Targeting ligand-independent CSF3R dimerization in chronic neutrophilic leukemia

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The modification of proteins with carbohydrates, called glycosylation, is commonly dysregulated in cancer. One example is chronic neutrophilic leukemia (CNL), a rare disease characterized by the uncontrolled growth of neutrophils. Over 80% of published CNL cases result from a mutation in the colony-stimulating factor 3 receptor (CSF3R). This mutation results in decreased O-linked glycosylation as well as increased ligand-independent dimerization. However, the relationship between these two findings is not known and there are no therapies that target the mutated epitope. The central hypothesis of this project is the loss of glycosylation decreases steric hindrance for CSF3R dimerization and reveals a cancer-specific epitope for targeted therapy. In Aim 1, mass spectrometry will identify the glycans that are present on the wild-type protein but missing in the mutated variant. Additionally, crystal structures of mutant CSF3R will reveal how the loss of glycosylation remodels the receptor interface and promotes ligand-independent dimerization. In Aim 2, screens of yeast-displayed proteins will identify candidates which bind to mutated, but not wild-type CSF3R. Biologics specific to the mutated protein will be validated for labeling cells and blocking dimerization in vitro, and selectively eliminating cancer cells in a mouse model of CNL. Combined, this work will demonstrate how understanding the structural effects of glycosylation facilitates drug discovery. This work is co-sponsored at Stanford University by Drs. Jennifer Cochran and Carolyn Bertozzi, leaders in protein engineering and glycobiology, respectively. This project is also supported through a collaboration with Dr. Julia Maxson, who first discovered the CSF3R mutation. This doctoral work will provide essential training for a research career, bridging the gap between molecular mechanisms of cancer and drug development.