Limits...
Rapid screening of novel agents for combination therapy in sarcomas.

Cubitt CL, Menth J, Dawson J, Martinez GV, Foroutan P, Morse DL, Bui MM, Letson GD, Sullivan DM, Reed DR - Sarcoma (2013)

Bottom Line: Dose-response curves were analyzed for synergy using methods derived from Chou and Talalay (1984).We found that histone deacetylase inhibitors were synergistic with etoposide, dasatinib, and Akt inhibitors across cell lines.Sorafenib and topotecan demonstrated a mixed response.

View Article: PubMed Central - PubMed

Affiliation: Chemical Biology and Molecular Medicine, H. Lee Moffitt Cancer Center and Research Institute, 12902 Magnolia Drive, Tampa, FL 33612, USA ; Translational Research Lab, H. Lee Moffitt Cancer Center and Research Institute, 12902 Magnolia Drive, Tampa, FL 33612, USA.

ABSTRACT
For patients with sarcoma, metastatic disease remains very difficult to cure, and outcomes remain less than optimal. Treatment options have not largely changed, although some promising gains have been made with single agents in specific subtypes with the use of targeted agents. Here, we developed a system to investigate synergy of combinations of targeted and cytotoxic agents in a panel of sarcoma cell lines. Agents were investigated alone and in combination with varying dose ratios. Dose-response curves were analyzed for synergy using methods derived from Chou and Talalay (1984). A promising combination, dasatinib and triciribine, was explored in a murine model using the A673 cell line, and tumors were evaluated by MRI and histology for therapy effect. We found that histone deacetylase inhibitors were synergistic with etoposide, dasatinib, and Akt inhibitors across cell lines. Sorafenib and topotecan demonstrated a mixed response. Our systematic drug screening method allowed us to screen a large number of combinations of sarcoma agents. This method can be easily modified to accommodate other cell line models, and confirmatory assays, such as animal experiments, can provide excellent preclinical data to inform clinical trials for these rare malignancies.

No MeSH data available.


Related in: MedlinePlus

Drug combination effects in U2-OS osteosarcoma cells. The CellTiter-Blue assay was used to monitor viability in treated wells relative to untreated wells (% of untreated). Caspase 3/7 activation was measured after 24-hour concurrent treatments. (a) Vorinostat plus etoposide concurrent treatment. Cell viability was measured after 72-hour drug treatment. Vertical axis represents mean viability result for 5 independent experiments (n = 5). Apoptosis panel shows percent caspase 3/7 activation relative to untreated controls (100%). (b) Sorafenib plus topotecan concurrent and sequential drug treatment effects on cell viability. Sequential treatment consisted of a 24-hour pretreatment with sorafenib followed by the addition of topotecan. Cell viability was measured 48 hours after topotecan addition. (c) Dasatinib plus topotecan concurrent treatment. 72-hour cell viability assays were performed at constant drug ratios of 2 : 1 and 1 : 1. Caspase 3/7 activation was assayed after a 24-hour concurrent drug treatment (1 : 1 ratio) as in (a).
© Copyright Policy - open-access
Related In: Results  -  Collection


getmorefigures.php?uid=PMC3824404&req=5

fig2: Drug combination effects in U2-OS osteosarcoma cells. The CellTiter-Blue assay was used to monitor viability in treated wells relative to untreated wells (% of untreated). Caspase 3/7 activation was measured after 24-hour concurrent treatments. (a) Vorinostat plus etoposide concurrent treatment. Cell viability was measured after 72-hour drug treatment. Vertical axis represents mean viability result for 5 independent experiments (n = 5). Apoptosis panel shows percent caspase 3/7 activation relative to untreated controls (100%). (b) Sorafenib plus topotecan concurrent and sequential drug treatment effects on cell viability. Sequential treatment consisted of a 24-hour pretreatment with sorafenib followed by the addition of topotecan. Cell viability was measured 48 hours after topotecan addition. (c) Dasatinib plus topotecan concurrent treatment. 72-hour cell viability assays were performed at constant drug ratios of 2 : 1 and 1 : 1. Caspase 3/7 activation was assayed after a 24-hour concurrent drug treatment (1 : 1 ratio) as in (a).

Mentions: Topoisomerase II inhibitors such as etoposide also demonstrated broad activity. In the pediatric-type cell lines, IC50 results ranged from 0.5 μM in the SK-ES-1 cell line to 6.8 μM in the U2-OS cell line. In the adult-type sarcoma cell lines, IC50 results were lowest in the SK-UT-1 cell line at 2.5 μM and as high as 7.4 μM in HT-1080 cells (Table 2). Cells were also treated continuously with both agents for 72 hours at a constant 2 : 1 vorinostat: etoposide molar ratio. Using this combination, we found that 6 of 9 cell lines showed a synergistic interaction. The CI values for the pediatric cell lines ranged from 0.6 to 1.0 with a mean value of 0.8 (Table 2). The CI values for the adult-type cell lines ranged from 0.2 to 0.7 with a mean value of 0.5. The concurrent treatment of vorinostat and topotecan for 24 hours resulted in more than additive increases in caspase 3/7 activation, indicating that effects on viability are at least partially mediated through apoptosis (Figures 2(a) and 2(b)), as shown in the U2-OS cell line.


