The Effects Of Lentivirus-mediated ShRNA Interference Targeting Mcl-1on Growth Of Lymphoma Cell Line

Lymphoma is the most common hematologic malignancy and comprises a heterogeneous group of malignancies consisting of different clinic pathologic entities. NK/T-cell lymphoma is a distinct subtype of lymphoma relatively common in Asia and South America. Histological features of this disease are angiocentricity, angiodestruction, and massive necrosis and apoptosis. Although most cases initially present with localized disease, NK/T-cell lymphoma is highly aggressive and resistant to chemotherapy with a poor prognosis. Moreover, many traditional treatments usually lead to a lot of toxicities which could affect the therapeutic outcome and quality of life. Thus, seeking a new strategy for the treatment of NK/T-cell lymphoma is needed.In the last few years a number of promising new therapies including gene therapy have been developed. RNA interference (RNAi) by viral vector is a new form of gene therapy. There are many advantages about viral vectors for RNAi compared to non-viral vectors, for example, high transfection efficiency, broad host range and long-term expression of therapeutic gene. ShRNA expression viral vectors such as retroviral vectors, adenoviral vectors and, more recently, lentiviral vectors had been reported and proven to be safe and effective for humans with no obvious side effects in many solid tumors. It had been reported that lymphoma cells had a low infection rate with most viral vectors. Thus finding an appropriate viral vector for lymphoma cells is the first step in gene therapy. Our previous researches had demonstrated that lentivirus could be an ideal vector for transducing heterologous gene into lymphoma cells.Mcl-1is a pro-survival member of Bcl-2family which could promote oncogenesis by inhibition of apoptosi. Many kinds of cancer cells have been shown to be reliant on Mcl-1for their survival and development. Substantial evidence showed that Mcl-1overexpression promoted oncogenesis recently, it had been reported in several haematological cancers and solid tumours, including leukemia and hepatocellular carcinomas. Forced overexpression of Mcl-1in transgenic mice led to a significantly high incidence of B-cell lymphoma, while forced Mcl-1down-regulation was able to induce apoptosis in a number of cancer cell types. Overexpression of Mcl-1also could lead to chemoresistance of certain cancers. Thus Mcl-1is an attractive and potential therapeutic target in a number of malignancies, and is the focus of a number of studies.OBJECTIVE:To explore the effects of lentivirus-mediated RNA interference (RNAi) targeting the Myeloid leukemia-1(Mcl-1) on proliferation and apoptosis of lymphoma cell line SNK-6.METHODS:Short hairpin RNA (shRNA) targeting Mcl-1was designed and the lentiviral vector carrying the shRNA was conducted. After infecttion of lentiviral vector in SNK-6cells, Real time PCR and western blot were used to analyze the expression of Mcl-1mRNA and protein. MTT assay and flow cytometry were employed to detect the status of proliferation and apoptosis.RESULTS:The shRNA targeting Mcl-1was successfully inserted into the lentiviral vector, which can effectively infect the lymphoma cell line SNK-6. After infecting the SNK-6cells, the expression of Mcl-1mRNA and protein was obviously decreased and MTT assay suggested that the proliferation of SNK-6cells was inhabited by RNAi targeting Mcl-1against control-shRNA group(31.6%±3.3%vs5.8%±2.7%, P〈001). Flow cytometry showed that the apoptosis rate of Mcl-1-shRNA-infected cells was significantly higher than the control group(28.9%±2.1%vs5.3%±1.5%, P<0.001).CONCLUSION:1、 The shRNA targeting Mcl-1was successfully inserted into the lentiviral vector, which can effectively infect the lymphoma cell line SNK-6.2、 Lentivirus-mediated shRNA targeting the Mcl-1can specifically block the Mcl-1gene expression, inhabit proliferation, and promote apoptosis of SNK-6cells.