Natural Killer (NK) cells are cytotoxic innate lymphocytes able to detect transformed cells through the balanced action of inhibitory and activating receptors. NKG2D is one of the main activating receptors involved in tumor surveillance thanks to its ability to recognize stress-induced ligands. Of note, the prolonged exposure to NKG2D ligands promotes receptor down-modulation that results in defective activation of NKG2D and other unrelated activating receptors, including DNAM-1 that is also involved in tumor clearance. However, further investigations are necessary to characterize how the NKG2D/DNAM-1 interplay affects NK cell anti-tumor function.

Primary cultured human NK cells were stimulated with the natural ligand MICA or an anti-NKG2D agonist antibody. The expression of activating and inhibitory receptors as well as DNAM-1-triggered signaling events and cytotoxicity were evaluated by flow cytometry. DNAM-1-mediated granule polarization was evaluated by confocal microscopy.

We showed that NKG2D crosslinking mediated by the natural ligand MICA or an agonist antibody had different consequences on primary cultured human NK cells. Indeed, MICA stimulation increases the expression of the checkpoint receptor TIGIT that is able to counteract DNAM-1 activation. Stimulation with the agonist antibody, without altering TIGIT expression, directly inhibits DNAM-1-mediated signal transduction and cytotoxic function with a mechanism that required NKG2D endocytosis.

Our findings contribute to shed light on the functional consequences of NKG2D engagement, demonstrating that a direct impact on DNAM-1-mediated signal transduction occurs independently from the modality of NKG2D crosslinking. Understanding the molecular mechanisms responsible for suppression of NK cell activation may help the development of therapeutic anti-cancer strategies aimed to prevent NK cell dysfunction or to reinvigorate an impaired cytotoxic activity.

As pivotal effectors of antiviral immunity, natural killer (NK) cells are crucial for controlling the spread of COVID-19. The nonstructural protein 13 of SARS-CoV-2 can encode a viral peptide (Nsp13232-240) preventing human leukocyte antigen E (HLA-E) from recognizing inhibitory receptor NKG2A, thereby activating NK cells. The underlying molecular mechanisms of Nsp13232-240 remain unclear. Therefore, we compared the interaction discrepancy between the self-peptide and Nsp13232-240, theoretically predicting its source. Results indicate that electrostatic interaction energy provides the main source of binding, and its attenuation greatly promotes binding affinity differences. Nsp13232-240 disrupts the hydrogen bond network between CD94 and HLA-E, impacting the binding of hot-spot residues, including Q112CD94 and E161HLA-E. Moreover, Nsp13232-240 breaks the salt bridges formed by K217NKG2A and K199NKG2A with HLA-E. Conformational changes induced by Nsp13232-240 lead to diminished atomic contacts and an unstable binding pattern. These findings provide novel insights into the immunomodulatory role of Nsp13232-240 and may inform future NK cell-mediated strategies targeting SARS-CoV-2.

Bone is a major site of metastasis in nasopharyngeal carcinoma (NPC). Recently, nuclear factor kappa-beta ligand (RANKL) inhibitors have garnered attention for their ability to inhibit osteoclast formation and bone resorption, as well as their potential to modulate immune functions and thereby enhance the efficacy of programmed cell death protein 1 (PD-1) inhibitor therapy.

We present a case of a patient with NPC who developed sternal stalk metastasis and multiple bone metastases with soft tissue invasion following radical chemoradiotherapy and targeted therapy. Prior to chemotherapy, the patient experienced severe bone marrow suppression and opted out of further chemotherapy sessions. However, the patient received combination therapy, including RANKL inhibitors (denosumab) alongside PD-1, radiotherapy, and granulocyte-macrophage colony-stimulating factor (PRaG) therapy (NCT05435768), and achieved 16 months of progression-free survival and more than 35 months of overall survival, without encountering any grade 2 or higher treatment-related adverse events.

Denosumab combined with PRaG therapy could be a new therapeutic approach for the second-line treatment in patients with bone metastases.

As important innate immune cells, natural killer (NK) cells play an essential role in resisting pathogen invasion and eliminating transformed cells. However, the hypoxic microenvironment caused by disease conditions is an important physicochemical factor that impairs NK cell function. With the increasing prominence of NK cells in immunotherapy, there has been a surge of interest in developing biological means through which NK cells may overcome the inhibition caused by hypoxia in disease conditions. Although the effects of hypoxic conditions in shaping the functions of NK cells have been increasingly recognized and investigated, reviews have been scantly. A comprehensive understanding of how NK cells adapt to hypoxia can provide valuable insights into how the functional capacity of NK cells may be restored. This review focuses on the functional alterations of NK cells in response to hypoxia. It delineates the mechanisms by which NK cells adapt to hypoxia at the transcriptional, metabolic, translational levels. Furthermore, given the complexity of the hypoxic microenvironment, we also elucidated the effects of key hypoxic metabolites on NK cells. Finally, this review discusses the current clinical therapies derived from targeting hypoxic NK cells. The study of NK cell adaptation to hypoxia has yielded new insights into immunotherapy. These insights may lead to development of novel strategies to improve the treatment of infectious diseases and cancer.

