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Background: Hepatocellular carcinoma (HCC) represents a significant global health challenge with high incidence and mortality rates. T cells and natural killer (NK) cells are pivotal in this context, yet HCC can evade immune surveillance. CD161 (KLRB1), a C-type lectin receptor, modulates immune responses and is expressed on NK cells and a subset of T cells. Its relevance in HCC remains poorly understood, with conflicting findings regarding its impact on patient prognosis. Methods: Utilizing TCGA data and single-cell analysis, we investigated the biological functions of KLRB1 in HCC. Peripheral blood samples from 126 HCC patients were collected to assess KLRB1 expression on NK and T cells. The diagnostic performance of KLRB1 on NK and CD8 + T cells was evaluated using receiver operating characteristic curve (ROC) analysis, while its prognostic significance was assessed using Kaplan-Meier analysis and COX regression models. Results: Analysis of TCGA data revealed a significant correlation between KLRB1 expression and immune activation, particularly T cell activation. Single-cell data further demonstrated elevated KLRB1 expression in tissue-resident NK and T cells within HCC, which co-expressed markers of immune activation. Clinical data showed downregulated KLRB1 expression on NK and T cells in HCC patients compared to health individuals, with lower expression levels correlating with poorer prognosis. Conclusion: KLRB1 emerges as a promising biomarker in HCC, with its downregulation on peripheral blood NK and T cells suggesting potential prognostic value. Further elucidation of KLRB1's role in HCC may pave the way for the development of targeted immunotherapies and the improvement of patient outcomes. [ABSTRACT FROM AUTHOR]

Metastasis causes greater than 90% of cancer-associated deaths, presenting huge challenges for detection and efficient treatment of cancer due to its high heterogeneity and widespread dissemination to various organs. Therefore, it is imperative to combat cancer metastasis, which is the key to achieving complete cancer eradication. Immunotherapy as a systemic approach has shown promising potential to combat metastasis. However, current clinical immunotherapies are not effective for all patients or all types of cancer metastases owing to insufficient immune responses. In recent years, immunological nanomaterials with intrinsic immunogenicity or immunomodulatory agents with efficient loading have been shown to enhance immune responses to eliminate metastasis. In this review, we would like to summarize various types of immunological nanomaterials against metastasis. Moreover, this review will summarize a series of immunological nanomaterial-mediated immunotherapy strategies to combat metastasis, including immunogenic cell death, regulation of chemokines and cytokines, improving the immunosuppressive tumour microenvironment, activation of the STING pathway, enhancing cytotoxic natural killer cell activity, enhancing antigen presentation of dendritic cells, and enhancing chimeric antigen receptor T cell therapy. Furthermore, the synergistic anti-metastasis strategies based on the combinational use of immunotherapy and other therapeutic modalities will also be introduced. In addition, the nanomaterial-mediated imaging techniques (e.g., optical imaging, magnetic resonance imaging, computed tomography, photoacoustic imaging, surface-enhanced Raman scattering, radionuclide imaging, etc.) for detecting metastasis and monitoring anti-metastasis efficacy are also summarized. Finally, the current challenges and future prospects of immunological nanomaterial-based anti-metastasis are also elucidated with the intention to accelerate its clinical translation. [ABSTRACT FROM AUTHOR]

Background: Photodynamic therapy (PDT) is an antitumor therapy and has traditionally been regarded as a localized therapy in itself. However, recent reports have shown that it not only exerts a direct cytotoxic effect on cancer cells but also enhances body's tumor immunity. We hypothesized that the immunological response induced by PDT could potentially enhance the efficacy of programmed death‐1 (PD‐1) / programmed death‐ligand 1 (PD‐L1) blockade. Methods: The cytotoxic effects of PDT on colon 26 cells were investigated in vitro using the WST assay. We investigated whether the antitumor effect of anti‐PD‐1 antibodies could be amplified by the addition of PDT. We performed combination therapy by randomly allocating tumor‐bearing mice to four treatment groups: control, anti‐PD‐1 antibodies, PDT, and a combination of anti‐PD‐1 antibodies and PDT. To analyze the tumor microenvironment after treatment, the tumors were resected and pathologically evaluated. Results: The viability rate of colon 26 cells decreased proportionally with the laser dose. In vivo experiments for combined PDT and anti‐PD‐1 antibody treatment, combination therapy showed an enhanced antitumor effect compared with the control. Immunohistochemical findings of the tumor microenvironment 10 days after PDT indicated that the number of CD8+ cells, the area of Iba‐1+ cells and the area expressing PD‐L1 were significantly higher in tumors treated with combination therapy than in tumors treated with anti‐PD‐1 antibody alone, PDT alone, or the control. Conclusions: PDT increased immune cell infiltration into the tumor microenvironment. The immunological response induced by PDT may enhance the efficacy of PD‐1/PD‐L1 blockade. [ABSTRACT FROM AUTHOR]

Invasive aspergillosis causes significant morbidity and mortality in immunocompromised patients. Natural killer (NK) cells are pivotal for antifungal defense. Thus far, CD56 is the only known pathogen recognition receptor on NK cells triggering potent antifungal activity against Aspergillus fumigatus. However, the underlying cellular mechanisms and the fungal ligand of CD56 have remained unknown. Using purified cell wall components, biochemical treatments, and ger mutants with altered cell wall composition, we herein found that CD56 interacts with the A. fumigatus cell wall carbohydrate galactosaminogalactan (GAG). This interaction induced NK cell activation, degranulation, and secretion of immune-enhancing chemokines and cytotoxic effectors. Supernatants from GAG-stimulated NK cells elicited antifungal activity and enhanced antifungal effector responses of polymorphonuclear cells. In conclusion, we identified A. fumigatus GAG as a ligand of CD56 on human primary NK cells, stimulating potent antifungal effector responses and activating other immune cells. Author summary: The opportunistic mold pathogen Aspergillus (A.) fumigatus possesses significant health risk, particularly for immunocompromised individuals. This ubiquitous fungus found in soil and decaying organic matter, causes a range of partially life-threatening infections known as aspergillosis. Natural killer (NK) cells, part of the innate immunity, recognize A. fumigatus by pathogen-associated molecular patterns (PAMPs) via their pattern recognition receptors (PRRs), leading to the elimination of infected cells, including direct cytotoxicity and cytokine production. Previously, we identified the NK-cell receptor CD56 as a PRR that recognizes A. fumigatus and triggers potent antifungal activity. However, the A. fumigatus ligand of CD56 has remained unknown. Here, we used different approaches to identify the A. fumigatus ligand of CD56 on NK cells. We found that CD56 directly binds to cell wall galactosaminogalactan. Specifically, deacetylated galactosamine residues of GAG played a role in the interaction with CD56 and triggered strong NK-cell activation, along with potent release of cytotoxic effectors and immune-enhancing chemokines. We also found that GAG-primed NK-cell supernatants activate and enhance effector responses of polymorphonuclear cells. Our study provides new insights on the A. fumigatus-NK cell interplay by identifying GAG as an activating ligand of CD56 on human NK cells, stimulating potent antifungal effector responses. [ABSTRACT FROM AUTHOR]

Background: p53, the most frequently mutated gene in cancer, lacks effective targeted drugs. Methods: We developed monoclonal antibodies (mAbs) that target a p53 hotspot mutation E285K without cross-reactivity with wild-type p53. They were delivered using lipid nanoparticles (LNPs) that encapsulate DNA plasmids. Western blot, BLI, flow cytometry, single-cell sequencing (scRNA-seq), and other methods were employed to assess the function of mAbs in vitro and in vivo. Results: These LNP-pE285K-mAbs in the IgG1 format exhibited a robust anti-tumor effect, facilitating the infiltration of immune cells, including CD8+ T, B, and NK cells. scRNA-seq revealed that IgG1 reduces immune inhibitory signaling, increases MHC signaling from B cells to CD8+ T cells, and enriches anti-tumor T cell and B cell receptor profiles. The E285K-mAbs were also produced in the dimeric IgA (dIgA) format, whose anti-tumor activity depended on the polymeric immunoglobulin receptor (PIGR), a membrane Ig receptor, whereas that of IgG1 relied on TRIM21, an intracellular IgG receptor. Conclusions: Targeting specific mutant epitopes using DNA-encoded and LNP-delivered mAbs represents a potential precision medicine strategy against p53 mutants in TRIM21- or PIGR-positive cancers. [ABSTRACT FROM AUTHOR]

Background JC polyomavirus (JCPyV) causes progressive multifocal leukoencephalopathy (PML), a potentially fatal complication of severe immune suppression with no effective treatment. Natural killer (NK) cells play critical roles in defense against viral infections; however, NK-cell response to JCPyV infection remains unexplored. Methods NK- and T-cell responses against the JCPyV VP1 were compared using intracellular cytokine staining upon stimulation with peptide pools. A novel flow cytometry-based assay was developed to determine NK-cell killing efficiency of JCPyV-infected astrocyte-derived SVG-A cells. Blocking antibodies were used to evaluate the contribution of NK-cell receptors in immune recognition of JCPyV-infected cells. Results In about 40% of healthy donors, we detected robust CD107a upregulation and IFN-γ production by NK cells, extending beyond T-cell responses. Next, using the NK-cell–mediated killing assay, we showed that coculture of NK cells and JCPyV-infected SVG-A cells leads to a 60% reduction in infection, on average. JCPyV-infected cells had enhanced expression of ULBP2—a ligand for the activating NK-cell receptor NKG2D, and addition of NKG2D blocking antibodies decreased NK-cell degranulation. Conclusions NKG2D-mediated activation of NK cells plays a key role in controlling JCPyV replication and may be a promising immunotherapeutic target to boost NK-cell anti-JCPyV activity. [ABSTRACT FROM AUTHOR]

