bims-imseme Biomed News
on Immunosenescence and T cell metabolism
Issue of 2020‒10‒04
thirty-two papers selected by
Pierpaolo Ginefra
Ludwig Institute for Cancer Research

  1. Front Immunol. 2020 ;11 1834
      Memory T cells persist for long term to mediate robust recall response upon rechallenging with previous encountered pathogens. The memory T cell pool is highly heterogeneous based on distinct phenotypic, functional, and locational properties, and contains discrete subsets, which contribute to diverse immune responses. In this mini-review, we will briefly discuss the distinct subsets of memory T cells and then focus on mitochondria-related metabolic and epigenetic regulations of CD8+ T cell memory formation. In particular, we discuss many aspects of mitochondrial quality control systems (biogenesis, dynamics, etc.) in regulating CD8+ T cell fate decision and antitumor immunity. Importantly, targeting mitochondrial metabolism to boost T cell memory formation and metabolic fitness might represent an attractive strategy to improve cancer immunotherapy including CAR-T therapy.
    Keywords:  CD8 T cell; cancer; immunotherapy; memory; mitochondria
  2. J Immunol. 2020 Sep 30. pii: ji2000459. [Epub ahead of print]
      CD8 T cell differentiation is orchestrated by dynamic metabolic changes that direct activation, proliferation, cytotoxic function, and epigenetic changes. We report that the BTB-ZF family transcriptional repressor Zbtb20 negatively regulates CD8 T cell metabolism and memory differentiation in mice. Effector and memory CD8 T cells with conditional Zbtb20 deficiency displayed enhanced mitochondrial and glycolytic metabolism, and memory CD8 T cells had enhanced spare respiratory capacity. Furthermore, Zbtb20-deficient CD8 T cells displayed increased flexibility in the use of mitochondrial fuel sources. Phenotypic and transcriptional skewing toward the memory fate was observed during the CD8 T cell response to Listeria monocytogenes Memory cells mounted larger secondary responses and conferred better protection following tumor challenge. These data suggest that inactivation of Zbtb20 may offer an approach to enhance metabolic activity and flexibility and improve memory CD8 T cell differentiation, useful attributes for T cells used in adoptive immunotherapy.
  3. Curr Opin Hematol. 2020 Nov;27(6): 353-359
      PURPOSE OF REVIEW: Controlling T cell activity through metabolic manipulation has become a prominent feature in immunology and practitioners of both adoptive cellular therapy (ACT) and haematopoietic stem cell transplantation (HSCT) have utilized metabolic interventions to control T cell function. This review will survey recent metabolic research efforts in HSCT and ACT to paint a broad picture of immunometabolism and highlight advances in each area.RECENT FINDINGS: In HSCT, recent publications have focused on modifying reactive oxygen species, sirtuin signalling or the NAD salvage pathway within alloreactive T cells and regulatory T cells. In ACT, metabolic interventions that bolster memory T cell development, increase mitochondrial density and function, or block regulatory signals in the tumour microenvironment (TME) have recently been published.
    SUMMARY: Metabolic interventions control immune responses. In ACT, efforts seek to improve the in-vivo metabolic fitness of T cells, while in HSCT energies have focused on blocking alloreactive T cell expansion or promoting regulatory T cells. Methods to identify new, metabolically targetable pathways, as well as the ability of metabolic biomarkers to predict disease onset and therapeutic response, will continue to advance the field towards clinically applicable interventions.
  4. Cancers (Basel). 2020 Sep 30. pii: E2828. [Epub ahead of print]12(10):
      The inability of tumor-infiltrating T lymphocytes to eradicate tumor cells within the tumor microenvironment (TME) is a major obstacle to successful immunotherapeutic treatments. Understanding the immunosuppressive mechanisms within the TME is paramount to overcoming these obstacles. T cell senescence is a critical dysfunctional state present in the TME that differs from T cell exhaustion currently targeted by many immunotherapies. This review focuses on the physiological, molecular, metabolic and cellular processes that drive CD8+ T cell senescence. Evidence showing that senescent T cells hinder immunotherapies is discussed, as are therapeutic options to reverse T cell senescence.
    Keywords:  CD8+ T cells; immunotherapy; metabolism; senescence
  5. Cancer Res. 2020 Oct 01. pii: canres.0524.2020. [Epub ahead of print]
      T cell exhaustion in cancer is linked to poor clinical outcomes and evidence suggests T cell metabolic changes precede functional exhaustion. Direct competition between tumor-infiltrating lymphocytes (TIL) and cancer cells for metabolic resources often renders T cells dysfunctional. Here, we report an epigenetic mechanism contributing to the development of metabolic exhaustion in TILs. Environmental stress produces epigenome remodeling events within tumor-infiltrating lymphocytes resulting from loss of the histone methyltransferase EZH2. Using a multi-omics approach, we have defined a Cdkn2a.Arf-mediated, p53-independent mechanism by which EZH2 inhibition leads to mitochondrial dysfunction and the resultant exhaustion. Reprogramming T cells to express a gain-of-function EZH2 mutant resulted in an enhanced ability of T cells to inhibit tumor growth in vitro and in vivo. Our data suggest manipulation of T cell EZH2 within the context of cellular therapies may yield lymphocytes which are able to withstand harsh tumor metabolic environments and collateral pharmacologic insults.
