The histone chaperone CAF-1 cooperates with the DNA methyltransferases to maintain Cd4 silencing in cytotoxic T cells.

The transcriptional repression of alternative lineage genes is critical for cell fate commitment. Mechanisms by which locus-specific gene silencing is initiated and heritably maintained during cell division are not clearly understood. To study the maintenance of silent gene states, we investigated how the Cd4 gene is stably repressed in CD8+ T cells. Through CRISPR and shRNA screening, we identified the histone chaperone CAF-1 as a critical component for Cd4 repression. We found that the large subunit of CAF-1, Chaf1a, requires the N-terminal KER domain to associate with the histone deacetylases HDAC1/2 and the histone demethylase LSD1, enzymes that also participate in Cd4 silencing. When CAF-1 was lacking, Cd4 derepression was markedly enhanced in the absence of the de novo DNA methyltransferase Dnmt3a but not the maintenance DNA methyltransferase Dnmt1. In contrast to Dnmt1, Dnmt3a deficiency did not significantly alter levels of DNA methylation at the Cd4 locus. Instead, Dnmt3a deficiency sensitized CD8+ T cells to Cd4 derepression mediated by compromised functions of histone-modifying factors, including the enzymes associated with CAF-1. Thus, we propose that the heritable silencing of the Cd4 gene in CD8+ T cells exploits cooperative functions among the DNA methyltransferases, CAF-1, and histone-modifying enzymes.

Polo-like kinase 1 regulates immune synapse assembly and cytotoxic T cell function by driving microtubule dynamics.

Elimination of virally infected or tumoral cells is mediated by cytotoxic T cells (CTL). Upon antigen recognition, CTLs assemble a specialized signaling and secretory domain at the interface with their target, the immune synapse (IS). During IS formation, CTLs acquire a transient polarity, marked by re-orientation of the centrosome and microtubule cytoskeleton toward the IS, thus directing the transport and delivery of the lytic granules to the target cell. Based on the implication that the kinase Aurora A has a role in CTL function, we hypothesized that its substrate, the mitotic regulator Polo-like kinase 1 (PLK1), might participate in CTL IS assembly. We demonstrate that PLK1 is phosphorylated upon TCR triggering and polarizes to the IS. PLK1 silencing or inhibition results in impaired IS assembly and function, as witnessed by defective synaptic accumulation of T cell receptors (TCRs), as well as compromised centrosome and lytic granule polarization to the IS, resulting in impaired target cell killing. This function is achieved by coupling early signaling to microtubule dynamics, a function pivotal for CTL-mediated cytotoxicity. These results identify PLK1 as a new player in CTL IS assembly and function.

Emerging roles of SARS-CoV-2 Spike-ACE2 in immune evasion and pathogenesis.

The COVID-19 pandemic, caused by SARS-CoV-2, has caused an estimated 5 billion infections and 20 million deaths by respiratory failure. In addition to the respiratory disease, SARS-CoV-2 infection has been associated with many extrapulmonary complications not easily explainable by the respiratory infection. A recent study showed that the SARS-CoV-2 spike protein, which mediates cell entry by binding to the angiotensin-converting enzyme 2 (ACE2) receptor, signals through ACE2 to change host cell behavior. In CD8+ T cells, spike-dependent ACE2-mediated signaling suppresses immunological synapse (IS) formation and impairs their killing ability, leading to immune escape of virus-infected cells. In this opinion article, we discuss the consequences of ACE2 signaling on the immune response and propose that it contributes to the extrapulmonary manifestations of COVID-19.

The levels of soluble CD137 are increased in tuberculosis patients and associated with disease severity and prognosis.

Tuberculosis (TB) was the leading cause of death from a single infectious agent before the coronavirus pandemic. Therefore, it is important to search for severity biomarkers and devise appropriate therapies. A total of 139 pulmonary TB (PTB) patients and 80 healthy controls (HCs) were recruited for plasma soluble CD137 (sCD137) detection through ELISA. Moreover, pleural effusion sCD137 levels were measured in 85 TB patients and 36 untreated lung cancer patients. The plasma cytokine levels in 64 patients with PTB and blood immune cell subpopulations in 68 patients with PTB were analysed via flow cytometry. Blood sCD137 levels were higher in PTB patients (p = 0.012) and correlated with disease severity (p = 0.0056). The level of sCD137 in tuberculous pleurisy effusion (TPE) was markedly higher than that in malignant pleurisy effusion (p = 0.018). Several blood cytokines, such as IL-6 (p = 0.0147), IL-8 (p = 0.0477), IP-10 (p ≤ 0.0001) and MCP-1 (p = 0.0057), and some laboratory indices were significantly elevated in severe PTB (SE) patients, but the percentages of total lymphocytes (p = 0.002) and cytotoxic T cells (p = 0.036) were significantly lower in SE patients than in non-SE patients. In addition, the sCD137 level was negatively correlated with the percentage of total lymphocytes (p = 0.0008) and cytotoxic T cells (p = 0.0021), and PTB patients with higher plasma sCD137 levels had significantly shorter survival times (p = 0.0041). An increase in sCD137 is a potential biomarker for severe TB and indicates a poor prognosis.

