(A) Schematic of experimental and computational pipeline. (B) cf-RNA yields and average fragment lengths from groups A, B, and C. (C) QC pipeline for screening degraded samples (3′ bias), ribosomal content, and DNA contamination. (D) Volcano plot:differentially expressed genes between ICI-treated patients with cancer (A+B+C, n = 22) and individuals acting as healthy controls (n = 30) (Benjamini-Hochberg –adjusted [BH-adjusted] P < 0.05), showing a higher number of differential genes. (E) Volcano plot: group C versus A (BH-adjusted P < 0.05) for cardiac pathways, with gene-type charts showing majority protein-coding genes. (F) IPA pathway analysis of upregulated differentially expressed genes in group C versus A patients. (G) Box plots: counts per million–trimmed mean of M values–normalized (CPM-TMM-normalized) counts of MYH6 and CCL5 cf-mRNA in group C versus A and B. (H and I) Box plots: CD4⁺/CD8⁺ T cell and Temra CD4⁺/CD8⁺ T cell proportions from single-cell RNA-Seq gene panels of ICI-treated patient PBMCs (3). (J and K) Heart and cardiomyocyte signature scores in group C versus A and B. *P < 0.05, **P < 0.01 by Mann-Whitney U test comparing groups. (L) Venn diagram: 6 genes (3 cardiac, 3 immune) were identified from C versus A and C versus B DEGs but not overlapping with B versus A DEGs to constitute an ICI-m–specific diagnostic panel. (M) ROC curves with metrics for C versus A (n = 10 vs 5), C versus B (n = 10 vs 7), and B versus A (n = 7 vs 5) using unsupervised panel (red) and ICI-m classifier (green). (N) ROC validation: both panels on separate group C (n = 6) versus groups A (n = 5) and B (n = 7).