Oliver Smithies and his collaborators did not make abetter mousetrap. They made a better mouse _ in fact,two strains of mice, both knockouts. Now the world ofdrug development is beating a path to their doors in NorthCarolina.
Smithies, who heads the department of pathology at theUniversity of North Carolina, in Chapel Hill, is seniorauthor of two back-to-back papers in the current Cell,dated Nov. 3, 1995. Each reports disruption of a mousegene that expresses an enzyme involved in prostaglandinsynthesis.
Prostaglandins are multi-purpose hormones in the body.Aspirin and similar analgesic drugs inhibit its synthesisindirectly. Directly, these so-called non-steroidal anti-inflammatory drugs (NSAID) target two forms of anenzyme responsible for producing prostaglandins.
Thus, these prostaglandin-precursor molecules _cyclooxygenase-1 and -2 _ (COX-1 and COX-2), arebull's eyes for aspirin's function _ and dysfunction.
Aspirin, and related over-the-counter NSAIDs, are themost widely used drugs in human medicine. They includesuch TV-commercial ache-and-pain alleviators asibuprofen (a.k.a. Advil, Motrin), naproxen (Naprosyn),indomethacin and sulindac (Clinoril), among others.
When a patient with a minor complaint phones thedoctor, his conventional advice has often been to "taketwo aspirins, and call me in the morning." Lately, though,aspirin has moved farther front and center in medicalpractice. Half a tablet a day of the familiar pill keepsheart trouble away, according to current wisdom.
But this benefit comes at a cost: Chronic aspirin-poppingcan bring on stomach irritation, even ulceration, andpossibly kidney damage. Two of Oliver Smithies'protoges, Robert Langenbach and Scott Morham, took upthe challenge of tracking the fine molecular line betweenaspirin's effects and side effects. That is, the twomolecular biologists set out to analyze the actions in thebody of COX-1 and COX-2.
Morham, in Smithies' lab at the University of NorthCarolina, chose to knock out the COX-2 gene from mice.His close collaborator, Langenbach, at the NationalInstitute of Environmental Health Sciences (NIEHS) innearby Research Triangle Park, N.C., disrupted the COX-1 gene in his rodents.
"What we did," Langenbach told BioWorld Today, "wasfirst clone a part of the mouse gene _ in my case, the oneencoding COX-1. I isolated that fragment of DNA froman embryonic stem cell, then modified a part of it so itwould disrupt the normal coding sequence, and thus nolonger make a viable protein.
"Then," he continued, "I put that back into a differentpopulation of cells from those in which I originallyisolated the DNA. What occurs is gene-targeting byhomologous recombination. Part of the normal region inthe cell's DNA recombined with the fragment I hadmodified, mutating that gene in the living cell. Finally,we injected that mutant cell into blastocyst-stageembryos, and put it into surrogate mouse mothers. Fromthen on, it was just breeding the mice."
Morham did likewise with COX-2. Their results surprisedthem.
His COX-2-minus mice shrugged off inflammatory drugsas nonchalantly as their normal controls. But theydeveloped severe kidney trouble and a tendency toabdominal infections.
Langenbach's COX-1-missing rodents, on the other hand,"survive well, have no gastric pathology, and show lessdrug-induced gastric ulceration than wild-type mice."This, even though their gastric prostaglandin levels werealmost nil.
This well-being, however, had a down-side: WhenLangenbach mated male and female COX-1 knockouts,nearly all their progeny were born dead.
"Previous to our papers in Cell," Morham told BioWorldToday, "the specific theory has been that the deleteriousside effects of COX-1 are caused by its inhibition, and thebeneficial anti-inflammatory effects by inhibiting COX-2.Our results tend to show that these ideas are not reallyquite correct."
To which Langenbach added, "Pharmaceutical companieshave consequently been working to develop drugs thatinhibit COX-2 without inhibiting COX-1. We've foundthat the situation is probably more complicated, andwe're now trying to determine what other factors areinvolved."
Morham picked up: "That's where our mice come in.New drugs, which inhibit either COX, can be developed.I'm now trying to cross-breed mice in order to knock outboth the enzymes in a single animal."
Between their two labs, he estimates, "we have around athousand knockout and control mice in our animalcolonies."
They intend to offer breeding pairs to basic-scienceresearchers gratis, and to pharmaceutical firms underlicense.
"We've just now received written and verbal expressionsof interest," Langenbach said, "from severalpharmaceutical companies. Even in the short time sincethe Cell papers, we've been contacted by about six firms,plus two biotechnology start-up companies."
The NIEHS researcher said that "These knockout animalstrains probably cost between $300,000 and half a milliondollars each to develop. That's taxpayers' money. If it canbe gotten back from industry, that will fund moreresearch at the University of North Carolina and NIEHS."n
-- David N. Leff Science Editor
(c) 1997 American Health Consultants. All rights reserved.