Yaxian Kong

PD-1(hi)TIM-3(+) T cells associate with and predict leukemia relapse in AML patients post allogeneic stem cell transplantation.

Prognosis of leukemia relapse post allogeneic stem cell transplantation (alloSCT) is poor and effective new treatments are urgently needed. T cells are pivotal in eradicating leukemia through a graft versus leukemia (GVL) effect and leukemia relapse is considered a failure of GVL. T-cell exhaustion is a state of T-cell dysfunction mediated by inhibitory molecules including programmed cell death protein 1 (PD-1) and T-cell immunoglobulin domain and mucin domain 3 (TIM-3). To evaluate whether T-cell exhaustion and inhibitory pathways are involved in leukemia relapse post alloSCT, we performed phenotypic and functional studies on T cells from peripheral blood of acute myeloid leukemia patients receiving alloSCT. Here we report that PD-1hiTIM-3+ cells are strongly associated with leukemia relapse post transplantation. Consistent with exhaustion, PD-1hiTIM-3+ T cells are functionally deficient manifested by reduced production of interleukin 2 (IL-2), tumor necrosis factor-α (TNF-α) and interferon-γ (IFN-γ). In addition, these cells demonstrate a phenotype consistent with exhausted antigen-experienced T cells by losing TN and TEMRA subsets. Importantly, increase of PD-1hiTIM-3+ cells occurs before clinical diagnosis of leukemia relapse, suggesting their predictive value. Results of our study provide an early diagnostic approach and a therapeutic target for leukemia relapse post transplantation.

Peritumoural neutrophils negatively regulate adaptive immunity via the PD-L1/PD-1 signalling pathway in hepatocellular carcinoma

BackgroundPD-L1 expression on neutrophils contributes to the impaired immune response in infectious disease, but the detailed role of PD-L1 expression on neutrophils in HCC remains unclear.MethodsWe investigated the phenotype and morphology of neutrophils infiltrated in tumour tissues from both patients with HCC and hepatoma-bearing mice.ResultsWe found that neutrophils dominantly infiltrated in the peritumoural region. The neutrophil-to-T cell ratio (NLR) was higher in peritumoural tissue than that in the intratumoural tissue and was negatively correlated with the overall survival of patients with HCC. Infiltrating neutrophils displayed a phenotype of higher frequency of programmed cell death ligand 1 (PD-L1) positive neutrophils. The ratio of PD-L1+ neutrophils-to-PD-1+ T cells was higher in peritumoural tissue and better predicted the disease-free survival of patients with HCC. We further confirmed a higher frequency of PD-L1+ neutrophils and PD-1+ T cells in hepatoma-bearing mice. Functionally, the PD-L1+ neutrophils from patients with HCC effectively suppressed the proliferation and activation of T cells, which could be partially reversed by the blockade of PD-L1.ConclusionsOur results indicate that the tumour microenvironment induces impaired antitumour immunity via the modulation of PD-L1 expression on tumour infiltrating neutrophils.

Expression of PD-1 on CD4+ T cells in peripheral blood associates with poor clinical outcome in non-small cell lung cancer.

Recent success of using agents inhibiting the major immune check point, programmed cell death-1 (PD-1) pathway, offers a great promise for effective cancer therapy. Two blocking antibodies for PD-1, nivolumab and pembrolizumab have recently been approved for treating advanced recurrent non-small cell lung cancer (NSCLC). Activation of PD-1 on T cells and PD-L1 on tumor cells or antigen presenting cells leads to T cell exhaustion and ultimately tumor growth. In this study, we performed flow cytometry analysis of peripheral blood samples collected from patients with advanced NSCLC at initial diagnosis. We report that surface expression of PD-1 on CD4+ T cells has a prognostic value in NSCLC patients, as high expression of PD-1 is associated with a shorter progression-free survival and overall survival. Importantly, we also found that high PD-1 expression on peripheral CD4+ T cells is associated with inferior clinical response in a subset of patients who received anti-PD-L1 treatment, indicating a potential predictive value of this marker. This work highlights the potential of a non-invasive and effective method to determine prognostic and predictive biomarkers for inhibiting the PD-1 pathway in NSCLC patients.

