Upload
dinhtuyen
View
216
Download
1
Embed Size (px)
Citation preview
www.cjcsysu.cn
Chinese Journal of Cancer Chinese Journal of Cancer
Establishment and biological characteristics of oxaliplatinresistant human colon cancer cell lines
Zhen Liu 1 , Meng Qiu 1 , QiuLin Tang 2 , Ming Liu 1 , Nan Lang 2 , Feng Bi 1
1 Department of Medical Oncology ,West China Hospital, Sichuan University, Chengdu, Sichuan 610041, P. R. China; 2 The Laboratory of Signal Transduction & Molecular Targeting Therapy, State Key Laboratory of Biotherapy, West China Hospital,Sichuan University, Chengdu, Sichuan 610041, P. R. China
揖Abstract铱 Background and Objective:
Methods:
Results:
Conclusions:
Key words:
Correspondence to: Feng Bi; Tel: +862885423609; Fax: +862885423609; Email: [email protected]
This paper was translated from Chinese into English by Medical Translation and edited by Wei Liu on 20100430.
Received: 20091116; Accepted: 20100210
Grant: National Natural Science Foundation of China (No. 30901719, No.
30971519)
Colon cancer is one of the common types of cancer in China. Chemotherapy is an important treatment for colon cancer [1] . Oxaliplatin (LOHP), a thirdgeneration platinum based drug, is one of the firstline drugs in colon cancer treatment. It has been reported in some clinical trials that as the standard firstline treatment of advanced colon cancer, the FOLFOX regimen [LOHP, 5fluorouracil (5FU) plus
leucovorin (LV)] can significantly increase the response rate up to 54% and the median survival to nearly two years. However, when switching to secondline treatment as the firstline chemotherapy fails, the response rate drops to only 4% [2] . Failure of chemotherapy greatly results from the acquired or congenital multidrug resistance (MDR) against various chemotherapeutic agents in a few cancer cells [3] . Certain cancer cells survive the chemotherapy and proliferate infinitely, which finally causes the death of the patient. Some researches showed that cancer stem cells may play an important role in this drugresistant course [4] . Hence, it is necessary to investigate the mechanism of LOHPresistance in colon cancer cells and provide new methods of overcoming this resistance in clinical practice. In this study, we used LOHP as an in vitro inducer to establish two LOHPresistant human colon cancer cell lines and explore the mechanism of this resistance.
窑 Original Article窑
661
2010; Vol. 29 Issue 7
Chinese Journal of Cancer
RPMI1640 culture medium and fetal bovine serum (FBS) were purchased from Gibco BRL company; CCK8 from Dojindo company; LOHP from SanofiAventis company; 5FU from Jinyao Amino Acid company (Tianjin); cisplatin (DDP) from Gejiu BioPharmaceutical (Yunnan); paclitaxel (PTX) from Taiji company; epirubicin (EPI) from Pfizer company; etoposide (VP16) from Sunnyhope Pharmaceutical company (Sichuan); and vincristine (VCR) from Minsheng Pharmaceutical company (Hangzhou).
The human colon cancer cell lines SW620 and lovo were preserved by our own lab. Oxaliplatinresistant cell lines SW620/LOHP and LoVo/LOHP were induced by continuous exposure to LOHP of low and gradually increased concentrations. When the cells were in logarithmic growth phase, LOHP was added to the medium to a final concentration of 0.01 mg/L. After a 24hour incubation, the old medium was discarded and fresh medium was added. Cells were passed when they were 80% confluent and LOHP of 0.01 mg/L was then added. The concentration of LOHP was gradually increased after the cells had grown stably. Finally, a cell line resistant to LOHP of 0.2 mg/L was derived from SW620 and kept in complete medium containing LOHP of 0.2 mg/L; a cell line resistant to LOHP of 2 mg/L was derived from lovo and kept in complete medium containing LOHP of 2 mg/L.
Single cell suspension (8 伊 10 3 /mL, 100 滋 L) was dispensed in a 96well plate. After a 24hour preincubation, the old medium was replaced by medium containing LOHP of 10 different concentrations, which was replaced in turn with fresh medium after another 24hour incubation. After 72 h, CCK8 was added for another 4hour incubation. Then the absorbance ( value) at 490/630 nm was measured using a UV spectrophotometer. An effectdose curve was drawn to calculate 50% inhibition concentration (IC50) and resistance index (RI). RI = IC 50 of drugresistant cell line / IC 50 of parent cell line.
