Fig 2: T Cells Deficient in Co-stimulatory Molecules Are Prevalent in FL and Predict Patient Outcome(A) The tSNE map of CD27 or CD28+ cells from CD3+ concatenated file (31 FL and 6 tonsils). CD4+ or CD8+ T cells were gated based on expression levels from CD4 or CD8 staining.(B) Graphs showing the percentages of CD27- or CD28- or CD27-CD28- cells from CD3+ T cells from 31 FL specimens and 6 tonsil tissues.(C) Graphs showing the percentages of CD27-, CD28-, or CD27-CD28- cells from CD25-, CD25+, PD-1high, PD-1low, PD-1neg, TN, TCM, TEM, or TEMRA cells from 31 FL specimens.(D) Graphs showing the percentages of surface markers from CD27-, CD27+, CD28-, or CD28+ cells from 31 FL specimens.(E and F) Kaplan-Meier curves for overall survival of FL patients (n = 31) by the number of CD3+CD27-, CD3+CD28-, CD3+CD27-CD28- (E), CD4+CD25+, CD4+CD25+CD27-, CD4+PD-1+, or CD4+PD-1+CD27- (F) T cells. Bottom: p value indicates a comparison between TN and TEMRA. See also Figure S5.

Fig 3: TV003 Vaccination Induces DENV-Specific Memory B Cells (MBCs)(A) IgM-CD27+ MBCs were isolated 6 months after TV003 immunization (n = 11 subjects) or after primary DENV2 infection (n = 3 subjects), immortalized with BCL6 + Bcl-xL (6XL), and supernatants from polyclonal (50 cells/well) cultures of 6XL-immortalized cells were screened for IgG binding to DENV2 (the challenge virus). The frequency of DENV2-reactive MBC was estimated from the total number of cells screened and based on the average of 1–2 reactive clones in a positive polyclonal culture, as previously shown.35 Statistical significance was determined using the non-parametric Mann-Whitney test. Bar graphs indicate the mean with error bars representing standard deviation across all individual subjects (open circles for TV003 vaccinees and closed squares for rDEN2?30-infected subjects).(B) Frequency of DENV-specific MBC at time of DENV2 challenge (day 180) in TV003 vaccinees with non-sterile protection (i.e., boosted post-challenge DENV2 neutralizing antibody levels) and those with sterile protection from DENV2 infection (i.e., non-boosted DENV2 antibody levels). Boxplot areas show 25th-75th percentiles with the median as a thick line and whiskers indicating the 95% confidence interval and closed circles indicate each data point from each subject (labeled with de-identified study identifiers).(C) DENV2-reactive MBC cultures from 9 TV003-vaccinated subjects (subject IDs on x axis) were screened individually for binding to DENV1, DENV3, and DENV4 virions by ELISA and the proportions of monovalent (DENV2-only); bivalent (DENV2 + 1 other DENV); trivalent (DENV2 + 2 other DENVs); and tetravalent (DENV1–DENV4) are expressed as a percentage of the total response. At least 2 technical replicates were performed for all ELISA-binding measurements.