Cytotoxic T lymphocytes require transcription for infiltration but not target cell lysis.

Effector cytotoxic T lymphocytes (CTLs) are critical for ridding the body of infected or cancerous cells. CTL T cell receptor (TCR) ligation not only drives the delivery and release of cytolytic granules but also initiates a new wave of transcription. In order to address whether TCR-induced transcriptomic changes impact the ability of CTLs to kill, we asked which genes are expressed immediately after CTLs encounter targets and how CTL responses change when inhibiting transcription. Our data demonstrate that while expression of cytokines/chemokines and transcriptional machinery depend on transcription, cytotoxic protein expression and cytolytic activity are relatively robust to transcription blockade, with CTLs lysing nearby target cells for several hours after actinomycin D treatment. Monitoring CTL movement among target cells after inhibiting transcription demonstrates an infiltration defect that is not rectified by provision of exogenous cytokine/chemokine gradients, indicating a cell-intrinsic transcriptional requirement for infiltration. Together, our results reveal differential molecular control of CTL functions, separating recruitment and infiltration from cytolysis.

Newly developed humanized anti-CKAP4 antibody suppresses pancreatic cancer growth by inhibiting DKK1-CKAP4 signaling.

Cytoskeleton-associated protein 4 (CKAP4) is a cell surface receptor for Dickkopf 1 (DKK1), a secreted protein. The DKK1-CKAP4 pathway is activated in various malignant tumors, including pancreatic, lung, esophageal, and liver cancers, to promote tumor growth. Thus, CKAP4 has been expected to represent a novel molecular target of cancer therapy. Recombinant mouse anti-CKAP4 antibodies were generated based on an original mouse antibody (3F11-2B10) and inhibited DKK1-CKAP4 signaling and xenograft tumor formation induced by pancreatic cancer cells, which was comparable with 3F11-2B10. From the 3F11-2B10 nucleotide sequence, humanized anti-CKAP4 antibody (Hv1Lt1) was subsequently developed. The binding affinity of Hv1Lt1 for CKAP4 was superior to that of 3F11-2B10. Hv1Lt1 inhibited DKK1 binding to CKAP4, AKT activity, and sphere formation of pancreatic cancer cells, which was comparable with 3F11-2B10. Hv1Lt1 also suppressed xenograft tumor formation induced by human pancreatic cancer cells and tumor growth in murine cancer models, in which murine pancreatic cancer organoids were orthotopically transplanted into the pancreas. In resected tumor samples from mice treated with Hv1Lt1, anti-tumor immune reactions were modulated and cytotoxic T cells were highly infiltrated in the tumor microenvironment. Additionally, combination of Hv1Lt1 and other chemotherapy drugs exhibited stronger effects compared with monotherapy. These results suggest that Hv1Lt1 represents a promising anti-cancer therapy.

Clonal haematopoiesis: A common progenitor for cytotoxic peripheral T-cell lymphoma and angioimmunoblastic T-cell lymphoma.

Recent studies have shown that follicular helper T-cell lymphoma of angioimmunoblastic type (AITL), the most common nodal peripheral T-cell lymphoma (PTCL), frequently arises in a background of clonal haematopoiesis (CH), a preneoplastic condition affecting up to 40% of elderly individuals. Data on a potential CH association are limited for other PTCL. We report a unique patient who sequentially developed both cytotoxic PTCL, not otherwise specified and AITL with distinct T-cell receptor rearrangements but shared somatic mutations originating from the same CH clone, thus providing convincing evidence that CH can give rise to T-cell neoplasms of different lineage.

Deubiquitinating enzymes: potential regulators of the tumor microenvironment and implications for immune evasion.

