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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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Effect of YM155, a survivin suppressant, in vitro and in vivoA) YM155 inhibited survivin expression. (Left) SiHa and ME-180 cells were treated with YM155 (0, 1, 10, 100, and 1000 nM) for 24 hrs. Survivin expression was analyzed by Western blotting. (Right) SiHa and ME-180 cells were treated with YM155 (50 nM) for the indicated period of time. Survivin expression was analyzed by Western blotting. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells treated with YM155 (0, 10, and 50 nM) for 24 h. After 14 days incubation, colonies were stained and counted. *P<0.05, bars represent mean ± SEM from triplicates. C) Luc-ME-180 cell subcutaneous xenografts were treated with YM155 (10mg/kg/day). Saline or YM155 was administered as a 3-day continuous infusion per week for 2 weeks. Tumor volume (mm3) was normalized to tumor volume at beginning of treatment (day 1) *P<0.05, bars represent mean (n=8/group) ± SEM. D) Metastatic lymph node analysis. (Left) Representative bioluminescence image. White arrows indicate metastatic lymph nodes. (Right) Quantification of the number metastatic lymph nodes identified in control (saline) vs. YM155 treated mice after 28 days implantation. Saline or YM155 (10 mg/kg/day) was administered as a 3-day continuous infusion per week for 2 weeks after 7 days implantation. Bars represent mean (n=8/groups) ± SEM.
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Figure 5: Effect of YM155, a survivin suppressant, in vitro and in vivoA) YM155 inhibited survivin expression. (Left) SiHa and ME-180 cells were treated with YM155 (0, 1, 10, 100, and 1000 nM) for 24 hrs. Survivin expression was analyzed by Western blotting. (Right) SiHa and ME-180 cells were treated with YM155 (50 nM) for the indicated period of time. Survivin expression was analyzed by Western blotting. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells treated with YM155 (0, 10, and 50 nM) for 24 h. After 14 days incubation, colonies were stained and counted. *P<0.05, bars represent mean ± SEM from triplicates. C) Luc-ME-180 cell subcutaneous xenografts were treated with YM155 (10mg/kg/day). Saline or YM155 was administered as a 3-day continuous infusion per week for 2 weeks. Tumor volume (mm3) was normalized to tumor volume at beginning of treatment (day 1) *P<0.05, bars represent mean (n=8/group) ± SEM. D) Metastatic lymph node analysis. (Left) Representative bioluminescence image. White arrows indicate metastatic lymph nodes. (Right) Quantification of the number metastatic lymph nodes identified in control (saline) vs. YM155 treated mice after 28 days implantation. Saline or YM155 (10 mg/kg/day) was administered as a 3-day continuous infusion per week for 2 weeks after 7 days implantation. Bars represent mean (n=8/groups) ± SEM.

Mentions: YM155 decreased survivin expression in both a concentration and time dependent manner (Figure 5A). As well, YM155 significantly reduced clonogenicity of SiHa and ME-180 cells, which was comparable to survivin siRNA (Figures 5B and 4B). In order to examine the effects of YM155 against tumor growth and lymph node metastasis in cervical cancer cells, we generated luciferase-expressing ME-180 cells (Luc-ME-180 cells), and evaluated in vivo anti-tumor activity. Mice treated with YM155 had significantly reduced tumor growth compared to control mice treated with saline (53% reduction at day 22, P<0.05, Figure 5C). We evaluated intratumoral survivin protein expression at days 0, 3, and 7, and confirmed inhibition of survivin expression during YM155 treatment (compared to saline controls; Figure S4A). Next, the inhibitory effect of YM155 on lymph node metastases was examined using an orthotopic xenograft model of cervical cancer [24]. After 28 days implantation, the number of lymph node metastases in YM155 vs. saline treated mice was evaluated using bioluminescence. All metastatic lymph nodes detected by bioluminescence were confirmed using histological analyses (Figure S4B). Importantly, YM155 significantly reduced the number of lymph node metastasis (P<0.05, Figure 5D), without affecting the large primary tumor transplanted into the cervix (data not shown).


