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Silencing invariant chains of dendritic cells enhances anti-tumor immunity using small-interfering rna

Chinese Medical Journal 2010年第22期123卷 3193-3199页

作者： KE Shan CHEN Xue-hua ZHU Zheng-gang LI Jian-fang YU Bei-qin GU Qin-long LIU Bing-ya Department of Hepatobiliary Surgery West Campus Bejing Chaoyang Hospital Capital Medical University Beijing 100043 China Department of Surgery Shanghai Institute of Digestive Surgery Ruijin Hospital School of Medicine Shanghai Jiao Tong University Shanghai 200025 China

Background Genetic modification of dendritic cells （DCs） has been used as an effective approach to enhance anti-tumor immunity. rna interference （rnai）, which can cause the degradation of any rna in a sequence-specific manner, is a post-transcriptional gene silencing mechanism. In this study, small-interfering rna （sirna） specific for the li gene was transfected into DCs, and the anti-tumor immunity of li-silenced DCs was assessed. Methods The silencing effect of sirna was evaluated by Western blotting and real-time PCR analyses. In vitro cytotoxic activity of T cells was evaluated using a Cytotox 96 non-radioactive cytotoxicity assay kit. The time to tumor onset and the tumor volumes were used as reliable indices to assess the anti-tumor immunity in vivo. To further examine the mechanisms underlying the anti-tumor immunity, flow cytometry analysis was used. Results The li expression of DCs was significantly reduced after li sirna transfection. Significant in vitro anti-tumor ability was exhibited when DCs were co-transfected with li sirna plus endogenous tumor antigen （P 〈0.05）. Furthermore tumor growth was greatly inhibited when mice were immunized with DCs transfected with li sirna plus tumor antigen prior to or subsequent to tumor implantation. Flow cytometry analysis in vitro and in vivo indicated that both CD4^= and CD8^＋ T cells were significantly activated in the li sirna group （P 〈0.05）. Conclusion Silencing of the li gene of DCs may offer a potential approach to enhance DC-based anti-tumor immunity.

关键词： small-interfering rna invariant chain dendritic cells tumor immunity

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Down-regulation amyloid β-protein 42 production by interfering with transcript of presenilin 1 gene with sirna

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Acta Pharmacologica Sinica 2004年第12期25卷 47-52页

作者： Huan-min LUO2, Hui DENG, Fei XIAO, Qin GAO, Wen WENG, Pei-fen ZHANG, Xiao-guang LI Neuropharmacological Research Laboratory, College ofPharmacy, Ji-nan University, Guangzhou 510632, China Neuropharmacological Research Laboratory College of Pharmacy Ji-nan University Guangzhou 510632 Chin

AIM: To investigate the pathogenesis of Aβ42yielding and new drug targets as well as the possibility of rna interference (rnai) technique for treatment of Alzheimer disease (AD). METHODS: Human AD presenilin 1 (PS1) cDNA sequence was obtained from NCBI website. The three sites of rnai action and one missense control site were selected in PS1 cDNA through online design of Ambion company. To confirm specificity of these sites, we conducted a BLAST search of the IMAGE EST library. The corresponding double-stranded DNA was used to construct pSilencer 3.1-H1 plasmid, which could transcribe small interference rna (sirna). Then, the pSliencer 3.1-H1 plasmids were transfected into CHO/PS1/APP cells with SuperFect transfection reagent. The cells have been transfected with the mutant PS1 and APP gene of AD. All the CHO/PS1/APP cells with pSliencer 3.1-H1 plasmids were screened out using G418. Transcripts of PS1 gene in CHO/PS1/APP were measured by RT-PCR, the contents of PS1 peptide and Aβ42 production inside CHO/PS1/APP cells were examined through Western blot and the Aβ42 change of secretion by CHO/PS1/APP was determined with ELISA. RESULTS: The site 3 of PS1 mrna was inhibited by rnai after 2 d. The effect was more obvious with the time. The peptide corresponding to PS1 gene and Aβ42 production in CHO/PS1/APP cells were both reduced after sirna interfere for 3 d. Aβ42 secretion by CHO/PS1/APP cells began to reduce on d 3, and reached the most significance on d 5. There was a time-dependent relationship between the transcript of PS1 gene and the production of Aβ42with rnai action. CONCLUSION: PS1 is essential for γ-secretase activity. Inhibition of the PS1 can decrease the levels of Aβ42. Some sites of PS1 mrna, for example, the site 3, may serve as a new drug target and rnai probably can be used for treatment of AD.

关键词： amyloid beta-protein amyloid beta-protein precursor secretase presenilin-1 rna interference small-interfering rna Alzheimer disease

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Attenuation of Telomerase Activity by sirna Targeted Telomerase rna Leads to Apoptosis and Inhibition of Proliferation in Human Renal Carcinoma Cells

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Chinese Journal of Clinical Oncology 2006年第5期3卷 326-331页

作者： Rumin Wen Junjie Liu Wang Li Wenfa Yang Lijun Mao Junnian Zheng Laboratory of Urology Affiliated Hospital of Xuzhou Medical College Xuzhou 221002 China.

OBJECTIVE Telomerase is an attractive molecular target for cancer therapy because the activation of telomerase is one of the key steps in cell immortalization and carcinogenesis. rna interference using small-interfering rna （sirna） has been demonstrated to be an effective method for inhibiting the expression of a given gene in human cells. The aim of the present study was to investigate whether inhibition of telomerase activity by sirna targeted against human telomerase rna （hTR） can inhibit proliferation and induce apoptotic cell death in human renal carcinoma cells （HRCCs）.METHODS The sirna duplexes for hTR were synthesized and 786-0 HRCCs were transfected with different concentrations of hTR-sirna. The influence on the hTR mrna level, telomerase activity, as well as the effect on cell proliferation and apoptosis was examined.RESULTS Anti-hTR sirna treatment of HRCCs resulted in specific reduction of hTR mrna and inhibition of telomerase activity. Additionally, significant inhibition of proliferation and induction of apoptosis were observed.CONCLUSION sirna agains： the hTR gene can inhibit proliferation and induce apoptosis by blocking telomerase activity of HRCCs. Specific hTR inhibition by sirna represents a promising new option for renal cancer treatment.

关键词： human telomerase rna telomerase small-interfering rna renal cell carcinoma proliferation.

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