Onset of nilotinib resistance in K562 and K562-Dox cells coincides with ABCB1 overexpression; ABCB1 levels decrease upon continued high dose nilotinib exposure.

<p>Expression levels of ABCB1 mRNA/protein were assessed in (<b>a</b>) K562 and (<b>b</b>) K562-Dox nilotinib resistant cells. K562 cells were also assessed for <i>LYN</i> mRNA expression. mRNA expression represents the mean of six independent experiments performed in triplicate. The histograms shown are representative of typical protein expression levels. The blue and black lines represent resistant and control cells respectively, the grey filled histograms represent cells stained with isotype control and percentage of cells staining positive for ABCB1 expression is indicated (<b>c</b>) K562-Dox cells were stained with the fluorescent ABCB1 substrate rhodamine-123 and assessed for their ability to effectively efflux rhodamine-123 in the absence and presence of the specific ABCB1 inhibitor PSC-833. K562 cells were used as a negative control for ABCB1 efflux. The histograms shown are representative of typical efflux patterns. The black lines represent fluorescent substrate alone while the blue lines represent PSC-833 mediated inhibition of ABCB1. Statistical analyses compared mRNA levels (as a percentage of house keeping genes) in each resistant intermediate to control cells. Analyses were performed using unpaired Student’s <i>t</i>-test (Welch’s correction was applied for data groups with unequal SD) or Mann-Whitney Rank Sum test. Statistically significant <i>p</i>-values are denoted by carets (^) or asterisks (* <i>p</i><0.05; ** <i>p</i><0.01; *** <i>p</i><0.001). Error bars represent SEM. NIL = nilotinib.