Ubiquitination and deubiquitination are important forms of posttranslational modification that govern protein homeostasis. Deubiquitinating enzymes (DUBs), a protein superfamily consisting of more than 100 members, deconjugate ubiquitin chains from client proteins to regulate cellular homeostasis. However, the dysregulation of DUBs is reportedly associated with several diseases, including cancer. The tumor microenvironment (TME) is a highly complex entity comprising diverse noncancerous cells (e.g., immune cells and stromal cells) and the extracellular matrix (ECM). Since TME heterogeneity is closely related to tumorigenesis and immune evasion, targeting TME components has recently been considered an attractive therapeutic strategy for restoring antitumor immunity. Emerging studies have revealed the involvement of DUBs in immune modulation within the TME, including the regulation of immune checkpoints and immunocyte infiltration and function, which renders DUBs promising for potent cancer immunotherapy. Nevertheless, the roles of DUBs in the crosstalk between tumors and their surrounding components have not been comprehensively reviewed. In this review, we discuss the involvement of DUBs in the dynamic interplay between tumors, immune cells, and stromal cells and illustrate how dysregulated DUBs facilitate immune evasion and promote tumor progression. We also summarize potential small molecules that target DUBs to alleviate immunosuppression and suppress tumorigenesis. Finally, we discuss the prospects and challenges regarding the targeting of DUBs in cancer immunotherapeutics and several urgent problems that warrant further investigation.

History of Hemophagocytic Lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is a severe cytokine storm syndrome (CSS), which until the turn of the century, was barely known but is now receiving increased attention. The history of HLH dates back to 1939 when it was first described in adults, to be followed in 1952 by the first description of its primary, familial form in children. Secondary forms of HLH are far more frequent and occur with infections, malignancies, metabolic diseases, iatrogenic immune suppression, and autoinflammatory/autoimmune diseases. Identification of the genetic defects leading to the defective function of natural killer (NK) cells and cytotoxic T cells as well as the corresponding mouse models have revolutionized our understanding of HLH and of immune function. Diagnosis relies on clinical and laboratory criteria; functional and genetic tests can help separate primary from secondary forms. Treatment with immunochemotherapy and hematopoietic stem cell transplantation has considerably improved survival in children with primary HLH, a formerly uniformly fatal disease.

Immunocyte profiling changes in patients received epidural versus intravenous analgesia after pancreatectomy: A randomized controlled trial.

BACKGROUND: Perioperative immunosuppressants, such as surgical stress and opioid use may downregulate anti-cancer immunocytes for patients undergoing pancreatectomy. Thoracic epidural analgesia (TEA) may attenuate these negative effects and provide better anti-cancer immunocyte profile change than intravenous analgesia using opioid. METHODS: We randomly assigned 108 adult patients undergoing pancreatectomy to receive one of two 72-h postoperative analgesia protocols: one was TEA, and the other was intravenous patient-controlled analgesia (IV-PCA). The perioperative proportional changes of immunocytes relevant to anticancer immunity-namely natural killer (NK) cells, cytotoxic T cells, helper T cells, mature dendritic cells, and regulatory T (Treg) cells were determined at 1 day before surgery, at the end of surgery and on postoperative day 1,4 and 7 using flow cytometry. In addition, the progression-free survival and overall survival between the two groups were compared. RESULTS: After surgery, the proportions of NK cells and cytotoxic T cells were significantly decreased; the proportion of B cells and mature dendritic cells and Treg cells were significantly increased. However, the proportions of helper T cells exhibited no significant change. These results were comparable between the two groups. Furthermore, there were no significant differences in progression-free survival (52.75 [39.96] and 57.48 [43.66] months for patients in the TEA and IV-PCA groups, respectively; p = 0.5600) and overall survival (62.71 [35.48] and 75.11 [33.10] months for patients in the TEA and IV-PCA groups, respectively; p = 0.0644). CONCLUSIONS: TEA was neither associated with favorable anticancer immunity nor favorable oncological outcomes for patients undergoing pancreatectomy.

Localized cytotoxic T cell-associated antigen 4 and antioxidant islet encapsulation alters macrophage signaling and induces regulatory and anergic T cells to enhance allograft survival.

