Oregon Health Sciences University

Scientists at Oregon Health Sciences University and the Portland Veterans Affairs Medical Center have observed a second barrier that apparently prevents passage of agents from the blood to the brain. The primary barrier is a good thing in a healthy person, but this barrier also keeps drugs such as cancer-fighting chemotherapy from reaching the brain in patients undergoing treatment for brain tumors and other brain malignancies.

Leslie L. Muldoon, Ph.D., assistant professor of neurology, and cell biology and developmental biology, and her colleagues in the Blood-Brain Barrier Program at OHSU report their finding in the February 1999 issue of the American Journal of Neuroradiology (which comes out in March). The group has 20 years of experience with a technique that opens the first blood-brain barrier - an anatomic structure composed of tight junctions in endothelial cells -- to deliver cancer-fighting drugs. But some agents that pass through the first barrier apparently get caught on the second barrier -- called the basement membrane -- and never reach the brain.

The Oregon group is able to get certain therapeutic agents inside the brain with a barrier disruption technique that involves injecting patients with a sugar solution. This solution causes the tight endothelial junctions to shrink and open temporarily. With the barrier down, physicians can get cancer-fighting drugs across the barrier and into the brain - a place drugs don't permeate well with conventional chemotherapy.

More recently, the group has been experimenting on ways to deliver genes across the blood-brain barrier in rodents. The genes are loaded onto altered viral vectors, such as the herpesvirus vector or the adenovirus vector. (Scientists are able to inactivate the dangerous parts of the virus and then use recombinant techniques to load genes onto the virus.

These recombinant viral vectors can then safely ferry therapeutic genes to their target tissues.) Using iron particles the same size as viruses, the Oregon researchers noticed that some agents crossed the endothelial junctions only to get stuck just beyond them. The authors inferred the existence of a secondary blood-brain barrier at the level of the basement membrane. The basement membrane is a protein layer surrounding capillaries.

Edward A. Neuwelt, M.D., professor of neurology at OHSU and the VAMC, describes the second barrier as a "spider web" because it seems to trap some viral particles, while others slip through. The agents that make it through he calls "stealth" and those that don't he terms "non-stealth."

Although their studies suggest the presence of a second barrier, the researchers don't yet fully understand how it works. If not an actual anatomic structure, it may be an electrically charged barrier like the one that exists in the kidney. In either case, Neuwelt says the group's next challenges are to find ways to defeat the second barrier and to learn more about the properties that allow the stealth agents to pass through both barriers.

The work with viral vectors is important because toxic genes can be targeted at tumor cells for killing. Researchers have been using a herpes virus gene to "infect" tumor cells, rendering them susceptible to the antiviral drug acyclovir or ganciclovir. This gene has been approved for clinical trials, and Phase I trials are under way at a few institutions by direct injection into the tumor. Oregon researchers hope to improve delivery by delivering these vectors from blood to brain.

Researchers also hope to use viral vectors to carry replacement genes into the brains of people with neurodegenerative disease caused by the absence of a single gene or by a defective gene. In Parkinson's disease, for example, it may be possible to deliver a functional gene to alleviate symptoms of the disease.

The research findings in the neuroradiology journal also point to problems with current magnetic resonance imaging. Blood-brain barrier disruption relies on MR imaging to assess the distribution of iron particles throughout the brain hemisphere. However, MR images cannot show incorporation of these agents at the cellular level. In other words, the MR image machines in clinical use today only show delivery across the first barrier, but not whether the agents have actually crossed the basement membrane, or second barrier. Clinicians may incorrectly assume that a drug is reaching its target in the brain when in fact it is being stopped at the second barrier.

With viral particles, which are much larger than chemotherapeutic molecules, it's important for clinicians to know whether a specific viral vector can pass through the secondary barrier. In an editorial in the same issue, Robert M. Quencer, M.D., editor of the American Journal of Neuroradiology, calls this "a distinct challenge for highly detailed MR imaging." He suggests the use of stronger magnets in MR imaging to provide adequate spatial resolution. The Oregon team has approval to assess the distribution of these viral-sized iron particles in patients with brain tumors to determine the feasibility of gene therapy.

In addition to Muldoon and Neuwelt, authors include Michael A. Pagel, B.A., VAMC, and Robert A. Kroll, D.V.M., Simon Roman-Goldstein, M.D., and Russell S. Jones, B.S. all of OHSU. The National Institutes of Health and the Veterans Administration fund the work.