New drug targets for fighting cancer could potentially slow the growth of cancer cells

Effectively fighting cancer often requires stopping cancer cells from multiplying. This requires understanding the proteins the cells need to survive. Protein profiles play a crucial role in this process, helping researchers identify proteins and their specific components that future drugs should target. However, when used alone, previous approaches are not detailed enough to capture all potential protein targets, resulting in some being missed.

By combining two methods of protein analysis, a team of chemists at Scripps Research has now mapped more than 300 small molecule-responsive cancer proteins, as well as their small molecule binding sites. The discovery of key protein targets that, when disrupted with specific chemical compounds (or small molecules), stop cancer cell growth could ultimately enable the development of more effective and precise cancer treatments. The results were published in Natural Chemistry on August 13, 2024.

“One method gave us a comprehensive view of which proteins were interacting with the chemicals, and the second method showed exactly where those interactions were taking place,” says co-senior author Benjamin Cravatt, PhD, the Norton B. Gilula Chair in Biology and Chemistry at Scripps Research.

Both methods are forms of activity-based protein profiling (ABPP), a technique Cravatt developed to capture protein activity on a global scale. The research team used their dual approach to label both the proteins and the protein sites that interacted with a library of stereoprobes – chemical compounds designed to bind permanently to proteins in a selective manner. Stereoprobes are used to study protein functions and identify potential drug targets.

“We made a conscious effort to design our stereoprobes with chemical features that tend to be underrepresented in compounds commonly used in drug discovery,” says co-senior author Bruno Melillo, PhD, an institute investigator in the Department of Chemistry at Scripps Research. “This strategy increases our chances of making discoveries that can advance biology and ultimately translate into improvements in human health.”

The research team's stereoprobes were electrophilic, meaning they were designed to bind irreversibly to proteins — specifically cysteine. This amino acid is ubiquitous in proteins, including those of cancer cells, and helps form important structural bonds. When chemicals react with cysteine, they can disrupt these bonds and cause proteins to malfunction, impairing cell growth. Many cancer drugs bind irreversibly to cysteines on proteins.

“We also focused on cysteine ​​because it is the most nucleophilic amino acid,” says lead author Evert Njomen, PhD, a HHMI Hanna H. Gray Fellow at Scripps Research and postdoctoral fellow in Cravatt’s lab.

To find out which specific proteins would bind to the stereoprobes, the team resorted to a method called protein-targeted ABPP. Using this approach, the researchers discovered more than 300 individual proteins that reacted with the stereoprobe compounds. But they wanted to dig even deeper and identify the exact locations of the reactions.

The second method, cysteine-targeted ABPP, pinpointed exactly where the stereoprobes bound to the proteins. This allowed the team to “zoom in” on a specific protein pocket and examine whether the cysteine ​​within it reacted with the stereoprobes, similar to focusing on a single point on a puzzle board to see if a particular piece fits.

Each stereoprobe molecule had two main components: the binding part and the electrophilic part. Once the binding component recognizes the cancer cell's protein pocket, the stereoprobe molecule can hopefully enter – just like a key must fit into a lock. When a stereoprobe remained in a pocket critical to the cancer cell's function, it blocked the protein from binding to other proteins – and ultimately prevented cell division.

“By attacking these very specific phases in the cell cycle, we can slow down the growth of cancer cells,” says Njomen. “A cancer cell would remain in what almost a two-cell condition, and your body's immune system would recognize them as defective and order their death.”

Identifying precise protein regions that are critical for cancer cell survival could help researchers develop more targeted treatments to stop cell proliferation.

Other key findings from the team included confirmation that their dual-pronged approach provided a more accurate picture of protein stereoprobe reactivity than a single method.

“We always knew that both methods have their drawbacks, but we didn't know exactly how much information was lost when we used only one technique,” says Njomen. “It was surprising to see that a significant number of protein targets were missed when we used one platform instead of the other.”

The team hopes that their findings will one day lead to new cancer therapies that target cell division. In the meantime, Njomen wants to design new stereoprobe libraries to uncover protein pockets involved in diseases other than cancer, including inflammatory diseases.

“Many proteins are associated with disease, but we don't have stereoprobes to study them,” she said. “In the future, I would like to find more protein pockets that we can study for drug discovery.”

Reference: Njomen E, Hayward RE, DeMeester KE, et al. Multilevel chemical proteome maps of tryptoline-acrylamide-protein interactions in cancer cells. Nat Chem. 2024:1-13. doi: 10.1038/s41557-024-01601-1

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