The loss of functional ß-cell mass is a hallmark of type 1 diabetes. Islet transplantation represents a promising alternative approach, but immune-mediated graft destruction remains a major challenge. We sought to use islet encapsulation technologies to improve graft survival and function without systemic immunosuppression. We hypothesized islet encapsulation with nanothin coatings consisting of tannic acid (TA), an antioxidant; poly(N-vinylpyrrolidone) (PVPON), a biocompatible polymer; and cytotoxic T cell-associated antigen 4 immunoglobulin (CTLA-4-Ig), an inhibitory immune receptor, will elicit localized immunosuppression to prolong islet allograft function and suppress effector T cell responses. In the absence of systemic immunosuppression, we demonstrated (PVPON/TA/CTLA-4-Ig)-encapsulated NOD.Rag islet grafts maintain function significantly longer than control IgG-containing (PVPON/TA/IgG) and nonencapsulated controls after transplantation into diabetic C57BL/6 mice. This protection coincided with diminished proinflammatory macrophage responses mediated by signal transducer and activator of transcription 1 signaling, decreased proinflammatory T cell effector responses, and CTLA-4-Ig-specific concomitant increases in anergic CD4+ T cells and regulatory T cells. Our results provide evidence that conjugation of CTLA-4-Ig to (PVPON/TA) coatings can suppress T cell activation, enhance regulatory T cell populations, prolong islet allograft survival, and induce localized immunosuppression after transplantation.

Interferon-Driven Immune Dysregulation in Common Variable Immunodeficiency-Associated Villous Atrophy and Norovirus Infection.

PURPOSE: About 15% of patients with common variable immunodeficiency (CVID) develop a small intestinal enteropathy, which resembles celiac disease with regard to histopathology but evolves from a distinct, poorly defined pathogenesis that has been linked in some cases to chronic norovirus (NV) infection. Interferon-driven inflammation is a prominent feature of CVID enteropathy, but it remains unknown how NV infection may contribute. METHODS: Duodenal biopsies of CVID patients, stratified according to the presence of villous atrophy (VA), IgA plasma cells (PCs), and chronic NV infection, were investigated by flow cytometry, multi-epitope-ligand cartography, bulk RNA-sequencing, and RT-qPCR of genes of interest. RESULTS: VA development was connected to the lack of intestinal (IgA+) PC, a T helper 1/T helper 17 cell imbalance, and increased recruitment of granzyme+CD8+ T cells and pro-inflammatory macrophages to the affected site. A mixed interferon type I/III and II signature occurred already in the absence of histopathological changes and increased with the severity of the disease and in the absence of (IgA+) PCs. Chronic NV infection exacerbated this signature when compared to stage-matched NV-negative samples. CONCLUSIONS: Our study suggests that increased IFN signaling and T-cell cytotoxicity are present already in mild and are aggravated in severe stages (VA) of CVID enteropathy. NV infection preempts local high IFN-driven inflammation, usually only seen in VA, at milder disease stages. Thus, revealing the impact of different drivers of the pathological mixed IFN type I/III and II signature may allow for more targeted treatment strategies in CVID enteropathy and supports the goal of viral elimination.

The presence of serum anti-SARS-CoV-2 IgA appears to protect primary health care workers from COVID-19.

The patterns of humoral and cellular responses to SARS-CoV-2 were studied in Swedish primary health care workers (n = 156) for 6 months during the Covid-19 pandemic. Serum IgA and IgG to SARS-CoV-2, T-cell proliferation and cytokine secretion, demographic and clinical data, PCR-verified infection, and self-reported symptoms were monitored. The multivariate method OPLS-DA was used to identify immune response patterns coupled to protection from Covid-19. Contracting Covid-19 was associated with SARS-CoV-2-specific neutralizing serum IgG, T cell, IFN-γ, and granzyme B responses to SARS-CoV-2, self-reported typical Covid-19 symptoms, male sex, higher BMI, and hypertension. Not contracting Covid-19 was associated with female sex, IgA-dominated, or no antibody responses to SARS-CoV-2, airborne allergy, and smoking. The IgG-responders had SARS-CoV-2-specific T-cell responses including a cytotoxic CD4+ T-cell population expressing CD25, CD38, CD69, CD194, CD279, CTLA-4, and granzyme B. IgA-responders with no IgG response to SARS-CoV-2 constituted 10% of the study population. The IgA responses were partially neutralizing and only seen in individuals who did not succumb to Covid-19. To conclude, serum IgG-dominated responses correlated with T-cell responses to SARS-CoV-2 and PCR-confirmed Covid-19, whereas IgA-dominated responses correlated with not contracting the infection.

