RNA Interference Against NDRG2 Gene In Cervical Cancer Cell Results In Increased Sencitivity To Cisplatin

Cervical cancer is the second largest cause of cancer mortality in women worldwide with more than 270,000 deaths per year. Current therapies for the treatment of advanced cervical cancer involve the use of cisplatin, often in combination with radiotherapy. Unfortunately, the current cisplatin-based treatment for cervical cancer does not lead to a high disease-free survival rate in patients with bulky or locally-advanced disease.Therefore, it is ugent to clarify the underlying molecular mechanisms that control chemosensitivity of cervical cancer cells and improve therapeutic effect.NDRG2 (also named SYLD/KIAA1248), a novel gene that lower expressing in glioma, was first discovered by our laboratory in 1999 with polymerase chain reaction (PCR)-based subtractive hybridization when looking for the differentially expressed genes between glioma and normal tissue(the GenBank accession No .AF159092), the entire mRNA of NDRG2 is 2024bp in length and it encodes a deduced 357 amino acids unkown protein with a calculated molecular mass of 41 kDa. The NDRG2 gene is located at chromosome 14q11.2, including 16 exons and 15 introns. For the highly homologous in amino acides to N-myc downstream-regulated gene 1(NDRG1), it was named NDRG2. Ndrg2 was strongly expressed in brain, heart and skeletal muscle, while weakly expressed or can not detected in other tissues. NDRG2 has been repeatedly isolated by other laboratories and given various names.Our previous study on the relationship between NDRG2 and radiosensitivity in Hela cells demonstrated that down-regulation of NDRG2 can enhanceγradiation-induced growth arrest and apoptosis,thus sensitizes Hela cells toγradiation ,while ectopic expression of NDRG2 promoted colony-forming efficiency of Hela cells underγradiation.All these data showed that NDRG2 is involved in the regulation of radiosensitivity of cervical cancer Hela cells,and NDRG2 might be involved in the regulation of stress response to DNA damage.As an important part of combined therapy to cervical cancer, chemotherapy is the standard therapies for the treatment of advanced cervical cancer, often in combination with radiotherapy. As one of most effective drug in chemotherapy to cervical cancer, cisplatin (CDDP) is believed to act via the formation of inter- and intrastrand cross-links in DNA, culminating in the initiation of cell death via caspases.If NDRG2 was involved in the regulation of stress response to DNA damage, we can postulate that NDRG2 may play a role in regulation of chemosensitivity to cisplatin. In order to prove above hypothesis, we treated Hela cell line with cisplatin, by semiquantitive RT-PCR and Western Blot, we found that NDRG2 gene could be obviously up-regulated both at mRNA and protein level with the time going .To further explore the effects of NDRG2 on chemosensitivity of cervical cancer cells, NDRG2 was genetically down-regulated in Hela cells by RNAi technique. We observed that the IC50 for cisplatin in Hela- negative control cells was much higher than stable cell lines down-regulating NDRG2.The colony-forming assay showed that suppression of NDRG2 could decrease the colony-forming ability of Hela cells and enhance the inhibitory effects of cisplatin on Hela cells. In addition, flow cytometry showed that inhibition of NDRG2 promoted cisplatin-induced apoptosis of Hela cells. Because this effect could be due to long-term changes in gene expression, we also examined the ic50 in Hela cells transiently transfected with a synthetic siRNA with the same result.In addition, we also found a very interesting phenomena that down-regulation of NDRG2 could inhibit Hela cells growth.Meanwhile, PCNA level significantly decreased in Hela cells expressing ndrg2-specific siRNA.In summary, the hypothesis that NDRG2 may play a role in regulation of chemosensitivity to cisplatin was first proposed and proved by us. And we proved that down-regulation of NDRG2 could enhance the sensitivity to cisplatin in cervical cancer Hela cells. Our previous study on the relationship between NDRG2 and radiosensitivity in Hela cells demonstrated that RNAi against NDRG2 gene enhances radiosensitivity in Hela cells. All these data showed NDRG2 may be a new therapeutic target for cervical cancer. It is a hot spot to learn the true function of a novel gene.As a candidate for tumor suppressor gene, NDRG2 maybe play more various physiological roles in cancer cells which deserves to research. We focused our studies on finding the role of NDRG2 from the aspect of chemosensitivity, which would enrich the understanding of this gene's function.