Researchers Unveil COOKIE-Pro for Enhanced Drug Discovery

A team of researchers at Baylor College of Medicine has developed a groundbreaking method for analyzing covalent inhibitors, which are drugs that form strong, permanent bonds with proteins. Published in Nature Communications, the study introduces a technique called COOKIE-Pro, designed to provide a comprehensive and unbiased understanding of how these inhibitors interact with proteins throughout the cell. This advancement aims to accelerate the development of safer and more effective therapeutics.

Covalent inhibitors are known for their effectiveness, exemplified by drugs such as aspirin and the cancer treatment ibrutinib. According to Jin Wang, PhD, director of the Center for NextGen Therapeutics at Baylor and corresponding author of the study, “These drugs can also bind to unintended off-target proteins, potentially leading to unwanted side effects.” The challenge lies in optimizing these drugs to achieve a balance between the strength of their binding and the speed at which they form that bond.

The limitations of existing methods to measure binding strength and reaction speed across the entire proteome have historically hindered drug development. Hanfeng Lin, the study’s first author and a graduate student in the Wang lab, noted, “We knew we needed to measure both affinity and reactivity, but doing it for one protein takes time, let alone thousands.” With COOKIE-Pro, researchers can visualize not only whether a drug binds to off-target proteins but also how effectively and quickly it does so, enabling drug designers to make more informed decisions.

Innovative Approach to Drug Profiling

COOKIE-Pro employs a specially designed “chaser” probe that attaches to protein-binding sites left unoccupied by the drug. Mass spectrometry is then utilized to measure the binding of this chaser probe, allowing researchers to deduce the extent of drug occupation across various proteins. This method enables the simultaneous calculation of binding affinity and inactivation rates for thousands of proteins.

The researchers validated COOKIE-Pro by applying it to two established drugs: spebrutinib and ibrutinib. Findings revealed that spebrutinib, known as a highly selective enzymatic inhibitor, is over 10 times more potent against the off-target protein TEC kinase than its intended target, Bruton’s tyrosine kinase (BTK). In contrast, for the less-selective ibrutinib, COOKIE-Pro successfully identified known off-targets, generating profiles that corroborated previously published data, thus affirming the method’s accuracy.

When researchers applied this high-throughput approach to a library of 16 covalent inhibitor fragments, they generated thousands of profiles, demonstrating the capability of COOKIE-Pro to efficiently guide the early stages of drug discovery.

A Step Towards Safer Medicines

The ultimate ambition of this research is to facilitate rational drug design. Wang emphasized the importance of distinguishing between intrinsic reactivity and true binding affinity. “A drug might appear potent because it binds quickly, but if that is simply because it has a ‘hot’ reactive group, it might cause side effects by binding everywhere.” COOKIE-Pro enables chemists to prioritize compounds that demonstrate potency by binding specifically to the appropriate target, rather than those that are merely reactive.

This innovative method represents a significant advancement in the field of drug discovery, paving the way for the next generation of highly selective and safer covalent medicines. By providing a clearer understanding of covalent inhibitor interactions at a proteome scale, COOKIE-Pro stands to enhance the therapeutic landscape and mitigate the risks associated with off-target interactions.