TNF-α is not required for birinapant response. (A) TNF-α expression by quantitative real-time PCR from 12 responsive PDX samples (birinapant IC₅₀ ≤ 100 nM) and 22 nonresponsive PDX samples (birinapant IC₅₀ > 100 nM). (B) Quantification of TNF-α by ELISA from cell lysates in WT and CRISPR-generated TNFko PDXs (3 independent experiments performed in duplicates). (C) Validation of TNFko MSCs by copy number variant quantitative PCR (2 experiments performed in triplicates). (D) Birinapant response curves for WT and TNFko PDX cells cocultured with TNFko MSCs (3 independent experiments performed in triplicates). (E) Gene expression of death receptor ligands and receptors for 17 primary B-ALL samples. (F) Birinapant response curves for PDX samples treated with neutralizing antibodies against lymphotoxin (LT; 0.5 μg/ml), TRAIL (10 ng/mL), and FasL (10 ng/mL) (3 independent experiments performed in triplicates). All quantifications are mean ± SEM. R, relapse.