By David N. Leff
A glance at the human skeleton reveals 26 separate bones in each hand and each foot. Adding to these numbers two ankles, knees, hips, elbows and shoulders gives a grand total of 114 peripheral joints.
Each is a moving target — literally — for inflammation and eventual bone destruction by rheumatoid arthritis (RA).
This mysterious malady — of unknown cause, dubious diagnosis and no cure — afflicts one in 100 people, on average, worldwide. Women are twice as likely as men to get RA, which usually kicks in between 20 and 50 years of age. While the disease is not death-dealing in itself, patients with advanced RA die 10 to 15 years earlier than normal life expectancy predicts, mainly from infections, lung or kidney disorders and gastrointestinal bleeding.
RA usually begins its insidious onslaught on one or more peripheral joints with a barrage of inflammation. Chief instigator of this painful process is a transcription factor called NF-kappaB, which normally lies low but at the ready, in cells throughout the body. Its safety lock is an inhibitory kappaB protein — I-kappaB.
When summoned to action, NF-kappaB turns on a squadron of genes that express pro-inflammatory proteins, notably cytokines and cell adhesion molecules. It gets that summons from a proteolytic enzyme complex, the proteasome-ubiquitin pathway, which — when it gets an RA signal — disarms that I-kappaB safety catch.
Proteasomes have been called "nature's cellular sanitation system." They make up 1 percent of the protein content of cells, and are the main site at which cellular proteins that have outlived their usefulness are chopped up to be recycled. Ubiquitin is a small busybody peptide that triggers proteasome activity. (See BioWorld Today, Feb. 4, 1997, p. 1.)
"The ubiquitin-proteasome pathway," explained molecular biologist Vito Palombella, "is a series of enzyme reactions, which take place when you link the ubiquitin molecule on to a substrate directed to undergo degradation. Once ubiquitin is attached to I-kappaB [that safety-catch for NF-kappaB], this serves as a substrate that amplifies 20 or more further ubiquitins."
Palombella is senior director of molecular biology at ProScript Inc., in Cambridge, Mass., and first author of a paper in the latest Proceedings of the National Academy of Sciences (PNAS), dated Dec. 22, 1998. Its title is "Role of the proteasome and NF-kappaB in streptococcal cell wall-induced polyarthritis."
Polyarthritis is to rats what rheumatoid arthritis is to people.
"Our rats make an excellent animal model of RA," Palombella told BioWorld Today. "We see a lot of histological changes in their bone and cartilage similar to what one sees in human RA. What's really nice about it," he added, "is that we get the same cartilage destruction, the bone erosion, the formation of pannus [inflammatory cells invading the joint space]. Also the same relapses and remissions as in chronic RA."
Modeling Rodents To Mimic Rheumatoid Arthritis
To wreak the symptoms and signs of full-blown polyarthritis on a cohort of rats, ProScript's collaborating co-author, pharmacologist Matthew Grisham at Louisiana State University, in Shreveport, injected them in the abdomen with a brew of potent pro-inflammatory streptococcal cell wall antigenic glycopeptides.
Then, for three or four weeks, the animals were dosed orally with PS341, ProScript's proprietary anti-inflammatory, proteasome-inhibiting small molecule. They developed an acute arthritis within two to four days, which lasted four or five days before partially remitting. About two weeks later, the disease flared up again in more joint-destructive form, which then followed the chronic relapse/remission pattern typical of RA.
Measuring the animals' swollen hind paws enabled the investigators to track subsidence of their arthritic symptoms.
The results of this experiment, their PNAS paper reported, "demonstrate that PS341 attenuates the acute phase and markedly inhibits the chronic phase of the inflammatory response."
"What PS341 does," Palombella explained, "is inhibit the degradation of I-kappaB, the inhibitor of NF-kappa B. If you block this degradation by the proteasome inhibitors, it's still around. It still binds up NF-kappaB, and prevents it from translocating into the nucleus, and activating transcription of pro-inflammatory genes. We discovered PS341 about three years ago, and have a patent on it."
Proteasome Inhibitor Makes Tumors Suicidal
Medicinal chemist Julian Adams, executive vice president of research and development at ProScript, told BioWorld Today: "We've surveyed other medical indications, and found more compelling data to pursue cancer rather than RA at the present time. So, looking for a preferred route toward clinical management of cancer indications, we've begun the first of a series of Phase I clinical trials with PS341 at the M.D. Anderson Cancer Center, in Houston.
"We are to start a second Phase I trial in January at Sloan-Kettering Cancer Center in New York," Adams said, "and have recently signed a CRADA [Cooperative Research and Development Agreement] with the National Cancer Institute. We are considering five or six additional Phase I studies to pursue a diverse range of cancers, trying to optimize the best mode utility for PS341."
Palombella said the trials "will give us a lot of experience on how to dose proteasome inhibitors in human patients. So, we may go back later with this class of compounds to treat inflammatory diseases, such as RA."
Describing the compound's mode of action in cancer, he explained, "The proteasome is also involved in regulating cell-cycle progression. A number of key regulatory proteins are degraded via the ubiquitin-proteasome pathway as the cell progresses through the cell cycle. And our proteasome inhibitor blocks degradation of these particular regulatory proteins, essentially causing the tumor cell to stop in the cell cycle, and eventually undergo apoptosis."
Meanwhile, he said, ProScript "has another class of proteasome inhibitors, which we are following up for stroke. For instance, when you get an ischemic event, there is an influx of neutrophils into the damaged or hypoxic region of the brain. There the neutrophils are activated, and they cause a lot of the damage.
"What we see," Palombella concluded, "is that this class of proteasome inhibitors will prevent that from happening. So, we are in preclinical development now for this proteasome inhibitor in the area of stroke." *