scholarly journals Licorice extract inhibits growth of non-small cell lung cancer by down-regulating CDK4-Cyclin D1 complex and increasing CD8+ T cell infiltration

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Jinglin Zhu ◽  
Ruifei Huang ◽  
Ruijie Yang ◽  
Yue Xiao ◽  
Jiangna Yan ◽  
...  
Keyword(s):  
T Cell ◽  
Cyclin D1 ◽  
In Vivo Studies ◽  
Cell Infiltration ◽  
Immunomodulatory Activity ◽  
Systems Pharmacology ◽  
T Cell Infiltration ◽  
Phase Cycle ◽  
Anti Tumor Activity

Abstract Background Targeting tumor microenvironment (TME) may provide therapeutic activity and selectivity in treating cancers. Therefore, an improved understanding of the mechanism by which drug targeting TME would enable more informed and effective treatment measures. Glycyrrhiza uralensis Fisch (GUF, licorice), a widely used herb medicine, has shown promising immunomodulatory activity and anti-tumor activity. However, the molecular mechanism of this biological activity has not been fully elaborated. Methods Here, potential active compounds and specific targets of licorice that trigger the antitumor immunity were predicted with a systems pharmacology strategy. Flow cytometry technique was used to detect cell cycle profile and CD8+ T cell infiltration of licorice treatment. And anti-tumor activity of licorice was evaluated in the C57BL/6 mice. Results We reported the G0/G1 growth phase cycle arrest of tumor cells induced by licorice is related to the down-regulation of CDK4-Cyclin D1 complex, which subsequently led to an increased protein abundance of PD-L1. Further, in vivo studies demonstrated that mitigating the outgrowth of NSCLC tumor induced by licorice was reliant on increased antigen presentation and improved CD8+ T cell infiltration. Conclusions Briefly, our findings improved the understanding of the anti-tumor effects of licorice with the systems pharmacology strategy, thereby promoting the development of natural products in prevention or treatment of cancers.

scholarly journals Single cell transcriptomics reveals the effect of PD-L1/TGF-β blockade on the tumor microenvironment

2020 ◽  
Author(s):  
Yoong Wearn Lim ◽  
Garry L. Coles ◽  
Savreet K. Sandhu ◽  
David S. Johnson ◽  
Adam S. Adler ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Microenvironment ◽  
Single Cell ◽  
Immune Cell ◽  
Single Cells ◽  
Cell Infiltration ◽  
Cytotoxic Lymphocytes ◽  
T Cell Infiltration ◽  
Anti Tumor Activity

AbstractBackgroundThe anti-tumor activity of anti-PD-1/PD-L1 therapies correlates with T cell infiltration in tumors. Thus, a major goal in oncology is to find strategies that enhance T cell infiltration and efficacy of anti-PD-1/PD-L1 therapy. TGF-β has been shown to contribute to T cell exclusion and anti-TGF-β improves anti-PD-L1 efficacy in vivo. However, TGF-β inhibition has frequently been shown to induce toxicity in the clinic, and the clinical efficacy of combination PD-L1 and TGF-β blockade has not yet been proven. To identify strategies to overcome resistance to PD-L1 blockade, the transcriptional programs associated with PD-L1 and/or TGF-β blockade in the tumor microenvironment should be further elucidated.ResultsFor the first time, we used single-cell RNA sequencing to characterize the transcriptomic effects of PD-L1 and/or TGF-β blockade on nearly 30,000 single cells in the tumor and surrounding microenvironment. Combination treatment led to upregulation of immune response genes, including multiple chemokine genes such as CCL5, in CD45+ cells, and down-regulation of extracellular matrix genes in CD45-cells. Analysis of publicly available tumor transcriptome profiles showed that the chemokine CCL5 was strongly associated with immune cell infiltration in various human cancers. Further investigation with in vivo models showed that intratumorally administered CCL5 enhanced cytotoxic lymphocytes and the anti-tumor activity of anti-PD-L1.ConclusionsTaken together, our data could be leveraged translationally to improve anti-PD-L1 plus anti-TGF-β combination therapy, for example through companion biomarkers, and/or to identify novel targets that could be modulated to overcome resistance.

scholarly journals Single-cell transcriptomics reveals the effect of PD-L1/TGF-β blockade on the tumor microenvironment

BMC Biology ◽  
2021 ◽  
Vol 19 (1) ◽  
Author(s):  
Yoong Wearn Lim ◽  
Garry L. Coles ◽  
Savreet K. Sandhu ◽  
David S. Johnson ◽  
Adam S. Adler ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Microenvironment ◽  
Single Cell ◽  
Immune Cell ◽  
Single Cells ◽  
Cell Infiltration ◽  
In Vivo Models ◽  
T Cell Infiltration ◽  
Anti Tumor Activity

Abstract Background The anti-tumor activity of anti-PD-1/PD-L1 therapies correlates with T cell infiltration in tumors. Thus, a major goal in oncology is to find strategies that enhance T cell infiltration and efficacy of anti-PD-1/PD-L1 therapy. TGF-β has been shown to contribute to T cell exclusion, and anti-TGF-β improves anti-PD-L1 efficacy in vivo. However, TGF-β inhibition has frequently been shown to induce toxicity in the clinic, and the clinical efficacy of combination PD-L1 and TGF-β blockade has not yet been proven. To identify strategies to overcome resistance to PD-L1 blockade, the transcriptional programs associated with PD-L1 and/or TGF-β blockade in the tumor microenvironment should be further elucidated. Results We used single-cell RNA sequencing in a mouse model to characterize the transcriptomic effects of PD-L1 and/or TGF-β blockade on nearly 30,000 single cells in the tumor and surrounding microenvironment. Combination treatment led to upregulation of immune response genes, including multiple chemokine genes such as CCL5, in macrophages, and downregulation of extracellular matrix genes in fibroblasts. Analysis of publicly available tumor transcriptome profiles showed that the chemokine CCL5 was strongly associated with immune cell infiltration in various human cancers. Further investigation with in vivo models showed that intratumorally administered CCL5 enhanced cytotoxic lymphocytes and the anti-tumor activity of anti-PD-L1. Conclusions Taken together, our data could be leveraged translationally to complement or find alternatives to anti-PD-L1 plus anti-TGF-β combination therapy, for example through companion biomarkers, and/or to identify novel targets that could be modulated to overcome resistance.

