SARS-CoV-2-specific T cells are rapidly expanded for therapeutic use and target conserved regions of the membrane protein.

T-cell responses to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have been described in recovered patients, and may be important for immunity following infection and vaccination as well as for the development of an adoptive immunotherapy for the treatment of immunocompromised individuals. In this report, we demonstrate that SARS-CoV-2-specific T cells can be expanded from convalescent donors and recognize immunodominant viral epitopes in conserved regions of membrane, spike, and nucleocapsid. Following in vitro expansion using a good manufacturing practice-compliant methodology (designed to allow the rapid translation of this novel SARS-CoV-2 T-cell therapy to the clinic), membrane, spike, and nucleocapsid peptides elicited interferon-γ production, in 27 (59%), 12 (26%), and 10 (22%) convalescent donors (respectively), as well as in 2 of 15 unexposed controls. We identified multiple polyfunctional CD4-restricted T-cell epitopes within a highly conserved region of membrane protein, which induced polyfunctional T-cell responses, which may be critical for the development of effective vaccine and T-cell therapies. Hence, our study shows that SARS-CoV-2 directed T-cell immunotherapy targeting structural proteins, most importantly membrane protein, should be feasible for the prevention or early treatment of SARS-CoV-2 infection in immunocompromised patients with blood disorders or after bone marrow transplantation to achieve antiviral control while mitigating uncontrolled inflammation.

Relationships among green space, ambient fine particulate matter, and cancer incidence in Taiwan: A 16-year retrospective cohort study.

INTRODUCTION: Green space and air pollution have been recognized as vital health determinants. There is a paucity of studies examining the interplay between green space, fine particulate matter (PM2.5), and the incidence of specific cancers. OBJECTIVE: We aimed to explore the contributions of green space and ambient PM2.5 to the risk of specific cancers in terms of the most common cancers based on incidence or mortality rate in Taiwan and to ascertain the interaction between green space and PM2.5 and their role in cancer risk. MATERIALS AND METHODS: This retrospective longitudinal cohort study included 407,415 participants. Data were obtained from the 2000-2015 Mei Jau Health Examination Database linked to the Taiwan Cancer Registry and Causes of Death datasets. All participants were aged ≥20 years and had no history of cancer. The environmental exposure were the normalized difference vegetation index (NDVI) and the 2-year average PM2.5 at baseline. Multivariate adjusted hazard ratios (HRs) were calculated using Cox proportional hazards models. We adjusted for covariates including demographics, anthropometrics, comorbidities, health behaviors, biochemical data, and environmental factors. RESULTS: During a median follow-up of 10.37 years, 11,576 cancer cases were reported. PM2.5 exposure increased the risk of all cancers (HR: 1.11, [95% CI: 1.06-1.15]), stomach cancer (HR: 1.27, [1.02-1.58]), endocrine gland cancer (HR: 2.13, [1.39-3.26]), breast cancer (HR: 1.12, [1.03-1.22]), and lung cancer (HR: 1.12, [1.01-1.24]). An increase in NDVI reduced the risk of prostate cancer (HR: 0.93, [0.88-0.99]) and lung cancer (HR: 0.95, [0.91-0.99]). NDVI influenced the incidence of prostate and all cancers by reducing PM2.5 concentrations. CONCLUSION: Long-term PM2.5 exposure is associated with an increased risk of some types of cancers. In contrast, an increase in environmental green space exposure is associated with lowering of the risk of prostate and lung cancer.

Identification of novel HLA-restricted preferentially expressed antigen in melanoma peptides to facilitate off-the-shelf tumor-associated antigen-specific T-cell therapies.

