{"id":{"repo_id":"bradford","oai_identifier":"oai:bradscholars.brad.ac.uk:10454/4420"},"canonical_url":"https://search.dev.ndltd.org/etd/bradford/oai:bradscholars.brad.ac.uk:10454/4420","repository":{"repo_id":"bradford","name":"University of Bradford","base_url":"https://bradscholars.brad.ac.uk/oai/request"},"display":{"title":"Synthesis of bespoke matrices to investigate a novel anti-tumour molecular target using affinity chromatography. The design, synthesis and evaluation of biotinylated biarylheterocycles used as novel affinity probes in the identification of anti-tumour molecular targets.","abstract":"Three novel, synthetic biarylheterocycles bearing imidazole terminal groups had previously been discovered with high cytotoxicity (IC50 16¿640 nM) against a number of human tumour cell lines. Notably, this biological activity was independent of duplex DNA binding affinity. The compounds were tested in the NCI 60-cell line panel and COMPARE analysis suggests they have a novel mechanism of action, targeting the product of a ¿gene-like sequence¿ of unidentified function. The identity of likely protein targets was explored using a chemical proteomic strategy. Bespoke affinity matrices for chromatography were prepared in which test compounds were attached to a solid support through a biotin tag. A synthetic route to hit compounds containing a biotin moiety in place of one of the imidazole sidechains was developed. Chemosensitivity studies confirmed that the biotinylated compounds retained their activity showing IC50 = 6.25 ¿M in a susceptible cell line, compared with > 100 ¿M for an insensitive cell line. The biotinylated ligands were complexed to a streptavidin-activated affinity column and exposed to cell lysates from the susceptible cell lines. Bound proteins were eluted from the column and separated using SDS-PAGE. Proteins were characterised by MALDI MS and MS/MS and identified using Mascot database searches. Heterogeneous nuclear ribonuclear protein A2/B1 was found to selectively bind to the affinity probes.","abstract_html":"Three novel, synthetic biarylheterocycles bearing imidazole terminal groups had previously been discovered with high cytotoxicity (IC50 16¿640 nM) against a number of human tumour cell lines. Notably, this biological activity was independent of duplex DNA binding affinity. The compounds were tested in the NCI 60-cell line panel and COMPARE analysis suggests they have a novel mechanism of action, targeting the product of a ¿gene-like sequence¿ of unidentified function. The identity of likely protein targets was explored using a chemical proteomic strategy. Bespoke affinity matrices for chromatography were prepared in which test compounds were attached to a solid support through a biotin tag. A synthetic route to hit compounds containing a biotin moiety in place of one of the imidazole sidechains was developed. Chemosensitivity studies confirmed that the biotinylated compounds retained their activity showing IC50 = 6.25 ¿M in a susceptible cell line, compared with > 100 ¿M for an insensitive cell line. The biotinylated ligands were complexed to a streptavidin-activated affinity column and exposed to cell lysates from the susceptible cell lines. Bound proteins were eluted from the column and separated using SDS-PAGE. Proteins were characterised by MALDI MS and MS/MS and identified using Mascot database searches. Heterogeneous nuclear ribonuclear protein A2/B1 was found to selectively bind to the affinity probes.","abstract_has_math":false,"creators":["Evans, Hayley R."],"institution":"University of Bradford","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":[],"advisors":["Wheelhouse, Richard T.","Sutton, Chris W.","Phillips, Roger M."],"committee_chairs":[],"committee_members":[],"year":2010,"date_issued":"2010-08-27T15:53:57Z","date_published":"2010-08-27T15:53:57Z","updated_at":"2026-07-24T01:14:32Z","subjects":["Anti-tumour molecular target","Affinity chromatography","Biotinylated biarylheterocycles","Affinity probes","Cytotoxicity","Human tumour cells","Chemosensitivity studies"],"languages":["en"],"rights":["\"Creative
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The University of Bradford theses are licenced under a Creative Commons Licence."],"dc:subject":["Anti-tumour molecular target","Affinity chromatography","Biotinylated biarylheterocycles","Affinity probes","Cytotoxicity","Human tumour cells","Chemosensitivity studies"],"dc:title":["Synthesis of bespoke matrices to investigate a novel anti-tumour molecular target using affinity chromatography. The design, synthesis and evaluation of biotinylated biarylheterocycles used as novel affinity probes in the identification of anti-tumour molecular targets."],"dc:type":["Thesis"],"dc:type.qualificationlevel":["doctoral"],"dc:type.qualificationname":["PhD"]},"updated_at":"2026-07-24T01:14:32Z"}