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Kinetics of MDR transport in tumor-initiating cells.

Koshkin V, Yang BB, Krylov SN - PLoS ONE (2013)

Bottom Line: In this way it was shown that activation of MDR in TICs occurs in two ways: through the increase of V max in one fraction of cells, and through decrease of K M in another fraction.In addition, kinetic data showed that heterogeneity of MDR parameters in TICs significantly exceeds that of bulk cells.Potential consequences of these findings for chemotherapy are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Centre for Research on Biomolecular Interactions, York University, Toronto, Ontario, Canada.

ABSTRACT
Multidrug resistance (MDR) driven by ABC (ATP binding cassette) membrane transporters is one of the major causes of treatment failure in human malignancy. MDR capacity is thought to be unevenly distributed among tumor cells, with higher capacity residing in tumor-initiating cells (TIC) (though opposite finding are occasionally reported). Functional evidence for enhanced MDR of TICs was previously provided using a "side population" assay. This assay estimates MDR capacity by a single parameter - cell's ability to retain fluorescent MDR substrate, so that cells with high MDR capacity ("side population") demonstrate low substrate retention. In the present work MDR in TICs was investigated in greater detail using a kinetic approach, which monitors MDR efflux from single cells. Analysis of kinetic traces obtained allowed for the estimation of both the velocity (V max) and affinity (K M) of MDR transport in single cells. In this way it was shown that activation of MDR in TICs occurs in two ways: through the increase of V max in one fraction of cells, and through decrease of K M in another fraction. In addition, kinetic data showed that heterogeneity of MDR parameters in TICs significantly exceeds that of bulk cells. Potential consequences of these findings for chemotherapy are discussed.

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Related in: MedlinePlus

Kinetic and immunophenotyping analysis of 4T1 TICs and bulk cells: (a) histograms of distribution of Michaelis parameters within CD44high/CD24low cells (gray bars) and bulk cells (white bars); (b) cumulative data summarizing Michaelis parameters in CD44high/CD24low cells (gray bars) and bulk cells (white bars) in 4 independent experiments (total 709 cells, p < 0.02 for Vmax).
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pone-0079222-g004: Kinetic and immunophenotyping analysis of 4T1 TICs and bulk cells: (a) histograms of distribution of Michaelis parameters within CD44high/CD24low cells (gray bars) and bulk cells (white bars); (b) cumulative data summarizing Michaelis parameters in CD44high/CD24low cells (gray bars) and bulk cells (white bars) in 4 independent experiments (total 709 cells, p < 0.02 for Vmax).

Mentions: Distribution of Vmax and KM values within bulk cells and TICs, as well as cumulative mean data are shown in Figure 4. Similarly to MCF-7 cells, TICs in 4T1 cell line show wider than bulk cells distribution of Vmax values with increased mean value. Range of KM variation in TICs is also broader than in bulk cells, but, in contrast to MCF-7 cells, KM mean values in both subpopulations of 4T1 cells are close to each other. However, due to broadened variation range TIC in 4T1 line, like in MCF-7 line, contain a significant fraction of cells with increased affinity (reduced KM). Thus, data on two cancer cell types suggest that MDR kinetics in TIC differ from these in bulk cells by: (i) broader variation of Vmax and KM parameters, (ii) increased Vmax mean value, (iii) formation within TIC of a fraction of cells with elevated MDR affinity (reduced KM).


Kinetics of MDR transport in tumor-initiating cells.

Koshkin V, Yang BB, Krylov SN - PLoS ONE (2013)

Kinetic and immunophenotyping analysis of 4T1 TICs and bulk cells: (a) histograms of distribution of Michaelis parameters within CD44high/CD24low cells (gray bars) and bulk cells (white bars); (b) cumulative data summarizing Michaelis parameters in CD44high/CD24low cells (gray bars) and bulk cells (white bars) in 4 independent experiments (total 709 cells, p < 0.02 for Vmax).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3815210&req=5

pone-0079222-g004: Kinetic and immunophenotyping analysis of 4T1 TICs and bulk cells: (a) histograms of distribution of Michaelis parameters within CD44high/CD24low cells (gray bars) and bulk cells (white bars); (b) cumulative data summarizing Michaelis parameters in CD44high/CD24low cells (gray bars) and bulk cells (white bars) in 4 independent experiments (total 709 cells, p < 0.02 for Vmax).
Mentions: Distribution of Vmax and KM values within bulk cells and TICs, as well as cumulative mean data are shown in Figure 4. Similarly to MCF-7 cells, TICs in 4T1 cell line show wider than bulk cells distribution of Vmax values with increased mean value. Range of KM variation in TICs is also broader than in bulk cells, but, in contrast to MCF-7 cells, KM mean values in both subpopulations of 4T1 cells are close to each other. However, due to broadened variation range TIC in 4T1 line, like in MCF-7 line, contain a significant fraction of cells with increased affinity (reduced KM). Thus, data on two cancer cell types suggest that MDR kinetics in TIC differ from these in bulk cells by: (i) broader variation of Vmax and KM parameters, (ii) increased Vmax mean value, (iii) formation within TIC of a fraction of cells with elevated MDR affinity (reduced KM).

Bottom Line: In this way it was shown that activation of MDR in TICs occurs in two ways: through the increase of V max in one fraction of cells, and through decrease of K M in another fraction.In addition, kinetic data showed that heterogeneity of MDR parameters in TICs significantly exceeds that of bulk cells.Potential consequences of these findings for chemotherapy are discussed.

View Article: PubMed Central - PubMed

Affiliation: Department of Chemistry and Centre for Research on Biomolecular Interactions, York University, Toronto, Ontario, Canada.

ABSTRACT
Multidrug resistance (MDR) driven by ABC (ATP binding cassette) membrane transporters is one of the major causes of treatment failure in human malignancy. MDR capacity is thought to be unevenly distributed among tumor cells, with higher capacity residing in tumor-initiating cells (TIC) (though opposite finding are occasionally reported). Functional evidence for enhanced MDR of TICs was previously provided using a "side population" assay. This assay estimates MDR capacity by a single parameter - cell's ability to retain fluorescent MDR substrate, so that cells with high MDR capacity ("side population") demonstrate low substrate retention. In the present work MDR in TICs was investigated in greater detail using a kinetic approach, which monitors MDR efflux from single cells. Analysis of kinetic traces obtained allowed for the estimation of both the velocity (V max) and affinity (K M) of MDR transport in single cells. In this way it was shown that activation of MDR in TICs occurs in two ways: through the increase of V max in one fraction of cells, and through decrease of K M in another fraction. In addition, kinetic data showed that heterogeneity of MDR parameters in TICs significantly exceeds that of bulk cells. Potential consequences of these findings for chemotherapy are discussed.

Show MeSH
Related in: MedlinePlus