Pharmacological and genetic modulation of Wnt-targeted Cre-Lox-mediated gene expression in colorectal cancer cells.
Wnt-targeted gene therapy has been proposed as a treatment for human colorectal cancer (CRC). The Cre-Lox system consists of methodology for enhancing targeted expression from tissue-specific or cancer-specific promoters. We analyzed the efficiency of Wnt-specific promoters as drivers of the Cre-mediated activity of a luciferase reporter gene or cell death effector gene in CRC cell lines in the presence and absence of two modulators of Wnt activity, sodium butyrate and lithium chloride. Butyrate is present in the colonic lumen after digestion of fiber-rich foods, whereas the colonic lumen is readily accessible to lithium chloride. In both SW620 and HCT-116 CRC cells, a physiologically relevant concentration of butyrate upregulated reporter and effector activity and altered the Wnt-specific expression pattern. Lithium chloride markedly enhanced Cre-Lox-mediated Wnt-specific reporter expression only in APC wild-type CRC cells. Possibilities for genetic modulation of the proposed CRC therapy included Wnt-specific expression of a floxed Lef1-VP16 fusion that enhanced Wnt-specific cell death and of a floxed dominant-negative Tcf4 that specifically downregulated endogenous Wnt activity. These findings demonstrated that the Cre-Lox system, in combination with pharmacological and genetic modulators, represents effective methodology for enhancing Wnt-targeted gene therapy.
Bordonaro
Michael
M
Department of Pharmacology and Developmental Therapeutics Section, Cancer Center, Yale University School of Medicine, New Haven, CT 06520, USA. michaelbordonaro@yahoo.com
Lazarova
Darina L
DL
Sartorelli
Alan C
AC
D017209:Apoptosis; D002087:Butyrates / Q000494:pharmacology; D045744:Cell Line, Tumor; D015179:Colorectal Neoplasms / Q000378:metabolism / Q000473:pathology / Q000628:therapy*; D003587:Cytomegalovirus / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics; D004167:Diphtheria Toxin / Q000235:genetics / Q000378:metabolism; D015536:Down-Regulation; D005786:Gene Expression Regulation; D017930:Genes, Reporter; D015316:Genetic Therapy / Q000379:methods*; D005822:Genetic Vectors; D049452:Green Fluorescent Proteins; D017405:Herpes Simplex Virus Protein Vmw65 / Q000235:genetics; D006801:Humans; D019426:Integrases / Q000235:genetics / Q000378:metabolism*; D058610:Isobutyrates; D018021:Lithium Chloride / Q000494:pharmacology; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008164:Luminescent Proteins / Q000032:analysis; D051821:Lymphoid Enhancer-Binding Factor 1; D010446:Peptide Fragments / Q000235:genetics / Q000378:metabolism; D011401:Promoter Regions, Genetic; D011518:Proto-Oncogene Proteins / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D014157:Transcription Factors / Q000235:genetics; D014764:Viral Proteins / Q000235:genetics / Q000378:metabolism*; D051153:Wnt Proteins; D029961:Zebrafish Proteins
D002087:Butyrates; D004268:DNA-Binding Proteins; D004167:Diphtheria Toxin; D017405:Herpes Simplex Virus Protein Vmw65; D058610:Isobutyrates; C497573:LEF1 protein, human; D008164:Luminescent Proteins; D051821:Lymphoid Enhancer-Binding Factor 1; D010446:Peptide Fragments; D011518:Proto-Oncogene Proteins; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D014764:Viral Proteins; D051153:Wnt Proteins; D029961:Zebrafish Proteins; C029795:diphtheria toxin fragment A; D049452:Green Fluorescent Proteins; C020380:isobutyric acid; D008156:Luciferases; C045073:Cre recombinase; D019426:Integrases; D018021:Lithium Chloride