STING agonists can reprogram the tumor microenvironment to induce immunological clearance within the central nervous system. Using multiplexed sequential immunofluorescence (SeqIF) and the Ivy Glioblastoma Atlas, STING expression was found in myeloid populations and in the perivascular space. The STING agonist 8803 increased median survival in multiple preclinical models of glioblastoma, including QPP8, an immune checkpoint blockade–resistant model, where 100% of mice were cured. Ex vivo flow cytometry profiling during the therapeutic window demonstrated increases in myeloid tumor trafficking and activation, alongside enhancement of CD8+ T cell and NK effector responses. Treatment with 8803 reprogrammed microglia to express costimulatory CD80/CD86 and iNOS, while decreasing immunosuppressive CD206 and arginase. In humanized mice, where tumor cell STING is epigenetically silenced, 8803 therapeutic activity was maintained, further attesting to myeloid dependency and reprogramming. Although the combination with a STAT3 inhibitor did not further enhance STING agonist activity, the addition of anti–PD-1 antibodies to 8803 treatment enhanced survival in an immune checkpoint blockade–responsive glioma model. In summary, 8803 as a monotherapy demonstrates marked in vivo therapeutic activity, meriting consideration for clinical translation. [ABSTRACT FROM AUTHOR]

Receptor Expression-Enhancing Protein 3 (REEP3) serves as a pivotal enzyme crucial for endoplasmic reticulum (ER) clearance during mitosis and is implicated in the advancement of diverse malignancies. Nonetheless, the biological role and mechanisms of REEP3 in pancreatic cancer patients, along with its interplay with immune infiltration, remain inadequately elucidated. In this study, we initially analyzed the differential expression of REEP3 between pancreatic cancer tissues and normal pancreas tissues using the Cancer Genome Atlas (TCGA), GTEx and Gene Expression Omnibus (GEO) databases. Subsequently, we utilized Kaplan–Meier analysis, Cox regression and ROC curve to determine the predictive value of REEP3 for the clinical outcomes of pancreatic cancer patients. Functional enrichment analyses, including Gene Ontology (GO), Kyoto Encyclopedia of Genes and Genomes (KEGG) and Gene Set Enrichment Analysis (GSEA), were conducted to explore the potential signaling pathways and biological functions associated with pancreatic cancer. Furthermore, we investigated the PPI network, miRNA, RBP and transcription factor interactions of REEP3 using databases such as GeneMania, STRING, StarBase, KnockTK, ENCODE, Jaspar and hTFtarget. Lastly, the "ssGSEA" algorithm and TIMER database were employed to investigate the correlation between REEP3 expression and immune infiltration as well as immune checkpoints. The expression of REEP3 in pancreatic cancer showed a significantly higher level compared to that in normal tissues. ROC curve analysis indicated that REEP3 holds substantial diagnostic potential for pancreatic cancer patients. Elevated REEP3 expression correlated with unfavorable outcomes in terms of both overall survival and relapse-free survival, establishing it as a notable adverse prognostic marker in pancreatic cancer. Moreover, both univariate and multivariate Cox regression analyses demonstrated that REEP3 maintained an independent association with overall survival. Functional enrichment analyses revealed pathways significantly linked to REEP3, including cytoplasmic translation, wound healing, viral processes, regulation of cellular component size and actin filament organization. Additionally, REEP3 expression displayed a significant positive correlation with CD8+ T cells, B cells, natural killer cells, dendritic cells and macrophages. REEP3 is a potential diagnostic, prognostic marker and immunotherapeutic target for pancreatic cancer. [ABSTRACT FROM AUTHOR]

Through Smad3-dependent signalings, transforming growth factor-β (TGF-β) suppresses the development, maturation, cytokine productions and cytolytic functions of NK cells in cancer. Silencing Smad3 remarkably restores the cytotoxicity of NK-92 against cancer in TGF-β-rich microenvironment, but its effects on the immunoregulatory functions of NK cells remain obscure. In this study, we identified Smad3 functioned as a transcriptional repressor for CSF2 (GM-CSF) in NK cells. Therefore, disrupting Smad3 largely mitigated TGF-β-mediated suppression on GM-CSF production by NK cells. Furthermore, silencing GM-CSF in Smad3 knockout NK cells substantially impaired their anti-lung carcinoma effects. In-depth study demonstrated that NK-derived GM-CSF strengthened T cell immune responses by stimulating dendritic cell differentiation and M1 macrophage polarization. Meanwhile, NK-derived GM-CSF promoted the survival of neutrophils, which in turn facilitated the terminal maturation of NK cells, and subsequently boosted NK-cell mediated cytotoxicity against lung carcinoma. Thus, Smad3-silenced NK-92 (NK-92-S3KD) may serve as a promising immunoadjuvant therapy with clinical translational value given its robust cytotoxicity against malignant cells and immunostimulatory functions to reinforce the therapeutic effects of other immunotherapies. [ABSTRACT FROM AUTHOR]

Alzheimer's disease (AD) is a serious brain disorder characterized by the presence of beta-amyloid plaques, tau pathology, inflammation, neurodegeneration, and cerebrovascular dysfunction. The presence of chronic neuroinflammation, breaches in the blood-brain barrier (BBB), and increased levels of inflammatory mediators are central to the pathogenesis of AD. These factors promote the penetration of immune cells into the brain, potentially exacerbating clinical symptoms and neuronal death in AD patients. While microglia, the resident immune cells of the central nervous system (CNS), play a crucial role in AD, recent evidence suggests the infiltration of cerebral vessels and parenchyma by peripheral immune cells, including neutrophils, T lymphocytes, B lymphocytes, NK cells, and monocytes in AD. These cells participate in the regulation of immunity and inflammation, which is expected to play a huge role in future immunotherapy. Given the crucial role of peripheral immune cells in AD, this article seeks to offer a comprehensive overview of their contributions to neuroinflammation in the disease. Understanding the role of these cells in the neuroinflammatory response is vital for developing new diagnostic markers and therapeutic targets to enhance the diagnosis and treatment of AD patients. [ABSTRACT FROM AUTHOR]

Although there is an association between abdominal aortic aneurysm (AAA) and circulating immune cell phenotypes, the exact causal relationship remains unclear. This study aimed to explore the causal relationships between immune cell phenotypes and AAA risk using a bidirectional two-sample Mendelian randomization approach. Data from genome-wide association studies pertaining to 731 immune cell traits and AAA were systematically analyzed. Using strict selection criteria, we identified 339 immune traits that are associated with at least 3 single nucleotide polymorphisms. A comprehensive MR analysis was conducted using several methods including Inverse Variance Weighted, Weighted Median Estimator, MR-Egger regression, Weighted Mode, and Simple Median methods. CD24 on switched memory cells (OR = 0.922, 95% CI 0.914–0.929, P = 2.62e−79) at the median fluorescence intensities level, and SSC-A on HLA-DR + natural killer cells (OR = 0.873, 95% CI 0.861–0.885, P = 8.96e−81) at the morphological parameter level, exhibited the strongest causal associations with AAA. In the reverse analysis, no significant causal effects of AAA on immune traits were found. The study elucidates the causal involvement of multiple circulating immune cell phenotypes in AAA development, signifying their potential as diagnostic markers or therapeutic targets. These identified immune traits may be crucial in modulating AAA-related inflammatory pathways. [ABSTRACT FROM AUTHOR]

Sanfilippo syndrome results from inherited mutations in genes encoding lysosomal enzymes that catabolise heparan sulfate (HS), leading to early childhood‐onset neurodegeneration. This study explores the therapeutic potential of photobiomodulation (PBM), which is neuroprotective and anti‐inflammatory in several neurodegenerative diseases; it is also safe and PBM devices are readily available. We investigated the effects of 10–14 days transcranial PBM at 670 nm (2 or 4 J/cm2/day) or 904 nm (4 J/cm2/day) in young (3 weeks) and older (15 weeks) Sanfilippo or mucopolysaccharidosis type IIIA (MPS IIIA) mice. Although we found no PBM‐induced changes in HS accumulation, astrocyte activation, CD206 (an anti‐inflammatory marker) and BDNF expression in the brains of Sanfilippo mice, there was a near‐normalisation of microglial activation in older MPS IIIA mice by 904 nm PBM, with decreased IBA1 expression and a return of their morphology towards a resting state. Immune cell immunophenotyping of peripheral blood with mass cytometry revealed increased pro‐inflammatory signalling through pSTAT1 and p‐p38 in NK and T cells in young but not older MPS IIIA mice (5 weeks of age), and expansion of NK, B and CD8+ T cells in older affected mice (17 weeks of age), highlighting the importance of innate and adaptive lymphocytes in Sanfilippo syndrome. Notably, 670 and 904 nm PBM both reversed the Sanfilippo‐induced increase in pSTAT1 and p‐p38 expression in multiple leukocyte populations in young mice, while 904 nm reversed the increase in NK cells in older mice. In conclusion, this is the first study to demonstrate the beneficial effects of PBM in Sanfilippo mice. The distinct reduction in microglial activation and NK cell pro‐inflammatory signalling and number suggests PBM may alleviate neuroinflammation and lymphocyte activation, encouraging further investigation of PBM as a standalone, or complementary therapy in Sanfilippo syndrome. [ABSTRACT FROM AUTHOR]