Drugresistant cells were dispended in a 96well plate (800 cells/well). Three wells were taken each day and CCK8 was added in for a 4hour incubation. A value at 490/630 nm was measured using a UV spectrophotometer. This procedure was repeated for 6 continuous days. Growth curves were drawn to calculate population doubling time according to the Patterson formula.
Single cell suspension was collected and washed with cold PBS for 3 times, then fixed with 70% ethanol at 4益 overnight. After centrifugation, the supernatant was discarded and the cells were washed with cold PBS twice. Then the cells were stained with propidium iodide (PI). Flow cytometry (FCM) was performed to analyze cell cycle.
After 8% sodium dodecyl sulfate polyacrylamide gel electropheresis (SDSPAGE), proteins were transferred onto a PVDF membrane. The membrane was blocked at room temperature for 2 h and then washed. Then the membrane was incubated with 1 颐 500 diluted P glycoprotein (Pgp) antibody, 1 颐 500 diluted multidrugresistant protein 1 (MRP1) antibody and 1颐 500 diluted MRP2 antibody at 4益 overnight, washed with TBST and then incubated with secondary antibody at room temperature for 2 h. After washed with TBST, the membrane was analyzed using an Odyssey Infrared Fluorescence Detector.
After dissociation and centrifugation, the cells were washed with PBS once and incubated with FITClabeled antiCD133 and PElabeled antiCD44 in an ice bath in dark for 45 min. Then the cells were washed with FACS buffer for 3 times. FCM was performed, with antimouse IgG being the isotype control.
The significance of difference between two paired groups was determined by the Student爷s test using the
662
www.cjcsysu.cn
Chinese Journal of Cancer Chinese Journal of Cancer
Figure 1 Morphologic appearance of parental cells and L鄄 OHP鄄 resistant cells SW620 cells (A) and lovo cells (C) have homogeneous size and are polygonal; SW620/L鄄 OHP cells (B) and lovo/L鄄 OHP cells (D) are irregular, with large nuclei, and some of them are enlarged ( 伊40).
SPSS17.0 software. A value of < 0.05 was considered significant.
Two LOHPresistant cell lines SW620/LOHP (resistant to LOHP of 0.2 mg/L) and LoVo/LOHP (resistant to LOHP of 2 mg/L) were successfully induced by continuous exposure to LOHP of low and gradually increased concentrations after 10 months for more than 100 cell passages. The cell lines were used for experiments after another 2 months of culture in drugfree medium. Figure 1 shows the morphologic differences between the resistant cells and their parent cells under light microscope.
According to the results of CCK8 assay, the IC 50 of
LOHP for SW620/LOHP cells was 0.337 mg/L, and RI was 21.06. After 2 months of drugfree culture, the IC 50 was 0.316 mg/L and RI was 19.750. Thus 93.78% drugresistance was preserved. The IC 50 for lovo/LOHP was 3.513 mg/L, and RI was 13.780. After 2 months of drugfree culture, the IC 50 was 3.162 mg/L and RI was 12.4. Thus 90.00% resistance was preserved. In addition, the SW620/LOHP and LoVo/LOHP cells grew stably after they had been frozen for 3 months and then thawed, and the IC 50 then was 0.337 mg/L and 3.513 mg/L, respectively. Thus freezing and thawing did not influence the drugresistance of these cell lines.
The SW620/LOHP and LoVo/LOHP cells also showed crossresistance to 5FU, VP16, DDP, VCR and EPI to various degrees, but were still sensitive to PTX (Table 1).
The growth curves of the parent cells and the drugresistant cells (Figure 2) show that the drugresistant
cells grew more slowly. The population doubling time of SW620/LOHP cells was significantly longer than that of SW620 cells by 5.41 h ( = 0.006), and the population doubling time of LoVo/LOHP cells was significantly longer than that of lovo cells by 3.34 h ( = 0.005).