Pancreatic and hepatobiliary cancers are increasing in prevalence and contribute significantly to cancer-related mortality worldwide. Emerging therapeutic approaches, particularly immunotherapy, are gaining attention for their potential to harness the patient's immune system to combat these tumors. Understanding the role of immune cells in the tumor microenvironment (TME) and their metabolic reprogramming is key to developing more effective treatment strategies. This review aims to explore the relationship between immune cell function and glucose metabolism in the TME of pancreatic and hepatobiliary cancers.

This review synthesizes current research on the metabolic adaptations of immune cells, specifically focusing on glucose metabolism within the TME of pancreatic and hepatobiliary cancers. We examine the mechanisms by which immune cells influence tumor progression through metabolic reprogramming and how these interactions can be targeted for therapeutic purposes.

Immune cells in the TME undergo significant metabolic changes, with glucose metabolism playing a central role in modulating immune responses. These metabolic shifts not only affect immune cell function but also influence tumor behavior and progression. The unique metabolic features of immune cells in pancreatic and hepatobiliary cancers provide new opportunities for targeting immune responses to combat these malignancies more effectively.

Understanding the complex relationship between immune cell glucose metabolism and tumor progression in the TME of pancreatic and hepatobiliary cancers offers promising therapeutic strategies. By modulating immune responses through targeted metabolic interventions, it may be possible to improve the efficacy of immunotherapies and better combat these aggressive cancers.

Patients with microsatellite instability-high (MSI-H) colorectal cancer (CRC) have high tumor mutation burden and tumor immunogenicity, exhibiting a higher response rate to immunotherapy and better survival. However, a portion of MSI-H CRC patients still experience adverse disease outcomes. We aimed to identify the tumor-autonomous regulators determining these heterogeneous clinical outcomes. The Cancer Genome Atlas (TCGA) dataset was used to identify regulators in MSI-H CRC patients with unfavorable outcomes. Stable CRC tumor clones expressing targeted regulators were established to evaluate migratory and stemness properties, immune cell vulnerability, and cell-in-cell (CIC) structure formation. RNA-sequencing (RNA-seq) was used to identify enriched biological pathways in stable CRC tumor clones. Clinicopathological characterization of formalin-fixed paraffin-embedded (FFPE) MSI-H CRC specimens was performed to explore the underlying mechanisms involved. We showed that cancer/testis antigen family 45 member A1 (CT45A1) expression was upregulated in MSI-H CRC patients with poor survival outcomes. CT45A1-expressing microsatellite stable (MSS) CRC cells showed enhanced migratory ability. However, CT45A1-expressing MSI-H CRC cells, but not MSS CRC cells, showed higher resistance to natural killer (NK) cell cytotoxicity and served as outer cells in homotypic CIC structures, preventing exogenous or therapeutic antibody access to inner CRC cells. Inactivating RHO-ROCK/MLCK-MLC2 signaling with small-molecule inhibitors or short-hairpin RNAs (shRNAs) targeting myosin light chain kinase (MYLK) abolished NK cell resistance and reduced the outer cell fate of CT45A1-expressing MSI-H CRC cells. In MSI-H CRC patients, CT45A1-positive tumors exhibited increased MLC2 phosphorylation, increased outer cell fate, and decreased survival. We demonstrated that CT45A1 potentiates the advanced progression of MSI-H CRC, and targeting MLC2 phosphorylation may enhance immunotherapy efficacy in CT45A1-positive MSI-H CRC patients.

Diethylhexyl phthalate (DEHP) is the most abundant phthalate compound in the environment, and has been linked with multiple human diseases. The immune system is closely associated with the occurrence and progression of various diseases. However, minimal research has addressed the impact of DEHP on the immune system. In this study, single-cell RNA sequencing was performed using spleen tissue of mice to comprehensively determine alterations of the immune system in response to DEHP. The results showed that DEHP exposure reduced the absolute number of peripheral white blood cells (WBCs), including lymphocytes, monocytes, eosinophils, basophils, and neutrophils in mice. In addition, scRNA-seq analyses showed that inflammatory signaling and the expression of heat shock proteins (HSPs) were reduced in all peripheral immune cell populations. Furthermore, we established a mice cecal ligation and puncture (CLP) model, and showed that DEHP exacerbated sepsis-induced immunosuppression and organ damage. These results suggest that DEHP is an environmental immune disruptor that undermines the immune system, exacerbating acute infections and organ damage. Our findings offer a novel perspective on the hazards of DEHP to human health.

