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Surface α-enolase promotes extracellular matrix degradation and tumor metastasis and represents a new therapeutic target.

Hsiao KC, Shih NY, Fang HL, Huang TS, Kuo CC, Chu PY, Hung YM, Chou SW, Yang YY, Chang GC, Liu KJ - PLoS ONE (2013)

Bottom Line: Treatment with antibody against α-enolase in vitro suppressed cell-associated plasminogen and matrix metalloproteinase activation, collagen and gelatin degradation, and cell invasion.Examination of the effect of treatment with shRNA plasmids revealed that down regulation of α-enolase decreases extracellular matrix degradation by and the invasion capacity of lung cancer cells.Adoptive transfer of α-enolase-specific antibody to mice resulted in accumulation of antibody in subcutaneous tumor and inhibited the formation of tumor metastasis in lung and bone.

View Article: PubMed Central - PubMed

Affiliation: Institute of Clinical Pharmacy and Pharmaceutical Sciences, National Cheng Kung University, Tainan, Taiwan.

ABSTRACT
In previous research, we found α-enolase to be inversely correlated with progression-free and overall survival in lung cancer patients and detected α-enolase on the surface of lung cancer cells. Based on these findings, we hypothesized that surface α-enolase has a significant role in cancer metastasis and tested this hypothesis in the current study. We found that α-enolase was co-immunoprecipitated with urokinase-type plasminogen activator, urokinase-type plasminogen activator receptor, and plasminogen in lung cancer cells and interacted with these proteins in a cell-free dot blotting assay, which can be interrupted by α-enolase-specific antibody. α-Enolase in lung cancer cells co-localized with these proteins and was present at the site of pericellular degradation of extracellular matrix components. Treatment with antibody against α-enolase in vitro suppressed cell-associated plasminogen and matrix metalloproteinase activation, collagen and gelatin degradation, and cell invasion. Examination of the effect of treatment with shRNA plasmids revealed that down regulation of α-enolase decreases extracellular matrix degradation by and the invasion capacity of lung cancer cells. Adoptive transfer of α-enolase-specific antibody to mice resulted in accumulation of antibody in subcutaneous tumor and inhibited the formation of tumor metastasis in lung and bone. This study demonstrated that surface α-enolase promotes extracellular matrix degradation and invasion of cancer cells and that targeting surface α-enolase is a promising approach to suppress tumor metastasis.

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Tumor bone metastasis was suppressed by systemic administration of Ab against ENO1.(A) Serum level of Ab against ENO1 in mice adoptively transferred with an isotype-control or mENO1-specific Ab (mENO1 Ab) during the experimental period was determined by an ELISA. (B) Bone metastasis in mice (n = 5) after intracardiac injection of LLC/luc cells and treatment with an isotype-control or mENO1 Ab was detected by the IVIS System. The percentage of mice remained alive and free of bone metastasis after adoptive transfer of an isotype-control or mENO1 Ab was determined. Two mice treated with the isotype-control Ab died from lung metastasis by day 15 (#). The arrows indicated the period of Ab injection. (C) The tumor in the bone of mice treated with an isotype-control Ab was detected by the IVIS System (left 2 panels). Establishment of bone metastasis was confirmed by H&E staining (in the right 2 panels, scale bar is 1mm and 200 µm in each graph). **p<0.01. The error bars were defined as mean±SD.
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pone-0069354-g007: Tumor bone metastasis was suppressed by systemic administration of Ab against ENO1.(A) Serum level of Ab against ENO1 in mice adoptively transferred with an isotype-control or mENO1-specific Ab (mENO1 Ab) during the experimental period was determined by an ELISA. (B) Bone metastasis in mice (n = 5) after intracardiac injection of LLC/luc cells and treatment with an isotype-control or mENO1 Ab was detected by the IVIS System. The percentage of mice remained alive and free of bone metastasis after adoptive transfer of an isotype-control or mENO1 Ab was determined. Two mice treated with the isotype-control Ab died from lung metastasis by day 15 (#). The arrows indicated the period of Ab injection. (C) The tumor in the bone of mice treated with an isotype-control Ab was detected by the IVIS System (left 2 panels). Establishment of bone metastasis was confirmed by H&E staining (in the right 2 panels, scale bar is 1mm and 200 µm in each graph). **p<0.01. The error bars were defined as mean±SD.

Mentions: In lung cancer patients, the incidence of bone metastasis of cancer cells, a process that is also characterized by a series of ECM degradation and invasion events [33], [37], has been estimated at 30% to 40% [38]. We next investigated the effect of blocking ENO1 on tumor bone metastasis in the third animal model. This model was created by intracardiac injection of LLC/luc BM 2nd cells (see Methods for a description of the development of this cell line) into C57BL/6 mice to allow for formation of bone metastasis, followed by systemic administration of either an ENO1-specific Ab or isotype-control Ab in the same manner as previously described (Figure 7A). The mice administered the isotype-control Ab developed lung metastasis between days 6 and 9, whereas those administered the ENO1-specific Ab did so between days 12 and 21 (data not shown). Furthermore, 60% of mice administered the isotype-control Ab developed bone metastasis, as detected by a live-imaging system, with the remainder dying because of lung metastasis at day 15, and all of the mice treated with the ENO1-specific Ab remained free of bone metastasis until day 21 (Figure 7B). The establishment of bone metastasis was confirmed by pathological examination (Figure 7C).


