Search

Background and Objectives: Pancreatic ductal adenocarcinoma (PDAC) is an aggressive cancer with poor response to systemic therapies, including immunotherapy. Given the immunotherapeutic potential of natural killer (NK) cells, we evaluated intratumoral NK cell infiltrates along with cytotoxic T cells in PDAC to determine their association with patient outcomes., Methods: We analyzed tumors from 93 PDAC patients treated from 2012 to 2020. Predictor variables included tumor-infiltrating lymphocytes (TILs), T-cell markers (CD3, CD8, CD45RO), NK marker (NKp46), and NK inhibitory marker (major histocompatibility complex class I [MHC-I]) by immunohistochemistry. Primary outcome variables were recurrence-free survival (RFS) and overall survival (OS)., Results: Mean TILs, CD3, and NKp46 scores were 1.3 ± 0.63, 20.6 ± 17.5, and 3.1 ± 3.9, respectively. Higher expression of CD3 and CD8 was associated with higher OS, whereas NK cell infiltration was not associated with either RFS or OS. There was a tight positive correlation between MHC-I expression and all T-cell markers, but not with NKp46., Conclusions: Overall NK cell infiltrates were low in PDAC and did not predict clinical outcomes, whereas T-cell infiltrates did. Further characterization of the immune infiltrate in PDAC, including inhibitory signals and suppressive cell types, may yield better biomarkers of prognosis and immune targeting in this refractory disease., (© 2024 Wiley Periodicals LLC.)

Guillain-Barré syndrome (GBS) is a rare heterogenous disorder of the peripheral nervous system, which is usually triggered by a preceding infection, and causes a potentially life-threatening progressive muscle weakness 1 . Although GBS is considered an autoimmune disease, the mechanisms that underlie its distinct clinical subtypes remain largely unknown. Here, by combining in vitro T cell screening, single-cell RNA sequencing and T cell receptor (TCR) sequencing, we identify autoreactive memory CD4 + cells, that show a cytotoxic T helper 1 (T H 1)-like phenotype, and rare CD8 + T cells that target myelin antigens of the peripheral nerves in patients with the demyelinating disease variant. We characterized more than 1,000 autoreactive single T cell clones, which revealed a polyclonal TCR repertoire, short CDR3β lengths, preferential HLA-DR restrictions and recognition of immunodominant epitopes. We found that autoreactive TCRβ clonotypes were expanded in the blood of the same patient at distinct disease stages and, notably, that they were shared in the blood and the cerebrospinal fluid across different patients with GBS, but not in control individuals. Finally, we identified myelin-reactive T cells in the nerve biopsy from one patient, which indicates that these cells contribute directly to disease pathophysiology. Collectively, our data provide clear evidence of autoreactive T cell immunity in a subset of patients with GBS, and open new perspectives in the field of inflammatory peripheral neuropathies, with potential impact for biomedical applications., (© 2024. The Author(s).)

Giant cell arteritis (GCA) is a systemic vasculitis mediated by an aberrant immunological response against the blood vessel wall. Although the pathogenic mechanisms that drive GCA have not yet been elucidated, there is strong evidence that CD4 + T cells are key drivers of the inflammatory process occurring in this vasculitis. The aim of this study was to further delineate the role of CD4 + T cells in GCA by applying single-cell RNA sequencing and T cell receptor (TCR) repertoire profiling to 114.799 circulating CD4 + T cells from eight GCA patients in two different clinical states, active and in remission, and eight healthy controls. Our results revealed an expansion of cytotoxic CD4 + T lymphocytes (CTLs) in active GCA patients, which expressed higher levels of cytotoxic and chemotactic genes when compared to patients in remission and controls. Accordingly, differentially expressed genes in CTLs of active patients were enriched in pathways related to granzyme-mediated apoptosis, inflammation, and the recruitment of different immune cells, suggesting a role of this cell type in the inflammatory and vascular remodelling processes occurring in GCA. CTLs also exhibited a higher clonal expansion in active patients with respect to those in remission. Drug repurposing analysis prioritized maraviroc, which targeted CTLs, as potentially repositionable for this vasculitis. In addition, effector regulatory T cells (Tregs) were decreased in GCA and showed lower expression of genes involved in their suppressive activity. These findings provide further insights into the pathogenic role of CD4 + T cells in GCA and suggest targeting CTLs as a potential therapeutic option., Competing Interests: Declaration of competing interest None., (Copyright © 2023 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Actin beta-like 2 (ACTBL2) was recently identified as a new mediator of migration in ovarian cancer cells. Yet, its impact on tumor-infiltrating and thus migrating leukocytes (TILs) remains to date unknown. This study characterizes the subset of ACTBL2-expressing TILs in epithelial ovarian cancer (EOC) and elucidates their prognostic influence on the overall survival of EOC patients with special regard to different histological subtypes. Comprehensive immunohistochemical analyses of Tissue-Microarrays of 156 ovarian cancer patients revealed, that a tumor infiltration by ACTBL2-positive leukocytes was significantly associated with an improved overall survival (OS) (61.2 vs. 34.4 months; p = 0.006) and was identified as an independent prognostic factor (HR = 0.556; p = 0.038). This significant survival benefit was particularly evident in patients with low-grade serous carcinoma (OS: median not reached vs. 15.6 months, p < 0.001; HR = 0.058, p = 0.018). In the present cohort, ACTBL2-positive TILs were mainly composed of CD44-positive cytotoxic T-cells (CD8+) and macrophages (CD68+), as depicted by double-immunofluorescence and various immunohistochemical serial staining. Our results provide significant evidence of the prognostic impact and cellular composition of ACTBL2-expressing TILs in EOC. Complementary studies are required to analyze the underlying molecular mechanisms of ACTBL2 as a marker for activated migrating leukocytes and to further characterize its immunological impact on ovarian carcinogenesis., (© 2023. The Author(s).)

Photoactivated immunotherapy has promising therapeutic efficacy for treating malignancies, especially metastatic tumors. In this study, an erythrocyte membrane-encapsulated copper indium selenium (RCIS) semiconductor nanomaterial was developed to eliminate primary and metastatic tumors, in which copper ions can induce chemodynamic performance, and the narrow band gap endows RCIS with the properties of near-infrared (NIR) light-activated photothermal and photodynamic amplified immunotherapy. Furthermore, RCIS can be used as a nanocarrier to form RNCIS nanoparticles (NPs) by loading NLG919, which blocks the indoleamine 2,3-dioxygenase-1. Under NIR light irradiation, RNCIS NPs release NLG919 at tumor sites via photothermal properties, thereby promoting the recruitment of cytotoxic T lymphocytes and M1 polarization of macrophages, targeting the activation and amplification of immune responses. Herein, in vitro and in vivo studies showed that RNCIS NPs effectively kill cancer cells and eliminate primary and metastatic tumors. Therefore, this study suggests that semiconductor nanomaterials with narrow bandgaps have great potential as photoimmunotherapy agents and NIR light-responsive nanocarriers for controlled release, providing a great paradigm for synergetic tumor photoimmunotherapy. STATEMENT OF SIGNIFICANCE: The Erythrocyte membrane-coated, NLG919-loaded copper indium selenium (RNCIS) semiconductor was designed for eliminating primary and metastatic tumors. RNCIS exhibits chemodynamic, photodynamic, and photothermal activated immunotherapy by inhibiting indoleamine 2,3-dioxygenase-1. This can enhance the recruitment of cytotoxic T lymphocyte and M1 polarization of macrophage, leading to higher synergetic photo-immune therapeutic efficacy., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this article., (Copyright © 2023 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

The triggering receptor expressed on myeloid cell 1 (TREM1) plays a critical role in development of chronic inflammatory disorders and the inflamed tumor microenvironment (TME) associated with most solid tumors. We examined whether loss of TREM1 signaling can abrogate the immunosuppressive TME and enhance cancer immunity. To investigate the therapeutic potential of TREM1 in cancer, we used mice deficient in Trem1 and developed a novel small molecule TREM1 inhibitor, VJDT. We demonstrated that genetic or pharmacological TREM1 silencing significantly delayed tumor growth in murine melanoma (B16F10) and fibrosarcoma (MCA205) models. Single-cell RNA-Seq combined with functional assays during TREM1 deficiency revealed decreased immunosuppressive capacity of myeloid-derived suppressor cells (MDSCs) accompanied by expansion in cytotoxic CD8+ T cells and increased PD-1 expression. Furthermore, TREM1 inhibition enhanced the antitumorigenic effect of anti-PD-1 treatment, in part, by limiting MDSC frequency and abrogating T cell exhaustion. In patient-derived melanoma xenograft tumors, treatment with VJDT downregulated key oncogenic signaling pathways involved in cell proliferation, migration, and survival. Our work highlights the role of TREM1 in cancer progression, both intrinsically expressed in cancer cells and extrinsically in the TME. Thus, targeting TREM1 to modify an immunosuppressive TME and improve efficacy of immune checkpoint therapy represents what we believe to be a promising therapeutic approach to cancer.

