Implication of functional activity for determining therapeutic efficacy of suicide genes in vitro.
Gene-Directed Enzyme Prodrug Therapy (GDEPT), commonly known as suicide gene therapy, provides a selective approach to eradicate tumor cells that is currently considered as an alternate approach to conventional therapy for cancers due to its high efficacy. Herein, we have demonstrated functional activity of the cytosine deaminase (CD) and the hybrid cytosine deaminase-uracil phosphoribosyltransferase (CD-UPRT) suicide genes in transfected cell lines. We have monitored retention profiles of various metabolites that were formed during enzymatic conversion of the prodrug 5-flurocytosine (5-FC) using reverse phase HPLC method. Therapeutic effect of suicide genes was established by cell viability and toxicity assay, whereas apoptotic induction was confirmed by DNA fragmentation ELISA. Our results demonstrated that 5-FC/CD-UPRT-mediated apoptotic cell death was more than 5-FC/CD, which could be further potentiated with anticancer compound curcumin. Such results corroborated 5-FC/CD-UPRT in combination with curcumin as a better chemosensitization method.
Gopinath
P
P
Department of Biotechnology, Indian Institute of Technology Guwahati, Guwahati, 781039, India.
Ghosh
Siddhartha Sankar
SS
D000818:Animals; D017209:Apoptosis / Q000187:drug effects; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects; D002851:Chromatography, High Pressure Liquid; D003474:Curcumin / Q000494:pharmacology; D043525:Cytosine Deaminase / Q000235:genetics / Q000378:metabolism*; D053938:DNA Fragmentation / Q000187:drug effects; D004357:Drug Synergism; D018274:Electroporation; D004797:Enzyme-Linked Immunosorbent Assay; D005437:Flucytosine / Q000378:metabolism* / Q000494:pharmacology; D005472:Fluorouracil / Q000378:metabolism; D005786:Gene Expression Regulation / Q000187:drug effects; D041722:Genes, Transgenic, Suicide / Q000235:genetics*; D005822:Genetic Vectors / Q000235:genetics; D019073:HT29 Cells; D006801:Humans; D010430:Pentosyltransferases / Q000235:genetics / Q000378:metabolism*; D016133:Polymerase Chain Reaction; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection / Q000379:methods
D005437:Flucytosine; D010430:Pentosyltransferases; C023269:uracil phosphoribosyltransferase; D043525:Cytosine Deaminase; D003474:Curcumin; D005472:Fluorouracil