After gaining drugresistance by LOHP induction, the cells proliferated more slowly in the logarithmic phase. The results of FCM showed an increase in the proportion of cells in G0 /G1 phase (38.9% vs. 29.0% , = 0.003) and a decrease in the proportion in G 2 /M phase (15.6% vs. 31.5% , < 0.001) in SW620/LOHP cells compared with SW620 cells, and an increase in the proportion of cells in G0 /G1 phase (61.2% vs. 50.1%, = 0.001) and a decrease in the proportion in G 2 /M phase (10.6% vs. 18.1% , =
0.001) in LoVo/LOHP cells compared with LoVo cells. All these changes were of statistical significance (Figure 3).
Western blot analysis showed that comparing with their parental cells, the expression of MRP2 protein in the resistant cells was upregulated, while those of Pgp and MRP1 had no significant change (Figure 4).
The results of FCM showed that comparing with their parental cells, CD133 was overexpressed (9.6% vs. 4.6%)
A B C D
663
2010; Vol. 29 Issue 7
Chinese Journal of Cancer
L鄄 OHP 5鄄 FU VP鄄 16 PTX VCR DDP EPI
0.016 依 0.004 0.018 依 0.005 0.175 依 0.084 0.015 依 0.001 0.014 依 0.002 0.122 依 0.065 0.004 依 0.001
0.337 依 0.084 0.105 依 0.017 0.400 依 0.027 0.015 依 0.002 0.027 依 0.007 0.634 依 0.168 0.034 依 0.003
21.06 5.83 2.29 1.00 1.93 5.20 8.50
0.022 0.001 0.011 0.815 0.038 0.008
< 0.001
0.255 依 0.025 0.085 依 0.007 1.241 依 0.251 0.738 依 0.142 0.289 依 0.027 0.525 依 0.113 0.252 依 0.020
3.513 依 0.329 0.249 依 0.018 2.354 依 0.484 0.847 依 0.114 0.478 依 0.058 3.188 依 0.725 1.271 依 0.041
13.78 2.93 1.90 1.15 1.65 6.07 5.04
< 0.001 < 0.001
0.024 0.359 0.007 0.003
< 0.001
Drug SW620 SW620/L鄄 OHP
RI P IC50 (mg/L)
Figure 2 Growth curves of SW620, SW620/L鄄 OHP, LoVo and LoVo/L鄄 OHP cells The curves of L鄄 OHP鄄 resistant cells rise up slowly. The doubling time of SW620 cells (银) is 29.53 h and that of SW620/L鄄 OHP cells (伊) is 34.94 h, on the other hand, the doubling time of loVo cells (音) is 24.77 h and that of loVo/L鄄 OHP cells (姻) is 28.11 h.
L鄄 OHP, oxaliplatin; 5鄄 FU, 5鄄 fluorouracil; VP鄄 16, etoposide; PTX, paclitaxel; VCR, vincristine; DDP, cisplatin; EPI, epirubicin. The sensitivity of colon cancer cell lines to seven antitumor drugs was evaluated using the CCK8 assay as described in methods. The 50% inhibition concentration (IC50) of antitumor drugs was calculated. The IC50 values are presented as mean 依 standard deviation. SW620/L鄄 OHP and LoVo/L鄄 OHP cells exhibit moderate cross鄄 resistance to cisplatin and epirubicin, but are sensitive to paclitaxel.
while CD44 level did not significantly change (0.2% vs. 0.2% ) in SW620/LOHP cells, and similarly CD133 was overexpressed (0.9% vs. 0.3% ) while CD44 level did not significantly change (0.2% vs. 0.2% ) in loVo/LOHP cells (Figure 5).
Oxaliplatin, a thirdgeneration platinumbased drug, is
the only one p latinumbased drug that has antitumor activity in colon cancer. Its cytotoxicity is resulted from inhibition of DNA synthesis by crosslinking adjacent guanine bases or adjacent guanine and adenine [5] . Resistance occurs when this DNA damage is repaired by nucleotide excision repair (NER) mechanism [6] . However, recent studies on the mechanism of platinum drugresistance mainly focus on cisplatin, but seldom on LOHP. Studies on the establishment of in vitro LOHPresistant model are also rarely seen. Therefore, in
Figure 3 Cell cycle distribution of parental cells and L鄄 OHP鄄 resistant cells Cell cycle determined by FCM shows a decrease of the proportion of cells in G 2 /M phase and an increase of the proportion in G 0 /G 1 phase in SW620/L鄄 OHP cells (D) and LoVo/L鄄 OHP cells (B) compared with SW620 cells (C) and LoVo cells (A).
