Extended Data Fig. 4: Status of TAGLN2 in OvCa-infiltrating CD8⁺ T cells.

a, Volcano plot of differentially expressed genes in dysfunctional tumour-specific CD8⁺ T cells (TCR_TAG) compared to non-tumour-specific CD8⁺ T cells (TCR_OT1) from a mouse autochthonous liver tumor model (GSE126974). Selected differentially expressed genes with an adjusted P values < 0.05 and Log₂ Fold Change > 1 or −1 are highlighted. (Two-sided Student’s t-test, P < 0.05, FC > 2). b, Relative expression dot plots of the indicated genes in different CD8⁺ T cell populations infiltrating human HGSOC tumors. The colour of each dot represents the average normalized expression from high (red) to low (blue). The size of each dot represents the percentage (pct) of positive cells for each gene. c, FACS analysis of TAGLN2 expression by CD8⁺ T cells present in peripheral blood of cancer-free women (n = 14), malignant ascites (n = 15), or peripheral blood (n = 8) of HGSOC patients. d, TAGLN2 transcripts in the indicated CD8⁺ T cell populations from human peripheral blood mononuclear cells (PBMC). The resulting transcript expression values calculated as normalized transcript per million (nTPM), resulting from the internal normalized pipeline (The Human Protein Atlas; proteinatlas.org). e, TAGLN2 expression was quantified in the indicated CD8⁺ T cell populations from malignant ascites (n = 15). f, Correlation analysis for IFNG, TNFA or GZMB versus TAGLN2 mRNA in CD8⁺ T cells from ascites. Data were normalized to ACTB (n = 15). g, Correlation of IFN-γ concentration versus levels of TAGLN2 in the indicated CD8⁺ T cell subsets present in the ascites (n = 14). h, Naïve peripheral CD8⁺ T cells from cancer-free women were activated via CD3/CD28 stimulation for 32 h and then incubated for 16 h with increasing amounts of HGSOC ascites supernatants (n = 5). Expression of TAGLN2, IFNG, and GZMB was assessed by qRT-PCR. Data were normalized to ACTB. i, FACS histograms and quantitative analysis of TAGLN2 expression in the indicated CD8⁺ T cells from peritoneal lavage of cancer-free female mice (n = 8; dotted lines) or mice bearing ID8-Defb29/Vegfa OvCa (n = 6; solid lines). Grey peaks denote isotype control staining. j,k, FACS analysis for CD44 (j) and Ki-67 (k) expression in the indicated T cells from the ascites of female mice bearing ID8-Defb29/Vegfa OvCa (n = 6 per group). l-n, Frequency of CD44⁺IFN-γ⁺ (l), CD44⁺TNF-α⁺ (m), and CD44⁺GZMB⁺ (n) cells within the indicated CD8⁺ T cells from the same mice described in (j) and (k). Data are presented as mean ± s.e.m. c,i-k, Two-tailed unpaired Student’s t-test. e,h, One-way ANOVA with Tukey’s multiple comparisons test. f,g, Spearman’s rank correlation test, Spearman coefficient (r) with exact P-value (two-tailed). l-n, Two-tailed paired Student’s t-test. Exact P-values are shown. The ‘n’ values represent biologically independent samples. gMFI, Geometric mean fluorescence intensity.

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