  6. Clin Immunol. 2020 Sep 29. pii: S1521-6616(20)30762-2. [Epub ahead of print] 108602
      OBJECTIVE: This study performed an integrated analysis of the cellular and transcriptional differences in peripheral immune cells between patients with Systemic Lupus Erythematosus (SLE) and healthy controls (HC).METHODS: Peripheral blood was analyzed using standardized flow cytometry panels. Transcriptional analysis of CD4+ T cells was performed by microarrays and Nanostring assays.
    RESULTS: SLE CD4+ T cells showed an increased expression of oxidative phosphorylation and immunoregulatory genes. SLE patients presented higher frequencies of activated CD38+HLA-DR+ T cells than HC. Hierarchical clustering identified a group of SLE patients among which African Americans were overrepresented, with highly activated T cells, and higher frequencies of Th1, Tfh, and plasmablast cells. T cell activation was positively correlated with metabolic gene expression in SLE patients but not in HC.
    CONCLUSIONS: SLE subjects presenting with activated T cells and a hyperactive metabolic signature may represent an opportunity to correct aberrant immune activation through targeted metabolic inhibitors.
  7. Nat Immunol. 2020 Sep 28.
      T follicular helper (TFH) cells are critical in adaptive immune responses to pathogens and vaccines; however, what drives the initiation of their developmental program remains unclear. Studies suggest that a T cell antigen receptor (TCR)-dependent mechanism may be responsible for the earliest TFH cell-fate decision, but a critical aspect of the TCR has been overlooked: tonic TCR signaling. We hypothesized that tonic signaling influences early TFH cell development. Here, two murine TCR-transgenic CD4+ T cells, LLO56 and LLO118, which recognize the same antigenic peptide presented on major histocompatibility complex molecules but experience disparate strengths of tonic signaling, revealed low tonic signaling promotes TFH cell differentiation. Polyclonal T cells paralleled these findings, with naive Nur77 expression distinguishing TFH cell potential. Two mouse lines were also generated to both increase and decrease tonic signaling strength, directly establishing an inverse relationship between tonic signaling strength and TFH cell development. Our findings elucidate a central role for tonic TCR signaling in early TFH cell-lineage decisions.
  8. Mol Oncol. 2020 Sep 27.
      Senescence refers to a cellular state featuring a stable cell-cycle arrest triggered in response to stress. This response also involves other distinct morphological and intracellular changes including alterations in gene expression and epigenetic modifications, elevated macromolecular damage, metabolism deregulation, and a complex proinflammatory secretory phenotype. The initial demonstration of oncogene-induced senescence in vitro established senescence as an important tumour suppressive mechanism, in addition to apoptosis. Senescence not only halts the proliferation of premalignant cells but also facilitates the clearance of affected cells through immunosurveillance. Failure to clear senescent cells owing to deficient immunosurveillance may, however, lead to a state of chronic inflammation that nurtures a pro-tumorigenic microenvironment favouring cancer initiation, migration, and metastasis. In addition, senescence is a response to post-therapy genotoxic stress. Therefore, tracking the emergence of senescent cells becomes pivotal to detect potential pro-tumorigenic events. Current protocols for the in vivo detection of senescence require the analysis of fixed or deep-frozen tissues, despite a significant clinical need for real-time bioimaging methods. Accuracy and efficiency of senescence detection is further hampered by a lack of universal and more specific senescence biomarkers. Recently, in an attempt to overcome these hurdles, an assortment of detection tools have been developed. These strategies all have significant potential for clinical utilization, and include flow cytometry combined with histo- or cytochemical approaches, nanoparticle-based targeted delivery of imaging contrast agents, OFF-ON fluorescent senoprobes, positron emission tomography (PET) senoprobes, and analysis of circulating SASP factors, extracellular vesicles, and cell-free nucleic acids isolated from plasma. Here we highlight the occurrence of senescence in neoplasia and advanced tumours, assess the impact of senescence on tumorigenesis, and discuss how the ongoing development of senescence detection tools might improve early detection of multiple cancers and response to therapy in the near future.
    Keywords:  Cellular senescence; cancer; detection; senoprobes; tumour microenvironment
  9. Nat Immunol. 2020 Sep 28.
      Foxp3+ regulatory T (Treg) cells expressing the interleukin (IL)-33 receptor ST2 mediate tissue repair in response to IL-33. Whether Treg cells also respond to the alarmin IL-33 to regulate specific aspects of the immune response is not known. Here we describe an unexpected function of ST2+ Treg cells in suppressing the innate immune response in the lung to environmental allergens without altering the adaptive immune response. Following allergen exposure, ST2+ Treg cells were activated by IL-33 to suppress IL-17-producing γδ T cells. ST2 signaling in Treg cells induced Ebi3, a component of the heterodimeric cytokine IL-35 that was required for Treg cell-mediated suppression of γδ T cells. This response resulted in fewer eosinophil-attracting chemokines and reduced eosinophil recruitment into the lung, which was beneficial to the host in reducing allergen-induced inflammation. Thus, we define a fundamental role for ST2+ Treg cells in the lung as a negative regulator of the early innate γδ T cell response to mucosal injury.
