The functional role of long non-coding RNA PANDAR in promoting colorectal cancer metastasis and its mechanism / 中国癌症杂志

Background and purpose: Accumulating evidence has revealed that long non-coding RNA (lncRNA) is correlated with carcinogenesis and tumor development. Recent literature suggested that lncRNA promoter of CDKN1A antisense DNA damage activated RNA (PANDAR) was involved in the development of various cancers. However, the functional role of PANDAR in colorectal cancer (CRC) has not been elucidated yet. The present study aimed to explore the functional role of lncRNA PANDAR in promoting CRC metastasis and its mechanism.Methods: The expression of lncRNA PANDAR in CRC cell lines and tissues was detected by real-time fluorescent quantitative polymerase chain reaction (RTFQ-PCR), and the correlation between lncRNA PANDAR expression and CRC clinicopathological characteristics was statistically analyzed. Then, lncRNA PANDAR stably silencing CRC cells (HCT116-shPANDAR), overexpression cells (DLD1-PANDAR) and control vector cells (HCT116-shNC and DLD1-vector) were established using lentiviral vectors. Moreover, Transwell assay and Matrigel assay were performed to investigate the function of lncRNA PANDAR in CRC migration and invasion. Furthermore, the expression of transcriptional factors mediating epithelial-mesenchymal transition of lncRNA PANDAR overexpression cells were monitored by RTFQ-PCR assay, and the function of the target gene in modulating lncRNA PANDAR mediated CRC metastasis was also explored. Results: The expression levels of lncRNA PANDAR in normal colorectal epithelial cells were much lower than in CRC cell. The levels of lncRNA PANDAR in tumor-adjacent tissues were verified to be much lower than in CRC tissues [(171.52±97.80)% vs (100.00±63.18)%, P<0.05]. Moreover, the expression of lncRNA PANDAR was detected to be significantly correlated with CRC TNM stage, lymph node metastasis and distant metastasis (P<0.05). Besides, lncRNA PANDAR deficiency significantly reduced the migration [100.00% vs (42.08±4.77)%, P<0.05] and invasion [100.00% vs (39.14±3.81)%, P<0.05] capabilities in CRC cells, in contrast, the migration [100.00% vs (194.12±9.33)%, P<0.05] and invasion [100.00% vs (204.08±12.27)%, P<0.05] capa-bilities of CRC cells were obviously increased with lncRNA PANDAR overexpression. Furthermore, zinc-finger E-box binding homeobox 1 (ZEB1) expression was detected to be positively correlated with lncRNA PANDAR expression, and ZEB1 silencing could significantly reverse the increased migration and invasion capabilities induced by lncRNA PANDAR in CRC cells. Conclusion: LncRNA PANDAR could promote CRC metastasis by potentially targeting ZEB1. LncRNA PANDAR might be a promising diagnostic marker and therapeutic target for CRC patients.

Ovarian cancer related gene targeting with large circular antisense library / 대한산부인과학회지

OBJECTIVE: The aim of this study is to find out the genes which are related to ovarian cancer cell growth using large circular antisense library. METHODS: Clones for antisense library were uni-directionally sub-cloned into pBS SK (-) vector. LC-antisense molecules were then purified from the culture supernatants of the bacterial competent cells superinfected with M13K07 helper bacteriophages. The LC-antisense library to 240 unigene clone was constructed and utilized in the identification of genes functionally involved in the growth of ovarian cancer cells. RESULTS: The 17 numbers out of the 240 numbers of the antisense library exerted a marked inhibitory effect on the growth of SK-OV 3. CONCLUSION: The putative functional categorization of each gene was then conducted via public databases. These candidates may be used as target genes for drug development or adjuvant of conventional chemotherapeutic drugs.

Apoptosis of Prostate Cancer by Inhibition of Bcl-xL Expression / 대한비뇨기과학회지

PURPOSE: In this study, we evaluated in vitro whether the anti-sense transfection that was targeted against Bcl-xL would induce cytotoxicity via apoptosis in prostate cancer cells. MATERIALS AND METHODS: cDNA of the human Bcl-xL gene was obtained by RT-PCR amplification, and the anti-sense Bcl-xL mRNA plasmid was generated using the pCR 3.1 TOPO plasmid vector. The function of the cloned anti-sense Bcl-xL plasmid vector (pCR3.1-AS-Bcl-xL) was evaluated by the Western blot analysis. Using the MTT assay, the efficacy of growth inhibition by transfection with pCR3.1-AS-Bcl-xL was tested in vitro on PC-3 and DU145 human prostate cancer cell lines. Immunoblot analyses of Bax and caspase-9 were also performed. To evaluate the apoptosis, DNA fragmentation and caspase-3 assay were performed. RESULTS: Bcl-xL expression after transfection with pCR3.1-AS-Bcl-xL was gradually decreased in PC-3 cells and was continuously decreased in DU145 cells, compared to the parent cells. Bax protein was not expressed in DU145 cells, and the levels of Bax protein expression was not altered in the transfected PC-3 cells compared to the parent cells. The cytotoxicity of pCR3.1-AS-Bcl-xL on PC-3 and DU145 cells increased significantly compared to the empty vector, pCR3.1. This increased cytotoxicity was associated with enhanced apoptosis as assessed by the DNA fragmentation assay and the caspase-3 assay. The expression of the active caspase-9 was increased in the PC-3 cells but not in the DU145 cells after transfection with pCR3.1-AS-Bcl-xL. CONCLUSIONS: Our results showed that the suppression of Bcl-xL by anti-sense transfection efficiently inhibited the growth of PC-3 and DU145 human prostate cancer cell lines. The inhibition of Bcl-xL expression can possibly be a novel therapeutic alternative in the treatment of hormone refractory prostate carcinoma.

Construction of Recombinant HSU17714 Gene Antisense Expressive Plasmid and Its Influence on Colon Cancer Cells Growth / 中国肿瘤生物治疗杂志

HSU17714 gene is a novel human colorectal carcinoma related gene isolated by subtractive hybridization method. The recombinant antisense expressive plasmid of HSU17714 gene was constructed by inserting the partial sequence (2612 bp) of this gene into the multiple cloning site of the vector pREP9 inversely. Then, it was transferred into human colon adenocarcinoma cells (SW1116) through liposome mediation. Obtained from the two-layer soft agarose culture test, flowcytometry test and cell growth rate detection, the data indicated that the antisense HSU17714 could inhibit the growth of SW1116 cells at different degrees. So, a conclusion could be drawn that the antisense HSU17714 gene expressive vector plays a role in the inhibition of SW1116 cell growth.