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Pharmacological targeting of NF-kappaB potentiates the effect of the topoisomerase inhibitor CPT-11 on colon cancer cells.

Lagadec P, Griessinger E, Nawrot MP, Fenouille N, Colosetti P, Imbert V, Mari M, Hofman P, Czerucka D, Rousseau D, Berard E, Dreano M, Peyron JF - Br. J. Cancer (2008)

Bottom Line: NF-kappaB interferes with the effect of most anti-cancer drugs through induction of anti-apoptotic genes.Eighty-five and 75% decreases in tumour size were observed when mice were treated with, respectively, 20 or 5 mg kg(-1) AS602868 associated with 30 mg kg(-1) CPT-11 compared to 47% with CPT-11 alone.This study is the first demonstration that a pharmacological inhibitor of the IKK2 kinase can potentiate the therapeutic efficiency of antineoplasic drugs on solid tumours.

View Article: PubMed Central - PubMed

Affiliation: INSERM U526, Nice F-06000, France. lagadec@unice.fr

ABSTRACT
NF-kappaB interferes with the effect of most anti-cancer drugs through induction of anti-apoptotic genes. Targeting NF-kappaB is therefore expected to potentiate conventional treatments in adjuvant strategies. Here we used a pharmacological inhibitor of the IKK2 kinase (AS602868) to block NF-kappaB activation. In human colon cancer cells, inhibition of NF-kappaB using 10 microM AS602868 induced a 30-50% growth inhibitory effect and strongly enhanced the action of SN-38, the topoisomerase I inhibitor and CPT-11 active metabolite. AS602868 also potentiated the cytotoxic effect of two other antineoplasic drugs: 5-fluorouracil and etoposide. In xenografts experiments, inhibition of NF-kappaB potentiated the antitumoural effect of CPT-11 in a dose-dependent manner. Eighty-five and 75% decreases in tumour size were observed when mice were treated with, respectively, 20 or 5 mg kg(-1) AS602868 associated with 30 mg kg(-1) CPT-11 compared to 47% with CPT-11 alone. Ex vivo tumour analyses as well as in vitro studies showed that AS602868 impaired CPT-11-induced NF-kappaB activation, and enhanced tumour cell cycle arrest and apoptosis. AS602868 also enhanced the apoptotic potential of TNFalpha on HT-29 cells. This study is the first demonstration that a pharmacological inhibitor of the IKK2 kinase can potentiate the therapeutic efficiency of antineoplasic drugs on solid tumours.

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In vitro and in vivo effect of AS602868 combined with CPT-11/SN-38 on apoptosis. (A) Apoptosis was measured by ELISA. HT-29 cells were incubated for 5 days with AS602868±SN-38 at indicated concentrations. Data are expressed as mean OD±s.d. of duplicates of one representative experiment from 4. The positive control is a DNA–histone complex included in the kit. (B) Tumours were removed from mice treated with AS602868 vehicle (control group), with AS602868 (20 mg kg−1) alone or combined with CPT-11 (30 mg kg−1). Then, they were minced, put into liquid nitrogen, and stored at −80°C. Frozen tumour sections (7 μm) were mounted in Fluoromount-G solution and processed following the protocol described in the In situ Cell Death Detection Kit (Roche Diagnostics). Analyses were performed using an LSM 510 confocal laser-scanning microscope with an oil objective × 40. (C) HT-29 cells were treated with AS602868±SN-38±z-VADfmk (50 μM) for 5 days. Cell viability was evaluated using the MTT assay. Data are expressed as means±s.d. of quadruplicates of one representative experiment from 3.
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fig4: In vitro and in vivo effect of AS602868 combined with CPT-11/SN-38 on apoptosis. (A) Apoptosis was measured by ELISA. HT-29 cells were incubated for 5 days with AS602868±SN-38 at indicated concentrations. Data are expressed as mean OD±s.d. of duplicates of one representative experiment from 4. The positive control is a DNA–histone complex included in the kit. (B) Tumours were removed from mice treated with AS602868 vehicle (control group), with AS602868 (20 mg kg−1) alone or combined with CPT-11 (30 mg kg−1). Then, they were minced, put into liquid nitrogen, and stored at −80°C. Frozen tumour sections (7 μm) were mounted in Fluoromount-G solution and processed following the protocol described in the In situ Cell Death Detection Kit (Roche Diagnostics). Analyses were performed using an LSM 510 confocal laser-scanning microscope with an oil objective × 40. (C) HT-29 cells were treated with AS602868±SN-38±z-VADfmk (50 μM) for 5 days. Cell viability was evaluated using the MTT assay. Data are expressed as means±s.d. of quadruplicates of one representative experiment from 3.

Mentions: Quantification by ELISA (Figure 4A) of mono- and oligonucleosomes showed that AS602868 (3 μM) or SN-38 (3 and 10 nM) alone induced HT-29 cell apoptosis. A higher effect was produced by combining the two drugs: 3 μM AS602868 (two-fold medium OD), 3 and 10 nM SN-38 (1.5 and 2-fold medium OD, respectively), compared to (three-fold medium OD) obtained with 3 μM AS602868+3 nM SN-38 and (3.2-fold) with 3 μM AS602868+10 nM SN-38. Similar results were obtained in vivo (Figure 4B). TUNEL experiments revealed nearly no apotosis in tumours from mice of the control group, and an increasing level in tumours from mice treated with AS602868, CPT-11, and with the combined treatment.


