Stable and complete overcoming of MDR1/P-glycoprotein-mediated multidrug resistance in human gastric carcinoma cells by RNA interference

Abstract

Multidrug resistance (MDR) is the major cause of failure of effective chemotherapeutic treatment of disseminated neoplasms. The ''classical'' MDR phenotype of human malignancies is mediated by drug extrusion by the adenosine triphosphate binding cassette (ABC)-transporter P-glycoprotein (MDR1/P-gp). For stable reversal of ''classical'' MDR by RNA interference (RNAi) technology, an H1-RNA gene promoter-driven expression vector encoding anti-MDR1/P-gp short hairpin RNA (shRNA) molecules was constructed. By introduction of anti-MDR1/P-gp shRNA expression vectors into the extremely high drug-resistant human gastric carcinoma cell line EPG85-257RDB, the MDR phenotype was completely reversed. The reversal of MDR was accompanied by a complete suppression of MDR1/P-gp expression on mRNA and protein level, and by a considerable increased intracellular anthracyline accumulation in the anti-MDR1/P-gp shRNA-treated cells. The data indicate that stable shRNA-mediated RNAi can be tremendously effective in reversing MDR1/P-gp-mediated MDR and is therefore a promising strategy for overcoming MDR by gene therapeutic applications.

Calcium carbonate nanoparticle delivering vascular endothelial growth factor-C siRNA effectively inhibits lymphangiogenesis and growth of gastric cancer in vivo

“…29 Recent reports have described DNA vector-based strategies for delivery of small interfering RNA (siRNA) into mammalian cells. [30][31][32] These traits of RNA interference suggest that it could serve as an appropriate candidate for use as a gene therapy agent when coupled with an appropriate delivery system.…”

Silencing the Breast Cancer Resistance Protein Expression and Function in Caco-2 Cells Using Lentiviral Vector-Based Short Hairpin RNA

“…This technique has been under extensive investigation and has been applied recently to the inhibition of MDR1 (Stege et al, 2004;Yagüe et al, 2004) and BCRP expression (Ee et al, 2004;Li et al, 2005;Priebsch et al, 2006;Lv et al, 2007). Results from these studies have shown that RNAi is useful for down-regulation of BCRP.…”

Delivery of short hairpin RNAs by transkingdom RNA interference modulates the classical ABCB1-mediated multidrug-resistant phenotype of cancer cells

“…15 When a gene therapy-like approach had been used in conjunction with plasmid-or adenovirus-encoded shRNAs, an even more complete knock down of the ABCB1 mRNA expression was noted. 8,13 The extent of inhibition of drug transport activity and reversal of drug resistance by the tkRNAi platform was less pronounced compared to existing RNAi strategies, and the amount of anti-ABCB1 shRNA recovered from the tkRNAi-treated gastric carcinoma cells was lower compared to that in cells treated with shRNA-encoding adenoviruses. Relative drug resistance levels determined by a cytotoxicity assay for cell survival whereby the drug-resistance level of drug-resistant epG85-257RDB cells was set to be 100%.…”

“…7 This cell model has previously been evaluated for different RNAi strategies such as transient in vitro downregulation of ABCB1 by chemically synthesized siRNA molecules, 15 adenovirus-administered shRNAs, 13 as well as stable knock down by plasmid encoded shRNAs. 8 Identical shRNA sequences had also shown efficacy for in vivo targeting of ABCB1 by RNAi. 10 This study confirmed that the tkRNAi approach induces targeted gene-silencing resulting in downregulation of the ABCB1-encoding mRNA and the corresponding ABC-transporter molecule.…”

“…The RNAi targeting sequence of ABCB1 was 5'-ATG TTG TCT GGA CAA GCA CT-3'. 8 The design of the DNA template encoding for shRNA against ABCB1 was: BamHI site-sense sequenceloop (5'-TTC AAG AGA-3')-antisense sequence-SalI site. Two oligodeoxynucleotides were synthesized.…”