Vaccination reduces morbidity and mortality due to infections, but efficacy may be limited due to distinct immunogenicity at the extremes of age. This raises the possibility of employing adjuvants to enhance immunogenicity and protection. Early IFNγ production is a hallmark of effective vaccine immunogenicity in adults serving as a biomarker that may predict effective adjuvanticity. We utilized mass cytometry (CyTOF) to dissect the source of adjuvant-induced cytokine production in human blood mononuclear cells (BMCs) from newborns (~39-week-gestation), adults (~18-63 years old) and elders (>65 years of age) after stimulation with pattern recognition receptors agonist (PRRa) adjuvants. Dimensionality reduction analysis of CyTOF data mapped the BMC compartment, elucidated age-specific immune responses and profiled PRR-mediated activation of monocytes and DCs upon adjuvant stimulation. Furthermore, we demonstrated PRRa adjuvants mediated innate IFNγ induction and mapped NK cells as the key source of TLR7/8 agonist (TLR7/8a) specific innate IFNγ responses. Hierarchical clustering analysis revealed age and TLR7/8a-specific accumulation of innate IFNγ producing γδ T cells. Our study demonstrates the application of mass cytometry and cutting-edge computational approaches to characterize immune responses across immunologically distinct age groups and may inform identification of the bespoke adjuvantation systems tailored to enhance immunity in distinct vulnerable populations. An in vitro mass cytometry study focusing on age and adjuvant-specific activation of human blood mononuclear cells reveals NK cells as the key source of TLR7/8 agonist induced innate IFNγ responses. [ABSTRACT FROM AUTHOR]

Myeloid-derived suppressor cells (MDSCs) have played a significant role in facilitating tumor immune escape and inducing an immunosuppressive tumor microenvironment. Eliminating MDSCs and tumor cells remains a major challenge in cancer immunotherapy. A novel approach has been developed using gemcitabine-celecoxib twin drug-based nano-assembled carrier-free nanoparticles (GEM-CXB NPs) for dual depletion of MDSCs and tumor cells in breast cancer chemoimmunotherapy. The GEM-CXB NPs exhibit prolonged blood circulation, leading to the preferential accumulation and co-release of GEM and CXB in tumors. This promotes synergistic chemotherapeutic activity by the proliferation inhibition and apoptosis induction against 4T1 tumor cells. In addition, it enhances tumor immunogenicity by immunogenic cell death induction and MDSC-induced immunosuppression alleviation through the depletion of MDSCs. These mechanisms synergistically activate the antitumor immune function of cytotoxic T cells and natural killer cells, inhibit the proliferation of regulatory T cells, and promote the M2 to M1 phenotype repolarization of tumor-associated macrophages, considerably enhancing the overall antitumor and anti-metastasis efficacy in BALB/c mice bearing 4T1 tumors. The simplified engineering of GEM-CXB NPs, with their dual depletion strategy targeting immunosuppressive cells and tumor cells, represents an advanced concept in cancer chemoimmunotherapy. [ABSTRACT FROM AUTHOR]

The short-lived nature and heterogeneity of Natural Killer (NK) cells limit the development of NK cell-based therapies, despite their proven safety and efficacy against cancer. Here, we describe the biological basis, detailed phenotype and function of long-lived anti-tumour human NK cells (CD56highCD16+), obtained without cell sorting or feeder cells, after priming of peripheral blood cells with Bacillus Calmette-Guérin (BCG). Further, we demonstrate that survival doses of a cytokine combination, excluding IL18, administered just weekly to BCG-primed NK cells avoids innate lymphocyte exhaustion and leads to specific long-term proliferation of innate cells that exert potent cytotoxic function against a broad range of solid tumours, mainly through NKG2D. Strikingly, a NKG2C+CD57-FcεRIγ+ NK cell population expands after BCG and cytokine stimulation, independently of HCMV serology. This strategy was exploited to rescue anti-tumour NK cells even from the suppressor environment of cancer patients' bone marrow, demonstrating that BCG confers durable anti-tumour features to NK cells. [ABSTRACT FROM AUTHOR]

Circulating tumor cells (CTCs) shed from primary tumors must overcome the cytotoxicity of immune cells, particularly natural killer (NK) cells, to cause metastasis. The tumor microenvironment (TME) protects tumor cells from the cytotoxicity of immune cells, which is partially executed by cancer‐associated mesenchymal stromal cells (MSCs). However, the mechanisms by which MSCs influence the NK resistance of CTCs remain poorly understood. This study demonstrates that MSCs enhance the NK resistance of cancer cells in a gap junction‐dependent manner, thereby promoting the survival and metastatic seeding of CTCs in immunocompromised mice. Tumor cells crosstalk with MSCs through an intercellular cGAS‐cGAMP‐STING signaling loop, leading to increased production of interferon‐β (IFNβ) by MSCs. IFNβ reversely enhances the type I IFN (IFN‐I) signaling in tumor cells and hence the expression of human leukocyte antigen class I (HLA‐I) on the cell surface, protecting the tumor cells from NK cytotoxicity. Disruption of this loop reverses NK sensitivity in tumor cells and decreases tumor metastasis. Moreover, there are positive correlations between IFN‐I signaling, HLA‐I expression, and NK tolerance in human tumor samples. Thus, the NK‐resistant signaling loop between tumor cells and MSCs may serve as a novel therapeutic target. [ABSTRACT FROM AUTHOR]

Vps34 is the unique member of the class III phosphoinositide 3‐kinase family that performs both vesicular transport and autophagy. Its role in natural killer (NK) cells remains uncertain. In this study, a model without Vps34 (Vps34fl/fl/CD122Cre/+) is generated, deleting Vps34 during and after NK‐cell commitment. These mice exhibit a nearly 90% decrease in NK cell count and impaired differentiation. A mechanistic study reveals that the absence of Vps34 disrupts the transport of IL‐15 receptor subunit alpha CD122 to the cell membrane, resulting in reduced responsiveness of NK cells to IL‐15. In mice lacking Vps34 at the terminal stage of NK‐cell development (Vps34fl/fl/Ncr1Cre/+), NK cells gradually diminish during aging. This phenotype is associated with autophagy deficiency and the stress induced by reactive oxygen species (ROS). Therefore, terminally differentiated NK cells lacking Vps34 display an accelerated senescence phenotype, while the application of antioxidants effectively reverses the senescence caused by Vps34 deletion by neutralizing ROS. In summary, this study unveils the dual and unique activity of Vps34 in NK cells. Vps34‐mediated vesicular transport is crucial for CD122 membrane trafficking during NK cell commitment, whereas Vps34‐mediated autophagy can delay NK cell senescence. [ABSTRACT FROM AUTHOR]

This study investigates the longitudinal dynamic changes in immune cells in COVID-19 patients over an extended period after recovery, as well as the interplay between immune cells and antibodies. Leveraging single-cell mass spectrometry, we selected six COVID-19 patients and four healthy controls, dissecting the evolving landscape within six months post-viral RNA clearance, alongside the levels of anti-spike protein antibodies. The T cell immunophenotype ascertained via single-cell mass spectrometry underwent validation through flow cytometry in 37 samples. Our findings illuminate that CD8 + T cells, gamma-delta (gd) T cells, and NK cells witnessed an increase, in contrast to the reduction observed in monocytes, B cells, and double-negative T (DNT) cells over time. The proportion of monocytes remained significantly elevated in COVID-19 patients compared to controls even after six-month. Subpopulation-wise, an upsurge manifested within various T effector memory subsets, CD45RA + T effector memory, gdT, and NK cells, whereas declines marked the populations of DNT, naive and memory B cells, and classical as well as non-classical monocytes. Noteworthy associations surfaced between DNT, gdT, CD4 + T, NK cells, and the anti-S antibody titer. This study reveals the changes in peripheral blood mononuclear cells of COVID-19 patients within 6 months after viral RNA clearance and sheds light on the interactions between immune cells and antibodies. The findings from this research contribute to a better understanding of immune transformations during the recovery from COVID-19 and offer guidance for protective measures against reinfection in the context of viral variants. [ABSTRACT FROM AUTHOR]

Pancreatic cancer is a major cause of cancer-related death, but despondently, the outlook and prognosis for this resistant type of tumor have remained grim for a long time. Currently, it is extremely challenging to prevent or detect it early enough for effective treatment because patients rarely exhibit symptoms and there are no reliable indicators for detection. Most patients have advanced or spreading cancer that is difficult to treat, and treatments like chemotherapy and radiotherapy can only slightly prolong their life by a few months. Immunotherapy has revolutionized the treatment of pancreatic cancer, yet its effectiveness is limited by the tumor's immunosuppressive and hard-to-reach microenvironment. First, this article explains the immunosuppressive microenvironment of pancreatic cancer and highlights a wide range of immunotherapy options, including therapies involving oncolytic viruses, modified T cells (T-cell receptor [TCR]-engineered and chimeric antigen receptor [CAR] T-cell therapy), CAR natural killer cell therapy, cytokine-induced killer cells, immune checkpoint inhibitors, immunomodulators, cancer vaccines, and strategies targeting myeloid cells in the context of contemporary knowledge and future trends. Lastly, it discusses the main challenges ahead of pancreatic cancer immunotherapy. [ABSTRACT FROM AUTHOR]

