New technique shows in detail where drug molecules hit their targets in the body

A team at Scripps Research invented a new method, called CATCH, that shows how drugs hit their targets in the body. Cells targeted by a drug (pargyline shown in cyan) can be identified by multiple rounds of immunolabeling (red showing neurons; yellow showing dopaminergic/noradrenergic neurons; blue showing cell nuclei). Credit: Scripps Research

Scientists at Scripps Research have invented a way to image, across different tissues and with higher precision than ever before, where drugs bind to their targets in the body. The new method could become a routine tool in drug development.

Described in a paper in Cell on April 27, 2022, the new method, called CATCH, attaches fluorescent tags to molecules and uses chemical techniques to improve the fluorescent signal. The researchers demonstrated the method with several different experimental drugs, revealing where—even within —the drug molecules hit their targets.

“This method ultimately should allow us, for the first time, to see relatively easily why one drug is more potent than another, or why one has a particular side effect while another one doesn’t,” says study senior author Li Ye, Ph.D., assistant professor of neuroscience at Scripps Research and The Abide-Vividion Chair in Chemistry and Chemical Biology.

The study’s first author, Zhengyuan Pang, is a graduate student in the Ye lab. The study also was a close collaboration with the laboratory of Ben Cravatt, Ph.D., Gilula Chair of Chemical Biology at Scripps Research.

“The unique environment at Scripps Research, where biologists routinely work together with chemists, is what made the development of this technique possible,” Ye says.

Understanding where drug molecules bind their targets to exert their —and side effects—is a basic part of . However, drug-target interaction studies traditionally have involved relatively imprecise methods, such as bulk analyses of drug-molecule concentration in entire organs.

The CATCH method involves the insertion of tiny chemical handles into drug molecules. These distinct chemical handles don’t react with anything else in the body, but do allow the addition of fluorescent tags after the have bound to their targets. In part because human or animal tissue tends to diffuse and block the light from these fluorescent tags, Ye and his team combined the tagging process with a technique that makes tissue relatively transparent.

In this initial study, the researchers optimized and evaluated their method for “covalent drugs,” which bind irreversibly to their targets with stable chemical bonds known as covalent bonds. This irreversibility of binding makes it particularly important to verify that such drugs are hitting their intended targets.

The scientists first evaluated several covalent inhibitors of an enzyme in the brain called fatty acid amide hydrolase (FAAH). FAAH inhibitors have the effect of boosting levels of cannabinoid molecules, including the “bliss molecule” anandamide, and are being investigated as treatments for pain and mood disorders. The scientists were able to image, at the single-cell level, where these inhibitors hit their targets within large volumes of mouse brain tissue, and could easily distinguish their different patterns of target engagement.

In one experiment, they showed that an experimental FAAH inhibitor called BIA-10-2474, which caused one death and several injuries in a clinical trial in France in 2016, engages unknown targets in the midbrain of mice even when the mice lack the FAAH enzyme—offering a clue to the source of the inhibitor’s toxicity.

In other tests demonstrating the unprecedented precision and versatility of the new method, the scientists showed that they could combine drug-target imaging with separate fluorescent-tagging methods to reveal the cell types to which a drug binds. They also could distinguish drug-target engagement sites in different parts of neurons. Finally, they could see how modestly different doses of a drug often strikingly affect the degree of target engagement in different brain areas.

The proof-of-principle study is just the beginning, Ye emphasizes. He and his team plan to develop CATCH further for use on thicker tissue samples, ultimately perhaps whole mice. Additionally, they plan to extend the basic approach to more common, non-covalently-binding drugs and chemical probes. On the whole, Ye says, he envisions the new method as a basic tool not only for drug discovery but even for basic biology.

“In situ Identification of Cellular Drug Targets in Mammalian Tissue” was co-authored by Zhengyuan Pang, Michael Schafroth, Daisuke Ogasawara, Yu Wang, Victoria Nudell, Neeraj Lal, Dong Yang, Kristina Wang, Dylan Herbst, Jacquelyn Ha, Carlos Guijas, Jacqueline Blankman, Benjamin Cravatt and Li Ye—all of Scripps Research during the study.



