PXR: Switch to Body's Garbage Disposal

For some time now, scientists have known about a superfamily of 48 ligand-activated transcription factors that are the body's way of regulating the expression of genes inside cells in response to hormonal ligand signals. Ligands for these receptors are usually small lipid-soluble molecules that enter the cell either by diffusion or transport, and bind to and activate their respective receptors. Some of the more familiar ligands, or hormones,include estrogen, progesterone and thyroid hormones, each of which has its own cognate receptor in the cell. The ligand-receptor complex is then able to bind to specific DNA sequences located in the promoter region of target-genes and turn them on and off. For example, genes that are regulated by the estrogen receptor contain response elements that are specifically triggered by the liganded-estrogen receptor complex.

The newest assistant professor in the KU's Department of Pharmacology and Toxicology, Jeff Staudinger, has spent the last four years characterizing the ligand-activated transcription factor molecule called the pregnane-x-receptor, or PXR. PXR is a member of the nuclear receptor super-family of transcription factors whose function is activated by a wide array of drugs instead of by one specific ligand or hormone. "It can be thought of as a broad-specificity 'drug-receptor' as opposed to a 'hormone-receptor'," he explained. "PXR activation by drugs regulates the expression of genes in the liver and intestine that eliminate toxins we encounter from our environment, or in other words, PXR regulates the bodies 'garbage-disposal system.'"

Staudinger's research has focused primarily on PXR's role in liver cells. "Everyone knows that the liver is the filtration organ of the body," he said. "Using the tools of molecular biology, more specifically transgenic and knockout mouse technology, we've been able to show that PXR is a broad-specificity 'drug sensor' that regulates the expression of certain drug hydroxylases, or drug-metabolizing enzymes in liver cells. PXR also regulates the expression of specific molecular pumps that transport many of these drugs into and out of liver cells; eventually placing them into the bile and helping us eliminate them from our system. For example, if you wake up in the morning and you take a drug that activates PXR, PXR will then up-regulate the biochemical pathway in our bodies through which these drugs are metabolized and eventually disposed of."

For about 40 years toxicologists have known that, when pre-treated with certain drugs and steroids, rodents were protected from subsequent exposure to a wide variety of toxins. They discovered they would have to give these rodents 10 to 100 fold more of a toxin for them to succumb to it, though the molecular mech-anism behind this protective effect was not clear. With the discovery and characterization of PXR as a broad-specificity "drug-sensor," Staud-inger's research has been able to shine the light of understanding on the molecular mechanism of this protection. cont. on page 4

"The 'eureka' moment came when we identified these drugs as PXR activators," said Staudinger. "Previous research had shown that when you pre-treated rodents with these drugs, they were protected against subsequent challenge with toxic substances. We used transgenic and knockout mice to show that PXR is the master switch that regulates drug-inducible transcription of a group of genes that function in a coordinated manner to help our bodies eliminate a vast array of toxins. That is, mice that did not have this molecular master-switch were unable to regulate the expression of these genes in response to drugs."

Though certainly there are other receptors that play a similar role in the body, PXR appears to be the most promiscuous of these because it responds to a vast array of clinically prescribed drugs. This is why Staudinger's research could have a huge impact upon the process of bringing novel drug therapeutics to market in the future. Understanding PXR's role has already been vital to treating patients who have been prescribed a combination of drugs and may suffer the ill effects of drug-drug interactions.

A classic example of the problem of drug-drug interactions is found in HIV patients who are on combination therapy and protease inhibitors. Often these patients develop yeast infections in their lungs. To control this infection, they are prescribed a lipophilic anti-infective agent called rifampicin, a very potent and efficacious PXR activator. When HIV patients begin taking rifampicin, a large increase in PXR-mediated drug metabolism and elimination is induced, thereby reducing the serum levels of the anti-HIV medications including the protease inhibitors. "It is very important to know if a new drug coming on the market is a PXR activator," Staudinger said. "We have developed high-throughput screening tools in the lab that enable us to predict if a potential therapeutic is going to have a drug-drug interaction via this molecular mechanism."

PXR does not exist in just liver cells. The mechanism exists in cells in the intestine, the kidneys, and there's even some indication that it exists in brain cells as well. Staudinger believes PXR may be involved in the blood-brain barrier, something that has confounded many attempts to administer drugs.

"The blood brain barrier is a biological protection mechanism that has evolved to keep toxins from reaching brain tissue," Staudinger explained. "But when clinicians seek to get therapeutics into the brain they have this amorphous blood-brain barrier to deal with. There is some anecdotal evidence suggesting that- and I believe that-PXR may be involved in this process. "

Also, interestingly, both the placenta and the ovaries also express PXR. These tissues also express certain genes that protect them from toxicity. As you might imagine, it's very important to regulate the level of toxins in the fetus during pregnancy. That is another frontier of PXR research I'd like to pursue, understanding PXR's role in protecting the fetus."

At KU Staudinger will continue to use transgenic or gene knockout technology to further define the role of these PXR target genes in the body and determine which drugs they remove. He will also use gene array or gene chip technology to identify additional drug-inducible PXR-target genes. In 1998, while doing a post-doctorate at GlaxoWellcome (now Glaxo-SmithKline), a pharmaceutical company specializing in HIV-related drugs, Staudinger collaborated with researchers from the University of North Carolina to develop a genetically engineered mouse, called a knockout mouse, that lacks a functional PXR gene.

With this population of PXR knockout mice in place, Staudinger has been able to ask some very broad questions. For example, what are the changes in gene expression in a wild type mouse when you give them a drug vs. when you give a PXR knockout mouse the same drug? Staudinger said, "If you see changes in gene expression in the mouse that lacks the PXR switch, it's likely that the gene is not regulated by PXR. When you're asking questions about 30,000 genes at a time, it's nice to have that control mouse to weed out the non-PXR-specific responses."

Staudinger just received a research-development fund award from KU's Center for Research. This award provided the necessary funds to bring this innovative gene knockout technology to the University of Kansas. "The Center for Research saw that KU was lagging behind other major research institutions that already had developed this technology. To their credit, they funded our proposal to create a transgenic and knockout mouse facility in the Department of Pharmacology and Toxicology for use by all researchers on the Lawrence campus. We've had a lot of support from the Pharmacology and Toxicology Department and from (HBC Director) Eli Michaelis in particular. He was instrumental in bringing me here, and we're all really excited about the potential uses of this technology at the KU-Lawrence campus."

Staudinger received his Ph.D. at the School of Biomedical Sciences at the University of Texas-Houston.