Natural Killer (NK) cells are integral to the innate immune system, renowned for their ability to target and eliminate cancer cells without the need for antigen presentation, sparing normal tissues. These cells are crucial in cancer immunosurveillance due to their diverse array of activating and inhibitory receptors that modulate their cytotoxic activity. However, the tumor microenvironment can suppress NK cell function through various mechanisms. Over recent decades, research has focused on overcoming these tumor escape mechanisms. Initially, efforts concentrated on enhancing T cell activity, leading to impressive results with immunotherapeutic approaches aimed at boosting T cell responses. Nevertheless, a substantial number of patients do not benefit from these treatments and continue to seek effective alternatives. In this context, NK cells present a promising avenue for developing new treatments, given their potent cytotoxic capabilities, safety profile, and activity against T cell-resistant tumors, such as those lacking HLA-I expression. Recent advancements in immunotherapy include strategies to restore and amplify NK cell activity through immune checkpoint inhibitors, cytokines, adoptive NK cell therapy, and CAR-NK cell technology. This review provides a comprehensive overview of NK cell receptors, the tumor escape mechanisms that hinder NK cell function, and the evolving field of NK cell-based cancer immunotherapy, highlighting ongoing efforts to develop more effective and targeted cancer treatment strategies.

Immune rejection caused by mismatches in human leucocyte antigens (HLAs) remains a major obstacle to the success of allogeneic cell therapies. Current strategies for the generation of 'universal' immune-compatible cells, particularly the editing of HLA class I (HLA-I) genes or the modulation of proteins that inhibit natural killer cells, often result in genomic instability or cellular cytotoxicity. Here we show that a β2-microglobulin super-enhancer (B2M-SE) that is responsive to interferon-γ is a critical regulator of the expression of HLA-I on mesenchymal stromal cells (MSCs). Targeted epigenetic repression of B2M-SE in MSCs reduced the surface expression of HLA-I below the threshold required to activate allogenic T cells while maintaining levels sufficient to evade cytotoxicity mediated by natural killer cells. In a humanized mouse model, the epigenetically edited MSCs demonstrated improved survival by evading the immune system, allowing them to exert enhanced therapeutic effects on LPS-induced acute lung injury. Targeted epigenetic repression of B2M-SE may facilitate the development of off-the-shelf cell sources for allogeneic cell therapy.

Cellular phenotype and function are altered in different microenvironments. For targeted therapies it is important to understand site-specific cellular adaptations. Juvenile idiopathic arthritis (JIA) is characterized by autoimmune joint inflammation, with frequent inadequate treatment responses. To comprehensively assess the inflammatory immune landscape, we designed a 37-parameter spectral flow cytometry panel delineating mononuclear cells from JIA synovial fluid (SF) of autoimmune inflamed joints, compared to JIA and healthy control blood. Synovial monocytes and NK cells (CD56bright) lack Fc-receptor CD16, suggesting antibody-mediated targeting may be ineffective. B cells and DCs, both in small frequencies in SF, undergo maturation with high 4-1BB, CD71, CD39 expression, supporting T-cell activation. SF effector and regulatory T cells were highly active with newly described co-receptor combinations that may alter function, and suggestion of metabolic reprogramming via CD71, TNFR2, and PD-1. Most SF effector phenotypes, as well as an identified CD4-Foxp3+ T-cell population, were restricted to the inflamed joint, yet specific SF-predominant CD4+ Foxp3+ Treg subpopulations were increased in blood of active but not inactive JIA, suggesting possible recirculation and loss of immunoregulation at distal sites. This first comprehensive dataset of the site-specific inflammatory landscape at protein level will inform functional studies and the development of targeted therapeutics to restore immunoregulatory balance and achieve remission in JIA.

Cytotoxic CD8+ T lymphocyte (CTL) recognition of non-mutated tumor-associated antigens (TAA), present on cancer cells and also in healthy tissues, is an important element of cancer immunity, but the mechanism of its selectivity for cancer cells and opportunities for its enhancement remain elusive. In this study, we found that CTL expression of the NK receptors (NKR) DNAM1 and NKG2D was associated with the effector status of CD8+ tumor-infiltrating lymphocytes and long-term survival of patients with melanoma. Using MART1 and NY-ESO-1 as model TAAs, we demonstrated that DNAM1 and NKG2D regulate T-cell receptor (TCR) functional avidity and set the threshold for TCR activation of human TAA-specific CTLs. Superior co-stimulatory effects of DNAM1 over CD28 involved enhanced TCR signaling, CTL killer function, and polyfunctionality. Double transduction of human CTLs with TAA-specific TCR and NKRs resulted in strongly enhanced antigen sensitivity, without a reduction in antigen specificity and selectivity of killer function. In addition, the elevation of NKR ligand expression on cancer cells due to chemotherapy also increased CTL recognition of cancer cells expressing low levels of TAAs. Our data help explain the ability of self-antigens to mediate tumor rejection in the absence of autoimmunity and support the development of dual-targeting adoptive T-cell therapies that use NKRs to enhance the potency and selectivity of recognition of TAA-expressing cancer cells.

Cannabidiol (CBD), the major non-psychoactive component of cannabis, exhibits anti-inflammatory properties, but less is known about the immunomodulatory potential of CBD on activated natural killer (NK) cells and/or their targets. Many tumor cells present heat shock protein 70 (Hsp70) on their cell surface in a tumor-specific manner and although a membrane Hsp70 (mHsp70) positive phenotype serves as a target for Hsp70-activated NK cells, a high mHsp70 expression is associated with tumor aggressiveness. This study investigated the immuno-modulatory potential of CBD on NK cells stimulated with TKD Hsp70 peptide and IL-2 (TKD+IL-2) and also on HCT116 p53wt and HCT116 p53-/- colorectal cancer cells exhibiting high and low basal levels of mHsp70 expression.

