Novel Technique Designs Mini-Proteins That May Lead to New Types of Therapeutics

Researchers say they have developed a technique to generate many small, different proteins that can be designed to bind to therapeutic targets. They add that their methodology, which reportedly produces thousands of new drug candidates, may lead to protection against infectious diseases, such as influenza, and antidotes to nerve toxins.

The computer-designed proteins, which did not previously exist in nature, combine the stability and bioavailability of small-molecule drugs with the specificity and potency of larger biologics, according to David Baker, Ph.D., who led the multi-institutional research project and is professor of biochemistry at the University of Washington School of Medicine and director of the UW Institute for Protein Design.

“These mini-protein binders have the potential of becoming a new class of drugs that bridge the gap between small-molecule drugs and biologics. Like monoclonal antibodies, they can be designed to bind to targets with high selectivity, but they are more stable and easier to produce and to administer,” said Dr. Baker who with colleagues published their study in Nature (“Massively Parallel De Novo Protein Design for Targeted Therapeutics”).

The technique relies on the Rosetta computer platform, developed by Dr. Baker and colleagues at the University of Washington. They designed thousands of short proteins, about 40 amino acids in length that the Rosetta program predicted would bind tightly to the molecular target.

Because of their small size, these short proteins tend to be extremely stable. They can be stored without refrigeration, noted Dr. Baker, who added that they also are more easily administered than large protein drugs, such as monoclonal antibodies.

In the past short, protein-binder drugs were usually re-engineered versions of naturally occurring proteins. But studies showed they didn’t work much better than monoclonal antibodies. Because these mini-proteins binders are original designs, they can be tailored to fit their targets much more tightly and are simpler to modify and refine, continued Dr. Baker.

In this study, the team designed two sets of proteins: one to prevent the influenza virus from invading cells and another that would bind to and neutralize a deadly nerve toxin from botulism, a potential bioweapon.

Computer modeling identified the amino acid sequences of thousands of short proteins that would fit into and bind to the influenza and botulinum targets. The team created short pieces of DNA that coded each of these proteins, grew the proteins in yeast cells, and then looked at how tightly they bound to their targets. The targets were Influenza H1 hemagglutinin and botulinum neurotoxin B.

The technique allowed them to design and test 22,660 proteins in just a few months, with more than than 2000 of them bound to their targets with high affinity, according to Dr. Baker. He pointed out that of the best candidates found the anti-influenza proteins neutralized viruses in cell culture and other designed proteins prevented the botulinum toxin from entering brain cells.

A nasal spray containing one of the custom-designed proteins completely protected mice from the flu if administered before or as much as 72 hours after exposure. The protection that the treatment provides equaled or surpassed that seen with antibodies, the researchers report.

Testing of a subset of the proteins showed that they were extremely stable and, unlike antibodies, did not become inactivated by high temperatures. The small proteins also triggered little or no immune response, a problem that often renders larger protein drugs ineffective, said Dr. Baker.