BACKGROUND AIMS: Preferentially expressed antigen in melanoma (PRAME) is a cancer/testis antigen that is overexpressed in many human malignancies and poorly expressed or absent in healthy tissues, making it a good target for anti-cancer immunotherapy. Development of an effective off-the-shelf adoptive T-cell therapy for patients with relapsed or refractory solid tumors and hematological malignancies expressing PRAME antigen requires the identification of major histocompatibility complex (MHC) class I and II PRAME antigens recognized by the tumor-associated antigen (TAA) T-cell product. The authors therefore set out to extend the repertoire of HLA-restricted PRAME peptide epitopes beyond the few already characterized. METHODS: Peptide libraries of 125 overlapping 15-mer peptides spanning the entire PRAME protein sequence were used to identify HLA class I- and II-restricted epitopes. The authors also determined the HLA restriction of the identified epitopes. RESULTS: PRAME-specific T-cell products were successfully generated from peripheral blood mononuclear cells of 12 healthy donors. Ex vivo-expanded T cells were polyclonal, consisting of both CD4+ and CD8+ T cells, which elicited anti-tumor activity in vitro. Nine MHC class I-restricted PRAME epitopes were identified (seven novel and two previously described). The authors also characterized 16 individual 15-mer peptide sequences confirmed as CD4-restricted epitopes. CONCLUSIONS: TAA T cells derived from healthy donors recognize a broad range of CD4+ and CD8+ HLA-restricted PRAME epitopes, which could be used to select suitable donors for generating off-the-shelf TAA-specific T cells.

The transcription factor E74-like factor 4 suppresses differentiation of proliferating CD4+ T cells to the Th17 lineage.

The differentiation of CD4(+) T cells into different Th lineages is driven by cytokine milieu in the priming site and the underlying transcriptional circuitry. Even though many positive regulators have been identified, it is not clear how this process is inhibited at transcriptional level. In this study, we report that the E-twenty six (ETS) transcription factor E74-like factor 4 (ELF4) suppresses the differentiation of Th17 cells both in vitro and in vivo. Culture of naive Elf4(-/-) CD4(+) T cells in the presence of IL-6 and TGF-ß (or IL-6, IL-23, and IL-1ß) resulted in increased numbers of IL-17A-positive cells compared with wild-type controls. In contrast, the differentiation to Th1, Th2, or regulatory T cells was largely unaffected by loss of ELF4. The increased expression of genes involved in Th17 differentiation observed in Elf4(-/-) CD4(+) T cells suggested that ELF4 controls their programming into the Th17 lineage rather than only IL-17A gene expression. Despite normal proliferation of naive CD4(+) T cells, loss of ELF4 lowered the requirement of IL-6 and TGF-ß signaling for IL-17A induction in each cell division. ELF4 did not inhibit Th17 differentiation by promoting IL-2 production as proposed for another ETS transcription factor, ETS1. Elf4(-/-) mice showed increased numbers of Th17 cells in the lamina propria at steady state, in lymph nodes after immunization, and, most importantly, in the CNS following experimental autoimmune encephalomyelitis induction, contributing to the increased disease severity. Collectively, our findings suggest that ELF4 restrains Th17 differentiation in dividing CD4(+) T cells by regulating commitment to the Th17 differentiation program.

G0S2 modulates homeostatic proliferation of naïve CD8⁺ T cells and inhibits oxidative phosphorylation in mitochondria.

Since its discovery, diverse functions have been attributed to the G0/G1 switch gene 2 (G0S2), from lipid metabolism to control of cell proliferation. Our group showed for the first time that G0S2 promotes quiescence in hematopoietic stem cells by interacting with and retaining nucleolin around the nucleus. Herein, we report the role of G0S2 in the differentiation and function of CD8(+) T cells examined in mice with an embryonic deletion of the G0s2 gene. G0S2 expression in naïve CD8(+) T cells decreased immediately after T-cell receptor activation downstream of the mitogen-activated protein kinase, calcium/calmodulin, phosphatidylinositol 3'-kinase and mammalian target of rapamycin pathways. Surprisingly, G0S2-null naïve CD8(+) T cells displayed increased basal and spare respiratory capacity that was not associated with increased mitochondrial biogenesis but with increased phosphorylation of AMP-activated protein kinase α. Naïve CD8(+) T cells showed increased proliferation in response to in vitro activation and in vivo lymphopenia; however, naïve CD8(+) T cells expressing the OT-1 transgene exhibited normal differentiation of naïve cells to effector and memory CD8(+) T cells upon infection with Listeria monocytogenes in a wild-type or a G0s2-null environment, with increased circulating levels of free fatty acids. Collectively, our results suggest that G0S2 inhibits energy production by oxidative phosphorylation to fine-tune proliferation in homeostatic conditions.

