Tumor inhibition by genomically integrated inducible RNAi-cassettes.
RNA interference (RNAi) has emerged as a powerful tool to induce loss-of-function phenotypes by post-transcriptional silencing of gene expression. In this study we wondered whether inducible RNAi-cassettes integrated into cellular DNA possess the power to trigger neoplastic growth. For this purpose inducible RNAi vectors containing tetracycline (Tet)-responsive derivatives of the H1 promoter for the conditional expression of short hairpin RNA (shRNA) were used to target human polo-like kinase 1 (Plk1), which is overexpressed in a broad spectrum of human tumors. In the absence of doxycycline (Dox) HeLa clones expressing TetR, that carry the RNAi-cassette stably integrated, exhibited no significant alteration in Plk1 expression levels. In contrast, exposure to Dox led to marked downregulation of Plk1 mRNA to 3% and Plk1 protein to 14% in cell culture compared to mismatch shRNA/Plk1-expressing cells. As a result of Plk1 depletion cell proliferation decreased to 17%. Furthermore, for harnessing RNAi for silencing disease-related genes in vivo we transplanted inducible RNAi-HeLa cells onto nude mice. After administration of Dox knockdown of Plk1 expression was observed correlating to a significant inhibition of tumor growth. Taken together, our data revealed that genomically integrated RNAi-elements are suitable to hamper tumor growth by conditional expression of shRNA.
Kappel
Sven
S
Department of Gynecology and Obstetrics, School of Medicine, J.W. Goethe-University, Theodor-Stern-Kai 7, D-60590 Frankfurt, Germany.
Matthess
Yves
Y
Zimmer
Brigitte
B
Kaufmann
Manfred
M
Strebhardt
Klaus
K
D000818:Animals; D018797:Cell Cycle Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D049109:Cell Proliferation; D004318:Doxycycline / Q000494:pharmacology; D005786:Gene Expression Regulation; D005822:Genetic Vectors; D015894:Genome, Human; D006367:HeLa Cells; D006801:Humans; D008297:Male; D051379:Mice; D009369:Neoplasms / Q000201:enzymology / Q000473:pathology / Q000628:therapy*; D010957:Plasmids; D011401:Promoter Regions, Genetic; D017346:Protein Serine-Threonine Kinases / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D011518:Proto-Oncogene Proteins / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D034622:RNA Interference; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism; D023041:Xenograft Model Antitumor Assays; D000097869:Polo-Like Kinase 1
D018797:Cell Cycle Proteins; D004318:Doxycycline; D017346:Protein Serine-Threonine Kinases; D011518:Proto-Oncogene Proteins; D012330:RNA, Double-Stranded; D000097869:Polo-Like Kinase 1