Rapid screening of novel agents for combination therapy in sarcomas.

Cubitt CL, Menth J, Dawson J, Martinez GV, Foroutan P, Morse DL, Bui MM, Letson GD, Sullivan DM, Reed DR - Sarcoma (2013)

Drug combination effects in U2-OS osteosarcoma cells. The CellTiter-Blue assay was used to monitor viability in treated wells relative to untreated wells (% of untreated). Caspase 3/7 activation was measured after 24-hour concurrent treatments. (a) Vorinostat plus etoposide concurrent treatment. Cell viability was measured after 72-hour drug treatment. Vertical axis represents mean viability result for 5 independent experiments (n = 5). Apoptosis panel shows percent caspase 3/7 activation relative to untreated controls (100%). (b) Sorafenib plus topotecan concurrent and sequential drug treatment effects on cell viability. Sequential treatment consisted of a 24-hour pretreatment with sorafenib followed by the addition of topotecan. Cell viability was measured 48 hours after topotecan addition. (c) Dasatinib plus topotecan concurrent treatment. 72-hour cell viability assays were performed at constant drug ratios of 2 : 1 and 1 : 1. Caspase 3/7 activation was assayed after a 24-hour concurrent drug treatment (1 : 1 ratio) as in (a).
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3824404&req=5

fig2: Drug combination effects in U2-OS osteosarcoma cells. The CellTiter-Blue assay was used to monitor viability in treated wells relative to untreated wells (% of untreated). Caspase 3/7 activation was measured after 24-hour concurrent treatments. (a) Vorinostat plus etoposide concurrent treatment. Cell viability was measured after 72-hour drug treatment. Vertical axis represents mean viability result for 5 independent experiments (n = 5). Apoptosis panel shows percent caspase 3/7 activation relative to untreated controls (100%). (b) Sorafenib plus topotecan concurrent and sequential drug treatment effects on cell viability. Sequential treatment consisted of a 24-hour pretreatment with sorafenib followed by the addition of topotecan. Cell viability was measured 48 hours after topotecan addition. (c) Dasatinib plus topotecan concurrent treatment. 72-hour cell viability assays were performed at constant drug ratios of 2 : 1 and 1 : 1. Caspase 3/7 activation was assayed after a 24-hour concurrent drug treatment (1 : 1 ratio) as in (a).
Mentions: Topoisomerase II inhibitors such as etoposide also demonstrated broad activity. In the pediatric-type cell lines, IC50 results ranged from 0.5 μM in the SK-ES-1 cell line to 6.8 μM in the U2-OS cell line. In the adult-type sarcoma cell lines, IC50 results were lowest in the SK-UT-1 cell line at 2.5 μM and as high as 7.4 μM in HT-1080 cells (Table 2). Cells were also treated continuously with both agents for 72 hours at a constant 2 : 1 vorinostat: etoposide molar ratio. Using this combination, we found that 6 of 9 cell lines showed a synergistic interaction. The CI values for the pediatric cell lines ranged from 0.6 to 1.0 with a mean value of 0.8 (Table 2). The CI values for the adult-type cell lines ranged from 0.2 to 0.7 with a mean value of 0.5. The concurrent treatment of vorinostat and topotecan for 24 hours resulted in more than additive increases in caspase 3/7 activation, indicating that effects on viability are at least partially mediated through apoptosis (Figures 2(a) and 2(b)), as shown in the U2-OS cell line.

Bottom Line: Dose-response curves were analyzed for synergy using methods derived from Chou and Talalay (1984).We found that histone deacetylase inhibitors were synergistic with etoposide, dasatinib, and Akt inhibitors across cell lines.Sorafenib and topotecan demonstrated a mixed response.

View Article: PubMed Central - PubMed

Affiliation: Chemical Biology and Molecular Medicine, H. Lee Moffitt Cancer Center and Research Institute, 12902 Magnolia Drive, Tampa, FL 33612, USA ; Translational Research Lab, H. Lee Moffitt Cancer Center and Research Institute, 12902 Magnolia Drive, Tampa, FL 33612, USA.

ABSTRACT
For patients with sarcoma, metastatic disease remains very difficult to cure, and outcomes remain less than optimal. Treatment options have not largely changed, although some promising gains have been made with single agents in specific subtypes with the use of targeted agents. Here, we developed a system to investigate synergy of combinations of targeted and cytotoxic agents in a panel of sarcoma cell lines. Agents were investigated alone and in combination with varying dose ratios. Dose-response curves were analyzed for synergy using methods derived from Chou and Talalay (1984). A promising combination, dasatinib and triciribine, was explored in a murine model using the A673 cell line, and tumors were evaluated by MRI and histology for therapy effect. We found that histone deacetylase inhibitors were synergistic with etoposide, dasatinib, and Akt inhibitors across cell lines. Sorafenib and topotecan demonstrated a mixed response. Our systematic drug screening method allowed us to screen a large number of combinations of sarcoma agents. This method can be easily modified to accommodate other cell line models, and confirmatory assays, such as animal experiments, can provide excellent preclinical data to inform clinical trials for these rare malignancies.

No MeSH data available.


Related in: MedlinePlus