The cytokine Meteorin-like inhibits anti-tumor CD8+ T cell responses by disrupting mitochondrial function.

Tumor-infiltrating lymphocyte (TIL) hypofunction contributes to the progression of advanced cancers and is a frequent target of immunotherapy. Emerging evidence indicates that metabolic insufficiency drives T cell hypofunction during tonic stimulation, but the signals that initiate metabolic reprogramming in this context are largely unknown. Here, we found that Meteorin-like (METRNL), a metabolically active cytokine secreted by immune cells in the tumor microenvironment (TME), induced bioenergetic failure of CD8+ T cells. METRNL was secreted by CD8+ T cells during repeated stimulation and acted via both autocrine and paracrine signaling. Mechanistically, METRNL increased E2F-peroxisome proliferator-activated receptor delta (PPARδ) activity, causing mitochondrial depolarization and decreased oxidative phosphorylation, which triggered a compensatory bioenergetic shift to glycolysis. Metrnl ablation or downregulation improved the metabolic fitness of CD8+ T cells and enhanced tumor control in several tumor models, demonstrating the translational potential of targeting the METRNL-E2F-PPARδ pathway to support bioenergetic fitness of CD8+ TILs.

The CCR6-CCL20 axis promotes regulatory T cell glycolysis and immunosuppression in tumors.

Regulatory T cells (Tregs) are important players in the tumor microenvironment. However, the mechanisms behind their immunosuppressive effects are poorly understood. We found that CCR6-CCL20 activity in tumor-infiltrating Tregs is associated with greater glycolytic activity and ablation of Ccr6 reduced glycolysis and lactic acid production while increasing compensatory glutamine metabolism. Immunosuppressive activity towards CD8+ T cells was abrogated in Ccr6-/- Tregs due to reduction in activation-induced glycolysis. Furthermore, Ccr6-/- mice exhibited improved survival across multiple tumor models compared to wildtype mice, and Treg and CD8+ T-cell depletion abrogated the improvement. In addition, Ccr6 ablation further promoted the efficacy of anti-PD-1 therapy in a preclinical glioma model. Follow-up knockdown of Ccl20 with siRNA also demonstrated improvement in antitumor efficacy. Our results unveil CCR6 as a marker and regulator of Treg-induced immunosuppression and identify approaches to target the metabolic determinants of Treg immunosuppressive activity.

Calcium-Related Gene Signatures May Predict Prognosis and Level of Immunosuppression in Gliomas.

Gliomas are the most common primary brain cancer. While it has been known that calcium-related genes correlate with gliomagenesis, the relationship between calcium-related genes and glioma prognosis remains unclear. We assessed TCGA datasets of mRNA expressions with differentially expressed genes (DEGs) and enrichment analysis to specifically screen for genes that regulate or are affected by calcium levels. We then correlated the identified calcium-related genes with unsupervised/supervised learning to classify glioma patients into 2 risk groups. We also correlated our identified genes with immune signatures. As a result, we discovered 460 calcium genes and 35 calcium key genes that were associated with OS. There were 13 DEGs between Clusters 1 and 2 with different OS. At the same time, 10 calcium hub genes (CHGs) signature model were constructed using supervised learning, and the prognostic risk scores of the 3 cohorts of samples were calculated. The risk score was confirmed as an independent predictor of prognosis. Immune enrichment analysis revealed an immunosuppressive tumor microenvironment with upregulation of checkpoint markers in the high-risk group. Finally, a nomogram was generated with risk scores and other clinical prognostic independent indicators to quantify prognosis. Our findings suggest that calcium-related gene expression patterns could be applicable to predict prognosis and predict levels of immunosuppression.

The safety and efficacy of dexamethasone in the perioperative management of glioma patients.

