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Journal of Chinese Integrative Medicine ›› 2012, Vol. 10 ›› Issue (8): 901-910.doi: 10.3736/jcim20120811

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Effects of medicated serum prepared with Chinese herbal medicine Changweiqing on pharmacokinetics of oxaliplatin in colon cancer cells

Yong Zhang, Xiao-wen Sun, Jian-hua Xu(), Hai Lu, Zhong-ze Fan, Jue Sun, Xiao-xiao Zhang#br#   

  1. Department of Traditional Chinese Medicine Oncology, Putuo Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai 200062, China
  • Received:2012-01-02 Accepted:2012-01-18 Online:2012-08-20 Published:2018-08-15
  • Contact: Xu Jian-hua

Objective: To investigate the effects of Changweiqing-medicated serum, which was prepared with a compound traditional Chinese herbal medicine, on the reversal of oxaliplatin (L-OHP) resistance and the relationship between the reversal and cellular accumulation of platinum and proteins associated with copper transporter in HCT116/L-OHP cells.

Methods: For clarifying the reversal effect of Changweiqing, methyl thiazolyl tetrazolium was applied to determine the L-OHP resistance of HCT116/L-OHP cell line. The relationship between the cellular accumulation of platinum and the L-OHP resistance in HCT116/L-OHP cells, and the effects of drug-medicated serum on intracellular contents of platinum were detected by atomic absorption spectrophotometry. Western blot method was used to determine the expressions of human copper transporter 1 (hCTR1), ATPase Cu 2+ transporting alpha polypeptide (ATP7A), copper-transporting P-type adenosine triphosphatase (ATP7B), glutathione S-transferase-π (GST-π) and multidrug resistance-associated protein 2 (MRP2).

Results: The inhibitory concentration 50% values of different pairs of L-OHP-sensitive and -resistant cells were 7.2 and 89.00. The resistance index of HCT116/L-OHP cells was 12.36. The reverse index of drug serum on HCT116/L-OHP cells was 2.74. The platinum content in HCT116/L-OHP cells was decreased compared with HCT116 cells in condition of 7.2 μg/mL L-OHP. After treating by 7.5% Changweiqing-medicated serum, the intracellular platinum contents in L-OHP-sensitive and -resistant cells were increased. It was dose-dependent that drug-medicated serum promoted the uptake of L-OHP by HCT116 or HCT116/L-OHP cells and inhibited the discharge. The 7.5% Changweiqing-medicated serum increased the expression of hCTR1 and decreased the expressions of ATP7A and ATP7B in HCT116/L-OHP cells, but had no effects on GST-π and MRP2 protein expressions.

Conclusion: Changweiqing can reverse the L-OHP resistance of HCT116/L-OHP by increasing the cellular platinum-DNA accumulation. Down-regulation of expression of ATP7B and ATP7A, and up-regulation of hCTR1 may cause the increase of intracellular platinum content in HCT116/L-OHP cells.

Key words: colonic neoplasms, drug resistance, neoplasm, compounds (TCD), platinum compounds, carrier proteins, HCT116 cells

Figure 1

Effects of different treatments on content of Pt in HCT116 and HCT116/L-OHP cellsData are represented as $\bar{x}$±s, n=5. ★P<0.05, ★★P<0.01. CWQ: Changweiqing; L-OHP: oxaliplatin; Pt: platinum."

Figure 2

Effects of Changweiqing-medicated serum on Pt-DNA content in HCT116 and HCT116/L-OHP cellsData are represented as $\bar{x}$±s, n=5. *P<0.05, vs single-used oxaliplatin; △P<0.05, △△P<0.01, vs HCT116 under the same treatment. CWQ: Changweiqing; L-OHP: oxaliplatin; Pt: platinum."

Table 1

Effects of Changweiqing-medicated serum on uptake of L-OHP after 60 and 120 min ($\bar{x}$±s)"

Group n Uptake of L-OHP (ng Pt/mg prot)
60 min 120 min
HCT116 cells
L-OHP 5 1 742.20±37.32 2 463.87±66.62□■■
1.875% CWQ plus L-OHP 5 2 048.69±29.06**□ 2 681.46±64.85*□■■
3.75% CWQ plus L-OHP 5 2 361.88±92.18**△□ 2 799.18±93.25**□■■
7.5% CWQ plus L-OHP 5 2 600.82±57.44**△▲□ 3 096.01±132.08**△▲□■
HCT116/L-OHP cells
L-OHP 5 1 263.59±15.53 1 673.64±19.62
1.875% CWQ plus L-OHP 5 1 318.87±11.86* 1 879.47±36.71*■■
3.75% CWQ plus L-OHP 5 1 452.45±28.15* 1 981.50±59.77**■■
7.5% CWQ plus L-OHP 5 1 667.41±37.72**△▲ 2 193.96±20.48**△▲■■

Figure 3

Effects of Changweiqing-medicated serum on efflux of L-OHP in colon cancer cellsA: HCT116 cells Pt efflux at 30 and 60 min; B: HCT116/L-OHP cells Pt efflux at 30 and 60 min. CWQ: Changweiqing; L-OHP: oxaliplatin; Pt: platinum."

Table 2

Effects of Changweiqing-medicated serum on efflux of L-OHP after 30 and 60 min ($\bar{x}$±s)"

Group n Residual platinum (%)
30 min 60 min
L-OHP 5 0.677±0.010 0.658±0.007
1.875% CWQ plus L-OHP 5 0.693±0.011* 0.672±0.010*
3.75% CWQ plus L-OHP 5 0.729±0.003* 0.721±0.007*△
7.5% CWQ plus L-OHP 5 0.790±0.021**△▲ 0.759±0.032**△▲
L-OHP 5 0.608±0.006 0.590±0.009
1.875% CWQ plus L-OHP 5 0.613±0.008 0.597±0.005
3.75% CWQ plus L-OHP 5 0.664±0.003* 0.648±0.008*△
7.5% CWQ plus L-OHP 5 0.708±0.023*△▲ 0.671±0.030*△▲

Figure 4

Bands of copper-transporting protein and related protein expressions in HCT116 and HCT116/L-OHP cells tested by Western blottingExpressions of hCTR1, ATP7A, ATP7B, GST-π and MRP2 proteins observed by Western blotting. CWQ: Changweiqing; L-OHP: oxaliplatin; hCTR1: human copper transporter 1; ATP7A: ATPase Cu2+ transporting alpha polypeptide; ATP7B: copper-transporting P-type adenosine triphosphatase; GST-π: glutathione S-transferase-π; MRP2: multidrug resistance-associated protein 2. "

Figure 5

Copper-transporting protein and related protein expressions in HCT116 and HCT116/L-OHP cellsQuantity results of protein expressions of hCTR1 (A), ATP7A (B), ATP7B (C), GST-π (D) and MRP2 (E). *P<0.05, vs control group; △P<0.05, vs HCT116 cells; ▲P<0.05, vs single-used oxaliplatin. CWQ: Changweiqing; L-OHP: oxaliplatin; hCTR1: human copper transporter 1; ATP7A: ATPase Cu2+ transporting alpha polypeptide; ATP7B: copper-transporting P-type adenosine triphosphatase; GST-π: glutathione S-transferase-π; MRP2: multidrug resistance-associated protein 2."

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