Lymphocyte subsets are the most intuitive expression of the body's immune ability, and the lymphocyte-to-monocyte ratio (LMR) also clearly reflect the degree of chronic inflammation activity. The purpose of this study is to investigate their predictive value of lymphocyte subsets and LMR to neoadjuvant therapy (NAT) efficacy in breast cancer patients. In this study, lymphocyte subsets and LMR were compared between breast cancer patients (n = 70) and benign breast tumor female populations (n = 48). Breast cancer patients were treated with NAT, and the chemotherapy response of the breast was evaluated using established criteria. The differences in lymphocyte subsets and LMR were also compared between pathological complete response (pCR) and non-pCR patients before and after NAT. Finally, data were analyzed using SPSS. The analytical results demonstrated that breast cancer patients showed significantly lower levels of CD3 + T cells, CD4 + T cells, CD4 + /CD8 + ratio, NK cells, and LMR compared to benign breast tumor women (P < 0.05). Among breast cancer patients, those who achieved pCR had higher levels of CD4 + T cells, NK cells, and LMR before NAT (P < 0.05). NAT increased CD4 + /CD8 + ratio and decreased CD8 + T cells in pCR patients (P < 0.05). Additionally, both pCR and non-pCR patients exhibited an increase in CD3 + T cells and CD4 + T cells after treatment, but the increase was significantly higher in pCR patients (P < 0.05). Conversely, both pCR and non-pCR patients experienced a decrease in LMR after treatment. However, this decrease was significantly lower in pCR patients (P < 0.05). These indicators demonstrated their predictive value for therapeutic efficacy. In conclusion, breast cancer patients experience tumor-related immunosuppression and high chronic inflammation response. But this phenomenon can be reversed to varying degrees by NAT. It has been found that lymphocyte subsets and LMR have good predictive value for pCR. Therefore, these markers can be utilized to identify individuals who are insensitive to NAT early on, enabling the adjustment of treatment plans and achieving precise breast cancer treatment. [ABSTRACT FROM AUTHOR]

The mammalian placenta's interface with the parent is a richly vascularized tissue whose development relies upon communication between many different cell types within the uterine microenvironment. The uterine blood vessels of the interface are reshaped during pregnancy into wide‐bore, flaccid vessels that convey parental blood to the exchange region of the placenta. Invasive trophoblast as well as parental uterine macrophages and Natural Killer cells are involved in the stepwise remodeling of these vessels and their respective contributions to this crucial process are still being delineated. However, the earliest steps in arteriole remodeling are understudied as they are difficult to study in humans, and other species lack the deep trophoblast invasion that is so prominent a feature of placentation in humans. Here, we further characterize the rat, with deep hemochorial placentation akin to humans, as a model system in which to tease apart the earliest, relatively understudied events in spiral arteriole remodeling. We show that the rat uterine−placental interface increases in size and vascularity rapidly, before trophoblast invasion. The remodeling stages in the arterioles of the rat uterine–placental interface follow a sequence of anatomical changes similar to those in humans, and there are changes to the arterioles' muscular tunica media prior to the marked influx of immune cells. The rat is a tractable model in which to better understand the cell/cell interactions occurring in vivo in an intact tissue microenvironment over time. [ABSTRACT FROM AUTHOR]

Autoimmune hepatitis (AIH) is an immune disorder characterized by hypergammaglobulinemia, autoantibodies, and chronic active hepatitis on liver histology. However, immune cell population characteristics in AIH patients remain poorly understood. This study was designed to analyze peripheral blood mononuclear cell (PBMC) characteristics in AIH through single-cell RNA sequencing (scRNA-seq) and explore potential AIH-related molecular mechanisms. We generated 3690 and 3511 single-cell transcriptomes of PBMCs pooled from 4 healthy controls (HCs) and 4 AIH patients, respectively, by scRNA-seq. These pooled PBMC transcriptomes were used for cell cluster identification and differentially expressed gene (DEG) identification. GO functional enrichment analysis was performed on the DEGs to determine the most active AIH immune cell biological functions. Although the PCA-based uniform manifold approximation and projection (UMAP) algorithm was used to cluster cells with similar expression patterns in the two samples, 87 up- and 12 downregulated DEGs were retained in monocytes and 101 up- and 15 downregulated DEGs were retained in NK cells from AIH PBMCs. Moreover, enriched GO terms in the PBMC-derived monocyte and NK cell clusters were related mainly to antigen processing and presentation, IFN-γ-mediated signaling, and neutrophil degranulation and activation. These potential molecular mechanisms may be important targets for AIH treatment. [ABSTRACT FROM AUTHOR]

Adipose tissue was once known as a reservoir for energy storage but is now considered a crucial organ for hormone and energy flux with important effects on health and disease. Glucose-dependent insulinotropic polypeptide (GIP) is an incretin hormone secreted from the small intestinal K cells, responsible for augmenting insulin release, and has gained attention for its independent and amicable effects with glucagon-like peptide 1 (GLP-1), another incretin hormone secreted from the small intestinal L cells. The GIP receptor (GIPR) is found in whole adipose tissue, whereas the GLP-1 receptor (GLP-1R) is not, and some studies suggest that GIPR action lowers body weight and plays a role in lipolysis, glucose/lipid uptake/disposal, adipose tissue blood flow, lipid oxidation, and free-fatty acid (FFA) re-esterification, which may or may not be influenced by other hormones such as insulin. This review summarizes the research on the effects of GIP in adipose tissue (distinct depots of white and brown) using cellular, rodent, and human models. In doing so, we explore the mechanisms of GIPR-based medications for treating metabolic disorders, such as type 2 diabetes and obesity, and how GIPR agonism and antagonism contribute to improvements in metabolic health outcomes, potentially through actions in adipose tissues. [ABSTRACT FROM AUTHOR]

Gestational diabetes mellitus (GDM) is a frequent and serious complication of pregnancy, often associated with obesity. Metabolic dysfunction and metainflammation are evident in both obesity and GDM. In this cross-sectional study, we aimed at defining the direct contribution of the immune system in GDM, across the main metabolic tissues, specifically focussing on elucidating the roles of obesity and GDM to the clinical outcome. Using immunoassays and multicolour flow cytometry, cytokine profiles and immune cell frequencies were measured in maternal circulation and central metabolic tissues [placenta and visceral adipose tissue (VAT)] in GDM-diagnosed (n = 28) and normal glucose tolerant (n = 32) women undergoing caesarean section. Participants were sub-grouped as non-obese [body mass index (BMI) < 30 kg/m2] or obese (BMI ≥ 30 kg/m2). Unsupervised data analysis was performed on the flow cytometry data set to identify functional alterations. GDM obese participants had significantly elevated circulating IL-6 and IL-17A levels. GDM non-obese participants had elevated circulating IL-12p70, elevated placental IL-17A, and VAT IFN-γ production. Unsupervised clustering of immune populations across the three biological sites simultaneously, identified different NK- and T-cell phenotypes that were altered in NGT obese and GDM non-obese participants, while a classical tissue monocyte cluster was increased in GDM obese participants. In this study, there was significant evidence of subclinical inflammation, and significant alterations in clusters of NK cells, T cells, and tissue monocyte populations in GDM. While increased adiposity assimilates with increased inflammation in the non-pregnant state, this overt relationship may not be as evident during pregnancy and warrants further examination in future longitudinal studies. Gestational diabetes mellitus (GDM) is a frequent and serious complication of pregnancy, often associated with obesity. In this study, there was significant evidence of subclinical inflammation in maternal circulation and in both placental and visceral adipose tissue. However, no direct causal role could be attributed to any specific immune cell population in GDM. Graphical Abstract [ABSTRACT FROM AUTHOR]

Very recently, an interesting phenomenon was described in the common vole; vole parents with similar locomotor ability produced significantly larger litters. Positive assortative mating is a tendency to prefer individuals with similar phenotypes. We tested whether this also applies to smell similarity. Odour preference was tested in a T-maze, where each female was presented with two male odours, i.e. shavings together with feces and urine from home boxes. After female preference was established, the female was either paired with a preferred male (chosen) or paired with a non-preferred male (opposite choice). For analysis of the relationship to odour preference, genotyping of major histocompatibility complex (MHC) Class II DRB was done using amplicon sequencing. In the set of 45 individuals from two populations, we recovered 38 nucleotide haplotypes (alleles). Similarity of alleles in parent pairs according to the indexes of Sørensen–Dice (S–D) and Jaccard were calculated. Values of these indexes in parental pairs with preferred males were significantly higher (more similar) than in not preferred. The number of offspring in parental pairs with preferred males were significantly higher than in not preferred males. However, there is no correlation between the mentioned indexes and the number of offspring. The relationship between the success of reproduction and alleles is not clear-cut, this may be influenced by the measure of similarity we used, or by something that we could not detect. [ABSTRACT FROM AUTHOR]

There are no medical therapies to stop progression of pulmonary emphysema. In chronic obstructive pulmonary disease (COPD), lung NK cell cytotoxicity towards lung parenchymal cells is increased, lung dendritic cells are activated, and lung regulatory T cell (Treg) numbers are decreased. We propose that re-establishing the balance between these three cell types might be a crucial component to arresting emphysema progression. Developing Treg-based therapies for COPD may require strategies to ensure the stability of Treg phenotypes. An increased understanding of the presumed dysregulated immune interactions between dendritic cells (DCs), natural killer cells (NK), and regulatory T cells (Tregs) in COPD might lead to novel and potentially disease-modifying COPD therapies that target specific cell types. Chronic obstructive pulmonary disease (COPD) is characterized by infiltration of the airways and lung parenchyma by inflammatory cells. Lung pathology results from the cumulative effect of complex and aberrant interactions between multiple cell types. However, three cell types, natural killer cells (NK), dendritic cells (DCs), and regulatory T cells (Tregs), are understudied and underappreciated. We propose that their mutual interactions significantly contribute to the development of COPD. Here, we highlight recent advances in NK, DC, and Treg biology with relevance to COPD, discuss their pairwise bidirectional interactions, and identify knowledge gaps that must be bridged to develop novel therapies. Understanding their interactions will be crucial for therapeutic use of autologous Treg, an approach proving effective in other diseases with immune components. [ABSTRACT FROM AUTHOR]