  10. Front Cell Dev Biol. 2020 ;8 842
      The emerging epitranscriptome plays an essential role in autoimmune disease. As a novel mRNA modification, N4-acetylcytidine (ac4C) could promote mRNA stability and translational efficiency. However, whether epigenetic mechanisms of RNA ac4C modification are involved in systemic lupus erythematosus (SLE) remains unclear. Herein, we detected eleven modifications in CD4+ T cells of SLE patients using mass spectrometry (LC-MS/MS). Furthermore, using samples from four CD4+ T cell pools, we identified lower modification of ac4C mRNA in SLE patients as compared to that in healthy controls (HCs). Meanwhile, significantly lower mRNA acetyltransferase NAT10 expression was detected in lupus CD4+ T cells by RT-qPCR. We then illustrated the transcriptome-wide ac4C profile in CD4+ T cells of SLE patients by ac4C-RIP-Seq and found ac4C distribution in mRNA transcripts to be highly conserved and enriched in mRNA coding sequence regions. Using bioinformatics analysis, the 3879 and 4073 ac4C hyper-acetylated and hypoacetylated peaks found in SLE samples, respectively, were found to be significantly involved in SLE-related function enrichments, including multiple metabolic and transcription-related processes, ROS-induced cellular signaling, apoptosis signaling, and NF-κB signaling. Moreover, we demonstrated the ac4C-modified regulatory network of gene biological functions in lupus CD4+ T cells. Notably, we determined that the 26 upregulated genes with hyperacetylation played essential roles in autoimmune diseases and disease-related processes. Additionally, the unique ac4C-related transcripts, including USP18, GPX1, and RGL1, regulate mRNA catabolic processes and translational initiation. Our study identified novel dysregulated ac4C mRNAs associated with critical immune and inflammatory responses, that have translational potential in lupus CD4+ T cells. Hence, our findings reveal transcriptional significance and potential therapeutic targets of mRNA ac4C modifications in SLE pathogenesis.
    Keywords:  CD4+ T cells; N4-acetylcytidine (ac4C); NAT10; epitranscriptome; systemic lupus erythematosus
  11. Mol Ther Methods Clin Dev. 2020 Dec 11. 19 14-23
      T cells modified to co-express cytokine or other factors with chimeric antigen receptor (CAR) can induce substantial and persistent increases in antitumor capacity in vivo. However, the uncontrolled expression of cytokines or factors can lead to the overactivation of immune cells, causing severe adverse events such as cytokine release syndrome (CRS) and neurotoxicity by CAR T cells with excessive growth potential. Conventional promoters are unregulated, and their expression is unlimited in T cells. In this study, by connecting the cytomegalovirus (CMV) enhancer, core interferon gamma (IFN-γ) promoter, and a T-lymphotropic virus long terminal repeat sequence (TLTR), we constructed and screened the chimeric promoter CIFT, which was highly expressed in some cell lines secreting IFN-γ and silenced in others. We placed this promoter upstream of the anti-programmed cell death protein 1 (anti-PD-1) antibody gene, and this construct was co-transfected with the CAR construct into T cells. In vitro or in vivo, CAR T cells showed increased secretion of anti-PD-1 antibody under control of the chimeric promoter CIFT. pS-CIFT-αPD-1/CAR T also had similar or lower PD-1 expression, higher levels of T cell activation, more release of IFN-γ, and better antitumor activity specifically against mesothelin-positive and PD-1 ligand 1 (PD-L1)-positive cell lines. The chimeric promoter may be a promising strategy to manipulate the content of immune checkpoint inhibitors or other proteins in future therapeutic approaches for cancer treatment.
    Keywords:  CAR-T cells; IFN-γ; anti-PD-1 antibody; chimeric promoter; cytokine release syndrome; immune checkpoint inhibitors; piggyBac transposon system
  12. Annu Rev Pharmacol Toxicol. 2020 Sep 30.
      Senescence is the consequence of a signaling mechanism activated in stressed cells to prevent proliferation of cells with damage. Senescent cells (Sncs) often develop a senescence-associated secretory phenotype to prompt immune clearance, which drives chronic sterile inflammation and plays a causal role in aging and age-related diseases. Sncs accumulate with age and at anatomical sites of disease. Thus, they are regarded as a logical therapeutic target. Senotherapeutics are a new class of drugs that selectively kill Sncs (senolytics) or suppress their disease-causing phenotypes (senomorphics/senostatics). Since 2015, several senolytics went from identification to clinical trial. Preclinical data indicate that senolytics alleviate disease in numerous organs, improve physical function and resilience, and suppress all causes of mortality, even if administered to the aged. Here, we review the evidence that Sncs drive aging and disease, the approaches to identify and optimize senotherapeutics, and the current status of preclinical and clinical testing of senolytics. Expected final online publication date for the Annual Review of Pharmacology and Toxicology, Volume 61 is January 7, 2021. Please see for revised estimates.