Apart from an increase in the density of NTB-A and a reduced expression of LAMP-1, the expression of all other activatory NK cell receptors including NKp30, NKG2D and CD69 which are significantly up-regulated after stimulation with TKD+IL-2 remained unaffected after a co-treatment with CBD. However, the release of major pro-inflammatory cytokines by NK cells such as interferon-γ (IFN-γ) and the effector molecule granzyme B (GrzB) was significantly reduced upon CBD treatment. With respect to the tumor target cells, CBD significantly reduced the elevated expression of mHsp70 but had no effect on the low basal mHsp70 expression. Expression of other NK cell ligands such as MICA and MICB remained unaffected, and the NK cell ligands ULBP and B7-H6 were not expressed on these target cells. Consistent with the reduced mHsp70 expression, treatment of both effector and target cells with CBD reduced the killing of high mHsp70 expressing tumor cells by TKD+IL-2+CBD pre-treated NK cells but had no effect on the killing of low mHsp70 expressing tumor cells. Concomitantly, CBD treatment reduced the TKD+IL-2 induced increased release of IFN-γ, IL-4, TNF-α and GrzB, but CBD had no effect on the release of IFN-α when NK cells were co-incubated with tumor target cells.

Cannabidiol (CBD) may potentially diminish the anti-tumor effectiveness of TKD+IL-2 activated natural killer (NK) cells.

Diverse cellular insults converge on activation of the heat shock factor 1 (HSF1), which regulates the proteotoxic stress response to maintain protein homoeostasis. HSF1 regulates numerous gene programmes beyond the proteotoxic stress response in a cell-type- and context-specific manner to promote malignancy. However, the role(s) of HSF1 in immune populations of the tumour microenvironment remain elusive. Here, we leverage an in vivo model of HSF1 activation and single-cell transcriptomic tumour profiling to show that augmented HSF1 activity in natural killer (NK) cells impairs cytotoxicity, cytokine production and subsequent anti-tumour immunity. Mechanistically, HSF1 directly binds and regulates the expression of key mediators of NK cell effector function. This work demonstrates that HSF1 regulates the immune response under the stress conditions of the tumour microenvironment. These findings have important implications for enhancing the efficacy of adoptive NK cell therapies and for designing combinatorial strategies including modulators of NK cell-mediated tumour killing.

Trastuzumab has improved survival rates in human epidermal growth factor receptor 2 (HER2)-positive breast cancer (BC), but drug resistance leads to treatment failure. Natural killer (NK) cell-mediated antibody-dependent cell cytotoxicity (ADCC) represents an essential antitumor immune mechanism of trastuzumab. Traditional Chinese medicine (TCM) has been used for centuries to treat diseases because of its capacity to improve immune responses. Xianling Lianxia formula (XLLXF), based on the principle of "strengthening body and eliminating toxin", exhibits a synergistic effect in the trastuzumab treatment of patients with HER2-positive BC. Notably, this synergistic effect of XLLXF was executed by enhancing NK cells and ADCC, as demonstrated through in vitro co-culture of NK cells and BC cells and in vivo intervention experiments. Mechanistically, the augmented impact of XLLXF on NK cells is linked to a decrease in cytokine inducible Src homology 2 (SH2) containing protein (CISH) expression, which in turn activates the Janus kinase 1 (JAK1)/signal transducer and activator of transcription 5 (STAT5) pathway. Collectively, these findings suggested that XLLXF holds promise for enhancing NK cell function and sensitizing patients with HER2-positive BC to trastuzumab.

The promising results obtained with immunotherapeutic approaches for multiple myeloma (MM) call for a better stratification of patients based on immune components. The most pressing being cytotoxic lymphocytes such as natural killer (NK) cells that are mandatory for MM surveillance and therapy. Here, we performed a single-cell RNA sequencing analysis of NK cells from 10 patients with MM and 10 age/sex-matched healthy donors that revealed important transcriptomic changes in the NK cell landscape affecting both the bone marrow (BM) and peripheral blood compartment. The frequency of mature cytotoxic CD56dim NK cell subsets was reduced in patients with MM at the advantage of late-stage NK cell subsets expressing NF-κB and interferon-I inflammatory signatures. These NK cell subsets accumulating in patients with MM were characterized by low CD16 and CD226 expression and poor cytotoxic functions. MM CD16/CD226Lo NK cells also had adhesion defects with reduced lymphocyte function-associated antigen 1 (LFA-1) integrin activation and actin polymerization that may account for their limited effector functions in vitro. Finally, analysis of BM-infiltrating NK cells in a retrospective cohort of 177 patients with MM from the Intergroupe Francophone du Myélome (IFM) 2009 trial demonstrated that a high frequency of NK cells and their low CD16 and CD226 expression were associated with a shorter overall survival. Thus, CD16/CD226Lo NK cells with reduced effector functions accumulate along MM development and negatively affect patients' clinical outcomes. Given the growing interest in harnessing NK cells to treat myeloma, this improved knowledge around MM-associated NK cell dysfunction will stimulate the development of more efficient immunotherapeutic drugs against MM.

