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Farnesyl diphosphate synthase is involved in the resistance to zoledronic acid of osteosarcoma cells.

Ory B, Moriceau G, Trichet V, Blanchard F, Berreur M, Rédini F, Rogers M, Heymann D - J. Cell. Mol. Med. (2008)

Bottom Line: Furthermore, the Zol-resistant cells were sensitive to conventional anti-cancer agents demonstrating that this resistance process is independent of the multidrug resistance phenotype.The transfection of Zol-resistant cells with FPPS siRNA strongly increased their sensitivity to Zol.This study demonstrates for the first time the induction of metabolic resistance after prolonged Zol treatment of osteosarcoma cells confirming the therapeutic potential of Zol for the treatment of bone malignant pathologies, but points out the importance of the treatment regimen may be important in terms of duration and dose to avoid the development of drug metabolic resistance.

View Article: PubMed Central - PubMed

Affiliation: Institut National de la Santé et de la Recherche Médicale, ERI 7, Nantes, France.

ABSTRACT
We recently demonstrated original anti-tumor effects of zoledronic acid (Zol) on osteosarcoma cell lines independently of their p53 and Rb status. The present study investigated the potential Zol-resistance acquired by osteosarcoma cells after prolonged treatment. After 12 weeks of culture in the presence of 1 microm Zol, the effects of high doses of Zol (10-100 microm) were compared between the untreated rat (OSRGA, ROS) and human (MG63, SAOS2) osteosarcoma cells and Zol-pretreated cells in terms of cell proliferation, cell cycle analysis, migration assay and cytoskeleton organization. Long-term treatment with 1 microm Zol reduced the sensitivity of osteosarcoma cells to high concentrations of Zol. Furthermore, the Zol-resistant cells were sensitive to conventional anti-cancer agents demonstrating that this resistance process is independent of the multidrug resistance phenotype. However, as similar experiments performed in the presence of clodronate and pamidronate evidenced that this drug resistance was restricted to the nitrogen-containing bisphosphonates, we then hypothesized that this resistance could be associated with a differential expression of farnesyl diphos-phate synthase (FPPS) also observed in human osteosarcoma samples. The transfection of Zol-resistant cells with FPPS siRNA strongly increased their sensitivity to Zol. This study demonstrates for the first time the induction of metabolic resistance after prolonged Zol treatment of osteosarcoma cells confirming the therapeutic potential of Zol for the treatment of bone malignant pathologies, but points out the importance of the treatment regimen may be important in terms of duration and dose to avoid the development of drug metabolic resistance.

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Geranylgeraniol (GGO) reverses FPPS siRNA effects in osteosarcoma cell lines. (A) Rat (OSRGA) and human (MG63, SAOS2) osteosarcoma cell lines were transfected with FPPS siRNA and treated 24 hrs after with increasing concentrations of Zol (0.1–100 μm) for 72 hrs in the presence or not of 25 μm GGO. The number of viable cells was then determined using the XTT assay. Graphs represent the mean values of three independent experiments performed in triplicate. Error bars represent the standard deviation.***P < 0.001. Statistical evaluation of the data was performed using the ANOVA test.(B) Western blot analysis of unprenylated RAP1A (unRAP1A) form. Cells transfected with control siRNA or with FPPS siRNA combined with 25 μm GGO were treated with 1 and 10 μm Zol for 24 and 48 hrs. All experiments were repeated three times, and a representative blot is shown.
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fig05: Geranylgeraniol (GGO) reverses FPPS siRNA effects in osteosarcoma cell lines. (A) Rat (OSRGA) and human (MG63, SAOS2) osteosarcoma cell lines were transfected with FPPS siRNA and treated 24 hrs after with increasing concentrations of Zol (0.1–100 μm) for 72 hrs in the presence or not of 25 μm GGO. The number of viable cells was then determined using the XTT assay. Graphs represent the mean values of three independent experiments performed in triplicate. Error bars represent the standard deviation.***P < 0.001. Statistical evaluation of the data was performed using the ANOVA test.(B) Western blot analysis of unprenylated RAP1A (unRAP1A) form. Cells transfected with control siRNA or with FPPS siRNA combined with 25 μm GGO were treated with 1 and 10 μm Zol for 24 and 48 hrs. All experiments were repeated three times, and a representative blot is shown.

Mentions: To determine whether the effects previously demonstrated for the FPPS siRNA in osteosarcoma cells are reversible, FPPS siRNA transfected cells treated with increasing doses of Zol were cultured in the presence of 25 μm geranylgeraniol, the FPPS metabolic product (Fig. 5). GGO protected rat and human osteosarcoma cell lines from the effects of Zol in the FPPS siRNA-transfected cells and totally reversed FPPS siRNA effects (Fig. 5A). We therefore investigated by western blot the expression kinetic of unRAP1A in the presence of 25 μm GGO in FPPS siRNA-transfected cells (Fig. 5B). GGO totally abolished unRAP1A expression similar to what had been observed in Zol-resistant cell lines (Fig. 3B). Overall, these data then strengthen our conclusion that FPPS is involved in the Zol-resistance mechanism.


