Fig 2: Broad transcriptional dysregulation of NK cells following perinatal MCMV infection.a Principal component analysis (PCA) was performed for all samples using the normalized, rlog-transformed count data, and the variability over the first (PC1) and second (PC2) principal components is shown on the x and y-axis, respectively. Ellipses around each experimental group denote a 95% confidence level. b Bar plots showing a selection of significantly enriched gene ontology (GO) terms in a list of genes that are deregulated in KLRG1+ NK cells following MCMV infection of newborn mice. c Heat map illustrating the differences in expression of transcription factors in KLRG1+ and KLRG1- NK cells from MCMV-infected mice and KLRG1- from mock-infected mice. d, e Expression of TCF-1 and BLIMP-1 was analyzed in KLRG1- NK1.1+ cells from mock-infected mice and KLRG1- and KLRG1+ NK1.1+ cells from MCMV infected mice 21 days p.i. by flow cytometry. Representative histograms and quantification of TCF-1 and BLIMP-1 expression by NK cells are shown. Mean values ± SD are shown (n = 5 mice per group, One-way ANOVA test was used). P values are shown. Source data are provided as a Source Data file.

Fig 3: OKI-179 alters acetylation in TCF1/7+ subsets. (A) T cells were gated based on CD45RA and CCR7 expression and assessed for TCF1/7 and LEF1 expression as shown. Histograms are representative of 6 individual patients receiving OKI-179. (B) Flow cytometry plots display changes in Ac-H3K27 in patient PBMCs taken at indicated time points after the initial dose of OKI-179 in comparison to TCF1/7 expression. (C) T cells were gated based on TCF1/7 and/or LEF1 expression as shown. MFI of Ac-H3K27 was quantified in TCF1/7+, LEF+TCF1/7-, and total CD4+ populations. Graphs show cumulative data of timepoints normalized to baseline controls within each patient; n=6 patients.

Fig 4: Assessment of exhaustion/dysfunction status of cancer-specific CD8 T cells within the blood via high-dimensional flow cytometry(A) Expression of key exhaustion markers (PD-1, TOX, and TCF1) in the indicated CD8 T cell subsets from before immunotherapy. Populations with less than 10 cells were excluded. Red boxes indicate MCPyV-specific CD8 T cells.|(B) Representative fluorescence-activated cell sorting (FACS) plot of CD8 T cells, showing tetramer binding versus PD-1, TOX, and TCF1 expression.(C) UMAP plots of CD8 T cells. Shown are MCPyV-specific T cells in patients who did (top) or did not (bottom) achieve a pCR.(D) Expression of 8 differentiation-related proteins using the same projection as in (C) and (E). Cancer-specific T cells are grouped in regions positive (orange/yellow) for exhaustion (PD-1, TOX, and EOMES), proliferation (Ki67 and HLA-DR), and effector function (granzyme B).(E) UMAP plots of CD8 T cells from blood. Cells are grouped, colored, and labeled using a priori-defined phenotypes.(F) Comprehensive representation of expression of differentiation proteins across T cell subsets that were a priori-defined (naive, effector, precursor exhausted, and terminally exhausted) compared with MCPyV-specific CD8 T cells before and after immunotherapy (bottom two rows, red box). As indicated in the legends, the size of each circle represents the proportion of cells positive for that marker via flow cytometry. The color represents the median fluorescence intensity of each protein in each population. Of note, the rare cancer-specific T cells identified in patients without a pCR were present at fewer than 10 cells per sample. Therefore, these results reflect only cancer-specific T cells in patients with pCRs.MFI, median fluorescence intensity; AF488, Alexa Fluor 488; PE, phycoerythrin; APC, allophycoerythrin, UMAP; uniform manifold approximation and projection; GZMB, granzyme B; Prec Ex, precursor exhausted; Term Ex, terminal exhausted; MCPyV, Merkel cell polyomavirus; EMRA, effector-memory RA positive; Activ/exh, activated/exhausted; tx, treatment. See also Figures S4–S6 and Table S1.

Fig 5: ACCi promotes T cell immunity against tumors(A–D) (A) GSVA of RNA sequencing (n = 4/4), (B) spectral flow cytometry (n = 4/4), and (C and D) representative images of TEM with indicated quantifications of OT-1 T cells cultured ± ACCi (n = 80/80, 20/20, and 126/126). Scale bars indicate 1 μm and 2 μm in vehicle and ACCi low-magnification images, respectively, and 0.5 μm in high-magnification images.(E–H) (E) Tumor growth (n = 4/8/8), (F) engraftment (n = 10/9), (G) polyfunctionality (n = 5/5), and (H) TCF-1 expression (9/9) of OT-1 T cells cultured ± ACCi, then infused into B16-OVA melanoma-bearing mice.(I–K) (I) p-ACC expression in indicated CD8+ T cell subsets from n = 7 normal donors, and (J) CD8+ and (K) CD4+ T cell subsets in response to vehicle or ACCi in n = 7 and n = 5 normal donors, respectively.Data are represented as mean ± SEM. Statistical analysis was performed using unpaired two-sided Student’s t test (D and F–H), log-rank (Mantel-Cox) test (E), and paired two-sided Student’s t test (I–K). *p < 0.05, **p < 0.01, ***p < 0.001, ****p < 0.0001.See also Figure S5.