1,5-Pentanediamine detected in CRKP-colonized patients impairs CD19 CAR-T cell function in vitro

CD19-directed chimeric antigen receptor T cell (CAR-T) therapy is effective in relapsed or refractory B cell malignancies, but responses remain heterogeneous. Carbapenem-resistant Klebsiella pneumoniae (CRKP) colonization is common in heavily treated hematology patients. 1,5-Pentanediamine (PDA; cadaverine), a microbiota-associated metabolite produced by several Enterobacteriaceae, was detected in serum from CRKP-colonized patients. Its effects on CAR-T cells are unknown. Serum PDA was quantified by liquid chromatography–tandem mass spectrometry in 30 CRKP-colonized patients without documented bloodstream infection at sampling. Healthy-donor-derived CD19 CAR-T cells were exposed to PDA in vitro (0–12 mM; 9 mM for most functional assays). Metabolic activity, apoptosis, phenotype, target-cell killing, degranulation, cytokine secretion, and cytotoxic-molecule production were assessed. RNA sequencing, RT-qPCR, and PD-1 blockade were performed. PDA was detectable in all 30 patients. Without a non-colonized comparator cohort, these data demonstrate detectability but not CRKP-specific source attribution. At millimolar concentrations, PDA reduced CAR-T cell metabolic activity, increased apoptosis, upregulated activation and checkpoint-associated inhibitory markers, altered the CD4/CD8 distribution, and increased the regulatory T cell frequency. PDA-treated CAR-T cells showed reduced NALM-6 killing and lower cytokine, perforin, and granzyme B output despite preserved CD107a degranulation. Transcriptomic analysis showed enrichment of cell cycle, apoptosis, and stress-related pathways, suppression of immune pathways, and exhaustion-associated transcriptional features. PD-1 blockade alone did not restore function under the tested conditions. Acute millimolar PDA exposure produced a dysfunctional CAR-T cell state with impaired survival and effector output but preserved degranulation. Its physiological relevance requires validation in chronic low-dose models, controlled clinical cohorts, patient-derived CAR-T cells, and additional target-cell systems.