Use this URL to cite or link to this record in EThOS: http://ethos.bl.uk/OrderDetails.do?uin=uk.bl.ethos.555108
Title: The development and characterisation of peptides to Image αvβ6 in cancer
Author: Saha, Antonio
Awarding Body: Queen Mary University of London
Current Institution: Queen Mary, University of London
Date of Award: 2011
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Abstract:
Introduction: The epithelial-specific integrin αvβ6 usually is undetectable on normal, adult, human tissue but is upregulated during tissue re-modelling and carcinogenesis. Objective: To develop and characterise agents for targeting αvβ6 in cancer. Methods: A series of peptides was generated from known high-affinity αvβ6 ligands. These peptides were then assessed for their ability to inhibit αvβ6-dependent cell adhesion. The lead 20 mer peptide, A20FMDV2, was identified. Flow cytometry, with two genetically identical cell lines that differed only in their expression of αvβ6 – A375P puro(negative) and A375Pβ6(positive)-, confirmed specificity for binding of A20FMDV2 to v6. The A20FMDV2 sequence was synthesised with both a biotin and DTPA-chelator group added. This was radiolabelled with Indium-111 and purity and structural integrity analysed by HPLC. Prior to in vivo experiments the endogenous expression of αvβ6 in nu/nu athymic mice was determined by examining 20 different organs (in triplicate) by immunohistochemistry. For in vivo experiments, athymic mice were inoculated subcutaneously with A375Pβ6 (right shoulder) and A375P puro (left shoulder). A radiolabelled variant of A20FMDV2 (111In-DTPA-A20FMDV2) was injected into xenografted mice. Some mice were imaged on a NanoSPECT/CT imager and biodistribution confirmed by determining the radioactivity associated with excised organs. Results: Immunohistochemistry of mouse tissues revealed, as expected, strong expression of v6 in the A375Pβ6 tumour but none in the A375Ppuro tumour. In addition, expression of vβ6 was found in normal murine hair follicles, gallbladder, urinary bladder, secretory endometrium, the stomach and along the GI tract. Flow cytometry with A375Ppuro and A375Pβ6 cells confirmed that DTPA-chelate addition to A20FMDV2 did not affect its specificity or affinity for v6. HPLC confirmed that DTPA-biotinylated-A20FMDV2 was labelled successfully with Indium-111 resulting in a single radiolabelled peak. Ex-vivo gamma counts revealed seven-fold higher uptake of the radiolabelled-peptide in the A375Pβ6 tumour compared with the A375Ppuro tumour. Biodistribution data were consistent with the endogenous αvβ6 expression as determined by immunohistochemistry. NanoSPECT/CT imaging clearly identified that the A375P6 tumour retained more radioactive peptide than the A375Ppuro tumour at 30, 60 and 120 minutes after injection. Images also showed uptake in the GI tract. Conclusion: We have determined that the peptide DTPA-A20FMDV2 is specific for v6, can be labelled with Indium-111 and selectively localises in vivo to v6-positive tissues, including human cancer. This is the first peptide described for the successful imaging, by SPECT, of v6-positive cancers.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID: uk.bl.ethos.555108  DOI: Not available
Keywords: Medicine ; Cancer
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