BACKGROUNDSingle nucleotide polymorphisms (SNPs) in the deoxycytidine kinase (dCK) gene are associated with chemosensitivity to nucleoside analogs. 2',2'-Difluoro 2'-deoxycytidine (gemcitabine) is a first-line nucleoside analog drug in the treatment of pancreatic cancer. However, the association between SNPs in the dCK gene and chemosensitivity to gemcitabine has not been fully established. Therefore, the present study aimed to investigate the relationship between SNPs in the dCK gene and chemosensitivity to gemcitabine in human pancreatic cancer cell lines.

METHODSSeven SNPs in the dCK gene were sequenced in six human pancreatic cancer cell lines. The chemosensitivity of these six cell lines to gemcitabine were evaluated in vitro with a Cell Counting Kit-8 (CCK-8) test. Inhibition rates were used to express the chemosensitivity of pancreatic cancer cell lines to gemcitabine.

RESULTSThe genotype of the A9846G SNP in the dCK gene was determined in six human pancreatic cancer cell lines. The cell lines BxPC-3 and T3M4 carried the A9846G SNP genotype AG, whereas cell lines AsPC-1, Mia PaCa2, SW1990 and SU86.86 carried the GG genotype. Cell lines with the AG genotype (BxPC-3 and T3M4) were more sensitive to gemcitabine compared with cell lines with the GG genotype (AsPC-1, Mia PaCa2, SW1990 and SU86.86) and significantly different inhibition rates were observed between cell lines carrying the AG and GG genotypes (P < 0.01).

CONCLUSIONSVariants in the A9846G SNP of the dCK gene were associated with sensitivity to gemcitabine in pancreatic cancer cell lines. The dCK A9846G SNP may act as a genetic marker to predict chemotherapy efficacy of gemcitabine in pancreatic cancer.

Antimetabolites, Antineoplastic ; pharmacology ; Cell Line, Tumor ; Cell Survival ; drug effects ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Deoxycytidine Kinase ; genetics ; Genotype ; Humans ; Pancreatic Neoplasms ; enzymology ; genetics ; Polymorphism, Single Nucleotide ; genetics

OBJECTIVETo explore the effect of emodin combined gemcitabine (E&G) on human pancreatic cancer cell line BxPC-3 in vitro.

METHODSBxPC-3 cells were treated with emodin alone in different concentrations (0, 10, 20, 40, 80, and 160 micromol/L, respectively) for 24, 48, and 72 h, and E&G (emodin 40 micromol/L + gemcitabine 20 micromol/L) for 72 h. The inhibition on BxPC-3 cell proliferation was detected by Cell Counting Kit-8 assay and the cell apoptosis of BxPC-3 was determined using flow cytometry.

RESULTSEmodin obviously suppressed the proliferation of BxPC-3 cells in a dose- and time-dependent manner. The survival rates of BxPC-3 cells by 40 micromol/L emodin for 24, 48, and 72 h were 79. 39%, 46. 35%, and 45. 44%, respectively, while the survival rate of BxPC-3 cells acted by 72-h E&G was only 26. 62%, showing significant difference from that by gemcitabine alone (42.78%) and the emodin alone (47.18%). The early apoptotic ratio of BxPC-3 cells induced by 24 h emodin (40 micromol/L) and gemcitabine (20 micromol/L) were 4.70% +/- 1.54% and 11.20% +/- 1.41% respectively, while early apoptotic ratio of BxPC-3 cells induced by E&G was 20.60% +/-3.23%, showing significant difference from that induced by emodin or gemcitabine alone (P<0.05).

CONCLUSIONSEmodin could significantly inhibit BxPC-3 cell growth. It could act synergistically with gemcitabine to inhibit the tumor proliferation of BxPC-3 cells. Its synergistic action was achieved mainly through inducing pancreatic cancer cell apoptosis.