Pharmacological targeting of NF-kappaB potentiates the effect of the topoisomerase inhibitor CPT-11 on colon cancer cells.

Lagadec P, Griessinger E, Nawrot MP, Fenouille N, Colosetti P, Imbert V, Mari M, Hofman P, Czerucka D, Rousseau D, Berard E, Dreano M, Peyron JF - Br. J. Cancer (2008)

In vitro and in vivo effect of AS602868 combined with CPT-11/SN-38 on apoptosis. (A) Apoptosis was measured by ELISA. HT-29 cells were incubated for 5 days with AS602868±SN-38 at indicated concentrations. Data are expressed as mean OD±s.d. of duplicates of one representative experiment from 4. The positive control is a DNA–histone complex included in the kit. (B) Tumours were removed from mice treated with AS602868 vehicle (control group), with AS602868 (20 mg kg−1) alone or combined with CPT-11 (30 mg kg−1). Then, they were minced, put into liquid nitrogen, and stored at −80°C. Frozen tumour sections (7 μm) were mounted in Fluoromount-G solution and processed following the protocol described in the In situ Cell Death Detection Kit (Roche Diagnostics). Analyses were performed using an LSM 510 confocal laser-scanning microscope with an oil objective × 40. (C) HT-29 cells were treated with AS602868±SN-38±z-VADfmk (50 μM) for 5 days. Cell viability was evaluated using the MTT assay. Data are expressed as means±s.d. of quadruplicates of one representative experiment from 3.
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Related In: Results  -  Collection

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fig4: In vitro and in vivo effect of AS602868 combined with CPT-11/SN-38 on apoptosis. (A) Apoptosis was measured by ELISA. HT-29 cells were incubated for 5 days with AS602868±SN-38 at indicated concentrations. Data are expressed as mean OD±s.d. of duplicates of one representative experiment from 4. The positive control is a DNA–histone complex included in the kit. (B) Tumours were removed from mice treated with AS602868 vehicle (control group), with AS602868 (20 mg kg−1) alone or combined with CPT-11 (30 mg kg−1). Then, they were minced, put into liquid nitrogen, and stored at −80°C. Frozen tumour sections (7 μm) were mounted in Fluoromount-G solution and processed following the protocol described in the In situ Cell Death Detection Kit (Roche Diagnostics). Analyses were performed using an LSM 510 confocal laser-scanning microscope with an oil objective × 40. (C) HT-29 cells were treated with AS602868±SN-38±z-VADfmk (50 μM) for 5 days. Cell viability was evaluated using the MTT assay. Data are expressed as means±s.d. of quadruplicates of one representative experiment from 3.
Mentions: Quantification by ELISA (Figure 4A) of mono- and oligonucleosomes showed that AS602868 (3 μM) or SN-38 (3 and 10 nM) alone induced HT-29 cell apoptosis. A higher effect was produced by combining the two drugs: 3 μM AS602868 (two-fold medium OD), 3 and 10 nM SN-38 (1.5 and 2-fold medium OD, respectively), compared to (three-fold medium OD) obtained with 3 μM AS602868+3 nM SN-38 and (3.2-fold) with 3 μM AS602868+10 nM SN-38. Similar results were obtained in vivo (Figure 4B). TUNEL experiments revealed nearly no apotosis in tumours from mice of the control group, and an increasing level in tumours from mice treated with AS602868, CPT-11, and with the combined treatment.

Bottom Line: NF-kappaB interferes with the effect of most anti-cancer drugs through induction of anti-apoptotic genes.Eighty-five and 75% decreases in tumour size were observed when mice were treated with, respectively, 20 or 5 mg kg(-1) AS602868 associated with 30 mg kg(-1) CPT-11 compared to 47% with CPT-11 alone.This study is the first demonstration that a pharmacological inhibitor of the IKK2 kinase can potentiate the therapeutic efficiency of antineoplasic drugs on solid tumours.

View Article: PubMed Central - PubMed

Affiliation: INSERM U526, Nice F-06000, France. lagadec@unice.fr

ABSTRACT
NF-kappaB interferes with the effect of most anti-cancer drugs through induction of anti-apoptotic genes. Targeting NF-kappaB is therefore expected to potentiate conventional treatments in adjuvant strategies. Here we used a pharmacological inhibitor of the IKK2 kinase (AS602868) to block NF-kappaB activation. In human colon cancer cells, inhibition of NF-kappaB using 10 microM AS602868 induced a 30-50% growth inhibitory effect and strongly enhanced the action of SN-38, the topoisomerase I inhibitor and CPT-11 active metabolite. AS602868 also potentiated the cytotoxic effect of two other antineoplasic drugs: 5-fluorouracil and etoposide. In xenografts experiments, inhibition of NF-kappaB potentiated the antitumoural effect of CPT-11 in a dose-dependent manner. Eighty-five and 75% decreases in tumour size were observed when mice were treated with, respectively, 20 or 5 mg kg(-1) AS602868 associated with 30 mg kg(-1) CPT-11 compared to 47% with CPT-11 alone. Ex vivo tumour analyses as well as in vitro studies showed that AS602868 impaired CPT-11-induced NF-kappaB activation, and enhanced tumour cell cycle arrest and apoptosis. AS602868 also enhanced the apoptotic potential of TNFalpha on HT-29 cells. This study is the first demonstration that a pharmacological inhibitor of the IKK2 kinase can potentiate the therapeutic efficiency of antineoplasic drugs on solid tumours.

Show MeSH
Related in: MedlinePlus