Limits...
Rigosertib induces cell death of a myelodysplastic syndrome-derived cell line by DNA damage-induced G2/M arrest.

Hyoda T, Tsujioka T, Nakahara T, Suemori S, Okamoto S, Kataoka M, Tohyama K - Cancer Sci. (2015)

Bottom Line: We also found that rigosertib exerted growth inhibitory effects on two lymphoid cell lines, Jurkat and Ramos.We further examined the molecular pathways influenced by rigosertib from the gene expression profiling data of MDS-L cells and found a possible involvement of rigosertib treatment in the upregulation of the genes related to microtubule kinetics and the downregulation of the mRNA degradation system.The gene set enrichment analysis showed the suppression of "nonsense-mediated mRNA decay (NMD)" as the most significantly affected gene set.

View Article: PubMed Central - PubMed

Affiliation: Division of Medical Technology, Kawasaki College of Allied Health Professions, Okayama, Japan; Department of Medical Technology, Graduate School of Health Sciences, Okayama University, Okayama, Japan; Department of Laboratory Medicine, Kawasaki Medical School, Okayama, Japan.

Show MeSH

Related in: MedlinePlus

Rigosertib suppresses the growth of myeloid and lymphoid cells mainly by apoptosis. (a) HL-60, MDS-L, Jurkat and Ramos cells were treated with rigosertib (0–5000 nM) for indicated times and cell count was assessed by MTT assay. The value without rigosertib was adjusted to 100%. The data represent the mean values with SD from five independent experiments. (b) HL-60, MDS-L, Jurkat and Ramos cells were treated with different concentrations of rigosertib (0, 50, 100 and 200 nM) for 48 h and apoptosis was assessed by flow cytometry using annevin V/propiodium iodide (PI) staining. The single-positive fraction for annexin V implies early apoptosis, and the double-positive fraction for annexin V/PI implies late apoptosis. The values of the lower right area and the upper right area indicate the percentage of the cells in early apoptosis and late apoptosis, respectively. (c) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of Akt and phospho-Akt with each antibody. The amount of alpha-tubulin was shown as a loading control. (d) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of cleaved PARP. The amount of alpha-tubulin was shown as a loading control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4376437&req=5

fig01: Rigosertib suppresses the growth of myeloid and lymphoid cells mainly by apoptosis. (a) HL-60, MDS-L, Jurkat and Ramos cells were treated with rigosertib (0–5000 nM) for indicated times and cell count was assessed by MTT assay. The value without rigosertib was adjusted to 100%. The data represent the mean values with SD from five independent experiments. (b) HL-60, MDS-L, Jurkat and Ramos cells were treated with different concentrations of rigosertib (0, 50, 100 and 200 nM) for 48 h and apoptosis was assessed by flow cytometry using annevin V/propiodium iodide (PI) staining. The single-positive fraction for annexin V implies early apoptosis, and the double-positive fraction for annexin V/PI implies late apoptosis. The values of the lower right area and the upper right area indicate the percentage of the cells in early apoptosis and late apoptosis, respectively. (c) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of Akt and phospho-Akt with each antibody. The amount of alpha-tubulin was shown as a loading control. (d) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of cleaved PARP. The amount of alpha-tubulin was shown as a loading control.

Mentions: We added rigosertib into the culture of HL-60, MDS-L, Jurkat and Ramos cells and found that all four cell lines were susceptible to rigosertib and each cell viability was reduced at concentrations <200 nM (Fig.1a and data not shown). We determined dose-response curves and the half-maximal inhibitory concentration (IC50) of rigosertib in HL-60 cells (24 and 48 h), MDS-L cells (24, 48, 72 and 96 h), Jurkat cells (24 and 48 h) and Ramos cells (24 and 48 h) with MTT assay, respectively. The IC50 value of rigosertib for HL-60 cells, Jurkat cells and Ramos cells at 48 h was 58.5 ± 1.8, 95.4 ± 0.6 and 193.1 ± 2.5 nM, whereas that of rigosertib for MDS-L cells at 96 h was 52.1 ± 0.8 nM. Cell counting by trypan blue staining indicated a similar tendency to the result of MTT assay (data not shown).


Rigosertib induces cell death of a myelodysplastic syndrome-derived cell line by DNA damage-induced G2/M arrest.