Ribosomal RNA Processing 8 (RRP8) is a gene associated with RNA modification and has been implicated in the development of several types of tumors in recent research. Nevertheless, the biological importance of RRP8 in pan-cancer has not yet been thoroughly and comprehensively investigated.

In this study, we conducted an analysis of various public databases to investigate the biological functions of RRP8. Our analysis included examining its correlation with pan-cancer prognosis, heterogeneity, stemness, immune checkpoint genes, and immune cell infiltration. Furthermore, we utilized the GDSC and CTRP databases to assess the sensitivity of RRP8 to small molecule drugs.

Our findings indicate that RRP8 exhibits differential expression between tumor and normal samples, particularly impacting the prognosis of various cancers such as Adrenocortical carcinoma (ACC) and Kidney Chromophobe (KICH). The expression of RRP8 is intricately linked to tumor heterogeneity and stemness markers. Additionally, RRP8 shows a positive correlation with the presence of tumor-infiltrating cells, with TP53 being the predominant mutated gene in these malignancies.

Our findings suggest that RRP8 may serve as a potential prognostic marker and therapeutic target in a variety of cancer types.

Hexokinases (HKs) control the first step of glucose catabolism. A switch of expression from liver HK (glucokinase, GCK) to the tumor isoenzyme HK2 is observed in hepatocellular carcinoma progression. Our prior work revealed that HK isoenzyme switch in hepatocytes not only regulates hepatic metabolic functions but also modulates innate immunity and sensitivity to Natural Killer (NK) cell cytotoxicity. This study investigates the impact of HK2 expression and its mitochondrial binding on the resistance of human liver cancer cells to NK-cell-induced cytolysis. We have shown that HK2 expression induces resistance to NK cell cytotoxicity in a process requiring mitochondrial binding of HK2. Neither HK2 nor GCK expression affects target cells' ability to activate NK cells. In contrast, mitochondrial binding of HK2 reduces effector caspase 3/7 activity both at baseline and upon NK-cell activation. Furthermore, HK2 tethering to mitochondria enhances their resistance to cytochrome c release triggered by tBID. These findings indicate that HK2 mitochondrial binding in liver cancer cells is an intrinsic resistance factor to cytolysis and an escape mechanism from immune surveillance.

Immunotherapy has been a research hotspot due to its low side effects, long-lasting efficacy, and wide anti-tumor spectrum. Recently, NK cell-based immunotherapy has gained broad attention for its unique immunological character of tumor identification and eradication and low risk of graft-versus-host disease and cytokine storm. With the cooperation of a drug delivery system (DDS), NK cells activate tumoricidal activity by adjusting the balance of the activating and inhibitory signals on their surface after drug-loaded DDS administration. Moreover, NK cells or NK-derived exosomes can also be applied as drug carriers for distinct modification to promote NK activation and exert anti-tumor effects. In this review, we first introduce the source and classification of NK cells and describe the common activating and inhibitory receptors on their surface. Then, we summarize the strategies for activating NK cells in vivo through various DDSs. Finally, the application prospects of NK cells in tumor immunotherapy are also discussed.

Natural killer (NK) cells have gained considerable attention and hold great potential for their application in tumor immunotherapy. This is mainly due to their MHC-unrestricted and pan-specific recognition capabilities, as well as their ability to rapidly respond to and eliminate target cells. To artificially generate therapeutic NK cells, various materials can be utilized, such as peripheral blood mononuclear cells (PBMCs), umbilical cord blood (UCB), induced pluripotent stem cells (iPSCs), and NK cell lines. Exploiting the therapeutic potential of NK cells to treat tumors through in vivo and in vitro therapeutic modalities has yielded positive therapeutic results.

This review provides a comprehensive description of NK cell therapeutic approaches for tumors and discusses the current problems associated with these therapeutic approaches and the prospects of NK cell therapy for tumors.

Although current stem cell therapies exhibit promising potential, the extended process of employing autologous cells and the necessity for donor-host matching to avert the rejection of transplanted cells significantly limit the widespread applicability of these treatments. It would be highly advantageous to generate a pluripotent universal donor stem cell line that is immune-evasive and, therefore, not restricted by the individual's immune system, enabling unlimited application within cell replacement therapies. Before such immune-evasive stem cells can be moved forward to clinical trials, in vivo testing via transplantation experiments in immune-competent animals would be a favorable approach preceding preclinical testing. By using human stem cells in immune competent animals, results will be more translatable to a clinical setting, as no parts of the immune system have been altered, although in a xenogeneic setting. In this way, immune evasiveness, cell survival, and unwanted proliferative effects can be assessed before clinical trials in humans. The current study presents the generation and characterization of three human embryonic stem cell lines (hESCs) for xenogeneic transplantation in immune-competent mice. The major histocompatibility complexes I- and II-encoding genes, B2M and CIITA, have been deleted from the hESCs using CRISPR-Cas9-targeted gene replacement strategies and knockout. B2M was knocked out by the insertion of murine CD47. Human-secreted embryonic alkaline phosphatase (hSEAP) was inserted in a safe harbor site to track cells in vivo. The edited hESCs maintained their pluripotency, karyotypic normality, and stable expression of murine CD47 and hSEAP in vitro. In vivo transplantation of hESCs into immune-competent BALB/c mice was successfully monitored by measuring hSEAP in blood samples. Nevertheless, transplantation of immune-evasive hESCs resulted in complete rejection within 11 days, with clear immune infiltration of T-cells on day 8. Our results reveal that knockout of B2M and CIITA together with species-specific expression of CD47 are insufficient to prevent rejection in an immune-competent and xenogeneic context.