GPC3 and PEG10 peptides associated with placental gp96 elicit specific T cell immunity against hepatocellular carcinoma.

The placenta and tumors can exhibit a shared expression profile of proto-oncogenes. The basis of placenta-derived heat shock protein gp96, which induces prophylactic and therapeutic T cell responses against cancer including hepatocellular carcinoma (HCC), remains unknown. Here, we identified the associated long peptides from human placental gp96 using matrix-assisted laser desorption/ionization-time-of-flight and mass spectrometry and analyzed the achieved proteins through disease enrichment analysis. We found that placental gp96 binds to numerous peptides derived from 73 proteins that could be enriched in multiple cancer types. Epitope-harboring peptides from glypican 3 (GPC3) and paternally expressed gene 10 (PEG10) were the major antigens mediating anti-HCC T cell immunity. Molecular docking analysis showed that the GPC3- and PEG10-derived peptides, mainly obtained from the cytotrophoblast layer of the mature placenta, bind to the lumenal channel and client-bound domain of the gp96 dimer. Immunization with bone marrow-derived dendritic cells pulsed with recombinant gp96-GPC3 or recombinant gp96-PEG10 peptide complex induced specific T cell responses, and T cell transfusion led to pronounced growth inhibition of HCC tumors in nude mice. We demonstrated that the chaperone gp96 can capture antigenic peptides as an efficient approach for defining tumor rejection oncoantigens in the placenta and provide a basis for developing GPC3 and PEG10 peptide-based vaccines against HCC. This study provides insight into the underlying mechanism of the antitumor response mediated by embryonic antigens from fetal tissues, and this will incite more studies to identify potential tumor rejection antigens from placenta.

Engineered CD8+ T cell-derived extracellular vesicles induce enhanced anti-cancer efficacy and targeting to lung cancer cells.

Lung cancer is a common and highly malignant tumor. Although lung cancer treatments continue to advance, conventional therapies are limited and the response rate of patients to immuno-oncology drugs is low. This phenomenon raises an urgent need to develop effective therapeutic strategies for lung cancer. In this study, we genetically modified human primary CD8+ T cells and obtained antitumor extracellular vesicles (EVs) from them. The engineered EVs, containing interlekin-2 and the anti-epidermal growth factor receptor (EGFR) antibody cetuximab on their surfaces, exhibited direct cytotoxicity against A549 human lung cancer cells and increased cancer cell susceptibility to human peripheral blood mononuclear cell-mediated cytotoxicity. In addition, the engineered EVs specifically targeted the lung cancer cells in an EGFR-dependent manner. Taken together, these findings show that surface engineering of cytokines and antibodies on CD8+ T cell-derived EVs not only enhances their antitumor effects but also confers target specificity, suggesting a potential of modifying the immune cell-derived EVs in cancer treatment.

A Review of Immune Checkpoint Blockade for the General Surgeon.

The immune system is a complex and interconnected system that has evolved to protect its host from foreign pathogens. CD8+ T cells are a type of immune cell that can be directly lethal to tumor cells. However, their tumor killing capabilities can be inhibited by checkpoint molecules. During the last decade, the development of medications that block these checkpoint molecules has revolutionized treatment for some cancer types and indications for use continue to grow. As usage of immunotherapy increases, toxicities and adverse events unique to immunotherapy are becoming more prevalent. Here, we review the commonly targeted inhibitory molecules along with their food and drug administration-approved indications in various cancer therapeutic regimens, immunotherapy-related toxicities, and how this may impact surgical planning.

Combining PARP Inhibition and Immunotherapy in BRCA-Associated Cancers.

Poly (ADP-ribose) polymerase (PARP) inhibitors have significantly improved treatment outcomes of homologous recombination (HR) repair-deficient cancers. While the activity of these agents is largely linked to multiple mechanisms underlying the synthetic lethality of PARP inhibition and HR deficiency, emerging data suggest that their efficacy is also tied to their effects on the immune microenvironment and dependent upon cytotoxic T-cell activation. Effects observed in preclinical models are currently being validated in on-treatment biopsy samples procured from patients enrolled in clinical trials. Although this work has stimulated the development of combinations of PARP inhibitors with immunomodulatory agents, results to date have not demonstrated the superiority of combined PARP inhibition and immune checkpoint blockade compared with PARP inhibition alone. These results have stimulated a more comprehensive assessment of the immunosuppressive components of the tumor microenvironment that must be addressed so that the efficacy of PARP inhibitor agents can be maximized.