Simple Summary: Mammary gland tumors are prevalent in canines, posing significant threats to their health and survival. With the limited efficacy of traditional treatments due to the high heterogeneity of these tumors, there is a growing focus on exploring new therapeutic avenues. Natural Killer cell-based immunotherapy has emerged as a promising approach in anti-tumor treatment, prompting research into optimization strategies. Resveratrol, a natural polyphenol compound, shows potential in regulating Natural Killer cell immune activity. However, the specific impact of Resveratrol-activated Natural Killer cells on canine mammary tumors and their therapeutic potential remain unclear. This study investigated the therapeutic potential of Resveratrol-activated Natural Killer cells in treating canine mammary tumors. Using various assays, including wound healing and flow cytometry, the study examined the impact of Resveratrol-pretreated Natural Killer cells on tumor cells in vitro and in a mouse model. The results demonstrate that Resveratrol enhances Natural Killer cells' ability to induce tumor cell death and inhibit tumor growth, migration, and invasion. Moreover, Resveratrol boosts Natural Killer cell recruitment to other immune cells in the body. Overall, Resveratrol shows promise in enhancing Natural Killer cell-mediated anti-tumor effects, suggesting a potential avenue for optimizing immunotherapy for canine mammary tumors and improving treatment efficacy. In order to explore the therapeutic effect of Resveratrol (Res)-activated Natural Killer (NK) cells on canine mammary tumors, this study employed a range of assays, including wound healing, colony formation, Transwell, flow cytometry, and Western blot experiments, to investigate the impact of Res-pretreated NK cells on canine mammary tumor cells in vitro. Additionally, a tumor-bearing mouse model was utilized to further analyze the therapeutic effects of Res-pretreated NK cells in vivo. The results showed that Res enhances the capacity of NK cells to induce apoptosis, pyroptosis, and ferroptosis in canine breast tumor cells, while also augmenting their influence on the migration, invasion, and epithelial-mesenchymal transition of these cells. Furthermore, pretreatment of NK cells with Res significantly amplified their inhibitory effect on breast tumor growth in vivo and promoted tumor tissue apoptosis. Additionally, Res enhanced the recruitment of NK cells to other immune cells in the body. In summary, Res has been shown to enhance the anti-breast-tumor effect of NK cells both in vitro and in vivo, offering a new avenue for optimizing immunotherapy for canine breast tumors. [ABSTRACT FROM AUTHOR]

Background: Ozanimod showed efficacy and safety in the phase 2 STEPSTONE study conducted in patients with moderately to severely active Crohn's disease. Aims: This analysis assessed the effects of ozanimod on circulating lymphocytes in Crohn's disease. Methods: Patients received ozanimod 0.92 mg for 12 weeks. Lymphocyte subtypes were evaluated using multicolor flow analysis on blood samples collected before treatment and on Week 12. Absolute lymphocyte count changes were analyzed by Wilcoxon signed rank tests. Disease activity changes and efficacy outcomes were evaluated at Week 12, and associations with lymphocyte subtype levels were assessed using Spearman's correlation and logistic regression. Results: Reductions in median total T, Th, and cytotoxic T cells occurred at Week 12 (45.4%–76.8%), with reductions in most subtypes of 47.5% to 91.3% (P < 0.001). CD8+ terminally differentiated effector memory cells were largely unaffected (median change, − 19%; P = 0.44). Reductions in median total B cells occurred at Week 12 (76.7%), with reductions in subtypes of 71.4% to 81.7% (P < 0.001). Natural killer and monocyte cell counts were unchanged. Greater baseline levels and changes in nonswitched memory B cells were significantly associated with clinical, endoscopic, and histologic efficacy (P < 0.05, all comparisons). Conclusions: Ozanimod reduced circulating levels of all B-cell and most T-cell subsets but not monocytes or natural killer cells. Key subsets relevant to immune surveillance were not reduced, supporting the low risk of infection and malignancy with ozanimod in chronic inflammatory diseases. Levels of nonswitched memory B cells were associated with efficacy, providing a potential marker for ozanimod response. Trial Registration: ClinicalTrials.gov: NCT02531113, EudraCT: 2015–002025–19 [ABSTRACT FROM AUTHOR]

Breast cancer is influenced by factors such as diet, a sedentary lifestyle, obesity, and postmenopausal status, which are all linked to prolonged hormonal and inflammatory exposure. Physical activity offers protection against breast cancer by modulating hormones, immune responses, and oxidative defenses. This study aimed to assess how a prolonged high-fat diet (HFD) affects the effectiveness of physical activity in preventing and managing mammary tumorigenesis. Ovariectomised C57BL/6 mice were provided with an enriched environment to induce spontaneous physical activity while being fed HFD. After 44 days (short-term, ST HFD) or 88 days (long-term, LT HFD), syngenic EO771 cells were implanted into mammary glands, and tumour growth was monitored until sacrifice. Despite similar physical activity and food intake, the LT HFD group exhibited higher visceral adipose tissue mass and reduced skeletal muscle mass. In the tumour microenvironment, the LT HFD group showed decreased NK cells and TCD8+ cells, with a trend toward increased T regulatory cells, leading to a collapse of the T8/Treg ratio. Additionally, the LT HFD group displayed decreased tumour triglyceride content and altered enzyme activities indicative of oxidative stress. Prolonged exposure to HFD was associated with tumour growth despite elevated physical activity, promoting a tolerogenic tumour microenvironment. Future studies should explore inter-organ exchanges between tumour and tissues. [ABSTRACT FROM AUTHOR]

Atopic dermatitis (AD) is one of the most common chronic inflammatory skin diseases, with an increasing number of targeted therapies available. While biologics to treat AD exclusively target the key cytokines of type 2 immunity, Janus kinase inhibitors target a broad variety of cytokines, including IFN-γ. To better stratify patients for optimal treatment outcomes, the identification and characterization of subgroups, especially with regard to their IFNG expression, is of great relevance, as the role of IFNG in AD has not yet been fully clarified. This study aims to define AD subgroups based on their lesional IFNG expression and to characterize them based on their gene expression, T cell secretome and clinical attributes. RNA from the lesional and non-lesional biopsies of 48 AD patients was analyzed by RNA sequencing. Based on IFNG gene expression and the release of IFN-γ by lesional T cells, this cohort was categorized into three IFNG groups (high, medium, and low) using unsupervised clustering. The low IFNG group showed features of extrinsic AD with a higher prevalence of atopic comorbidities and impaired epidermal lipid synthesis. In contrast, patients in the high IFNG group had a higher average age and an activation of additional pro-inflammatory pathways. On the cellular level, higher amounts of M1 macrophages and natural killer cell signaling were detected in the high IFNG group compared to the low IFNG group by a deconvolution algorithm. However, both groups shared a common dupilumab response gene signature, indicating that type 2 immunity is the dominant immune shift in both subgroups. In summary, high and low IFNG subgroups correspond to intrinsic and extrinsic AD classifications and might be considered in the future for evaluating therapeutic efficacy or non-responders. [ABSTRACT FROM AUTHOR]

Enhancing immune cell functions in tumors remains a major challenge in cancer immunotherapy. Natural killer cells (NK) are major innate effector cells with broad cytotoxicity against tumors. Accordingly, NK cells are ideal candidates for cancer immunotherapy, including glioblastoma (GBM). Hypoxia is a common feature of solid tumors, and tumor cells and normal cells adapt to the tumor microenvironment by upregulating the transcription factor hypoxia-inducible factor (HIF)-1α, which can be detrimental to anti-tumor effector immune cell function, including that of NK cells. We knocked out HIF-1α in human primary NK cells using clustered regularly interspaced short palindromic repeat (CRISPR)-associated protein 9 (Cas9). Then, cellular characterizations were conducted in normoxic and hypoxic conditions. Electroporating two HIF-1α-targeting guide RNA–Cas9 protein complexes inhibited HIF-1α expression in expanded NK cells. HIF-1α knockout human NK cells, including populations in hypoxic conditions, enhanced the growth inhibition of allogeneic GBM cells and induced apoptosis in GBM-cell-derived spheroids. RNA-sequencing revealed that the cytotoxicity of HIF-1α knockout NK cells could be related to increased perforin and TNF expression. The results demonstrated that HIF-1α knockout human NK cells, including populations, enhanced cytotoxicity in an environment mimicking the hypoxic conditions of GBM. CRISPR–Cas9-mediated HIF-1α knockout NK cells, including populations, could be a promising immunotherapeutic alternative in patients with GBM. [ABSTRACT FROM AUTHOR]