Background: Acquired idiopathic generalized anhidrosis (AIGA) leads to heat intolerance due to the loss or reduction in thermoregulatory sweating over an extensive area of the body. The pathomechanism of AIGA is still unclear but is believed to be autoimmune., Objectives: We investigated the clinical and pathological features of inflammatory AIGA (InfAIGA) and noninflammatory AIGA (non-InfAIGA) within the skin., Methods: We compared anhidrotic and normohidrotic skin samples from 30 patients with InfAIGA and non-InfAIGA, as well as skin samples of melanocytic nevus as a negative control. We conducted morphometric analysis and immunohistochemical analysis of cell types and expression of inflammatory molecules (TIA1, CXCR3 and MxA). MxA expression was used as a proxy for type 1 interferon activity., Results: We found that tissue samples from patients with InfAIGA exhibited inflammation within the sweat duct and atrophy of the sweat coil, whereas patients with non-InfAIGA exhibited only atrophy of the sweat coil. Cytotoxic T lymphocyte infiltration and MxA expression were only observed in the sweat ducts of patients with InfAIGA., Conclusions: InfAIGA is associated with increased sweat duct inflammation and sweat coil atrophy, whereas non-InfAIGA is only associated with sweat coil atrophy. These data suggest that inflammation leads to epithelial destruction of sweat ducts associated with the sweat coil atrophy and subsequent loss of function. Non-InfAIGA may be regarded as a postinflammatory state of InfAIGA. These observations indicate the contribution of both type 1 and type 2 interferons to sweat gland injury. The mechanism involved is similar to the pathomechanism of alopecia areata (AA)., (© 2023 The Authors. Journal of the European Academy of Dermatology and Venereology published by John Wiley & Sons Ltd on behalf of European Academy of Dermatology and Venereology.)

Evasion from immunity is a major obstacle to the achievement of successful cancer immunotherapy. Hybrids derived from cell-cell fusion are theoretically associated with tumor heterogeneity and progression by conferring novel properties on tumor cells, including drug resistance and metastatic capacity; however, their impact on immune evasion remains unknown. Here, we investigated the potency of tumor-macrophage hybrids in immune evasion. Hybrids were established by co-culture of a melanoma cell line (A375 cells) and type 2 macrophages. The hybrids showed greater migration ability and greater tumorigenicity than the parental melanoma cells. The hybrids showed heterogeneous sensitivity to New York esophageal squamous cell carcinoma-1 (NY-ESO-1)-specific T-cell receptor-transduced T (TCR-T) cells and two out of four hybrid clones showed less sensitivity to TCR-T compared with the parental cells. An in vitro tumor heterogeneity model revealed that the TCR-T cells preferentially killed the parental cells compared with the hybrids and the survival rate of the hybrids was higher than that of the parental cells, indicating that the hybrids evade killing by TCR-T cells efficiently. Analysis of a single-cell RNA sequencing dataset of patients with melanoma revealed that a few macrophages expressed RNA encoding melanoma differentiation antigens including melan A, tyrosinase, and premelanosome protein, which indicated the presence of hybrids in primary melanoma. In addition, the number of potential hybrids was correlated with a poorer response to immune checkpoint blockade. These results provide evidence that melanoma-macrophage fusion has a role in tumor heterogeneity and immune evasion. © 2023 The Pathological Society of Great Britain and Ireland., (© 2023 The Pathological Society of Great Britain and Ireland.)

We present an individual-based model for the coevolutionary dynamics between CD8+ cytotoxic T lymphocytes (CTLs) and tumour cells. In this model, every cell is viewed as an individual agent whose phenotypic state is modelled by a discrete variable. For tumour cells, this variable represents a parameterization of the antigen expression profiles, while for CTLs it represents a parameterization of the target antigens of T-cell receptors (TCRs). We formally derive the deterministic continuum limit of this individual-based model, which comprises a non-local partial differential equation for the phenotype distribution of tumour cells coupled with an integro-differential equation for the phenotype distribution of CTLs. The biologically relevant homogeneous steady-state solutions of the continuum model equations are found. The linear-stability analysis of these steady-state solutions is then carried out in order to identify possible conditions on the model parameters that may lead to different outcomes of immune competition and to the emergence of patterns of phenotypic coevolution between tumour cells and CTLs. We report on computational results of the individual-based model, and show that there is a good agreement between them and analytical and numerical results of the continuum model. These results shed light on the way in which different parameters affect the coevolutionary dynamics between tumour cells and CTLs. Moreover, they support the idea that TCR-tumour antigen binding affinity may be a good intervention target for immunotherapy and offer a theoretical basis for the development of anti-cancer therapy aiming at engineering TCRs so as to shape their affinity for cancer targets., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Institute of Mathematics and its Applications. All rights reserved.)

Significance Statement: T-cell infiltration is a hallmark of crescentic GN (cGN), often caused by ANCA-associated vasculitis. Pathogenic T-cell subsets, their clonality, and downstream effector mechanisms leading to kidney injury remain to be fully elucidated. Single-cell RNA sequencing and T-cell receptor sequencing revealed activated, clonally expanded cytotoxic CD4 + and CD8 + T cells in kidneys from patients with ANCA-associated cGN. In experimental cGN, kidney-infiltrating CD8 + T cells expressed the cytotoxic molecule, granzyme B (GzmB), which induced apoptosis in renal tissue cells by activation of procaspase-3, and aggravated disease pathology. These findings describe a pathogenic function of (clonally expanded) cytotoxic T cells in cGN and identify GzmB as a mediator and potential therapeutic target in immune-mediated kidney disease., Background: Crescentic GN (cGN) is an aggressive form of immune-mediated kidney disease that is an important cause of end stage renal failure. Antineutrophilic cytoplasmic antibody (ANCA)-associated vasculitis is a common cause. T cells infiltrate the kidney in cGN, but their precise role in autoimmunity is not known., Methods: Combined single-cell RNA sequencing and single-cell T-cell receptor sequencing were conducted on CD3 + T cells isolated from renal biopsies and blood of patients with ANCA-associated cGN and from kidneys of mice with experimental cGN. Functional and histopathological analyses were performed with Cd8a-/- and GzmB-/- mice., Results: Single-cell analyses identified activated, clonally expanded CD8 + and CD4 + T cells with a cytotoxic gene expression profile in the kidneys of patients with ANCA-associated cGN. Clonally expanded CD8 + T cells expressed the cytotoxic molecule, granzyme B (GzmB), in the mouse model of cGN. Deficiency of CD8 + T cells or GzmB ameliorated the course of cGN. CD8 + T cells promoted macrophage infiltration and GzmB activated procaspase-3 in renal tissue cells, thereby increasing kidney injury., Conclusions: Clonally expanded cytotoxic T cells have a pathogenic function in immune-mediated kidney disease., (Copyright © 2023 by the American Society of Nephrology.)

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

Objective: Liver cancer is a deadly malignancy associated with high mortality and morbidity. Less than 20% of patients with advanced liver cancer respond to a single anti-PD-1 treatment. The high heterogeneity of neutrophils in the tumor immune microenvironment in liver cancer may contribute to resistance to immune checkpoint blockade (ICB). However, the underlying mechanism remains largely unknown., Methods: We established an orthotopic liver cancer model by using transposable elements to integrate the oncogenes Myc and Kras G12D into the genome in liver cells from conditional Trp53 null/null mice (pTMK/Trp53 -/- ). Flow cytometry and immunohistochemistry were used to assess the changes in immune cells in the tumor microenvironment. An ex vivo coculture assay was performed to test the inhibitory effects of tumor-associated neutrophils (TANs) on CD8 + T cells. The roles of neutrophils, T cells, and NK cells were validated through antibody-mediated depletion. The efficacy of the combination of neutrophil depletion and ICB was evaluated., Results: Orthotropic pTMK/Trp53 -/- mouse liver tumors displayed a moderate response to anti-Ly6G treatment but not PD-1 blockade. Depletion of neutrophils increased the infiltration of CD8 + T cells and decreased the number of exhausted T cells in the tumor microenvironment. Furthermore, depletion of either CD8 + T or NK cells abrogated the antitumor efficacy of anti-Ly6G treatment. Moreover, the combination of anti-Ly6G with anti-PD-L1 enhanced the infiltration of cytotoxic CD8 + T cells and thereafter resulted in a significantly greater decrease in tumor burden., Conclusions: Our data suggest that TANs may contribute to the resistance of liver cancer to ICB, and combining TAN depletion with T cell immunotherapy synergistically increases antitumor efficacy., Competing Interests: Ning Zhang is the CSO of Yunnan Baiyao Group. The other authors declare no competing interests., (Copyright © 2023 Cancer Biology & Medicine.)

The tumor microenvironment is a result of dynamic interaction between different cellular and non-cellular components. In its essence it is not a solo performer, but an ensemble of performers that includes cancer cells, fibroblasts, myo-fibroblasts, endothelial cells and immune cells. The short review highlights important immune infiltrates within the tumor microenvironment that shape cytotoxic t lymphocyte (CTL)-rich immune hot and CTL-deficient immune cold tumors and novel strategies that have potential role in enhancing our immune responses in both immune hot and immune cold tumors., (© 2023. The Author(s).)