Apoptosis ; drug effects ; Cell Line, Tumor ; Cell Proliferation ; drug effects ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Emodin ; pharmacology ; Humans ; Pancreatic Neoplasms ; pathology

This study was aimed to investigate the effect of methylation transferase inhibitor 5-aza-2'-deoxycytidine (5-aza-2dC) of different concentrations on the apoptosis of human acute myeloid leukemia (AML) cell line HL-60 and the expression of DAPK gene in HL-60 cells, as well as to explore the possible anti-AML mechanism of 5-aza-2dC. HL-60 cells were treated by 5-aza-2dC of different concentrations. The effect of 5-aza-2dC on the HL-60 cell morphology was observed by Wright's staining. The effect of 5-aza-2dC on HL-60 cell apoptosis and DAPK mRNA expression was detected by flow cytometry and reverse transcription-polymerize chain reaction (RT-PCR) respectively. The results showed that the 5-aza-2dC induced the apoptosis of HL-60 cells in a concentration-dependent manner; the 5-aza-2dC increased the expression levels of DAPK mRNA in HL-60 cells in a concentration-dependent manner. It is concluded that the apoptosis rate of HL-60 cells and DAPK mRNA expression level displayed a rising trend with 5-aza-2dC concentration increasing. Therefore, DAPK gene may participate in HL-60 cell apoptosis induced by 5-aza-2dC.
Apoptosis ; drug effects ; Death-Associated Protein Kinases ; genetics ; Deoxycytidine ; pharmacology ; Gene Expression ; drug effects ; HL-60 Cells ; Humans

OBJECTIVE: To investigate the apoptosis of human breast cell line MCF-7 cells induced by gemcitabine and radiation. METHODS: The MTT method was applied to study the growth inhibition of MCF-7 treated with gemcitabine, radiation, gemcitabine and radiation. The apoptosis index (AI) was analyzed by flow cytometry. The morphology of the MCF-7 cells apoptosis was observed by transmission electron microscopy. RESULTS: When MCF-7 cells were treated with gemcitabine at different concentrations for 24 h, the cell growth inhibition rate was increased in a concentration-dependent manner. The apoptotic indexes (AI) of MCF-7 of four groups by flow cytometry revealed. The AI of (R+D) group was significantly different from those of the radiation group and the gemcitabine group (P<0.05). Condensed chromation, nuclear fragmentation and apoptotic body of MCF-7 cells were found by transmission electron microscope. CONCLUSION The apoptosis of human breast cancer cell line, MCF-7 cells, could be induced by gemcitabine. Gemcitabine can significantly enhance the radiation-induced apoptosis of MCF-7 cells.
Antimetabolites, Antineoplastic ; pharmacology ; Apoptosis ; drug effects ; radiation effects ; Breast Neoplasms ; pathology ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Female ; Flow Cytometry ; Humans ; Radiation ; Radiation-Sensitizing Agents ; pharmacology ; Tumor Cells, Cultured

PURPOSE: This study was aimed to investigate the effect of pseudolaric acid B (PAB) on proliferation, invasion and epithelial-to-mesenchymal transition (EMT) in pancreatic cancer cells and to explore the possible mechanism. MATERIALS AND METHODS: The pancreatic cancer cell line SW1990 was cultured and treated with PAB dose- and time-dependent manners. Cell proliferation and invasion ability were measured by MTT assay and Matrigel/Transwell test, respectively. Semi-quantitative real-time polymerase chain reaction and Western blotting were conducted to detect the expression of EMT markers and the key molecules. Finally, nude mice subcutaneous transplantation tumor model was used to confirm the therapy efficacy of PAB. RESULTS: PAB could inhibit SW1990 cell proliferation and invasion in time- and dose-dependent manners. Vimentin, fibronectin, N-cadherin, Snail, Slug, YAP, TEAD1, and Survivin were down-regulated (p < 0.01), while E-cadherin, caspase-9, MST1, and pYAP were up-regulated (p < 0.05). Combined PAB and gemcitabine treatment markedly restricted the tumor growth compared with gencitabin or PAB alone groups. CONCLUSION: PAB could inhibit the proliferation and invasion ability of pancreatic cancer cells through activating Hippo-YAP pathway and inhibiting the process of EMT.
Animals ; Antineoplastic Agents/pharmacology ; Antineoplastic Agents/therapeutic use ; Biomarkers, Tumor/metabolism ; Cadherins ; Cell Line, Tumor ; Cell Movement ; Cell Proliferation/drug effects ; Cytokines ; Deoxycytidine/analogs & derivatives ; Deoxycytidine/pharmacology ; Deoxycytidine/therapeutic use ; Diterpenes/pharmacology ; Diterpenes/therapeutic use ; Epithelial-Mesenchymal Transition/drug effects ; Female ; Humans ; Mice, Nude ; Neoplasm Invasiveness ; Pancreatic Neoplasms/diet therapy ; Pancreatic Neoplasms/pathology ; Real-Time Polymerase Chain Reaction ; Signal Transduction/drug effects ; Vimentin/metabolism