LoVo cells LoVo/L鄄 OHP cells SW620 cells SW620/L鄄 OHP cells
2.5 2.0 1.5 1.0 0.5
0 1 2 3 4 5 6
Time (/d)
10080604020
0 0 64 128 192 256 320 384 448 512 DNA Content
10080604020 0 0 64 128 192 256 320 384 448 512 DNA Content
353025201510
5 0 0 64 128 192 256 320 384 448 512 DNA Content
120 10080604020
0 0 64 128 192 256 320 384 448 512 DNA Content
A B C D CV G21S0 CV G21S0 CV G21S0 CV G21S0 CELL CYCLE
DATA Mean G1=118.2 CV G1 = 7 .9 % G1 = 38 .9
Mean G2=286.9 CV G2 = 7 .9 % G2 = 15 .6
% S = 45. 5 G2/G1 =1 .750 Chi Sq. = 0 .9
CELL CYCLE DATA Mean G1=122.2 CV G1 = 8.6 % G1 = 29.0
Mean G2=213.9 CV G2 = 8.6 % G2 = 31.5
% S = 39.5 G2/G1 =1.750 Chi Sq. = 1.0
CELL CYCLE DATA Mean G1=137.7 CV G1 = 5.5 % G1 = 61.2
Mean G2=241.0 CV G2 = 5.5 % G2 = 10.6
% S = 28.2 G2/G1 =1.750 Chi Sq. = 1.5
CELL CYCLE DATA Mean G1=84.9 CV G1 = 8.4 % G1 = 50.1
Mean G2=148.6 CV G2 = 8.3 % G2 = 18.1
% S = 31.8 G2/G1 =1.750 Chi Sq. = 1.8
LoVo LoVo/L鄄 OHP RI P
IC50 (mg/L)
664
www.cjcsysu.cn
Chinese Journal of Cancer Chinese Journal of Cancer
the current study, we established two in vitro LOHPresistant cell models to provide the basis for further studies on the mechanism of LOHPresistance in colon cancer.
One of the major approaches of studying the mechanism of MDR in tumor cells is to establish drugresistant cell lines in vitro. In this study, it took 10 months for us to successfully establish two LOHPresistant human colon cancer cell lines SW620/LOHP and LoVo/LOHP through continuous exposure to LOHP of low
and gradually increased concentrations. These two cell lines have shown stable drugresistance and consistent biological characteristics.
Cell cycle is arrested in G2 /M phase by platinumbased drugs through their inhibitory effect on cyclindependent kinase (CDK) activity. Carole Voland . reported that cells were arrested in G 2 phase by LOHP platinum as a result of inhibition of G2 /M phase transition and thus inhibition of cell division [7] , an increase in cells in G 2 /M phase finally led to apoptosis, which was showed as
Figure 4 Expressions of P鄄 glycoprotein (P鄄 gp), multidrug鄄 resistant protein 1 (MRP1) and MRP2 in SW620, SW620/L鄄 OHP, LoVo and LoVo/L鄄 OHP cells. Up regulation of MRP2 is observed in SW620/L鄄 OHP and LoVo/L鄄 OHP cells compared with SW620 and LoVo cells, while the expressions of P鄄 gp and MRP1 had no differences between parental cells and oxaliplatin鄄 resistant cells.
Figure 5 Expression of CD133 and CD44 in SW620 and SW620/L鄄 OHP cells The positive rates of CD133 were 4.6% in SW620 cells (A), 9.6% in SW620/L鄄 OHP cells (B), 0.9% in LoVo/L鄄 OHP cells (E), and 0.3% in LoVo cells (F); the positive rates of CD44 were 0.2% in SW620 cells (C), SW620/L鄄 OHP cells (D), LoVo/L鄄 OHP cells (G), and LoVo cells (H). The expression of CD133 is increased in oxaliplatin鄄 resistant cell lines compared with parental cells while CD44 expression shows no difference between parental cells and oxaliplatin鄄 resistant cells.