Type 2 innate lymphoid cells (ILC2s) play a pivotal role in type 2 asthma. CD226 is a costimulatory molecule involved in various inflammatory diseases.

We aimed to investigate CD226 expression and function within human and mouse ILC2s, and to assess the impact of targeting CD226 on ILC2-mediated airway hyperreactivity (AHR).

We administered IL-33 intranasally to wild-type mice, followed by treatment with anti-CD226 antibody or isotype control. Pulmonary ILC2s were sorted for ex vivo analyses through RNA sequencing and flow cytometry. Next, we evaluated the effects of CD226 on AHR and lung inflammation in wild-type and Rag2-/- mice. Additionally, we compared peripheral ILC2s from healthy donors and asthmatic patients to ascertain the role of CD226 in human ILC2s.

Our findings demonstrated an inducible expression of CD226 in activated ILC2s, enhancing their cytokine secretion and effector functions. Mechanistically, CD226 alters intracellular metabolism and enhances PI3K/AKT and MAPK signal pathways. Blocking CD226 ameliorates ILC2-dependent AHR in IL-33 and Alternaria alternata-induced models. Interestingly, CD226 is expressed and inducible in human ILC2s, and its blocking reduces cytokine production. Finally, we showed that peripheral ILC2s in asthmatic patients exhibited elevated CD226 expression compared to healthy controls.

Our findings underscore the potential of CD226 as a novel therapeutic target in ILC2s, presenting a promising avenue for ameliorating AHR and allergic asthma.

CD155 is an immunoglobulin-like protein overexpressed in almost all the tumor cells, which not only promotes proliferation, adhesion, invasion, and migration of tumor cells, but also regulates immune responses by interacting with TIGIT, CD226 or CD96 receptors expressed on several immune cells, thereby modulating the functionality of these cellular subsets. As a novel immune checkpoint, the inhibition of CD155/TIGIT, either as a standalone treatment or in conjunction with other immune checkpoint inhibitors, has demonstrated efficacy in managing advanced solid malignancies. In this review, we summarize the intricate relationship between on tumor surface CD155 and its receptors, with further discussion on how they regulate the occurrence of tumor immune escape. In addition, novel therapeutic strategies and clinical trials targeting CD155 and its receptors are summarized, providing a strong rationale and way forward for the development of next-generation immunotherapies.

The etiology of various neurodegenerative disorders that mainly affect the central nervous system including (but not limited to) Alzheimer's disease, Parkinson's disease and Huntington's disease has classically been attributed to neuronal defects that culminate with the loss of specific neuronal populations. However, accumulating evidence suggests that numerous immune effector cells and the products thereof (including cytokines and other soluble mediators) have a major impact on the pathogenesis and/or severity of these and other neurodegenerative syndromes. These observations not only add to our understanding of neurodegenerative conditions but also imply that (at least in some cases) therapeutic strategies targeting immune cells or their products may mediate clinically relevant neuroprotective effects. Here, we critically discuss immunological mechanisms of central neurodegeneration and propose potential strategies to correct neurodegeneration-associated immunological dysfunction with therapeutic purposes.

Poliovirus receptor-related immunoglobulin domain-containing protein, or PVRIG, is a newly discovered immune checkpoint that has emerged as a promising target for cancer immunotherapy. It is primarily expressed on activated T and natural killer (NK) cells, and once engaged with its ligand, PVRL2, it induces inhibitory signaling in T cells, thereby promoting the functional exhaustion of tumor-infiltrating lymphocytes (TILs). Here, we characterized IBI352g4a, a novel humanized anti-PVRIG antibody with Fc-competent function, explored the mechanism of its antitumor activity in preclinical models, and systemically evaluated the contribution of FcrR engagement to PVRIG blockade-induced antitumor activity. IBI352g4a binds to the extracellular domain of human PVRIG with high affinity (Kd = 0.53 nM) and specificity, and fully blocks the interaction between PVRIG and its ligand PVRL2. Unlike other immune checkpoints, IBI352g4a significantly induced NK cell activation and degranulation, but had a minimal effect on T-cell activation in in vitro functional assays. IBI352g4a induced strong antitumor effect in several preclinic models, through in vivo mechanism analysis we found that both NK and T cells contribute to the antitumor effect, but NK cells play predominant roles. Specifically, a single dose of IBI352g4a induced significant NK cell activation in TILs, but T-cell activation was observed only after the second dose. Moreover, the Fc effector function is critical for both NK cell activation and treatment efficacy in vitro and in vivo. Our study, for the first time, demonstrates that both NK activation and FcrR engagement are required for antitumor efficacy induced by PVRIG blockade.