Effects of personal noise exposure, sleep quality, and burnout on quality of life: An online participation cohort study in Taiwan.

Chronic noise exposure in daily life not only causes physical and mental illness but also reduces quality of life. However, collecting objective data on sound exposure and subjective acoustic comfort through a traditional one-shot survey is difficult. This study applied online chatbots in social media to explore the effects of daily sound exposure, personal characteristics, noise sensitivity, burnout status, and sleep quality on quality of life using a short-term participatory cohort design. During the two-month survey in 2022, 207 participants completed at least 15 days of collection of data on sound exposure and perception, as well as periodic structural questionnaires during the follow-ups. Linear regression and generalized linear models were applied to explore the factors influencing personal burnout, the Pittsburgh Sleep Quality Index, and quality of life. A chain mediation model was applied to explore the direct and indirect effects of noise exposure on quality of life. The results showed a better quality of life among respondents who rated their home environment better, were in good health, had better daily acoustic comfort, and were less exposed to noise during the week. In contrast, respondents with lower daily acoustic comfort and a higher frequency of noise-induced sleep disturbances and mood disorders were more likely to have poorer sleep quality. A higher personal burnout was associated with poorer health, longer exposure to noise during the week, a higher frequency of noise-induced illnesses, and neurotic traits. In the mediation analyses, noise-induced sleep disturbance and better daily acoustic comfort also had important direct influences on quality of life compared to the indirect pathway through sleep quality and personal burnout. In conclusion, noise exposure in daily life not only exacerbated poor sleep quality and personal burnout but also reduced quality of life.

Exploring the spatial association between the distribution of temperature and urban morphology with green view index.

Urban heat islands will occur if city neighborhoods contain insufficient green spaces to create a comfortable environment, and residents' health will be adversely affected. Current satellite imagery can only effectively identify large-scale green spaces and cannot capture street trees or potted plants within three-dimensional building spaces. In this study, we used a deep convolutional neural network semantic segmentation model on Google Street View to extract environmental features at the neighborhood level in Taipei City, Taiwan, including the green vegetation index (GVI), building view factor, and sky view factor. Monthly temperature data from 2018 to 2021 with a 0.01° spatial resolution were used. We applied a linear mixed-effects model and geographically weighted regression to explore the association between pedestrian-level green spaces and ambient temperature, controlling for seasons, land use information, and traffic volume. Their results indicated that a higher GVI was significantly associated with lower ambient temperatures and temperature differences. Locations with higher traffic flows or specific land uses, such as religious or governmental, are associated with higher ambient temperatures. In conclusion, the GVI from street-view imagery at the community level can improve the understanding of urban green spaces and evaluate their effects in association with other social and environmental indicators.

Associations between community green view index and fine particulate matter from Airboxes.

Urban greenery can help to improve air quality, reduce health risks and create healthy livable urban communities. This study aimed to explore the role of urban greenery in reducing air pollution at the community level in Tainan City, Taiwan, using air quality sensors and street-view imagery. We also collected the number of road trees around each air quality sensor site and identified the species that were best at absorbing PM2.5. Three greenness metrics were used to assess community greenery in this study: two Normalized Difference Vegetation Indices (NDVI) from different satellites and the Green View Index (GVI) from Google Street View (GSV) images. Land-use Regression (LUR) was used for statistical analysis. The results showed that a higher GVI within a 500 m buffer was significantly associated with decreased PM2.5. Neither NDVI metrics within a 500 m circular buffer were significantly associated with decreased PM2.5. Evergreen trees were significantly associated with lower ambient PM2.5, compared with deciduous and semi-deciduous trees. Because localized changes in air quality profoundly affect public health and environmental equity, our findings provide evidence for future urban community greenspace planning and its beneficial impacts on reducing air pollution.

