Dual responsive promoters to target therapeutic gene expression to radiation-resistant hypoxic tumour cells

Chadderton, Naomi, Cowen, Rachel L., Sheppard, Freda C.D, Robinson, Suzanne, Greco, Olga, Scott, Simon D., Stratford, Ian J., Patterson, Adam V. and Williams, Kaye J. (2005) Dual responsive promoters to target therapeutic gene expression to radiation-resistant hypoxic tumour cells. International Journal of Radiation Oncology*Biology*Physics, 62 (1). pp. 213-222. ISSN 0360-3016

Abstract

Purpose: Tumor hypoxia is unequivocally linked to poor radiotherapy outcome. This study aimed to identify enhancer sequences that respond maximally to a combination of radiation and hypoxia for use in genetic radiotherapy approaches.

Methods and materials: The influence of radiation (5 Gy) and hypoxia (1% O2) on reporter-gene expression driven by hypoxia (HRE) and radiation (Egr-1) responsive elements was evaluated in tumor cells grown as monolayers or multicellular spheroids. Hypoxia-inducible factor-1α (HIF-1α) and HIF-2α protein expression was monitored in parallel.

Results: Of the sequences tested, an HRE from the phosphoglycerate kinase-1 gene (PGK-18[5+]) was maximally induced in response to hypoxia plus radiation in all 5 cell lines tested. The additional radiation treatment afforded a significant increase in the induction of PGK-18[5+] compared with hypoxia alone in 3 cell lines. HIF-1α/2α were induced by radiation but combined hypoxia/radiation treatment did not yield a further increase. The dual responsive nature of HREs was maintained when spheroids were irradiated after delivery of HRE constructs in a replication-deficient adenovirus.

Conclusions: Hypoxia-responsive enhancer element sequences are dually responsive to combined radiation and hypoxic treatment. Their use in genetic radiotherapy in vivo could maximize expression in the most radio-resistant population at the time of radiation and also exploit microenvironmental changes after radiotherapy to yield additional switch-on.

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