Lung cancer is a leading cause of cancer death in Australia and the world [1, 2]. There are two types of lung cancers, non small cell (NSCLC) and small cell (SCLC). NSCLC accounts for 75–80% of all lung cancers. Overall, NSCLC has a low five-year survival rate of only 8–14%. Furthermore, approximately 75% of all NSCLC patients present with advanced cancers . The goals to manage this group of patients are no longer curative, but instead, palliative, to prolong the survival time through palliation chemotherapy or best supportive care . The median survival of a patient with advanced or metastasis NSCLC is approximately six to eight months [4, 5]. Clearly, the future of therapy depends on the development of new 'target agents' that explore methods of inhibiting tumor growth, or sensitizing tumors to chemotherapy or radiation to add to current unsatisfactory therapeutic armament.
Gene therapy is one such strategy being considered. Several genes have been explored, including tumour necrosis factor , P53 tumour suppresser gene , Herpes Simplex Virus Type-1 (HSV-1) thymidine kinase (TK), and bacterial cytosine deaminase (CD) gene . These approaches generally tried to deliver the gene(s) to cancer cells, and hoped that the transgene would be translated into a protein to provide a therapeutic effect. However, clinical trials have shown that the therapeutic outcome was severely limited by the poor efficiency of current gene transfer vector systems, inadequate weak promoters to drive transgene expression. Therefore, so far, only three cancer gene therapy protocols had reached phase III trials before being terminated.
Epidermal growth factor receptor (EGFR) is a glycoprotein with a molecular weight of 170,000 to 180,000. It is an intrinsic tyrosine-specific protein kinase, which is stimulated upon epidermal growth factor (EGF) binding. The known downstream effectors of EGFR include PI3-K, RAS-RAF-MAPK P44/P42, and protein kinase C signaling pathways. EGFR signaling involved in cell growth, angiogenesis, DNA repair, and autocrine growth regulation in NSCLC as well as in a wide spectrum of human cancer cells . Thus, it has recently emerged as an innovative target for the development of new cancer therapy, particularly for NSCLC .
Recently, a monoclonal antibody against EGFR called cetuximab has been developed. It has shown excellent clinical effects for the treatment of lung and head and neck cancer in a clinical trial in humans [11, 12]. Other small chemical inhibitors, such as ZD-1839 have also been developed and demonstrated anti-tumor effects in in vitro and in vivo . However, clinical use of ZD-1839 in humans has not been very successful. Although long term evaluation of the drug is still needed, ZD-1839, as a monoclonal antibody drug (in a protein form), and as with any other drug therapies, was disappointing, demonstrating the need for the development of new and effective technologies .
Other novel products based on short DNA and RNA are also currently being developed. These include LY900003 (Affinitac™), OST-774 (Tarceva™), and trastuzumab (Herceptin). LY900003 is an antisense oligonucleotide, known to modify gene expression by interacting with the mRNA involved in the production of disease-specific proteins . However, antisense therapeutics have shortcomings in specificity and consistency.
RNA interference (RNAi) is an evolutionarily conserved process in which recognition of double-stranded RNA (dsRNA) ultimately leads to posttranscriptional suppression of gene expression. This suppression is mediated by short double stranded RNA (dsRNA), also called small interfering RNA (siRNA), which induces specific degradation of mRNA through complementary base pairing. In several model systems, ie.: mostly lower order animals, this natural response has been developed into a powerful tool for the investigation of gene function [16, 17]. More recently it was discovered that introducing synthetic 21-nucleotide dsRNA duplexes into mammalian cells could efficiently silence gene expression. Although the precise mechanism is still unclear, RNAi offers a new way to inactivate genes of interest. When compared with traditional antisense knockout technologies, it provides a potential new approach for modulation of oncogenic gene function in cancer cells . In this study, we investigated the possibility whether RNAi could silence EGFR gene in commonly used NSCLC cancer cell lines, A549 and SPC-A1. We also assessed the degree of EGFR gene silencing and its functional outcome in terms of effects on cell proliferation and growth inhibition in vitro and in vivo. Our results suggest that RNAi-mediated silencing of EGFR may provide an opportunity to develop a new treatment strategy for NSCLC.