Background: Effective anti-tumor immune responses are mediated by T cells and require organized, spatially coordinated interactions within the tumor microenvironment (TME). Understanding coordinated T-cell-behavior and deciphering mechanisms of radiotherapy resistance mediated by tumor stem cells will advance risk stratification of oropharyngeal cancer (OPSCC) patients treated with primary chemoradiotherapy (RCTx)., Methods: To determine the role of CD8 T cells (CTL) and tumor stem cells for response to RCTx, we employed multiplex immunofluorescence stains on pre-treatment biopsy specimens from 86 advanced OPSCC patients and correlated these quantitative data with clinical parameters. Multiplex stains were analyzed at the single-cell level using QuPath and spatial coordination of immune cells within the TME was explored using the R-package Spatstat., Results: Our observations demonstrate that a strong CTL-infiltration into the epithelial tumor compartment (HR for overall survival, OS: 0.35; p<0.001) and the expression of PD-L1 on CTL (HR: 0.36; p<0.001) were both associated with a significantly better response and survival upon RCTx. As expected, p16 expression was a strong predictor of improved OS (HR: 0.38; p=0.002) and correlated with overall CTL infiltration (r: 0.358, p<0.001). By contrast, tumor cell proliferative activity, expression of the tumor stem cell marker CD271 and overall CTL infiltration, regardless of the affected compartment, were not associated with response or survival., Conclusion: In this study, we could demonstrate the clinical relevance of the spatial organization and the phenotype of CD8 T cells within the TME. In particular, we found that the infiltration of CD8 T cells specifically into the tumor cell compartment was an independent predictive marker for response to chemoradiotherapy, which was strongly associated with p16 expression. Meanwhile, tumor cell proliferation and the expression of stem cell markers showed no independent prognostic effect for patients with primary RCTx and thus requires further study., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Haist, Kaufmann, Kur, Zimmer, Grabbe, Schmidberger, Weigert and Mayer.)

CD4 + T cells have the ability to differentiate into relatively specialized effector subsets after exposure to innate immune signals. The remarkable plasticity of CD4 + T cells is required to achieve immune responses in different tissues and against various pathogens. Numerous studies have shown that CD4 + T cells can play direct and indispensable roles in protective immunity by killing infected or transformed cells. Although the lineage decision of commitment to the CD4 + or CD8 + cell lineage is once thought to be inflexible, the identification of antigen-experienced CD4 + T cells with cytotoxic activity suggests the existence of unexpected plasticity for these cells. The recognition of CD4 + cytotoxic T lymphocytes (CTLs) and the mechanisms driving the differentiation of this particular cell subset create opportunities to explore the roles of these effector cells in protective immunity and immune-related pathology. CD4 + CTLs are proven to play a protective role in antiviral immunity. Here, the latest investigations on the phenotypic and functional features of CD4 + CTLs and their roles in antitumor immunity and immunotherapy are briefly reviewed., (© 2022 Wiley-VCH GmbH.)

Focal T lymphocyte aggregates commonly occur in colorectal cancer; however, their biological significance is unknown. To study focal aggregates of T lymphocytes, a deep learning-based framework for automated identification of T cell accumulations (T cell nests) was developed using CD8, PD-1, CD112R, and Ki67 multiplex fluorescence immunohistochemistry. To evaluate the clinical significance of these parameters, a cohort of 523 colorectal cancers with clinical follow-up data was analyzed. Spatial analysis of locally enriched CD8 + T cell density and cell-to-cell contacts identified T cell nests in the tumor microenvironment of colorectal cancer. CD112R and PD-1 expressions on CD8 + T cells located in T cell nests were found to be elevated compared with those on CD8 + T cells in all other tumor compartments (P < .001 each). Although the highest mean CD112R expression on CD8 + T cells was observed at the invasive margin, the PD-1 expression on CD8 + T cells was elevated in the center of the tumor (P < .001 each). Across all tissue compartments, proliferating CD8 + T cells showed higher relative CD112R and PD-1 expressions than those shown by non-proliferating CD8 + T cells (P < .001 each). Integration of all available spatial and immune checkpoint expression parameters revealed a superior predictive performance for overall survival (area under the curve, 0.65; 95% CI, 0.60-0.70) compared with the commonly used CD8 + tumor-infiltrating lymphocyte density (area under the curve, 0.57; 95% CI, 0.53-0.61; P < .001). Cytotoxic T cells with elevated CD112R and PD-1 expression levels are orchestrated in T cell nests of colorectal cancer and predict favorable patient outcomes, and the spatial nonredundancy underlies fundamental differences between both inhibitory immune checkpoints that provide a rationale for dual anti-CD112R/PD-1 immune checkpoint therapy., (Copyright © 2022 The Authors. Published by Elsevier Inc. All rights reserved.)

Backgroud: Immune impairment, marked by increased expression of cytotoxic T lymphocyte antigen (CTLA)-4, promotes the disease progression of chronic hepatitis B., Objective: This study aimed to determine the association between serum CTLA-4 level and disease progression in patients with chronic hepatitis B., Methods: A cross-sectional study was conducted at Haji Adam Malik General Hospital Medan, Indonesia between October 2021 to September 2022. A total of 150 participants were enrolled. Patients aged 18 years or older with evidence of chronic hepatitis B, HBV-related liver cirrhosis, and HBV-related hepatocellular carcinoma (HCC) were enrolled. Exclusion criteria were history of chronic hepatotoxic drug consumption, underlying liver abnormalities other than HBV infection, and liver injury due to metastasized malignancy from other sites. Serum CTLA-4 level was determined from serum using human CTLA-4 enzyme linked immunosorbent assay kit., Results: Most participants were males and aged between 40 and 60 years. Serum CTLA-4 level was positively associated with chronic hepatitis B progression (P<0.001). Serum CTLA-4 level was negatively correlated with serum platelet (P<0.001) and albumin levels (P<0.001) but positively correlated with serum ALT (P=0.045) and total bilirubin levels (P<0.001)., Conclusions: Serum CTLA-4 level is associated with disease progression in patients with chronic hepatitis B., Competing Interests: authors declare that there are no conflicts of interest., (© 2023 Darmadi Darmadi, Dharma Lindarto, Jelita Siregar, Tri Widyawati, Muhammad Rusda, Mustafa Mahmud Amin, Fauzi Yusuf, Putri Chairani Eyanoer, Masrul Lubis, Imelda Rey.)

Background/purpose: The pathogenesis of chronic actinic dermatitis (CAD) is more complicated than other photodermatoses. However, the relationship between the clinical severity of CAD and the offending photocontact or contact allergens or both, and the correlations of CAD immunopathogenesis with the immunoregulatory molecules involved in adaptive immunity are yet to be investigated., Methods: We performed phototesting with broad-spectrum ultraviolet (UV) B, UVA, and visible light to establish the presence of photosensitivity in 121 patients with CAD, together with photopatch and contact patch testing. Nine patients with CAD were selected according to their clinical severity score for CAD (CSS-CAD), and triple direct immunofluorescence analysis was performed with paraffin-embedded skin biopsy samples., Results: As CSS-CAD was closely correlated with the multiplicity of photo(contact) allergens, particularly photoallergens, three or more photoallergens were detected in the severe CAD group (52.5%); less in the moderate group (32.8%); and only one in the mild group (14.8%; P = .025). In the groups showing greater severity of disease, the absolute numbers of IFN-γ + , IL-17 + , CD4+, CD8+, common-γ chain receptor (common-γCR) + , and CD69 + tissue-resident memory cells increased on average; there was also an increase in the CD4+/CD8+ cell ratio, with the more severely affected groups. However, the levels of TNF-α + and FoxP3 + regulatory T (Treg) cells and the mean IL-17/IFN-γ cell ratio decreased in the more severely affected CSS-CAD subgroups., Conclusions: Based on the clinical analysis and immunopathogenic results, avoidance of excessive sun exposure, and topical and systemic blocking agents for photo(contact) allergens are recommended. Additionally, conventional immunomodulators and emerging agents including JAK-STAT inhibitors may be administered for CAD treatment in the future., (© 2022 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Human papillomavirus (HPV)-driven oropharyngeal carcinoma (OPSCC) is distinct from tobacco- or alcohol-associated OPSCC and has a unique immune landscape. Studies have supported the heterogeneity of T cells, accompanied by a broad repertoire of T-cell responses, within tumors driven by HPV infection. However, the phenotype and function of these HPV-related T cells remain unclear. Using a combination of single-cell RNA sequencing, flow cytometry, pharmacologic inhibition, and immunofluorescence staining, we explored the prognostic implication of HPV-related T cells and further validated our findings in two independent cohorts. Cytotoxic T lymphocytes (CTL) within OPSCC displayed a spectrum of transcriptional signatures. Among which, we identified CD161 receptor, encoded by KLRB1, as a potential marker to distinguish the CTL subsets in HPV-positive OPSCC with a divergent evolutionary trajectory. In-depth analysis revealed that CD161+ CTLs exhibited a more robust immune response over the CD161- counterparts and a T cell-inflamed phenotype that could be further reinvigorated by immune-checkpoint blockade. Despite the high expression of exhaustion markers, reinforcement of CD161+ CTL reactivity was expected to boost immune responses, considering their functional reversibility. We further confirmed that the high level of intratumoral CD161+ CTLs associated with a favorable treatment response and prolonged overall survival. Therefore, our research not only provides an insight into the immune landscape of HPV-driven OPSCC but also sheds light on a special subset of CTLs with prognostic and therapeutic significance., (©2023 The Authors; Published by the American Association for Cancer Research.)