Cutting edge: Expression of the transcription factor E74-like factor 4 is regulated by the mammalian target of rapamycin pathway in CD8+ T cells.

T cell receptor activation inhibits expression of the E74-like factor (ELF) 4 and Krüppel-like factor 4 genes to release naive CD8(+) T cells from their quiescent state. In this study, we show that ELF4 controls the ERK-mediated proliferative response by maintaining normal levels of dual-specificity phosphatases 1 and 5 in CD8(+) T cells. In activated CD8(+) T cells, the mammalian target of rapamycin pathway inhibits ELF4 and Krüppel-like factor 4 expression downstream of ERK and PI3K signaling. Our findings demonstrate that rapamycin could be used to modulate expression of this transcriptional network involved in cell-cycle regulation.

Virus-Specific T Cell Therapies for HIV: Lessons Learned From Hematopoietic Stem Cell Transplantation.

Human immunodeficiency virus (HIV) has caused millions of deaths and continues to threaten the health of millions of people worldwide. Despite anti-retroviral therapy (ART) substantially alleviating severity and limiting transmission, HIV has not been eradicated and its persistence can lead to other health concerns such as cancer. The only two cases of HIV cure to date are HIV+ cancer patients receiving an allogeneic hematopoietic stem cell transplantation (allo-HSCT) from a donor with the CCR5 Δ32 mutation. While this approach has not led to such success in other patients and is not applicable to HIV+ individuals without cancer, the encouraging results may point toward a breakthrough in developing a cure strategy for HIV. Adoptive transfer of virus-specific T cells (VSTs) post HSCT has been effectively used to treat and prevent reactivation of latent viral infections such as cytomegalovirus (CMV) and Epstein-Barr virus (EBV), making VSTs an attractive therapeutic to control HIV rebound. Here we will discuss the potential of using adoptive T cell therapies in combination with other treatments such as HSCT and latency reversing agents (LRAs) to achieve a functional cure for HIV.

Host conditioning with IL-1ß improves the antitumor function of adoptively transferred T cells.

Host conditioning has emerged as an important component of effective adoptive cell transfer-based immunotherapy for cancer. High levels of IL-1ß are induced by host conditioning, but its impact on the antitumor function of T cells remains unclear. We found that the administration of IL-1ß increased the population size and functionality of adoptively transferred T cells within the tumor. Most importantly, IL-1ß enhanced the ability of tumor-specific T cells to trigger the regression of large, established B16 melanoma tumors in mice. Mechanistically, we showed that the increase in T cell numbers was associated with superior tissue homing and survival abilities and was largely mediated by IL-1ß-stimulated host cells. In addition, IL-1ß enhanced T cell functionality indirectly via its actions on radio-resistant host cells in an IL-2- and IL-15-dependent manner. Our findings not only underscore the potential of provoking inflammation to enhance antitumor immunity but also uncover novel host regulations of T cell responses.

T cells genetically engineered to overcome death signaling enhance adoptive cancer immunotherapy.

