Fig. 4: PDL-1 and FLT3 increased expression during disease progression.

a The canonical pathway “PD-1, PD-L1 cancer immunotherapy pathway” according to IPA. Green indicates downregulated genes, red indicates overexpressed ones in the comparison T3 vs. T1 MEP_2 cluster. Blue genes are predicted inhibited, while orange indicates predicted activated ones. b Includes dot plots representing the expression of genes involved in the PD-1/PD-L1 pathway in cells belonging to the MEP_2 cluster. c, d The percentages of activated CD4⁺ and CD8⁺ T cells assessed in HD (n = 7), T1, T2, and T3 samples, respectively. T cell activation was evaluated by flow cytometry; the gating strategy used to identify activated subpopulations, defined as CD38⁺HLA-DR⁺, is reported in Supplementary Fig. 10. Percentages are reported as mean ± standard error of the mean (s.e.m.). In e, cells in the tSNE plot are colored in red scale according to FLT3 expression. Dot plot in f represents the distribution of FLT3 expression values. Percentages represent the frequency of FLT3 expressing cells in each sample. In g, cells in the tSNE plot are colored based on AUC values for VELTEN_FLT3_SATB1_IND1 gene module¹⁸. h The distribution of AUC values. Dots are colored based on the sample name. Based on this distribution, we identified cells where the gene signature is active with an AUC > 0.29. Two-sided Fisher’s exact test was used to compare the proportion of cells expressing FLT3 or with active VELTEN_FLT3_SATB1_IND1 gene module in three timepoints (****p ≤ 0.0001).