Bi-Coastal Collaboration Uncovers Novel Activators of Key Cell Receptors

By Eric Sauter

Working in close collaboration, scientists at The Scripps Research Institute in La Jolla, California, and in Jupiter, Florida, have identified several new molecules that activate cell receptors responsible for the regulation of a number of key physiological processes. The molecules have the potential to advance drug development.

The study was published as an ASAP Article on July 1, 2008, by the journal ACS Chemical Biology.

The newly published agonists (an agonist is a molecule that improves the activity of another molecule) interact selectively with one of two subtypes of the Sphingosine 1-phosphate (S1P) receptor system, which helps regulate heart rate, the size of coronary arteries, vessel wall integrity, and lymphocyte trafficking.

"Understanding the role of individual receptors, something that could help in understanding physiology and pathology that underlies successful therapeutics, has been limited by the lack of selective chemical probes for the various S1P sub-types," said Hugh Rosen, a Scripps Research scientist and professor in the Department of Chemical Physiology. "These new selective probes will be of broad use in understanding cell functions—and will help us validate therapeutic targets."

The two receptor subtypes, S1P1 and S1P3, are involved in signaling response to extracellular ligands (small molecules that bind specifically to larger ones), and play an important role in various physiological functions. S1P1 has been linked to immunosuppression in multiple sclerosis, while S1P3 on dendritic cells was recently shown by Wolfram Ruf (Scripps Research Immunology and Microbial Sciences Department) and collaborators to be a potential therapeutic target for sepsis syndrome.

"We chose these receptors as a model system because of their clear medical importance," Rosen said. "Uncovering some of the first validated agonists for S1P3 might serve as the basis for the development of tools for further study of this receptor and its role in septic shock."

The S1P receptors belong to the largest subfamily of G protein-coupled receptors for which high-resolution x-ray structures are available. S1P1 and S1P3 are the most closely related members of this family, sharing approximately 50 percent identity and more than 70 percent similar amino acids.

"The key question is why is S1P3 a more difficult receptor to find good chemical starting points for than S1P1, its close relative." Rosen said.

Using ultra-high-throughput screening, the Scripps Research Molecular Screening Center, a national small molecule discovery center in the National Institutes of Health (NIH) Screening Centers Network, tested more than 80,000 compounds on both receptors to find some answers.

The most remarkable difference was in small molecule agonist recognition, as demonstrated by the very different validated hit rates for the high-throughput screening of the respective receptors. Assays developed by Steven Brown, head of the assay development lab in the Screening Center, and other colleagues in La Jolla were implemented at Scripps Florida by Peter Hodder, lead identification scientific director, and other members of the team. Then in a combined effort across the campuses, Stephan Schürer, Informatics group leader in Florida, used computer modeling ligand docking, while La Jolla colleagues contributed receptor mutagenesis to create models that revealed both similarities and important differences in the binding pockets of the two receptors.

The high-throughput screening process was more effective in finding good agonists with S1P1 than S1P3, Rosen said, and fewer and less potent agonists were identified for S1P3. Chemical residues in the binding pockets caused changes in agonist accommodation. As a result, the S1P1 binding pocket may accommodate more rigid and perhaps larger ligands compared to S1P3. The study also reported the first S1P3 agonist with selectivity against S1P1.

The new study depended on a seamless collaboration in a unified data environment linking both campuses of The Scripps Research Institute. The study focuses primarily on identifying those mechanisms that may become appropriate targets for exploration as modulators of biological function and possible drug candidates.

"What we have done is generate chemical tools that allow us to uncover how human physiology and pathophysiology are wired," Rosen said. "We open up the basic science to gain a deeper understanding, which in turn creates opportunities and multiple approaches that can lead to benefits for patients."

The authors of the study, Ligand-Binding Pocket Shape Differences between S1P1 and S1P3 Determine Efficiency of Chemical Probe Identification by Ultrahigh-Throughput Screening, were Stephan C. Schürer, Steven J. Brown, Pedro Gonzales Cabrera, Marie-Therese Schaeffer, Jacqueline Chapman, Euijung Jo, Peter Chase, Tim Spicer, Peter Hodder, and Hugh Rosen, all of The Scripps Research Institute. See http://pubs.acs.org/cgi-bin/abstract.cgi/acbcct/asap/abs/cb800051m.html .

The study was supported by the Molecular Libraries Initiative of the National Institutes of Health Roadmap.

Send comments to: mikaono[at]scripps.edu

"These new selective probes will be of broad use in understanding cell functions—and will help us validate new therapeutic targets."

—Hugh Rosen