Extracellular deposition of amyloid-β as neuritic plaques and intracellular accumulation of hyperphosphorylated, aggregated tau as neurofibrillary tangles are two of the characteristic hallmarks of Alzheimer's disease 1,2 . The regional progression of brain atrophy in Alzheimer's disease highly correlates with tau accumulation but not amyloid deposition 3-5 , and the mechanisms of tau-mediated neurodegeneration remain elusive. Innate immune responses represent a common pathway for the initiation and progression of some neurodegenerative diseases. So far, little is known about the extent or role of the adaptive immune response and its interaction with the innate immune response in the presence of amyloid-β or tau pathology 6 . Here we systematically compared the immunological milieux in the brain of mice with amyloid deposition or tau aggregation and neurodegeneration. We found that mice with tauopathy but not those with amyloid deposition developed a unique innate and adaptive immune response and that depletion of microglia or T cells blocked tau-mediated neurodegeneration. Numbers of T cells, especially those of cytotoxic T cells, were markedly increased in areas with tau pathology in mice with tauopathy and in the Alzheimer's disease brain. T cell numbers correlated with the extent of neuronal loss, and the cells dynamically transformed their cellular characteristics from activated to exhausted states along with unique TCR clonal expansion. Inhibition of interferon-γ and PDCD1 signalling both significantly ameliorated brain atrophy. Our results thus reveal a tauopathy- and neurodegeneration-related immune hub involving activated microglia and T cell responses, which could serve as therapeutic targets for preventing neurodegeneration in Alzheimer's disease and primary tauopathies., (© 2023. The Author(s), under exclusive licence to Springer Nature Limited.)

Background/aim: Melanoma is a skin cancer without effective therapy, showing high immunogenicity and mostly partial spontaneous regression (SR). The exact mechanisms of SR are still not well understood; therefore, the use of animal melanoma models is necessary to unravel the immunological processes during SR., Materials and Methods: Skin melanoma samples (n=57) and peripheral blood samples (n=57) from the same animals were collected. Melanoma-bearing Libechov Minipigs (MeLiM) aged 3, 4, 6, 8, 10, 12, 20, and 32 weeks were used, and samples were analysed by flow cytometry for detection of immune cell subpopulations., Results: The proportion of CD3-CD8+ (NK) cells in melanoma samples was found to be higher compared to blood samples at 6-8 weeks of age and then at 12 weeks of age. The population of CD4+CD8+ (effector/memory T helper) cells and CD4-CD8+ (cytotoxic T and NK) cells was also increased in melanoma compared to blood samples in 10-32-week-old pigs. The proportion of CD4-CD8+ cells in melanoma samples, then augmented until the 32 nd week. On the contrary, the proportion of CD4+CD8- (naive T helper) cells was lower in melanoma samples versus blood samples in 6-32-week-old animals., Conclusion: Cytotoxic T cells were the most abundant population of tumour infiltrating immune cells found in MeLiM melanomas of animals aged 10-32 weeks, probably causing the destruction of melanoma cells. Furthermore, the development of specific (adaptive) immune response represented mainly by cytotoxic T cells seems to be crucial for the successful SR of porcine melanoma., (Copyright © 2023 International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.)

Purpose: Cisplatin-containing combination chemotherapy has been the standard of care since the late 1980s, but the response rate is <50%. Studies have shown that the efficiency of chemotherapy differs among molecular subtypes of bladder cancer. In this study, we aimed to correlate FOXA1, a marker for differentiation of the basal and luminal subtypes, with tumor immune cell infiltration and the effect of chemotherapy in bladder cancer., Materials and Methods: Eighty-three patients with bladder cancer treated with chemotherapy were reviewed. Clinicopathological variables for each case were recorded. FOXA1, M2 tumor-associated macrophage (TAM), dendritic cell (DC), and cytotoxic T lymphocyte (CTL) were examined by immunohistochemistry. The relationship between FOXA1, immune cell infiltration, and clinical response to chemotherapy was assessed., Results: The overall objective response rate was 34%. The objective response rate for tumors with lower FOXA1 expression was 58% and for tumors with higher FOXA1 expression was 12%. Tumors with infiltrated M2 TAM proportion <3% had a higher objective response rate compared with infiltrated M2 TAM proportion >3% tumors (46% vs. 21%, p = 0.02). Tumors with infiltrated CTL proportion >5% had a higher objective response rate compared with infiltrated CTL proportion <5% tumors (50% vs. 17%, p = 0.002). DCs showed no significant differences. We found that the objective response rate for tumors with lower FOXA1 expression, proportion <3% M2 TAM infiltration, and proportion >5% CTL infiltration is 82%. Lower FOXA1 expression was associated with low M2 TAM infiltration and high CTL infiltration., Conclusions: Thus, we showed that in patients with bladder cancer who received chemotherapy, the higher clinical response rate is associated with low FOXA1 expression, low M2 TAM infiltration, and high CTL infiltration., (© 2021 The Author(s) Published by S. Karger AG, Basel.)

Our understanding of the molecular mechanisms underlying postsurgical recurrence of non-small cell lung cancer (NSCLC) is rudimentary. Molecular and T cell repertoire intratumor heterogeneity (ITH) have been reported to be associated with postsurgical relapse; however, how ITH at the cellular level impacts survival is largely unknown. Here we report the analysis of 2880 multispectral images representing 14.2% to 27% of tumor areas from 33 patients with stage I NSCLC, including 17 cases (relapsed within 3 years after surgery) and 16 controls (without recurrence ≥5 years after surgery) using multiplex immunofluorescence. Spatial analysis was conducted to quantify the minimum distance between different cell types and immune cell infiltration around malignant cells. Immune ITH was defined as the variance of immune cells from 3 intratumor regions. We found that tumors from patients having relapsed display different immune biology compared with nonrecurrent tumors, with a higher percentage of tumor cells and macrophages expressing PD-L1 (P =.031 and P =.024, respectively), along with an increase in regulatory T cells (Treg) (P =.018), antigen-experienced T cells (P =.025), and effector-memory T cells (P =.041). Spatial analysis revealed that a higher level of infiltration of PD-L1 + macrophages (CD68 + PD-L1 + ) or antigen-experienced cytotoxic T cells (CD3 + CD8 + PD-1 + ) in the tumor was associated with poor overall survival (P =.021 and P =.006, respectively). A higher degree of Treg ITH was associated with inferior recurrence-free survival regardless of tumor mutational burden (P =.022), neoantigen burden (P =.021), genomic ITH (P =.012) and T cell repertoire ITH (P =.001). Using multiregion multiplex immunofluorescence, we characterized ITH at the immune cell level along with whole exome and T cell repertoire sequencing from the same tumor regions. This approach highlights the role of immunoregulatory and coinhibitory signals as well as their spatial distribution and ITH that define the hallmarks of tumor relapse of stage I NSCLC., (Copyright © 2022 United States & Canadian Academy of Pathology. Published by Elsevier Inc. All rights reserved.)

Objective: Asthma is a chronic lung disease comprising multiple endotypes and characterized by periodic exacerbations. A diverse array of T cells has been found to contribute to all endotypes of asthma in pathogenic and regulatory roles. Here, we review the contributions of CD4+, CD8+, and unconventional T cells in allergic and nonallergic asthma., Data Sources: Review of published literature pertaining to conventional and unconventional T-cell types in asthma., Study Selections: Recent peer-reviewed articles pertaining to T cells in asthma, with additional peer-reviewed studies for context., Results: Much research in asthma has focused on the roles of CD4+ T H cells. Roles for T H 2 cells in promoting allergic asthma pathogenesis have been well-described, and the recent description of pathogenic T H 2A cells provides additional insight into these responses. Other T H types, notably T H 1 and T H 17, have been linked to neutrophilic and steroid-resistant asthma phenotypes. Beyond CD4+ T cells, CD8+ Tc2 cells are also strongly associated with allergic asthma. An emerging area for study is unconventional T-cell types, including γδT, invariant natural killer T, and mucosal-associated invariant T cells. Although data in asthma remain limited for these cells, their ability to bridge innate and adaptive responses likely makes them key players in asthma. A number of asthma therapies target T-cell responses, and, although data are limited, they seem to modulate T-cell populations., Conclusion: Given the diversity and heterogeneity of asthma and T-cell responses, there remain many rich avenues for research to better understand the pathogenesis of asthma. Despite the breadth of T cells in asthma, approved therapeutics remain limited to T H 2 networks., (Copyright © 2022 American College of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: Lymphocytic thyroiditis (LT) is frequently seen in the tumor microenvironment (TME) of papillary thyroid carcinomas (PTCs). However, the characteristic of these tumor-infiltrating lymphocytes (TILs) is not well understood., Objective: We aim to define the TME of PTC cases by characterizing the TILs., Design: This is a cross-sectional observational study., Patients: We enrolled 29 PTC (23 having concurrent LT), 14 LT, and 13 hyperplastic nodules with LT (HN) patients from January 2016 to December 2020., Measurements: Immunohistochemical (IHC) expression of CD8, FoxP3, PD-1, and PD-L1 was studied in PTC with LT and compared with HN. PD-1 and PD-L1 expression was correlated at the mRNA level by quantitative real-time PCR. Immunophenotyping of TILs was done in FNAC samples of PTC and LT by flow cytometry., Results: IHC revealed the presence of CD8 + cytotoxic T lymphocytes (CTLs) and FoxP3 + T regulatory cells (Tregs) in 83% and 52% of PTC with LT cases, respectively. Flow cytometric analysis of the PTC samples revealed a significant abundance of CTL compared with Treg and a higher CTL with lower Treg counts compared with LT. On IHC, PD-1 positivity was noted in 56.5% of PTC with LT cases, while intermediate PD-L1 positivity was found in 70% of the cases. There was a significant upregulation of PD-1 mRNA in PTC with LT. A significant correlation was noted with PD-L1 expression with lymph node metastasis and presence of Treg cells., Conclusions: Increased expression of PD-1 and PD-L1 in the TME of PTC may provide a potential molecular mechanism for tumor survival despite the predominance of CTLs, possibly through their inactivation or exhaustion., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Banerjee, Nahar, Dahiya, Mukherjee, Dey, Gupta, Radotra, Sachdeva, Sood, Bhadada and Bhansali.)

