A17
Regulation of AKT by DNA-PK mediates platinum resistance in ovarian cancer
Michelle Chen, Hani Gabra, Euan Stronach
Imperial College London, London, UK
Background
Ovarian cancer is characterised by typically advanced stage at diagnosis, frequent relapse following chemotherapy and the development of resistance to platinum-based chemotherapy.
Activation of oncogenic pro-survival factors, such as AKT, are known to play a role in resistance to chemotherapeutic agents, including cisplatin. Previously, we demonstrated that AKT acts as a key modulator of platinum resistance, using matched ovarian cell line pairs. Bozulic et al (2008) showed that DNA-PK phosphorylates AKT at S473 in human endothelial cells following DNA double strand breaks after irradiation and promotes survival.
Here, we showed that DNA-PK phosphorylates AKT at S473 in response to cisplatin and that inhibition of DNA-PK resulted in re-sensitization to platinum-induced apoptosis. To our knowledge this is the first report of a role for DNA-PK in the activation of AKT in cancer cells and in chemoresistance.
Method
Western blot analysis showed protein expression changes following treatment with cisplatin, irradiation or insulin. Immunoprecipitation and immunoflourescence was carried out to investigate interaction and co-localization between DNA-PK and AKT. Effects of DNA-PK inhibition with inhibitor, NU7026, or siRNA were measured for apoptosis with caspase 3/7 activation.
Results
Western blot analysis showed that pre-treatment with DNA-PK inhibitor, NU7026, inhibits P-AKT S473, but not P-AKT T308 in response to DNA damage induced by cisplatin. The interaction between DNA-PK and AKT was confirmed by immunoprecipitation, while immunofluorescence showed co-localization of DNA-PK and P-AKT S473. These platinum-resistant cell lines were re-sensitized to apoptosis when DNA-PK was down-regulated with NU7026 or siRNA.
Conclusion
This is the first report of DNA-PK activating AKT specifically in response to DNA damaging agents, such as cisplatin, but not insulin, in cancer cells. AKT inhibitors have been found to cause hyperglycaemia clinically. This study suggests that specific targeting of DNA damage based activation of AKT offers a mechanistic route to bypass potential clinical side effects of direct AKT inhibitors.
