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A dominant-negative FGF1 mutant (the R50E mutant) suppresses tumorigenesis and angiogenesis.

Mori S, Tran V, Nishikawa K, Kaneda T, Hamada Y, Kawaguchi N, Fujita M, Saegusa J, Takada YK, Matsuura N, Zhao M, Takada Y - PLoS ONE (2013)

Bottom Line: R50E is defective in inducing ternary complex formation, cell proliferation, and cell migration, and suppresses FGF signaling induced by WT FGF1 (a dominant-negative effect) in vitro.Taken together, our results suggest that R50E suppresses angiogenesis induced by FGF1 or FGF2, and thereby indirectly suppresses tumorigenesis, in addition to its possible direct effect on tumor cell proliferation in vivo.We propose that R50E has potential as an anti-cancer and anti-angiogenesis therapeutic agent ("FGF1 decoy").

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pathology, Osaka University Graduate School of Medicine, Division of Health Sciences, Suita, Osaka, Japan.

ABSTRACT
Fibroblast growth factor-1 (FGF1) and FGF2 play a critical role in angiogenesis, a formation of new blood vessels from existing blood vessels. Integrins are critically involved in FGF signaling through crosstalk. We previously reported that FGF1 directly binds to integrin αvβ3 and induces FGF receptor-1 (FGFR1)-FGF1-integrin αvβ3 ternary complex. We previously generated an integrin binding defective FGF1 mutant (Arg-50 to Glu, R50E). R50E is defective in inducing ternary complex formation, cell proliferation, and cell migration, and suppresses FGF signaling induced by WT FGF1 (a dominant-negative effect) in vitro. These findings suggest that FGFR and αvβ3 crosstalk through direct integrin binding to FGF, and that R50E acts as an antagonist to FGFR. We studied if R50E suppresses tumorigenesis and angiogenesis. Here we describe that R50E suppressed tumor growth in vivo while WT FGF1 enhanced it using cancer cells that stably express WT FGF1 or R50E. Since R50E did not affect proliferation of cancer cells in vitro, we hypothesized that R50E suppressed tumorigenesis indirectly through suppressing angiogenesis. We thus studied the effect of R50E on angiogenesis in several angiogenesis models. We found that excess R50E suppressed FGF1-induced migration and tube formation of endothelial cells, FGF1-induced angiogenesis in matrigel plug assays, and the outgrowth of cells in aorta ring assays. Excess R50E suppressed FGF1-induced angiogenesis in chick embryo chorioallantoic membrane (CAM) assays. Interestingly, excess R50E suppressed FGF2-induced angiogenesis in CAM assays as well, suggesting that R50E may uniquely suppress signaling from other members of the FGF family. Taken together, our results suggest that R50E suppresses angiogenesis induced by FGF1 or FGF2, and thereby indirectly suppresses tumorigenesis, in addition to its possible direct effect on tumor cell proliferation in vivo. We propose that R50E has potential as an anti-cancer and anti-angiogenesis therapeutic agent ("FGF1 decoy").

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R50E suppresses WT FGF1-induced endothelial cell migration.Lower side of the filter in the modified Boyden chamber was coated with fibronectin (10 µg/ml). The lower chamber was filled with serum-free medium with WT FGF1 (5 ng/ml) or the mixture of WT FGF1 and excess R50E (5 and 250 ng/ml, respectively). HUVECs were plated on the filter and incubated for 6 h. Chemotaxed cells were counted from the digital images of the stained cells. Data is shown as means +/− SE per field. Statistical analysis was done by one-way ANOVA plus Tukey analysis.
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pone-0057927-g002: R50E suppresses WT FGF1-induced endothelial cell migration.Lower side of the filter in the modified Boyden chamber was coated with fibronectin (10 µg/ml). The lower chamber was filled with serum-free medium with WT FGF1 (5 ng/ml) or the mixture of WT FGF1 and excess R50E (5 and 250 ng/ml, respectively). HUVECs were plated on the filter and incubated for 6 h. Chemotaxed cells were counted from the digital images of the stained cells. Data is shown as means +/− SE per field. Statistical analysis was done by one-way ANOVA plus Tukey analysis.

Mentions: Endothelial cell migration is a critical feature of tumor angiogenesis. We tested the effect of R50E on migration of HUVECs. Lower side of the filter in the modified Boyden chamber was coated with fibronectin (10 µg/ml). The lower chamber was filled with serum-free EBM-2 medium with WT FGF1 (5 ng/ml) and/or R50E (5 and 250 ng/ml, respectively). HUVECs were plated on the filter and incubated for 6 h, and cells were stained with crystal violet. Chemotaxed cells were counted from the digital images of the stained cells. We found that R50E did not induce cell migration at 5 and 250 ng/ml concentration (Fig. 2). Excess R50E significantly suppressed migration of HUVECs induced by WT FGF1 (Fig. 2). This suggests that R50E acts as an antagonist of FGF1 in migration of HUVECs.