OBJECTIVETo establish gemcitabine-resistant pancreatic cancer cell strain and study the role of thioredoxin reductase (TrxR) in drug-resistant process.

METHODSGemcitabine-resistant pancreatic cancer cell strain SW1990/GZ was induced by increasing drug dosage intermittently, then the changes of its biological features and the activity of TrxR were examined.

RESULTSStable drug-resistant SW1990/GZ cell strain was established by culturing with gemcitabine for 9 months. The morphology and growth characteristics of the cell strain changed remarkably. The cells shrunk and became rounder; its endoplasm expanded; granular substances increased; and the doubling-time was prolonged. Resistance of the cell line to gemcitabine, fluorouracil, adriamycin, and mitomycin significantly increased. The TrxR activity of the drug-resistant cells was increased markedly.

CONCLUSIONSW1990/GZ has certain multidrug resistance to some chemotherapy drugs, and TrxR plays a role in the drug-resistant process.

Antimetabolites, Antineoplastic ; pharmacology ; Cell Line, Tumor ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Drug Resistance, Neoplasm ; drug effects ; Humans ; Pancreatic Neoplasms ; enzymology ; pathology ; Thioredoxin-Disulfide Reductase ; drug effects ; metabolism

OBJECTIVETo evaluate the effect of gemcitabine in enhancing the radiosensitivity of hepatoma cell line HepG2 and explore its mechanisms.

METHODSClonogenic survival assay is employed to calculate the ratios of L-Q model radiation biology parameters and radiosensitization after different doses of irradiation. Flow cytometry was used to detect the changes in HepG2 cell cycle and apoptosis rate after gemcitabine treatment and radiation exposure.

RESULTSThe survival fraction at 2 Gy of HepG2 cells treated with gemcitabine was significantly lower, and the value of alpha was significantly higher than those of untreated cells. GEM treatment increased the percentage of radiation-induced G0/G1 phase cells and cell apoptosis.

CONCLUSIONGemcitabine can significantly enhance the radiosensitivity of HepG2 cells by enhancing radiation-induced cell cycle arrest in G0/G1 phase and cell apoptosis.

Apoptosis ; drug effects ; Cell Cycle Checkpoints ; drug effects ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Hep G2 Cells ; Humans ; Radiation Tolerance ; drug effects ; Radiation-Sensitizing Agents ; pharmacology

OBJECTIVETo investigate the relationship between the expression of thymidine phosphorylase (TP) and the sensitivity of gastric carcinoma to 5-fluorouracil (5-FU) and its prodrugs.

METHODSGastric carcinoma cell line AGS was transfected with recombinant plasmid pEGFP-N1-TP or control plasmid pEGFP-N1 by lipofectamin 2000. The expression of green fluorescence labeled protein was observed under fluorescence microscope. Positive clones AGS-p and AGS-pTP were selected by G418 treatment. Expression of TP protein and mRNA was detected by immunocytochemistry and RT-PCR, respectively. Drug sensitivity to 5-FU and its prodrugs was assessed by MTT assay.

RESULTSCell clones with the expression of green fluorescent protein (AGS-p) and a clone with TP and green fluorescent fusion protein (AGS-pTP) were established. Immunostaining of TP protein was strongly positive in AGS-pTP and negative in AGS-p and AGS. The expression of TP mRNA was significantly higher in AGS-pTP (0.8090 ± 0.0450) than that in AGS (0.0490 ± 0.0046) and AGS-p (0.0610 ± 0.0069; P < 0.01). The sensitivity to doxifluridine and capecitabine in AGS-pTP was significantly increased, as compared with that in AGS-p. IC50 values of AGS-pTP to doxifluridine and capecitabine were estimated 1.7 folds and 2.2 folds as much as that of AGS-p, respectively. The sensitivity to 5-FU was not different between AGS-pTP and AGS-p.

