Abstract

Antibody–based approaches have provided vast improvements in the detection and treatment of many cancers. However, progress is currently limited in prostate cancer due to a lack of clinically-relevant biomarkers. Novel biomarker discovery may be achieved with antibody technology, which utilises cancer cells to pan antibody libraries/sources, to identify antibodies against non-predetermined antigens. The major limitation currently encountered is the difficulty in establishing the identities of such antigens, due to the need for extensive post-selection antibody characterisation. The work embodied in this thesis utilised protein array technology to monitor the generation of a large recombinant antibody library directed towards nonpredetermined antigens present in a prostate cancer cell line. Protein arrays were used to profile the immune response, mounted by the avian host following cancer cell immunisation. Investigations of the antigen profile revealed a trend in the immune response towards orthologs that were conserved between human and chicken. Similar protein array-profiling was utilised to characterise the resulting avian scFv-phage library generated, as well as 384 scFv isolated following enrichment of the library towards xxviii prostate cancer cells. Evaluation of identified scFv-antigens allowed for the selection of an antigen subset to be screened against the scFv library. This screening resulted in the isolation of the first recombinant antibody to Zinc Finger Protein 358, an antigen currently uncharacterised in prostate cancer. This antibody was isolated and characterised. In conclusion, the incorporation of protein array technology into recombinant antibodybased biomarker discovery enabled antigen characterisation at an earlier stage and on a larger scale than possible with traditional methods, thus circumventing the bottleneck associated with the current strategies in biomarker discovery using recombinant antibody technology. The high-throughput approach developed in this study sheds light on our understanding of immunisation processes and substantially enhances the potential of recombinant antibody technology for cancer biomarker discovery and antibody development.