A new era in drug development

For decades, a significant number of proteins, essential for the treatment of various diseases, have remained inaccessible to oral drug treatments. Traditional small molecules often have difficulty binding to proteins with flat surfaces or require specificity for particular protein homologs. Typically, larger biologics that can target these proteins require an injection, limiting patient comfort and accessibility.

In a new study published in Nature Chemistry Biology, Scientists from the laboratory of Professor Christian Heinis at EPFL have reached an important milestone in drug development. Their research opens the door to a new class of orally available drugs, addressing a long-standing challenge for the pharmaceutical industry.

“There are many diseases for which targets have been identified, but drugs to reach and reach them have not been able to be developed,” says Heinis. “Most of these are cancer types, and many of the targets in these cancers are protein-protein interactions that are important for tumor growth but cannot be inhibited.”

The study focused on cyclic peptides, which are versatile molecules known for their high affinity and specificity in binding to difficult pathological targets. At the same time, the development of cyclic peptides as oral drugs has proven difficult because they are quickly digested or poorly absorbed from the gastrointestinal tract.

“Cyclic peptides are of great interest for drug development because these molecules can bind to difficult targets for which it has been difficult to generate drugs using established methods,” says Heinis. “But cyclic peptides generally cannot be administered orally, in pill form, which greatly limits their application.”

Cycling breakthrough

The research team targeted the enzyme thrombin, which is a critical disease target due to its central role in blood clotting; Thrombin regulation is essential for the prevention and treatment of thrombotic disorders such as strokes and heart attacks.

To generate cyclic peptides capable of targeting thrombin and sufficiently stable, scientists developed a two-step combinatorial synthesis strategy to synthesize a large library of cyclic peptides with thioether linkages, which improve their metabolic stability when taken by oral route.

“We have now succeeded in generating cyclic peptides that bind to a pathological target of our choice and can also be administered orally,” explains Heinis. “To this end, we developed a new method in which thousands of small cyclic peptides with random sequences are chemically synthesized at the nanoscale and examined in a high-throughput process.”

Two steps, one pot

The new process has two steps and takes place in the same reagent container, a feature chemists call “a pot.”

The first step involves synthesizing linear peptides, which then undergo a chemical process to form a ring-shaped structure, in technical terms, being “cyclized”. This is done by using “bis-electrophilic linkers” – chemical compounds used to connect two molecular groups – to form stable thioether bonds.

In the second phase, the cyclized peptides undergo acylation, a process that attaches carboxylic acids to them, further diversifying their molecular structure.

The technique eliminates the need for intermediate purification steps, enabling high-throughput screening directly in synthesis plates, combining the synthesis and screening of thousands of peptides to identify candidates with high affinity for specific disease targets, in this case, thrombin.

Using the method, Ph.D. The student leading the project, Manuel Merz, was able to generate a comprehensive library of 8,448 cyclic peptides with an average molecular mass of approximately 650 Daltons (Da), slightly above the maximum limit of 500 Da recommended for small molecules available orally.

Cyclic peptides also showed strong affinity for thrombin.

When tested on rats, the peptides showed an oral bioavailability of up to 18%, meaning that when the cyclic peptide drug is taken orally, 18% of it successfully enters the blood circulation and has a therapeutic effect. Since orally administered cyclic peptides typically have a bioavailability of less than 2%, increasing this figure to 18% is a substantial advancement for drugs in the biologics category, which includes peptides.

Define aims

By enabling the oral availability of cyclic peptides, the team has opened up possibilities for treating a range of diseases that are difficult to treat with conventional oral medications. The versatility of the method means it can be adapted to target a wide range of proteins, which could lead to advances in areas where medical need is currently unmet.

“To apply the method to more difficult disease targets, such as protein-protein interactions, it will likely be necessary to synthesize and study larger libraries,” explains Manuel Merz. “By automating other steps of the methods, libraries containing more than a million molecules appear to be within our reach.”

In the next stage of this project, researchers will target several intracellular protein-protein interaction targets for which it has been difficult to develop inhibitors based on conventional small molecules. They are confident that orally applicable cyclic peptides can be developed for at least some of them.