transfer, which represents the science behind gene therapy. It involves packaging a gene of interest (1; which codes for the therapeutic protein) into a suitable vector (2). The vector is (using an appropriate delivery device; see below) brought near to cells in the target tissue (i.e., the host's tissue in which we want the therapeutic protein to be produced or "synthesized"), and gains entry into these cells (3). This entry may be aided by contact with a receptor and/or a coreceptor found (i.e., "expressed") on the target cell surface. The vector eventually makes its way into the nucleus of the host cell (4); this region of the cell houses the host's genomic (or "native") DNA. Depending on the type of vector used, the cDNA in the vector may or may not become one with the host's native DNA. Regardless of this event, using the transcriptional and translational machinery of the host, the vector (without replicating itself) allows the host's cell to produce large amounts of the therapeutic protein (5). This protein may stay within the cell (e.g., an enzyme), or may move out of the cell (i.e., a secreted protein or protein fragment known as a polypeptide). Either way, the protein's presence should have a beneficial (or therapeutic) effect on the host. Depending on the vector used, the host may mount an inflammatory response to the vector, which can limit the effectiveness of this technology. Much progress is being made regarding the reduction of the inflammatory (immune-stimulatory) effects of vectors.

4. How does gene transfer to blood vessels work?

In accordance with the gene transfer mechanism described above, gene transfer to blood vessels works as follows (see Figure 3, below). Say certain brain arteries in a patient are known to be diseased in that they are not producing enough of a protein such as eNOS or HO-1 ( visit the Key References section for more information). This can occur, e.g., in the setting of cerebral vasospasm following rupture of an aneurysm ( I want to find out more about the mechanism of cerebral vasospasm, so take me to Cerebral Vasospasm now). In such scenarios, a therapeutic gene of interest (e.g., cDNA for eNOS or HO-1) is packaged into a suitable "clinical-grade" vector (e.g., a safe and highly purified adenovirus that has been engineered to be unable to reproduce itself, but still able to carry the therapeutic gene and allow its conversion into a therapeutic protein). The vector is then delivered to the target vessel(s) using a suitable delivery device (see 4. below). The vector particles now make their way into the cells of the target artery in the host [cell-entry in the case of an adenovirus typically involves attachment of outer parts of the adenovirus to a certain host-cell receptor known as the Coxsackie virus-adenovirus receptor (CAR) and an "integrin" coreceptor]. The vector particles eventually reach the nucleus of the host's cell, and there they allow the host's molecular machinery to produce the therapeutic protein (e.g., "recombinant" eNOS or HO-1). Note that in the case of an adenovirus, when the vector enters the nucleus, the adenovirus vector's own DNA does not incorporate itself into (i.e., does not fuse or "integrate" with) the host cell's original (or "native") DNA (i.e., it remains "episomal"). Other vectors, on the other hand, do fuse. The act of fusion versus nonfusion has interesting consequences; these are elaborated elsewhere [ for a thorough and colorful review of this topic, see Khurana & Meyer (2003) in the Key References section of this Website].

Mechanism of Vascular Gene Transfer:

Figure 3 shows how gene transfer works in a blood vessel. A vector (1) incorporating a gene of interest is transferred into a target blood vessel (2). This event is called "transduction". If we look at a cross-section (3) through the same region of the blood vessel wall that the vector has entered, we can see that the vector has caused the production of the therapeutic protein (yellow circles in 3) encoded by the gene of interest. Of course, the vector may not enter all of the cells of the target tissue, and so the therapeutic protein is produced only in cells in which all of the above have taken place (namely, accurate delivery, efficient transduction, and effective biosynthesis).

5. How do you deliver genes to blood vessels?

There are several ways in which vectors containing therapeutic genes can be delivered to a blood vessel (see Figure 4, below). First, note that the "route of delivery" may be through the central opening (lumen) of the artery (i.e., "intraluminal" delivery) or via the outer wall (adventitia) of the artery (i.e., "adventitial" or "perivascular" delivery). The structure of an artery is discussed elsewhere ( take me to Brain Artery Structure now). Second, the "technique of delivery" may be associated with a physical method (mechanical approach) or a nonphysical method (chemical or molecular approach) aimed at either enhancing targeting of the vector to a particular site (or tissue) in the body (i.e., increasing the accuracy of the transduction process) or improving the chances of the vector entering the cells of the target tissue (i.e., increasing the efficiency of the transduction process), or both. Third, there are different devices that can be used for the delivery of vectors. For example, in the intraluminal approach, a catheter (essentially a small, flexible hollow tube) can be threaded through the arterial tree to (or close to) the specific artery into which the vector [now in the form of a liquid ("solution" or "suspension")] can be "squirted". Alternatively, a catheter can be used to release (or "deploy") a stent (essentially a tube that expands against the innermost surface of the artery); it is a special type of stent of course (a bioactive stent), namely, it has been impregnated with a vector, or may contain previously transduced cells capable of producing the therapeutic product. In the perivascular approach, a small paintbrush can be used to gently, safely, and very effectively massage a vector into the wall of an artery from its outer surface. This is a form of mechanical transduction that has the advantages of being direct, intuitive (i.e., easy to understand and easy to use), and relatively rapid and efficient [Khurana et al., Gene Therapy (2003), Khurana and Meyer (2003), U.S. Patent No. 6,821,264; Visit the Key References or Genebrush Sections). Another type of perivascular approach is to use a needle to inject a vector into the space surrounding the artery. This can be done close to the artery itself (i.e., local delivery), or at some distance from the artery (i.e., remote delivery). If the vector is injected close to the outer wall of the target artery, it typically has to be contained in an outer catheter or sheath; if it is injected remote to the target artery (say, into the cerebrospinal fluid that circulates in and around the brain and spinal cord - like a "spinal" or "intrathecal" injection), then it will diffuse throughout these structures in a more nonspecific manner. All of these approaches have been shown in gene transfer studies to be effective in causing successful transduction of arteries. They are all subject to ongoing development.

Regardless of the delivery approach, the main goal is to safely, accurately, and efficiently get vector particles into cells of the target artery, where the particles can then use the host's molecular machinery to produce the desired therapeutic protein.

Techniques of Vascular Gene Delivery:

Figure 4 illustrates different techniques (1 - 4) that can be used to deliver genes to an artery. (1) A hollow catheter (blue tube in the figure); note the vector (yellow) particles being squirted into the lumen of the artery. There are other types of catheters with special chambers that intentionally trap the vector in a confined space and thereby allow transduction to occur along only a relatively small (finite) length of the artery. (2) A stent (yellow hashed cylindrical sheet in the figure; only one-half of the stent is shown) can be used; this is in fact deployed by special catheters, and expands against the inner surface of the artery. Vector particles or vector-transduced cells are impregnated into the stent. (3) A needle can be used to inject a vector solution (yellow droplets in the figure) into the space surrounding an artery. (4) A small surgical paintbrush can be used to gently and directly massage a vector-containing solution or paste (yellow streak in figure) into the wall of an artery. Note that a "gene gun" (not shown here) has also been developed to fire vectors such as naked DNA particles directly into tissues.

6. What progress has been made in gene therapy for diseases affecting brain blood vessels?

Since the first gene delivery to brain arteries carried out in 1995, there have been dozens of studies that have characterized the mechanism and benefits of this approach in the brain circulation.

This area has been recently reviewed by Khurana & Meyer (2003), and an interested reader is referred to this article and others in the Gene Therapy Key References section of this Website.