Kinetics of MDR Transport in Tumor-Initiating Cells

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.

Metabolic Suppression of a Drug‐Resistant Subpopulation in Cancer Spheroid Cells

“…To prevent alteration of mitoxantrone content, further manipulations were performed at 4°C. In our [Koshkin et al, ] and others' [ Kars et al, ; Zhang et al, ] previous studies, it was found that intrinsic MDR in MCF‐7 cells is supported by the MRP1 transporter. Therefore, loaded cells were incubated for 60 min with an antibody for the extracellular domain of MRP1 (IU2H10 1:100 50 μg/mL, Novus Biologicals, Littleton, CO) [Binyamin et al, ; Chen et al, ].…”

“…Cell viability was determined using a standard colorimetric MTT (3‐4,5‐dimethylthiazol‐2‐yl‐2, 5‐diphenyl‐tetrazolium bromide) assay. As previously described [Koshkin et al, ], cells were seeded at 10 4 cells/well in 96‐well plates and grown for 48 h at 37°C, with or without cytotoxic agent, then supplemented with MTT (0.5 mg/mL) and incubated for 4 h. Redox activity in viable cells converted the oxidized form of MTT into the reduced formazan form. Formazan crystals were released from cells and dissolved using SDS (3% final content).…”

Dasatinib self-assembled nanoparticles decorated with hyaluronic acid for targeted treatment of tumors to overcome multidrug resistance

“…Multidrug resistance (MDR) is one of the main obstacles to effective chemotherapy of tumor. Mechanisms of MDR production include increased efflux of drugs to reduce intracellular accumulation of drugs, promoting antiapoptotic mechanisms, and repairing DNA damage (Almalik et al., 2013 ; Koshkin et al., 2013 ; Singh et al., 2017 ; Trujillo-Nolasco et al., 2019 ). Among them, P-glycoprotein (P-gp), as a membrane transporter of the ATP binding cassette family, can pump out substrate drugs through an ATP-dependent mechanism, thereby reducing intracellular drug accumulation, which is one of the main causes of MDR (Chen et al., 2016 ; Gupta et al., 2017 ; Li et al., 2017 ).…”

Cutaneous metastases of breast cancer during adjuvant chemotherapy correlates with increasing CD44+/CD24− and ALDH-1 expression: a case report and literature review

“…However, the latter study (19) also reported that patients with tumors containing high numbers of CD44 + /CD24 − cells more frequently developed bone metastases in the course of the disease. The molecular mechanisms underlying the roles of breast CSCs in chemoresistance have been reported in several studies (20,21) and proposed clinical implication for breast cancer therapeutics (22)(23)(24)(25). The occurrence of a fraction of breast CSCs with increased multidrug resistance (MDR) affinity clinically may be an additional mechanism for the positive selection of breast CSCs during chemotherapy (20).…”