RESUMEN
Early expansion and long-term persistence predict efficacy of chimeric antigen receptor T cells (CARTs)1-7, but mechanisms governing effector versus memory CART differentiation and whether asymmetric cell division induces differential fates in human CARTs remain unclear. Here we show that target-induced proximity labelling enables isolation of first-division proximal-daughter and distal-daughter CD8 CARTs that asymmetrically distribute their surface proteome and transcriptome, resulting in divergent fates. Target-engaged CARs remain on proximal daughters, which inherit a surface proteome resembling activated-undivided CARTs, whereas the endogenous T cell receptor and CD8 enrich on distal daughters, whose surface proteome resembles resting CARTs, correlating with glycolytic and oxidative metabolism, respectively. Despite memory-precursor phenotype and in vivo longevity, distal daughters demonstrate transient potent cytolytic activity similar to proximal daughters, uncovering an effector-like state in distal daughters destined to become memory CARTs. Both partitioning of pre-existing transcripts and changes in RNA velocity contribute to asymmetry of fate-determining factors, resulting in diametrically opposed transcriptional trajectories. Independent of naive, memory or effector surface immunophenotype, proximal-daughter CARTs use core sets of transcription factors known to support proliferation and effector function. Conversely, transcription factors enriched in distal daughters restrain differentiation and promote longevity, evidenced by diminished long-term in vivo persistence and function of distal-daughter CARTs after IKZF1 disruption. These studies establish asymmetric cell division as a framework for understanding mechanisms of CART differentiation and improving therapeutic outcomes.
Asunto(s)
División Celular Asimétrica , Linfocitos T CD8-positivos , Diferenciación Celular , Receptores Quiméricos de Antígenos , Animales , Humanos , Ratones , Linfocitos T CD8-positivos/citología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Linaje de la Célula , Glucólisis , Memoria Inmunológica , Inmunoterapia Adoptiva , Oxidación-Reducción , Proteoma/metabolismo , Receptores Quiméricos de Antígenos/metabolismo , Receptores Quiméricos de Antígenos/inmunología , ARN Mensajero/genética , ARN Mensajero/metabolismo , Factores de Transcripción/metabolismo , TranscriptomaRESUMEN
Chimeric antigen receptor (CAR) T cell dysfunction is a major barrier to achieving lasting remission in hematologic cancers, especially in chronic lymphocytic leukemia (CLL). We have shown previously that Δ133p53α, an endogenous isoform of the human TP53 gene, decreases in expression with age in human T cells, and that reconstitution of Δ133p53α in poorly functional T cells can rescue proliferation [A. M. Mondal et al., J. Clin. Invest. 123, 5247-5257 (2013)]. Although Δ133p53α lacks a transactivation domain, it can form heterooligomers with full-length p53 and modulate the p53-mediated stress response [I. Horikawa et al., Cell Death Differ. 24, 1017-1028 (2017)]. Here, we show that constitutive expression of Δ133p53α potentiates the anti-tumor activity of CD19-directed CAR T cells and limits dysfunction under conditions of high tumor burden and metabolic stress. We demonstrate that Δ133p53α-expressing CAR T cells exhibit a robust metabolic phenotype, maintaining the ability to execute effector functions and continue proliferating under nutrient-limiting conditions, in part due to upregulation of critical biosynthetic processes and improved mitochondrial function. Importantly, we show that our strategy to constitutively express Δ133p53α improves the anti-tumor efficacy of CAR T cells generated from CLL patients that previously failed CAR T cell therapy. More broadly, our results point to the potential role of the p53-mediated stress response in limiting the prolonged antitumor functions required for complete tumor clearance in patients with high disease burden, suggesting that modulation of the p53 signaling network with Δ133p53α may represent a translationally viable strategy for improving CAR T cell therapy.