Extended Data Fig. 10: Compensatory metabolism upon IDH2 inhibition influences histone modification and memory T cell differentiation.

a, KDM5 activating potential represented by the ratio of fold increase in K_m value for α-KG versus the sum of fold increased IC₅₀ concentrations of inhibitory metabolites (2-HG, succinate, fumarate and malate), calculated as: (concentration α-KG/K_m α-KG)/((concentration 2-HG/IC₅₀ 2-HG) + (concentration succinate/IC₅₀ succinate) + (concentration fumarate/IC₅₀ fumarate) + (concentration malate/IC₅₀ malate)). Ratio based on cellular metabolite concentrations calculated from absolute metabolite quantifications. (n = 6 biological replicates, pooled data from 2 independent experiments). b, Immunoblot quantification of H3K4me3 in DMSO- and IDH2i-treated CD8⁺ T cells, supplemented with α-KG and/or KDM5 inhibitor. (n = 2 biological replicates, pooled data from 2 independent experiments). c,d, Immunoblot (c) and quantifications (d) of H3K27ac and H3K14ac in DMSO or IDH2i-conditioned CD8⁺ T cells, treated with etomoxir (FAOi) and supplemented with acetate. (n = 3 biological replicates). e, Immunoblot quantification of H3K27ac and H3K14ac in DMSO or IDH2i-conditioned CD8⁺ T cells, treated with ACLYi. (n = 3 biological replicates). f, Percentage of CD62L⁺ T cells in DMSO or IDH2i-conditioned CD8⁺ T cells, treated with ACLYi. (n = 5 biological replicates, pooled data from 2 independent experiments). g,h, Immunoblot (g) and quantification (h) of H3K27ac in DMSO, IDH2i-, DMSO+HATi-and IDH2i+HATi-conditioned CD8⁺ T cells. (n = 3 biological replicates, pooled data from 3 independent experiments). i, CD62L expression in DMSO, IDH2i-, DMSO+HATi- and IDH2i+HATi-conditioned cells. (HATi = histone acetyltranferase p300/CBP inhibitor, 8 μM C646). (Representative blot and histogram from 3 independent experiments with 3 biological replicates). j, Tumour growth curve of B16-OVA tumours from mice treated with either DMSO, IDH2i-, DMSO+HATi- or IDH2i+HATi-conditioned OVA-specific CD8⁺ T cells. (n = 13 (IDH2i), n = 14 (DMSO+HATi, IDH2i+HATi) and n = 15 (DMSO) biological replicates per group, pooled data from 2 independent experiments). k, Pluripotent naive CD8⁺ T cells are characterized by low metabolic activity and permissive H3K4me3 deposition at pro-memory genes, including Sell, Ccr7 and Tcf7. After activation, T_E cells increase glutamine metabolism along both the oxidative and reductive pathways (represented by two red arrows on the left panel of the figure). RC generates citrate and acetyl-CoA, which is a key metabolite supporting cell growth and function. This specific T_E cell metabolism creates a unique metabolite composition (represented by balanced levels of α-KG on one side and succinate, fumarate, malate and 2-HG on the other side) enabling the activity of the α-KG-dependent histone demethylase KDM5. KDM5-mediated demethylation of H3K4me3 in activated T cells induces a repressive chromatin state at pro-memory genes, which prevents the maintenance of pluripotency and facilitates terminal effector differentiation. Blocking RC through inhibition of IDH2 does not hinder T cell proliferation and function, as compensatory fatty acid oxidation can refuel acetyl-CoA pools, with the support of glutamine anaplerosis, which is now redirected entirely in the oxidative branch of the TCA cycle (represented by the larger red arrow on the right panel of the figure), altogether enhancing cellular OCR. This leads to a perturbation of the metabolite balance by increasing the levels of the TCA intermediates succinate, fumarate, malate and 2-HG (represented by a shift in the balance upon IDH2 inhibition). The accumulation of these inhibitory metabolites impairs KDM5 demethylase activity, and elevated H3K4me3 is likely to foster a permissive chromatin state at pro-memory genes, allowing for the maintenance of pluripotency and memory differentiation. Data represent mean ± s.e.m. and were analysed by unpaired two-tailed Students t-test (a,j) or two-way ANOVA using the original FDR test of Benjamini and Hochberg (b,d–f,h). Only relevant statistical comparisons are shown. For gel source data, see Supplementary Fig. 1.

Source data