Hyoda T, Tsujioka T, Nakahara T, Suemori S, Okamoto S, Kataoka M, Tohyama K - Cancer Sci. (2015)

Rigosertib suppresses the growth of myeloid and lymphoid cells mainly by apoptosis. (a) HL-60, MDS-L, Jurkat and Ramos cells were treated with rigosertib (0–5000 nM) for indicated times and cell count was assessed by MTT assay. The value without rigosertib was adjusted to 100%. The data represent the mean values with SD from five independent experiments. (b) HL-60, MDS-L, Jurkat and Ramos cells were treated with different concentrations of rigosertib (0, 50, 100 and 200 nM) for 48 h and apoptosis was assessed by flow cytometry using annevin V/propiodium iodide (PI) staining. The single-positive fraction for annexin V implies early apoptosis, and the double-positive fraction for annexin V/PI implies late apoptosis. The values of the lower right area and the upper right area indicate the percentage of the cells in early apoptosis and late apoptosis, respectively. (c) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of Akt and phospho-Akt with each antibody. The amount of alpha-tubulin was shown as a loading control. (d) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of cleaved PARP. The amount of alpha-tubulin was shown as a loading control.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4376437&req=5

fig01: Rigosertib suppresses the growth of myeloid and lymphoid cells mainly by apoptosis. (a) HL-60, MDS-L, Jurkat and Ramos cells were treated with rigosertib (0–5000 nM) for indicated times and cell count was assessed by MTT assay. The value without rigosertib was adjusted to 100%. The data represent the mean values with SD from five independent experiments. (b) HL-60, MDS-L, Jurkat and Ramos cells were treated with different concentrations of rigosertib (0, 50, 100 and 200 nM) for 48 h and apoptosis was assessed by flow cytometry using annevin V/propiodium iodide (PI) staining. The single-positive fraction for annexin V implies early apoptosis, and the double-positive fraction for annexin V/PI implies late apoptosis. The values of the lower right area and the upper right area indicate the percentage of the cells in early apoptosis and late apoptosis, respectively. (c) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of Akt and phospho-Akt with each antibody. The amount of alpha-tubulin was shown as a loading control. (d) HL-60 and MDS-L cells were treated with indicated concentrations of rigosertib for 24 and 48 h, respectively, and protein lysates were analyzed by immunoblotting analysis for the detection of cleaved PARP. The amount of alpha-tubulin was shown as a loading control.
Mentions: We added rigosertib into the culture of HL-60, MDS-L, Jurkat and Ramos cells and found that all four cell lines were susceptible to rigosertib and each cell viability was reduced at concentrations <200 nM (Fig.1a and data not shown). We determined dose-response curves and the half-maximal inhibitory concentration (IC50) of rigosertib in HL-60 cells (24 and 48 h), MDS-L cells (24, 48, 72 and 96 h), Jurkat cells (24 and 48 h) and Ramos cells (24 and 48 h) with MTT assay, respectively. The IC50 value of rigosertib for HL-60 cells, Jurkat cells and Ramos cells at 48 h was 58.5 ± 1.8, 95.4 ± 0.6 and 193.1 ± 2.5 nM, whereas that of rigosertib for MDS-L cells at 96 h was 52.1 ± 0.8 nM. Cell counting by trypan blue staining indicated a similar tendency to the result of MTT assay (data not shown).

Bottom Line: We also found that rigosertib exerted growth inhibitory effects on two lymphoid cell lines, Jurkat and Ramos.We further examined the molecular pathways influenced by rigosertib from the gene expression profiling data of MDS-L cells and found a possible involvement of rigosertib treatment in the upregulation of the genes related to microtubule kinetics and the downregulation of the mRNA degradation system.The gene set enrichment analysis showed the suppression of "nonsense-mediated mRNA decay (NMD)" as the most significantly affected gene set.

View Article: PubMed Central - PubMed

Affiliation: Division of Medical Technology, Kawasaki College of Allied Health Professions, Okayama, Japan; Department of Medical Technology, Graduate School of Health Sciences, Okayama University, Okayama, Japan; Department of Laboratory Medicine, Kawasaki Medical School, Okayama, Japan.

Show MeSH
Related in: MedlinePlus