OBJECTIVE: In this single-institution retrospective cohort study, the authors evaluated the effect of dexamethasone on postoperative complications and overall survival in patients with glioma undergoing resection. METHODS: A total of 435 patients who underwent resection of a primary glioma were included in this retrospective cohort study. The inclusion criterion was all patients who underwent resection of a primary glioma at a tertiary medical center between 2014 and 2019. RESULTS: The use of both pre- and postoperative dexamethasone demonstrated a trend toward the development of postoperative wound infections (3% vs 0% in single use or no use, p = 0.082). No association was detected between dexamethasone use and the development of new-onset hyperglycemia (p = 0.149). On multivariable Cox proportional hazards analysis, dexamethasone use was associated with a greater hazard of death (overall p = 0.017); this effect was most pronounced for preoperative (only) dexamethasone use (hazard ratio 3.0, p = 0.062). CONCLUSIONS: Combined pre- and postoperative dexamethasone use may increase the risk of postoperative wound infection, and dexamethasone use, specifically preoperative use, may negatively impact survival. These findings highlight the potential for serious negative consequences with dexamethasone use.

Synergy between glutamate modulation and anti-programmed cell death protein 1 immunotherapy for glioblastoma.

OBJECTIVE: Immune checkpoint inhibitors such as anti-programmed cell death protein 1 (anti-PD-1) have shown promise for the treatment of cancers such as melanoma, but results for glioblastoma (GBM) have been disappointing thus far. It has been suggested that GBM has multiple mechanisms of immunosuppression, indicating a need for combinatorial treatment strategies. It is well understood that GBM increases glutamate in the tumor microenvironment (TME); however, the significance of this is not well understood. The authors posit that glutamate upregulation in the GBM TME is immunosuppressive. The authors utilized a novel glutamate modulator, BHV-4157, to determine synergy between glutamate modulation and the well-established anti-PD-1 immunotherapy for GBM. METHODS: C57BL/6J mice were intracranially implanted with luciferase-tagged GL261 glioma cells. Mice were randomly assigned to the control, anti-PD-1, BHV-4157, or combination anti-PD-1 plus BHV-4157 treatment arms, and median overall survival was assessed. In vivo microdialysis was performed at the tumor site with administration of BHV-4157. Intratumoral immune cell populations were characterized with immunofluorescence and flow cytometry. RESULTS: The BHV-4157 treatment arm demonstrated improved survival compared with the control arm (p < 0.0001). Microdialysis demonstrated that glutamate concentration in TME significantly decreased after BHV-4157 administration. Immunofluorescence and flow cytometry demonstrated increased CD4+ T cells and decreased Foxp3+ T cells in mice that received BHV-4157 treatment. No survival benefit was observed when CD4+ or CD8+ T cells were depleted in mice prior to BHV-4157 administration (p < 0.05). CONCLUSIONS: In this study, the authors showed synergy between anti-PD-1 immunotherapy and glutamate modulation. The authors provide a possible mechanism for this synergistic benefit by showing that BHV-4157 relies on CD4+ and CD8+ T cells. This study sheds light on the role of excess glutamate in GBM and provides a basis for further exploring combinatorial approaches for the treatment of this disease.

Comparison of POCIS and grab sampling techniques for monitoring PPCPs in vernal pools in central Pennsylvania.

