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Antitumor effects and molecular mechanisms of figitumumab, a humanized monoclonal antibody to IGF-1 receptor, in esophageal carcinoma.

Zhang T, Shen H, Dong W, Qu X, Liu Q, Du J - Sci Rep (2014)

Bottom Line: Our results showed that CP not only inhibited IGF-1 induced receptor autophosphorylation and downstream signaling, but also triggered β-arrestin1 and G protein-coupled receptor kinases (GRKs) mediated ERK1/2 activation, indicating CP as a biased agonist for IGF-1R.Inhibition of ERK1/2 enhanced the antitumor activity of CP.Furthermore, CP was a more powerful agonist for IGF-1R down-regulation than IGF-1, and dysregulation of β-arrestin1 and GRKs affected this down-regulation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Oncology, Shandong Provincial Hospital Affiliated to Shandong University, Shandong University, Jinan, P.R. China.

ABSTRACT
The insulin-like growth factor type 1 receptor (IGF-1R) plays an essential role in the development of numerous cancers. Figitumumab (CP) is not only a monoclonal antibody, it also has agonist activity on IGF-1R. The antitumor activity of CP in esophageal squamous cell carcinoma (ESCC) is still unclear. In our study, we identified IGF-1R as an independent prognostic factor in ESCC patients, and investigated the antitumor effects of CP in ESCC cell lines. CP suppressed tumor growth and sensitized cells to chemotherapeutic drugs. In addition, CP inhibited cell proliferation, migration, colony forming activity and anti-apoptosis induced by IGF-1. Our results showed that CP not only inhibited IGF-1 induced receptor autophosphorylation and downstream signaling, but also triggered β-arrestin1 and G protein-coupled receptor kinases (GRKs) mediated ERK1/2 activation, indicating CP as a biased agonist for IGF-1R. Inhibition of ERK1/2 enhanced the antitumor activity of CP. Furthermore, CP was a more powerful agonist for IGF-1R down-regulation than IGF-1, and dysregulation of β-arrestin1 and GRKs affected this down-regulation. Thus, we demonstrated antitumor activities of CP on ESCC, and as a biased agonist, CP induced ERK1/2 activation and receptor down-regulation required β-arrestin1 and GRKs, suggesting a promising role for targeting IGF-1R in ESCC.

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Related in: MedlinePlus

Antitumor activity of CP on ESCC cell lines.(a) Cells were plated, allowed to adhere and serum starvation for 12 hours, and then treated with indicated concentration of IGF-1 for 48 h. Number of viable cells are displayed as percentage of untreated cells. The SEM are displayed from 3 independent replicates. (b) Cells were handled as above, and then treated with 50 ng/ml IGF-1 and different concentrations of CP for 48 h. Number of cells are displayed as percentage of untreated cells. (c) Cells were plated, allowed to adhere overnight and then treated with indicated concentrations of CP in either SFM or with serum for 96 h. Number of cells are displayed as percentage of untreated cells. (d) Cells were plated, allowed to adhere, and serum starvation for 12 h. Cells were first treated with 1 ng/ml CP for 24 h, and then treated with paclitaxel (PTX), cisplatin (DBP) and 5-fluorouracil (5-FU) for another 24 h. Number of cells are displayed as percentage of untreated cells. (e) Cells (103/plate) were seeded onto 100 mm culture plates and incubated with indicated treatment for 14 days, then cell layer was methanol fixed and stained with 0.05% crystal violet solution containing 25% of methanol. (f) Cells were plated in six-well chambers, grown normally for 24 h and starved overnight. Cells were cut with a cell scraper and five images were captured along the cut surface. Additional images were captured 48 h later. (g) Cells were plated, allowed to adhere and serum starvation for 12 h. After 48 h of indicated treatment, cells were collected with LDS sample buffer and protein samples were analyzed by western blot for PARP and GAPDH. The student's t-test or one-way ANOVA were used for statistical analysis properly. *p<0.05; **p<0.01; ***p<0.001.
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f2: Antitumor activity of CP on ESCC cell lines.(a) Cells were plated, allowed to adhere and serum starvation for 12 hours, and then treated with indicated concentration of IGF-1 for 48 h. Number of viable cells are displayed as percentage of untreated cells. The SEM are displayed from 3 independent replicates. (b) Cells were handled as above, and then treated with 50 ng/ml IGF-1 and different concentrations of CP for 48 h. Number of cells are displayed as percentage of untreated cells. (c) Cells were plated, allowed to adhere overnight and then treated with indicated concentrations of CP in either SFM or with serum for 96 h. Number of cells are displayed as percentage of untreated cells. (d) Cells were plated, allowed to adhere, and serum starvation for 12 h. Cells were first treated with 1 ng/ml CP for 24 h, and then treated with paclitaxel (PTX), cisplatin (DBP) and 5-fluorouracil (5-FU) for another 24 h. Number of cells are displayed as percentage of untreated cells. (e) Cells (103/plate) were seeded onto 100 mm culture plates and incubated with indicated treatment for 14 days, then cell layer was methanol fixed and stained with 0.05% crystal violet solution containing 25% of methanol. (f) Cells were plated in six-well chambers, grown normally for 24 h and starved overnight. Cells were cut with a cell scraper and five images were captured along the cut surface. Additional images were captured 48 h later. (g) Cells were plated, allowed to adhere and serum starvation for 12 h. After 48 h of indicated treatment, cells were collected with LDS sample buffer and protein samples were analyzed by western blot for PARP and GAPDH. The student's t-test or one-way ANOVA were used for statistical analysis properly. *p<0.05; **p<0.01; ***p<0.001.

