Figure 1. ABC Transporter Upregulation Mediates THZ1 Resistance in NB and Lung Cancer Cells.

(A) Dose-responsecurvesforTHZ1^Sand THZ1^R NBcellstreated with THZ1 for72 hr(left). Forall dose-response curves, fractionsofviablecells relativeto DMSO-treated cells are shown. Western blot (WB) analysis of the indicated proteins in THZ1^S and THZ1^R cells treated with DMSO or THZ1 (right) (Kelly, 1 μM; NGP, LAN5, SK-N-BE2 [BE2], 300 nM) for 6 hr. (B) qRT-PCR analysis ofABC transporter expression in THZ1^S and THZ1^R cells (left). Expression was normalized to levels in THZ1^S cells, with GAPDH used as an internal control. WB analysis of ABC proteins in THZ1^S versus THZ1^R cells (right). (C).Dose-response curves for THZ1^S versus THZ1^R lung cancer cells treated with THZ1 for 72 hr (left). WB analysis of the indicated proteins in THZ1^S and THZ1^R cells treated with DMSO or THZ1 (NCI-H82, H3122, 300 nM; PC9, 1 μM) for 6 hr (right). (D) qRT-PCR (left) and WB (right) analyses of ABCG2 in THZ1^Sand THZ1^R cells. (E).Viability ofTHZ1^R Kelly NB cellstreated with THZ1 in combination with tariquidar at the indicated doses for72 hr (left). WB ofthe indicated proteins in THZ1^R Kelly cells treated with THZ1 at the indicated concentrations in combination with tariquidar (125 nM) for 6 hr (right). (F).Dose-response curves forTHZ1^SversusTHZ1^R H82 SCLC cells treated with THZ1 in combination with KO-143 (200 nM) ortariquidar (125 nM) for72 hr(left). Viability of THZ1^R PC9 cells treated with THZ1 in combination with the indicated doses of KO-143 for 72 hr (right). (G).Viability of THZ1^R Kelly cells expressing either a control shRNA or two individual shRNAs against ABCB1 and treated with THZ1 (left). WB ofthe indicated proteins in THZ1^R cells expressing either control (shCtrl) or ABCB1 shRNAs and treated with THZ1 alone (1 μM) or in combination with tariquidar (125 nM) for 6 hr (right). Error bars on all dose-response curves indicate mean values ± SD for three experiments. See also Figures S1-S3.