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Fig. 1
CD271 is upregulated by chemo- and MAPKi therapy. A, Melanoma cell lines were treated with carmustine (BCNU), cisplatin (CISP), and dacarbazine (DTIC) and CD271 expression was evaluated 48 hours later by FACS. B, Melanoma patient–derived cells were treated with cisplatin, PLX4032 (BRAFi), and/or MEK162 (MEKi). CD271 levels were analyzed by FACS 72 and 168 hours later. C, Paraffin blocks derived from 13 patients were collected before and after treatment with BRAFi + MEKi and stained with S100 and CD271 Abs. Scale bar, 100 μm. D, Quantification of melanoma CD271+ cells was performed by QuPath. The average of 10 areas was calculated and normalized to S100. Wilcoxon matched-pairs signed rank test was used (**, P < 0.01). E, Melanoma cells were cultured in 3D for 72 hours. Spheroids were treated with trypsin to obtain a single cell suspension and CD271 expression was evaluated by FACS. F, BRAF (WM793B) and NRAS (M130425) mutated targeted therapy sensitive (S) cell lines were cultured with increasing concentration of BRAFi or MEKi to make the cells resistant (R) in vitro. CD271 expression was evaluated by Western blot and FACS comparing resistant versus sensitive cells.