Date of Award

Winter 2005

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Program/Concentration

Biomedical Sciences

Committee Director

Richard R. Drake

Committee Member

Richard Britten

Committee Member

Richard Ciavarra

Committee Member

John Semmes

Abstract

Mutations in the adenomatous polyposis coli gene are frequently associated with progression of colon carcinoma and most other types of epithelial carcinomas. This usually results in stabilization of β-catenin protein levels, followed by transactivation of Tcf-4/β-catenin responsive genes. The effectiveness of a Tcf-4/β-catenin transcriptional enhancer element in combination with a c-fos or carcinoembryonic antigen promoter was tested for its ability to act as a tumor specific regulator of gene expression in a panel of human tumor and normal cell lines. Luciferase reporter assays indicated enhanced activity of the Tcf-4/β-catenin transcriptional element only in tumor cell lines, with minimal activities in normal colon cell lines. The Tcf-4/β-catenin enhancer and c-fos promoter linked with the herpes virus thymidine kinase suicide gene in combination with ganciclovir was further evaluated in the normal and tumor cell lines. The Tcf-4/β-catenin elements conferred tumor specific expression of HSV thymidine kinase (HSV-TK) gene, resulting in selective metabolism of ganciclovir and cell killing of only tumor cell lines. There was no detectable expression of HSV thymidine kinase expression in normal colon cell lines. Additionally, recombinant adenoviral constructs were made to deliver the gene expression cassette, containing the HSV-TK expressed from a Tcf-4/c-fos enhancer/promoter combination, to the tumor cells. No expression was detected in the normal colon cells as opposed to significant levels of HSV-TK gene expression observed in tumor cells. Furthermore, various chemical and genetic modulators were also screened in an effort to identify newer methods to regulate the activity of the proposed recombinant enhancer/promoter combinations. These data suggest that Tcf-4/β-catenin enhancer can be effectively coupled with a suitable tumor-specific or tumor-related mammalian promoter for selective expression of therapeutic genes for gene therapy of epithelial cancers. This approach also offers a potential promoter cassette approach linked with the Tcf-4/β-catenin enhancer to better individualize treatment to cancer patients.

Comments

Dissertation submitted to the Faculty of Eastern Virginia Medical School and Old Dominion University in Partial Fulfillment of the Requirement for the Degree of Doctor of Philosophy in Biomedical Sciences.

DOI

10.25777/jt8j-5m12

ISBN

9780542580048

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