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. Author manuscript; available in PMC: 2023 Dec 2.
Published in final edited form as: Cancer Discov. 2023 Jun 2;13(6):1478–1497. doi: 10.1158/2159-8290.CD-22-0825

Figure 1: (R)-2HG likely has other functionally relevant targets besides TET2 in IDH-mutant AML.

Figure 1:

(A) Pie-charts illustrating the relative frequencies of TET2 (left) and IDH (right) mutations in clonal myeloid disorders (see Supplementary Table S1 for details). Low-grade clonal myeloid disorders: clonal hematopoiesis of indeterminate potential (CHIP), idiopathic cytopenia of undetermined significance (ICUS), refractory anemia (RA), refractory anemia with ring sideroblasts (RARS), and myeloproliferative neoplasms (MPN). Intermediate-grade clonal myeloid disorder: refractory cytopenia with multilineage dysplasia (RCMD). High-grade clonal myeloid disorders: refractory anemia with excess blasts (RAEB) and acute myeloid leukemias (AML) secondary to myelodysplastic disorder (MDS-AML) or myeloproliferative neoplasm (MPN-AML). (B) Immunoblot analysis of IDH1 expression in TF-1 cells expressing empty vector or HA-tagged R132H-mutant IDH1 (IDH1R132H). (C) GC-MS analysis of intracellular concentrations of 2HG in TF-1 cells expressing empty vector or IDH1R132H. Shown are mean values of duplicate experiments. (D) Proliferation under cytokine-poor conditions of TF-1 cells expressing empty vector or IDH1R132H. (E) Total 5hmC content in the TF-1 cells from (B-D) collected on day 0 of cytokine withdrawal, as measured by LC-MS. The average (± SD) of two independent biological replicates is shown. (F) GC-MS analysis of intracellular concentrations of 2HG under the following conditions: cultured in DMSO (0 μM R-2HG); cultured in 100 μM TFMB-(R)-2HG (100 μM R-2HG); cultured in 250 μM TFMB-(R)-2HG (250 μM R-2HG); and cultured in 500 μM TFMB-(R)-2HG (500 μM R-2HG). Shown are mean values of duplicate experiments. (G) Total 5hmC content in TF-1 cells following pre-treatment for 21 days with the indicated concentrations of TFMB-(R)-2-HG or DMSO-control (0 μM (R)-2HG), as measured by LC-MS. The average (± SD) of two independent biological replicates is shown. (H) Proliferation under cytokine-poor conditions of the TFMB-(R)-2-HG-treated TF-1 cells from (F-G). Representative results from at least three independent experiments are shown.