Single chain variable fragments: Novel applications as selection tools and therapeutic agents in drug discovery

dc.contributor.advisorHall, J.C.
dc.contributor.authorWeisser, Nina Elizabeth of Environmental Biologyen_US of Guelphen_US of Philosophyen_US
dc.description.abstractSingle chain variable fragment (scFv) antibodies (Abs) were used in two novel applications to broaden their utility for use in drug discovery. In the first application, scFvs were used to improve the selection of peptides that mimic carbohydrate antigens ('i.e.' peptide "mimotopes") for vaccination. A bottleneck in peptide mimotope discovery includes the inability of initial screening regimes to differentiate between mimotopes and non-mimotopes, thus resulting in inefficient 'in vivo' analysis. We demonstrated a rapid screening method to identify putative mimotopes using a scFv phage-display library. The use of this screening method may increase the probability of identifying peptides that will elicit a carbohydrate cross-reactive response ' in vivo'. A human naïve scFv library was screened against both an established peptide mimotope and a non-mimotope of the Group B ' Streptococcus' (GBS) type III polysaccharide to determine if selected antibodies cross-reacted with the original GBS polysaccharide. We were able to differentiate between these two peptides because peptide-binding Abs that cross-reacted to GBS were isolated only with the peptide mimotope. In the second application, we describe a strategy that we hypothesized would increase the therapeutic utility of scFvs by increasing their 'in vivo' half-life and giving them the ability to enable Fc[gamma]R-mediated effector functions towards the target antigen specified by the scFv. This increase in therapeutic utility was achieved by the non-covalent binding between epitope-tagged scFvs (e.g. 6xHis-tagged scFv) and an anti-epitope tag IgG (e.g. anti-Penta-His) that resulted in the formation of a bivalent scFv-IgG complex. To test our hypothesis we used two different murine anti-epitope tag IgG1 Abs (' i.e.' anti-c-Myc and anti-Penta-His) in combination with an epitope-tagged ('i.e.' c-Myc and 6xHis) murine anti-'Salmonella enterica ' serovar Typhimurium ('S'. Typhimurium) scFv, to examine both 'in vivo' persistence in mice and FcR-mediated complement recruitment and phagocytosis of 'S'. Typhimurium by the murine macrophage (M[Phi])-like cell line J774. Compared to the monovalent scFv controls we demonstrated that bivalent scFv-IgG complexes can bind C1q complement, increase bacterial phagocytosis in J774 cells, and increase scFv ' in vivo' [beta]-phase half-life by ca. 3-fold in a mouse model. The potential of both of these applications to increase the therapeutic utility of scFvs antibodies are discussed.en_US
dc.publisherUniversity of Guelphen_US
dc.rights.licenseAll items in the Atrium are protected by copyright with all rights reserved unless otherwise indicated.
dc.subjectsingle chain variable fragmenten_US
dc.subjectdrug discoveryen_US
dc.subjectselection toolsen_US
dc.subjectcarbohydrate antigensen_US
dc.titleSingle chain variable fragments: Novel applications as selection tools and therapeutic agents in drug discoveryen_US


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