SAN DIEGO—The use of biologic factors that stimulate nerve growth or repair has long been considered a potential means to reduce neurologic injury or enhance recovery following stroke. But while animal studies have often found nerve growth factors to have beneficial effects, clinical trials have had disappointing results—and investigators are beginning to reconsider their methods for evaluating the effectiveness of new agents for the treatment of stroke.

FIT FOR BEAST, NOT MAN

Neurotrophic factors—such as nerve growth factor (NGF), brain-derived neurotrophic factor (BDNF), and basic fibroblast growth factor (bFGF)—regulate the growth, survival, and connectivity of neurons, especially in the developing nervous system. Although the normal physiologic roles of these substances in the adult nervous system are less well established, neurotrophic factors have been shown to support neuronal survival in response to various forms of injury in cell culture systems and in animal stroke models. However, clinical investigators are having difficulty in translating these laboratory studies into treatments that are effective for patients with stroke.

These difficulties were illustrated by a recent clinical trial of bFGF, the results of which were presented at the 52nd Annual Meeting of the American Academy of Neurology. Wayne M. Clark, MD, reported that animal studies had suggested that intravenous bFGF reduces neurologic injury following experimentally induced stroke. Dr. Clark is Professor of Neurology at Oregon Health Sciences University in Portland and Director of the Oregon Stroke Center.

In addition to its direct effect on neuronal survival, Dr. Clark noted that bFGF is believed to promote vascular growth, potentially increasing blood flow to ischemic tissue. Dr. Clark added that in a previous phase 1 study of 66 stroke patients, bFGF appeared to be well tolerated; and therefore, a large phase 2/3 clinical trial was initiated to examine bFGF for the treatment of acute stroke.

In this placebo-controlled, double-blind clinical trial, patients presenting within six hours of onset of an acute ischemic stroke were randomized to receive one of two doses of bFGF (5 or 10 mg intravenously) or placebo. The planned enrollment for the study was 900 patients: however, the trial was stopped by the study's safety committee after enrollment of only about 300 patients because the results of an interim analysis showed that the bFGF-treated patients actually had worse outcomes than patients who were treated with placebo. The bFGF-treated patients had significantly worse scores than placebo-treated patients on the Rankin stroke scale (the study's primary end point) and also exhibited an increase in mortality within 90 days of treatment (29% in the bFGF 5-mg group, 25% in the 10-mg group, and 13% in the placebo group).

The reasons for the poor outcome in this study, Dr. Clark said, are unclear at present. He noted that hypotension was reported somewhat more often among patients who received bFGF than among patients who received placebo, but he said that it is uncertain whether this contributed to the outcomes observed in the study. However, he said, investigators are considering the possibility that the observed effects of bFGF in animal studies might have been more closely related to changes in blood pressure or blood flow than to a direct neuroprotective effect.

ANIMAL MODELS AND HUMAN DISEASE—AN UNBREACHABLE GAP?

Dr. Clark offered several possible explanations for the observation that animal studies of new stroke medications are frequently positive, but subsequent clinical investigations typically fail. First, he said, the delay between the onset of ischemia and the initiation of treatment is probably a critical factor.

"In most animal studies, the agent is administered as a pretreatment or, at worst, perhaps an hour after ischemia," Dr. Clark said. "Almost all the clinical trials have enrolled most patients between five and six hours after stroke." It might therefore be the case that patients are simply being treated too late. Although great efforts have been made during the past decade to treat stroke patients more quickly, "we're still treating less than 5% of all stroke patients in under three hours," he said. Dr. Clark noted that although some newer trials are designed to study patients who can be treated in less than three hours from stroke onset, so few patients are available to meet this enrollment criterion that these trials take a very long time to complete.

A second important factor may be that human patients are rarely able to tolerate experimental medications in dose levels comparable to those that are administered in animal studies. In addition, Dr. Clark pointed out that human strokes tend to be much more variable than the experimental infarctions that are produced in most animal models.

THE SEARCH FOR NEUROPROTECTION CONTINUES

Despite these difficulties, preclinical studies of potential new therapies for stroke continue. Dr. Clark explained that in addition to neurotrophic factors, several growth-regulating cytokines may also be important in the development of neurologic injury following stroke, perhaps through their effects on the influx of leukocytes into sites of injury or through regulation of the release of tissue-damaging free radicals. Some of the best characterized of these cytokines are tumor necrosis factor-a (TNF-a), interleukin (IL)-1ß, and IL-6.

For example, Dr. Clark described the results of animal studies demonstrating that both systemic and intraventricular IL-1ß receptor antagonists have reduced infarct volumes following experimental stroke. With TNF-a, the situation is somewhat more complex: receptor blockers have been shown to reduce infarct size, although knockout animals actually had larger infarct volumes than control animals. "That's where the story gets complicated," Dr. Clark said. "Maybe you need a little bit; but if it's too much, then that's bad."

Dr. Clark also noted that although some studies have suggested that IL-6 levels are correlated with stroke severity in patients, studies of IL-6 knockout animals found no difference in lesion size between control and knockout animals. Investigation continues to clarify the potential role of cytokine modulation as a potential stroke therapy.

Researchers are also exploring alternative approaches to stimulating neuronal growth and repair. One avenue of investigation is the use of purine nucleosides, such as adenosine or inosine, for the treatment of stroke. Recently, investigators at the biotechnology company Boston Life Sciences, Inc, reported that inosine, which has been shown to stimulate nerve growth in vitro, improved the neurologic status of rats that were subjected to experimental cerebral ischemia.

According to the researchers, infusion of inosine into the cerebrospinal fluid of rats significantly reduced the degree of neurologic impairment (hemiparesis) following experimental stroke. "Inosine appears to be effective even when given after the damage has been done and the stroke is complete," said Marc Lanser, MD, Chief Scientific Officer of Boston Life Sciences, Inc.

RECONSIDERING STUDY DESIGNS

Justin A. Zivin, MD, PhD, who is a Professor of Neuroscience at the University of California, San Diego, and has been an investigator in numerous preclinical and clinical stroke studies, agreed that although neurotrophic agents have promise for the treatment of stroke, clinical progress has been slow. According to Dr. Zivin, investigators are reconsidering clinical trial design. "The main thing is that many of the trials that failed were designed before we had adequate information about how to design better trials. These first-generation trials were all testing a variety of drugs in essentially the same way, and it's probably the strategy that was bad rather than the drugs," he said.

Dr. Zivin noted that, in the future, clinical trials will probably be conducted differently. "For one thing, there will unquestionably be greater emphasis on reproducing protocols that are effective in animal models in people," he said. In addition, preclinical investigators are currently attempting to develop animal models that more closely resemble the clinical situation. Finally, he said, most clinical trials will be carried out in the context of combination therapy.

Dr. Zivin noted that, of the large number of clinical trials conducted in stroke patients over the past several years, only three have proven successful, and all of those were trials of thrombolytic agents. He added that newer agents probably will, for the most part, be tested in combination with thrombolytic therapy. "One of the thoughts is that until you reverse the primary problem—and, in stroke, in the overwhelming majority of cases the primary problem is an occluded vessel—other drugs that might be useful have a hard time getting to the tissue at risk." Thus, he said, the combined use of neuroprotective and thrombolytic agents may be much more effective than the use of either agent alone, although he conceded that this concept requires further testing in controlled trials.

NR