TCR clonotype expansion and tumor-specific T-cell responses are induced during therapy. (A) Scatterplot relating average change in frequency of multiplet TCR clones from pretreatment (pre Tx) to day 8 (both sites combined) to percentage distant tumor reduction. (B) Examples of 4 TCR clonotypes and the location of cells with those clonotypes in the T-cell UMAP. (C) Phenotype distribution of TCR clones that increased by week 6 after treatment (left) and those that decreased by week 6 (right). (D) Schematic workflow of experimental in vitro immune response assay. T-cell reactivity against tumors were measured before treatment and on treatment (weeks 1, 3, 6, and 12). (E) Representative flow cytometry plots measuring autologous tumor-reactive granzyme B–positive memory (CD45RO⁺) CD8 T-cells under the indicated conditions. Most granzyme B–positive/CD45RO⁺ CD8_M T-cells were also Ki-67-positive and perforin-positive (mean, 67%; range, 2.3%-96%). (F) Unsupervised clustering of patients by percentage distant tumor reduction and percentage change in tumor-reactive activated memory CD4 and CD8 T-cells at week 3. PBMC, peripheral blood mononuclear cells. max, maximum; min, minimum.