  13. Clin Transl Oncol. 2020 Sep 30.
      CAR-T cell therapy, as a novel immunotherapy approach, has indicated successful results in the treatment of hematological malignancies; however, distinct results have been achieved regarding solid tumors. Tumor immunosuppressive microenvironment has been identified as the most critical barrier in CAR-T cell therapy of solid tumors. Developing novel strategies to augment the safety and efficacy of CAR-T cells could be useful to overcome the solid tumor hurdles. Similar to other cancer treatments, CAR-T cell therapy can cause some side effects, which can disturb the healthy tissues. In the current review, we will discuss the practical breakthroughs in CAR-T cell therapy using the multi-targeted and programmable CARs instead of conventional types. These superior types of CAR-T cells have been developed to increase the function and safety of T cells in a controllable manner, which would diminish the incidence of relevant side effects. Moreover, we will describe the capability of these powerful CARs in targeting multiple tumor antigens, redirecting the CAR-T cells to specific target cells, incrementing the safety of CARs, and other advantages that lead to promising outcomes in cancer CAR-T cell therapy.
    Keywords:  Adoptive cell therapy; Cancer; Conventional CARs; Immunotherapy; Multi-targeted CARs; Programmable CARs
  14. Cell Immunol. 2020 Sep 04. pii: S0008-8749(20)30372-5. [Epub ahead of print]357 104212
      Regulation of the adaptive immune response is critical for health. Regulatory activity can be found in multiple components of the immune system, however, the focus on particular components of the immune regulatory network has left many aspects of this critical immune component understudied. Here we review the evidence for activities of CD8+ T cells in immune homeostasis and regulation of autoimmune reactivity. The heterogeneous nature of identified CD8+ cell types are examined, and common phenotypes associated with functional activities are defined. The varying types of antigen signal crucial for CD8+ T cell regulatory activity are identified and the implications of these activation pathways for control of adaptive responses is considered. Finally, the promising capacity for transgenic antigen receptor directed cytotoxicity as a mechanism for modulation of autoimmunity is detailed.
    Keywords:  Autoimmunity; CD8; Immune regulation; Regulatory; T cells
  15. Blood Adv. 2020 Oct 13. 4(19): 4653-4664
      Engineered T-cell therapies have demonstrated impressive clinical responses in patients with hematologic malignancies. Despite this efficacy, many patients have a transient persistence of T cells, which can be correlated with transient clinical response. Translational data on T cells from pediatric cancer patients shows a progressive decline in chimeric antigen receptor (CAR) suitability with cumulative chemotherapy regardless of regimen. We investigated the effects of chemotherapy on surviving T cells in vitro, describing residual deficits unique to each agent including mitochondrial damage and metabolic alterations. In the case of cyclophosphamide but not doxorubicin or cytarabine, these effects could be reversed with N-acetylcysteine. Specifically, we observed that surviving T cells could be stimulated, expanded, and transduced with CARs with preserved short-term cytolytic function but at far lower numbers and with residual metabolic deficits. These data have implications for understanding the effects of chemotherapy on mature T cells later collected for adoptive cell therapy, as chemotherapy-exposed T cells may have lingering dysfunction that affects ex vivo adoptive cell therapy manufacturing techniques and, ultimately, clinical efficacy.
  16. Mol Cell. 2020 Sep 22. pii: S1097-2765(20)30614-6. [Epub ahead of print]
      Metabolism reprogramming is critical for both cancer progression and effective immune responses in the tumor microenvironment. Amino acid metabolism in different cells and their cross-talk shape tumor immunity and therapy efficacy in patients with cancer. In this review, we focus on multiple amino acids and their transporters, solute carrier (SLC) members. We discuss their involvement in regulation of immune responses in the tumor microenvironment and assess their associations with cancer immunotherapy, chemotherapy, and radiation therapy, and we review their potential as targets for cancer therapy. We stress the necessity to understand individual amino acids and their transporters in different cell subsets, the molecular intersection between amino acid metabolism, and effective T cell immunity and its relevance in cancer therapies.
    Keywords:  CD8(+) T cell; amino acid; cancer; checkpoint blockade; immunotherapy; metabolism; solute carriers
  17. Nat Cell Biol. 2020 Oct;22(10): 1170-1179
      SIRT1 (Sir2) is an NAD+-dependent deacetylase that plays critical roles in a broad range of biological events, including metabolism, the immune response and ageing1-5. Although there is strong interest in stimulating SIRT1 catalytic activity, the homeostasis of SIRT1 at the protein level is poorly understood. Here we report that macroautophagy (hereafter referred to as autophagy), a catabolic membrane trafficking pathway that degrades cellular components through autophagosomes and lysosomes, mediates the downregulation of mammalian SIRT1 protein during senescence and in vivo ageing. In senescence, nuclear SIRT1 is recognized as an autophagy substrate and is subjected to cytoplasmic autophagosome-lysosome degradation, via the autophagy protein LC3. Importantly, the autophagy-lysosome pathway contributes to the loss of SIRT1 during ageing of several tissues related to the immune and haematopoietic system in mice, including the spleen, thymus, and haematopoietic stem and progenitor cells, as well as in CD8+CD28- T cells from aged human donors. Our study reveals a mechanism in the regulation of the protein homeostasis of SIRT1 and suggests a potential strategy to stabilize SIRT1 to promote productive ageing.
