Non-viral targeted delivery system mediates transfection of thymidine kinase gene of herpes simplex virus into ovarian cancer cells:a comparison between one time and continuous mediation

Objective: To compare the transferring efficiency and killing effects of one time and continuous mediation with GE7,a non-viral targeted delivery system,in transfection of thymidine kinase gene of herpes simplex virus (HSV-tk) into ovarian cancer cells. Methods: GE7 was used to prepare recombinants with β-galactosidase (β-gal) and HSV1-tk; the re-combinants were then used to transfect human ovarian cancer line CaOV3 once and continuously. β-gal staining was used to compare the efficiencies of one time and continuous mediation with GE7 system. Ganciclovior (GCV) was introduced into HSV1-tk transfected ovarian cells. Through drawing the cell growth curve and flow cytometry,the killing effects of GCV on once and continuously GE7/HSV1-tk transfected cells were observed. Results: We found that the one time and continuous exogenous gene transfer efficiencies were about 80% and 85%,respectively. When 1μg/mL GCV was used to treat ovarian cell transfected with HSV1-tk gene,growth inhibiting rates of ovarian cells of one time and continuous transferring were 82% and 90%,respectively; their apoptosis indices were 15 and 30,respectively. Under same GCV concentration,continuous me-diation of GE7/pCMV-tk transfection into ovarian cancer cells had more significant inhibitory effect than one time mediation (P<0.05). Conclusion: Compared with one time mediation,continuous mediation of transfection with GE7 gene delivery system has higher efficiency. Continuous mediation of GE7/HSV1-tk/GCV therapeutic gene system has more powerful killing effect.

Two less common human microRNAs miR-875 and miR-3144 target a conserved site of E6 oncogene in most high-risk human papillomavirus subtypes

Human papillomaviruses （HPVs） including high.risk （HR） and low-risk （LR） subtypes have distinguishable variation on both genotypes and phenotypes. The co- infection of multiple HR-HPVs, headed by HPV16, is common in cervical cancer in female. Recently accu- mulating reports have focused on the interaction be- tween virus and host, particularly the role of human microRNAs （miRNAs） in anti-viral defense by targeting viral genome. Here, we found a well-conserved target site of miRNAs in the genomes of most HR-HPVs, not LR-HPVs, by scanning all potential target sites of human miRNAs on 24 HPVs of unambiguous subtypes of risk. The site is targeted by two less common human miR- NAs, miR-875 and miR-3144, and is located in E6 onco- gene open reading frame （ORF） and overlap with the first alternative splice exon of viral early transcripts. In validation tests, miR-875 and miR-3144 were identified to suppress the target reporter activity markedly and inhibit the expression of both synthetically exogenous E6 and endogenous E6 oncogene. High level of two miRNAs can inhibit cell growth and promote apoptosisin HPV16-positive cervical cancer cells. This study pro- vides a promising common target of miRNAs for most HR-HPVs and highlights the effects of two low ex- pressed human miRNAs on tumour suppression.

RNA-binding protein LIN28B inhibits apoptosis through regulation of the AKT2/FOXO3A/BIM axis in ovarian cancer cells

LIN28B is an evolutionarily conserved RNA-binding protein that regulates mRNA translation and miRNA let-7 maturation in embryonic stem cells and developing tissues.Increasing evidence demonstrates that LIN28B is activated in cancer and serves as a critical oncogene.However,the underlying molecular mechanisms of LIN28B function in tumorigenesis are still largely unknown.Here we report that LIN28B was expressed in over half of the patients with epithelial ovarian cancer who were examined(n=584).Functional experiments demonstrated that LIN28B inhibited ovarian cancer cell apoptosis.Furthermore,we showed that the proapoptotic factor BIM played an essential role in the antiapoptotic function of LIN28B.RNA-IP microarray analysis suggested that LIN28B binds to mRNAs that are associated with the DNA damage pathway,such as AKT2,in ovarian cancer cells.By binding to AKT2 mRNA and enhancing its protein expression,LIN28B regulated FOXO3A protein phosphorylation and decreased the transcriptional level of BIM,which antagonized the antiapoptosis activity of LIN28B.Taken together,these results mechanistically linked LIN28B and the AKT2/FOXO3A/BIM axis to the apoptosis pathway.The findings may have important implications in the diagnosis and therapeutics of ovarian cancer.