Mass cytometry defines differences in lymphocyte populations between responders and nonresponders, before treatment. Patient PBMC samples were assessed before cycles 1, 2, and 6 by mass cytometry. Using viSNE analysis, CD45⁺ cells were clustered and lymphocyte populations defined by lineage-marker–informed manual gating. (A) Representative viSNE plot depicting the major lymphocyte populations identified. (B) Representative density plot of 1 patient, over time. Numbers indicate the frequency of cells within the gate. (C) Summary data showing the frequency of NK cells between responders (R, n = 20; red) and nonresponders (NR, n = 13; blue). (D) Summary of total NK cells as determined by ALC and percentage of NK cells. (E) Summary expression of PD-1 and GzmB on NK cells from R (red) vs NR (blue). (F) Summary data of total CD8⁺ T cells from R (red) vs NR (blue). (G-H) FlowSOM was performed on CD8⁺ CD3⁺ T cells; 10 metaclusters were identified. (G) Heat map depicting the relative expression of the markers used to generate the FlowSOM clusters. Metaclusters 2 and 3 are within boxes. (H) Total CD8⁺ T cells within each Metacluster from R (red) vs NR (blue) at cycle 1. Data were compared in a type 3 mixed-effects model (C-F) or by 2-way analysis of variance (H). *P ≤ .05; **P ≤ .01; ***P ≤ .001. n.s. not significant.