Active ingredients in pharmaceuticals and personal care products (PPCPs) can persist through wastewater treatment plants and be released into the environment where they can inadvertently pose risks to non-target organisms. Emerging contaminants (ECs), including PPCPs, are commonly detected in wastewater effluent. With the increasing beneficial re-use of treated wastewater globally, there is a need to understand how spray-irrigation activities affect the occurrence and persistence of ECs in the environment to which they are introduced. Here, we explore the impacts of wastewater spray-irrigation on nearby ephemeral wetlands (e.g., vernal pools) through the use of grab and Polar Organic Chemical Integrative Sampling (POCIS) techniques. This study sought to determine whether integrative sampling techniques are better suited than traditional grab sampling techniques in assessing the presence and concentrations of ECs in vernal pools by evaluating 34 ECs in six vernal pools in central Pennsylvania. Three pools were impacted by wastewater spray-irrigation activities and three were in a nearby forested area. Results of this study found that POCIS detected a wide range of 25 ECs (log Kow between -2.6 and 9.37) more or, in some cases, equally frequently, relative to grab samples. Additionally, grab samples were found to best capture short-lived elevated inputs of ECs (from irrigation events) while POCIS were found to best capture ECs that were present in vernal pools over a longer period of time (weeks to months). For ECs detected more frequently in grab samples, concentrations were higher compared to time weighted average aqueous concentrations estimated from POCIS. This study advances understanding of the potential impact of wastewater beneficial reuse on vernal pools and informs how best to monitor the presence of ECs in vernal pools using integrative and grab sampling techniques.

Combination checkpoint therapy with anti-PD-1 and anti-BTLA results in a synergistic therapeutic effect against murine glioblastoma.

Clinical trials involving anti-programmed cell death protein-1 (anti-PD-1) failed to demonstrate improved overall survival in glioblastoma (GBM) patients. This may be due to the expression of alternative checkpoints such as B- and T- lymphocyte attenuator (BTLA) on several immune cell types including regulatory T cells. Murine GBM models indicate that there is significant upregulation of BTLA in the tumor microenvironment (TME) with associated T cell exhaustion. We investigate the use of antibodies against BTLA and PD-1 on reversing immunosuppression and increasing long-term survival in a murine GBM model. C57BL/6 J mice were implanted with the murine glioma cell line GL261 and randomized into 4 arms: (i) control, (ii) anti-PD-1, (iii) anti-BTLA, and (iv) anti-PD-1 + anti-BTLA. Kaplan-Meier curves were generated for all arms. Flow cytometric analysis of blood and brains were done on days 11 and 16 post-tumor implantation. Tumor-bearing mice treated with a combination of anti-PD-1 and anti-BTLA therapy experienced improved overall long-term survival (60%) compared to anti-PD-1 (20%) or anti-BTLA (0%) alone (P = .003). Compared to monotherapy with anti-PD-1, mice treated with combination therapy also demonstrated increased expression of CD4+ IFN-γ (P < .0001) and CD8+ IFN-γ (P = .0365), as well as decreased levels of CD4+ FoxP3+ regulatory T cells on day 16 in the brain (P = .0136). This is the first preclinical investigation into the effects of combination checkpoint blockade with anti-PD-1 and anti-BTLA treatment in GBM. We also show a direct effect on activated immune cell populations such as CD4+ and CD8 + T cells and immunosuppressive regulatory T cells through this combination therapy.

Sustained localized delivery of immunotherapy to lymph nodes reverses immunosuppression and increases long-term survival in murine glioblastoma.

Introduction: Despite the advent of immunotherapy as a promising therapeutic, glioblastoma (GBM) remains resistant to using checkpoint blockade due to its highly immunosuppressive tumor milieu. Moreover, current anti-PD-1 treatment requires multiple infusions with adverse systemic effects. Therefore, we used a PCL:PEG:PCL polymer gel loaded with anti-PD-1 and implanted at the site of lymph nodes in an attempt to maximize targeting of inactivated T cells as well as mitigate unnecessary systemic exposure. Methods: Mice orthotopically implanted with GL261 glioma cells were injected with hydrogels loaded with anti-PD-1 in one of the following locations: cervical lymph nodes, inguinal lymph nodes, and the tumor site. Mice treated systemically with anti-PD-1 were used as comparative controls. Kaplan-Meier curves were generated for all arms, with ex vivo flow cytometric staining for L/D, CD45, CD3, CD4, CD8, TNF-α and IFN-y and co-culture ELISpots were done for immune cell activation assays. Results: Mice implanted with PCL:PEG:PCL hydrogels carrying anti-PD-1 at the site of their lymph nodes showed significantly improved survival outcomes compared to mice systemically treated with anti-PD-1 (P = .0185). Flow cytometric analysis of brain tissue and co-culture of lymph node T cells from mice implanted with gels demonstrated increased levels of IFN-y and TNF-α compared to mice treated with systemic anti-PD-1, indicating greater reversal of immunosuppression compared to systemic treatment. Conclusions: Our data demonstrate proof of principle for using localized therapy that targets lymph nodes for GBM. We propose an alternative treatment paradigm for developing new sustained local treatments with immunotherapy that are able to eliminate the need for multiple systemic infusions and their off-target effects.

