By David N. Leff

Obesity packs a double whammy — mental as well as physical.

In Western society, being overweight, or downright fat, can cause severe social and emotional anxiety in girls and women, leading in extremis to life-threatening anorexia nervosa — self-inflicted starvation.

Grossly corpulent men suffer perhaps fewer and milder mental traumas, but they share with obese females the very high risk of contracting familial diabetes mellitus and its horrendous down-the-road complications.

Mice too get fat, and get diabetes. This makes them convenient models for studying the human maladies. One obese murine surrogate, the ob/ob mouse, is born bereft of the gene that encodes the appetite-suppressing hormone, leptin.

So far, only a single human family has surfaced with this particular cause of excess weight, a leptin deficiency. "They were identified only a couple of months ago," observed molecular biologist Varavani Dwarki, "and already their kids are starting to show weight increase."

He pointed out that not all human, or murine, obesity is due to a shortage of leptin, "but if a human case does occur because of leptin deficiency, I think it will follow the ob/ob mouse model." Dwarki is associate director of gene delivery and targeting at Chiron Corp., in Emeryville, Calif.

As such, he is senior author of a paper in the current Proceedings of the National Academy Of Sciences (PNAS), dated Dec. 9, 1997. Its title: "Long-term correction of obesity and diabetes in genetically obese mice by a single intramuscular injection of recombinant adeno-associated virus encoding mouse leptin."

"The way this experiment was done," Dwarki told BioWorld Today, "we wanted to do two things: One, take ob/ob mice that had not yet become obese, and demonstrate by gene therapy whether we can prevent them from becoming fat, and also from getting diabetes."

His second goal was to construct, test and prove a gene delivery vehicle that could correct a chronic genetic disorder for a long period of time.

For starters, he and his co-authors at Chiron selected as their vector a recombinant adeno-associated virus (rAAV), "because it is nonpathogenic, can infect both dividing and nondividing cells, and lacks potentially immunogenic virally encoded genes."

Into this delivery vehicle they bundled a normal mouse leptin gene, poised to express the 16- or 17-amino-acid protein.

Early this year, they injected this packaged virus into the leg muscles of ob/ob mice. Control animals received simple saline injections.

A New Home For Leptin Genes

Targeting muscle cells "was a very important thing," Dwarki observed, "because gene therapists tend to assume that they have to replace the protein at the site in the body where it is normally made. Leptin is normally made in adipose tissues, in the fat cells."

He pursued this point: "In theory, if you say, OK, you'll put back the gene only in the fat cells, here we have shown that you can put it in the muscle, as a depot or store for continuous leptin secretion.

"Remember," Dwarki continued, "this hormone, which is being made in the muscles of our ob/ob mice right now, has to get into the circulation, go to the brain and act in the hypothalamus, which is where the leptin receptor is located."

At eight weeks into their post-therapy, the treated cohort passed their glucose tolerance tests on a par with perfectly normal, non-obese mice.

"The reason it has an effect on diabetes," Dwarki continued, "is that the hormone also goes to the pancreas, and their islet cells, which also have receptors for leptin."

The mice got their single vector shot at the age of four to six weeks, just at the point in life when ob/ob mice begin to put on weight. Over the first three weeks, they went up from an initial 25 grams to 35 grams. So did the control rodents. The latter kept right on overeating and adding grams, while the treated rodents dipped down, and stayed down, to an average 30 grams at six months after their gene therapy. Since then, now eight months out, they maintain that slenderizing body weight and, Dworki said, "are living happily, running around, looking healthy." But the controls tip their scales at a chubby-plus 80 grams.

Similar Study Also Succeeded

Chiron's just-published experiment is not the first attempt to treat incipient obesity and diabetes in ob/ob mice with gene therapy. Exactly one year ago, in PNAS dated Dec. 10, 1996, pioneer gene therapist Savio Woo, then at Baylor College of Medicine, in Houston, reported treating the obesity and diabetes-challenged rodents with recombinant adenovirus vectors (rAVV) freighting in the leptin cDNA.

Sure enough, the one-shot therapy dropped their weight "dramatically," and normalized insulin and glucose levels. But the improvements were short-lived. (See BioWorld Today, Dec. 18, 1996, p. 1.)

Other groups have tried injecting the leptin protein itself into ob/ob mice. This therapy too reduced body weight, food intake and fat mass, but like insulin, the injections would have to be repeated continuously and for life.

"We already know that our delivery system is manageable," Dwarki said. "One thing we need to figure out is how do we scale this up for large-animal, subhuman primate trials, which we want to do for our next study?

"But obviously," he pointed out, "working for a company, we have to have all the rights. We have quite a few research targets from which we'd like to choose."

Beyond obesity and diabetes, he sees his gene therapy construct as treating "any disease in which you need therapeutic protein constantly in the circulation. For single-gene defects," he concluded, "such as lipid storage diseases and clotting-factor deficiencies, where you want to replace the hormone or factor, this will be a fantastic way to go about it." *

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