Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 3. Effects of gefitinib and GGTI‑298 on the EGFR signaling pathway and cyclin D1. Protein levels for the EGFR signaling pathway and cyclin D1 were measured by immunoblotting analysis. HCC827 and A549 cells were treated with DMSO, gefitinib (HCC827, 20 nM; A549, 5 µM), 5 µM GGTI‑298 or a combination of gefitinib and GGTI‑298 for 48 h. Representative results are shown (n=3 biological replicates). GGTI/GGTI‑298, geranylgeranyl transferase 1 inhibitor; Gef, gefitinib; DMSO, dimethylsulfoxide; p‑, phos phorylated; EGFR, epidermal growth factor receptor; AKT, protein kinase B; ERK, extracellular signal‑regulated kinase.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Western Blot

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 4. GGTI‑298 inhibits the EGFR‑AKT signaling pathway through RhoA. (A) GGTI‑298 inhibited the phosphorylation of AKT and EGFR induced by EGF. A549 cells were treated with 5 µM GGTI‑298 in serum‑free medium for 48 h. Prior to protein extraction, cultures were exposed to 30 ng EGF for 30 min, as indicated in the figure. (B) PTEN levels were measured by immunoblotting analysis. A549 cells were treated with DMSO, 5 µM gefitinib, 5 µM GGTI‑298 or a combination of gefitinib and GGTI‑298 for 48 h. GAPDH was used as an internal control. (C) Activity of RhoA was measured in A549 cells treated with DMSO and GGTI‑298 for 48 h. (D) A549 cells were transfected with control siRNA or siRhoA, and the activity of RhoA and the phosphorylation of EGFR were measured 72 h following transfection. GST was used as a control in the pull‑down assays. Representative results are shown (n=3 biological replicates). GGTI/GGTI‑298, geranylgeranyl transferase 1 inhibitor; Gef, gefitinib; DMSO, dimethylsulfoxide; p‑, phosphorylated; EGFR, epidermal growth factor receptor; AKT, protein kinase B; PTEN, phosphatase and tensin homolog; RhoA, Ras homolog family member A; si‑/siRNA, small interfering RNA; GST, glutathione S‑transferase.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Phospho-proteomics, Protein Extraction, Western Blot, Control, Activity Assay, Transfection, Small Interfering RNA

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 5. Schematic representation of the effect of the combined treatment of gefitinib and GGTI‑298 on the inhibition of EGFR signaling. In vitro combination treatments of gefitinib and GGTI‑298 inhibited the EGFR signaling pathway more thoroughly than either agent alone, resulting in the inhibition of cell proliferation and migration, and the promotion of apop tosis. GGTI‑298, geranylgeranyl transferase 1 inhibitor; RhoA, Ras homolog family member A; EGFR, epidermal growth factor receptor; AKT, protein kinase B.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Inhibition, In Vitro, Migration

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 4. GGTI‑298 inhibits the EGFR‑AKT signaling pathway through RhoA. (A) GGTI‑298 inhibited the phosphorylation of AKT and EGFR induced by EGF. A549 cells were treated with 5 µM GGTI‑298 in serum‑free medium for 48 h. Prior to protein extraction, cultures were exposed to 30 ng EGF for 30 min, as indicated in the figure. (B) PTEN levels were measured by immunoblotting analysis. A549 cells were treated with DMSO, 5 µM gefitinib, 5 µM GGTI‑298 or a combination of gefitinib and GGTI‑298 for 48 h. GAPDH was used as an internal control. (C) Activity of RhoA was measured in A549 cells treated with DMSO and GGTI‑298 for 48 h. (D) A549 cells were transfected with control siRNA or siRhoA, and the activity of RhoA and the phosphorylation of EGFR were measured 72 h following transfection. GST was used as a control in the pull‑down assays. Representative results are shown (n=3 biological replicates). GGTI/GGTI‑298, geranylgeranyl transferase 1 inhibitor; Gef, gefitinib; DMSO, dimethylsulfoxide; p‑, phosphorylated; EGFR, epidermal growth factor receptor; AKT, protein kinase B; PTEN, phosphatase and tensin homolog; RhoA, Ras homolog family member A; si‑/siRNA, small interfering RNA; GST, glutathione S‑transferase.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Phospho-proteomics, Protein Extraction, Western Blot, Control, Activity Assay, Transfection, Small Interfering RNA

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 5. Schematic representation of the effect of the combined treatment of gefitinib and GGTI‑298 on the inhibition of EGFR signaling. In vitro combination treatments of gefitinib and GGTI‑298 inhibited the EGFR signaling pathway more thoroughly than either agent alone, resulting in the inhibition of cell proliferation and migration, and the promotion of apop tosis. GGTI‑298, geranylgeranyl transferase 1 inhibitor; RhoA, Ras homolog family member A; EGFR, epidermal growth factor receptor; AKT, protein kinase B.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Inhibition, In Vitro, Migration

