Ferreira, Catia and Missailidis, Sotiris
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Mucins are high molecular weight glycoproteins that provide a protective layer on epithelial surfaces and are involved in cell-cell interactions, signalling, and metastasis. The membrane-bound MUC1 mucin is expressed in normal mucosas and the aberrant expression of its under-glycosylated forms has been reported in various carcinomas of the epithelium. MUC1 is a human tumour antigen expressed in breast, pancreatic and ovarian cancers. Agents able to bind tightly and specifically to the surface of malignant cells would greatly benefit cancer diagnosis and treatment, whereas the targeting of cell surface receptors would have significant implications on inflammation and immunity. While antibodies have the ability to specifically recognise some tumour cell markers, their large size and own immunogenicity markedly limit their pharmacological value. The development of nuclease resistant DNA molecules, termed aptamers, has provided a new alternative to antibodies. Using the Systematic Evolution of Ligands by Exponential Enrichment (SELEX) methodology, one can generate vast libraries of oligonucleotide ligands (DNA, RNA, or unnatural products), screened rapidly for specific sequences that have appropriate binding affinities and specificities to the clinically relevant marker.
We have identified synthetic DNA oligonucleotide aptamers that bind to the MUC1 tumour marker with low nanomolar affinity via the 20 tandem repeat sequence of the MUCI. These specific aptamers were selected from an initial library that contained a degenerate region of 25 bases to result in 425 random- sequence DNA molecules. Ten rounds of in vitro selection and amplification were performed, to confer affinity maturation of aptamers for MUCI. Selected aptamers were cloned, sequenced and found to be sharing some unique consensus sequences. The affinity of each aptamer for MUC1 was studied by qualitative and quantitated methods such as ELISA, BIAcore, and EMSA. Affinities on the nanomolar range have been identified and confirmed.
Efforts in our laboratory are now focusing on optimization of their delivery, functionality and structural properties. Fluorescent labelled aptamers have been successfully used in the identification of MUC1 expressing tumour cell lines such as the MCF7 breast tumour cell line and can be used in future diagnostic assays, whereas radiolabelled aptamers can be used clinically to enable imaging and therapy of the tumour marker bearing cancer cells.
|Item Type:||Journal Article|
|Copyright Holders:||Not known|
|Keywords:||aptamers; MUC1; SELEX; radiolabeling; radiopharmaceuticals; tumour imaging; cancer therapy|
|Academic Unit/Department:||Science > Life, Health and Chemical Sciences|
|Interdisciplinary Research Centre:||Biomedical Research Network (BRN)|
|Depositing User:||Sotiris Missailidis|
|Date Deposited:||05 Jul 2006|
|Last Modified:||10 Mar 2014 10:31|
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