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MicroRNA-192 suppresses liver metastasis of colon cancer.

Geng L, Chaudhuri A, Talmon G, Wisecarver JL, Are C, Brattain M, Wang J - Oncogene (2013)

Bottom Line: Alterations associated with the metastatic phenotype in the primary tumors include increased apoptosis, decreased proliferation and angiogenesis.Finally, studies performed on human colonic adenocarcinoma show that expression of miR-192 is significantly reduced in neoplastic cells as compared with normal colonic epithelium.Importantly, there is a significant decrease in miR-192 expression in stage IV tumors when compared with stage I or II lesions.

View Article: PubMed Central - PubMed

Affiliation: Eppley Institute for Research in Cancer and Allied Diseases, Omaha, NE, USA.

ABSTRACT
Metastasis causes most deaths from colon cancer yet mechanistic understanding and therapeutic options remain limited. Here we show that expression of microRNA (miR)-192 is inversely correlated with metastatic potential of colon cancer cells. Ectopic expression of miR-192 sensitizes colon cancer cells to growth factor deprivation stress-induced apoptosis, whereas inhibition of miR-192 confers resistance. Overexpression of miR-192 inhibits metastatic colonization to the liver in an orthotopic mouse model of colon cancer. Alterations associated with the metastatic phenotype in the primary tumors include increased apoptosis, decreased proliferation and angiogenesis. Further studies indicate that miR-192 downregulates expression of Bcl-2, Zeb2 and VEGFA in vitro and in vivo, which is responsible for enhanced apoptosis, increased expression of E-cadherin and decreased angiogenesis in vivo, respectively. Finally, studies performed on human colonic adenocarcinoma show that expression of miR-192 is significantly reduced in neoplastic cells as compared with normal colonic epithelium. Importantly, there is a significant decrease in miR-192 expression in stage IV tumors when compared with stage I or II lesions. These findings indicate that miR-192 has an important role in colon cancer development and progression. Our studies underscore the clinical relevance and prognostic significance of miR-192 expression in colon cancer. Therefore, a major implication of our studies is that restoration of miR-192 expression or antagonism of its target genes (Bcl-2, Zeb2 or VEGFA) may have considerable therapeutic potential for anti-metastatic therapy in patients with colon cancer.

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miRNA-192 contributes to GFDS-induced apoptosisThe miR-192 precursor was overexpressed in HCT116 (A and B) and RCA cells (C and D) and chemically synthesized miR-192 inhibitor (Inh) or negative control (Con) was transfected into FET cells (E and F). Western blot analysis of cleaved caspase 3 or PARP was performed in HCT116 (A), RCA (C) and FET cells (E) under GFDS for indicated time periods. DNA fragmentation assays were performed in HCT116 cells under GFDS for 1 or 2 days (B, *P < 0.01, **P < 0.05), in RCA cells under GFDS for 2 days (D, *P < 0.1) and in FET cells under GFDS for 2 days (F, *P < 0.05). The data are presented as the mean ± SE of triplicate experiments.
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Figure 2: miRNA-192 contributes to GFDS-induced apoptosisThe miR-192 precursor was overexpressed in HCT116 (A and B) and RCA cells (C and D) and chemically synthesized miR-192 inhibitor (Inh) or negative control (Con) was transfected into FET cells (E and F). Western blot analysis of cleaved caspase 3 or PARP was performed in HCT116 (A), RCA (C) and FET cells (E) under GFDS for indicated time periods. DNA fragmentation assays were performed in HCT116 cells under GFDS for 1 or 2 days (B, *P < 0.01, **P < 0.05), in RCA cells under GFDS for 2 days (D, *P < 0.1) and in FET cells under GFDS for 2 days (F, *P < 0.05). The data are presented as the mean ± SE of triplicate experiments.

Mentions: Although the percentages of cells in G1 and S phase were slightly increased and decreased respectively in miR-192-expressing cells relative to the control cells (Fig. S2), the differences were not statistically significant. However, miR-192-expressing cells displayed significantly increased sensitivity to GFDS-induced apoptosis as reflected by enhanced caspase 3 cleavage as compared to the control cells (Fig. 2A). These observations were further confirmed by DNA fragmentation ELISA assays, which showed 2.5-fold or 80% increase of apoptosis in miR-192-expressing cells when compared to the control cells after exposing to GFDS for one or two days respectively (Fig. 2B, * P < 0.05, ** P < 0.01). To determine whether miR-192 has similar function in other colon cancer cells, miR-192 was ectopically expressed in RCA cells that show low endogenous miR-192 expression (Fig. 1B). The suppressive activity of exogenous miR-192 was indicated by the decrease of luciferase activity of the miR-192 reporter (pMiRluc-192) (Fig. S3A). Similar to HCT116 cells, there was an increase of cleaved caspase 3 in miR-192-expressing RCA cells under GFDS as compared to the control cells (Fig. 2C), which was confirmed by DNA fragmentation assays showing 45% increase of apoptosis (Fig. 2D, *P < 0.05). Of note, miR-192 mimic showed same effect as stably expressed miR-192 in both cell lines (data not shown). These results indicate that ectopic expression of miR-192 sensitizes colon cancer cells to GFDS-induced apoptosis.


MicroRNA-192 suppresses liver metastasis of colon cancer.

