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The microRNA-218~Survivin axis regulates migration, invasion, and lymph node metastasis in cervical cancer.

Kogo R, How C, Chaudary N, Bruce J, Shi W, Hill RP, Zahedi P, Yip KW, Liu FF - Oncotarget (2015)

Bottom Line: Lower miR-218 expression was associated significantly with worse overall survival (OS), disease-free survival (DFS), and pelvic/aortic lymph node recurrence.In vitro, miR-218 over-expression decreased clonogenicity, migration, and invasion.YM155, a small molecule survivin inhibitor, significantly suppressed tumor growth and lymph node metastasis in vivo.

View Article: PubMed Central - PubMed

Affiliation: Ontario Cancer Institute, University Health Network (UHN), Toronto, Ontario, Canada.

ABSTRACT
Cervical cancer is the third most common cancer in women worldwide. In the present study, global microRNA profiling for 79 cervical cancer patient samples led to the identification of miR-218 down-regulation in cervical cancer tissues compared to normal cervical tissues. Lower miR-218 expression was associated significantly with worse overall survival (OS), disease-free survival (DFS), and pelvic/aortic lymph node recurrence. In vitro, miR-218 over-expression decreased clonogenicity, migration, and invasion. Survivin (BIRC5) was subsequently identified as an important cervical cancer target of miR-218 using in silico prediction, mRNA profiling, and quantitative real-time PCR (qRT-PCR). Concordant with miR-218 over-expression, survivin knockdown by siRNA decreased clonogenicity, migration, and invasion. YM155, a small molecule survivin inhibitor, significantly suppressed tumor growth and lymph node metastasis in vivo. Our findings demonstrate that the miR-218~survivin axis inhibits cervical cancer progression by regulating clonogenicity, migration, and invasion, and suggest that the inhibition of survivin could be a potential therapeutic strategy to improve outcome in this disease.

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Survivin knockdown reduced survival, migration, and invasion in cervical cancer cellsA) mRNA (left) and protein (right) survivin expression after 48 hrs negative control siRNA or survivin siRNAs (10 nM each) transfection in SiHa and ME-180 cells. Survivin mRNA expression levels were normalized to GAPDH, relative to cells transfected with negative control siRNA. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each). At 48 hrs post-transfection, cells were re-seeded at low density in 6-well plates. After 10-14 days incubation, colonies were stained and counted. C) Representative image (left) and quantification bar graph (right) of migrated SiHa and ME-180 cells. Migration assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in trans-well chambers. After 48 hrs incubation, migrated cells were stained and counted. D) Representative image (left) and quantification bar graph (right) of invaded SiHa and ME-180 cells. Invasion assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in the Matrigel invasion chambers. After 48 hrs incubation, invaded cells were stained and counted. A-D: *P<0.05, bars represent mean ± SEM from triplicates.
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Figure 4: Survivin knockdown reduced survival, migration, and invasion in cervical cancer cellsA) mRNA (left) and protein (right) survivin expression after 48 hrs negative control siRNA or survivin siRNAs (10 nM each) transfection in SiHa and ME-180 cells. Survivin mRNA expression levels were normalized to GAPDH, relative to cells transfected with negative control siRNA. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each). At 48 hrs post-transfection, cells were re-seeded at low density in 6-well plates. After 10-14 days incubation, colonies were stained and counted. C) Representative image (left) and quantification bar graph (right) of migrated SiHa and ME-180 cells. Migration assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in trans-well chambers. After 48 hrs incubation, migrated cells were stained and counted. D) Representative image (left) and quantification bar graph (right) of invaded SiHa and ME-180 cells. Invasion assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in the Matrigel invasion chambers. After 48 hrs incubation, invaded cells were stained and counted. A-D: *P<0.05, bars represent mean ± SEM from triplicates.

Mentions: Survivin is the smallest member of the inhibitor of apoptosis (IAP) family, and is mainly associated with the regulation of mitosis and inhibition of apoptosis [21]. Some functions of survivin still remain unknown because survivin interacts with numerous proteins. In order to characterize survivin function in cervical cancer cells, SiHa and ME-180 cells were transfected with survivin siRNAs. Survivin knockdown was confirmed both at the mRNA and protein level (Figure 4A). This down-regulation was accompanied by a significant reduction in clonogenicity compared to control cells transfected with negative control siRNA (P<0.05; Figure 4B). Additionally, survivin knockdown also reduced migration and invasion capacities, phenocopying miR-218 over-expression (P<0.05; Figures 4C and 4D). Overall, these data support the postulate that the miR-218~survivin axis regulates clonogenicity, migration, and invasion in cervical cancer.


The microRNA-218~Survivin axis regulates migration, invasion, and lymph node metastasis in cervical cancer.

