By David N. Leff

Coroners faced with a violent death of uncertain cause often have two options: murder or suicide.

In insults to neurons in the brain, such as ischemic stroke, the answer is both of the above: intentional death and self-destruction.

In stroke, the "intent" is sudden blockage of blood to the brain. This critical oxygen starvation of sensitive neurons brings on the suicidal programmed cell death now known as apoptosis.

In North America, stroke strikes more than 600,000 people a year, and leaves 400,000 of them permanently disabled from irreversible damage to cerebral cells.

"Certain neurons in the brain's hippocampus," explained neuroscientist George Robertson, professor of cellular and molecular medicine at the University of Ottawa, in Ontario, "play a very important role in memory consolidation. And they're extremely sensitive to ischemia — interrupted blood flow to the brain.

"When these cells die," he continued, the stroke victim can't remember anything; memories are sort of lost in space."

This slow neuronal death, he pointed out, "takes two or three days to occur. So there's a window of opportunity for salvaging them by some form of therapy, if the mechanism by which those neurons die could be interrupted."

Robertson is senior author of an article in the September 1997 issue of Nature Medicine that describes experiments pointing toward near-future therapeutic prospects not only for stroke but potentially for Parkinson's disease, and eventually cancer. Its title: "Elevation of neuronal expression of NAIP [Neuronal Apoptosis Inhibitory Protein] reduces ischemic damage in the rat hippocampus."

"In these studies," he told BioWorld Today, "we have demonstrated for the first time that pharmacologically modulating the expression of the NAIP protein can protect brain cells by inhibiting the apoptotic process."

Robertson's other day job is as director of the neuroprotection program at Ottawa-based Apoptogen Inc., located on the university campus. Two of the neuroscientists who co-founded the company last year are molecular geneticists Robert Korneluk and Alexander MacKenzie, both co-authors of the Nature Medicine paper.

It was they who reported in 1995 that the NAIP gene is missing or altered in patients with spinal muscular atrophy (see BioWorld Today, Nov. 20, 1996, p. 1), and that this mutation produces a protein that causes uncontrolled cell death. The normal protein, on the contrary, appears to protect neurons from premature apoptosis.

If Some Cells Survive, Why Not All?

In a rat model of human cerebral stroke, Robertson and his co-authors found some neurons somehow survive the ischemia. "Not all cells have equal susceptibility to injury after oxygen starvation," he observed. "Some cells are very resistant; nobody knows why."

The experiments they report, he added, "show that some of these cells, known to be very resistant, have high levels of this anti-death, anti-apoptotic protein, NAIP. This suggested to us," he went on, "that if we could somehow boost production of NAIP in neurons that do die after experimental stroke, they might survive."

The team produced experimental stroke in rats by tying off for 10 minutes the four major arteries — two carotid, two vertebral — that supply blood to the brain. During this ischemic interlude, they treated the animals with one of two NAIP-raising strategies.

One was to deliver to them the cDNA sequence for NAIP in an adenoviral package. The other was to inject the rodents with a bacterial alkaloid compound that jump-starts NAIP production.

Both approaches duly increased NAIP, and largely protected the threatened neurons from programmed cell death. But while successful as experimental prototypes, Robertson pointed out, neither has a future in human therapy for stroke.

"I think that the gene therapy approach," he said, "is going to be useful for brain trauma, or spinal cord injury, for example, in cases where you want to rescue neurons that are degenerating slowly. It probably won't be useful in a stroke situation, where you're dealing with virtually hours rather than days."

As for the alkaloid compound, he observed, "unfortunately, it's a blunt instrument, not something that could be used in humans because it has so many other properties."

When such a drug is discovered, "its use in acute stroke is the least likely of all events," Robertson pointed out. "First off, the time window is very narrow."

Rather, he foresees such a compound "as a prophylactic treatment in people who are at risk of a stroke — namely, those who've already had one, people going into a high-risk situation during surgery, which may involve bleeding, or the possibility of a cardiac infarct occurring. You want to jack the system up as much as you can before they go into such a dangerous setting."

From Stroke To Parkinson's To Cancer

But beyond stroke, he and his colleagues predict broader applications in neurodegenerative afflictions. "It may have relevance for Parkinson's disease [PD]," he said, and added: "We actually have some evidence now in animal models of PD that treatments that up-regulate NAIP are effective."

Co-author Stephen Crocker will present an abstract at the annual meeting of the Society for Neurosciences in New Orleans next month, Robertson went on, "showing that if we over-express this protein in dopaminergic neurons, we can increase their resistance to damage."

"Obviously we're a company that's developing drug screens," Apoptogen's president, John Gillard, told BioWorld Today. "We're looking for partnerships in drug screening and, in the longer term, with major pharma companies in the area of neuroprotection."

One such firm with which he is in active drug-screening discussion, Gillard added, is PanLabs International, of Bothell, Wash. "PanLabs," he pointed out, "is owned by Toronto-based MDS Capital Corp., a fairly large Canadian venture-capital and health care company.

MDS, Gillard observed, is one of four investment groups which contributed C$8 million last Thursday to support Apoptogen's research programs, including cancer.

Anent the cancer connection, Robertson observed "there is evidence" that these NAIP-family genes may be important in maintaining the life of tumor cells. In cancer, he concluded, "you obviously want to promote cell death, whereas in the brain we want to prevent apoptosis." *