Non-small-cell lung carcinoma (NSCLC) is the most common lung cancer and one of the pioneer tumors in which immunotherapy has radically changed patients' outcomes. However, several issues are emerging and their implementation is required to optimize immunotherapy-based protocols. In this work, we investigate the ability of the Bromodomain and Extra-Terminal protein inhibitors (BETi) to stimulate a proficient anti-tumor immune response toward NSCLC. By using in vitro, ex-vivo, and in vivo models, we demonstrate that these epigenetic drugs specifically enhance Natural Killer (NK) cell cytotoxicity. BETi down-regulate a large set of NK inhibitory receptors, including several immune checkpoints (ICs), that are direct targets of the transcriptional cooperation between the BET protein BRD4 and the transcription factor SMAD3. Overall, BETi orchestrate an epigenetic reprogramming that leads to increased recognition of tumor cells and the killing ability of NK cells. Our results unveil the opportunity to exploit and repurpose these drugs in combination with immunotherapy.

Natural killer (NK) cells are unique from other immune cells in that they can rapidly kill multiple neighboring cells without the need for antigenic pre-sensitization once the cells display surface markers associated with oncogenic transformation. Given the dynamic role of NK cells in tumor surveillance, NK cell-based immunotherapy is rapidly becoming a "new force" in tumor immunotherapy. However, challenges remain in the use of NK cell immunotherapy in the treatment of solid tumors. Many metabolic features of the tumor microenvironment (TME) of solid tumors, including oxygen and nutrient (e.g., glucose, amino acids) deprivation, accumulation of specific metabolites (e.g., lactate, adenosine), and limited availability of signaling molecules that allow for metabolic reorganization, multifactorial shaping of the immune-suppressing TME impairs tumor-infiltrating NK cell function. This becomes a key barrier limiting the success of NK cell immunotherapy in solid tumors. Restoration of endogenous NK cells in the TME or overt transfer of functionally improved NK cells holds great promise in cancer therapy. In this paper, we summarize the metabolic biology of NK cells, discuss the effects of TME on NK cell metabolism and effector functions, and review emerging strategies for targeting metabolism-improved NK cell immunotherapy in the TME to circumvent these barriers to achieve superior efficacy of NK cell immunotherapy.

Immune checkpoints (ICs) play a pivotal role in orchestrating immune regulation, crucial for the maintenance of immune tolerance and prevention of autoimmune diseases. One noteworthy example among these immune regulators is T cell immunoglobulin (Ig) and immunoreceptor tyrosine-based inhibitory motif (ITIM) domain (TIGIT). The TIGIT pathway's inhibition or the absence of TIGIT has been linked to the hyperactivation and excessive proliferation of T cells, rendering individuals more susceptible to autoimmune diseases and exacerbating inflammatory responses. Conversely, the activation of TIGIT has exhibited promising outcomes in ameliorating autoimmune disorders, as observed in murine models of systemic lupus erythematosus (SLE). Consequently, a judicious exploration of the co-inhibitory axis appears warranted for the effective management of pathogenic immune responses in SLE. In light of compelling evidence, this review undertakes a comprehensive examination of TIGIT's characteristics within the context of autoimmunity, offering insights into its potential as a therapeutic target for SLE.

Immunosuppression increases the risk of nosocomial infection in patients with chronic critical illness. This exploratory study aimed to determine the immunometabolic signature associated with nosocomial infection during chronic critical illness. We prospectively recruited patients who were admitted to the respiratory care center and who had received mechanical ventilator support for more than 10 days in the intensive care unit. The study subjects were followed for the occurrence of nosocomial infection until 6 weeks after admission, hospital discharge, or death. The cytokine levels in the plasma samples were measured. Single-cell immunometabolic regulome profiling by mass cytometry, which analyzed 16 metabolic regulators in 21 immune subsets, was performed to identify immunometabolic features associated with the risk of nosocomial infection. During the study period, 37 patients were enrolled, and 16 patients (43.2%) developed nosocomial infection. Unsupervised immunologic clustering using multidimensional scaling and logistic regression analyses revealed that expression of nuclear respiratory factor 1 (NRF1) and carnitine palmitoyltransferase 1a (CPT1a), key regulators of mitochondrial biogenesis and fatty acid transport, respectively, in natural killer (NK) cells was significantly associated with nosocomial infection. Downregulated NRF1 and upregulated CPT1a were found in all subsets of NK cells from patients who developed a nosocomial infection. The risk of nosocomial infection is significantly correlated with the predictive score developed by selecting NK cell-specific features using an elastic net algorithm. Findings were further examined in an independent cohort of COVID-19-infected patients, and the results confirm that COVID-19-related mortality is significantly associated with mitochondria biogenesis and fatty acid oxidation pathways in NK cells. In conclusion, this study uncovers that NK cell-specific immunometabolic features are significantly associated with the occurrence and fatal outcomes of infection in critically ill population, and provides mechanistic insights into NK cell-specific immunity against microbial invasion in critical illness.

