FOXP1 and KLF2 reciprocally regulate checkpoints of stem-like to effector transition in CAR T cells.

In cancer and infections, self-renewing stem-like CD8+ T cells mediate the response of immunotherapies and replenish terminally exhausted T cells and effector-like T cells. However, the programs governing the lineage choice in chimeric antigen receptor (CAR) T cells are unclear. Here, by simultaneously profiling single-cell chromatin accessibility and transcriptome in the same CAR T cells, we identified heterogeneous chromatin states within CD8+ T cell subsets that foreshadowed transcriptional changes and were primed for regulation by distinct transcription factors. Transcription factors that controlled each CD8+ T cell subset were regulated by high numbers of enhancers and positioned as hubs of gene networks. FOXP1, a hub in the stem-like network, promoted expansion and stemness of CAR T cells and limited excessive effector differentiation. In the effector network, KLF2 enhanced effector CD8+ T cell differentiation and prevented terminal exhaustion. Thus, we identified gene networks and hub transcription factors that controlled the differentiation of stem-like CD8+ CAR T cells into effector or exhausted CD8+ CAR T cells.

BACH2 enforces the transcriptional and epigenetic programs of stem-like CD8+ T cells.

During chronic infection and cancer, a self-renewing CD8+ T cell subset maintains long-term immunity and is critical to the effectiveness of immunotherapy. These stem-like CD8+ T cells diverge from other CD8+ subsets early after chronic viral infection. However, pathways guarding stem-like CD8+ T cells against terminal exhaustion remain unclear. Here, we show that the gene encoding transcriptional repressor BACH2 is transcriptionally and epigenetically active in stem-like CD8+ T cells but not terminally exhausted cells early after infection. BACH2 overexpression enforced stem-like cell fate, whereas BACH2 deficiency impaired stem-like CD8+ T cell differentiation. Single-cell transcriptomic and epigenomic approaches revealed that BACH2 established the transcriptional and epigenetic programs of stem-like CD8+ T cells. In addition, BACH2 suppressed the molecular program driving terminal exhaustion through transcriptional repression and epigenetic silencing. Thus, our study reveals a new pathway that enforces commitment to stem-like CD8+ lineage and prevents an alternative terminally exhausted cell fate.

[Screening and verification of proteins of Salvia miltiorrhiza polyphenol oxidase interaction].

Polyphenol oxidase(PPO) is an important antioxidant enzyme in plants. It has the functions of scavenging active oxygen and synthesizing phenols, lignin, and plant protection factors, and can enhance the plant's resistance to stress and resistance to pests and diseases. Our previous research found that Salvia miltiorrhiza PPO gene can positively regulate salvianolic acid B synthesis. In order to further explore the mechanism, a pGBKT7-PPO bait vector was constructed using the cloned S. miltiorrhiza polyphenol oxidase gene(SmPPO, GenBank accession number: KF712274.1), and verified that it had no self-activation and no toxicity. The titer of S. miltiorrhiza cDNA library constructed by our laboratory was 4.75 × 107 cfu·mL~(-1), which met the requirements for library construction. Through yeast two-hybrid test, 22 proteins that could interact with SmPPO were screened. Only yeast PAL1 and TAT interacted with SmPPO through yeast co-transformation verification. Further verification was performed by bimolecular fluorescence complementary detection(BiFC). Only TAT and SmPPO interacted, so it meant that TAT and SmPPO interacted. TAT and SmPPO were truncated according to the domain, respectively. The first 126 amino acids of SmPPO and tyrosine amino transferase(TAT) were obtained to interact on the cell membrane and chloroplast. SmPPO was obtained by subcellular localization test, which was mainly loca-lized on the nucleus and cell membrane; TAT was localized on the cell membrane. Real-time quantitative PCR results showed that the SmPPO gene was mainly expressed in roots and stems; the TAT gene was expressed in roots, and the expression level in stems and flowers was low. This article lays a solid foundation for the in-depth study of the molecular mechanism of the interaction of S. miltiorrhiza SmPPO and TAT to regulate the synthesis of phenolic substances.

Nilotinib Effects on Safety, Tolerability, and Biomarkers in Alzheimer's Disease.

OBJECTIVE: Preclinical evidence with nilotinib, a US Food and Drug Administration (FDA)-approved drug for leukemia, indicates improvement in Alzheimer's disease phenotypes. We investigated whether nilotinib is safe, and detectable in cerebrospinal fluid, and alters biomarkers and clinical decline in Alzheimer's disease. METHODS: This single-center, phase 2, randomized, double-blind, placebo-controlled study investigated the safety, tolerability, and pharmacokinetics of nilotinib, and measured biomarkers in participants with mild to moderate dementia due to Alzheimer's disease. The diagnosis was supported by cerebrospinal fluid or amyloid positron emission tomography biomarkers. Nilotinib 150 mg versus matching placebo was taken orally once daily for 26 weeks followed by nilotinib 300 mg versus placebo for another 26 weeks. RESULTS: Of the 37 individuals enrolled, 27 were women and the mean (SD) age was 70.7 (6.48) years. Nilotinib was well-tolerated, although more adverse events, particularly mood swings, were noted with the 300 mg dose. In the nilotinib group, central nervous system (CNS) amyloid burden was significantly reduced in the frontal lobe compared to the placebo group. Cerebrospinal fluid Aß40 was reduced at 6 months and Aß42 was reduced at 12 months in the nilotinib group compared to the placebo. Hippocampal volume loss was attenuated (-27%) at 12 months and phospho-tau-181 was reduced at 6 months and 12 months in the nilotinib group. INTERPRETATION: Nilotinib is safe and achieves pharmacologically relevant cerebrospinal fluid concentrations. Biomarkers of disease were altered in response to nilotinib treatment. These data support a larger, longer, multicenter study to determine the safety and efficacy of nilotinib in Alzheimer's disease. ANN NEUROL 2020 ANN NEUROL 2020;88:183-194.