Method Profile
Combine short-term autologous tumor killing with receptor-repertoire and single-cell analysis of lymphocyte states.
Functional comparison of expanded patient TILs against matched tumor and normal epithelial targets.
Twelve lung-cancer organoid cultures from eight adenocarcinoma and four squamous-cell carcinoma patients; distinct patient subsets used for repertoire, killing and single-cell experiments.
Expanded autologous TILs at E:T 5:1 or 10:1 for 48 hours, with images every two to three hours and orthogonal injury/viability assays. Separate day-0/day-14 receptor analysis and five-donor single-cell profiling.
Functional readouts and controls
CFSE/DAPI target-cell flow cytometry, caspase-3/7 imaging, LDH and cytokines; organoids alone, blood mononuclear cells, matched normal organoids and selected cross-patient targets.
Immune profiling
TCR clone diversity/overlap and single-cell transcriptomes. Cytotoxic and exhaustion-associated signals are assessed together; pathway enrichment is not a blocking experiment.
Keep patient, technical-replicate and profiling denominators separate. Compare functional autologous selectivity and profile changes without inferring lineage transitions or pathway causality from cluster enrichment.
TCR comparison two patients; representative killing includes three technical replicates and a separate two-replicate blood-cell control; single-cell analysis five independent patients.
Target organoids alone, blood mononuclear cells, matched normal and other-patient organoids; before/after expansion or coculture as specified.
Selected autologous combinations showed target injury and greater LDH killing than normal or other-patient controls. GZMB increased but GZMH did not; exhaustion-associated transcripts also increased. Five-donor population comparisons used one-tailed paired tests.
Primary study comparisons of the stated models, treatments and measurements.
Patient-matched lung cancer organoids and tumor-infiltrating lymphocytes supported short-term killing assays and single-cell profiling. Cytotoxic changes appeared alongside exhaustion-associated signals, while pathway enrichment remained associative and clinical response prediction was untested.
