Tolerance to self-antigens prevents the elimination of cancer by the immune system1,2. We used synthetic chimeric antigen receptors (CARs) to overcome immunological tolerance and mediate tumor rejection in patients with chronic lymphocytic leukemia (CLL). Remission was induced in a subset of subjects, but most did not respond. Comprehensive assessment of patient-derived CAR T cells to identify mechanisms of therapeutic success and failure has not been explored. We performed genomic, phenotypic and functional evaluations to identify determinants of response. Transcriptomic profiling revealed that CAR T cells from complete-responding patients with CLL were enriched in memory-related genes, including IL-6/STAT3 signatures, whereas T cells from nonresponders upregulated programs involved in effector differentiation, glycolysis, exhaustion and apoptosis. Sustained remission was associated with an elevated frequency of CD27+CD45RO- CD8+ T cells before CAR T cell generation, and these lymphocytes possessed memory-like characteristics. Highly functional CAR T cells from patients produced STAT3-related cytokines, and serum IL-6 correlated with CAR T cell expansion. IL-6/STAT3 blockade diminished CAR T cell proliferation. Furthermore, a mechanistically relevant population of CD27+PD-1CD8+ CAR T cells expressing high levels of the IL-6 receptor predicts therapeutic response and is responsible for tumor control. These findings uncover new features of CAR T cell biology and underscore the potential of using pretreatment biomarkers of response to advance immunotherapies.

Key Points Ibrutinib treatment of CLL enhances the generation of CAR T cells for adoptive immunotherapy. Concurrent ibrutinib therapy improves the engraftment and therapeutic efficacy of anti-CD19 CAR T cells in mouse models.

The success of chimeric antigen receptor (CAR)-mediated immunotherapy in acute lymphoblastic leukemia (ALL) highlights the potential of T-cell therapies with directed cytotoxicity against specific tumor antigens. The efficacy of CAR T-cell therapy depends on the engraftment and persistence of T cells following adoptive transfer. Most protocols for T-cell engineering routinely expand T cells for 9 to 14 days. Because the potential for engraftment and persistence is related to the state of T-cell differentiation, we hypothesized that reducing the duration of culture would limit differentiation and enhance the efficacy of CAR T-cell therapy. We demonstrated that T cells with a CAR-targeting CD19 (CART19) exhibited less differentiation and enhanced effector function when harvested from cultures at earlier (day 3 or 5) compared with later (day 9) timepoints. We then compared the therapeutic potential of early versus late harvested CART19 in a murine xenograft model of ALL and showed that the antileukemic activity inversely correlated with culture time: day 3 harvested cells showed robust tumor control despite using a 6-fold lower dose of CART19, whereas day 9 cells failed to control leukemia at limited cell doses. We also demonstrated the feasibility of an abbreviated culture in a large-scale current good manufacturing practice-compliant process. Limiting the interval between T-cell isolation and CAR treatment is critical for patients with rapidly progressing disease. Generating CAR T cells in less time also improves potency, which is central to the effectiveness of these therapies. .

Proteome patterns in peach fruit (Prunus persica L.) stored at different low temperatures were examined in order to gain a better understanding why peach fruit is less prone to chilling injury when stored at 0 degrees C than at 5 degrees C. Some differently expressed proteins in peach fruit stored at 0 and 5 degrees C were identified using electrospray ionization quadrupole time-of-flight tandem mass spectrometry. Among these proteins, four membrane stability related proteins, i.e., enolase, temperature-induced lipocalin, major allergen Pru p 1, and type II SK2 dehydrin were enhanced, but three proteins related to phenolic compounds metabolization, cinnamyl-alcohol dehydrogenase 5, cinnamyl-alcohol dehydrogenase 1, and chorismate mutase, were repressed in peach fruit at 0 degrees C as compared to that at 5 degrees C. The abundance of glucose-6-phosphate dehydrogenase, NADP-dependent isocitrate dehydrogenase, and NADP-dependent malic enzyme, which catalyze the reactions during sugar metabolism and energy pathways, was found to decrease in peach fruit stored at 0 degrees C. In addition, our data revealed that low temperature of 0 degrees C might regulate the endogenous H(2)O(2) level, resulting in activating the transcriptional level of genes encoding the proteins related to membrane stability. These results provide a comprehensive knowledge to understand the mechanisms by which peach fruit stored at 0 degrees C showed a higher chilling tolerance than that at 5 degrees C.