Rational Engineering of Erythropoietin for Targeted Red Blood Cell Production

In tuning the quantitative parameters of protein-protein interactions, evolution often employs combinations of weak physical contacts to achieve specificity. This natural phenomenon can be applied to the engineering of targeted, protein-based drugs with improved cell-type specificity relative to their existing counterparts. Our approach uses mutagenesis to weaken a signaling protein’s affinity for its receptor on non-target cells, combined with its fusion to a cell-specific targeting element via a flexible linker. Targeting should be driven by the targeting element, and despite the mutation, subsequent binding of the mutated protein should occur due to its high local concentration at the cell surface. We have applied this approach to the development of an erythropoietin (EPO)-based therapeutic that targets human glycophorin A (huGYPA) on red blood cell precursors (RBCs). Recombinant EPO has been used for two decades to treat forms of anemia associated with end-stage renal failure, AIDS, chemotherapy, and hemoglobinopathies. Clinical use of EPO has recently decreased due to concerns over the drug’s off-target effects, including tumor recurrence and platelet formation or activation, which may lead to coronary disease or thrombosis. Using a transgenic mouse strain that expresses huGYPA, we have demonstrated that our targeted version of EPO stimulates RBC production without activating platelet formation. We also show that our compound has a similar serum half-life to Aranesp, a long-acting EPO derivative that is used clinically. In patients, targeting EPO to RBCs should allow higher doses to fully restore RBC levels without increasing the risk of cardiovascular events or cancer progression. In addition, the targeted EPO developed here should have an extended serum half-life and reduced immunogenicity relative to existing EPO drugs. More broadly, this work establishes the utility of our approach as a general platform for the rational design of targeted therapeutics.

Grant acknowledgements: NIH NRSA Fellowship, Wyss Institute, DARPA