Across clinical trials, T cell expansion and persistence following adoptive cell transfer (ACT) have correlated with superior patient outcomes. Herein, we undertook a pan-cancer analysis to identify actionable ligand-receptor pairs capable of compromising T cell durability following ACT. We discovered that FASLG, the gene encoding the apoptosis-inducing ligand FasL, is overexpressed within the majority of human tumor microenvironments (TMEs). Further, we uncovered that Fas, the receptor for FasL, is highly expressed on patient-derived T cells used for clinical ACT. We hypothesized that a cognate Fas-FasL interaction within the TME might limit both T cell persistence and antitumor efficacy. We discovered that genetic engineering of Fas variants impaired in the ability to bind FADD functioned as dominant negative receptors (DNRs), preventing FasL-induced apoptosis in Fas-competent T cells. T cells coengineered with a Fas DNR and either a T cell receptor or chimeric antigen receptor exhibited enhanced persistence following ACT, resulting in superior antitumor efficacy against established solid and hematologic cancers. Despite increased longevity, Fas DNR-engineered T cells did not undergo aberrant expansion or mediate autoimmunity. Thus, T cell-intrinsic disruption of Fas signaling through genetic engineering represents a potentially universal strategy to enhance ACT efficacy across a broad range of human malignancies.

T cell stemness and dysfunction in tumors are triggered by a common mechanism.

A paradox of tumor immunology is that tumor-infiltrating lymphocytes are dysfunctional in situ, yet are capable of stem cell-like behavior including self-renewal, expansion, and multipotency, resulting in the eradication of large metastatic tumors. We find that the overabundance of potassium in the tumor microenvironment underlies this dichotomy, triggering suppression of T cell effector function while preserving stemness. High levels of extracellular potassium constrain T cell effector programs by limiting nutrient uptake, thereby inducing autophagy and reduction of histone acetylation at effector and exhaustion loci, which in turn produces CD8+ T cells with improved in vivo persistence, multipotency, and tumor clearance. This mechanistic knowledge advances our understanding of T cell dysfunction and may lead to novel approaches that enable the development of enhanced T cell strategies for cancer immunotherapy.

Mg2+ regulation of kinase signaling and immune function.

Mg2+ is required at micromolar concentrations as a cofactor for ATP, enzymatic reactions, and other biological processes. We show that decreased extracellular Mg2+ reduced intracellular Mg2+ levels and impaired the Ca2+ flux, activation marker up-regulation, and proliferation after T cell receptor (TCR) stimulation. Reduced Mg2+ specifically impairs TCR signal transduction by IL-2-inducible T cell kinase (ITK) due to a requirement for a regulatory Mg2+ in the catalytic pocket of ITK. We also show that altered catalytic efficiency by millimolar changes in free basal Mg2+ is an unrecognized but conserved feature of other serine/threonine and tyrosine kinases, suggesting a Mg2+ regulatory paradigm of kinase function. Finally, a reduced serum Mg2+ concentration in mice causes an impaired CD8+ T cell response to influenza A virus infection, reduces T cell activation, and exacerbates morbidity. Thus, Mg2+ directly regulates the active site of specific kinases during T cell responses, and maintaining a high serum Mg2+ concentration is important for antiviral immunity in otherwise healthy animals.

A cleavage product of Polycystin-1 is a mitochondrial matrix protein that affects mitochondria morphology and function when heterologously expressed.

Recent studies have reported intrinsic metabolic reprogramming in Pkd1 knock-out cells, implicating dysregulated cellular metabolism in the pathogenesis of polycystic kidney disease. However, the exact nature of the metabolic changes and their underlying cause remains controversial. We show herein that Pkd1 k o /ko renal epithelial cells have impaired fatty acid utilization, abnormal mitochondrial morphology and function, and that mitochondria in kidneys of ADPKD patients have morphological alterations. We further show that a C-terminal cleavage product of polycystin-1 (CTT) translocates to the mitochondria matrix and that expression of CTT in Pkd1 ko/ko cells rescues some of the mitochondrial phenotypes. Using Drosophila to model in vivo effects, we find that transgenic expression of mouse CTT results in decreased viability and exercise endurance but increased CO2 production, consistent with altered mitochondrial function. Our results suggest that PC1 may play a direct role in regulating mitochondrial function and cellular metabolism and provide a framework to understand how impaired mitochondrial function could be linked to the regulation of tubular diameter in both physiological and pathological conditions.