  18. Nat Immunol. 2020 Sep 28.
      Antiviral CD8+ T cell responses are characterized by an initial activation/priming of T lymphocytes followed by a massive proliferation, subset differentiation, population contraction and the development of a stable memory pool. The transcription factor BATF3 has been shown to play a central role in the development of conventional dendritic cells, which in turn are critical for optimal priming of CD8+ T cells. Here we show that BATF3 was expressed transiently within the first days after T cell priming and had long-lasting T cell-intrinsic effects. T cells that lacked Batf3 showed normal expansion and differentiation, yet succumbed to an aggravated contraction and had a diminished memory response. Vice versa, BATF3 overexpression in CD8+ T cells promoted their survival and transition to memory. Mechanistically, BATF3 regulated T cell apoptosis and longevity via the proapoptotic factor BIM. By programing CD8+ T cell survival and memory, BATF3 is a promising molecule to optimize adoptive T cell therapy in patients.
  19. Mech Ageing Dev. 2020 Sep 26. pii: S0047-6374(20)30162-7. [Epub ahead of print] 111366
      Remyelination is a physiological response to demyelinating events aiming to restore saltatory conduction and preserve axonal integrity. Resident oligodendrocyte precursor cells (OPC) of the CNS tissue under appropriate conditions are mobilized to proliferate, migrate, and differentiate, in order to produce new myelin sheaths in the demyelinated lesion. In multiple sclerosis (MS), the most common immune-mediated demyelinating disease, remyelination efficiency declines with increasing age and disease duration. As myelin regeneration attempts in clinical trials so far are scarce, and have been met with limited success, the need to explore new remyelinating strategies is more compelling. Recently, ageing and cellular senescence have been implicated to the pathophysiology of a number of neurodegenerative diseases, including multiple sclerosis. Evidence on OPC senescence brings forward the possibility of exploiting cellular senescence as a possible target for promoting the endogenous remyelinating capacity of the CNS. Here we discuss the data indicating how cellular senescence affects remyelination, and the putative benefits to be drawn through the use of senolytic or senomorphic therapies targeting senescent cell populations in MS.
    Keywords:  cellular senescence; multiple sclerosis; oligodendrocyte; oligodendrocyte progenitor; remyelination
  20. Oncol Lett. 2020 Nov;20(5): 258
      As previously reported, hydrogen gas improves the prognosis of patients with cancer by restoring exhausted CD8+ T cells into active CD8+ T cells, possibly by activating mitochondria. As mitochondrial activators exhibit synergistic effects with nivolumab, the current study investigated whether hydrogen gas also affects the clinical outcomes of nivolumab. A total of 42 of 56 patients with lung cancer treated with nivolumab received hydrogen gas. Exhausted markers (PD-1 and Tim-3) on cell populations in the CD8+ T cell differentiation pathway were analyzed using flow cytometry. The concentration of coenzyme Q10 (CoQ10) was measured as a marker of mitochondrial function. The 42 patients treated with hydrogen gas and nivolumab (HGN) indicated a significantly longer overall survival (OS) compared with those treated with nivolumab only (n=14). In multivariate analysis, PD-1+Tim-3+terminal CD8+ T cells (PDT+) were an independent poor prognostic factor in OS, and CoQ10 showed a tendency to be associated with improved OS. The change in the rate of PDT+ and CoQ10 after vs. before HGN (PDT+ ratio and CoQ10 ratio, respectively) revealed that patients with low PDT+ ratio (<0.81) and high CoQ10 ratio (>1.175) had significantly longer OS compared with those with high PDT+ ratio and low CoQ10 ratio. Furthermore, PDT+, with a significant reverse correlation with CoQ10, was significantly lower in patients with high CoQ10 and/or CoQ10 ratio than in those low CoQ10 and/or CoQ10. Hydrogen gas has been suggested to enhance the clinical efficacy of nivolumab by increasing CoQ10 (mitochondria) to reduce PDT+, with PDT+ and CoQ10 as reliable negative and positive biomarkers of nivolumab, respectively.
    Keywords:  Nivolumab; T cell immunoglobulin and mucin domain-containing protein 3; coenzyme Q10; hydrogen gas; lung cancer
  21. Nat Cancer. 2020 May;1(5): 533-545
      Cancer cells express high levels of PD-L1, a ligand of the PD-1 receptor on T cells, allowing tumors to suppress T cell activity. Clinical trials utilizing antibodies that disrupt the PD-1/PD-L1 checkpoint have yielded remarkable results, with anti-PD-1 immunotherapy approved as first-line therapy for lung cancer patients. We used CRISPR-based screening to identify regulators of PD-L1 in human lung cancer cells, revealing potent induction of PD-L1 upon disruption of heme biosynthesis. Impairment of heme production activates the integrated stress response (ISR), allowing bypass of inhibitory upstream open reading frames in the PD-L1 5' UTR, resulting in enhanced PD-L1 translation and suppression of anti-tumor immunity. We demonstrated that ISR-dependent PD-L1 translation requires the translation initiation factor eIF5B. eIF5B overexpression, which is frequent in lung adenocarcinomas and associated with poor prognosis, is sufficient to induce PD-L1. These findings illuminate mechanisms of immune checkpoint activation and identify targets for therapeutic intervention.
