Figure 6. FGF/RAS-dependent resistance is reversible.
(A) Removal of HRAS(G12V) restores LDE225 sensitivity. SMB(HRAS) cells were infected with lentivirus expressing Flp or tdTomato (control), then assayed by relative survival at indicated LDE225 concentrations (mean ± s.e.m., n = 3).
(B–C) Shh signaling assayed by Immunoblot and qRT-PCR. mean ± s.d., n = 3, ** p < 0.01, *** p < 0.001, one-way ANOVA, Bonferroni correction.
(D–E) SMB21(GF) and SMB55(GF) cells were cultured in media without GF for 3 weeks. (D) Relative survival was then analyzed in indicated LDE225 concentrations (72 hrs). (mean ± s.e.m., n > 9). (E) qRT-PCR analysis of Atoh1, Gli1. mean ± s.d., n = 6, *** p < 0.001, one-way ANOVA with Bonferroni.
(F) Relative survival for SMB21 and SMB21(HRAS) cells treated with MEK inhibitor PD325901 (120 hrs) (mean ± s.e.m., n = 4).
(G) MEK inhibitor PD325901 (0, 10, 100, 1000 nM for 24 hrs) reduce phospho-Erk analyzed by immunoblot.