Memory-phenotype CD4+ T cells spontaneously generated under steady-state conditions exert innate TH1-like effector function.

Conventional CD4+ T cells are composed of naïve, pathogen-specific memory, and pathogen-independent memory-phenotype (MP) cells under steady state. Naïve and pathogen-specific memory cells play key roles in adaptive immunity, whereas the homeostatic mechanisms regulating the generation of MP cells and their biological functions are unclear. We show that MP cells are autonomously generated from peripheral naïve cells in the absence of infectious stimulation in a T cell receptor (TCR)- and CD28-dependent manner. We further demonstrate that MP cells contain a T-bethi subpopulation that is continuously generated by environmental interleukin-12 (IL-12) and rapidly produces interferon-γ (IFN-γ) in response to IL-12 in the absence of pathogen recognition. These cells can provide nonspecific host resistance against Toxoplasma gondii infection while enhancing the adaptive CD4+ T cell responses. Together, these findings reveal that MP cells are continuously generated from naïve precursors and have a previously undescribed innate immune function by which they produce an early, T helper cell type 1 (TH1)-like protective response against pathogens.

Differential roles of KLF4 in the development and differentiation of CD8+ T cells.

The transcription factor Krüppel-like factor 4 (KLF4) can activate or repress gene expression in a cell-context dependent manner. We have previously shown that KLF4 inhibits the proliferation of naïve CD8(+) T cells in vitro downstream of the transcription factor ELF4. In this work, we describe a novel role of KLF4 in the differentiation of CD8(+) T cells upon infection. Loss of KLF4 had minimal effect on thymic T cell development and distribution of mature T cells in the spleen, blood, and lymph nodes. KLF4-deficient naïve CD8(+) T cells also displayed normal homeostatic proliferation upon adoptive transfer into lymphopenic hosts. However, activation of KLF4-deficient naïve CD8(+) T cells by in vitro TCR crosslink and co-stimulation resulted in increased proliferation. Furthermore, naïve KLF4-deficient OT-I CD8(+) T cells generated increased numbers of functional memory CD8(+) T cells compared to wild type OT-I CD8(+) T cells co-injected in the same recipient in both primary and recall responses to Listeria monocytogenes-OVA. Collectively, our data demonstrate that KLF4 regulates differentiation of functional memory CD8(+) T cells while sparing development and homeostasis of naïve CD8(+) T cells.

Kruppel-like factor 4 (KLF4) promotes the survival of natural killer cells and maintains the number of conventional dendritic cells in the spleen.

The development and survival of NK cells rely on a complex, spatiotemporal gene expression pattern regulated by specific transcription factors in NK cells and tissue-specific microenvironments supported by hematopoietic cells. Here, we show that somatic deletion of the KLF4 gene, using inducible and lineage-specific cre-transgenic mice, leads to a significant reduction of NK cells (NK1.1(+) TCR-ß(-)) in the blood and spleen but not in the BM, liver, or LNs. Functional and immunophenotypic analyses revealed increased apoptosis of CD27(+/-) CD11b(+) NK cells in the spleen of KLF4-deficient mice, although remaining NK cells were able to lyse tumor target cells and produce IFN-γ. A normal recovery of adoptively transferred KLF4-deficient NK cells in WT hosts suggested that the survival defect was not intrinsic of NK cells. However, BM chimeras using KLF4-deficient mice as donors indicated that reduced survival of NK cells depended on BM-derived hematopoietic cells in the spleen. The number of CD11c(hi) DCs, which are known to support NK cell survival, was reduced significantly in the spleen of KLF4-deficient mice, likely a result of a lower number of precDC progenitor cells in this tissue. Taken together, our data suggest that the pluripotency-associated gene KLF4 is required for the maintenance of DCs in the spleen and consequently, survival of differentiated NK cells in this tissue.