Adaptive immune cells prevent solid tumor progression by targeting and killing tumor cells. However, there are no comprehensive studies on peripheral circulating adaptive immune cell characterization in colorectal cancer (CRC) patients or the effect of tumor-node-metastasis (TNM) stages on these cells. In this study, the number, phenotype, and function of different subsets of circulating adaptive immune cells in peripheral blood of CRC patients were analyzed. We found remarkable differences in CRC patients compared with those in healthy controls, including reduced absolute counts of total T cells, helper T lymphocytes (Th), cytotoxic T lymphocytes (Tc), and double-negative T lymphocytes, a decreased proportion of INF-γ + cells in total T cells and Th, and increased percentages of B cells, plasmablasts, and activated T cells. Compared with early-stage CRC patients, advanced-stage CRC patients showed more severe immunosenescence, which manifested as decreased proportions of CD8 + naive T cells with strong proliferative ability and CD8 + central memory T cells with immune surveillance function. Proportions and absolute counts of CD8 + and CD4 + terminally differentiated effector memory T cells were increased, indicating immunosenescence. The immune cell characteristics analyzed in this study serve as a starting point for further research to determine potential clinical implications., (© 2022. The Author(s).)

Sjögren's syndrome is a chronic autoimmune disorder whose pathogenesis is poorly understood and that lacks effective therapies. Detailed quantitative and spatial analyses of tissues affected by Sjögren's syndrome were undertaken, including the quantitation of the frequency of selected cell-cell interactions in the disease milieu. Quantitative analyses of CD4 + T cell subsets and of CD8 + T cells in the labial salivary glands from untreated patients with primary Sjögren's syndrome revealed that activated CD8 + cytotoxic T cells (CD8 + CTLs) were the most prominent T cells in these infiltrates. An accumulation of apoptotic glandular epithelial cells, mainly ductal and acinar cells, was observed, consistent with the impaired salivary secretion often observed in patients with this disease. FasL expressing activated CD8 + T cells were seen to accumulate around Fas expressing apoptotic epithelial cells. Quantitative analyses of apoptotic cell types and of conjugates between cytotoxic T cells and epithelial cells undergoing apoptosis suggest that Sjögren's syndrome is primarily driven by CD8 + CTL mediated execution of epithelial cells mainly represented by ductal and acinar cells., (© 2022. The Author(s).)

The CD169 + macrophages in lymph nodes are implicated in cytotoxic T lymphocyte (CTL) activation and are associated with improved prognosis in several malignancies. Here, we investigated the significance of CD169 + macrophages in oral squamous cell carcinoma (OSCC). Further, we tested the anti-tumor effects of naringenin, which has been previously shown to activate CD169 + macrophages, in a murine OSCC model. Immunohistochemical analysis for CD169 and CD8 was performed on lymph node and primary tumor specimens from 89 patients with OSCC. We also evaluated the effects of naringenin on two murine OSCC models. Increased CD169 + macrophage counts in the regional lymph nodes correlated with favorable prognosis and CD8 + cell counts within tumor sites. Additionally, naringenin suppressed tumor growth in two murine OSCC models. The mRNA levels of CD169, interleukin (IL)-12, and C-X-C motif chemokine ligand 10 (CXCL10) in lymph nodes and CTL infiltration in tumors significantly increased following naringenin administration in tumor-bearing mice. These results suggest that CD169 + macrophages in lymph nodes are involved in T cell-mediated anti-tumor immunity and could be a prognostic marker for patients with OSCC. Moreover, naringenin is a new potential agent for CD169 + macrophage activation in OSCC treatment., (© 2022. The Author(s).)

An approach of decorating bacteria with triple immune nanoactivators is reported to develop tumor-resident living immunotherapeutics. Under cytocompatible conditions, tumor-specific antigens and checkpoint blocking antibodies are simultaneously conjugated onto bacterial surface and then polydopamine nanoparticles are formed via in situ dopamine polymerization. In addition to serving as a linker, polydopamine with its photothermal effect can repolarize tumor-associated macrophages to a pro-inflammatory phenotype. The linked antigens promote the maturation of dendritic cells and generate tumor-specific immune responses, while the anchored antibodies block immune checkpoints and activate cytotoxic T lymphocytes. Decorated bacteria show spatiotemporal tumor retention and proliferation-dependent drug release, achieving potent antitumor effects in two antigen-overexpressing tumor models. This work provides a versatile platform to prepare multimodal and long-acting therapeutics for cancer immunotherapy., (© 2022 Wiley-VCH GmbH.)

Immunosuppressive elements within the tumor microenvironment, such as tumor-associated macrophages (TAM), can present a barrier to successful antitumor responses by cytolytic T cells. Here we employed preclinical syngeneic p53 null mouse models of triple-negative breast cancer (TNBC) to develop a treatment regimen that harnessed the immunostimulatory effects of low-dose cyclophosphamide coupled with the pharmacologic inhibition of TAMs using either a small-molecule CSF1R inhibitor or an anti-CSF1R antibody. This therapeutic combination was effective in treating several highly aggressive TNBC murine mammary tumor and lung metastasis models. Single-cell RNA sequencing characterized tumor-infiltrating lymphocytes including Th cells and antigen-presenting B cells that were highly enriched in responders to combination therapy. In one model that exhibited long-term posttreatment tumor regression, high-dimensional imaging techniques identified the close spatial localization of B220+/CD86+-activated B cells and CD4+ T cells in tertiary lymphoid structures that were present up to 6 weeks posttreatment. The transcriptional and metabolic heterogeneity of TAMs was also characterized in two closely related claudin-low/mesenchymal subtype tumor models with differential treatment responses. A murine TAM signature derived from the T12 model was highly conserved in human claudin-low breast cancers, and high expression of the TAM signature correlated with reduced overall survival in patients with breast cancer. This TAM signature may help identify human patients with claudin-low breast cancer that will benefit from the combination of cyclophosphamide and anti-CSF1R therapy. These studies illustrate the complexity of the tumor immune microenvironment and highlight different immune responses that result from rational immunotherapy combinations., Significance: Immunostimulatory chemotherapy combined with pharmacologic inhibition of TAMs results in durable treatment responses elicited by Th cells and B cells in claudin-low TNBC models., (©2022 American Association for Cancer Research.)

Background: Receptor-interacting protein kinase 2 (RIPK2, also known as RIP2) was reported to be associated with bacterial infections as well as inflammatory responses. However, the role of RIPK2 in prognosis and immunotherapy response is yet to be elucidated in human pan-cancer., Methods: In this study, we investigated the expression, gene alteration landscape and prognostic value of RIPK2 in 33 cancers through various databases including Ualcan, cBioportal and Gene Expression Profiling Interactive Analysis 2 (GEPIA2). Then, the correlation between RIPK2 and immune infiltration, immune score, stromal score, and ESTIMATE score was investigated in the Cancer Genome Atlas (TCGA) and tumor immune estimation resource (TIMER) databases. Independent cohorts were utilized to explore the role of RIPK2 in tumor immunotherapy response. Furthermore, Gene set enrichment analysis (GSEA) was conducted to explore the mechanisms by which RIPK2 regulates immune therapy resistance. Single-cell RNA-seq datasets were used to analyze the expression level of RIPK2 on different immune cells. Moreover, CellMiner database was used to explore the relationship between RIPK2 expression with drug response., Result: Compared with normal tissue, tumor tissue had a higher expression level of RIPK2 in various cancers. Survival analysis showed that high expression of RIPK2 associated with poor prognosis in numerous cancers. RIPK2 was found to promote a series of immune cell infiltration and B cells, macrophages, and neutrophils were significantly positively correlated with the expression of RIPK2. Moreover, RIPK2 affected immune score, stromal score and ESTIMATE score for a wide range of cancers. In the vast majority of 33 cancers, gene co-expression analysis showed that RIPK2 was positively correlated with the expression of immune checkpoint markers, such as PDCD1 (PD-1), CD274 (PD-L1), CTLA4 and TIGIT. RIPK2 aggravated cytotoxic T lymphocyte (CTL) dysfunction and related to the poor efficacy of immune checkpoint blockade in skin cutaneous melanoma (SKCM) and kidney renal clear cell carcinoma (KIRC). High expression of RIPK2 promoted innate immunotherapy resistance and adaptive immunotherapy resistance through IL-6/JAK/STAT3 signaling, interferon-gamma response, and interferon-alpha response pathway., Conclusions: These results confirmed that RIPK2 could serve as a prognostic biomarker and promoted immune therapy resistance via triggering cytotoxic T lymphocytes dysfunction., (© 2022. The Author(s).)