MRP2
MRP1
P鄄 gp
茁 鄄 actin
A B C C G: SSvsFS G: SSvsFS G: SSvsFS G: SSvsFS
.1 1000 PMT3 LOG .1 1000 PMT3 LOG .1 1000 PMT3 LOG .1 1000 PMT3 LOG
1 2
3 4 D
1 2
3 4 C
1 2
3 4 B
1 2
3 4 A
E F G H G: SSvsFS G: SSvsFS G: SSvsFS G: SSvsFS
.1 1000 PMT3 LOG .1 1000 PMT3 LOG .1 1000 PMT3 LOG .1 1000 PMT3 LOG
1 2
B
1 2
3 4 A
1
4
2
3
1 2
3 4 A
3 4 B
665
2010; Vol. 29 Issue 7
Chinese Journal of Cancer
cytotoxicity of the drug [8] . In the current study, an increase in the proportion of cells in G 0 /G 1 phase and S phase as well as a decrease in the proportion in G2 /M phase were observed in the two LOHPresistant cell lines. Activation of DNA repair mechanisms in the resistant cells might be the possible reason of these changes, which helped the cells pass through G 2 phase to M phase and subsequently divide and proliferate, which in turn resulted in a shift of cell cycle distribution. These findings may provide the basis for the selection of cell cyclespecific chemotherapeutic drugs for LOHPresistant patients in clinical treatment.
The LOHPresistant cell lines also showed crossresistance to platinumbased drugs such as DDP, as well as to some other anticancer drugs that act via different mechanisms. Some researchers have suggested a grading system depending on RI: low (< 5), medium (515) and high (> 15) [9] . According to this system, SW620/LOHP and lovo/LOHP cells both showed medium resistance to DDP and EPI and low resistance to VP16 and VCR, but no resistance to PTX. In addition, SW620/LOHP and LoVo/LOHP cells showed medium and low resistance, respectively, to 5FU.
One common mechanism of drugresistance is a decrease in the accumulation of platinum compounds in the cells, which is thought to be closely related to the exporter ATP7B but not associated with Pgp and MRP1 [10] . Liedert . [7] demonstrated an increase in both MRP2 mRNA and
protein levels in cells resistant to platinumbased drugs, but no significant change in mRNA and protein levels of other ATPbinding cassette transporters, which is consistent with the LOHPresistant cell lines in our present study. It has also been reported that it is the ATPdependent GSX pump instead of Pgp that accounts for the enhanced exportation of platinum drugs in the platinum drugresistant cells.
MRP (ABCC) proteins are membrane transporters that can use the energy of ATP hydrolysis to carry out their 野drug pump冶 function [11] . These proteins are 15% homologous to Pgp in amino sequence but do not act synergistically with Pgp in term of their 野drug pump冶 function. Pgp acts on a variety of drugs, including doxorubicin (ADM), VCR, VP16, PTX, and anthracene derivatives [12] , whereas MRP transfers cytotoxic drugs that are able to bind reduced glutathione (GSH), such as ADM, VP16, and VCR [13] , and alters their intracellular distribution [14] . Resistance to EPI, VP16, and VCR [1517] is Pgp/MRP mediated; however, neither Pgp nor MRP1 was upregulated in LoVo/LOHP and SW620/LOHP cells, which suggests that the crossresistance to EPI, VP16, and VCR in these two cell lines was possibly MRP2mediated. Some researches showed that overexpression of MRP2 resulted in crossresistance to VP16, DDP, ADM, and EPI [18] . MRP2mediated resistance to certain drugs is associated with GSH level, and the MRP2mediated resistance to
VP16, ADM, and DDP can be reversed by the GSH inhibitor BSA. Moreover, in our present study, the LOHPresistant cells also demonstrated low to medium resistance to 5FU. 5FUresistance is believed to result from several mechanisms [19] , including changes in the activities of thymidylate synthetase (TS), dihydropyrimidine dehydrogenase (DPD) and folic acid metabolismrelated enzymes, DNA mismatch repair (MMR) [20] , and so on, but not associated with the expressions of Pgp and MRP. These imply that aberration of DNA repair mechanisms may also play an important role in 5FUresistance. In addition, the two LOHPresistant cell lines in our study showed no crossresistance to PTX, which may be because PTX resistance is mediated by Pgp rather than by MRP2 [21,22] . This provides an important basis for selecting sensitive secondline drugs when patients fail to respond to LOHP as the firstline drug in clinical treatment.
