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Miniproteins: The next big thing in medicine?

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Autoimmune diseases are caused by our own immune system reacting when it is not supposed to. The results can range from mild to life threatening, but are usually a huge burden to patients, as long-term treatment is hard. The main way to diminish the symptoms is by inactivating the immune system, or part of it, to stop it from reacting against its own body. One of the best and most common ways to do so nowadays, is using antibodies.

Antibodies are proteins produced by immune system cells (specifically B cells), and they bind other proteins. In autoimmune diseases, antibodies can be used as treatment, being produced to bind the parts of the immune system that are deregulated, stopping them from working. But there are some problems with their use, such as requiring regular injections, side effects, and a high economic cost.

Miniproteins as a possible antibody replacement

Researchers have now developed miniproteins. They are just what the name indicates, very tiny proteins, made of up to 60 amino acids, while antibodies are usually over 1000. Doctors and researchers are hoping that these miniproteins can be used for autoimmune disease treatment in the same way as antibodies. That can be done if they are blocking the immune system proteins responsible for the disease.

The researchers chose two proteins that are linked with autoinflammatory diseases. Interleukin 23 (IL-23), which, if inhibited, helps treat psoriasis among others, and interleukin 17 (IL-17), linked with inflammatory bowel disease.

Why miniproteins?

These small proteins may have many advantages over antibodies. Firstly, they can be engineered to have certain properties, such as good absorption in the gut, which could mean that instead of injections, a pill could be taken. Miniproteins are also more stable, cheaper to produce, and can be engineered easily using computational models. This last property means that if they work, it would be easy to target other problematic proteins in autoimmune diseases, cancer, etc.

Engineering miniproteins using computational biology

The hard part was obviously engineering these miniproteins for the job of inhibiting the IL-17 and IL-23 responses. To do this, they took data from IL-23 binding to the IL-23 receptor. They also used computer analysis to model how a miniprotein could bind effectively to the IL-23 receptor. Using a combination of known binding spots and computer-generated likely binding spots, they created miniproteins to bind effectively the IL-23 receptor. Similarly, they created miniproteins to bind IL-17

Analysis of the final miniprotein binding the IL23 receptor. The real structure of the crystallized protein (purple and blue) overlaps closely to its computational model predicted structure (gray). This shows the accuracy of the prediction, and suggests computational modeling is a powerful design tool for future miniproteins. The boxed region indicates the binding region of the miniprotein. Image source: Stephanie Berger et al (2024)

After generating many relevant miniproteins, they analyzed them to find the best ones at binding the IL-23 receptor and IL-17. Researchers also analyzed miniprotein stability in the gut, as well as their heat resistance. Then, they worked to change the miniproteins in subtle ways to improve these parameters. They managed to do so, after many small changes in each one, and the results in cells were impressive.

Treating colitis in mice with miniproteins

To test the efficacy of the miniproteins in an actual disease model, mice with colitis were treated with the IL-23 inhibiting miniproteins. Oral administration worked fine, but the free (unbound) miniproteins were broken down fast within the mice. However, only free miniproteins could be broken down, and due to their high affinity, the ones that bound their targets inhibited the IL-23 response for long periods of time. This meant that mice could be treated with daily oral doses of the miniproteins to reduce the symptoms of the disease. The miniproteins had a higher efficacy than antibodies used in humans.

Next steps

The power of miniproteins comes from how easily they can be engineered and tailored for each specific disease. If they work as well in humans as they do in mice, soon we could have many new treatments that could be not only cheaper, but also better, more available and easier to administer and take than antibody-based treatments. Many people with chronic conditions and autoimmune/autoinflammatory diseases could have an easier time and life, dealing with their disease. Of course, clinical trials and further testing will be needed, but so far, it seems a new avenue has been opened for the development of new treatments.

However, miniproteins can help with more than autoimmune diseases. In fact, miniproteins have been engineered to target other harmful toxins, for example from bacteria, as highlighted by recent research, which shows that developed miniproteins can neutralize toxins from Clostridioides difficile. During the recent COVID pandemic, efforts were made to develop miniproteins against the SARS-CoV-2 virus itself, and miniproteins could perhaps be very useful in the case of another viral pandemic. Of course, clinical trials and further testing will be needed, but so far, it seems a new avenue has been opened for the development of new treatments for many types of diseases and infections.

Link to the original post: Berger, S., Seeger, F., Yu, T. Y., Aydin, M., Yang, H., Rosenblum, D., … & Baker, D. (2024). Preclinical proof of principle for orally delivered Th17 antagonist miniproteins. Cell, 187(16), 4305-4317. DOI: 10.1016/j.cell.2024.05.052

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