Fig. 7

Multiplex immunofluorescence (mIF) validation of TOX2, RUNX3 and PRL-3 expression in an independent cohort of clinical samples from 42 NKTL patients (NUH). A Representative images of protein expression of CD3, RUNX3, TOX2 and PRL-3 in NKTL patient samples using mIF method. Left columns represented protein expression of CD3 (membrane, magenta), PRL-3 (cytoplasm, red), RUNX3 (nuclear, cyan), and TOX2 (nuclear, green) in NKTL with multiplexed immunofluorescence staining. Right columns indicated the corresponding image analysis masks. Double positive cells were in white; single positive cells were marked in the corresponding immunofluorescence staining color; while negative cells were in blue. The scale bars indicate 50 µm. B Correlation between TOX2 expression with PRL-3 expression in NKTL patients (n = 42) was determined by mean intensity of staining quantified with Visiopharm program. A significant positive correlation was determined by Pearson's p < 0.001, and R = 0.65. C Correlation between TOX2 expression with RUNX3 expression in NKTL patients (n = 42) was determined by mean intensity of staining quantified with Visiopharm program. A significant positive correlation was determined by Pearson's p = 0.001, and R = 0.50. D Kaplan–Meier analysis was performed on the overall survival between patients (n = 30) expressing higher TOX2 expression (TOX2-High, ≥ median expression) and lower (TOX2-Low, < median expression). E Kaplan–Meier analysis was performed on the overall survival between patients (n = 30) expressing higher PRL-3 expression (PRL-3-High, ≥ median expression) and lower (PRL-3-Low, < median expression). In D and E statistical significance (p) was evaluated by Log-rank test and p < 0.05 was considered as significant. HR: hazard ratio