Extended Data Fig. 8: Depletion of ELP3 or CTU2 strongly reduces tumour growth of melanoma tumours with acquired drug resistance in a HIF1α-dependent manner.
From: Codon-specific translation reprogramming promotes resistance to targeted therapy
a, Eight-week-old NOD/SCID mice were injected with A375 drug-resistant cells (RES; vemurafenib 0.3 μM) depleted of ELP3 using an inducible shRNA against ELP3 or control. After 10 days, all mice were treated daily with IPTG to induce depletion for two weeks. Mice were also daily treated or not with vemurafenib (25 mg kg−1) for two weeks. Indicated protein levels were assessed in tumours after death by western blot. n = 3 independent animals. b, Eight-week-old NOD/SCID mice were injected with A375 drug-resistant cells (vemurafenib 0.3 μM) depleted of CTU2 expression using an inducible shRNA against CTU2 or control. After 10 days, mice were treated daily with IPTG to induce depletion for two weeks. Mice were also daily treated or not with vemurafenib (25 mg kg−1) for two weeks. Indicated protein levels were assessed in tumours after death by western blot. n = 3 independent animals. c, Indicated primary melanoma lines were depleted of HIF1α by esiRNA and treated or not with 15 μM vemurafenib for 24 h. Cell death was quantified by FACS. n = 2 independent experiments, data are mean + s.d. d, A2058 cells were depleted of HIF1A and treated for 24 h as indicated. Cell death was quantified by FACS. n = 2 independent experiments, data are mean + s.d. e, f, MM029 (e) or A2058 (f) cells expressing the empty vector (Empty), HIF1AWT–Flag or HIF1AMut–Flag constructs were depleted of ELP3 or CTU1 using shRNA and treated as indicated for 24 h. Cell death was quantified by FACS. n = 2 independent experiments, data are mean + s.d. g, NOD/SCID mice were injected with A2058 cells expressing the HIF1AWT–Flag or HIF1AMut–Flag constructs and infected with inducible shRNA control or ELP3 shRNA (1 × 106 cells per condition). After 10 days, mice received daily IPTG (100 μl; 25 mM) and vemurafenib (25 mg kg−1) treatment for 26 days. ELP3 and Flag–HIF1α protein levels were detected after death in three different tumours by western blot.