Primary Epstein–Barr virus (EBV) infection which can manifest as infectious mononucleosis (IM) is commonly acquired during childhood. EBV primarily invades B cells leading to a lytic reaction; the control of the infection is handled by natural killer and T cells in immunocompetent individuals. The infection has a wide spectrum of clinical findings and can lead to serious complications in patients with certain underlying immunological dysfunctions. We retrospectively investigated peripheral white blood cell populations' surface marker characteristics in IM using a comprehensive flow cytometry marker panel. Twenty-one cases of IM and seventeen EBV-seropositive cases without IM serving as controls were included. We observed novel alterations in lymphocyte, neutrophil, and monocyte populations. In addition to increased activated cytotoxic T cells and low B cells, we demonstrated high T-large granular lymphocyte (T-LGL) populations in IM cases. Furthermore, despite T cells' increased HLA-DR expression, another activation marker, CD11b, was lower in T-LGL populations. Monocytes showed increased CD16 expression; CD64 was higher in neutrophils. Our findings point to monocyte and neutrophil activation which may account for acute clinical features and may contribute to the understanding of IM immunobiology. Furthermore, they may serve as a useful tool in investigating inherited and post-transplant conditions characterized by deficiencies in controlling EBV infection. [ABSTRACT FROM AUTHOR]

Plasma gelsolin (pGSN) overexpression in ovarian cancer (OVCA) disarms immune function, contributing to chemoresistance. The aim of this study was to investigate the immunoregulatory effects of pGSN expression on natural killer (NK) cell function in OVCA. OVCA tissues from primary surgeries underwent immunofluorescent staining of pGSN and the activated NK cell marker natural cytotoxicity triggering receptor 1 to analyze the prognostic impact of pGSN expression and activated NK cell infiltration. The immunoregulatory effects of pGSN on NK cells were assessed using apoptosis assay, cytokine secretion, immune checkpoint-receptor expression, and phosphorylation of STAT3. In OVCA tissue analyses, activated NK cell infiltration provided survival advantages to patients. However, high pGSN expression attenuated the survival benefits of activated NK cell infiltration. In the in vitro experiment, pGSN in OVCA cells induced NK cell death through cell-to-cell contact. pGSN increased T-cell immunoglobulin and mucin-domain-containing-3 expression (TIM-3) on activated NK cells. Further, it decreased interferon-γ production in activated TIM-3+ NK cells, attenuating their anti-tumor effects. Thus, increased pGSN expression suppresses the anti-tumor functions of NK cells. The study provides insights into why immunotherapy is rarely effective in patients with OVCA and suggests novel treatment strategies. [ABSTRACT FROM AUTHOR]

This document, titled "In This Issue," provides a summary of several articles published in the journal International Immunology. The first article discusses the reprogramming of T-cell metabolism to enhance T-cell immunotherapy, particularly in the context of solid tumors with an immunosuppressive metabolic microenvironment. The second article focuses on the crystal structure of the C-type lectin receptor CLEC12A and its potential diagnostic and therapeutic implications. The third article explores a novel approach to identify statin-mediated anti-cancer effects by analyzing the effects of inhibiting HMGCR variants on lipid metabolism and immunotherapy. The fourth article investigates the role of glycan-mediated lymphocyte homing in experimental autoimmune encephalomyelitis (EAE) and its potential as a therapeutic target in multiple sclerosis (MS). Lastly, the fifth article examines the N-glycosylation of CD155 and its impact on NK cell activation, suggesting that therapeutic antibodies targeting CD155 may be more effective against tumors expressing N105-glycosylated CD155. [Extracted from the article]

The cluster of differentiation 155 (CD155) is highly expressed on tumor cells and augments or inhibits the cytotoxic activities of natural killer (NK) cells and T cells through its receptor ligands DNAX accessory molecule 1 (DNAM-1) and T-cell immunoglobulin (Ig) and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT), respectively. Although CD155 is heavily glycosylated, the role of glycosylation of CD155 in the cytotoxic activity of effector lymphocytes remains unknown. Here, we show that the N-linked glycosylation at residue 105 (N105 glycosylation) in the first Ig-like domain of CD155 is involved in the binding of CD155 to both DNAM-1 and TIGIT. The N105 glycosylation also plays an essential role to induce signaling in both DNAM-1 and TIGIT reporter cells. Moreover, we show that the N105 glycosylation of CD155 contributes preferentially to the DNAM-1-mediated activating signal over the TIGIT-mediated inhibitory signal in NK cells. Our results demonstrated the important role of the N105 glycosylation of CD155 in DNAM-1 and TIGIT functions and shed new light on the understanding of tumor immune responses. [ABSTRACT FROM AUTHOR]

Previous observational and experimental studies have suggested a relationship between statin treatments and the augmentation of immunotherapy effects; however, the causal role of statin usage in promoting antitumor immunity remains largely unexplored. Utilizing large-scale genome-wide association studies, we conducted a Mendelian Randomization (MR) analysis to examine the association between genetically proxied inhibition of the gene for 3-hydroxy-3-methylglutaryl coenzyme A reductase (HMGCR), a specific target of statins, and 524 immunotherapy-related profiles, encompassing immune cells, inflammatory cytokines, immune checkpoints, and gut microbiota. Our findings indicated a suggestive association between statin therapy and proinflammatory as well as antitumor effects; notably, inhibition of HMGCR demonstrated a robust link with increased susceptibility of various immune cell types, including basophil cells, white blood cells, eosinophil cells, neutrophil cells, activated CD8+ T cells, dendritic cells, and natural killer cells; furthermore, a causal relationship was observed between statin use and a decrease in terminal CD8+ T cells, granulocytes, monocytes, and myeloid-derived suppressor cells; genetically proxied statin usage was also significantly associated with elevated levels of proinflammatory cytokines and immunotherapy-related gut microbiota; importantly, the potential inhibition of HMGCR in influencing the response to immunotherapy was confirmed in the real-world cohorts. This study provides novel insights into the regulatory role of HMGCR inhibition in antitumor immunity, suggesting that strategies targeting HMGCR or lipid regulation may hold therapeutic potential for enhancing the efficacy of immunotherapy. [ABSTRACT FROM AUTHOR]

Objectives Because of its low frequency in adult populations and clinical and laboratory overlap with hemophagocytic lymphohistiocytosis and other T-cell lymphomas, T-cell/natural killer (NK) cell systemic, chronic, active Epstein-Barr virus (EBV) (T/NK sCAEBV) infection remains underdiagnosed, preventing critical, prompt therapeutic interventions. Methods We report a 5-case series that included 2 adult patients with T/NK sCAEBV and 3 additional adult patients with T/NK lymphomas with concomitant systemic EBV infection to review these entities' overlapping diagnostic and clinical features. Results Approximately 95% of the world population has been infected with EBV during their lifetime, and infection is usually asymptomatic, with symptomatic cases eventually resolving spontaneously. A small subset of immunocompetent patients develops CAEBV, a life-threatening complication resulting from EBV-infected T-cell or NK cell neoplastic lymphocytes. The sites of end-organ damage in T/NK sCAEBV demonstrate pathologic findings such as reactive lymphoid proliferations, making the diagnosis difficult to establish, with the only curative option being an allogeneic hematopoietic stem cell transplant. Conclusions This diagnosis is most prevalent in Asia, with few cases reported in Western countries. Adult age is an independent risk factor for poor outcomes, and most cases are diagnosed in pediatric populations. [ABSTRACT FROM AUTHOR]

Multiple myeloma (MM) is a hematological malignancy caused by the clonal expansion of malignant plasma cells in the bone marrow. Myeloma cells are susceptible to killing by natural killer (NK) cells, but NK cells fail to control disease progression, suggesting immunosuppression. The activation threshold of NK-effector function is regulated by interaction between KIRs and self-HLA class I, during a process called "education" to ensure self-tolerance. NK cells can respond to diseased cells based on the absence of HLA class I expression ("Missing-self" hypothesis). The HLA and KIR repertoire is extremely diverse; thus, the present study aimed to characterize potential variances in genotypic composition of HLA Class I NK-epitopes and KIRs between MM patients and healthy controls. Genotypic expression of KIR and HLA (HLA-C group-C1/C2 and Bw4 motifs (including HLA-A*23, A*24, A*32) were analyzed in 172 MM patients and 195 healthy controls. Compared to healthy controls, we did not observe specific KIR genes or genotypes, or HLA NK-epitopes with higher prevalence among MM patients. The presence of all three HLA NK-epitopes (C1+C2+Bw4+) was not associated with MM occurrence. However, MM patients were more likely to be C1-/C2+/Bw4+ (p = 0.049, OR 1.996). In line with this, there was a trend of increased genetic co-occurrence of Bw4 and KIR3DL1 in MM patients (p = 0.05, OR 1.557). Furthermore, MM patients were more likely to genetically express both C2/KIR2DL1 and Bw4/KIR3DL1 (p = 0.019, OR 2.453). Our results reveal an HLA NK-epitope combination that is associated with the occurrence of MM. No specific KIR genotypes were associated with MM. [ABSTRACT FROM AUTHOR]

Rheumatoid arthritis (RA) is a type of autoimmune disease that results in immune disorder and excessive inflammatory response due to a reduction of self-tolerance. Invariant natural killer T (iNKT) cells can effectively alleviate clinical symptoms and hyper-inflammation in RA, but their mechanism of action is not well-defined. This study aims to investigate the mechanism of iNKT cell therapy for RA. We established a DBA/1 mouse model for RA and treated it with specific iNKT cells. A cytometric bead array was used to measure the amounts of cytokines in the serum. Flow cytometry was then employed to identify different subsets of helper T cells (Th), the frequency of conventional dendritic cells (cDC), the expression of CD80, CD86, programmed cell death ligand 1 (PD-L1), and PD-L2 on cDC surfaces, and associated pathway proteins. iNKT cell treatment reduced Th1/Th2 and Th17/ regulatory T (Treg) cell ratios while increasing interleukin-4 (IL-4) and IL-10. It enhanced the generation of immature cDCs, and it upregulated the level of PD-L2 by stimulating the signal transducer and activator of transcription 3 (STAT3) signaling pathway. Meanwhile, it activated the extracellular signal-regulated kinase 1/2 (ERK1/2) pathway and inhibited the nuclear factor kappa B (NF-κB) pathway. According to our findings, iNKT cell treatment increased the expression of phosphates STAT3 in lymph node cDC, causing them to upregulate PD-L2 molecules. While activating the ERK1/2 pathway and inhibiting the NF-κB pathway, tolerogenic cDC was produced, restoring immune homeostasis and correcting excessive inflammation. These results deliver new insights into the treatment of RA by iNKT cells. [ABSTRACT FROM AUTHOR]