Aims: Angioimmunoblastic T-cell lymphoma (AITL) is genetically characterized by TET2 and DNMT3A mutations occurring in haematopoietic progenitor cells, and late events (e.g. the RHOA-G17V mutation) associated with malignant transformation. As TET2/DNMT3A-mutated progenitor cells can differentiate into multilineage progenies and give rise to both AITL and myeloid neoplasms, they may also have the potential to lead to other metachronous/synchronous neoplasms. We report two cases showing parallel evolution of two distinct potentially neoplastic lymphoid proliferations from a common mutated haematopoietic progenitor cell population., Methods and Results: Both cases presented with generalized lymphadenopathy. In case 1 (a 67-year-old female), an initial lymph node (LN) biopsy was dismissed as reactive, but a repeat biopsy showed a nodal marginal zone lymphoma (NMZL)-like proliferation with an increase in the number of T-follicular helper (TFH) cells. Immunohistochemistry, and clonality and mutational analyses by targeted sequencing of both whole tissue sections and microdissected NMZL-like lesions, demonstrated a clonal B-cell proliferation that harboured the BRAF-G469R mutation and shared TET2 and DNMT3A mutations with an underlying RHOA-G17V-mutant TFH proliferation. Review of the original LN biopsy showed histological and immunophenotypic features of AITL. In case 2 (a 66-year-old male), cytotoxic T-cell lymphoma with an increase in the number of Epstein-Barr virus-positive large B cells was diagnosed on initial biopsy. On review together with the relapsed biopsy, we identified an additional occult neoplastic TFH proliferation/smouldering AITL. Both T-cell proliferations shared TET2 and DNMT3A mutations while RHOA-G17V was confined to the smouldering AITL., Conclusions: In addition to demonstrating diagnostic challenges, these cases expand the potential of clonal haematopoiesis in the development of different lineage neoplastic proliferations., (© 2022 The Authors. Histopathology published by John Wiley & Sons Ltd.)

Purpose: Patients with glioblastoma (GBM) are treated with radiotherapy (RT) and temozolomide (TMZ). These treatments may cause prolonged systemic lymphopenia, which itself is associated with poor outcomes. NT-I7 is a long-acting IL7 that expands CD4 and CD8 T-cell numbers in humans and mice. We tested whether NT-I7 prevents systemic lymphopenia and improves survival in mouse models of GBM., Experimental Design: C57BL/6 mice bearing intracranial tumors (GL261 or CT2A) were treated with RT (1.8 Gy/day × 5 days), TMZ (33 mg/kg/day × 5 days), and/or NT-I7 (10 mg/kg on the final day of RT). We followed the mice for survival while serially analyzing levels of circulating T lymphocytes. We assessed regulatory T cells (Treg) and cytotoxic T lymphocytes in the tumor microenvironment, cervical lymph nodes, spleen, and thymus, and hematopoietic stem and progenitor cells in the bone marrow., Results: GBM tumor-bearing mice treated with RT+NT-I7 increased T lymphocytes in the lymph nodes, thymus, and spleen, enhanced IFNγ production, and decreased Tregs in the tumor which was associated with a significant increase in survival. NT-I7 also enhanced central memory and effector memory CD8 T cells in lymphoid organs and tumor. Depleting CD8 T cells abrogated the effects of NT-I7. Furthermore, NT-I7 treatment decreased progenitor cells in the bone marrow., Conclusions: In orthotopic glioma-bearing mice, NT-I7 mitigates RT-related lymphopenia, increases cytotoxic CD8 T lymphocytes systemically and in the tumor, and improves survival. A phase I/II trial to evaluate NT-I7 in patients with high-grade gliomas is ongoing (NCT03687957)., (©2022 American Association for Cancer Research.)

Immunotherapeutic treatment approaches are now an integral part of the treatment of many solid tumors. However, attempts to integrate immunotherapy into the treatment of prostate cancer have been disappointing so far. This is due to a highly immunosuppressive, "cold" tumor microenvironment, which is characterized, for example, by the absence of cytotoxic T cells, an increased number of myeloid-derived suppressor cells or regulatory T cells, a decreased number of tumor antigens, or a defect in antigen presentation. The consequence is a reduced efficacy of many established immunotherapeutic treatments such as checkpoint inhibitors. However, a growing understanding of the underlying mechanisms of tumor-immune system interactions raises hopes that immunotherapeutic strategies can be optimized in the future. The aim of this review is to provide an overview of the current status and future directions of immunotherapy development in prostate cancer. Background information on immune response and tumor microenvironment will help to better understand current therapeutic strategies under preclinical and clinical development.

Muscle cell death in polymyositis is induced by CD8 + cytotoxic T lymphocytes. We hypothesized that the injured muscle fibers release pro-inflammatory molecules, which would further accelerate CD8 + cytotoxic T lymphocytes-induced muscle injury, and inhibition of the cell death of muscle fibers could be a novel therapeutic strategy to suppress both muscle injury and inflammation in polymyositis. Here, we show that the pattern of cell death of muscle fibers in polymyositis is FAS ligand-dependent necroptosis, while that of satellite cells and myoblasts is perforin 1/granzyme B-dependent apoptosis, using human muscle biopsy specimens of polymyositis patients and models of polymyositis in vitro and in vivo. Inhibition of necroptosis suppresses not only CD8 + cytotoxic T lymphocytes-induced cell death of myotubes but also the release of inflammatory molecules including HMGB1. Treatment with a necroptosis inhibitor or anti-HMGB1 antibodies ameliorates myositis-induced muscle weakness as well as muscle cell death and inflammation in the muscles. Thus, targeting necroptosis in muscle cells is a promising strategy for treating polymyositis providing an alternative to current therapies directed at leukocytes., (© 2022. The Author(s).)

Background: A limited number of studies have characterized genomic properties of hepatocellular carcinoma (HCC) patients in response to anti-PD-1 immunotherapy., Methods: Herein, we performed comprehensive molecular characterization of immediate (D-42 to D-1) pre-treatment tumor biopsy specimens from 60 patients with sorafenib-failed HCC in a single-arm prospective phase II trial of pembrolizumab. Objective response rate was the primary efficacy endpoint. We used whole-exome sequencing, RNA sequencing, and correlative analysis. In addition, we performed single-cell RNA sequencing using peripheral blood mononuclear cells., Results: The overall response rate of pembrolizumab in sorafenib-failed HCC patients was 10% ([6/60] 95% CI, 2.4-17.6). In a univariate analysis using clinicopathological features, female gender, PD-L1 positivity, and low neutrophil-to-lymphocyte ratio (NLR) were identified as contributing factors to pembrolizumab response. Somatic mutations in CTNNB1 and genomic amplifications in MET were found only in non-responders. Transcriptional profiles through RNA sequencing identified that pembrolizumab responders demonstrated T cell receptor (TCR) signaling activation with expressions of MHC genes, indicating increased levels of T cell cytotoxicity. In single-cell sequencing from 10 pre- and post-treatment peripheral blood mononuclear cells (PBMCs), patients who achieved a partial response or stable disease exhibited immunological shifts toward cytotoxic CD8+ T cells. Conversely, patients with progressive disease showed an increased number of both CD14+ and CD16+ monocytes and activation of neutrophil-associated pathways., Conclusions: Taken together, HCC patients with infiltration of cytotoxic T cells, along with increased active circulating CD8+ T cells during pembrolizumab treatment and down-regulation of neutrophil-associated markers, significantly benefited from pembrolizumab treatment., Trial Registration: NCT#03163992 (first posted: May 23, 2017)., (© 2021. The Author(s).)

Hemophagocytic lymphohistiocytosis (HLH) is a hyperinflammatory disorder characterized by the inability to properly terminate an immune response. Familial HLH (FHLH) and related immune dysregulation syndromes are associated with mutations in the genes PRF1, UNC13D, STX11, STXBP2, LYST, AP3B1 , and RAB27A , all of which are required for the assembly, exocytosis, and function of cytotoxic granules within CD8+ T cells and natural killer (NK) cells. Loss-of-function mutations in these genes render the cytotoxicity pathway ineffective, thereby failing to eradicate immune stimuli, such as infectious pathogens or malignant cells. The resulting persistent immune system stimulation drives hypercytokinemia, ultimately leading to severe tissue inflammation and end-organ damage. Traditionally, a diagnosis of FHLH requires the identification of biallelic loss-of-function mutations in one of these degranulation pathway genes. However, this narrow definition fails to encompass patients with other genetic mechanisms underlying degranulation pathway dysfunction. In particular, mounting clinical evidence supports a potential digenic mode of inheritance of FHLH in which single loss-of-function mutations in two different degranulation pathway genes cooperate to impair pathway activity. Here, we review the functions of the FHLH-associated genes within the degranulation pathway and summarize clinical evidence supporting a model in which cumulative defects along this mechanistic pathway may underlie HLH., Competing Interests: MH is a consultant for Novartis and Sobi. KN receives research funding from Incyte. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Steen, Hermiston, Nichols and Meyer.)

Abstract: Erythema nodosum leprosum (ENL) occurs as an immunological complication of multibacillary leprosy (MBL). The pathogenesis of ENL is long considered to be a T-cell-mediated process. The role of B cells and plasma cells in ENL is not well described in the literature. Therefore, we investigated the B-cell and plasma cell infiltrates in the skin biopsies of biopsy-proven cases of ENL by immunohistochemistry and image morphometry and compared the result with paucibacillary leprosy and MBL. Moreover, we sought a correlation of the B-cell and plasma cell infiltrates with different clinical, hematological, histopathological, and bacteriological parameters as well as the T-cell subsets in the skin biopsies. Our study highlighted a significant reduction in the number of B cells from paucibacillary leprosy to MBL to ENL, although there was no significant variation in the plasma cell infiltrate. The plasma cell infiltrate correlated with absolute neutrophilia in the blood and the presence of eosinophils in the ENL lesions. Both B cells and plasma cells positively correlated with CD4-positive T-helper cells and the CD8-positive cytotoxic T cells. Besides, the B cells also correlated positively with the CD3-positive pan T cells in the biopsy and negatively correlated with the T-regulatory:T-cell ratio. Our results suggested the role of B cells and plasma cells even at the tissue level in the pathobiogenesis of ENL., Competing Interests: The authors declare no conflicts of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

During the last decade, we have persistently addressed the question, "how can the innate immune system be used as a therapeutic tool to eliminate cancer?" A cancerous tumor harbors innate immune cells such as macrophages, which are held in the tumor-promoting M2 state by tumor-cell-released cytokines. We have discovered that these tumor-associated macrophages (TAM) are repolarized into the nitric oxide (NO)-generating tumoricidal M1 state by the dietary agent curcumin (CC), which also causes recruitment of activated natural killer (NK) cells and cytotoxic T (Tc) cells into the tumor, thereby eliminating cancer cells as well as cancer stem cells. Indications are that this process may be NO-dependent. Intriguingly, the maximum blood concentration of CC in mice never exceeds nanomolar levels. Thus, our results submit that even low, transient levels of curcumin in vivo are enough to cause repolarization of the TAM and recruitment NK cells as well as Tc cells to eliminate the tumor. We have observed this phenomenon in two cancer models, glioblastoma and cervical cancer. Therefore, this approach may yield a general strategy to fight cancer. Our mechanistic studies have so far implicated induction of STAT-1 in this M2→M1 switch, but further studies are needed to understand the involvement of other factors such as the lipid metabolites resolvins in the CC-evoked anticancer pathways.