Geng L, Chaudhuri A, Talmon G, Wisecarver JL, Are C, Brattain M, Wang J - Oncogene (2013)

miRNA-192 contributes to GFDS-induced apoptosisThe miR-192 precursor was overexpressed in HCT116 (A and B) and RCA cells (C and D) and chemically synthesized miR-192 inhibitor (Inh) or negative control (Con) was transfected into FET cells (E and F). Western blot analysis of cleaved caspase 3 or PARP was performed in HCT116 (A), RCA (C) and FET cells (E) under GFDS for indicated time periods. DNA fragmentation assays were performed in HCT116 cells under GFDS for 1 or 2 days (B, *P < 0.01, **P < 0.05), in RCA cells under GFDS for 2 days (D, *P < 0.1) and in FET cells under GFDS for 2 days (F, *P < 0.05). The data are presented as the mean ± SE of triplicate experiments.
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Related In: Results  -  Collection

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getmorefigures.php?uid=PMC4016997&req=5

Figure 2: miRNA-192 contributes to GFDS-induced apoptosisThe miR-192 precursor was overexpressed in HCT116 (A and B) and RCA cells (C and D) and chemically synthesized miR-192 inhibitor (Inh) or negative control (Con) was transfected into FET cells (E and F). Western blot analysis of cleaved caspase 3 or PARP was performed in HCT116 (A), RCA (C) and FET cells (E) under GFDS for indicated time periods. DNA fragmentation assays were performed in HCT116 cells under GFDS for 1 or 2 days (B, *P < 0.01, **P < 0.05), in RCA cells under GFDS for 2 days (D, *P < 0.1) and in FET cells under GFDS for 2 days (F, *P < 0.05). The data are presented as the mean ± SE of triplicate experiments.
Mentions: Although the percentages of cells in G1 and S phase were slightly increased and decreased respectively in miR-192-expressing cells relative to the control cells (Fig. S2), the differences were not statistically significant. However, miR-192-expressing cells displayed significantly increased sensitivity to GFDS-induced apoptosis as reflected by enhanced caspase 3 cleavage as compared to the control cells (Fig. 2A). These observations were further confirmed by DNA fragmentation ELISA assays, which showed 2.5-fold or 80% increase of apoptosis in miR-192-expressing cells when compared to the control cells after exposing to GFDS for one or two days respectively (Fig. 2B, * P < 0.05, ** P < 0.01). To determine whether miR-192 has similar function in other colon cancer cells, miR-192 was ectopically expressed in RCA cells that show low endogenous miR-192 expression (Fig. 1B). The suppressive activity of exogenous miR-192 was indicated by the decrease of luciferase activity of the miR-192 reporter (pMiRluc-192) (Fig. S3A). Similar to HCT116 cells, there was an increase of cleaved caspase 3 in miR-192-expressing RCA cells under GFDS as compared to the control cells (Fig. 2C), which was confirmed by DNA fragmentation assays showing 45% increase of apoptosis (Fig. 2D, *P < 0.05). Of note, miR-192 mimic showed same effect as stably expressed miR-192 in both cell lines (data not shown). These results indicate that ectopic expression of miR-192 sensitizes colon cancer cells to GFDS-induced apoptosis.

Bottom Line: Alterations associated with the metastatic phenotype in the primary tumors include increased apoptosis, decreased proliferation and angiogenesis.Finally, studies performed on human colonic adenocarcinoma show that expression of miR-192 is significantly reduced in neoplastic cells as compared with normal colonic epithelium.Importantly, there is a significant decrease in miR-192 expression in stage IV tumors when compared with stage I or II lesions.

View Article: PubMed Central - PubMed

Affiliation: Eppley Institute for Research in Cancer and Allied Diseases, Omaha, NE, USA.

ABSTRACT
Metastasis causes most deaths from colon cancer yet mechanistic understanding and therapeutic options remain limited. Here we show that expression of microRNA (miR)-192 is inversely correlated with metastatic potential of colon cancer cells. Ectopic expression of miR-192 sensitizes colon cancer cells to growth factor deprivation stress-induced apoptosis, whereas inhibition of miR-192 confers resistance. Overexpression of miR-192 inhibits metastatic colonization to the liver in an orthotopic mouse model of colon cancer. Alterations associated with the metastatic phenotype in the primary tumors include increased apoptosis, decreased proliferation and angiogenesis. Further studies indicate that miR-192 downregulates expression of Bcl-2, Zeb2 and VEGFA in vitro and in vivo, which is responsible for enhanced apoptosis, increased expression of E-cadherin and decreased angiogenesis in vivo, respectively. Finally, studies performed on human colonic adenocarcinoma show that expression of miR-192 is significantly reduced in neoplastic cells as compared with normal colonic epithelium. Importantly, there is a significant decrease in miR-192 expression in stage IV tumors when compared with stage I or II lesions. These findings indicate that miR-192 has an important role in colon cancer development and progression. Our studies underscore the clinical relevance and prognostic significance of miR-192 expression in colon cancer. Therefore, a major implication of our studies is that restoration of miR-192 expression or antagonism of its target genes (Bcl-2, Zeb2 or VEGFA) may have considerable therapeutic potential for anti-metastatic therapy in patients with colon cancer.

Show MeSH
Related in: MedlinePlus