Blimp-1 impairs T cell function via upregulation of TIGIT and PD-1 in patients with acute myeloid leukemia

BackgroundT cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif (ITIM) domain (TIGIT) and programmed cell death protein 1 (PD-1) are important inhibitory receptors that associate with T cell exhaustion in acute myeloid leukemia (AML). In this study, we aimed to determine the underlying transcriptional mechanisms regulating these inhibitory pathways. Specifically, we investigated the role of transcription factor B lymphocyte-induced maturation protein 1 (Blimp-1) in T cell response and transcriptional regulation of TIGIT and PD-1 in AML.MethodsPeripheral blood samples collected from patients with AML were used in this study. Blimp-1 expression was examined by flow cytometry. The correlation of Blimp-1 expression to clinical characteristics of AML patients was analyzed. Phenotypic and functional studies of Blimp-1-expressing T cells were performed using flow cytometry-based assays. Luciferase reporter assays and ChIP assays were applied to assess direct binding and transcription activity of Blimp-1. Using siRNA to silence Blimp-1, we further elucidated the regulatory role of Blimp-1 in the TIGIT and PD-1 expression and T cell immune response.ResultsBlimp-1 expression is elevated in T cells from AML patients. Consistent with exhaustion, Blimp-1+ T cells upregulate multiple inhibitory receptors including PD-1 and TIGIT. In addition, they are functionally impaired manifested by low cytokine production and decreased cytotoxicity capacity. Importantly, the functional defect is reversed by inhibition of Blimp-1 via siRNA knockdown. Furthermore, Blimp-1 binds to the promoters of PD-1 and TIGIT and positively regulates their expression.ConclusionsOur study demonstrates an important inhibitory effect of Blimp-1 on T cell response in AML; thus, targeting Blimp-1 and its regulated molecules to improve the immune response may provide effective leukemia therapeutics.

Sepsis-Induced Thymic Atrophy Is Associated with Defects in Early Lymphopoiesis.

Impaired T lymphopoiesis is associated with immunosuppression of the adaptive immune response and plays a role in the morbidity and mortality of patients and animal models of sepsis. Although previous studies examined several intrathymic mechanisms that negatively affect T lymphopoiesis, the extrathymic mechanisms remain poorly understood. Here, we report a dramatic decrease in the percentage of early T lineage progenitors (ETPs) in three models of sepsis in mice (cecal ligation and puncture, lipopolysaccharide continuous injection, and poly I:C continuous injection). However, septic mice did not show a decrease in the number of bone marrow (BM) precursor cells. Instead, the BM progenitors for ETPs expressed reduced mRNA levels of CC chemokine receptor (CCR) 7, CCR9 and P‐selectin glycoprotein ligand 1, and exhibited impaired homing capacity in vitro and in vivo. Furthermore, RNA‐Seq analysis and real‐time PCR showed a marked downregulation of several lymphoid‐related genes in hematopoietic stem and progenitor cells. Hematopoietic stem and progenitor cells differentiated into myeloid cells but failed to generate T lymphocytes in vitro and in vivo. Our results indicate that the depletion of ETPs in septic mice might be a consequence of an impaired migration of BM progenitors to the thymus, as well as a defect in lymphoid lineage commitment. Stem Cells 2016;34:2902–2915

Expansion of activated regulatory T cells by myeloid-specific chemokines via an alternative pathway in CSF of bacterial meningitis patients

Previous studies have demonstrated that activation/expansion by certain cytokines as well as recruitment by specific chemokines is involved in enrichment of regulatory T (Treg) cells in local tissues or organs under pathological conditions. Recent evidence indicates that human Treg cells are a heterogeneous population that comprises three distinct subpopulations: CD25+CD45RA+ resting Treg (rTreg) cells, CD25hiCD45RA− activated Treg (aTreg) cells, which are both suppressive, and CD25+CD45RA− cytokine‐secreting T cells with proinflammatory capacity. Moreover, rTreg cells can proliferate and convert to aTreg cells. Here, we found an increase in aTreg‐cell frequency in the cerebrospinal fluid (CSF) of patients with postneurosurgery bacterial meningitis. We revealed that such an increased aTreg‐cell frequency in the CSF was not due to enhanced chemotaxis. Instead of a classic conversion pathway from rTreg to aTreg cells, we identified an alternative route of Treg‐cell conversion from cytokine‐secreting cells to aTreg cells induced by myeloid‐specific chemokine CXC chemokine receptor (CXCR) ligand 5 via CXCR1 and CXCR2 receptors, or by CSF myeloid cells in a cell–cell contact manner. Our results reveal a different view of how the immune system controls overwhelming local immune responses during infection, and provide evidence of how innate immunity negatively regulates adaptive immunity.