CONCLUSIONSEnhancement of TP expression improves the sensitivity of gastric carcinoma cells to doxifluridine and capecitabine. But it does not affect the sensitivity to 5-FU.

Antimetabolites, Antineoplastic ; pharmacology ; Capecitabine ; Cell Line, Tumor ; drug effects ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Floxuridine ; pharmacology ; Fluorouracil ; analogs & derivatives ; pharmacology ; Humans ; Plasmids ; RNA, Messenger ; metabolism ; Recombinant Proteins ; genetics ; metabolism ; Sensitivity and Specificity ; Stomach Neoplasms ; metabolism ; pathology ; Thymidine Phosphorylase ; biosynthesis ; genetics ; Transfection

OBJECTIVETo observe the synergistic effects of paclitaxel and gemcitabine on prostate cancer cell line PC-3 in vitro.

METHODSCell morphology, MTU, flow cytometer and immunocytochemical method were used to observe the effects of 10(-6), 10(-7), 10(-8) mol/L paclitaxel and 10(-7), 10(-8), 10(-9) mol/L gemcitabine on prostate cancer cell line PC-3 by single or synergistic administration for 48 hours in vitro.

RESULTSGemcitabine above 10(-8) mol/L enhanced the growth suppression [suppression ratio > or = (50.8 +/- 4.2)%, P < 0.05] and apoptosis [apoptosis ratio > or = (22.9 +/- 2.3)%, P < 0.05] and down-regulation of the expression of cyclin D1 [expression ratio < or = (9.6 +/- 1.6)%, P < 0.01] induced by paclitaxel above 10(-7) mol/L in PC-3 cells. Gemcitabine changed the ratio of G2/M cell arrest induced by paclitaxel from (70.3 +/- 9.7)% to (38.2 +/- 4.2)%, and reversed the G2/M arrest partially (P < 0.01).

CONCLUSIONPaclitaxel and gemcitabine can enhance the growth suppression and apoptosis induced by paclitaxel in a synergistic way. They show great potential in the treatment of androgen-independent carcinoma of the prostate.

Antimetabolites, Antineoplastic ; pharmacology ; Antineoplastic Agents, Phytogenic ; pharmacology ; Apoptosis ; drug effects ; Cell Line, Tumor ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Dose-Response Relationship, Drug ; Down-Regulation ; Drug Synergism ; Flow Cytometry ; Humans ; Male ; Paclitaxel ; pharmacology ; Prostatic Neoplasms ; pathology

OBJECTIVETo investigate the impact of the drug resistance on the radioresistance in human pancreatic cancer cell lines.

METHODSThree drug resistant pancreatic cancer cell sublines induced by fluorouracil (5-FU), adriamycin (ADM) and gemcitabine respectively, SW1990/FU, SW1990/ADM and SW1990/Gz, were tested for the cell cycle and radio-sensitivity with flow cytometry and clonogenic assay.

RESULTSCompared with SW1990, the cell cycle assay indicated higher G(0)/G(1) period percentage in SW1990/FU and SW1990/Gz, but the G(2)/M period percentage decreased; SW1990/FU had the same while SW1990/Gz had lower S period percentage. SW1990/ADM almost had a similar cell cycle with SW1990. Clonogenic assay showed both SW1990/FU and SW1990/Gz had greater survival fraction (SF(2)) than SW1990, but SW1990/ADM had seemingly similar SF(2) as SW1990.

CONCLUSIONDrug resistant pancreatic cancer cell lines have reduced G(2)/M period percentage and increased radioresistance.

Antineoplastic Agents ; pharmacology ; Cell Cycle ; drug effects ; Cell Line, Tumor ; Deoxycytidine ; analogs & derivatives ; pharmacology ; Doxorubicin ; pharmacology ; Drug Resistance, Neoplasm ; drug effects ; genetics ; Fluorouracil ; pharmacology ; Humans ; Pancreatic Neoplasms ; pathology ; Radiation Tolerance ; drug effects