Fig. 4: EOS301984 inhibits ENT1 to restore T cell expansion in high-adenosine environments.

a, Percent inhibition of ENT1 in a transporter assay by dose ranges of EOS301984 or dilazep. Mean values from technical replicates (n = 2 and 4, respectively) are displayed; error bars, s.d. b, Percent inhibition of adenosine uptake by T cells preactivated in the presence of EOS301984 or dilazep. Mean values from n = 3 donors are displayed; error bars, s.d. c, Rescue of T cell proliferation by EOS301984 (300 nM) in the presence of adenosine (100 µM). d, Rescue of T cell proliferation by EOS301984 and dilazep in the presence of ATP (100 µM). Mean values from technical duplicates, representative of nine donors. e, Pyrimidine metabolite levels in activated human T cells treated with adenosine (50 µM) and EOS301984 relative to DMSO-treated cells. f, Seahorse Mito Stress Test analysis of T cell oxygen consumption rate (OCR) following 24 h CD3/CD28 activation in the presence or absence of adenosine (50 µM) and EOS301984 (100 nM). Mean values from n = 4 technical replicates are displayed; error bars, s.d. Representative example of n = 3 donors. g, Summary of basal and maximal OCR from experiments performed as in f. h, Seahorse Mito Stress Test analysis of T cell OCR following 24 h CD3/CD28 activation in the presence of adenosine (50 µM) and inupadenant (300 nM), as indicated. i, CMV pp65 antigen recall assay performed in the presence of exogenous ATP as a source of adenosine and the indicated reagents. Representative of three donors. j, Summary data from i. k, FluoroSpot analysis of IFNγ production from PBMC cultures as performed in i. l, Antigen-specific killing activity of PBMC cultures as performed in i and normalized to the levels observed in the absence of ATP as a reference control. Symbols represent each of the n = 3 donors (biological replicates); bars are group means; error bars, s.d.; P values are from two-way ANOVA with Tukey’s multiple comparisons test (c,e,g–h,j–l).

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