  22. Biomedicines. 2020 Sep 30. pii: E390. [Epub ahead of print]8(10):
      Background: Patients with HIV (PWH) develop geriatric comorbidities, including functional and cognitive decline at a younger age. However, contributing mechanisms are unclear and interventions are lacking. We hypothesized that deficiency of the antioxidant protein glutathione (GSH) contributes to multiple defects representing premature aging in PWH, and that these defects could be improved by supplementing the GSH precursors glycine and N-acetylcysteine (GlyNAC). Methods: We conducted an open label clinical trial where eight PWH and eight matched uninfected-controls were studied at baseline. PWH were studied again 12-weeks after receiving GlyNAC, and 8-weeks after stopping GlyNAC. Controls did not receive supplementation. Outcome measures included red-blood cell and muscle GSH concentrations, mitochondrial function, mitophagy and autophagy, oxidative stress, inflammation, endothelial function, genomic damage, insulin resistance, glucose production, muscle-protein breakdown rates, body composition, physical function and cognition. Results: PWH had significant defects in measured outcomes, which improved with GlyNAC supplementation. However, benefits receded after stopping GlyNAC. Conclusions: This open label trial finds that PWH have premature aging based on multiple biological and functional defects, and identifies novel mechanistic explanations for cognitive and physical decline. Nutritional supplementation with GlyNAC improves comorbidities suggestive of premature aging in PWH including functional and cognitive decline, and warrants additional investigation.
    Keywords:  Glutathione; GlyNAC; HIV; cognition; mitochondrial function; oxidative stress; physical function; premature aging
  23. J Virol. 2020 Sep 30. pii: JVI.01627-20. [Epub ahead of print]
      Protection of a majority of viral vaccines is mediated by CD4 T cell-dependent humoral immunity. The methyltransferase enhancer of zeste homolog 2 (EZH2) dictates the differentiation of naïve CD4 T cells into distinct effector T helper subsets at the onset of acute viral infection. However, whether and how EZH2 manipulates differentiated virus-specific CD4 T cell expansion remains to be elucidated. Here, we found EZH2 is integral for virus-specific CD4 T cell expansion in a mouse model of acute viral infection. By a mechanism that involves fine-tuning the mechanistic target of rapamycin (mTOR) signaling, EZH2 participates in integrating metabolic pathways to support cell expansion. The genetic ablation of EZH2 leads to impaired cellular metabolism and consequent poor CD4 T cell response to acute viral infection. Thus, we identified EZH2 as a novel regulator in virus-specific CD4 T cell expansion during acute viral infection.IMPORTANCECD4 T cell response is critical in curtailing viral infection or eliciting efficacious viral vaccination. Highly efficient expansion of virus-specific CD4 T cells culminates qualified CD4 T cell response. Here, we found that the epigenetic regulator EZH2 is prerequisite for virus-specific CD4 T cell response, with a mechanism coupling of cell expansion and metabolism. Thus, our study provides valuable insights for strategies targeting EZH2 to improve efficacy of CD4 T cell-based viral vaccine and to help treat diseases associated with aberrant CD4 T cell response.
  24. Cancer Cell. 2020 Sep 15. pii: S1535-6108(20)30432-3. [Epub ahead of print]
      PD-1/PD-L1-checkpoint blockade therapy is generally thought to relieve tumor cell-mediated suppression in the tumor microenvironment but PD-L1 is also expressed on non-tumor macrophages and conventional dendritic cells (cDCs). Here we show in mouse tumor models that tumor-draining lymph nodes (TDLNs) are enriched for tumor-specific PD-1+ T cells which closely associate with PD-L1+ cDCs. TDLN-targeted PD-L1-blockade induces enhanced anti-tumor T cell immunity by seeding the tumor site with progenitor-exhausted T cells, resulting in improved tumor control. Moreover, we show that abundant PD-1/PD-L1-interactions in TDLNs of nonmetastatic melanoma patients, but not those in corresponding tumors, associate with early distant disease recurrence. These findings point at a critical role for PD-L1 expression in TDLNs in governing systemic anti-tumor immunity, identifying high-risk patient groups amendable to adjuvant PD-1/PD-L1-blockade therapy.
    Keywords:  PD-1; PD-L1; T cells; checkpoint immunotherapy; dendritic cells; lymph node; tumor immunology
  25. J Biochem. 2020 Oct 01. pii: mvaa109. [Epub ahead of print]
      Cellular senescence is an important tumor suppression mechanism that inhibits the proliferation of damaged cells. In senescent cells, irreparable DNA damage causes accumulation of genomic DNA fragments in the cytoplasm, which are recognized by the cyclic GMP-AMP synthase (cGAS)-stimulator of interferon gene (STING) pathway, resulting in secretion of numerous inflammatory proteins. This phenomenon is called senescence-associated secretory phenotype (SASP), and results in multiple physiological or pathological processes in the body. In addition, DNA damage also increases small extracellular vesicle (EV) release from senescent cells. This review presents recent insights into the molecular mechanisms and biological functions of senescence-associated extracellular vesicle (SA-EV) release that is associated with age-related diseases, particularly cancer.