IQGAP2 acts as an independent prognostic factor and is related to immunosuppression in DLBCL.

BACKGROUND: Almost one-third of patients with diffuse large B-cell lymphoma (DLBCL) cannot be cured with initial therapy and will eventually succumb to the disease. Further elaboration of prognostic markers of DLBCL will provide therapeutic targets. IQ motif-containing GTPase activating protein 2 (IQGAP2) acts as a tumour suppressor in hepatocellular, prostate, and gastric cancers. However, the role of IQGAP2 in DLBCL remains unclear. METHODS: We collected mRNA expression data from 614 samples and the corresponding clinical information. The survival time of patients was compared between groups according to the mRNA expression level of IQGAP2. Survival analyses were performed in different subgroups when considering the effect of age, tumour stage, serum lactate dehydrogenase (LDH) concentration, performance status, and the number of extra nodal disease sites. The biological processes associated with IQGAP2-associated mRNAs were analysed to predict the function of IQGAP2. The correlation of IQGAP2 mRNA with immunosuppressive genes and leukocyte infiltration were analysed. RESULTS: The overall survival of patients with increased IQGAP2 mRNA levels was reduced even after aggressive treatment independent of age, tumour stage, serum LDH concentration, performance status, and the number of extra nodal disease sites. Furthermore, the biological processes of IQGAP2-associated mRNAs were mainly immune processes. IQGAP2 mRNA expression was correlated with the expression of immunosuppressive genes and leukocyte infiltration. CONCLUSION: IQGAP2 mRNA is an independent prognostic factor and is related to immunosuppression in DLBCL. This discovery may provide a promising target for further development of therapy.

PD-1+ Monocytes Mediate Cerebral Vasospasm Following Subarachnoid Hemorrhage.

BACKGROUND: Cerebral vasospasm is a major source of morbidity and mortality following aneurysm rupture and has limited treatment options. OBJECTIVE: To evaluate the role of programmed death-1 (PD-1) in cerebral vasospasm. METHODS: Endovascular internal carotid artery perforation (ICAp) was used to induce cerebral vasospasm in mice. To evaluate the therapeutic potential of targeting PD-1, programmed death ligand-1 (PD-L1) was administered 1 h after ICAp and vasospasm was measured histologically at the level of the ICA bifurcation bilaterally. PD-1 expressing immune cell populations were evaluated by flow cytometry. To correlate these findings to patients and evaluate the potential of PD-1 as a biomarker, monocytes were isolated from the peripheral blood and analyzed by flow cytometry in a cohort of patients with ruptured cerebral aneurysms. The daily frequency of PD-1+ monocytes in the peripheral blood was correlated to transcranial Doppler velocities as well as clinical and radiographic vasospasm. RESULTS: We found that PD-L1 administration prevented cerebral vasospasm by inhibiting ingress of activated Ly6c+ and CCR2+ monocytes into the brain. Human correlative studies confirmed the presence of PD-1+ monocytes in the peripheral blood of patients with ruptured aneurysms and the frequency of these cells corresponded with cerebral blood flow velocities and clinical vasospasm. CONCLUSION: Our results identify PD-1+ monocytes as mediators of cerebral vasospasm and support PD-1 agonism as a novel therapeutic strategy.