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 1. Synergic antiproliferation effect of gefitinib and GGTI‑298 in NSCLC cell lines. (A) The relative protein expression of HMGCR compared with GAPDH in each of the five NSCLC cell lines following immunoblotting analysis. GAPDH was used as an internal control; the relative amount of HMGCR was calculated as, HMGCR expression/GAPDH expression in each cell line, with the relative HMGCR expression of H1299 cells set as 1. Cell prolif eration was measured by Cell Counting Kit‑8 in (B) HCC827 and (C) A549 cells treated with gefitinib (HCC827, 10 nM; A549, 5 µM) and/or GGTI‑298 (concentration indicated) for 48 h. The results were presented as the mean ± standard error of the mean (n=3 biological replicates). **P<0.01, as indicated. HMGCR, 3‑hydroxy‑3‑methylglutaryl‑co‑enzyme A reductase; GGTI‑298, geranylgeranyl transferase 1 inhibitor; NSCLC, non‑small cell lung cancer; DMSO, dimethylsulfoxide.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Expressing, Western Blot, Control, CCK-8 Assay, Concentration Assay

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 4. GGTI‑298 inhibits the EGFR‑AKT signaling pathway through RhoA. (A) GGTI‑298 inhibited the phosphorylation of AKT and EGFR induced by EGF. A549 cells were treated with 5 µM GGTI‑298 in serum‑free medium for 48 h. Prior to protein extraction, cultures were exposed to 30 ng EGF for 30 min, as indicated in the figure. (B) PTEN levels were measured by immunoblotting analysis. A549 cells were treated with DMSO, 5 µM gefitinib, 5 µM GGTI‑298 or a combination of gefitinib and GGTI‑298 for 48 h. GAPDH was used as an internal control. (C) Activity of RhoA was measured in A549 cells treated with DMSO and GGTI‑298 for 48 h. (D) A549 cells were transfected with control siRNA or siRhoA, and the activity of RhoA and the phosphorylation of EGFR were measured 72 h following transfection. GST was used as a control in the pull‑down assays. Representative results are shown (n=3 biological replicates). GGTI/GGTI‑298, geranylgeranyl transferase 1 inhibitor; Gef, gefitinib; DMSO, dimethylsulfoxide; p‑, phosphorylated; EGFR, epidermal growth factor receptor; AKT, protein kinase B; PTEN, phosphatase and tensin homolog; RhoA, Ras homolog family member A; si‑/siRNA, small interfering RNA; GST, glutathione S‑transferase.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Phospho-proteomics, Protein Extraction, Western Blot, Control, Activity Assay, Transfection, Small Interfering RNA

Journal: Molecular medicine reports

Article Title: Geranylgeranyl transferase 1 inhibitor GGTI‑298 enhances the anticancer effect of gefitinib.

doi: 10.3892/mmr.2018.9371

Figure Lengend Snippet: Figure 3. Effects of gefitinib and GGTI‑298 on the EGFR signaling pathway and cyclin D1. Protein levels for the EGFR signaling pathway and cyclin D1 were measured by immunoblotting analysis. HCC827 and A549 cells were treated with DMSO, gefitinib (HCC827, 20 nM; A549, 5 µM), 5 µM GGTI‑298 or a combination of gefitinib and GGTI‑298 for 48 h. Representative results are shown (n=3 biological replicates). GGTI/GGTI‑298, geranylgeranyl transferase 1 inhibitor; Gef, gefitinib; DMSO, dimethylsulfoxide; p‑, phos phorylated; EGFR, epidermal growth factor receptor; AKT, protein kinase B; ERK, extracellular signal‑regulated kinase.

Article Snippet: The membranes were incubated with the following primary antibodies: Phosphorylated (p)-EGFR (cat. no. 3777), cyclin D1 (cat. no. 2922), phosphatase and tensin homolog (PTEN; cat. no. 9552), GAPDH (cat. no. 97166), glutathione S-transferase (GST; all Cell Signaling Technology, MA, USA), p-extracellular signal-regulated kinase (ERK; cat. no. EP197Y), ERK (cat. no. ab 17942), p-protein kinase B (AKT; cat. no. EP2109Y), AKT (cat. no. EPR16798; all Abcam, Cambridge, UK), EGFR (cat. no. sc-03-G), 3-hydroxy-3-methylglutaryl-co-enzyme A reductase (HMGCR; cat. no. sc-271595) and RhoA (cat. no. sc-418; all Santa Cruz Biotechnology, Inc., Dallas, TX, USA).

Techniques: Western Blot