    Keywords:  Cellular senescence; DNA damage; cGAS-STING; extracellular vesicle; senescence-associated secretory phenotype
  26. Front Immunol. 2020 ;11 1859
      Previous studies suggest that the presence of antigen-specific polyfunctional T cells is correlated with improved pathogen clearance, disease control, and clinical outcomes; however, the molecular mechanisms responsible for the generation, function, and survival of polyfunctional T cells remain unknown. The study of polyfunctional T cells has been, in part, limited by the need for intracellular cytokine staining (ICS), necessitating fixation and cell membrane permeabilization that leads to unacceptable degradation of RNA. Adopting elements from prior research efforts, we developed and optimized a modified protocol for the isolation of high-quality RNA (i.e., RIN > 7) from primary human T cells following aldehyde-fixation, detergent-based permeabilization, intracellular cytokines staining, and sorting. Additionally, this method also demonstrated utility preserving RNA when staining for transcription factors. This modified protocol utilizes an optimized combination of an RNase inhibitor and high-salt buffer that is cost-effective while maintaining the ability to identify and resolve cell populations for sorting. Overall, this protocol resulted in minimal loss of RNA integrity, quality, and quantity during cytoplasmic staining of cytokines and subsequent flourescence-activated cell sorting. Using this technique, we obtained the transcriptional profiles of functional subsets (i.e., non-functional, monofunctional, bifunctional, polyfunctional) of CMV-specific CD8+T cells. Our analyses demonstrated that these functional subsets are molecularly distinct, and that polyfunctional T cells are uniquely enriched for transcripts involved in viral response, inflammation, cell survival, proliferation, and metabolism when compared to monofunctional cells. Polyfunctional T cells demonstrate reduced activation-induced cell death and increased proliferation after antigen re-challenge. Further in silico analysis of transcriptional data suggested a critical role for STAT5 transcriptional activity in polyfunctional cell activation. Pharmacologic inhibition of STAT5 was associated with a significant reduction in polyfunctional cell cytokine expression and proliferation, demonstrating the requirement of STAT5 activity not only for proliferation and cell survival, but also cytokine expression. Finally, we confirmed this association between CMV-specific CD8+ polyfunctionality with STAT5 signaling also exists in immunosuppressed transplant recipients using single cell transcriptomics, indicating that results from this study may translate to this vulnerable patient population. Collectively, these results shed light on the mechanisms governing polyfunctional T cell function and survival and may ultimately inform multiple areas of immunology, including but not limited to the development of new vaccines, CAR-T cell therapies, and adoptive T cell transfer.
    Keywords:  CMV; RNA quality; RNAseq; fixation; permeabilization; polyfunctional T cells
  27. Cell Signal. 2020 Sep 23. pii: S0898-6568(20)30267-9. [Epub ahead of print]76 109790
      The primary activating receptor for T cells is the T cell receptor (TCR), which is stimulated upon binding to an antigen/MHC complex. TCR activation results in the induction of regulated signaling pathways vital for T cell differentiation, cellular adhesion and cytokine release. A critical TCR-induced signaling protein is the adaptor protein LAT. Upon TCR stimulation, LAT is phosphorylated on conserved tyrosines, which facilitates the formation of multiprotein complexes needed for propagation of signaling pathways. Although the role of the conserved tyrosines in LAT-mediated signaling has been investigated, few studies have examined the role of larger regions of LAT in TCR-induced pathways. In this study, a sequence alignment of 97 mammalian LAT proteins was used to identify several "functional" domains on LAT. Using LAT mutants expressed in Jurkat E6.1 cells, we observed that the membrane proximal, proline-rich region of LAT and the correct order of domains containing conserved tyrosines are necessary for optimal TCR-mediated early signaling, cytokine production, and cellular adhesion. Together, these data show that LAT contains distinct regions whose presence and correct order are required for the propagation of TCR-mediated signaling pathways.