Mentions: As 77.27% of the patients had expression of IGF-1R whereby the expression of IGF-1R was a strong predictor of patients' overall survival, the IGF-1R could be an interesting target in treating ESCC. Next, we utilized the CP to explore the antitumor effect of IGF-1R inhibition. First, we investigated the proliferation of the two ESCC cell lines, Eca-109 and TE-1, with addition of the IGF-1. Cells were incubated with varying doses of IGF-1 for 48 hours, and the effects on proliferation were measured using the CCK-8 assay. Both of the cell lines were responsive to IGF-1 stimulation, and Eca-109 cells seemed to be more sensitive to IGF-1 stimulation (Figure 2a). Cells were then pretreated with various doses of CP for 1 hour and then stimulated with 50 ng/ml IGF-1. CP was able to completely block the proliferation and growth effect induced by IGF-1 in both of the cell lines (Figure 2b). In addition, CP itself inhibited the cell proliferation of ESCC cells under both serum and serum free conditions (Figure 2c). It is indicated that IGF-1R activation mediates the resistance of chemotherapeutic therapies in various cancers1727. Our results showed that CP increased the sensitivity of chemotherapeutic drugs such as paclitaxel (PTX), cisplatin (DBP) and 5-fluorouracil (5-FU) to both of the ESCC cell lines (Figure 2d).


Antitumor effects and molecular mechanisms of figitumumab, a humanized monoclonal antibody to IGF-1 receptor, in esophageal carcinoma.

Zhang T, Shen H, Dong W, Qu X, Liu Q, Du J - Sci Rep (2014)

Antitumor activity of CP on ESCC cell lines.(a) Cells were plated, allowed to adhere and serum starvation for 12 hours, and then treated with indicated concentration of IGF-1 for 48 h. Number of viable cells are displayed as percentage of untreated cells. The SEM are displayed from 3 independent replicates. (b) Cells were handled as above, and then treated with 50 ng/ml IGF-1 and different concentrations of CP for 48 h. Number of cells are displayed as percentage of untreated cells. (c) Cells were plated, allowed to adhere overnight and then treated with indicated concentrations of CP in either SFM or with serum for 96 h. Number of cells are displayed as percentage of untreated cells. (d) Cells were plated, allowed to adhere, and serum starvation for 12 h. Cells were first treated with 1 ng/ml CP for 24 h, and then treated with paclitaxel (PTX), cisplatin (DBP) and 5-fluorouracil (5-FU) for another 24 h. Number of cells are displayed as percentage of untreated cells. (e) Cells (103/plate) were seeded onto 100 mm culture plates and incubated with indicated treatment for 14 days, then cell layer was methanol fixed and stained with 0.05% crystal violet solution containing 25% of methanol. (f) Cells were plated in six-well chambers, grown normally for 24 h and starved overnight. Cells were cut with a cell scraper and five images were captured along the cut surface. Additional images were captured 48 h later. (g) Cells were plated, allowed to adhere and serum starvation for 12 h. After 48 h of indicated treatment, cells were collected with LDS sample buffer and protein samples were analyzed by western blot for PARP and GAPDH. The student's t-test or one-way ANOVA were used for statistical analysis properly. *p<0.05; **p<0.01; ***p<0.001.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4215295&req=5

