Use of the radiation‐inducible WAF1 promoter to drive iNOS gene therapy as a novel anti‐cancer treatment

Worthington, Jenny; McCarthy, Helen; Barrett, E.; Adams, Catherine; Hirst, David

doi:10.1002/jgm.567

Cited by 50 publications

(64 citation statements)

References 29 publications

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“…16 We have previously used the WAF1 promoter to drive expression of iNOS, resulting in modification of vascular tone in rat arteries ex vivo, and to achieve radiosensitization of tumour cells in vitro and in vivo. [17][18][19] The WAF1 promoter also restricted iNOS transgene expression to the radiation field. 19 Here, we further characterize the WAF1 promoter in a range of normal and human tumour cell lines of differing p53 status using a WAF1-driven green fluorescent protein (GFP) reporter construct.…”

Section: Introductionmentioning

confidence: 99%

p21(WAF1)-mediated transcriptional targeting of inducible nitric oxide synthase gene therapy sensitizes tumours to fractionated radiotherapy

et al. 2006

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Section: Introductionmentioning

confidence: 99%

p21(WAF1)-mediated transcriptional targeting of inducible nitric oxide synthase gene therapy sensitizes tumours to fractionated radiotherapy

et al. 2006

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“…50 The WAF-1 promoter has to date been the only inducible system reported activated in response to both radiation and hypoxia. 37 Acute hypoxic exposure (2 h) reportedly induced a 5.4 and 4.3-fold increased in reporter gene expression within 24 hours in RIF-1 and HT29 cells in vitro. The effect of combined radiation and hypoxic exposure were however not evaluated.…”

Section: Combination With Hypoxiamentioning

confidence: 91%

“…Indeed, when the E9 promoter was used to drive the iNOS gene, basal levels of iNOS increased progressively up to 8 h in transfected human microvascular endothelial cells (HMEC-1) and resulted in inhibition of the vasoconstrictor phenylephrine by 83% 36 and a seven-day growth delay in transfected murine sarcoma (RIF1) in vivo. 37 Other promoters. The X-rays responsive elements of the GADD45a have been reported to be induced by radiation doses as low as 0.5 Gy with expression levels reaching a maximum 0.5 to 1 h after exposure.…”

Section: Radio-enhancibility Of the Promotersmentioning

confidence: 99%

Radiation to control transgene expression in tumors

Marignol¹,

Coffey²,

Hollywood³

et al. 2007

Cancer Biology & Therapy

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Significant evidence has accumulated indicating that certain genes are induced by ionising radiation. An implication of this observation is that their promoter regions include radiation-responsive sequences. These sequences have been isolated in the promoter of several genes including Erg-1, p 21/WAF-1 , GADD45a and t-PA. The mechanism by which radiation induces gene expression remains unclear but involves putative binding sites for selected transcription factors and/or p53. Consensus CC(A/T) 6 GG sequences have been localized in the Erg-1 promoter and are referred to as serum response elements or CArG elements. The tandem combination of CArG elements has been shown to improve gene expression levels, with a 9-copy motif conferring maximum inducibility. The response of these genes to ionising radiation appears to follow a sigmoid relationship with time and dose. Therapeutic induction of suicide genes and significant cytotoxicity can be achieved at clinically relevant x-rays doses both in vitro and in vivo but was found to be cell-type dependent. Radiation-inducible gene therapy can be potentially enhanced by exploiting hypoxia through the inclusion of hypoxia-response element motifs in the expression cassette, the use of the anaerobic bacteria or the use of neutron irradiation. These results are encouraging and provide significant evidence that gene therapy targeted to the radiation field is a reasonably attractive therapeutic option and could help overcome hypoxic radioresistant tumors.

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“…However, toxicity to normal tissue or differential sensitization among malignant versus normal cells are the issues that have not yet been addressed. On the other hand, the precision of radiation treatment provides a potential opportunity to utilize tumorspecific and inducible promoter systems with therapeutic genes that kill tumor cells when expressed (3,4).…”

Section: Introductionmentioning

confidence: 99%

Radiation-inducible silencing of uPA and uPAR in vitro and in vivo in meningioma

Rao¹

2010

Int J Oncol

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Abstract. Stereospecific radiation treatment offers a distinct opportunity for temporal and spatial regulation of gene expression at tumor sites by means of inducible promoters. To this end, a plasmid, pCArG-U2, was constructed by incorporating nine CArG elements (in tandem) of EGR1 gene upstream to uPA and uPAR siRNA oligonucleotides in a pCi-neo vector. Radiation-induced siRNA expression was detected in a meningioma cell line (IOMM-Lee). Immunoblotting and RT-PCR analyses confirmed downregulation of uPA and uPAR. A similar effect was observed in transfected cells followed by H 2 O 2 treatment. Moreover, pre-treatment of transfected cells with N-acetyl L-cysteine blocked the silencing of uPA and uPAR, which further confirmed the oxidative damage-mediated downregulation. Cell proliferation assays and Western blot analysis for apoptotic molecules confirmed cell death in a radiation-inducible fashion. Migration and Matrigel invasion assays also revealed a marked decrease in migration and invasion. Immunocytochemistry showed a marked decrease in uPA and uPAR levels in transfected and irradiated cells. H&E staining revealed a decrease in the preestablished tumor volume among the animals treated with pCArG-U2 and radiation. Immunohistochemistry of the brain sections established with intracranial tumors also revealed a marked decrease in uPA and uPAR in a radiation-inducible fashion. Taken together, our data suggest pCArG-U2 as a suitable candidate for radiation-inducible gene therapy.

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Use of the radiation‐inducible WAF1 promoter to drive iNOS gene therapy as a novel anti‐cancer treatment

Abstract: iNOS gene therapy in combination with an inducible promoter results in significant tumour cell radiosensitisation. The WAF1 promoter may be a good candidate for a gene therapy as it is silent in normal tissue yet can be induced by the tumour environment.

Cited by 50 publications

References 29 publications

p21(WAF1)-mediated transcriptional targeting of inducible nitric oxide synthase gene therapy sensitizes tumours to fractionated radiotherapy

p21(WAF1)-mediated transcriptional targeting of inducible nitric oxide synthase gene therapy sensitizes tumours to fractionated radiotherapy

Radiation to control transgene expression in tumors

Radiation-inducible silencing of uPA and uPAR in vitro and in vivo in meningioma

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