P-04 Short-interfering RNA for targeting IGFs in tumour cells

Insulin-like growth factor I (IGF-I) and its receptor (IGF-IR) play essential role in several cellular processes, and their deregulation leads to tumor progression. Since few years, it has been shown that antisense RNA targeted against IGF-I produced from a episomal plasmid stably transfected in cells inhibited in vivo tumor progression and also induced antitumoral immune response. In order to clarify the role of IGF-I and IGF-IR in tumorigenesis, an alternative strategy to antisense RNA using small interfering RNA was developed. These small molecules will be more easily delivered to target cells and avoid the utilisation of EBV-based episomal plasmids. Our study addresses the effect of targeting IGF-IR and the IGF-I mRNA with RNA interference on mouse breast cancer cells and hepatoma cancer cells. In order to quantify rapidly the effect of our designed siRNA on their target site, we used plasmid constructs, which contains renilla luciferase gene fused to eitheir IGF-I or IGF-IR coding sequences. We have cotransfected our plasmidic construct with IGF-IR cDNA in breast cancer cells and IGF-I construct in hepatocarcinoma cells. An efficient inhibition of the luciferase signal was observed in both cases, allowing us to identify the most inhibitory siRNA sequence. We then decided to investigate these siRNAs directly on the endogenous transcripts. Using real time RT-PCR, our preliminary studies have shown high inhibition of IGF-IR mRNA in mouse breast cancer cells. Concerning hepatoma cancer cells, we demonstrated for the first time an efficient inhibition of IGF-I mRNA with RNA interference approach. These results encourage us to investigate whether IGF-IR and IGF-I targeting by siRNA strategies are able to produce an antitumoral immune response in syngenic animal models. We will also present preliminary results obtained with deoxyribozyme approach to inhibit IGF-I and IGF-IR expression in cancer cells. This data could permit us to better understand the interaction between the immune and endocrine system.