Kogo R, How C, Chaudary N, Bruce J, Shi W, Hill RP, Zahedi P, Yip KW, Liu FF - Oncotarget (2015)

Survivin knockdown reduced survival, migration, and invasion in cervical cancer cellsA) mRNA (left) and protein (right) survivin expression after 48 hrs negative control siRNA or survivin siRNAs (10 nM each) transfection in SiHa and ME-180 cells. Survivin mRNA expression levels were normalized to GAPDH, relative to cells transfected with negative control siRNA. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each). At 48 hrs post-transfection, cells were re-seeded at low density in 6-well plates. After 10-14 days incubation, colonies were stained and counted. C) Representative image (left) and quantification bar graph (right) of migrated SiHa and ME-180 cells. Migration assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in trans-well chambers. After 48 hrs incubation, migrated cells were stained and counted. D) Representative image (left) and quantification bar graph (right) of invaded SiHa and ME-180 cells. Invasion assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in the Matrigel invasion chambers. After 48 hrs incubation, invaded cells were stained and counted. A-D: *P<0.05, bars represent mean ± SEM from triplicates.
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Figure 4: Survivin knockdown reduced survival, migration, and invasion in cervical cancer cellsA) mRNA (left) and protein (right) survivin expression after 48 hrs negative control siRNA or survivin siRNAs (10 nM each) transfection in SiHa and ME-180 cells. Survivin mRNA expression levels were normalized to GAPDH, relative to cells transfected with negative control siRNA. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each). At 48 hrs post-transfection, cells were re-seeded at low density in 6-well plates. After 10-14 days incubation, colonies were stained and counted. C) Representative image (left) and quantification bar graph (right) of migrated SiHa and ME-180 cells. Migration assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in trans-well chambers. After 48 hrs incubation, migrated cells were stained and counted. D) Representative image (left) and quantification bar graph (right) of invaded SiHa and ME-180 cells. Invasion assays were performed by seeding SiHa and ME-180 cells transfected with negative control siRNA or survivin siRNAs (10 nM each) in the Matrigel invasion chambers. After 48 hrs incubation, invaded cells were stained and counted. A-D: *P<0.05, bars represent mean ± SEM from triplicates.
Mentions: Survivin is the smallest member of the inhibitor of apoptosis (IAP) family, and is mainly associated with the regulation of mitosis and inhibition of apoptosis [21]. Some functions of survivin still remain unknown because survivin interacts with numerous proteins. In order to characterize survivin function in cervical cancer cells, SiHa and ME-180 cells were transfected with survivin siRNAs. Survivin knockdown was confirmed both at the mRNA and protein level (Figure 4A). This down-regulation was accompanied by a significant reduction in clonogenicity compared to control cells transfected with negative control siRNA (P<0.05; Figure 4B). Additionally, survivin knockdown also reduced migration and invasion capacities, phenocopying miR-218 over-expression (P<0.05; Figures 4C and 4D). Overall, these data support the postulate that the miR-218~survivin axis regulates clonogenicity, migration, and invasion in cervical cancer.

Bottom Line: Lower miR-218 expression was associated significantly with worse overall survival (OS), disease-free survival (DFS), and pelvic/aortic lymph node recurrence.In vitro, miR-218 over-expression decreased clonogenicity, migration, and invasion.YM155, a small molecule survivin inhibitor, significantly suppressed tumor growth and lymph node metastasis in vivo.

View Article: PubMed Central - PubMed

Affiliation: Ontario Cancer Institute, University Health Network (UHN), Toronto, Ontario, Canada.

ABSTRACT
Cervical cancer is the third most common cancer in women worldwide. In the present study, global microRNA profiling for 79 cervical cancer patient samples led to the identification of miR-218 down-regulation in cervical cancer tissues compared to normal cervical tissues. Lower miR-218 expression was associated significantly with worse overall survival (OS), disease-free survival (DFS), and pelvic/aortic lymph node recurrence. In vitro, miR-218 over-expression decreased clonogenicity, migration, and invasion. Survivin (BIRC5) was subsequently identified as an important cervical cancer target of miR-218 using in silico prediction, mRNA profiling, and quantitative real-time PCR (qRT-PCR). Concordant with miR-218 over-expression, survivin knockdown by siRNA decreased clonogenicity, migration, and invasion. YM155, a small molecule survivin inhibitor, significantly suppressed tumor growth and lymph node metastasis in vivo. Our findings demonstrate that the miR-218~survivin axis inhibits cervical cancer progression by regulating clonogenicity, migration, and invasion, and suggest that the inhibition of survivin could be a potential therapeutic strategy to improve outcome in this disease.

Show MeSH
Related in: MedlinePlus