Radiofluorinated Antisense Oligodeoxynucleotides for in vivo Molecular Imaging of Cellular Response to Radiotherapy

INTRODUCTION: In tumours expressing the wild-type p53 gene activation of the p53-dependent DNA damage response pathway is a critical determinant of the cellular response to ionizing radiation. The p21Waf1/Cip1 gene transcriptionally trans-activated by p53 post-irradiation has been associated with resistance to radiation therapy and its presence or absence in tumours may be related to treatment response. Non-invasive monitoring of changes in the levels of this transcript in situ by positron emission tomography would potentially provide a major tool for evaluating tumour and normal tissue responses to radiation therapy during treatment. AIM: The aim of this study will be to explore the potential of antisense oligodeoxyoligonucleotides (as-ODNs) directed against p21Waf1/Cip1 mRNA as PET-imaging agents for in vivo molecular imaging. METHODS: Radiofluorination of the as-ODNs will be performed by radiolabeling of a trifluoromethanesulfonyl precursor with 18F and conjugation of the radiolabeled precursor to the ODNs. Analysis and subsequent purification will be performed using thin layer chromatography and high performance liquid chromatography. The structure and purity of non-radioactive fluorinated ODNs will be determined by nuclear magnetic resonance and electron impact mass spectrometry. Cell uptake studies of 18F-labeled ODNs and radiofluorinated ODNs labelled with a fluorescent dye will be performed with tumor cells that are known either to express (parental HCT116 cells) or not to express (p21-deficient HCT116/p21 -/- cells) p21Waf1/Cip1 mRNA in vitro following induction with ionizing radiation. As a control, also the accumulation of a radiolabeled random ODN sequence in these two cell lines will be examined. DISSEMINATION: Developing this approach will also set the table for the design, synthesis and implementation of other as-ODN PET probes for any mRNA species of interest that might provide insight into the processes of carcinogenesis and treatment response. KEY WORDS: radiation therapy, gene activation, antisense oligodeoxynucleotides, molecular imaging, positron emission tomography, cellular response.