Fig. 2.

Treatment resistant BC cells overexpress ROR1 have a higher sphere forming efficiency: (a) Administration of therapy leads to the increase of ROR1 expression intensity in HER2⁺ treatment naïve and treatment exposed BC patient (p = 3.021e-02. unpaired two-sided t-test). (b) Tumor cells were dissociated from treatment naïve, sensitive and resistant HER2⁺ BC patients and equal numbers of cells were cultured in the supplemented CSC medium and stained for ROR1 expression in the spheres. Matched HER2⁺ BC patient's tissues were stained for ROR1 expression by immunofluorescence (Scale bar: 50 μM). (c) FACS analysis of ROR1⁺ and ROR1⁻ population in treatment naïve, sensitive and resistant cells from primary HER2⁺ BC patient tumors. (d) Equal number of sorted ROR1⁺ and ROR1⁻ cells from primary BC patients (n = 2) were cultured in the supplemented CSC medium and analyzed for sphere forming efficiency (p < .0002, paired two-sided t-test). (e) Comparison of epithelial-to-mesenchymal transition related genes showing mRNA expression from treatment naïve, sensitive and resistant ROR1⁺ and ROR1⁻ primary HER2⁺ BC patients cell fractions (results are mean fold change +/− SE, n = 3). (f) Comparison of ROR1 mRNA expression in primary HER2⁺ treatment naïve, sensitive and resistant BC patient tumors (results are mean fold change +/− SE, n = 3). (g) In vitro treatment of HER2⁺ primary tumors cells treated with T-DM1 (5 nM) and assessed for sphere forming efficiency (black arrowhead). Below is shown immunoblotting of ROR1 from each group of patients' protein samples (results are mean +/− SE, n = 3 run in triplicate; Scale bar: 100 μM).