Farnesyl diphosphate synthase is involved in the resistance to zoledronic acid of osteosarcoma cells.

Ory B, Moriceau G, Trichet V, Blanchard F, Berreur M, Rédini F, Rogers M, Heymann D - J. Cell. Mol. Med. (2008)

Geranylgeraniol (GGO) reverses FPPS siRNA effects in osteosarcoma cell lines. (A) Rat (OSRGA) and human (MG63, SAOS2) osteosarcoma cell lines were transfected with FPPS siRNA and treated 24 hrs after with increasing concentrations of Zol (0.1–100 μm) for 72 hrs in the presence or not of 25 μm GGO. The number of viable cells was then determined using the XTT assay. Graphs represent the mean values of three independent experiments performed in triplicate. Error bars represent the standard deviation.***P < 0.001. Statistical evaluation of the data was performed using the ANOVA test.(B) Western blot analysis of unprenylated RAP1A (unRAP1A) form. Cells transfected with control siRNA or with FPPS siRNA combined with 25 μm GGO were treated with 1 and 10 μm Zol for 24 and 48 hrs. All experiments were repeated three times, and a representative blot is shown.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4401135&req=5

fig05: Geranylgeraniol (GGO) reverses FPPS siRNA effects in osteosarcoma cell lines. (A) Rat (OSRGA) and human (MG63, SAOS2) osteosarcoma cell lines were transfected with FPPS siRNA and treated 24 hrs after with increasing concentrations of Zol (0.1–100 μm) for 72 hrs in the presence or not of 25 μm GGO. The number of viable cells was then determined using the XTT assay. Graphs represent the mean values of three independent experiments performed in triplicate. Error bars represent the standard deviation.***P < 0.001. Statistical evaluation of the data was performed using the ANOVA test.(B) Western blot analysis of unprenylated RAP1A (unRAP1A) form. Cells transfected with control siRNA or with FPPS siRNA combined with 25 μm GGO were treated with 1 and 10 μm Zol for 24 and 48 hrs. All experiments were repeated three times, and a representative blot is shown.
Mentions: To determine whether the effects previously demonstrated for the FPPS siRNA in osteosarcoma cells are reversible, FPPS siRNA transfected cells treated with increasing doses of Zol were cultured in the presence of 25 μm geranylgeraniol, the FPPS metabolic product (Fig. 5). GGO protected rat and human osteosarcoma cell lines from the effects of Zol in the FPPS siRNA-transfected cells and totally reversed FPPS siRNA effects (Fig. 5A). We therefore investigated by western blot the expression kinetic of unRAP1A in the presence of 25 μm GGO in FPPS siRNA-transfected cells (Fig. 5B). GGO totally abolished unRAP1A expression similar to what had been observed in Zol-resistant cell lines (Fig. 3B). Overall, these data then strengthen our conclusion that FPPS is involved in the Zol-resistance mechanism.

Bottom Line: Furthermore, the Zol-resistant cells were sensitive to conventional anti-cancer agents demonstrating that this resistance process is independent of the multidrug resistance phenotype.The transfection of Zol-resistant cells with FPPS siRNA strongly increased their sensitivity to Zol.This study demonstrates for the first time the induction of metabolic resistance after prolonged Zol treatment of osteosarcoma cells confirming the therapeutic potential of Zol for the treatment of bone malignant pathologies, but points out the importance of the treatment regimen may be important in terms of duration and dose to avoid the development of drug metabolic resistance.

View Article: PubMed Central - PubMed

Affiliation: Institut National de la Santé et de la Recherche Médicale, ERI 7, Nantes, France.

ABSTRACT
We recently demonstrated original anti-tumor effects of zoledronic acid (Zol) on osteosarcoma cell lines independently of their p53 and Rb status. The present study investigated the potential Zol-resistance acquired by osteosarcoma cells after prolonged treatment. After 12 weeks of culture in the presence of 1 microm Zol, the effects of high doses of Zol (10-100 microm) were compared between the untreated rat (OSRGA, ROS) and human (MG63, SAOS2) osteosarcoma cells and Zol-pretreated cells in terms of cell proliferation, cell cycle analysis, migration assay and cytoskeleton organization. Long-term treatment with 1 microm Zol reduced the sensitivity of osteosarcoma cells to high concentrations of Zol. Furthermore, the Zol-resistant cells were sensitive to conventional anti-cancer agents demonstrating that this resistance process is independent of the multidrug resistance phenotype. However, as similar experiments performed in the presence of clodronate and pamidronate evidenced that this drug resistance was restricted to the nitrogen-containing bisphosphonates, we then hypothesized that this resistance could be associated with a differential expression of farnesyl diphos-phate synthase (FPPS) also observed in human osteosarcoma samples. The transfection of Zol-resistant cells with FPPS siRNA strongly increased their sensitivity to Zol. This study demonstrates for the first time the induction of metabolic resistance after prolonged Zol treatment of osteosarcoma cells confirming the therapeutic potential of Zol for the treatment of bone malignant pathologies, but points out the importance of the treatment regimen may be important in terms of duration and dose to avoid the development of drug metabolic resistance.

Show MeSH
Related in: MedlinePlus