Fig. 2: Active T-PLL samples present a significant enrichment of pathways involved in MYC signaling, cell cycle regulation, and energy metabolism.

A Volcano plots: pairwise differential gene expression analyses between T-PLL cells from active vs indolent stages. Genes with an absolute log2 expression fold-change ≥0.5 and adjusted p < 0.05 were considered significantly different. Colored points indicate differentially expressed genes (DEGs, n = 24 genes) detected in at least 5/11 sample pairs. Positive fold-changes indicate elevated expression in active-disease T-PLL. sign.: significant. Bar chart: number of recurring DEGs. A significant proportion of DEGs were detected across multiple sample pairs (p < 0.0001, permutation test, B = 10,000, one-sided). B Differential expression heatmap of recurrent DEGs. Color indicates the log2 fold-change between active and indolent T-PLL cells (red: upregulated in active T-PLL, white: no change, blue: downregulated in active T-PLL). Font of gene labels indicates the number of patients that showed deregulation in the same direction (bold black: DEG in 5 pairs, regular black: DEG in 4 pairs, regular gray: DEG in 3 pairs). C Experimental design for the identification of shrinking and expanding clusters. For each longitudinal sample pair, SNN-based clustering on integrated T-PLL cell gene expression was performed³⁴. The relative contributions of each time point to the individual clusters were quantified and compared to the global indolent/active cell ratio using permutation testing (B = 10,000, two-sided). Clusters that significantly differed from the overall sample distribution (false discovery rate (FDR) < 0.05, observed log2 difference in proportion ≥1) were identified as expanding or shrinking. Remaining clusters were labeled as stable. Created in BioRender. Herling, M. (2026) https://BioRender.com/eih43igD GSEA enrichment of most abundant HALLMARK pathways between expanding and shrinking clusters³⁶. Dot colors represent the Normalized Enrichment Score (NES, red: enrichment in expanding clusters, blue: enrichment in shrinking clusters). Dot sizes indicate the FDR. E Total number and predicted type of short variants derived from WGS. F Mean variant allele frequencies (VAFs) of functional coding variants in genes previously identified as recurrently mutated in T-PLL (n = 6 T-PLL samples)¹³. G Dot plot comparing WGS-derived VAFs of ATM mutations and copy number alterations (CNAs) between three longitudinal T-PLL sample pairs. H WGS-derived CNAs of MYC in three sequential T-PLL patients. Source data and complete summaries of statistical analyses are provided in the Source Data file.