Cancer stem cells are stem celllike cells that exist in the hematopoietic system or some solid tumors. The cancer stem cell hypothesis suggests that during chemotherapy tumor size reduces but a small number of stem cells survive, which cause relapse and metastasis [23] . The selfprotective mechanism of stem cells against cytotoxic drugs remains to be elucidated. But it is currently believed that the major mechanisms involve the drug pump activity of ABC transporters [24] , overexpression of antiapoptotic genes, DNA repair, increase in cells in G 0 /G 1 phase, and slowdown of the proliferation of stem cells to evade the action of chemotherapeutic drugs [25] . Stem cells enriched from U87MG cells overexpress CD133 and are crossresistant against ADM, VP16, carboplatin and BCNU [26] . In our present study, MRP2 upregulation, slowdown of cell proliferation, and an increase in G0/G1 phase cells were observed in the LOHPresistant cell lines, which implied a stem cell phenotype. Furthermore, the proportion of CD133 +
cells also increased as the cells became resistant to LOHP. These all may suggest an important role of cancer stem cells in the development of MDR in the LOHPresistant cell lines. Thus it is possible to sensitize the tumor tissue to LOHP and reverse its resistance by using stem celltargeting therapy.
In the current study, we successfully established two LOHPresistant human colon cancer cell lines SW620/LOHP and lovo/LOHP. They are stable in cell passage and drugresistance, and their biological characteristics are highly consistent. These cell lines may serve as ideal models for the study of the mechanism and the way of reversal of LOHPresistance in colon cancer.
Segal NH, Saltz LB. Evolving treatment of advanced colon cancer 咱1暂
666
www.cjcsysu.cn
Chinese Journal of Cancer Chinese Journal of Cancer
[J]. Annu Rev Med, 2009,60:207-219. Tourniqand C, Andre T, Achille E, et al. FOLFIRI followed by FOLFOX6 or the reverse sequence in advanced colorectal cancer: a randomized GERCOR study [J]. J Clin Oncol, 2004,22 (2):229- 237. Wu DL, Huang F, Lu HZ. Drug鄄 resistant proteins in breast cancer: recent progress in multidrug resistance [J]. Chin J Cancer, 2003,22(4):441-4. [in Chinese] Hong SP, Wen J, Park S, et al. CD44鄄 positive cells are responsible for gemcitabine resistance in pancreatic cancer cells [J]. Int J Cancer, 2009,125(10):2323-2331. Rabik CA, Dolan ME. Molecular mechanisms of resistance and toxicity associated with platinating agents [J]. Cancer Treat Rev, 2007,33(1):9-23. Gossage L, Madhusudan S. Current status of excision repair cross complementing鄄 group 1 (ERCC1) in cancer [J]. Cancer Treat Rev, 2007,33(6):565-577. Liedert B, Materna V, Schadendorf D, et al. Overexpression of cMOAT (MRP2/ABCC2) is associated with decreased formation of platinum鄄 DNA adducts and decreased G2鄄 arrest in melanoma cells resistant to cisplatin [J]. J Invest Dermatol, 2003,121(1):172-176 Voland C, Bord A, P佴 leraux A, et al. Repression of cell cycle鄄 related proteins by oxaliplatin but not cisplatin in human colon cancer cells [J]. Mol Cancer Ther, 2006,5(9):2149-2157. Snow K, Judd Wl. Characterisation of adriamycin and amsacrine鄄 resistant human leukaemic T cell lines [J]. Br J Cancer, 1991,63 (1):17-28. Rabik CA, Dolan ME. Molecular mechanisms of resistance and toxicity associated with