Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

New Inhibitors of Elusive Enzymes Promise to Be Valuable Scientific Tools

01.11.2012
The work has implications for developing approaches to inflammation

Scientists at The Scripps Research Institute (TSRI) have discovered the first selective inhibitors of an important set of enzymes.

The new inhibitors, and chemical probes based on them, now can be used to study the functions of enzymes known as diacylglycerol lipases (DAGL), their products, and the pathways they regulate. Early tests in mouse macrophages suggest that DAGL-inhibiting compounds might also have therapeutic uses, for they suppress the production of a pro-inflammatory molecule that has been implicated in rheumatoid arthritis and related conditions.

“We’ve developed the first set of chemical probes that effectively allows one to study these DAGL enzymes in living cell and animal models,” said Benjamin F. Cravatt, chairman of the Department of Chemical Physiology, professor in the Dorris Neuroscience Center and member of the Skaggs Institute for Chemical Biology at TSRI. Cravatt and his laboratory conducted the new study, published in the current issue of the journal Nature Chemical Biology.

Important But Poorly Understood
DAGL enzymes have been of interest mainly because of their role in making 2-AG (2-Arachidonoylglycerol), an important cannabinoid that is naturally produced in humans and other mammals. Cannabinoids are named for Cannabis (marijuana) plants, because they stimulate the same cellular receptors that are hit by marijuana’s active ingredients. Drugs that can enhance 2-AG’s signaling in the nervous system are being developed as treatments for pain, depression and anxiety.
But 2-AG exists in various tissues throughout the body, and on the whole, its functions are not well understood. Until now researchers have lacked enzyme inhibitors that can usefully probe those functions by selectively shutting off 2-AG’s production. “Existing DAGL inhibitors block many other enzymes, are not very potent, and do a poor job of getting into cells,” Cravatt said. “There has been a need for better chemical tools in this area.”

Cravatt’s laboratory had previously developed a set of compounds that act as potent inhibitors of serine hydrolases—the broad enzyme family to which DAGL enzymes belong. In the new study, Cravatt’s team, including first author Ken Hsu, a Hewitt Foundation postdoctoral researcher in the Cravatt laboratory, screened a library of these compounds for specific activity as DAGL inhibitors.

A Big Improvement

After finding a promising lead compound, Hsu and his colleagues chemically optimized it to obtain KT109 and KT172. The former selectively inhibits DAGLâ, the main enzymatic producer of 2-AG outside the nervous system. KT172 inhibits both DAGLâ and DAGLá, which is principally responsible for making 2-AG within the nervous system.

In a big improvement over previously described DAGL inhibitors, KT109 and KT172 are highly selective (i.e., they do not block many other, non-DAGL enzymes) and active in cells and animals. By analyzing the structures of their initial DAGL inhibitors, the team was also able to devise a new DAGL-tailored activity-based probe that binds to the active site of DAGLs and fluorescently labels these low-abundance and difficult-to-detect enzymes in cell or tissue samples. “Without the DAGL-specific probe, we would have found it very difficult to develop, optimize and confirm target engagement for our DAGL inhibitors,” Hsu said.

In neuron-like mouse cells, human prostate cancer cells, and mouse liver cells and macrophages (a type of immune cell that is frequently involved in inflammatory conditions), the DAGL inhibitors were able to inactivate DAGLâ activity. “At the optimal doses used, we were able to achieve selective and near-complete inhibition of the enzyme,” said Hsu. In these cell and animal studies, the inhibitors also reduced levels of 2-AG as well as arachidonic acid, another bioactive lipid that DAGL enzymes can regulate.

New Questions

2-AG is known to have an anti-inflammatory effect when it activates cannabinoid receptors on macrophages. Thus, one might expect that knocking down 2-AG production with a DAGL inhibitor would have a pro-inflammatory effect. Instead, Hsu, Cravatt and their colleagues found that blocking DAGL in mouse macrophages that had been stimulated with pro-inflammatory agents markedly lowered their secretion of TNFá, a major inflammatory signaling molecule.
Blocking DAGL has potential effects on multiple lipid signaling pathways in cells, and the researchers aren’t yet certain which of these effects explains the surprising suppression of TNFá. “The effect is dependent on DAGLâ, though, because we see the same result in DAGLâ knockout mice,” said Hsu. Cravatt added that their observations of the unexpected DAGL-inhibition effects in mouse macrophages could be due to the suppression of pro-inflammatory eicosanoids that derive from downstream metabolites regulated by DAGLâ.

TNFá is a key instigator of the inflammation seen in rheumatoid arthritis, and antibodies directed against TNFá are now front-line therapies for the condition. “What we’ve done so far is just early-stage cell biology, but conceivably the further optimization of our DAGL inhibitors could result in a new type of anti-inflammatory drug that also works against arthritis and related conditions,” Cravatt said.