A dominant-negative FGF1 mutant (the R50E mutant) suppresses tumorigenesis and angiogenesis.

Mori S, Tran V, Nishikawa K, Kaneda T, Hamada Y, Kawaguchi N, Fujita M, Saegusa J, Takada YK, Matsuura N, Zhao M, Takada Y - PLoS ONE (2013)

R50E suppresses WT FGF1-induced endothelial cell migration.Lower side of the filter in the modified Boyden chamber was coated with fibronectin (10 µg/ml). The lower chamber was filled with serum-free medium with WT FGF1 (5 ng/ml) or the mixture of WT FGF1 and excess R50E (5 and 250 ng/ml, respectively). HUVECs were plated on the filter and incubated for 6 h. Chemotaxed cells were counted from the digital images of the stained cells. Data is shown as means +/− SE per field. Statistical analysis was done by one-way ANOVA plus Tukey analysis.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3585250&req=5

pone-0057927-g002: R50E suppresses WT FGF1-induced endothelial cell migration.Lower side of the filter in the modified Boyden chamber was coated with fibronectin (10 µg/ml). The lower chamber was filled with serum-free medium with WT FGF1 (5 ng/ml) or the mixture of WT FGF1 and excess R50E (5 and 250 ng/ml, respectively). HUVECs were plated on the filter and incubated for 6 h. Chemotaxed cells were counted from the digital images of the stained cells. Data is shown as means +/− SE per field. Statistical analysis was done by one-way ANOVA plus Tukey analysis.
Mentions: Endothelial cell migration is a critical feature of tumor angiogenesis. We tested the effect of R50E on migration of HUVECs. Lower side of the filter in the modified Boyden chamber was coated with fibronectin (10 µg/ml). The lower chamber was filled with serum-free EBM-2 medium with WT FGF1 (5 ng/ml) and/or R50E (5 and 250 ng/ml, respectively). HUVECs were plated on the filter and incubated for 6 h, and cells were stained with crystal violet. Chemotaxed cells were counted from the digital images of the stained cells. We found that R50E did not induce cell migration at 5 and 250 ng/ml concentration (Fig. 2). Excess R50E significantly suppressed migration of HUVECs induced by WT FGF1 (Fig. 2). This suggests that R50E acts as an antagonist of FGF1 in migration of HUVECs.

Bottom Line: R50E is defective in inducing ternary complex formation, cell proliferation, and cell migration, and suppresses FGF signaling induced by WT FGF1 (a dominant-negative effect) in vitro.Taken together, our results suggest that R50E suppresses angiogenesis induced by FGF1 or FGF2, and thereby indirectly suppresses tumorigenesis, in addition to its possible direct effect on tumor cell proliferation in vivo.We propose that R50E has potential as an anti-cancer and anti-angiogenesis therapeutic agent ("FGF1 decoy").

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular Pathology, Osaka University Graduate School of Medicine, Division of Health Sciences, Suita, Osaka, Japan.

ABSTRACT
Fibroblast growth factor-1 (FGF1) and FGF2 play a critical role in angiogenesis, a formation of new blood vessels from existing blood vessels. Integrins are critically involved in FGF signaling through crosstalk. We previously reported that FGF1 directly binds to integrin αvβ3 and induces FGF receptor-1 (FGFR1)-FGF1-integrin αvβ3 ternary complex. We previously generated an integrin binding defective FGF1 mutant (Arg-50 to Glu, R50E). R50E is defective in inducing ternary complex formation, cell proliferation, and cell migration, and suppresses FGF signaling induced by WT FGF1 (a dominant-negative effect) in vitro. These findings suggest that FGFR and αvβ3 crosstalk through direct integrin binding to FGF, and that R50E acts as an antagonist to FGFR. We studied if R50E suppresses tumorigenesis and angiogenesis. Here we describe that R50E suppressed tumor growth in vivo while WT FGF1 enhanced it using cancer cells that stably express WT FGF1 or R50E. Since R50E did not affect proliferation of cancer cells in vitro, we hypothesized that R50E suppressed tumorigenesis indirectly through suppressing angiogenesis. We thus studied the effect of R50E on angiogenesis in several angiogenesis models. We found that excess R50E suppressed FGF1-induced migration and tube formation of endothelial cells, FGF1-induced angiogenesis in matrigel plug assays, and the outgrowth of cells in aorta ring assays. Excess R50E suppressed FGF1-induced angiogenesis in chick embryo chorioallantoic membrane (CAM) assays. Interestingly, excess R50E suppressed FGF2-induced angiogenesis in CAM assays as well, suggesting that R50E may uniquely suppress signaling from other members of the FGF family. Taken together, our results suggest that R50E suppresses angiogenesis induced by FGF1 or FGF2, and thereby indirectly suppresses tumorigenesis, in addition to its possible direct effect on tumor cell proliferation in vivo. We propose that R50E has potential as an anti-cancer and anti-angiogenesis therapeutic agent ("FGF1 decoy").

Show MeSH
Related in: MedlinePlus