platinating agents [J]. Cancer Treat Rev, 2007,33(1):9-23. Yu ZC, Ding J, Bi F, et al. Reversal effects of mrp antisense RNA on the resistance of gastric cancer cell line SGC7901 to VCR [J]. Chin J Cancer Biother, 2000,7(3):174-176. [in Chinese] Chen LM, Liang YJ, Zhang X, et al. Reversal of P鄄 gp鄄 mediated multidrug resistance by bromotetrandrine in vivo is associated with enhanced accumulation of chemotherapeutical drug in tumor tissue [J]. Anticancer Res. 2009,29(11):4597-4604. Woodahl EL, Crouthamel MH, Bui T, et al. MDR1 (ABCB1) G1199A (Ser400Asn) polymorphism alters transepithelial permeability and sensitivity to anticancer agents [J]. Cancer Chemother Pharmacol. 2009 ,64(1):183-188. Koehn J, Fountoulakis M, Krapfenbauer K. Multiple drug resistance associated with function of ABC鄄 transporters in diabetes mellitus:
molecular mechanism and clinical relevance [J]. Infect Disord Drug Targets. 2008,8(2):109-118. Yuan SQ, Zhou ZW, Liang YJ, et al. Correlation of chemosensitivity measured by histoculture drug response assay to expression of multidrug resistance genes and proteins in gastric cancer [J]. Chin J Cancer, 2009,28(4):337-43. [in Chinese] Zhai BJ, Wu F, Shao ZY, et al. Establishment of in vivo adriamycin鄄 induced multidrug resistance models of subcutaneous and hepatic transplanted human liver cancer in nude mice [J]. Chin J Cancer, 2004,23(8):905-9. [in Chinese] Boh佗 cov佗 V, Sulov佗 Z, Dovinov佗 I, et al. L1210 cells cultivated under the selection pressure of doxorubicin or vincristine express common mechanisms of multidrug resistance based on the overexpression of P鄄 glycoprotein [J]. Toxicol In Vitro. 2006,20 (8): 1560-1568. Cui Y, K觟 nig J, Buchholz JK, et al. drug resistance and ATP鄄 Dependent conjugate transport mediated by the apical multidrug resistance protein,MRP2,permanently expressed in human and canine cells[J]. Mol Pharmacol. 1999,55(5):929-937. Yamada T, Tanaka N, Yokoi K,et al. Prediction of sensitivity to 5鄄 fluorouracil (5鄄 fu) by metabolic and target enzyme activities in colon cancer[J]. Gan To Kagaku Ryoho. 2006,33(11):1603-1609. Oda S, Kuraoka I, Maehara Y. DNA repair as a determinant of tumour chemosensitivity [J]. Gan To Kagaku Ryoho. 2007,34 (3): 347-357. Takano M, Otani Y, Tanda M,et al. Paclitaxel鄄 resistance conferred by altered expression of efflux and influx transporters for paclitaxel in the human hepatoma cell line, HepG2 [J]. Drug Metab Pharmacokinet. 2009,24(5):418-427. Zhou J, Cheng SC, Luo D,et al. Study of multi鄄 drug resistant mechanisms in a taxol鄄 resistant hepatocellular carcinoma QGY鄄 TR 50 cell line [J]. Biochem Biophys Res Commun. 2001, 280 (5): 1237-1242. Mittal S, Mifflin R, Powell DW. Cancer stem cells: the other face of Janus[J]. Am J Med Sci. 2009 ,338(2):107-112. Dean M. ABC transporters, drug resistance, and cancer stem cells [J]. Mammary Gland Biol Neoplasia. 2009,14(1):3-9. Gangemi R, Paleari L, Orengo AM. Cancer stem cells: a new paradigm for understanding tumor growth and progression and drug resistance[J]. Curr Med Chem. 2009,16(14):1688-1703. Nakai E, Park K, Yawata T, et al. Enhanced MDR1 Expression and Chemoresistance of Cancer Stem Cells Derived from Glioblastoma [J]. Cancer Invest袁. 2009,27(9):901-908.
咱2暂
咱3暂
咱4暂
咱5暂
咱6暂
咱7暂
咱8暂
咱9暂
咱10暂
咱11暂
咱12暂
咱13暂
咱14暂
咱15暂
咱16暂
咱17暂
咱18暂
咱19暂
咱20暂
咱21暂
咱22暂
咱23暂 咱24暂 咱25暂
咱26暂
667