Type 2 DCs (DC2s) comprise the majority of conventional DCs within most tumors; however, little is known about their ability to initiate and sustain antitumor immunity, as most studies have focused on antigen cross-presenting DC1s. Here, we report that DC2 infiltration identified by analysis of multiple human cancer data sets showed a significant correlation with survival across multiple human cancers, with the benefit being seen in tumors resistant to cytotoxic T cell control. Characterization of DC subtype infiltration into an immunotherapy-resistant model of breast cancer revealed that impairment of DC1s through 2 unique models resulted in enhanced DC2 functionality and improved tumor control. BATF3 deficiency depleted intratumoral DC1s, which led to increased DC2 lymph node migration and CD4+ T cell activation. Enhancing DC2 stimulatory potential by genetic deletion of Hsp90b1 (encoding molecular chaperon GP96) led to a similar enhancement of T cell immunity and improved survival in a spontaneous breast cancer model. These data highlight the therapeutic and prognostic potential of DC2s within checkpoint blockade-resistant tumors.

Chronic rhinosinusitis with nasal polyps (CRSwNP) is characterized by a chronic inflammatory process often associated with comorbid asthma. In this study, we analyzed the transcriptomes of single T helper (T H ) cells from nasal polyps of patients with CRSwNP and validated these findings using multiparameter flow cytometry. Polyp tissue contained suppressive T regulatory (T reg ) cells, T H 2 cells, type 2 innate lymphoid cells, and three transcriptionally distinct subsets of cytotoxic CD4 + T cells (CD4 + CTL). GATA3 expression was a feature of polyp T reg cells, whereas T H 2 cells highly expressed TCN1 , CD200R , and HPGDS and were enriched for genes involved in lipid metabolism. Only a portion of polyp T H 2 cells expressed the prostaglandin D2 receptor CRTH2, whereas a subpopulation of CD109 + CRTH2 - T H 2 cells expressed mRNA for common inhibitor receptors including LAG3 and TIM3 and produced IL-10. Together, we resolved the complexity of T H cells in patients with CRSwNP, identifying several distinct clusters of CD4 + CTL and a population of CD109 + CRTH2 - T H 2 cells with putative regulatory potential., (Copyright © 2021 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Purpose: Expansion of CD8 + cytotoxic Tlymphocytes is a prerequisite for anti-cancer immune activity and has gained interest in the era of immune checkpoint therapy., Methods: To understand the CD8 + T cell dynamics in the tumor microenvironment, we used multiplex fluorescence immunohistochemistry to quantitate CD8 + proliferation (Ki67 co-expression) in tissue microarrays from 1107 colorectal, 642 renal cell, 1066 breast, 375 ovarian, 451 pancreatic and 347 gastric cancer samples., Results: The density and the percentage of proliferating (Ki67 + ) CD8 + T cells were both highly variable between tumor types as well as between patients with the same tumor type. Elevated density and percentage of proliferating CD8 + cytotoxic T cells were significantly associated with favorable tumor parameters such as low tumor stage, negative nodal stage (p ≤ 0.0041 each), prolonged overall survival (p ≤ 0.0028 each) and an inflamed immune phenotype (p = 0.0025) in colorectal cancer and, in contrast, linked to high tumor stage, advanced ISUP/Fuhrman/Thoenes grading (each p ≤ 0.003), shorter overall survival (p ≤ 0.0330 each) and an immune inflamed phenotype (p = 0.0094) in renal cell cancer. In breast, ovarian, pancreatic and gastric cancer the role of (Ki67 + )CD8 + Tcells was not linked to clinicopathological data., Conclusion: Our data demonstrate a tumor type dependent prognostic impact of proliferating (Ki67 + )CD8 + Tcells and an inverse impact in colorectal and renal cell cancer., (© 2021. The Author(s).)

LINC01355 has been demonstrated to be dysregulated in several cancers. However, the exact molecular function of LINC01355 in the pathogenesis of OSCC remains unstudied. Here, we reported the effect of LINC01355 in OSCC and investigated the mechanisms. Firstly, we found that the results indicated LINC01355 was increased in OSCC cells. Knockdown of LINC01355 repressed OSCC cell proliferation, migration, and invasion. Recently, immunotherapy is a significant method for the treatment of cancers, in which CD8+ T cells exhibit a significant role. The influence of LINC01355 on the antitumor activity of CD8+ T cells was also focused in this study. As shown, the silence of LINC01355 could repress OSCC tumor growth via inducing CD8+ T cell immune responses. In addition, we found that downregulation of LINC01355 significantly restrained CD8+ T cell apoptosis, induced CD8+ T cell percentage, and enhanced the cytolysis activity when cocultured with OSCC cells. It has been reported that the Notch pathway represses CD8+ T cell activity in cancer patients. In our present study, we displayed that lack of LINC01355 suppressed OSCC malignant behaviors and enhanced the antitumor activity of CD8+ T cells via inactivating Notch signaling. We showed that decreased LINC01355 significantly restrained the Notch signal via a decrease of Notch-1, JAG-1, and HES-1. Repression of Notch1 reversed the effect of LINC01355 in OSCC cells. In conclusion, it was implied that LINC01355 might induce the development of OSCC via modulating the Notch signal pathway, which could provide a candidate therapeutic target for OSCC., Competing Interests: The authors declare that there is no conflict of interest regarding the publication of this paper., (Copyright © 2021 Chen Zou et al.)

Background: Adoptive T cell transfer-based immunotherapy yields unsatisfactory results in the treatment of solid tumors, partially owing to limited tumor infiltration and the immunosuppressive microenvironment in solid tumors. Therefore, strategies for the noninvasive tracking of adoptive T cells are critical for monitoring tumor infiltration and for guiding the development of novel combination therapies., Methods: We developed a radiolabeling method for cytotoxic T lymphocytes (CTLs) that comprises metabolically labeling the cell surface glycans with azidosugars and then covalently conjugating them with 64 Cu-1,4,7-triazacyclononanetriacetic acid-dibenzo-cyclooctyne ( 64 Cu-NOTA-DBCO) using bioorthogonal chemistry. 64 Cu-labeled control-CTLs and ovalbumin-specific CTLs (OVA-CTLs) were tracked using positron emission tomography (PET) in B16-OVA tumor-bearing mice. We also investigated the effects of focal adhesion kinase (FAK) inhibition on the antitumor efficacy of OVA-CTLs using a poly(lactic-co-glycolic) acid (PLGA)-encapsulated nanodrug (PLGA-FAKi)., Results: CTLs can be stably radiolabeled with 64 Cu with a minimal effect on cell viability. PET imaging of 64 Cu-OVA-CTLs enables noninvasive mapping of their in vivo behavior. Moreover, 64 Cu-OVA-CTLs PET imaging revealed that PLGA-FAKi induced a significant increase in OVA-CTL infiltration into tumors, suggesting the potential for a combined therapy comprising OVA-CTLs and PLGA-FAKi. Further combination therapy studies confirmed that the PLGA-FAKi nanodrug markedly improved the antitumor effects of adoptive OVA-CTLs transfer by multiple mechanisms., Conclusion: These findings demonstrated that metabolic radiolabeling followed by PET imaging can be used to sensitively profile the early-stage migration and tumor-targeting efficiency of adoptive T cells in vivo. This strategy presents opportunities for predicting the efficacy of cell-based adoptive therapies and for guiding combination regimens.

Tumors arising in the context of Lynch Syndrome or constitutional mismatch repair deficiency are hypermutated and have a good response towards immune-checkpoint inhibitors (ICIs), including α-PD-L1 antibodies. However, in most cases, resistance mechanisms evolve. To improve outcomes and prevent resistance development, combination approaches are warranted. Herein, we applied a combined regimen with an α-PD-L1 antibody and gemcitabine in a preclinical tumor model to activate endogenous antitumor immune responses. Mlh1 -/- mice with established gastrointestinal tumors received the α-PD-L1 antibody (clone 6E11; 2.5 mg/kg bw, i.v., q2wx3) and gemcitabine (100 mg/kg bw, i.p., q4wx3) in mono- or combination therapy. Survival and tumor growth were recorded. Immunological changes in the blood were routinely examined via multi-color flow cytometry and complemented by ex vivo frameshift mutation analysis to identify alterations in Mlh1 -/- -tumor-associated target genes. The combined therapy of α-PD-L1 and gemcitabine prolonged median overall survival of Mlh1 -/- mice from four weeks in the untreated control group to 12 weeks, accompanied by therapy-induced tumor growth inhibition, as measured by [18F] -FDG PET/CT. Plasma cytokine levels of IL13, TNFα, and MIP1β were increased and also higher than in mice receiving either monotherapy. Circulating splenic and intratumoral myeloid-derived suppressor cells (MDSCs), as well as M2 macrophages, were markedly reduced. Besides, residual tumor specimens from combi-treated mice had increased numbers of infiltrating cytotoxic T-cells. Frameshift mutations in APC , Tmem60 , and Casc3 were no longer detectable upon treatment, likely because of the successful eradication of single mutated cell clones. By contrast, novel mutations appeared. Collectively, we herein confirm the safe application of combined chemo-immunotherapy by long-term tumor growth control to prevent the development of resistance mechanisms.

