High-Level, β-Catenin/TCF-Dependent Transgene Expression in Secondary Colorectal Cancer Tissue

Abstract

There is an urgent need for improved therapies for inoperable metastatic colon cancer. Gene-directed enzyme prodrug therapy (GDEPT) using adenovirus vectors works well in preclinical models of this disease, but successful clinical application is hampered by an inability to construct vectors that express at high levels in infected tumor cells but not in infected normal cells. Constitutive activation of beta-catenin-dependent gene expression is almost certainly a key causative event in the genesis of colon and some other cancers. Here we have exploited this oncogenic defect to design a synthetic promoter, CTP1, that, in contrast to currently available tumor-selective promoters, is both highly active in cancer cells and highly cancer-cell-specific. CTP1 directs high-level beta-galactosidase expression in freshly isolated biopsies of secondary colon cancer, but is not detectably active in associated normal liver tissue. We also demonstrate that CTP1 can direct high-level, tumor-specific therapeutic gene expression in vivo. Intratumoral injection of an adenovirus vector encoding Escherichia coli nitroreductase driven by CTP1 efficiently sensitized SW480 xenografts to the prodrug CB1954, whereas systemic vector and prodrug administration produced no apparent signs of toxicity. CTP1 may form the basis for effective, targeted gene therapy of metastatic colon cancer and other tumors with deregulated beta-catenin/T cell factor.

5-Fluorocytosine increases the toxicity of Wnt-targeting replicating adenoviruses that express cytosine deaminase as a late gene

“…43 Yet another option for colon cancer is to express nitroreductase, which converts CB1954 to a bifunctional alkylating agent. 44 It remains to be seen whether yCD expression from our Tcf viruses will increase their efficacy in tumour xenograft models. 5-FC treatment alone of xenografts injected with a replicating E1B-deficient adenovirus expressing a bacterial CD gene fused to HSV tk (Ad5-CD/TKrep) was unable to eradicate tumours.…”

Adenoviruses with Tcf binding sites in multiple early promoters show enhanced selectivity for tumour cells with constitutive activation of the wnt signalling pathway

“…Other groups have noted similar differences in wnt activity in colon cancer cell lines. 18,19 All of the cell lines tested (except Co115, where the defect is not known) and virtually all colon tumours possess APC or ␤-catenin mutations resulting in activation of the wnt pathway. This indicates that wnt activation is a critical step in colon cancer tumorigenesis.…”

“…[14][15][16][17] Other groups have used Tcf promoters to regulate the expression of suicide genes. 18,19 We placed Tcf binding sites in the adenovirus E2 promoter, which regulates expression of the viral replication genes, because mutations elsewhere in the virus or cell cannot bypass the absolute requirement for E2 gene products in viral replication. In order to achieve tight regulation of E2 transcription, the adjacent E3 enhancer was mutated.…”

Escherichia coli nitroreductase plus CB1954 enhances the effect of radiotherapy in vitro and in vivo

“…The potential for a bystander effect associated with such virally directed enzyme prodrug therapy (VDEPT) strategies offers the prospect of also sensitizing adjacent untransduced malignant (and normal) cells to irradiation. 3 In these studies, we seek to evaluate the ability of replication-defective adenovirus (E1 and E3 deleted) containing the E. coli NTR gene under the control of either a constitutively active viral cytomegalovirus or a tissue-specific (CTP1) 10 promoter to enhance the cytotoxic effect of ionizing radiation against colorectal cancer cells both in vitro and in vivo.…”