As far as our digestive system is concerned, peptides are foodstuffs to be broken down, not therapeutics to be kept intact until they enter the bloodstream. Peptide drugs, then, must run a gauntlet of countless degradative enzymes. Few have what it takes to stand the punishment. To give peptide drugs a sturdier backbone, researchers at the Ecole Polytechnique Fédérale de Lausanne (EPFL) have been developing double-bridged peptides, that is, peptides that are cyclized by two chemical bridges.

More rigid and stable than linear peptides, the double-bridged peptides better resist digestive enzymes. Still, the cyclized structures haven’t been rolling toward pill form. The ones that have been evaluated thus far have given into enzymatic pressure. Undeterred, the EPFL researchers decided to explore the double-bridged peptide space more widely—so widely, in fact, that the researchers realized that they needed a new search method.

Led by Christian Heinis, PhD, associate professor, the EPFL team developed a phage display method to screen large libraries of genetically encoded double-bridged peptides. The method, which identifies among billions of double-bridged peptides those that bind a disease target of interest and survive enzymes of the gastrointestinal tract, was described in a paper (“De novo development of proteolytically resistant therapeutic peptides for oral administration”) that appeared May 11 in the journal Nature Biomedical Engineering.

The method involves three steps:

  • First, billions of genetically encoded random peptide sequences are cyclized by two chemical bridges that impose conformational constraints onto the peptides’ backbones so that they are more difficult to attack by enzymes.
  • Second, this library of peptides is exposed to enzymes from cow intestine to eliminate all those peptides that are not stable.
  • Third, the scientists dip target proteins into the pool of surviving peptides to fish out those that bind to the wanted disease target.

Using this method, Heinis and colleagues managed to find two promising peptides. In their article, the scientists wrote, “[We] generated a peptide inhibitor of the coagulation Factor XIa with nanomolar affinity that resisted gastrointestinal proteases in all regions of the gastrointestinal tract of mice after oral administration, enabling more than 30% of the peptide to remain intact, and small quantities of it to reach the blood circulation. We also developed a gastrointestinal-protease-resistant peptide antagonist for the interleukin-23 receptor, which has a role in the pathogenesis of Crohn’s disease and ulcerative colitis.”

Using their phage display method, the researchers succeeded for the first time in evolving target-specific peptides that can resist breakdown in the gastrointestinal tract. For example, they gave mice a lead peptide that inhibits thrombin—an important anti-thrombosis target—in the form of a pill. The peptide remained intact in the stomach and intestines, and even though it reached the bloodstream in rather small quantities, most of it remained fully intact across the entire gastrointestinal tract. This is a key step toward engineering oral peptide drugs.

Peptides are short chains of amino acids that occur in our body, in plants or bacteria to control diverse functions. Several peptides are used as drugs such as insulin, which controls the metabolism of sugar, and cyclosporine, which suppresses organ rejection after transplants. More than 40 peptides are already approved as drugs, generating revenues in the billions. There are several hundreds of peptide-based medications currently in clinical trials. But almost none of these drug-peptides can be taken orally.

Peptide drugs that can be administered in pill form may become more common if target-specific, protease-resistant peptides could be generated and screened in large numbers. Exploring this possibility could become easier with the Heinis group’s new method. “It’s a bit like searching a needle in a haystack, and this method makes this easy,” Heinis explained.

His group is developing oral peptides that act directly on gastrointestinal targets, meaning that they don’t need to travel into the bloodstream. “We are focusing on chronic inflammatory diseases of the gastrointestinal tract like Crohn’s disease and ulcerative colitis as well as bacterial infections,” Heinis noted. “We have already succeeded in generating enzyme-resistant peptides against the interleukin-23 receptor, an important target of these diseases, which affect millions of patients worldwide without any oral drug available.”

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