Melanoma differentiation associated gene-9 (MDA-9), Syntenin-1, or syndecan binding protein is a differentially regulated prometastatic gene with elevated expression in advanced stages of melanoma. MDA-9/Syntenin expression positively associates with advanced disease stage in multiple histologically distinct cancers and negatively correlates with patient survival and response to chemotherapy. MDA-9/Syntenin is a highly conserved PDZ-domain scaffold protein, robustly expressed in a spectrum of diverse cancer cell lines and clinical samples. PDZ domains interact with a number of proteins, many of which are critical regulators of signaling cascades in cancer. Knockdown of MDA-9/Syntenin decreases cancer cell metastasis, sensitizing these cells to radiation. Genetic silencing of MDA-9/Syntenin or treatment with a pharmacological inhibitor of the PDZ1 domain, PDZ1i, also activates the immune system to kill cancer cells. Additionally, suppression of MDA-9/Syntenin deregulates myeloid-derived suppressor cell differentiation via the STAT3/interleukin (IL)-1β pathway, which concomitantly promotes activation of cytotoxic T lymphocytes. Biologically, PDZ1i treatment decreases metastatic nodule formation in the lungs, resulting in significantly fewer invasive cancer cells. In summary, our observations indicate that MDA-9/Syntenin provides a direct therapeutic target for mitigating aggressive breast cancer and a small-molecule inhibitor, PDZ1i, provides a promising reagent for inhibiting advanced breast cancer pathogenesis., Competing Interests: Competing interest statement: W.K.C. and P.B.F. are cofounders and have ownership interest in InVaMet Therapeutics, Inc. Virginia Commonwealth University and the Sanford Burnham Prebys Medical Discovery Institute have ownership interest in InVaMet Therapeutics, Inc.

The mechanism by which anti-cancer immunity shapes early carcinogenesis of lung adenocarcinoma (ADC) is unknown. In this study, we characterize the immune contexture of invasive lung ADC and its precursors by transcriptomic immune profiling, T cell receptor (TCR) sequencing and multiplex immunofluorescence (mIF). Our results demonstrate that anti-tumor immunity evolved as a continuum from lung preneoplasia, to preinvasive ADC, minimally-invasive ADC and frankly invasive lung ADC with a gradually less effective and more intensively regulated immune response including down-regulation of immune-activation pathways, up-regulation of immunosuppressive pathways, lower infiltration of cytotoxic T cells (CTLs) and anti-tumor helper T cells (Th), higher infiltration of regulatory T cells (Tregs), decreased T cell clonality, and lower frequencies of top T cell clones in later-stages. Driver mutations, chromosomal copy number aberrations (CNAs) and aberrant DNA methylation may collectively impinge host immune responses and facilitate immune evasion, promoting the outgrowth of fit subclones in preneoplasia into dominant clones in invasive ADC.

Dendritic cells (DC) and T-cells are mediators of CTL-responses. Autologous (from patients with acute myeloid leukaemia (AML) or myelodysplasia (MDS)) or allogeneic (donor)-T-cells stimulated by DC leu , gain an efficient lysis of naive blasts, although not in every case. CXCL8, -9, -10, CCL2, -5 and Interleukin (IL-12) were quantified by Cytometric Bead Array (CBA) in supernatants from 5 DC-generating methods and correlated with AML-/MDS-patients' serum-values, DC-/T-cell-interactions/antileukemic T-cell-reactions after mixed lymphocyte culture (MLC) and patients' clinical course. The blast-lytic activity of T-cells stimulated with DC or mononuclear cells (MNC) was quantified in a cytotoxicity assay. Despite great variations of chemokine-levels, correlations with post-stimulation (after stimulating T-cells with DC in MLC) improved antileukemic T-cell activity were seen: higher released chemokine-values correlated with improved T-cells' antileukemic activity (compared to stimulation with blast-containing MNC) - whereas with respect to the corresponding serum values higher CXCL8-, -9-, and -10- but lower CCL5- and -2-release correlated with improved antileukemic activity of DC-stimulated (vs. blast-stimulated) T-cells. In DC-culture supernatants higher chemokine-values correlated with post-stimulation improved antileukemic T-cell reactivity, whereas higher serum-values of CXCL8, -9, and -10 but lower serum-values of CCL5 and -2 correlated with post-stimulation improved antileukemic T-cell-reactivity. In a context of 'DC'-stimulation (vs serum) this might point to a change of (CCL5 and -2-associated) functionality from a more 'inflammatory' or 'tumor-promoting' to a more 'antitumor'-reactive functionality. This knowledge could contribute to develop immune-modifying strategies that promote antileukemic (adaptive) immune-responses., (Copyright © 2021 Elsevier GmbH. All rights reserved.)

The beta-coronavirus SARS-CoV-2 induces severe disease (COVID-19) mainly in elderly persons with risk factors, whereas the majority of patients experience a mild course of infection. As the circulating common cold coronaviruses OC43 and HKU1 share some homologous sequences with SARS-CoV-2, beta-coronavirus cross-reactive T-cell responses could influence the susceptibility to SARS-CoV-2 infection and the course of COVID-19. To investigate the role of beta-coronavirus cross-reactive T-cells, we analyzed the T-cell response against a 15 amino acid long peptide (SCoV-DP15: DLSPRWYFYYLGTGP) from the SARS-CoV-2 nucleoprotein sequence with a high homology to the corresponding sequence (QLLPRWYFYYLGTGP) in OC43 and HKU1. SCoV-DP15-specific T-cells were detected in 4 out of 23 (17.4%) SARS-CoV-2-seronegative healthy donors. As HIV-1 infection is a potential risk factor for COVID-19, we also studied a cohort of HIV-1-infected patients on antiretroviral therapy. 44 out of these 116 HIV-1-infected patients (37.9%) showed a specific recognition of the SCoV-DP15 peptide or of shorter peptides within SCoV-DP15 by CD4 + T-cells and/or by CD8 + T-cells. We could define several new cross-reactive HLA-I-restricted epitopes in the SARS-CoV-2 nucleoprotein such as SPRWYFYYL (HLA-B*07, HLA-B*35), DLSPRWYFYY (HLA-A*02), LSPRWYFYY (HLA-A*29), WYFYYLGTGP and WYFYYLGT. Epitope specific CD8 + T-cell lines recognized corresponding epitopes within OC43 and HKU1 to a similar degree or even at lower peptide concentrations suggesting that they were induced by infection with OC43 or HKU1. Our results confirm that SARS-CoV-2-seronegative subjects can target SARS-CoV-2 not only by beta-coronavirus cross-reactive CD4 + T-cells but also by cross-reactive CD8 + cytotoxic T-cells (CTL). The delineation of cross-reactive T-cell epitopes contributes to an efficient epitope-specific immunomonitoring of SARS-CoV-2-specific T-cells. Further prospective studies are needed to prove a protective role of cross-reactive T-cells and their restricting HLA alleles for control of SARS-CoV-2 infection. The frequent observation of SARS-CoV-2-reactive T-cells in HIV-1-infected subjects could be a reason that treated HIV-1 infection does not seem to be a strong risk factor for the development of severe COVID-19., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Schmidt, Nganou-Makamdop, Tenbusch, El Kenz, Maier, Lapuente, Überla, Spriewald, Bergmann, Harrer and Harrer.)

Exocytosis of cytotoxic granules (CG) by lymphocytes is required for the elimination of infected and malignant cells. Impairments in this process underly a group of diseases with dramatic hyperferritinemic inflammation termed hemophagocytic lymphohistiocytosis (HLH). Although genetic and functional studies of HLH have identified proteins controlling distinct steps of CG exocytosis, the molecular mechanisms that spatiotemporally coordinate CG release remain partially elusive. We studied a patient exhibiting characteristic clinical features of HLH associated with markedly impaired cytotoxic T lymphocyte (CTL) and natural killer (NK) cell exocytosis functions, who beared biallelic deleterious mutations in the gene encoding the small GTPase RhoG. Experimental ablation of RHOG in a model cell line and primary CTLs from healthy individuals uncovered a hitherto unappreciated role of RhoG in retaining CGs in the vicinity of the plasma membrane (PM), a fundamental prerequisite for CG exocytotic release. We discovered that RhoG engages in a protein-protein interaction with Munc13-4, an exocytosis protein essential for CG fusion with the PM. We show that this interaction is critical for docking of Munc13-4+ CGs to the PM and subsequent membrane fusion and release of CG content. Thus, our study illuminates RhoG as a novel essential regulator of human lymphocyte cytotoxicity and provides the molecular pathomechanism behind the identified here and previously unreported genetically determined form of HLH., (© 2021 by The American Society of Hematology.)