Poster Presentation 26th Lorne Cancer Conference 2014

The interaction of cisplatin and related novel drug analogues with DNA containing telomeric repeat sequences  (#176)

Ben Johnson 1 , Janice Aldrich-Wright 1 , Vincent Murray 2 , Nial Wheate 3 , Mark Temple 1
  1. School of Science and Health , University of Western Sydney, Campbelltown, NSW, Australia
  2. School of Biotechnology and Biomolecular Sciences , University of New South Wales , Sydney , NSW, Australia
  3. Faculty of Pharmacy , University of Sydney , Sydney , NSW, Australia

Cisplatin (cis-diamminedichloroplatinum(II)) is a platinum-based drug that has been recognized as an effective anti-cancer drug following investigations by Barnett Rosenberg in 1965. Its pharmacology is determined by the drugs ability to form covalent bonds with the DNA at purine bases. This forms kinks in the DNA, known as DNA adducts. The DNA adducts inhibit DNA replication within the cell, which leads to cell death and subsequent tumour regression. Although these DNA adducts induce cancer cell death, there are associated cytotoxicity and drug resistance issues that still need to be resolved. Therefore, a range of novel drug analogues are in development with the aim to circumvent these issues.

In this study, the sequence specificity of DNA adducts induced by cisplatin and related drug analogues have been characterized in a DNA sequence containing seven human telomeric repeats of GGGTTA. Previous studies have shown this sequence to be a major target for cisplatin and it is anticipated that this DNA sequence is a good candidate sequence to test other novel covalent binding drugs.    

In this investigation, we have analysed the DNA-drug interactions at the level of base-pair resolution, through the application of a polymerase stop assay – the Linear Amplification Reaction. This reaction involves the annealing of a fluorescently-labelled primer and the extension of single stranded DNA until the DNA polymerase is inhibited by a DNA adduct. Hence, the sequence specificity of DNA adducts can be determined from sites of polymerase inhibition.  

The results demonstrate that cisplatin, as a positive control, preferentially damages the DNA at guanine bases in each telomeric repeat sequence, which is consistent with previous cisplatin studies. The sequence specificity of two groups of novel drug analogues were also analysed –  monoadduct-forming complexes and diadduct-forming complexes. It was found that compounds from both groups preferentially formed DNA adducts at the guanines in the telomeric repeats, with the diadduct-forming complexes producing the most intense damage at these sites.