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ow, thanks to a recent convergence of advances in the field of basic vascular cell biology and clinical medicine, several new therapeutic approaches to these diseases are being evaluated.

Both of these vision threatening eye diseases are characterized by the development of abnormal blood vessel growth in the eye, a process known as angiogenesis. In the case of ARMD, new blood vessels grow under the retina, while diabetic retinopathy is caused by the growth of abnormal vessels on top of the retina. The effect is much the same; the vessels interfere with normal structures or the transmission of light to the back of the eye, impeding vision.

Martin Friedlander, M.D., Ph.D., Associate Professor, Department of Cell Biology, and a retina specialist in the Division of Ophthalmology at Scripps Clinic, is conducting key research aimed both at understanding these disease processes and at developing treatments for them.

"The proliferation of new blood vessels is a common feature in many ocular diseases including not only age-related macular degeneration and proliferative diabetic retinopathy but also rubeotic glaucoma, interstitial keratitis and retinopathy of prematurity. It also is a leading factor contributing to corneal graft failure. If we can selectively target new vessels involved in these diseases while leaving old vessels alone, then we would have a way to shut down these angiogenic processes," said Friedlander.

FROM ONCOLOGY TO OPHTHALMOLOGY

Many forms of cancer also depend on the development of new blood vessels to survive and grow. Thanks to work done in the field of oncology, several anti-angiogenic compounds are now in clinical trials. These include compounds targeting a variety of growth factors including vascular endothelial growth factor (VEGF) and fibroblast growth factor (FGF).

As in the oncology field, VEGF is one of several targets for reducing angiogenesis in ocular disease. Recent studies indicate that VEGF may play an important role in stimulating angiogenesis in diabetic eye disease. Clinical studies with anti-VEGF antibodies and other VEGF-blocking molecules are now underway to evaluate the potential of this approach for treatment of diabetic retinopathy, he noted.

But there is more to the story than VEGF. For many years, TSRI researcher David Cheresh, Ph.D., has been studying another aspect of angiogenesis involving a class of molecules called integrins. About eight years ago, he discovered that one particular integrin, a_vb₃ was activated on newly sprouting blood vessels and that the same integrin is expressed on new blood vessels feeding tumors. Even more exciting, he and his colleagues at Merck KGaA were able to create other molecules to block a_vb₃ and shut down tumor growth. The two scientists met shortly after Friedlander arrived at TSRI in 1993, having moved his lab from UCLA.

"I had been interested in the potential of antiangiogenesis for treating eye diseases for many years. We knew that abnormal growth of new blood vessels played a major role in these blinding diseases. The problem was, we didn't have a rational approach to treating angiogenesis. At a dinner for new faculty, Richard Lerner brought my attention to an upcoming journal article by David Cheresh on the integrin research. This research showed me that they knew something about the mechanism of angiogenesis, providing the basis for a rational therapeutic approach, something I had been looking for for years," Friedlander recalled.

The two have been collaborating ever since. Friedlander's focus is on how integrins function during cytokine-driven angiogenesis processes. In his early work, he confirmed that the a_vb₃ integrin was indeed expressed on new vessels growing in the eye. He also found that he could shut down new vessel growth in animal eye models by using an antibody antagonist of a_vb₃ integrins developed in the Cheresh lab. The surprising finding was that this antibody did not shut down VEGF-driven angiogenesis. An antibody to a distinct but related integrin, a_vb₅, did shut down the VEGF-, but not FGF-driven angiogenesis. Furthermore, a peptide antagonist of both integrins shut down both types of angiogenesis. From this work emerged the concept of at least two distinct cytokine-driven, integrin-mediated pathways of angiogenesis.

Subsequent research correlating data from human disease specimens with the animal model results revealed that different integrins were activated depending on where in the eye the new vessels were growing. In some cases VEGF was involved, while in other cases FGF was a more important factor. The researchers showed that patients with ARMD, who have new vessels growing under the retina, expressed mostly a_vb₃. Patients with diabetic retinopathy, in contrast, activated two integrins, a_vb₃ and a_vb₅.

"This has profound implications for understanding both the cause of these diseases and potential treatment approaches. The findings suggest that targeting VEGF alone would not produce any significant benefits for ARMD patients, whereas it may be very effective for treating diabetic retinopathy. Clinical trials are underway in oncology using humanized antibodies to a_vb₃ integrin. Studies with retinal tissue from patients with ARMD indicate that the antibodies may have a significant effect on angiogenesis. Clinical trials in ARMD are planned with this approach," he said.

