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Increased sensitivity to gemcitabine of P-glycoprotein and multidrug resistance-associated protein-overexpressing human cancer cell lines.

Bergman AM, Pinedo HM, Talianidis I, Veerman G, Loves WJ, van der Wilt CL, Peters GJ - Br. J. Cancer (2003)

Bottom Line: Gemcitabine was tested against human melanoma, non-small-cell lung cancer, small-cell lung cancer, epidermoid carcinoma and ovarian cancer cells with an MDR phenotype as a result of selection by drug exposure or by transfection with the mdr1 gene.In 2R120 and 2R160 cells, dCK activities were seven- and four-fold higher than in SW1573, respectively, which was associated with an increased dCK mRNA and dCK protein.P-glycoprotein and MRP1 overexpression possibly caused a cellular stress resulting in increased gemcitabine metabolism and sensitivity, while reversal of collateral gemcitabine sensitivity by verapamil also suggests a direct relation between the presence of membrane efflux pumps and gemcitabine sensitivity.

View Article: PubMed Central - PubMed

Affiliation: Department Medical Oncology, VU University Medical Center, PO Box 7057, 1007 MB Amsterdam, The Netherlands.

ABSTRACT
Gemcitabine (2',2'-difluorodeoxycytidine) is a deoxycytidine analogue that is activated by deoxycytidine kinase (dCK) to its monophosphate and subsequently to its triphosphate dFdCTP, which is incorporated into both RNA and DNA, leading to DNA damage. Multidrug resistance (MDR) is characterised by an overexpression of the membrane efflux pumps P-glycoprotein (P-gP) or multidrug resistance-associated protein (MRP). Gemcitabine was tested against human melanoma, non-small-cell lung cancer, small-cell lung cancer, epidermoid carcinoma and ovarian cancer cells with an MDR phenotype as a result of selection by drug exposure or by transfection with the mdr1 gene. These cell lines were nine- to 72-fold more sensitive to gemcitabine than their parental cell lines. The doxorubicin-resistant cells 2R120 (MRP1) and 2R160 (P-gP) were nine- and 28-fold more sensitive to gemcitabine than their parental SW1573 cells, respectively (P<0.01), which was completely reverted by 25 micro M verapamil. In 2R120 and 2R160 cells, dCK activities were seven- and four-fold higher than in SW1573, respectively, which was associated with an increased dCK mRNA and dCK protein. Inactivation by deoxycytidine deaminase was 2.9- and 2.2-fold decreased in 2R120 and 2R160, respectively. dFdCTP accumulation was similar in SW1573 and its MDR variants after 24 h exposure to 0.1 micro M gemcitabine, but dFdCTP was retained longer in 2R120 (P<0.001) and 2R160 (P<0.003) cells. 2R120 and 2R160 cells also incorporated four- and six-fold more [(3)H]gemcitabine into DNA (P<0.05), respectively. P-glycoprotein and MRP1 overexpression possibly caused a cellular stress resulting in increased gemcitabine metabolism and sensitivity, while reversal of collateral gemcitabine sensitivity by verapamil also suggests a direct relation between the presence of membrane efflux pumps and gemcitabine sensitivity.

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Sensitivity factors to gemcitabine, without 25 μM verapamil (▪) or with 25 μM verapamil () (IC50 of the MDR variants relative to IC50 of SW1573 was set at 1), of the MRP1-overexpressing 2R120, the P-gP-overexpressing 2R160, the mdr1-transfected S1(1.1) and the MRP1-transfected S1(MRP). Verapamil itself did not affect cellular growth. Values are means±s.d. of at least three experiments. *Sensitivity with 25 μM verapamil statistically significantly different from gemcitabine alone (t-test, independent samples, P<0.05).
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fig2: Sensitivity factors to gemcitabine, without 25 μM verapamil (▪) or with 25 μM verapamil () (IC50 of the MDR variants relative to IC50 of SW1573 was set at 1), of the MRP1-overexpressing 2R120, the P-gP-overexpressing 2R160, the mdr1-transfected S1(1.1) and the MRP1-transfected S1(MRP). Verapamil itself did not affect cellular growth. Values are means±s.d. of at least three experiments. *Sensitivity with 25 μM verapamil statistically significantly different from gemcitabine alone (t-test, independent samples, P<0.05).

Mentions: The MDR phenotype of doxorubicin resistance can be reversed by the calcium-channel blocker verapamil. This phenomenon has been used as a characteristic of the efflux pump. In order to determine whether the activity of the pump was associated with the collateral sensitivity to gemcitabine, we exposed SW1573 and its variants to gemcitabine and verapamil. Verapamil almost completely reversed the gemcitabine sensitivity to the level of the parental SW1573 cells (Figure 2Figure 2


Increased sensitivity to gemcitabine of P-glycoprotein and multidrug resistance-associated protein-overexpressing human cancer cell lines.