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)

Effect of YM155, a survivin suppressant, in vitro and in vivoA) YM155 inhibited survivin expression. (Left) SiHa and ME-180 cells were treated with YM155 (0, 1, 10, 100, and 1000 nM) for 24 hrs. Survivin expression was analyzed by Western blotting. (Right) SiHa and ME-180 cells were treated with YM155 (50 nM) for the indicated period of time. Survivin expression was analyzed by Western blotting. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells treated with YM155 (0, 10, and 50 nM) for 24 h. After 14 days incubation, colonies were stained and counted. *P<0.05, bars represent mean ± SEM from triplicates. C) Luc-ME-180 cell subcutaneous xenografts were treated with YM155 (10mg/kg/day). Saline or YM155 was administered as a 3-day continuous infusion per week for 2 weeks. Tumor volume (mm3) was normalized to tumor volume at beginning of treatment (day 1) *P<0.05, bars represent mean (n=8/group) ± SEM. D) Metastatic lymph node analysis. (Left) Representative bioluminescence image. White arrows indicate metastatic lymph nodes. (Right) Quantification of the number metastatic lymph nodes identified in control (saline) vs. YM155 treated mice after 28 days implantation. Saline or YM155 (10 mg/kg/day) was administered as a 3-day continuous infusion per week for 2 weeks after 7 days implantation. Bars represent mean (n=8/groups) ± SEM.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
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Figure 5: Effect of YM155, a survivin suppressant, in vitro and in vivoA) YM155 inhibited survivin expression. (Left) SiHa and ME-180 cells were treated with YM155 (0, 1, 10, 100, and 1000 nM) for 24 hrs. Survivin expression was analyzed by Western blotting. (Right) SiHa and ME-180 cells were treated with YM155 (50 nM) for the indicated period of time. Survivin expression was analyzed by Western blotting. B) Clonogenic assays were performed by seeding SiHa and ME-180 cells treated with YM155 (0, 10, and 50 nM) for 24 h. After 14 days incubation, colonies were stained and counted. *P<0.05, bars represent mean ± SEM from triplicates. C) Luc-ME-180 cell subcutaneous xenografts were treated with YM155 (10mg/kg/day). Saline or YM155 was administered as a 3-day continuous infusion per week for 2 weeks. Tumor volume (mm3) was normalized to tumor volume at beginning of treatment (day 1) *P<0.05, bars represent mean (n=8/group) ± SEM. D) Metastatic lymph node analysis. (Left) Representative bioluminescence image. White arrows indicate metastatic lymph nodes. (Right) Quantification of the number metastatic lymph nodes identified in control (saline) vs. YM155 treated mice after 28 days implantation. Saline or YM155 (10 mg/kg/day) was administered as a 3-day continuous infusion per week for 2 weeks after 7 days implantation. Bars represent mean (n=8/groups) ± SEM.
Mentions: YM155 decreased survivin expression in both a concentration and time dependent manner (Figure 5A). As well, YM155 significantly reduced clonogenicity of SiHa and ME-180 cells, which was comparable to survivin siRNA (Figures 5B and 4B). In order to examine the effects of YM155 against tumor growth and lymph node metastasis in cervical cancer cells, we generated luciferase-expressing ME-180 cells (Luc-ME-180 cells), and evaluated in vivo anti-tumor activity. Mice treated with YM155 had significantly reduced tumor growth compared to control mice treated with saline (53% reduction at day 22, P<0.05, Figure 5C). We evaluated intratumoral survivin protein expression at days 0, 3, and 7, and confirmed inhibition of survivin expression during YM155 treatment (compared to saline controls; Figure S4A). Next, the inhibitory effect of YM155 on lymph node metastases was examined using an orthotopic xenograft model of cervical cancer [24]. After 28 days implantation, the number of lymph node metastases in YM155 vs. saline treated mice was evaluated using bioluminescence. All metastatic lymph nodes detected by bioluminescence were confirmed using histological analyses (Figure S4B). Importantly, YM155 significantly reduced the number of lymph node metastasis (P<0.05, Figure 5D), without affecting the large primary tumor transplanted into the cervix (data not shown).

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