Slug (SNAI2) down-regulation by RNA interference facilitates apoptosis and inhibits invasive growth in neuroblastoma preclinical models.
OBJECTIVE
We assessed the relevance of Slug (SNAI2) for apoptosis resistance and invasion potential of neuroblastoma cells in vitro and in vivo.


METHODS
We evaluated the effect of imatinib mesylate on invasion and analyzed the genes modulated by imatinib mesylate treatment in neuroblastoma cells. Slug expression, inhibited by imatinib mesylate treatment, was knocked down in neuroblastoma cells by RNA interference, and the effects on invasion and apoptosis were evaluated in vitro. A pseudometastatic model of neuroblastoma in severe combined immunodeficient mice was used to assess the effects of Slug silencing alone or in combination with imatinib mesylate treatment on metastasis development.


RESULTS
Microarray analysis revealed that several genes, including Slug, were down-regulated by imatinib mesylate. Slug expression was detectable in 8 of 10 human neuroblastoma cell lines. Two Slug-expressing cell lines were infected with a vector encoding a microRNA to Slug mRNA. Infected cells with reduced levels of Slug were tested for the expression of apoptosis-related genes (p53, Bax, and Bcl-2) identified previously as Slug targets. Bcl-2 was down-regulated in Slug-interfered cells. Slug down-regulation increased sensitivity to apoptosis induced by imatinib mesylate, etoposide, or doxorubicin. Invasion of Slug-silenced cells was reduced in vitro. Animals injected with Slug-silenced cells had fewer tumors than controls and the inhibition of tumor growth was even higher in animals treated with imatinib mesylate.


CONCLUSIONS
Slug down-regulation facilitates apoptosis induced by proapoptotic drugs in neuroblastoma cells and decreases their invasion capability in vitro and in vivo. Slug inhibition, possibly combined with imatinib mesylate, may represent a novel strategy for treatment of metastatic neuroblastoma.
Vitali
Roberta
R
Section of Toxicology and Biomedical Sciences, ENEA Research Center Casaccia, Rome, Italy.
Mancini
Camillo
C
Cesi
Vincenzo
V
Tanno
Barbara
B
Mancuso
Mariateresa
M
Bossi
Gianluca
G
Zhang
Ying
Y
Martinez
Robert V
RV
Calabretta
Bruno
B
Dominici
Carlo
C
Raschellà
Giuseppe
G
D000818:Animals; D000970:Antineoplastic Agents / Q000494:pharmacology; D017209:Apoptosis / Q000502:physiology*; D001549:Benzamides; D015153:Blotting, Western; D045744:Cell Line, Tumor; D049109:Cell Proliferation; D015536:Down-Regulation; D004591:Electrophoresis, Polyacrylamide Gel; D005434:Flow Cytometry; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D000068877:Imatinib Mesylate; D051379:Mice; D009361:Neoplasm Invasiveness; D009447:Neuroblastoma / Q000188:drug therapy / Q000378:metabolism*; D020411:Oligonucleotide Array Sequence Analysis; D010879:Piperazines / Q000494:pharmacology; D011743:Pyrimidines / Q000494:pharmacology; D034622:RNA Interference; D020133:Reverse Transcriptase Polymerase Chain Reaction; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors / Q000378:metabolism*
D000970:Antineoplastic Agents; D001549:Benzamides; D010879:Piperazines; D011743:Pyrimidines; C000604407:SNAI1 protein, human; C000604408:SNAI2 protein, human; C000604449:Snai2 protein, mouse; D000071250:Snail Family Transcription Factors; D014157:Transcription Factors; D000068877:Imatinib Mesylate