More information:
Zhengyuan Pang et al, In situ identification of cellular drug targets in mammalian tissue, Cell (2022). DOI: 10.1016/j.cell.2022.03.040

Journal information:
Cell



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Hexbyte Glen Cove New technique builds super-hard metals from nanoparticles

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This gold “coin” was made from nanoparticle building blocks, thanks to a new technique developed by Brown University researchers. Making bulk metals this way allows for precise of the metal’s microstructure, which enhances its mechanical properties. Credit: Chen Lab / Brown University

Metallurgists have all kinds of ways to make a chunk of metal harder. They can bend it, twist it, run it between two rollers or pound it with a hammer. These methods work by breaking up the metal’s grain structure—the microscopic crystalline domains that form a bulk piece of metal. Smaller grains make for harder metals.

Now, a group of Brown University researchers has found a way to customize metallic grain structures from the bottom up. In a paper published in the journal Chem, the researchers show a method for smashing individual nanoclusters together to form solid macro-scale hunks of solid metal. Mechanical testing of the metals manufactured using the technique showed that they were up to four times harder than naturally occurring metal structures.

“Hammering and other hardening methods are all top-down ways of altering , and it’s very hard to control the you end up with,” said Ou Chen, an assistant professor of chemistry at Brown and corresponding author of the new research. “What we’ve done is create nanoparticle building blocks that fuse together when you squeeze them. This way we can have uniform grain sizes that can be precisely tuned for enhanced properties.”

For this study, the researchers made centimeter-scale “coins” using nanoparticles of gold, silver, palladium and other metals. Items of this size could be useful for making high-performance coating materials, electrodes or thermoelectric generators (devices that convert heat fluxes into electricity). But the researchers think the process could easily be scaled up to make super-hard metal coatings or larger industrial components.

The key to the process, Chen says, is the chemical treatment given to the nanoparticle building blocks. Metal nanoparticles are typically covered with organic molecules called ligands, which generally prevent the formation of metal-metal bonds between particles. Chen and his team found a way to strip those ligands away chemically, allowing the clusters to fuse together with just a bit of pressure.

The metal coins made with the technique were substantially harder than standard metal, the research showed. The gold coins, for example, were two to four times harder than normal. Other properties like electrical conduction and light reflectance were virtually identical to standard metals, the researchers found.

Researchers from Brown University have demonstrated a way to make bulk metals from nanoparticle building blocks. For a new study, the team made metal “coins” from nanoparticles of gold, silver, palladium and other metals . Credit: Chen lab / Brown University

The optical properties of the gold coins were fascinating, Chen says, as there was a dramatic color change when the nanoparticles were compressed into bulk metal.

“Because of what’s known as the plasmonic effect, gold nanoparticles are actually purplish-black in color,” Chen said. “But when we applied pressure, we see these purplish clusters suddenly turn to a bright gold color. That’s one of the ways we knew we had actually formed bulk gold.”

In theory, Chen says, the technique could be used to make any kind of metal. In fact, Chen and his team showed that they could make an exotic form of metal known as a . Metallic glasses are amorphous, meaning they lack the regularly repeating crystalline structure of normal metals. That gives rise to remarkable properties. Metallic glasses are more easily molded than traditional metals, can be much stronger and more crack-resistant, and exhibit superconductivity at low temperatures.

“Making metallic glass from a single component is notoriously hard to do, so most metallic glasses are alloys,” Chen said. “But we were able to start with amorphous palladium and use our technique to make a palladium metallic glass.”

Chen says he’s hopeful that the technique could one day be widely used for commercial products. The chemical treatment used on the nanoclusters is fairly simple, and the pressures used to squeeze them together are well within the range of standard industrial equipment. Chen has patented the technique and hopes to continue studying it.

“We think there’s a lot of potential here, both for industry and for the scientific research community,” Chen said.



More information:
Yasutaka Nagaoka et al, Bulk Grain-Boundary Materials from Nanocrystals, Chem (2021). DOI: 10.1016/j.chempr.2020.12.026

Journal information:
Chem


Citation:
New technique builds super-hard metals from nanoparticles (2021, January 23)
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