Objectives: Gout is characterized by hyperuricemia and recurrent inflammatory episodes caused by intra-articular crystal deposition of monosodium urate (MSU). There is a clear relationship between gout and metabolic syndrome. Recent evidence indicates that perforin plays a role in regulating glucose homeostasis and provides protection in diet-induced non-alcoholic steatohepatitis models. However, the impact of perforin on immune inflammation in gout remains unclear. Methods: We induced acute gout models in both wild-type (WT) mice and Prf1null mice by administering intra-articular injections of MSU crystals. We compared the ankle joint swelling and the histological score between the two groups. Furthermore, we investigated underlying mechanisms through in vitro co-culture experiments involving CD8 T cells and macrophages. Results: In this study, Prf1null mice showed significantly more pronounced ankle swelling with increased inflammatory cell infiltrations compared with WT mice 24 h after local MSU injection. Moreover, MSU-induced Prf1null mice exhibited increased accumulation of CD8 T cells but not NK cells. Perforin-deficient CD8 T cells displayed reduced cytotoxicity towards bone marrow–derived M0 and M1 macrophages and promoted TNF-α secretion from macrophage. Conclusions: Perforin from CD8 T cells limits joint inflammation in mice with acute gout by downregulating macrophage-mediated inflammation. Key Points • Perforin deficiency increased swelling in the ankle joints of mice upon MSU injection. • Perforin deficiency is associated with increased immune cell recruitment and severe joint damage in gout. • Perforin regulated CD8 T cell accumulation in gout and promoted CD8 T cell cytotoxicity towards M0 and M1 macrophages. • CD8 T cell-derived perforin regulated pro-inflammatory cytokine secretion of macrophage. [ABSTRACT FROM AUTHOR]

Although Multiple Sclerosis (MS) is primarily thought to be an autoimmune condition, its possible viral etiology must be taken into consideration. When mice are administered neurotropic viruses like mouse hepatitis virus MHV‐A59, a murine coronavirus, or its isogenic recombinant strain RSA59, neuroinflammation along with demyelination are observed, which are some of the significant manifestations of MS. MHV‐A59/RSA59 induced neuroinflammation is one of the best‐studied experimental animal models to understand the viral‐induced demyelination concurrent with axonal loss. In this experimental animal model, one of the major immune checkpoint regulators is the CD40‐CD40L dyad, which helps in mediating both acute‐innate, innate‐adaptive, and chronic‐adaptive immune responses. Hence, they are essential in reducing acute neuroinflammation and chronic progressive adaptive demyelination. While CD40 is expressed on antigen‐presenting cells and endothelial cells, CD40L is expressed primarily on activated T cells and during severe inflammation on NK cells and mast cells. Experimental evidences revealed that genetic deficiency of both these proteins can lead to deleterious effects in an individual. On the other hand, interferon‐stimulated genes (ISGs) possess potent antiviral properties and directly or indirectly alter acute neuroinflammation. In this review, we will discuss the role of an ISG, ISG54, and its tetratricopeptide repeat protein Ifit2; the genetic and experimental studies on the role of CD40 and CD40L in a virus‐induced neuroinflammatory demyelination model. [ABSTRACT FROM AUTHOR]

Thymoma is frequently correlated with various autoimmune diseases. However, unequivocal therapeutic targets for thymoma remain undefined, and the role of immune checkpoints in the development of thymoma-related autoimmune illnesses is unclear. We examined 39 thymoma samples and 44 normal control samples from the GEO database. Following batch correction, we identified 224 Differentially Expressed Genes (DEGs) using the Limma package. We employed Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses to enrich for functional pathways of DEGs. We utilized a Protein–Protein Interaction (PPI) network to identify hub genes and determine their correlation with immune cell infiltration using CIBERSORT. Real-time quantitative polymerase chain reaction (RT-qPCR), western blot, and immunohistochemical staining were implemented to verify identified hub genes in vivo. Simultaneously, we evaluated the prognostic relevance of the hub gene using clinical data. We determined COL1A1, COL1A2, and BGN to be the central hub genes in thymoma. Validation via RT-qPCR, Western blot, and immunohistochemical staining established significant statistical divergence between thymoma tissue and the normal thymus for only BGN. Expression levels of BGN showed strong negative correlation with the infiltration level of B cells and CD4+ T cells, yet a significant positive correlation with the level of neutrophil infiltration. We found high immune infiltration levels of macrophages, NK cells, and Th1 cells in the thymoma microenvironment in patients with a high expression of BGN. Co-localization of BGN and macrophages within thymoma tissue was discerned via tissue staining. Clinical data dictated that thymoma patients exhibiting elevated BGN expression underwent longer hospital stays, longer lengths in intensive care units, greater hospitalization costs, and extended ventilator usage; our study, augmented by clinical information, recognized BGN as possessive of diagnostic and prognostic significance in thymoma through in silico and molecular verification experiments. Our findings offered an important objective for thymoma-treated autoimmune disease comprehension, supplemented by the strong association with immune infiltration. [ABSTRACT FROM AUTHOR]

Objective: This retrospective study aimed to investigate the prevalence of chronic endometritis, diagnosed using CD138 immunohistochemistry, among infertile women and to assess the association between chronic endometritis and recurrent implantation failure (RIF). Methods: In total, 266 patients who underwent hysteroscopy due to infertility between 2019 and 2020 were included in the analysis. Of these, 136 patients with RIF and 130 non-RIF patients were included in the study. CD138 immunohistochemistry test results, blood biomarkers (including natural killer cells, white blood cells, and the lymphocyte-to-neutrophil ratio), and data on pregnancy outcomes were obtained. If the CD138 test yielded a positive result, the patients received antibiotic treatment. Results: The overall proportion of CD138-positive patients was 32.7% (87/266). The CD138 positivity rate was not related to the number of cycles with implantation failure. In the RIF patient group, no significant associations were found between CD138 positivity and peripheral blood markers. The clinical pregnancy rates were similar between infertile women treated with antibiotics for chronic endometritis and those without chronic endometritis. Conclusion: To improve the pregnancy rate in infertile patients, it may be helpful to combine CD138 testing with other laboratory tests and administer antibiotic treatment if the result is positive. [ABSTRACT FROM AUTHOR]

The success of mRNA vaccines against COVID-19 has enhanced the potential of lipid nanoparticles (LNPs) as a system for the delivery of mRNA. In this review, we describe our progress using a lipid library to engineer ionizable lipids and promote LNP technology from the viewpoints of safety, controlled biodistribution, and mRNA vaccines. These advancements in LNP technology are applied to cancer immunology, and a potential nano-DDS is constructed to evaluate immune status that is associated with a cancer-immunity cycle that includes the sub-cycles in tumor microenvironments. We also discuss the importance of the delivery of antigens and adjuvants in enhancing the cancer-immunity cycle. Recent progress in NK cell targeting in cancer immunotherapy is also introduced. Finally, the impact of next-generation DDS technology is explained using the MITO-Porter membrane fusion-based delivery system for the organelle targeting of the mitochondria. We introduce a successful example of the MITO-Porter used in a cell therapeutic strategy to treat cardiomyopathy. [Display omitted] • The efficiency of lipid nanoparticles is discussed from the viewpoint of a 3-dimensinal structure of ionizable lipids. • A potential nano-DDS is designed based on a cancer immunity cycle that includes sub-cycles in a tumor microenvironment. • MITO-Porter was introduced as an organelle-targeting DDS that was successfully applied to cell transplantation therapy. [ABSTRACT FROM AUTHOR]

Although various types of mRNA-based vaccines have been explored, the optimal conditions for induction of both humoral and cellular immunity remain rather unknown. In this study, mRNA vaccines of nucleoside-modified mRNA in lipoplexes (LPXs) or lipid nanoparticles (LNPs) were evaluated after administration in mice through different routes, assessing mRNA delivery, tolerability and immunogenicity. In addition, we investigated whether mRNA vaccines could benefit from the inclusion of the adjuvant alpha-galactosylceramide (αGC), an invariant Natural Killer T (iNKT) cell ligand. Intramuscular (IM) vaccination with ovalbumin (OVA)-encoding mRNA encapsulated in LNPs adjuvanted with αGC showed the highest antibody- and CD8+ T cell responses. Furthermore, we observed that addition of signal peptides and endocytic sorting signals of either LAMP1 or HLA-B7 in the OVA-encoding mRNA sequence further enhanced CD8+ T cell activation although reducing the induction of IgG antibody responses. Moreover, mRNA LNPs with the ionizable lipidoid C12–200 exhibited higher pro-inflammatory- and reactogenic activity compared to mRNA LNPs with SM-102, correlating with increased T cell activation and antitumor potential. We also observed that αGC could further enhance the cellular immunity of clinically relevant mRNA LNP vaccines, thereby promoting therapeutic antitumor potential. Finally, a Listeria monocytogenes mRNA LNP vaccine supplemented with αGC showed synergistic protective effects against listeriosis, highlighting a key advantage of co-activating iNKT cells in antibacterial mRNA vaccines. Taken together, our study offers multiple insights for optimizing the design of mRNA vaccines for disease applications, such as cancer and intracellular bacterial infections. [Display omitted] [ABSTRACT FROM AUTHOR]

