Synergistic Reversal of Multidrug-Resistance Phenotype in Acute Myeloid Leukemia Cells by Cyclosporin A and Cremophor EL

Abstract

AbstractCremophor (Crem) EL, the vehicle for intravenous delivery of cyclosporin A (CsA), has been reported to counteract multidrug resistance (MDR) in P-glycoprotein (Pgp)-over-expressing cell lines. Because of this, we sought to determine whether Crem functions independently as a modulator of MDR in blast cells obtained from acute myelogenous leukemia (AML) patients, and the nature of its interaction in combination with CsA in reversing an MDR phenotype. In the phenotypically classical MDR AML cell lines HL-60/Vine (overexpresses Pgp) or HL-60/AR (does not overex-press Pgp), the dose causing half-maximum enhancement (D50) of daunorubicin (DNR, 1 µg/mL, 3 hours) accumulation was achieved by the combination of CsA and Crem (CsA/Crem) at 1.2 µmol/L CsA. In contrast, the D50 for Crem alone was approached at an amount that would be needed to suspend 6.2 µmol/L CsA for HL-60/Vinc, and 81 µmol/L CsA for HL-60/AR. The D50 concentrations for CsA alone (dissolved in ethanol, which does not alter DNR accumulation) were also higher than those for CsA/Crem, being 6.5 µmol/L for HL-60/Vinc, and 3.1 µmol/L for HL-60/AR. The maximum absolute level of enhancement of DNR accumulation (Emax) in each cell line was approximately equivalent for CsA/Crem or CsA alone, and was equal to the 3 hr intracellular DNR accumulation observed in parental, drug sensitive HL-60/W cells. For Crem alone, HL-60/AR and HL-60/Vinc cells showed markedly different responses: HL-60/Vinc cells attained intracellular DNR content comparable to HL-60/W, whereas HL-60/AR cells achieved only approximately 35% of this level. Multiple-drug effects were analyzed by calculation of the Combination Index (Chou and Talalay, Adv Enzyme Regul 22:27, 1984), which indicated that CsA and Crem are synergistic in causing enhancement of DNR accumulation in these MDR HL-60 cell lines. In blasts from AML patients, 5 µmol/L CsA/Crem or an equivalent amount of Crem alone each caused significant (P < .001) enhancement of DNR accumulation (60 AML-patient marrow samples) or DNR retention (51 AML-patient marrows). Similarly, CsA/Crem or Crem alone caused significant (P < .01) enhancement of the cytotoxicity of DNR in 36 AML blast cell specimens. The degree of enhancement of accumulation/retention or cytotoxicity by CsA/Crem was approximately equivalent to that obtained with Crem alone. These studies indicate that Crem can reverse an MDR phenotype in patient AML blast cells. The equivalent degree of enhancement observed for Crem alone versus CsA/Crem also suggests that the Emax for DNR accumulation or retention was reached in AML cells by Crem alone (relative to 5 µmol/L CsA). Hence, synergism of CsA and Crem, indicated by the HL-60 studies, may permit the plasma concentrations obtained in recent phase I/II clinical trials (1 to 4 µmol/L CsA in Crem), to achieve maximal reversal of MDR in AML blast cells.