Bergman AM, Pinedo HM, Talianidis I, Veerman G, Loves WJ, van der Wilt CL, Peters GJ - Br. J. Cancer (2003)

Sensitivity factors to gemcitabine, without 25 μM verapamil (▪) or with 25 μM verapamil () (IC50 of the MDR variants relative to IC50 of SW1573 was set at 1), of the MRP1-overexpressing 2R120, the P-gP-overexpressing 2R160, the mdr1-transfected S1(1.1) and the MRP1-transfected S1(MRP). Verapamil itself did not affect cellular growth. Values are means±s.d. of at least three experiments. *Sensitivity with 25 μM verapamil statistically significantly different from gemcitabine alone (t-test, independent samples, P<0.05).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2741118&req=5

fig2: Sensitivity factors to gemcitabine, without 25 μM verapamil (▪) or with 25 μM verapamil () (IC50 of the MDR variants relative to IC50 of SW1573 was set at 1), of the MRP1-overexpressing 2R120, the P-gP-overexpressing 2R160, the mdr1-transfected S1(1.1) and the MRP1-transfected S1(MRP). Verapamil itself did not affect cellular growth. Values are means±s.d. of at least three experiments. *Sensitivity with 25 μM verapamil statistically significantly different from gemcitabine alone (t-test, independent samples, P<0.05).
Mentions: The MDR phenotype of doxorubicin resistance can be reversed by the calcium-channel blocker verapamil. This phenomenon has been used as a characteristic of the efflux pump. In order to determine whether the activity of the pump was associated with the collateral sensitivity to gemcitabine, we exposed SW1573 and its variants to gemcitabine and verapamil. Verapamil almost completely reversed the gemcitabine sensitivity to the level of the parental SW1573 cells (Figure 2Figure 2

Bottom Line: Gemcitabine was tested against human melanoma, non-small-cell lung cancer, small-cell lung cancer, epidermoid carcinoma and ovarian cancer cells with an MDR phenotype as a result of selection by drug exposure or by transfection with the mdr1 gene.In 2R120 and 2R160 cells, dCK activities were seven- and four-fold higher than in SW1573, respectively, which was associated with an increased dCK mRNA and dCK protein.P-glycoprotein and MRP1 overexpression possibly caused a cellular stress resulting in increased gemcitabine metabolism and sensitivity, while reversal of collateral gemcitabine sensitivity by verapamil also suggests a direct relation between the presence of membrane efflux pumps and gemcitabine sensitivity.

View Article: PubMed Central - PubMed

Affiliation: Department Medical Oncology, VU University Medical Center, PO Box 7057, 1007 MB Amsterdam, The Netherlands.

ABSTRACT
Gemcitabine (2',2'-difluorodeoxycytidine) is a deoxycytidine analogue that is activated by deoxycytidine kinase (dCK) to its monophosphate and subsequently to its triphosphate dFdCTP, which is incorporated into both RNA and DNA, leading to DNA damage. Multidrug resistance (MDR) is characterised by an overexpression of the membrane efflux pumps P-glycoprotein (P-gP) or multidrug resistance-associated protein (MRP). Gemcitabine was tested against human melanoma, non-small-cell lung cancer, small-cell lung cancer, epidermoid carcinoma and ovarian cancer cells with an MDR phenotype as a result of selection by drug exposure or by transfection with the mdr1 gene. These cell lines were nine- to 72-fold more sensitive to gemcitabine than their parental cell lines. The doxorubicin-resistant cells 2R120 (MRP1) and 2R160 (P-gP) were nine- and 28-fold more sensitive to gemcitabine than their parental SW1573 cells, respectively (P<0.01), which was completely reverted by 25 micro M verapamil. In 2R120 and 2R160 cells, dCK activities were seven- and four-fold higher than in SW1573, respectively, which was associated with an increased dCK mRNA and dCK protein. Inactivation by deoxycytidine deaminase was 2.9- and 2.2-fold decreased in 2R120 and 2R160, respectively. dFdCTP accumulation was similar in SW1573 and its MDR variants after 24 h exposure to 0.1 micro M gemcitabine, but dFdCTP was retained longer in 2R120 (P<0.001) and 2R160 (P<0.003) cells. 2R120 and 2R160 cells also incorporated four- and six-fold more [(3)H]gemcitabine into DNA (P<0.05), respectively. P-glycoprotein and MRP1 overexpression possibly caused a cellular stress resulting in increased gemcitabine metabolism and sensitivity, while reversal of collateral gemcitabine sensitivity by verapamil also suggests a direct relation between the presence of membrane efflux pumps and gemcitabine sensitivity.

Show MeSH
Related in: MedlinePlus