RNA viral infections seriously endanger human health. Src homology 2 (SH2) domain-containing protein tyrosine phosphatase 2 (SHP2) suppresses innate immunity against influenza A virus, and pharmacological inhibition of SHP2 provokes hepatic innate immunity. SHP2 binds and catalyzes tyrosyl dephosphorylation of protein zero-related (PZR), but the regulatory effect of PZR on innate immune response to viral infection is unclear. In this study, the transcription and protein level of PZR in host cells were found to be decreased with RNA viral infection, and high level of PZR was uncovered to inhibit interferon (IFN) signaling mediated by RIG-I and MDA5. Through localizing in mitochondria, PZR targeted and interacted with MAVS (also known as IPS-1/VISA/Cardif), suppressing the aggregation and activation of MAVS. Specifically, Y263 residue in ITIM is critical for PZR to exert immunosuppression under RNA viral infection. Moreover, the recruited SHP2 by PZR that modified with tyrosine phosphorylation under RNA viral infection might inhibit phosphorylation activation of MAVS. In conclusion, PZR and SHP2 suppress innate immune response to RNA viral infection through inhibiting MAVS activation. This study reveals the regulatory mechanism of PZR-SHP2-MAVS signal axis on IFN signaling mediated by RIG-I and MDA5, which may provide new sight for developing antiviral drugs.

Secreted proteins play critical roles in regulating immune responses, exerting cytotoxic effects on tumor cells, promoting inflammatory processes, and influencing cellular metabolism. Deciphering the intricate relationship between the heterogeneity of secreted proteins and their transcriptional states is pivotal in the study of cellular heterogeneity. Here we proposed a cell-antibody conjugate-based sequencing methodology (Cellab-seq) for joint characterization of secreted proteins and transcriptome. Cellab-seq utilizes a chemoenzymatic strategy to construct cell-antibody conjugates, which enables the capture of secreted proteins and their signal transduction with the incorporation of barcode detection antibodies. We applied Cellab-seq to investigate how gene expression influences the activity of secreted proteins in NK cells. Altogether, this strategy facilitates a nuanced understanding of cellular dynamics under diverse physiological conditions, ultimately contributing to the prevention, diagnosis and treatment of diseases.

The immune responses to cancer cells involve both innate and acquired immune cells. In the meantime, the most attention has been drawn to the adaptive immune cells, especially T cells, while, it is now well known that the innate immune cells, especially natural killer (NK) cells, play a vital role in defending against malignancies. While the immune cells are trying to eliminate malignant cells, cancer cells try to prevent the function of these cells and suppress immune responses. The suppression of NK cells in various cancers can lead to the induction of an exhausted phenotype in NK cells, which will impair their function. Recent studies have shown that the occurrence of this phenotype in various types of leukemic malignancies can affect the prognosis of the disease, and targeting these cells may be considered a new immunotherapy method in the treatment of leukemia. Therefore, a detailed study of exhausted NK cells in leukemic diseases can help both to understand the mechanisms of leukemia progression and to design new treatment methods by creating a deeper understanding of these cells. Here, we will comprehensively review the immunobiology of exhausted NK cells and their role in various leukemic malignancies. Video Abstract.

This study aimed to investigate the role of regulatory T cells in patients with unexplained recurrent pregnancy loss (URPL).

We retrospectively analyzed 136 women who had experienced two or more miscarriages before 24 weeks of gestation for no obvious reason from May 2018 to October 2021. The basic clinical data of the patients and expression of lymphocyte subsets such as regulatory T cells (Tregs) and natural killer cells (NKs) by flow cytometry were collected to explore the risk factors of pregnancy outcome in URPL patients.

A total of 136 URPL patients were enrolled in this study. Eventually, 50 patients attained clinical pregnancy. The median age was 31.8 ± 4.6 years in patients with clinical pregnancy. The univariate and multivariate logistic regression analyses indicated that Tregs was associated with the pregnancy outcomes of patients with URPL (odds ratio 0.63, 95% confidence interval 0.50-0.80). More importantly, a U-shaped association was found between Tregs and pregnancy outcome (p < 0.001), with either higher or lower Tregs levels adversely affecting pregnancy outcome.

Tregs levels that are either too high or too low can harm pregnancy outcomes. It was expected to be a very promising quantitative biomarker for predicting pregnancy outcomes in URPL patients.

We have developed a 31-color panel to define the steady-state phenotype of T cells in human peripheral blood (Table 1). The panel presented here was optimized using cryopreserved peripheral blood mononuclear cells (PBMC). The markers included in this panel were chosen in order to characterize the steady-state phenotype of T cells and includes markers (CD45RA, CD45RO, CCR7, CD95) to distinguish the main subsets (e.g., naïve, TEM , TCM , TEMRA , TSCM etc.) of CD4, CD8, and γδ T cells. This panel also includes markers for the identification of differentiation status (CD27, CD28), activation/antigen experience status (CD11a, CD49d, CD38, HLA-DR, CD56, and CD39), co-inhibitory marker expression (PD-1, TIM-3), and CD4 T helper subsets (CXCR3, CXCR5, CCR4, CCR6, Foxp3, CD25, and CD127). This optimized panel provides a broad assessment of the steady-state phenotype of human T cells.