Surface α-enolase promotes extracellular matrix degradation and tumor metastasis and represents a new therapeutic target.

Hsiao KC, Shih NY, Fang HL, Huang TS, Kuo CC, Chu PY, Hung YM, Chou SW, Yang YY, Chang GC, Liu KJ - PLoS ONE (2013)

Tumor bone metastasis was suppressed by systemic administration of Ab against ENO1.(A) Serum level of Ab against ENO1 in mice adoptively transferred with an isotype-control or mENO1-specific Ab (mENO1 Ab) during the experimental period was determined by an ELISA. (B) Bone metastasis in mice (n = 5) after intracardiac injection of LLC/luc cells and treatment with an isotype-control or mENO1 Ab was detected by the IVIS System. The percentage of mice remained alive and free of bone metastasis after adoptive transfer of an isotype-control or mENO1 Ab was determined. Two mice treated with the isotype-control Ab died from lung metastasis by day 15 (#). The arrows indicated the period of Ab injection. (C) The tumor in the bone of mice treated with an isotype-control Ab was detected by the IVIS System (left 2 panels). Establishment of bone metastasis was confirmed by H&E staining (in the right 2 panels, scale bar is 1mm and 200 µm in each graph). **p<0.01. The error bars were defined as mean±SD.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3716638&req=5

pone-0069354-g007: Tumor bone metastasis was suppressed by systemic administration of Ab against ENO1.(A) Serum level of Ab against ENO1 in mice adoptively transferred with an isotype-control or mENO1-specific Ab (mENO1 Ab) during the experimental period was determined by an ELISA. (B) Bone metastasis in mice (n = 5) after intracardiac injection of LLC/luc cells and treatment with an isotype-control or mENO1 Ab was detected by the IVIS System. The percentage of mice remained alive and free of bone metastasis after adoptive transfer of an isotype-control or mENO1 Ab was determined. Two mice treated with the isotype-control Ab died from lung metastasis by day 15 (#). The arrows indicated the period of Ab injection. (C) The tumor in the bone of mice treated with an isotype-control Ab was detected by the IVIS System (left 2 panels). Establishment of bone metastasis was confirmed by H&E staining (in the right 2 panels, scale bar is 1mm and 200 µm in each graph). **p<0.01. The error bars were defined as mean±SD.
Mentions: In lung cancer patients, the incidence of bone metastasis of cancer cells, a process that is also characterized by a series of ECM degradation and invasion events [33], [37], has been estimated at 30% to 40% [38]. We next investigated the effect of blocking ENO1 on tumor bone metastasis in the third animal model. This model was created by intracardiac injection of LLC/luc BM 2nd cells (see Methods for a description of the development of this cell line) into C57BL/6 mice to allow for formation of bone metastasis, followed by systemic administration of either an ENO1-specific Ab or isotype-control Ab in the same manner as previously described (Figure 7A). The mice administered the isotype-control Ab developed lung metastasis between days 6 and 9, whereas those administered the ENO1-specific Ab did so between days 12 and 21 (data not shown). Furthermore, 60% of mice administered the isotype-control Ab developed bone metastasis, as detected by a live-imaging system, with the remainder dying because of lung metastasis at day 15, and all of the mice treated with the ENO1-specific Ab remained free of bone metastasis until day 21 (Figure 7B). The establishment of bone metastasis was confirmed by pathological examination (Figure 7C).

Bottom Line: Treatment with antibody against α-enolase in vitro suppressed cell-associated plasminogen and matrix metalloproteinase activation, collagen and gelatin degradation, and cell invasion.Examination of the effect of treatment with shRNA plasmids revealed that down regulation of α-enolase decreases extracellular matrix degradation by and the invasion capacity of lung cancer cells.Adoptive transfer of α-enolase-specific antibody to mice resulted in accumulation of antibody in subcutaneous tumor and inhibited the formation of tumor metastasis in lung and bone.

View Article: PubMed Central - PubMed

Affiliation: Institute of Clinical Pharmacy and Pharmaceutical Sciences, National Cheng Kung University, Tainan, Taiwan.

ABSTRACT
In previous research, we found α-enolase to be inversely correlated with progression-free and overall survival in lung cancer patients and detected α-enolase on the surface of lung cancer cells. Based on these findings, we hypothesized that surface α-enolase has a significant role in cancer metastasis and tested this hypothesis in the current study. We found that α-enolase was co-immunoprecipitated with urokinase-type plasminogen activator, urokinase-type plasminogen activator receptor, and plasminogen in lung cancer cells and interacted with these proteins in a cell-free dot blotting assay, which can be interrupted by α-enolase-specific antibody. α-Enolase in lung cancer cells co-localized with these proteins and was present at the site of pericellular degradation of extracellular matrix components. Treatment with antibody against α-enolase in vitro suppressed cell-associated plasminogen and matrix metalloproteinase activation, collagen and gelatin degradation, and cell invasion. Examination of the effect of treatment with shRNA plasmids revealed that down regulation of α-enolase decreases extracellular matrix degradation by and the invasion capacity of lung cancer cells. Adoptive transfer of α-enolase-specific antibody to mice resulted in accumulation of antibody in subcutaneous tumor and inhibited the formation of tumor metastasis in lung and bone. This study demonstrated that surface α-enolase promotes extracellular matrix degradation and invasion of cancer cells and that targeting surface α-enolase is a promising approach to suppress tumor metastasis.

Show MeSH
Related in: MedlinePlus