Tumour-specific therapeutic adenovirus vectors: repression of transgene expression in healthy cells by endogenous p53

Ks, Lipinski; Djeha, A.Hakim; Krausz, Eberhard; Lane, David P.; Pf, Searle; Mountain, Andrew; Cj, Wrighton

doi:10.1038/sj.gt.3301403

Cited by 14 publications

(16 citation statements)

References 23 publications

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“…The ptkGC 3 p53luc-bsd HeLa p53 reporter cells (6,23) were plated in triplicate into 96-well plates (f3.5 Â 10 3 cells/well). The number of living cells in each well after treatment was estimated colorimetrically with the WST-1 assay (Roche, Mannheim, Germany), and the luciferase activity was determined with the luciferase assay reagent Bright-Glo (Promega, Madison, WI).…”

Section: Methodsmentioning

confidence: 99%

Activation of p53 in Cervical Cancer Cells by Human Papillomavirus E6 RNA Interference Is Transient, but Can Be Sustained by Inhibiting Endogenous Nuclear Export–Dependent p53 Antagonists

Koivusalo

Mialon²,

Pitkänen³

et al. 2006

Cancer Research

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Abstractp53 is degraded in cervical cancer cells by the human papillomavirus E6 and can be stabilized with short interfering RNA (siRNA) molecules targeting E6 mRNA. In this in vitro study, we show that E6 siRNA-induced p53 activation is transient in HeLa cervical cancer cells despite continuous suppression of E6 mRNA; activation can be sustained if the endogenous p53 antagonists COP1, MDM2, Pirh2, and c-Jun-NH 2 -kinase are also targeted by siRNAs or by inhibiting the nuclear export of p53 with leptomycin B. The direct targeting of any one of these four cellular p53 antagonists had no effect on p53 activity when E6 was intact, but inhibited the fading off of E6 siRNA-induced p53 activation in nonstress conditions. The effect was additive when multiple cellular antagonists were concomitantly inhibited, indicating that all these proteins degrade p53 when E6 is inactivated. The antiproliferative effect induced by E6 silencing was enhanced when the endogenous p53 antagonists were additionally targeted. In conclusion, if human papillomavirus E6 is inhibited under nonstress conditions, the subsequent p53 activation is quickly reversed by the endogenous p53 degenerative machinery. The present results indicate that several cellular p53 antagonists must be inhibited for sustained p53 activity if E6 siRNA therapy is attempted and if no combined genotoxic therapy is applied. (Cancer Res 2006; 66(24): 11817-24)

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Section: Methodsmentioning

confidence: 99%

Activation of p53 in Cervical Cancer Cells by Human Papillomavirus E6 RNA Interference Is Transient, but Can Be Sustained by Inhibiting Endogenous Nuclear Export–Dependent p53 Antagonists

Koivusalo

Mialon²,

Pitkänen³

et al. 2006

Cancer Research

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“…The cells were cultured in Dulbecco's modified Eagle's medium (Euroclone, Wetherby, UK) containing 10% FBS, 2 mM l-glutamine, 1x non essential amino acids (Euroclone) and 50 µg/ml gentamycin (Calbiochem, San Diego, CA). The SiHa p53 reporter cell line, carrying the p53 reporter plasmid ptkGC 3 p53-luc, 20 has been described previously. 19 The SiHa DDp53 and SiHa E6 cell lines carry plasmids that express a truncated dominant negative p53 and the HPV16 E6 protein, respectively, under the control of the CMV promoter.…”

Section: Methodsmentioning

confidence: 99%

The cytotoxicity of chemotherapy drugs varies in cervical cancer cells depending on the p53 status

Koivusalo¹,

Hietanen²

2004

Cancer Biology & Therapy

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“…Deleting the E1B-55K adenoviral gene permits the selective replication of these viruses in cells lacking functional p53 [Bischoff et al, 1996]. A p53-dependent Ad expression cassette was similarly effective [Lipinski et al, 2001]. This latter consisted of two components; a lac repressor (lacI) inducible by wild-type p53, and a therapeutic transgene under control of the heat shock 70 (hsp70) promoter incorporating LacI binding sites.…”

Section: Promoters For Cancer Gene Therapymentioning

confidence: 99%

Promoters and Control Elements: Designing Expression Cassettes for Gene Therapy

Papadakis

Nicklin²,

Baker³

et al. 2004

CGT

145

121

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It has become apparent that the clinical success anticipated in the field of gene therapy has been limited by progress in several of the fundamental areas of genetics, molecular and cellular biology relevant to its application. Whilst a great deal of effort has been made in the evaluation of transgenes, it is only more recently with the advance of vector systems that attention has begun to be focused upon the means and control of transgene expression. Until recently, the majority of constructs have employed ubiquitous viral promoters to drive expression from simple gene expression cassettes using viral promoters and lacking introns, 3' untranslated regions (UTRs), locus control regions (LCR's), matrix attachment regions (MAR's) and other such genetic components. It has consequently emerged that these elements may have a key role in determining the levels and longevity of gene expression attainable in vivo, irrespective of the vector system utilised. The majority of gene therapy applications would also benefit from the specific optimisation of 'tailormade' expression cassettes to optimise their therapeutic efficacy. In conjunction with modification of vector tropism and strategies to limit their immunogenicity, this should create vectors suitable for the clinical application of gene therapy. This review aims to highlight some of the principle considerations of gene expression in vivo, and the means by which it may most effectively be achieved, whether this is via the minimal modification of an existing eukaryotic promoter or by the more extensive design of a novel promoter and associated elements.

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Tumour-specific therapeutic adenovirus vectors: repression of transgene expression in healthy cells by endogenous p53

Cited by 14 publications

References 23 publications

Activation of p53 in Cervical Cancer Cells by Human Papillomavirus E6 RNA Interference Is Transient, but Can Be Sustained by Inhibiting Endogenous Nuclear Export–Dependent p53 Antagonists

Activation of p53 in Cervical Cancer Cells by Human Papillomavirus E6 RNA Interference Is Transient, but Can Be Sustained by Inhibiting Endogenous Nuclear Export–Dependent p53 Antagonists

The cytotoxicity of chemotherapy drugs varies in cervical cancer cells depending on the p53 status

Promoters and Control Elements: Designing Expression Cassettes for Gene Therapy

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