Research could lead to ways to prevent arrhythmias

Aug 20 2004

The worm C. elegans seems an unlikely candidate for studies related to cardiac arrhythmias. After all, the microscopic organism doesn't even have a heart.

That fact did not deter Christina I. Petersen, Ph.D., research assistant professor of Anesthesiology at Vanderbilt University Medical Center. Petersen and colleagues, including Jeffrey R. Balser, M.D., Ph.D., associate vice chancellor for Research at Vanderbilt, have now developed a C. elegans-based screening tool to identify novel protein regulators of a potassium channel involved in the rhythmic heartbeat.

They report the success of their approach in the Aug. 10 issue of the Proceedings of the National Academy of Sciences. The tool could lead to ways to prevent arrhythmias and sudden death associated with block of a potassium channel called HERG (Human ether-a-go-go related gene).

Many therapeutic drugs -- antihistamines, antidepressants, antibiotics -- block HERG, Petersen said. This undesired blockade can cause acquired long QT syndrome, a disorder that puts patients at risk for life-threatening arrhythmias. It is not clear why some individuals receiving a HERG-blocking drug develop the syndrome, Petersen said.

"It's a serious problem," she said. "The FDA now requires that every drug be assayed for HERG block before it is approved." Even medicines that might be beneficial for the vast majority of patients do not make it to the market – or have been pulled from the market -- if they block HERG, Petersen said.

"The pharmaceutical industry is very interested in finding ways to screen for susceptibility to acquired long QT syndrome and in discovering how to prevent it."

Other investigators have demonstrated that variations in HERG itself do not account for the majority of interindividual differences. Petersen and colleagues suspected that other proteins -- modulating factors -- are influencing HERG channel susceptibility to drug block.

That's where the worm came in. "This paper is really a proof of principle of the utility of using C. elegans to search for these HERG-interacting proteins," Petersen said.

Even though the worm doesn't have a heart, it does have a pharynx -- a muscular tube that "pumps" food inside. The pharynx, Petersen said, is similar in many ways to a heart. It has an intrinsic beating -- like a heartbeat -- that responds to neuronal input by speeding up or slowing down.

The investigators were able to take advantage of a worm line with a mutation in the worm version of HERG. The mutation, unc-103, causes profound neuromuscular defects, including a pharyngeal pumping defect that Petersen noticed as she learned to work with the worms.

"I remember the first time I saw it (the pumping defect)," she said. "It looked just like a cardiac arrhythmia." Whereas the normal worm pharynx has a rhythmic "beat" and rarely pauses, Petersen said, the unc-103 pharynx has long stretches of pauses, "as if the worms are having an arrhythmia."

The team developed a method to quantitate the pauses and demonstrated that "knocking out" the unc-103 protein using a technique called RNA interference restored normal pharyngeal pumping. The investigators reasoned that if they knocked out important HERG modulators, the effect would be the same: a restoration of rhythmic pharyngeal pumping.

To test this concept, they chose several candidate genes known to interact with HERG or other related potassium channels, knocked them out, and looked for improvements in the pumping defect. For two of the candidates, pumping improved -- the arrhythmia got better. One of these was a protein called KCR1, which Sabina Kupershmidt, Ph.D., assistant professor of Anesthesiology, and colleagues at Vanderbilt recently demonstrated prevents drug block of HERG in cardiac cells grown in the laboratory.

"We were really excited," Petersen said. "We already knew that KCR1 affected drug block of HERG in cells, and now we had data in vivo, in a living animal, that KCR1 was important for HERG function."

The investigators then turned to Ping Yang, Ph.D., research instructor and Dan M. Roden, M.D., director of the division of Clinical Pharmacology, both at Vanderbilt, to look for evidence that KCR1 impacts acquired long QT syndrome. Using a database of patients who developed acquired long QT syndrome after drug treatment, Yang was able to detect a variant in the KCR1 gene that occurred more often in control patients -- those who did not develop the syndrome.

"We suspect that this variant is protective, that patients who have it are somehow more protected against drug block of HERG compared to patients who don't have it," Petersen said. By engineering the variant into KCR1 and studying it in cells, the researchers confirmed that drugs do not block HERG as readily when the variant KCR1 is present.

"We're very encouraged because these studies demonstrate that you can identify a protein in the worm – which seems totally unrelated to humans -- get a phenotype in vivo, and find evidence in humans that suggests this protein is relevant to cardiac potassium channel function," Petersen said. "We speculate that KCR1 might be useful in a therapeutic context to prevent HERG block." The investigators are now using the C. elegans tool to search for novel interacting proteins that modulate HERG drug block. Any new proteins they find may have therapeutic potential for preventing acquired long QT syndrome.

In addition to Petersen, Balser, Yang, and Roden, authors of the PNAS paper include Toni R. McFarland, Svetlana Z. Stepanovic, Kenshi Hayashi, M.D., Ph.D., and Alfred L. George, M.D., at Vanderbilt and David J. Reiner, Ph.D., and James H. Thomas, Ph.D., at the University of Washington. The research was supported by the National Institutes of Health.

Balser is the James Taloe Gwathmy Clinician-Scientist and professor of Anesthesiology at Vanderbilt. Roden is the William Stokes Professor of Experimental Therapeutics and professor of Medicine and Pharmacology at Vanderbilt.

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