f2: Antitumor activity of CP on ESCC cell lines.(a) Cells were plated, allowed to adhere and serum starvation for 12 hours, and then treated with indicated concentration of IGF-1 for 48 h. Number of viable cells are displayed as percentage of untreated cells. The SEM are displayed from 3 independent replicates. (b) Cells were handled as above, and then treated with 50 ng/ml IGF-1 and different concentrations of CP for 48 h. Number of cells are displayed as percentage of untreated cells. (c) Cells were plated, allowed to adhere overnight and then treated with indicated concentrations of CP in either SFM or with serum for 96 h. Number of cells are displayed as percentage of untreated cells. (d) Cells were plated, allowed to adhere, and serum starvation for 12 h. Cells were first treated with 1 ng/ml CP for 24 h, and then treated with paclitaxel (PTX), cisplatin (DBP) and 5-fluorouracil (5-FU) for another 24 h. Number of cells are displayed as percentage of untreated cells. (e) Cells (103/plate) were seeded onto 100 mm culture plates and incubated with indicated treatment for 14 days, then cell layer was methanol fixed and stained with 0.05% crystal violet solution containing 25% of methanol. (f) Cells were plated in six-well chambers, grown normally for 24 h and starved overnight. Cells were cut with a cell scraper and five images were captured along the cut surface. Additional images were captured 48 h later. (g) Cells were plated, allowed to adhere and serum starvation for 12 h. After 48 h of indicated treatment, cells were collected with LDS sample buffer and protein samples were analyzed by western blot for PARP and GAPDH. The student's t-test or one-way ANOVA were used for statistical analysis properly. *p<0.05; **p<0.01; ***p<0.001.
Mentions: As 77.27% of the patients had expression of IGF-1R whereby the expression of IGF-1R was a strong predictor of patients' overall survival, the IGF-1R could be an interesting target in treating ESCC. Next, we utilized the CP to explore the antitumor effect of IGF-1R inhibition. First, we investigated the proliferation of the two ESCC cell lines, Eca-109 and TE-1, with addition of the IGF-1. Cells were incubated with varying doses of IGF-1 for 48 hours, and the effects on proliferation were measured using the CCK-8 assay. Both of the cell lines were responsive to IGF-1 stimulation, and Eca-109 cells seemed to be more sensitive to IGF-1 stimulation (Figure 2a). Cells were then pretreated with various doses of CP for 1 hour and then stimulated with 50 ng/ml IGF-1. CP was able to completely block the proliferation and growth effect induced by IGF-1 in both of the cell lines (Figure 2b). In addition, CP itself inhibited the cell proliferation of ESCC cells under both serum and serum free conditions (Figure 2c). It is indicated that IGF-1R activation mediates the resistance of chemotherapeutic therapies in various cancers1727. Our results showed that CP increased the sensitivity of chemotherapeutic drugs such as paclitaxel (PTX), cisplatin (DBP) and 5-fluorouracil (5-FU) to both of the ESCC cell lines (Figure 2d).

Bottom Line: Our results showed that CP not only inhibited IGF-1 induced receptor autophosphorylation and downstream signaling, but also triggered β-arrestin1 and G protein-coupled receptor kinases (GRKs) mediated ERK1/2 activation, indicating CP as a biased agonist for IGF-1R.Inhibition of ERK1/2 enhanced the antitumor activity of CP.Furthermore, CP was a more powerful agonist for IGF-1R down-regulation than IGF-1, and dysregulation of β-arrestin1 and GRKs affected this down-regulation.

View Article: PubMed Central - PubMed

Affiliation: Institute of Oncology, Shandong Provincial Hospital Affiliated to Shandong University, Shandong University, Jinan, P.R. China.

ABSTRACT
The insulin-like growth factor type 1 receptor (IGF-1R) plays an essential role in the development of numerous cancers. Figitumumab (CP) is not only a monoclonal antibody, it also has agonist activity on IGF-1R. The antitumor activity of CP in esophageal squamous cell carcinoma (ESCC) is still unclear. In our study, we identified IGF-1R as an independent prognostic factor in ESCC patients, and investigated the antitumor effects of CP in ESCC cell lines. CP suppressed tumor growth and sensitized cells to chemotherapeutic drugs. In addition, CP inhibited cell proliferation, migration, colony forming activity and anti-apoptosis induced by IGF-1. Our results showed that CP not only inhibited IGF-1 induced receptor autophosphorylation and downstream signaling, but also triggered β-arrestin1 and G protein-coupled receptor kinases (GRKs) mediated ERK1/2 activation, indicating CP as a biased agonist for IGF-1R. Inhibition of ERK1/2 enhanced the antitumor activity of CP. Furthermore, CP was a more powerful agonist for IGF-1R down-regulation than IGF-1, and dysregulation of β-arrestin1 and GRKs affected this down-regulation. Thus, we demonstrated antitumor activities of CP on ESCC, and as a biased agonist, CP induced ERK1/2 activation and receptor down-regulation required β-arrestin1 and GRKs, suggesting a promising role for targeting IGF-1R in ESCC.

Show MeSH
Related in: MedlinePlus