Non-specific cytotoxic cell receptor protein-1 (NCCRP-1) is involved in anti-parasite innate CD8+ T cell-mediated cytotoxicity in ginbuna crucian carp.

CD8+ cytotoxic T cells (CTLs) are a main cellular component of adaptive immunity. Our previous research has shown that CD8+ cells demonstrate spontaneous cytotoxic activity against the parasite Ichthyophthirius multifiliis in ginbuna crucian carp, suggesting that CD8+ cells play an important role in innate immunity. Herein, we investigated the molecules and cellular signal pathways involved in the cytotoxic response of ginbuna crucian carp. We considered non-specific cytotoxic receptor protein-1 (NCCRP-1) as candidate molecule for parasite recognition. We detected NCCRP-1 protein in CD8+ cells and the thymus as well as in other cells and tissues. CD8+ cells expressed mRNA for NCCRP-1, Jak2, and T cell-related molecules. In addition, treatment with a peptide containing the presumed antigen recognition site of ginbuna NCCRP-1 significantly inhibited the cytotoxic activity of CD8+ cells against the parasites. The cytotoxic activity of CD8+ cells was significantly inhibited by treatment with the JAK1/2 inhibitor baricitinib. These results suggest that teleost CTLs recognize I. multifiliis through NCCRP-1 and are activated by JAK/STAT signaling.

Mixed lineage kinase 3 inhibition induces T cell activation and cytotoxicity.

Mixed lineage kinase 3 (MLK3), also known as MAP3K11, was initially identified in a megakaryocytic cell line and is an emerging therapeutic target in cancer, yet its role in immune cells is not known. Here, we report that loss or pharmacological inhibition of MLK3 promotes activation and cytotoxicity of T cells. MLK3 is abundantly expressed in T cells, and its loss alters serum chemokines, cytokines, and CD28 protein expression on T cells and its subsets. MLK3 loss or pharmacological inhibition induces activation of T cells in in vitro, ex vivo, and in vivo conditions, irrespective of T cell activating agents. Conversely, overexpression of MLK3 decreases T cell activation. Mechanistically, loss or inhibition of MLK3 down-regulates expression of a prolyl-isomerase, Ppia, which is directly phosphorylated by MLK3 to increase its isomerase activity. Moreover, MLK3 also phosphorylates nuclear factor of activated T cells 1 (NFATc1) and regulates its nuclear translocation via interaction with Ppia, and this regulates T cell effector function. In an immune-competent mouse model of breast cancer, MLK3 inhibitor increases Granzyme B-positive CD8+ T cells and decreases MLK3 and Ppia gene expression in tumor-infiltrating T cells. Likewise, the MLK3 inhibitor in pan T cells, isolated from breast cancer patients, also increases cytotoxic CD8+ T cells. These results collectively demonstrate that MLK3 plays an important role in T cell biology, and targeting MLK3 could serve as a potential therapeutic intervention via increasing T cell cytotoxicity in cancer.

Inhibitory Effect of Zinc on Colorectal Cancer by Granzyme B Transcriptional Regulation in Cytotoxic T Cells.

Zinc is one of the essential trace elements and is involved in various functions in the body. Zinc deficiency is known to cause immune abnormalities, but the mechanism is not fully understood. Therefore, we focused our research on tumor immunity to elucidate the effect of zinc on colorectal cancer and its mechanisms. Mice were treated with azoxymethane (AOM) and dextran sodium sulfate (DSS) to develop colorectal cancer, and the relationship between zinc content in the diet and the number and area of tumors in the colon was observed. The number of tumors in the colon was significantly higher in the no-zinc-added group than in the normal zinc intake group, and about half as many in the high-zinc-intake group as in the normal-zinc-intake group. In T-cell-deficient mice, the number of tumors in the high-zinc-intake group was similar to that in the normal-zinc-intake group, suggesting that the inhibitory effect of zinc was dependent on T cells. Furthermore, we found that the amount of granzyme B transcript released by cytotoxic T cells upon antigen stimulation was significantly increased by the addition of zinc. We also showed that granzyme B transcriptional activation by zinc addition was dependent on calcineurin activity. In this study, we have shown that zinc exerts its tumor-suppressive effect by acting on cytotoxic T cells, the center of cellular immunity, and increases the transcription of granzyme B, one of the key molecules in tumor immunity.