    Keywords:  Domain order; LAT; Signal transduction; Structure/function study; T cells; TCR
  28. Front Oncol. 2020 ;10 1361
      Immune checkpoint inhibitors (ICPi) targeting the PD-1/PD-L1 pathway have shown marked success in patients with advanced melanoma. However, 60-70% of patients fail to respond, warranting a therapeutic intervention that could increase response rates. We and others have shown that S-adenosylmethionine (SAM), a universal methyl donor, has significant anticancer effects in numerous cancers previously; however, its effect on melanoma progression has not been evaluated. Interestingly, SAM was reported to be essential for T cell activation and proliferation and, thus, could potentially cooperate with ICPi and block melanoma progression. In this study, we examined the antitumor effects of SAM and ICPi alone and in combination in a well-established melanoma mouse model wherein syngeneic C57BL/6 mouse were subcutaneously (orthotopic) injected with B16-F1 cells. Treatment of mice with either SAM or anti-PD-1 antibody alone resulted in significant reduction in tumor volumes and weights; effects that were highest in mice treated with a combination of SAM+anti-PD-1. RNA-sequencing analysis of the primary tumors showed numerous differentially expressed genes (DEGs) following treatment with SAM+anti-PD-1, which was shown to downregulate cancer, MAPK, and tyrosine kinase pathways. Indeed, SAM+anti-PD-1 reversed the aberrant expression of some known melanoma genes. Tumor immunophenotyping revealed the SAM+anti-PD-1 combination was significantly more effective than either SAM or anti-PD-1 as the CD8+ T cells had higher activation, proliferation, and cytokine production compared to all other groups. This study shows that the combination of currently approved agents SAM and ICPi can effectively block melanoma via alteration of key genes/pathways implicated in cancer and immune response pathways, providing the rationale for the initiation of clinical trials with SAM and ICPi.
    Keywords:  DNA methylation; S-adenosylmethionine; anti-PD-1; immune checkpoint inhibitors; immunity; melanoma
  29. Cancers (Basel). 2020 Sep 24. pii: E2756. [Epub ahead of print]12(10):
      Pancreatic ductal adenocarcinoma (PDAC) is characterized by a mostly immunosuppressive microenvironment. Tumor-draining lymph nodes (TDLN) are a major site for priming of tumor-reactive T cells and also tumor metastasis. However, the phenotype and function of T cells in TDLNs from PDAC patients is unknown. In this study, lymph nodes from the pancreatic head (PH), the hepatoduodenal ligament (HDL) and the interaortocaval (IAC) region were obtained from 25 patients with adenocarcinoma of the pancreatic head. Additionally, tumors and matched blood were analyzed from 16 PDAC patients. Using multicolor flow cytometry, we performed a comprehensive analysis of T cells. CD4+ T cells were the predominant T cell subset in PDAC-draining lymph nodes. Overall, lymph node CD4+ and CD8+ T cells had a similar degree of activation, as measured by CD69, inducible T cell co-stimulator (ICOS) and CD137 (4-1BB) expression and interferon-γ (IFNγ) secretion. Expression of the inhibitory receptor programmed death 1 (PD-1) by lymph node and tumor-infiltrating regulatory T cells (Tregs) correlated with lymph node metastasis. Collectively, Treg cells and PD-1 are two relevant components of the immunosuppressive network in PDAC-draining lymph nodes and may be particularly attractive targets for combinatorial immunotherapeutic strategies in selected patients with node-positive PDAC.
    Keywords:  PD-1; T cells; Treg cells; pancreatic cancer; tumor-draining lymph nodes
  30. Nat Commun. 2020 Oct 02. 11(1): 4946
      Immune-related adverse events (irAEs), caused by anti-PD-1/PD-L1 antibodies, can lead to fulminant and even fatal consequences and thus require early detection and aggressive management. However, a comprehensive approach to identify biomarkers of irAE is lacking. Here, we utilize a strategy that combines pharmacovigilance data and omics data, and evaluate associations between multi-omics factors and irAE reporting odds ratio across different cancer types. We identify a bivariate regression model of LCP1 and ADPGK that can accurately predict irAE. We further validate LCP1 and ADPGK as biomarkers in an independent patient-level cohort. Our approach provides a method for identifying potential biomarkers of irAE in cancer immunotherapy using both pharmacovigilance data and multi-omics data.
  31. Cancer Res. 2020 Sep 30. pii: canres.1094.2020. [Epub ahead of print]
      The aggressive primary brain tumor glioblastoma (GBM) is characterized by aberrant metabolism that fuels its malignant phenotype. Diverse genetic subtypes of malignant glioma are sensitive to selective inhibition of the NAD+ salvage pathway enzyme nicotinamide phosphoribosyltransferase (NAMPT). However, the potential impact of NAD+ depletion on the brain tumor microenvironment has not been elaborated. In addition, systemic toxicity of NAMPT inhibition remains a significant concern. Here we show that microparticle-mediated intratumoral delivery of NAMPT inhibitor GMX1778 induces specific immunological changes in the tumor microenvironment of murine GBM, characterized by upregulation of immune checkpoint PD-L1, recruitment of CD3+, CD4+, and CD8+ T cells, and reduction of M2-polarized immunosuppressive macrophages. NAD+ depletion and autophagy induced by NAMPT inhibitors mediated the upregulation of PD-L1 transcripts and cell surface protein levels in GBM cells. NAMPT inhibitor modulation of the tumor immune microenvironment was therefore combined with PD-1 checkpoint blockade in vivo, significantly increasing the survival of GBM bearing animals. Thus, the therapeutic impacts of NAMPT inhibition extended beyond neoplastic cells, shaping surrounding immune effectors. Microparticle delivery and release of NAMPT inhibitor at the tumor site offers a safe and robust means to alter an immune tumor microenvironment that could potentiate checkpoint immunotherapy for glioblastoma.