Biotechnology Letters

, Volume 30, Issue 11, pp 1913–1921 | Cite as

Implication of functional activity for determining therapeutic efficacy of suicide genes in vitro

  • P. Gopinath
  • Siddhartha Sankar GhoshEmail author
Original Research Paper


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.


Curcumin Cytosine deaminase Fluorocytosine Suicide gene therapy Tumor cells Uracil phosphoribosyltransferase 



This work was financially supported by the Department of Biotechnology [BT/PR9988/NNT/28/76/2007] and Council of Scientific and Industrial Research [No.37 (1248)/06/EMRII] Government of India.


  1. Bernt KM, Steinwaerder DS, Ni S et al (2002) Enzyme-activated prodrug therapy enhances tumor-specific replication of adenovirus vectors. Cancer Res 62:6089–6098PubMedGoogle Scholar
  2. Chu RL, Post DE, Khuri FR et al (2004) Use of replicating oncolytic adenoviruses in combination therapy for cancer. Clin Cancer Res 10:5299–5312PubMedCrossRefGoogle Scholar
  3. Ghosh SS, Gopinath P, Ramesh A (2006) Adenoviral vectors: a promising tool for gene therapy. Appl Biochem Biotechnol 133:9–29PubMedCrossRefGoogle Scholar
  4. Gopinath P, Ghosh SS (2007) Monitoring green fluorescent protein for functional delivery of E. coli cytosine deaminase suicide gene and the effect of curcumin in vitro. Gene Ther Mol Biol 11:219–228Google Scholar
  5. Gopinath P, Ghosh SS (2008) Apoptotic induction with bifunctional E. coli cytosine deaminase-uracil phosphoribosyltransferase mediated suicide gene therapy is synergized by curcumin treatment in vitro. Mol Biotechnol (DOI  10.1007/s12033-007-9026-3)
  6. Harris JD, Gutierrez AA, Hurst HC et al (1994) Gene therapy for cancer using tumour-specific prodrug activation. Gene Ther 1:170–175PubMedGoogle Scholar
  7. Hermiston T (2000) Gene-delivery from replication-selective viruses: arming guided missiles in the war against cancer. J Clin Invest 105:1169–1172PubMedCrossRefGoogle Scholar
  8. Holder JW, Elmore E, Barrett JC (1993) Gap junction function and cancer. Cancer Res 53:3475–3485PubMedGoogle Scholar
  9. Hotz-Wagenblatt A, Shalloway D (1993) Gap junctional communication and neoplastic transformation. Crit Rev Oncog 4:541–558PubMedGoogle Scholar
  10. Howells LM, Mitra A, Manson MM (2007) Comparison of oxaliplatin- and curcumin-mediated antiproliferative effects in colorectal cell lines. Int J Cancer 121:2929–2937CrossRefGoogle Scholar
  11. Kambara H, Tamiya T, Ono Y et al (2002) Combined radiation and gene therapy for brain tumors with adenovirus-mediated transfer of cytosine deaminase and uracil phosphoribosyltransferase genes. Cancer Gene Ther 9:840–845PubMedCrossRefGoogle Scholar
  12. Kanai F, Kawakami T, Hamada H et al (1998) Adenovirus-mediated transduction of Escherichia coli uracil phosphoribosyltransferase gene sensitizes cancer cells to low concentrations of 5-fluorouracil. Cancer Res 58:1946–1951PubMedGoogle Scholar
  13. Kawamura K, Tasaki K, Hamada H et al (2000) Expression of Escherichia coli uracil phosphoribosyltransferase gene in murine colon carcinoma cells augments the antitumoral effect of 5-fluorouracil and induces protective immunity. Cancer Gene Ther 7:637–643PubMedCrossRefGoogle Scholar
  14. Khatri A, Zhang B, Doherty E et al (2006) Combination of cytosine deaminase with uracil phosphoribosyl transferase leads to local and distant bystander effects against RM1 prostate cancer in mice. J Gene Med 8:1086–1096PubMedCrossRefGoogle Scholar
  15. Marais R, Spooner RA, Light Y et al (1996) Gene-directed enzyme prodrug therapy with a mustard prodrug/carboxypeptidase G2 combination. Cancer Res 56:4735–4742PubMedGoogle Scholar
  16. Miyagi T, Koshida K, Hori O et al (2003) Gene therapy for prostate cancer using the cytosine deaminase/uracil phosphoribosyltransferase suicide system. J Gene Med 5:30–37PubMedCrossRefGoogle Scholar
  17. Moolten FL (1986) Tumor chemosensitivity conferred by inserted herpes thymidine kinase genes: paradigm for a prospective cancer control strategy. Cancer Res 46:5276–5281PubMedGoogle Scholar
  18. Moolten FL (1994) Drug sensitivity (“suicide”) genes for selective cancer chemotherapy. Cancer Gene Ther 1:279–287PubMedGoogle Scholar
  19. Mullen CA, Kilstrup M, Blaese RM (1992) Transfer of the bacterial gene for cytosine deaminase to mammalian cells confers lethal sensitivity to 5-fluorocytosine: a negative selection system. Proc Natl Acad Sci USA 89:33–37PubMedCrossRefGoogle Scholar
  20. Pabbruwe MB, Stewart K, Chaudhuri JB (2005) A comparison of colorimetric and DNA quantification assays for the assessment of meniscal fibrochondrocyte proliferation in microcarrier culture. Biotechnol Lett 27:1451–1455PubMedCrossRefGoogle Scholar
  21. Pan J, Xu G, Yeung SJ (2001) Cytochrome C release is upstream to activation of Caspase-9, Caspase-8, and Caspase-3 in the enhanced apoptosis of anaplastic thyroid cancer cells induced by manumycin and paclitaxel. J Clin Endocr Metab 86:4731–4740PubMedCrossRefGoogle Scholar
  22. Pillai GR, Srivastava AS, Hassanein TI et al (2004) Induction of apoptosis in human lung cancer cells by curcumin. Cancer Lett 208:163–170CrossRefGoogle Scholar
  23. Rowley S, Lindauer M, Gebert JF et al (1996) Cytosine deaminase gene as a potential tool for the genetic therapy of colorectal cancer. J Surg Oncol 61:42–48PubMedCrossRefGoogle Scholar
  24. Shpitz B, Giladi N, Sagiv E et al (2006) Celecoxib and curcumin additively inhibit the growth of colorectal cancer in a rat model. Digestion 74:140–144PubMedCrossRefGoogle Scholar
  25. Springer CJ, Duvaz IN (2000) Prodrug-activating systems in suicide gene therapy. J Clin Invest 105:1161–1167PubMedCrossRefGoogle Scholar
  26. Tiraby M, Cazaux C, Baron M et al (1998) Concomitant expression of E. coli cytosine deaminase and uracil phosphoribosyltransferase improves the cytotoxicity of 5-fluorocytosine. FEMS Microbiol Lett 167:41–49PubMedCrossRefGoogle Scholar
  27. Xia K, Liang D, Tang A et al (2004) A novel fusion suicide gene yeast CDglyTK plays a role in radio-gene therapy of nasopharyngeal carcinoma. Cancer Gene Ther 11:790–796PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2008

Authors and Affiliations

  1. 1.Department of BiotechnologyIndian Institute of Technology GuwahatiGuwahatiIndia
  2. 2.Centre for NanotechnologyIndian Institute of Technology GuwahatiGuwahatiIndia

Personalised recommendations