T-cell Immunoglobulin and ITIM Domain Contributes to CD8+ T-cell Immunosenescence

Aging is associated with immune dysfunction, especially T‐cell defects, which result in increased susceptibility to various diseases. Previous studies showed that T cells from aged mice express multiple inhibitory receptors, providing evidence of the relationship between T‐cell exhaustion and T‐cell senescence. In this study, we showed that T‐cell immunoglobulin and immunoreceptor tyrosine‐based inhibitory motif (ITIM) domain (TIGIT), a novel co‐inhibitory receptor, was upregulated in CD8+ T cells of elderly adults. Aged TIGIT+ CD8+ T cells expressed high levels of other inhibitory receptors including PD‐1 and exhibited features of exhaustion such as downregulation of the key costimulatory receptor CD28, representative intrinsic transcriptional regulation, low production of cytokines, and high susceptibility to apoptosis. Importantly, their functional defects associated with aging were reversed by TIGIT knockdown. CD226 downregulation on aged TIGIT+ CD8+ T cells is likely involved in TIGIT‐mediated negative immune suppression. Collectively, our findings indicated that TIGIT acts as a critical immune regulator during aging, providing a strong rationale for targeting TIGIT to improve dysfunction related to immune system aging.

A Novel CD48-Based Analysis of Sepsis-Induced Mouse Myeloid-Derived Suppressor Cell Compartments

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous subset of cells that expands dramatically in many disease states and can suppress T-cell responses. MDSCs mainly include monocytic and granulocytic subpopulations that can be distinguished in mice by the expression of Ly6G and Ly6C cell surface markers. This identification system has been validated in experimental tumor models, but not in models of inflammation-associated conditions such as sepsis. We challenged growth factor independent 1 transcription repressor green fluorescent protein (Gfi1:GFP) knock-in reporter mice with cecal ligation and puncture surgery and found that CD11b+Ly6GlowLy6Chigh MDSCs in this sepsis model comprised both monocytic and granulocytic MDSCs. The evidence that conventional Ly6G/Ly6C marker analysis may not be suited to study of inflammation-induced MDSCs led to the development of a novel strategy of distinguishing granulocytic MDSCs from monocytic MDSCs in septic mice by expression of CD48. Application of this novel model should help achieve a more accurate understanding of the inflammation-induced MDSC activity.

Phylogenomic analysis unravels evolution of yellow fever virus within hosts

The yellow fever virus (YFV) recently reemerged in the large outbreaks in Africa and Brazil, and the first imported patients into Asia have recalled the concerns of YFV evolution. Here we show phylogenomics of YFV with serial clinical samples of the 2016 YFV infections. Phylogenetics exhibited that the 2016 strains were close to Angola 1971 strains and only three amino acid changes presented new to other lineages. Deep sequencing of viral genomes discovered 101 intrahost single nucleotide variations (iSNVs) and 234 single nucleotide polymorphisms (SNPs). Analysis of iSNV distribution and mutated allele frequency revealed that the coding regions were under purifying selection. Comparison of the evolutionary rates estimated by iSNV and SNP showed that the intrahost rate was ~2.25 times higher than the epidemic rate, and both rates were higher than the long-term YFV substitution rate, as expected. In addition, the result also hinted that short viremia duration of YFV might further hinder the evolution of YFV.

Tumor‐activated liver stromal cells regulate myeloid‐derived suppressor cells accumulation in the liver

Regulating mechanisms underlying hepatic myeloid‐derived suppressor cell (MDSC) accumulation remain to be described. Here, we provide evidence for the involvement of tumour‐activated liver stromal cells in the process of hepatic MDSCs migration and accumulation. Our data showed an elevated frequency of MDSCs in the liver of tumour‐bearing mice. Moreover, tumour‐activated liver stromal cells promote MDSC migration into the liver site. Further investigation indicated higher levels of cytokine and chemokine expression in liver stromal cells after exposure to the tumour‐conditioned supernatant. Notably, the expression levels of proinflammatory factors, mainly including macrophage colony stimulating factor (M‐CSF), transforming growth factor‐β (TGF‐β), monocyte chemotactic protein‐1 (MCP‐1) and stromal‐derived factor‐1 (SDF‐1), increased after treatment with tumour‐conditioned supernatant, and blockade of MCP‐1 or SDF‐1 decreased the proportion of tumour infiltrated MDSCs in mice co‐transplanted with liver stromal cells and tumour cells, but not in mice with only tumour cells injection. These findings demonstrate that tumour‐activated liver stromal cells produce higher levels of chemokines and cytokines, which may contribute to MDSC accumulation into the liver site in patients with liver cancer.