Figure 4.

Monitoring of regulatory T cell (T_reg) T cell receptor (TCR) repertoire changes after T_reg infusion. (a) Comparative frequency analysis of T_reg clonotypes using high‐throughput TCR‐α chain sequencing. Scatterplots were generated to compare relative abundance of T_reg TCR sequences at different time‐points post‐T_reg transfer to their relative frequencies (in %) pretreatment (first row) or to the relative frequencies (in %) of the sequences within the expanded T_reg pool (second row). Each dot represents a unique sequence. Sequences that were found only in one of the two compared samples are shown as light grey symbols. (b) Correlation analysis of the T_reg repertoires. Shown are Pearson's correlation coefficients as measures of the strength of the association between the TCR repertoire at different time‐points post‐T_reg transfer versus the TCR repertoire pre‐T_reg therapy (grey bars) or versus the TCR repertoire of the expanded T_reg pool (black bars). Bar graph on left = patient 1, bar graph on the right = patient 2. (c) Tracking of the 20 most frequent T_reg clones of each sample over time. Every coloured line represents a particular clone. Plots on left: the 20 most abundant T_reg clones within the expanded T_reg pool followed throughout the indicated timepoints pre‐ and post‐adoptive transfer for patient 1 (first row) and patient 2 (bottom row). Middle column: the 20 most frequent T_reg clones pre therapy and their frequencies over time (first row, patient 1; bottom row, patient 2). Second and third rows show the same analysis for CD8⁺ T cells and CD4⁺ effector memory T cells [CD4⁺ T effector (T_eff)] isolated from patient 1. Plots on right: the 20 most abundant T_reg clones at the last visit post‐T_reg transfer and their frequencies before (first row: patient 1, last row: patient 2). The same analysis was conducted for CD8⁺ (second row) and CD4⁺ T_eff (third row) from patient 1.