Scramblase Xk-related protein 8 (Xkr8) regulates the externalization of phosphatidylserine (PS) during apoptosis and holds a pivotal role in fostering tumor immunosuppression. Targeting Xkr8 in conjunction with chemotherapy demonstrated a novel avenue for amplifying antitumor immune response and overcoming chemo-immune resistance. Here we further evaluated this strategy by using a clinically relevant orthotopic model and elucidated the mechanism through in-depth single-cell RNA sequencing (scRNA-seq). We found that Xkr8 knockdown exhibited the potential to lead to immunogenic cell death (ICD) by impeding the normal clearance of apoptotic cells. Co-delivery of Xkr8 small interference RNA (siRNA) and a prodrug conjugate of 5-fluorouracil (5-Fu) and oxoplatin (FuOXP) showed remarkable therapeutic efficacy in an orthotopic pancreatic tumor model with increased infiltration of proliferative NK cells and activated macrophages in the tumor microenvironment (TME). Single-cell trajectory analysis further unveiled that tumor infiltrating CD8+ T cells are differentiated favorably to cytotoxic over exhausted phenotype after combination treatment. Our study sheds new light on the impact of Xkr8 knockdown on TME and solidifies the rationale of combining Xkr8 knockdown with chemotherapy to treat various types of cancers. [Display omitted] [ABSTRACT FROM AUTHOR]

Adoptive cell therapy with tumor-infiltrating T cells (TILs) has generated exciting clinical trial results for the treatment of unresectable solid tumors. However, solid tumors remain difficult targets for adoptively transferred T cells, due in part to poor migration of TILs to the tumor, physical barriers to infiltration, and active suppression of TILs by the tumor. Furthermore, a highly skilled team is required to obtain tumor tissue, isolate and expand the TILs ex vivo, and reinfuse them into the patient, which drives up costs and limits patient access. Here, we describe a cell-free polymer implant designed to recruit, genetically reprogram and expand host T cells at tumor lesions in situ. Importantly, the scaffold can be fabricated on a large scale and is stable to lyophilization. Using a mouse breast cancer model, we show that the implants quickly and efficiently amass cancer-specific host lymphocytes at the tumor site in quantities sufficient to bring about long-term tumor regression. Given that surgical care is the mainstay of cancer treatment for many patients, this technology could be easily implemented in a clinical setting as an add-on to surgery for solid tumors. Furthermore, the approach could be broadened to recruit and genetically reprogram other therapeutically desirable host cells, such as macrophages, natural killer cells or dendritic cells, potentially boosting the antitumor effectiveness of the implant even more. Our group developed an implantable device that (1) recruits T cells in situ, (2) genetically equips them with tumor-specific receptors, and (iii) rapidly expands and disperses them into solid tumor lesions. More specifically, the implant is based on a porous collagen scaffold (inherently supporting rapid T-cell migration), which is functionalized with a potent chemoattractant (CCL21) to recruit host T cells. To efficiently reprogram and expand recruited T cells within the scaffold, vectors encoding a tumor-specific receptor (chimeric antigen receptor in our study) and stimulatory anti- CD3/CD28 antibodies were co-encapsulated. All these components can be manufactured at large-scale and without the need to prepare a cellular product for each individual patient, and the final device can be frozen and stored until use. [Display omitted] • Rationally designed biomaterial scaffolds recruit, reprogram, expand and release human T cells. • Biomaterial scaffolds can efficiently transduce host T cells with tumor-specific CAR transgenes. • Scaffold-mediated in situ programming of CAR T-cells can improve survival of mice with established disease. [ABSTRACT FROM AUTHOR]

Lonicera japonica Thunb. (LJT) is known for its valuable medicinal properties that highlight its potential application in the pharmaceutical and health food industry. We predict that LJT polyphenols by network pharmacology may be involved in immunomodulation, and the study of LJT polyphenols regulating immunity is still insufficient; therefore, we experimentally found that LJT enhances immunity by promoting the proliferation and phagocytic activity of RAW246.7 cells. A model of an immunosuppressed mouse was constructed using cyclophosphamide‐induced, and LJT was extracted for the intervention. We found that LJT restored immune homeostasis in immune deficiency mice by inhibiting the abnormal apoptosis in lymphocytes, enhancing natural killer cell cytotoxicity, promoting T lymphocyte proliferation, and increasing the CD4+ and CD8+ T lymphocytes in quantity. Moreover, LJT treatment modulates immunity by significantly downregulating lipopolysaccharide‐induced inflammation and oxidative stress levels. We verified the immunomodulatory function of LJT through both cell and animal experiments. The combination of potential‐protein interactions and molecular docking later revealed that LJT polyphenols were associated with immunomodulatory effects on MAPK1; together, LJT intervention significantly modulates the immune, with the activation of MAPK1 as the underlying mechanism of action, which provided evidence for the utilization of LJT as a nutraceutical in immune function. [ABSTRACT FROM AUTHOR]

Management of lymphoid malignancies requires substantial health system resources. Total national health expenditure might influence population-based lymphoid malignancy survival. We studied the long-term survival of patients with 12 lymphoid malignancy types and examined whether different levels of national health expenditure might explain differences in lymphoid malignancy prognosis between European countries and regions. For this observational, retrospective, population-based study, we analysed the EUROCARE-6 dataset of patients aged 15 or older diagnosed between 2001 and 2013 with one of 12 lymphoid malignancies defined according to International Classification of Disease for Oncology (third edition) and WHO classification, and followed up to 2014 (Jan 1, 2001–Dec 31, 2014). Countries were classified according to their mean total national health expenditure quartile in 2001–13. For each lymphoid malignancy, 5-year and 10-year age-standardised relative survival (ASRS) was calculated using the period approach. Generalised linear models indicated the effects of age at diagnosis, gender, and total national health expenditure on the relative excess risk of death (RER). 82 cancer registries (61 regional and 21 national) from 27 European countries provided data eligible for 10-year survival estimates comprising 890 730 lymphoid malignancy cases diagnosed in 2001–13. Median follow-up time was 13 years (IQR 13–14). Of the 12 lymphoid malignancies, the 10-year ASRS in Europe was highest for hairy cell leukaemia (82·6% [95% CI 78·9–86·5) and Hodgkin lymphoma (79·3% [78·6–79·9]) and lowest for plasma cell neoplasms (29·5% [28·9–30·0]). RER increased with age at diagnosis, particularly from 55–64 years to 75 years or older, for all lymphoid malignancies. Women had higher ASRS than men for all lymphoid malignancies, except for precursor B, T, or natural killer cell, or not-otherwise specified lymphoblastic lymphoma or leukaemia. 10-year ASRS for each lymphoid malignancy was higher (and the RER lower) in countries in the highest national health expenditure quartile than in countries in the lowest quartile, with a decreasing pattern through quartiles for many lymphoid malignancies. 10-year ASRS for non-Hodgkin lymphoma, the most representative class for lymphoid malignancies based on the number of incident cases, was 59·3% (95% CI 58·7–60·0) in the first quartile, 57·6% (55·2–58·7) in the second quartile, 55·4% (54·3–56·5) in the third quartile, and 44·7% (43·6–45·8) in the fourth quartile; with reference to the European mean, the RER was 0·80 (95% CI 0·79–0·82) in the first, 0·91 (0·90–0·93) in the second, 0·94 (0·92–0·96) in the third, and 1·45 (1·42–1·48) in the fourth quartiles. Total national health expenditure is associated with geographical inequalities in lymphoid malignancy prognosis. Policy decisions on allocating economic resources and implementing evidence-based models of care are needed to reduce these differences. Italian Ministry of Health, European Commission, Estonian Research Council. [ABSTRACT FROM AUTHOR]

Effective diagnosis and understanding of the mechanism of intrapulmonary metastasis (IM) from multiple primary lung cancers (MPLC) aid clinical management. However, the actual detection panels used in the clinic are variable. Current research on tumor microenvironment (TME) of MPLC and IM is insufficient. Therefore, additional investigation into the differential diagnosis and discrepancies in TME between two conditions is crucial. Two hundred and fourteen non-small cell lung cancer patients with multiple tumors were enrolled and 507 samples were subjected to DNA sequencing (NGS 10). Then, DNA and RNA sequencing (master panel) were performed on the specimens from 32 patients, the TME profiles between tumors within each patient and across patients and the differentially expressed genes were compared. Four patients were regrouped with NGS 10 results. Master panel resolved the classifications of six undetermined patients. The TME in MPLC exhibited a high degree of infiltration by natural killer (NK) cells, CD56dim NK cells, endothelial cells, etc. P  < 0.05. Conversely, B cells, activated B cells, regulatory cells, immature dendritic cells, etc. P  < 0.001, were heavily infiltrated in the IM. NECTIN4 and LILRB4 mRNA were downregulated in the MPLC (P  < 0.0001). Additionally, NECTIN4 (P  < 0.05) and LILRB4 were linked to improved disease-free survival in the MPLC. In conclusion, IM is screened from MPLC by pathology joint NGS 10 detections, followed by a large NGS panel for indistinguishable patients. A superior prognosis of MPLC may be associated with an immune-activating TME and the downregulation of NECTIN4 and LILRB4 considered as potential drug therapeutic targets. [ABSTRACT FROM AUTHOR]