(270a) Engineering and Analysis of Antibody Fragments for Prion Disease Therapy

Prion diseases are propagated by the conversion of the native form of the prion protein (PrP^C) to an alternatively folded, disease causing form (PrP^Sc) catalyzed by the interaction of PrP^Sc with PrP^C. Due to its centrality in the pathogenesis of prion diseases, several antibodies have been developed for therapy that target PrP^C; however these antibodies have shown limited success in animal models. We have engineered a suite of six scFvs based on the most promising anti-prion antibody therapeutic, ICSM 18.  The antibodies vary in affinity from 1 nM to 320 nM and have expression levels in yeast that cover two log orders, implying differential stability. Characterization of the antibodies in cell culture models of prion is ongoing. Additionally, to gain insight into the molecular level antibody-antigen interactions, we have developed a methodology to provide residue level epitope identification using yeast surface display and next generation DNA sequencing. We have validated this technology by comparing our findings for ICSM18 to the antibody-complex crystal structure, and applied the new method to identify critical residues that form the epitopes of three additional anti-Prion antibodies.  Finally, we are gaining molecular level insight into the protein-protein interactions that mediate prion formation and propagation using the same technology.  Such knowledge may lead to the identification of epitopes for more effect therapeutic antibodies for prion diseases.  Combined, our work enables a quantitative analysis of the effects of affinity, stability, and epitope specificity on therapeutic efficacy and provides insight into the molecular basis of prion pathogenesis.