Novel lentiviral-inducible transgene expression systems and versatile single-plasmid reporters for in vitro and in vivo cancer biology studies
| Autor: | R Kan, Maria Li Lung, Wai Ho Shuen, Z Yu, H L Lung |
|---|---|
| Rok vydání: | 2015 |
| Předmět: |
Transcriptional Activation
Cancer Research Carcinogenesis Transgene Genetic Vectors Gene Expression Mice Nude Biology Transduction (genetics) Genes Reporter Transduction Genetic In vivo Cell Line Tumor Neoplasms Chlorocebus aethiops Gene expression Animals Humans Luciferase Transgenes Promoter Regions Genetic Molecular Biology Luciferases Renilla Mice Inbred BALB C Lentivirus HEK 293 cells Genetic Therapy Transfection Molecular biology In vitro Cell biology HEK293 Cells COS Cells Molecular Medicine Female Neoplasm Transplantation Plasmids |
| Zdroj: | Cancer Gene Therapy. 22:207-214 |
| ISSN: | 1476-5500 0929-1903 |
| DOI: | 10.1038/cgt.2015.9 |
| Popis: | Many of the cancer cell lines derived from solid tumors are difficult to transfect using commonly established transfection approaches. This hurdle for some DNA transfection systems has hindered cancer biology studies. Moreover, there are limited tools for studying pathway activities. Therefore, highly efficient improved gene transfer and versatile genetic tools are required. In this study, we established and developed a comprehensive set of new lentiviral tools to study gene functions and pathway activities. Using the optimized conditions, cancer cell lines achieved >90% transduction efficiency. Novel lentiviral doxycycline-regulated pTet-IRES-EGFP (pTIE) systems for transgene expression and TRE reporters used for pathway activity determination were developed and tested. The pTIE Tet-Off system showed in vitro doxycycline-sensitive responses with low or undetectable leakage of protein expression and in vivo tumor suppression as illustrated using candidate tumor suppressors, Fibulin-2 and THY1. In contrast, the Tet-On system showed dose-dependent responses. The pTRE-EGFP (pTE) and pTRE-FLuc-EF1α-RLuc (pT-FER) reporters with the NFκB p65 subunit consensus sequence showed GFP and firefly luciferase responses, which were directly correlated with TNFα stimulation, respectively. Taken together, these newly developed lentiviral systems provide versatile in vitro and in vivo platforms to strengthen our capabilities for cancer biology studies. |
| Databáze: | OpenAIRE |
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