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. Author manuscript; available in PMC: 2021 May 15.
Published in final edited form as: Oncogene. 2020 Nov 15;40(3):564–577. doi: 10.1038/s41388-020-01552-0

Figure 2.

Figure 2.

HHT inhibits protein synthesis and reduces the level of the RRM2 protein in Ewing sarcoma cell lines. (A) Comparison of protein expression (RPPA) in Ewing sarcoma cell lines treated with DMSO or HHT (10 nM) for 24 h. (B, C) EW8 and TC71 cell lines were treated with different concentrations of HHT for 24 h and then RRM2 levels were assessed by immunoblotting. (D) shRRM2 cells, grown in the presence or absence of doxycycline, were treated with HHT (10 nM) for 24 h. (E) EW8 and TC71 cells were treated with HHT (50 nM), MG132 (10 μM), or the drug combination for 6 h. Lysates were then collected for immunoblotting. (F) EW8 and TC71 were treated with HHT (50 nM) for 6 h and then DNA replication was quantified using EdU. (G) Ewing sarcoma cell lines were treated with HHT (10 nM or 50 nM) for 24 h. Cells were labeled with puromycin to quantify protein synthesis and then lysates were collected for immunoblotting. (H, I) Sarcoma and leukemia cell lines were treated with HHT (10 nM or 50 nM) for 24 h. Cellular lysates were then collected for immunoblotting. Protein loading for all of the immunoblots was normalized using cell number.