Cravatt and his team are now studying the pathways through which the new inhibitors have this anti-inflammatory effect. They also plan to develop new inhibitors that will selectively block DAGLá and 2-AG production in the central nervous system.

The other co-authors of the study, “DAGLâ inhibition perturbs a lipid network involved in macrophage inflammatory responses,” were Katsunori Tsuboi, Alexander Adibekian, Holly Pugh and Kim Masuda, all of the Department of Chemical Physiology at TSRI.

The research was supported by grants from the National Institutes of Health (DA009789, DA033760, MH084512) and a Hewitt Foundation Postdoctoral Fellowship.

About The Scripps Research Institute

The Scripps Research Institute is one of the world's largest independent, not-for-profit organizations focusing on research in the biomedical sciences. Over the past decades, Scripps Research has developed a lengthy track record of major contributions to science and health, including laying the foundation for new treatments for cancer, rheumatoid arthritis, hemophilia, and other diseases. The institute employs about 3,000 people on its campuses in La Jolla, CA, and Jupiter, FL, where its renowned scientists—including three Nobel laureates—work toward their next discoveries. The institute's graduate program, which awards Ph.D. degrees in biology and chemistry, ranks among the top ten of its kind in the nation. For more information, see www.scripps.edu.
For information:
Office of Communications
Tel: 858-784-8134
Fax: 858-784-8136
press@scripps.edu

Mika Ono | EurekAlert!
Further information:
http://www.scripps.edu

More articles from Life Sciences:

nachricht New type of photosynthesis discovered
17.06.2018 | Imperial College London

nachricht New ID pictures of conducting polymers discover a surprise ABBA fan
17.06.2018 | University of Warwick

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: AchemAsia 2019 will take place in Shanghai

Moving into its fourth decade, AchemAsia is setting out for new horizons: The International Expo and Innovation Forum for Sustainable Chemical Production will take place from 21-23 May 2019 in Shanghai, China. With an updated event profile, the eleventh edition focusses on topics that are especially relevant for the Chinese process industry, putting a strong emphasis on sustainability and innovation.

Founded in 1989 as a spin-off of ACHEMA to cater to the needs of China’s then developing industry, AchemAsia has since grown into a platform where the latest...

Im Focus: First real-time test of Li-Fi utilization for the industrial Internet of Things

The BMBF-funded OWICELLS project was successfully completed with a final presentation at the BMW plant in Munich. The presentation demonstrated a Li-Fi communication with a mobile robot, while the robot carried out usual production processes (welding, moving and testing parts) in a 5x5m² production cell. The robust, optical wireless transmission is based on spatial diversity; in other words, data is sent and received simultaneously by several LEDs and several photodiodes. The system can transmit data at more than 100 Mbit/s and five milliseconds latency.

Modern production technologies in the automobile industry must become more flexible in order to fulfil individual customer requirements.

Im Focus: Sharp images with flexible fibers

An international team of scientists has discovered a new way to transfer image information through multimodal fibers with almost no distortion - even if the fiber is bent. The results of the study, to which scientist from the Leibniz-Institute of Photonic Technology Jena (Leibniz IPHT) contributed, were published on 6thJune in the highly-cited journal Physical Review Letters.

Endoscopes allow doctors to see into a patient’s body like through a keyhole. Typically, the images are transmitted via a bundle of several hundreds of optical...

Im Focus: Photoexcited graphene puzzle solved

A boost for graphene-based light detectors

Light detection and control lies at the heart of many modern device applications, such as smartphone cameras. Using graphene as a light-sensitive material for...

Im Focus: Water is not the same as water

Water molecules exist in two different forms with almost identical physical properties. For the first time, researchers have succeeded in separating the two forms to show that they can exhibit different chemical reactivities. These results were reported by researchers from the University of Basel and their colleagues in Hamburg in the scientific journal Nature Communications.

From a chemical perspective, water is a molecule in which a single oxygen atom is linked to two hydrogen atoms. It is less well known that water exists in two...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

VideoLinks
Industry & Economy
Event News

Munich conference on asteroid detection, tracking and defense

13.06.2018 | Event News

2nd International Baltic Earth Conference in Denmark: “The Baltic Sea region in Transition”

08.06.2018 | Event News

ISEKI_Food 2018: Conference with Holistic View of Food Production

05.06.2018 | Event News

 
Latest News

A sprinkle of platinum nanoparticles onto graphene makes brain probes more sensitive

15.06.2018 | Materials Sciences

100 % Organic Farming in Bhutan – a Realistic Target?

15.06.2018 | Ecology, The Environment and Conservation

Perovskite-silicon solar cell research collaboration hits 25.2% efficiency

15.06.2018 | Power and Electrical Engineering

VideoLinks
Science & Research
Overview of more VideoLinks >>>