MATRIX METALLOPROTEINASES

Matrix metalloproteinases (MMPs) are enzymes involved in regulating the extracellular matrix, the glue-like structure that holds our bodies together. They are involved in many cellular processes including angiogenesis. Friedlander's lab is examining the relationship between integrin signaling and the extracellular matrix, looking for a way to block MMPs involved in angiogenesis selectively, without interfering with other vital processes.

"These MMPs are all over the body. If you throw in an antagonist you will affect many systems in the body. In fact, one MMP antagonist was pulled from the market several years ago after patients developed serious side effects," he said.

The researchers may have found a more specific MMP. Recent research showed that actively growing endothelial cells have a way of localizing a specific MMP, called MMP2, to the tips of new vessels. The research showed that MMP2 can be degraded into another molecule called PEX (the carboxy-terminal, non-catalytic domain of MMP2). PEX in turn can bind to the a_vb₃ integrin, providing another way to shut down angiogenesis.

"This demonstrates a wonderful trait of nature. It appears that nature evolved a mechanism for using molecules for one purpose, and then recycle and use them to shut down the very same process," he noted.

OLD DRUGS, NEW USES

The antiangiogenic properties of the class of drugs known as corticosteroids have been known for many years. However, the severe side effects that accompany long-term use of these drugs have limited their use in treating eye disease. A new potent angiostatic steroid, anecortave acetate, that is devoid of the glucocorticoid activity that causes the side effects, may be the answer. Phase II clinical trials of this agent (sponsored by Alcon Laboratories) for treating ARMD are now well along; Friedlander is one of 15 principal investigators nationwide.

"I'm very excited about this trial. We have worked with this compound in the laboratory and have been impressed with the results in animal models," he said.

He is also a principal investigator at one of 30 centers worldwide participating in a clinical study (sponsored by Novartis) of another drug that was not originally developed for treatment of eye disease, called octreatide (Sandostatin LAR).

TOPOGENESIS

During medical school, and then as a junior faculty member, Friedlander worked in the laboratory of Gunter Blobel, M.D., Ph.D. Blobel, who won the 1999 Nobel Prize in Physiology or Medicine, described the general principles underlying the sorting and targeting of proteins to particular cell compartments. He determined that the protein itself carries the information that specifies its proper destination in the cell. These cellular zip codes are known as topogenic signals.

Friedlander participated in that early research first describing the presence of internal topogenic signals in polytopic membrane proteins like rhodopsin. He continues to apply the principles discovered in the Blobel lab to his current research. Part of his research group continues to study the role that topogenic signals play in targeting visually important polytopic membrane proteins to their proper places in normal and diseased eyes. He has studied the role of topogenic signals in the function of rhodopsin, a protein essential for sight. He is also studying the role that topogenesis plays in sodium and calcium exchange in photoreceptors. It appears that when these two proteins are mutated and display the wrong cellular 'zip code', this can result in degenerative changes in the eye of the type seen in certain inherited and acquired retinal and macular degenerations.

SPECIAL DELIVERY

An issue that continues to elude researchers developing treatments for eye diseases like ARMD and diabetic retinopathy is how to get the drugs to the intended destination. While molecular biology continues to provide sophisticated molecules with therapeutic potential, the technology to deliver these drugs lags behind. Noting that the drug companies "just aren't moving fast enough," Friedlander joined this effort and has accepted the challenge of developing practical drug delivery devices in his lab.

"My lab is interested in basic science. But we are also interested in clinical applications. So we are collaborating with a bioengineering group at Brown University to develop bio-erodable polymers -- tiny microspheres in which to incorporate some of these novel compounds.

It is helpful that the eye is a self-contained system. We would like to be able to inject a treatment behind the eye and then have it be released slowly over time. The idea would be to inhibit the progression of disease by injecting only a couple of times per year," he explained.

Cell-based delivery is another option his lab is exploring. In collaboration with Drs. Glen Nemerow and Peter Ghazal, associate professors in the Department of Immunology, the Ocular Gene Therapy Program at TSRI investigates ways to incorporate useful compounds directly into the cells involved. One approach involves using genetic engineering to introduce a gene into cells that will then produce a protein. Another approach they are looking at involves putting genetically engineered cells into a semipermeable membrane, and putting that membrane inside an inert container. This type of implant would allow slow release of a medicine or protein, while minimizing the risk of triggering an immune response. Still another approach involves using the outer coating of the eyeball, known as the sclera, as a kind of sponge. It may be possible to inject the sclera with a therapeutic compound and then let that compound seep into the eye over time.

"I cannot imagine a better place in the world to combine basic science and clinical research than The Scripps Research Institute. Now we can combine a major focus on visual cell biology with the clinical facilities to offer our patients the latest available therapeutic options," he said.