Many human diseases are tricky to target, especially those like Alzheimer’s disease or autoinflammatory diseases that involve proteins gone haywire (1). Sometimes, cells degrade proteins too quickly, which can lead to disease. To solve this problem, Entact Bio has built a class of small molecules that leverage a cell’s own protein regulation machinery to protect a specific protein from degradation.
Victoria Richon, chief executive officer of Entact Bio, explained that boosting the levels of certain proteins has its distinct advantages. “Gene therapy is one approach to try and increase that protein, but it’s limited because we only have certain cell tropisms, and we can only go in and fix certain genes,” she said. “This approach is a small molecule approach, so you can penetrate all tissues of the body including the brain. You can target specifically that one protein and do it in a very dose-dependent way.”
Rerouting proteins
Proteins travel throughout the cell to carry out critical functions like providing structural support and catalyzing reactions. One of the cell’s mechanisms of controlling protein localization and expression is through ubiquitination, a post-translational modification where the cell tags proteins for degradation. During ubiquitination, three enzymes participate in adding a ubiquitin tag to be transferred to a protein of interest (2). Originally, scientists thought that proteins tagged with ubiquitin were just degraded through the proteasome. Now, researchers know that ubiquitin can change where the protein is trafficked, essentially acting like “Google Maps” to get the protein where it needs to go.

Ubiquitination is reversible, and the reverse reaction (called deubiquitination) is mediated by deubiquitinating enzymes (DUB). These DUB can cleave the ubiquitin tag from a protein of interest, essentially saving it from degradation or tracking to a faraway location (3). The human genome encodes more than one hundred DUB, and some are highly specific to whatever protein they target (3).
According to Gustavo Silva, a biologist at Duke University who is not associated with Entact Bio, perturbing the ubiquitination/deubiquitination system can prove useful in a variety of diseases. “You can bump up the [ubiquitination] system if proteins are damaged and misfolded to try and establish a healthy cell state faster,” he said. “On the other side, especially in cancer, you can potentially stop specific cancer cells from being able to move out proteins, which lead to toxicity and cancer cell death.”
Richon and the Entact Bio team plan to identify and harness DUB produced by the body and then use small molecules to push the DUB towards the protein of interest. By using this approach, they can rewrite a faulty protein tracking code, enabling protein specific perturbations to fix disease states.
Building molecular matchmakers
To better harness the deubiquitinase pathway, Richon and her team started by building a computational and experimental platform to sort through the available DUB and their target proteins in the human body, then designed a small molecule (aptly named an ENTAC, or enhancement-targeting-chimera) that binds to both the DUB and the protein of interest. Critically, the scientists had to find a ligand that would bind to the DUB without inhibiting its activity.

They added a linker that connected to another ligand that would bind to the target protein. The ENTAC looks like a barbell: A ligand on one side of the linker binds to the DUB, and a ligand on the other side binds to the protein target.
“Most DUB have multiple substrates, so they are not specifically impacting or targeting your protein of interest,” Richon said. “By actually having the affinity of the two ends of your molecule for the DUB and the target protein, you are bringing them together in a way that they may not normally be brought together.”
To test if the ENTAC triggered deubiquitination in the target protein, Richon and her team developed strategies for measuring the levels of ubiquitin tag on the protein after small molecule treatment. While the company has not yet released any data on the deubiquitination capabilities of its ENTAC candidates, Richon noted a few key questions that need to be answered: “Does the drug deubiquitinate the protein? Then, can we show enhancement of protein activity, whether it’s an increase in the abundance of the protein or enhancement of the pathway the protein is affecting?”
They plan to first test the activity of their ENTAC molecules in cell-free models and then move towards cell and in vivo animal models. It’s a long process, but one that Richon hopes will be fruitful. “We often refer to our drug discovery process as an audit trail of how the drug works,” she said.
Drugging the undruggable
One of the exciting promises of ubiquitination/deubiquitination-associated therapeutics is the potential for targeting previously “undruggable” proteins. Certain proteins do not have a well defined structure, making it very difficult to build a traditional drug molecule that can bind tightly to the protein itself. By using DUB, which naturally have an affinity for their protein targets, there is an increased likelihood of reaching the protein to deubiquitinate it.
Silva described this approach as “hijacking the system in a way that you use small molecules that serve as a glue — a molecular glue — which will bring DUB that are already inside your cell to your target protein.” He plans to reprogram DUB to respond more appropriately during disease.
What’s most exciting is that we have the potential to treat a disease that hasn’t been treated before.
- Victoria Richon, Entact Bio
Entact Bio’s technology joins that of several other biotechnology companies targeting the ubiquitin space such as KSQ Therapeutics, which has a candidate that inhibits a DUB implicated in cancer, and Mission Therapeutics, which inhibits a DUB implicated in mitochondrial dysfunction. While all these companies perturb ubiquitination/deubiquitination pathways in different ways, they ultimately want to correct the protein dysregulation found in human disease.
Richon noted that Entact Bio has not yet publicly released which specific diseases they hope to target with their ENTAC molecules. She is excited, though, to potentially apply their technology to several specific areas — cancer among them. “Oncology has oncogenes, which are overactive proteins that drive cancer and also has tumor suppressor genes, which are lost in cancer,” she said. “By increasing the activity of a tumor suppressing pathway, you would be driving death in a cancer cell.”
Richon can also see applications for the team’s technology for neurodegenerative, inflammatory, and metabolic diseases. In the future, the team envisions developing strategies for regulating multiple proteins by using one small molecule, perhaps by targeting a master protein that controls the activity of many downstream proteins. They are also excited about the artificial intelligence and machine learning technologies in their drug discovery platform. “It’s the next generation tool for us, not only in the making of compounds themselves, but in identifying new targets and thinking about pathways differently,” she said.
“What’s most exciting is that we have the potential to treat a disease that hasn’t been treated before,” Richon said. “We need to give doctors more in their medicine chest in order to treat patients for what is actually disease causing for them.”
References
- Popovic, D., Vucic, D. & Dikic, I. Ubiquitination in disease pathogenesis and treatment. Nat Med 20, 1242– 1253 (2014).
- Bedford, L., Lowe, J., Dick, L.R., Mayer, R.J. & Brownell, J.E. Ubiquitin-like protein conjugation and the ubiquitin– proteasome system as drug targets. Nat Rev Drug Discov 10, 29–46 (2011).
- Snyder, N.A. & Silva, G.M. Deubiquitinating enzymes (DUBs): Regulation, homeostasis, and oxidative stress response. J Biol Chem 297, 101077 (2021).