By David N. Leff

"Innocent sleep," Shakespeare called it, "that knits up the ravell'd sleeve of care." Now we confront the not-so-innocent sleep - narcolepsy - which ties that slumberous sleeve into a hard knot of lifelong misery.

"Narcolepsy ruins lives," observed neuropsychopharmacologist and molecular geneticist Emmanuel Mignot, at Stanford University in California. "It's especially true," he added, because an average of 14 years pass before the sleep disorder is diagnosed."

Narcolepsy (derived evocatively from the Greek words for "stupor" and "seizure") is a rare but devastating affliction in which its victims fall suddenly and frequently asleep throughout the day. Besides this overpowering inappropriate drowsiness, its hallmarks include sleep fragmentation, abnormal REM (rapid eye movements) during sleep, visual hallucinatory dreams, and sudden cataplexy. The last item, brief but extreme muscle paralysis, is the disorder's signature symptom.

When a diagnosis of narcolepsy includes cataplexy, as it does in the Europe, Mignot pointed out, its prevalence runs around one case per 2,000, or some 135,000 people in the population, In the U.S., where this symptom tends to be excluded, the numbers are much higher, well above 150,000. Like its uncertain epidemiology, narcolepsy's cause is unclear. By and large, it doesn't run in families, but arises sporadically in adolescents and young adults, male or female alike, and lasts for life.

"Constant drowsiness," observed Mignot, who heads Stanford's Center for Narcolepsy, "often impairs performance at school, and makes it difficult to hold down a job. Not surprisingly, narcoleptics are also accident-prone, with 10 times the rate of auto accidents as the general population. And because their cataplexy bouts often occur in public, they try to suppress their emotions or shun social situations. Yet, ironically, emotions - especially laughter, anger or joy - are often the triggering event of cataplexic attacks."

Stimulants have been the only therapy for narcolepsy; they treat symptoms, not cause, and often have severe side effects. Early this year, the biotech firm Cephalon Inc., of West Chester, Pa., brought to market a drug trade-named Provigil, which is reportedly free of major adverse reactions. (See BioWorld Today, Dec. 29, 1998, p. 1.)

But a true cure for narcolepsy awaits discovery of a gene or genes of which the mutations are responsible for the disorder. Now a paper in the twice-monthly journal Cell promotes this prospect from possible to probable. Its title: "The sleep disorder canine narcolepsy is caused by a mutation in the hypocretin (orexin) receptor 2 gene." Mignot is its senior author.

Monique Started It All

"This project began about 10 years ago," he told BioWorld Today, "when I realized that because we had this canine model, which showed all the symptoms of narcolepsy as a single gene, it might be a good way to find the intimate causes of the disease, as opposed to fishing and looking at known neurotransmitter levels."

How man's best friend became the animal model of choice for studying narcolepsy is a story that goes back a quarter century. "In the 1970s," Mignot recalled, "my mentor, Dr. William Dement, made the discovery. I think he was giving a lecture on narcolepsy to some congress, and a veterinarian in the audience said, 'I have a dog with exactly that problem. She falls asleep all the time and when excited, goes into REM sleep, and gets paralyzed.' This dog, Monique by name," Mignot went on, "was a French poodle from Canada. They brought her to Stanford, found she had all the symptoms, then did a national search for dogs with narcolepsy, and started the present canine colony."

Mignot brought the saga up to date: "Initially, the scientists tried to breed their narcoleptic animals, but couldn't really establish a genetic transmission. For most of the dog cases, as in most of the human ones, the disorder arises sporadically."

In 1977 they got a litter of Doberman pinschers with several puppies affected. When they bred those, they found a single recessive gene, which a little later turned up in Labradors. "Then," Mignot recounted, "crossbreeding the Labradors with the Dobermans showed the same cistron [DNA coding sequence], most likely the same gene, but they didn't know if it was the same mutation. It turned out not to be, because the mixed breeds did develop narcolepsy."

Mignot joined that scene in 1986, and started the genetic project three years later. For most of the ensuing decade, he and his co-workers tried every trick of genetic linkage analysis then available to hunt down that gene in the canine genome.

"We decided about two years ago," he related, "that the only way we could move this project forward was to build a bacterial artificial chromosome (BAC) library for the dog genome, then clone the region, do real linkage, and narrow down the gene locus."

Different Breeds, Different Defects

"Eventually we started to suspect that the canine chromosome region containing our marker was probably syntenic [partly homologous] to human chromosome 6. We narrowed down to 800 kilobase segments, a region known to contain only one gene. We knew that gene was a good candidate. In May 1999 we found that it had differences in many RFLPs, which led us to suspect that there was something wrong with the gene.

"This turned out to be due to the insertion of a retrotransposon in the hypocretin receptor2 gene in narcoleptic Dobermans, that was producing exon skipping. A different mutation, also producing exon skipping was also observed in Labradors. The resulting transcript was grossly abnormal. And then we wrote this Cell paper."

Their candidate gene turned out to be "very interesting," Mignot said, "because it's a G-protein-coupled receptor for a natural transmitter. Its ligand, named hypocretin, is a peptide in the hypothalamus, with homology to the sequence of the hormone secretin.

"These receptors are probably the most important targets for the pharmaceutical industry," he pointed out, "because I think as many as 30 percent of the medications taken today are proteins acting on these receptors. About 1,000 are known to exist in the genome, so the pharmaceutical industry has been very interested in isolating them all." Mignot made the point: "The researchers know that the same gene exists in humans, and they plan to search for defective versions in narcoleptic individuals.

"With the gene in hand," he continued, "it should be possible to design a drug that can compensate for the failure of the hyposecretin system, and perhaps," he concluded, "produce sleeping pills against insomnia that more closely mimic natural brain chemistry."

The forthcoming issue of Cell, dated Aug. 20, 1999, carries the narcolepsy story from canines to rodents in a paper titled, "Narcolepsy in orexin knockout mice: Molecular genetics of sleep regulation."