Wisconsin Alumni Research Foundation

Therapeutics & Vaccines
Therapeutics Vaccines
Transferrin-Based Lysosome Targeting Degraders
WARF: P210432US02

Inventors: Weiping Tang, Yaxian Zhou

The Invention
UW researchers have developed a novel means of degrading extracellular proteins as a potential cancer treatment. The researchers recognized that transferrin is a protein ligand that carries iron into cells through binding to a transferrin receptor membrane protein. Their strategy involves attaching a small molecule or protein/peptide binder of the protein of interest to transferrin creating a transferrin-ligand shuttle. Transferrin acts as a shuttle to bring the protein of interest into the cell. Once inside the cell, transferrin enters an endosome where it deposits the protein of interest payload analogous to how it deposits iron in the endosome. Transferrin leaves the endosome and encounters its receptor. The transferrin receptor transports the transferrin-ligand shuttle back outside the cell where it can bind another protein and shuttle it inside the cell. The released target protein will be degraded in the lysosome. The inventors are utilizing a natural endocytotic cycle present in all cells which allows for recycling of the transferrin leading to a catalytic protein degradation process. Cancer cells overexpress the transferrin receptor, so this method should specifically target cancer cells. The inventors have tested two of the three general strategies they believe will work. The first covalently links a ligand that binds a protein of interest to the transferrin protein. In the test case, they used biotin to target an anti-biotin antibody. They incubated the transferrin-biotin and anti-biotin antibody with various cells for various time periods. Using Western blot analysis to detect the proteins inside the cell and outside the cell, they showed that the antibody was internalized in a transferrin-dependent manner. They also showed that over time the transferrin molecule shuttled back and forth inside and outside the cell demonstrating the catalytic nature of this method. The second method involved transfecting cells with a plasmid expressing transferrin linked to a binding protein capable of binding a protein of interest (Flag tag and an anti-Flag antibody as the proof of concept). They used a fluorescently labeled anti-Flag antibody and measured the fluorescence in the cell lysate over time. They confirmed the presence of the proteins inside and outside the cells over time using a Western blot analysis showing that the antibody entered the cell in a transferrin dependent manner. The third method has not been tested but would use a transferrin receptor antibody in place of the transferrin as a shuttle. The inventors will need to show the antibody can be recycled back out of the cell to preserve the catalytic nature of the transferrin-based methods.
Additional Information
For More Information About the Inventors
For current licensing status, please contact Rafael Diaz at [javascript protected email address] or 608-960-9847