We reported that curcumin in combination with ERRP, a pan erbB inhibitor causes a greater inhibition of the development of colon cancer cells that both agent alone 28. We have also reported that curcumin acts synergistically with FOLFOX in inhibiting growth of colon cancer cells in vitro. These and other related observations have prompted us to undertake the recent investigation. Our working hypothesis, as a result, is that a mixture of dasatinib and curcumin will be an efficient therapeutic approach for colorectal neoplasia and/or cancer. We even more hypothesize that this improved usefulness is the end result of an attenuation of numerous signaling pathways foremost to inhibition of transformation properties of colon cancer cells.
Human colon cancer HCT 116 p53 wild PH-797804 type, HT 29, and HCT 116 p53 null and SW 620 cells have been employed to investigate efficacy of combined remedy of dasatinib in and curcumin in development inhibition. HCT 116, HT 29 and SW 620 cells have been obtained from American Type Culture Collection, whereas HCT 116 p53 null cells, initially produced in Dr. Bert Vogelstein laboratory at John Hopkins University, Baltimore, MD, were obtained from Dr Ping Dou at Karmanos Cancer Institute. The cells have been maintained in tissue culture flasks in Dulbeccos modified Eagle medium in a humidified incubator at 37 C in an environment of 95% air and 5% CO2. The cell culture medium was supplemented with 5% FBS and 1% antibiotic/ antimycotic. Human umbilical vein endothelial cells, a kind present from Dr.
Fazlul Sarkar at the Karmanos Cancer Institute, Detroit, MI, have been used for angiogenesis assay. Endothelial growth medium with nutrient dietary supplements have been purchased from Lonza Walkersville Inc.. Furthermore, c-Met Inhibitors the cell culture medium was supplemented with 5% FBS and 1% antibiotic/antimycotic. Medium was altered 3 times a week and cells had been passaged using trypsin/EDTA. Dulbeccos modified Eagle medium, fetal bovine serum, and antibiotic/ antimycotic had been obtained from GIBCO BRL. Dasatinib was ordered from LC laboratories. Protease inhibitor cocktail, 3 2,5 diphenyltetrazolium bromide, and all other chemicals have been obtained from Sigma. Anti p EGFRs, p HER2, p HER3, p Src, Src, p Akt, p Erk, BclXL and Cox 2 p IGF 1R, IGF 1, IGFBP3 and Rb have been ordered from Cell Signaling. Antibodies to B actin antibody was purchased from Sigma.
Chemiluminescence detection of proteins was conducted with the use of a kit from Amersham Biosciences/Amersham Pharmacia Biotech. Recombinant TGF was bought from Oncogene. Inhibition of cell growth in response to dasatinib and or curcumin was examined by 3 2,5 diphenyl tetrazolium bromide assay as described previously 30. Briefly, cells had been dispersed by trypsin EDTA remedy and resuspended in suitable culture medium containing 5% of FBS and 5,000 cells/effectively have been seeded into 96 properly culture plates with six replicates. Following 24 hrs of plating, incubation was ongoing for another 48 h in the absence or presence of distinct drugs as described in the legends to the figures.
Human colon cancer HCT 116 p53 wild PH-797804 type, HT 29, and HCT 116 p53 null and SW 620 cells have been employed to investigate efficacy of combined remedy of dasatinib in and curcumin in development inhibition. HCT 116, HT 29 and SW 620 cells have been obtained from American Type Culture Collection, whereas HCT 116 p53 null cells, initially produced in Dr. Bert Vogelstein laboratory at John Hopkins University, Baltimore, MD, were obtained from Dr Ping Dou at Karmanos Cancer Institute. The cells have been maintained in tissue culture flasks in Dulbeccos modified Eagle medium in a humidified incubator at 37 C in an environment of 95% air and 5% CO2. The cell culture medium was supplemented with 5% FBS and 1% antibiotic/ antimycotic. Human umbilical vein endothelial cells, a kind present from Dr.
Fazlul Sarkar at the Karmanos Cancer Institute, Detroit, MI, have been used for angiogenesis assay. Endothelial growth medium with nutrient dietary supplements have been purchased from Lonza Walkersville Inc.. Furthermore, c-Met Inhibitors the cell culture medium was supplemented with 5% FBS and 1% antibiotic/antimycotic. Medium was altered 3 times a week and cells had been passaged using trypsin/EDTA. Dulbeccos modified Eagle medium, fetal bovine serum, and antibiotic/ antimycotic had been obtained from GIBCO BRL. Dasatinib was ordered from LC laboratories. Protease inhibitor cocktail, 3 2,5 diphenyltetrazolium bromide, and all other chemicals have been obtained from Sigma. Anti p EGFRs, p HER2, p HER3, p Src, Src, p Akt, p Erk, BclXL and Cox 2 p IGF 1R, IGF 1, IGFBP3 and Rb have been ordered from Cell Signaling. Antibodies to B actin antibody was purchased from Sigma.
Chemiluminescence detection of proteins was conducted with the use of a kit from Amersham Biosciences/Amersham Pharmacia Biotech. Recombinant TGF was bought from Oncogene. Inhibition of cell growth in response to dasatinib and or curcumin was examined by 3 2,5 diphenyl tetrazolium bromide assay as described previously 30. Briefly, cells had been dispersed by trypsin EDTA remedy and resuspended in suitable culture medium containing 5% of FBS and 5,000 cells/effectively have been seeded into 96 properly culture plates with six replicates. Following 24 hrs of plating, incubation was ongoing for another 48 h in the absence or presence of distinct drugs as described in the legends to the figures.
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