Fig 2: HBV-specific as well as global IL-27 expressing TFH cells were augmented in PD-1+ fraction of cells rather than PD-1-. (A) HBV-specific as well as global cytokine producing TFH cells were assessed in PD-1+ and PD-1- TFH cell population in CHB and HC-vacc (n=14 subjects in each group). (B) To identify the functional status of PD-1+ TFH cells, expression of CD69, an activation marker, was analyzed on both PD-1+ and PD-1- cells after HBs, HBc and PMA/ionomycin stimulation. Statistics were calculated using 2-tailed non-parametric, Mann-Whitney U test. Bars graphs indicate median with range. * indicates p < 0.05 and ***p < 0.001.

Fig 3: Scatter plot showing the distribution of the MFI of PD1 on CD4+CD25+ before and after vaccination.

Fig 4: Functional inhibition of LAG3/MHC-II and PD1/PD-L1 interactions.Cell-based assessment of T cell activation measured by induction of an NFAT response element-driven luminescent reporter gene. Relative light units (arbitrary scale) are reported. a Relief of LAG3/MHC-II-mediated suppression. Standard errors of the mean are indicated. Approximate half-maximal stimulation concentrations: CB213: 20 nM, Lg-Lg-Pd (LAG3-LAG3-PD1): 22 nM, relatlimab: 9.7 nM, aLAG3 17B4 (anti LAG3 mouse monoclonal): 1.6 nM. MSA-MSA-MSA is three interlinked VH recognising mouse serum albumin as a negative control. b Relief of PD1/PD-L1-mediated suppression. Standard errors of the means are indicated. Approximate half-maximal stimulation concentrations: CB213: 105 nM, Lg-Lg-Pd (LAG3-LAG3-PD1): 115 nM, nivolumab: 6 nM, DART (MGD-013): 1.4 nM. MSA-MSA-MSA is three interlinked VH recognising mouse serum albumin as a negative control.

Fig 5: LAG3, PD1, and NKG2D expression in breast cancer from TCGA data. (A, B) Expression of LAG3, PD1, and NKG2D on DNT cells and its ligands on MDA-MB-231 and SK-BR-3 cells. Experiments were repeated twice with similar results. (C) Box-whisker plots showing the expression of LAG3 (lag3) in subgroups of breast cancer samples from the TCGA database, which was tested by UALCAN. (D) Dataset analyzed by TIMER 2.0 revealed the correlation of lag3 expression with the overall survival of different breast cancer subtype patients. (E) Spearman's correlation of LAG3 (lag3), PD1 (pdcd1), and NKG2D (klrk1) with immune cell infiltrations across breast cancer (TIMER). p < .05 was considered statistically significant. DNT, double negative T; LAG3:, lymphocyte-associated gene 3; PD1, programmed cell death protein 1; TIMER, tumor immune estimation resource