Tumor-derived GDF-15 to suppress t-lymphocyte recruitment to the tumor microenvironment resulting in resistance to ANTI-PD-1 treatment.

2021 ◽  
Vol 39 (15_suppl) ◽  
pp. e14532-e14532
Author(s):  
Joerg Wischhusen ◽  
Markus Haake ◽  
Neha Vashist ◽  
Sabrina Genßler ◽  
Kilian Wistuba-Hamprecht ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Immune Cell ◽  
Serum Levels ◽  
Cell Infiltration ◽  
Immune Cell Infiltration ◽  
T Cell Infiltration ◽  
Melanoma Patients

e14532 Background: Growth and differentiation factor 15 (GDF-15) is a divergent member of the TGF-β superfamily with low to absent expression in healthy tissue. GDF-15 has been linked to feto-maternal immune tolerance, to prevention of excessive immune cell infiltration during tissue damage, and to anorexia. Various major tumor types secrete high levels of GDF-15. In cancer patients, elevated GDF-15 serum levels correlate with poor prognosis and reduced overall survival (OS). Methods: Impact of a proprietary GDF-15 neutralizing antibody (CTL-002) regarding T cell trafficking was analyzed by whole blood adhesion assays, a HV18-MK melanoma-bearing humanized mouse model and a GDF-15-transgenic MC38 model. Additionally, patient GDF-15 serum levels were correlated with clinical response and overall survival in oropharyngeal squamous cell carcinoma (OPSCC) and melanoma brain metastases. Results: In whole blood cell adhesion assays GDF-15 impairs adhesion of T and NK cells to activated endothelial cells. Neutralization of GDF-15 by CTL-002 rescued T cell adhesion. In HV18-MK-bearing humanized mice CTL-002 induced a strong increase in TIL numbers. Subset analysis revealed an overproportional enrichment of T cells, in particular CD8+ T cells. As immune cell exclusion is detrimental for checkpoint inhibitor (CPI) therapy, a GDF-15-transgenic MC38 model was tested for anti-PD-1 therapy efficacy. In GDF-15 overexpressing MC38 tumors response to anti PD-1 therapy was reduced by 90% compared to wtMC38 tumors. Combining aPD-1 with CTL-002 resulted in 50% of the mice rejecting their GDF-15 overexpressing tumors. Clinically, inverse correlations of GDF-15 levels with CD8+ T cell infiltration were shown for HPV+ OPSCC and for melanoma brain metastases. GDF-15 serum levels were significantly higher in HPV- than in HPV+ OPSCC patient (p < 0.0001). Low GDF-15 levels corresponded to longer OS in both HPV- and HPV+ OPSCC. In two independent melanoma patient cohorts treated with nivolumab or pembrolizumab low baseline serum GDF-15 levels were predictive for clinical response to anti-PD1 treatment and superior OS. Bivariate analysis including LDH indicates that GDF-15 independently predicts poor survival in aPD-1 treated melanoma patients. Conclusions: Taken together our in vitro and in vivo data show that elevated GDF-15 levels block T-cell infiltration into tumor tissues. Neutralizing GDF-15 with CTL-002 restores the ability of T cells to extravasate blood vessels and enter tumor tissue both in vitro and in vivo. In melanoma, patients with higher GDF-15 levels have significantly shorter survival and are less likely to respond to anti-PD1 therapy. GDF-15 may thus serve as a new predictive biomarker for anti-PD1 response, but most importantly also represents a novel target for cancer immunotherapy to improve tumor immune cell infiltration and response to anti-PD1 therapy.

scholarly journals EMP3 mediates glioblastoma‐associated macrophage infiltration to drive T cell exclusion

2021 ◽  
Vol 40 (1) ◽  
Author(s):  
Qun Chen ◽  
Jing Jin ◽  
Xin Huang ◽  
Fan Wu ◽  
Hongguang Huang ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Modulation ◽  
Tumour Progression ◽  
Critical Factor ◽  
Bioinformatic Analysis ◽  
Cell Infiltration ◽  
Effective Response ◽  
T Cell Infiltration

Abstract Background The immunosuppressive tumour microenvironment is a critical factor in the initiation and progression of glioblastoma (GBM), which is characterized by an abundance of tumour-associated macrophages (TAMs) but a paucity of infiltrating T cells. In this research, we studied whether epithelial membrane protein 3 (EMP3) plays a crucial role in immune modulation in GBM. Methods TCGA and CGGA transcriptomic profiles of wild-type IDH1 GBM were used for bioinformatic analysis. The role of EMP3 in GBM was validated through in vivo and in vitro experiments. Human GBM specimens were collected and evaluated using immunofluorescence analysis. Results EMP3 was associated with immunosuppression in GBM. Elevated EMP3 in GBM areas was accompanied by high expression of PD-L1 and abundant M2 TAM recruitment but a lake of T cell infiltration. We found that EMP3 was a potent protein in M2 TAM polarization and recruitment that impaired the ability of GBM cells to secrete CCL2 and TGF-β1. Furthermore, EMP3 suppressed T cell infiltration into GBM tumours by inhibiting the secretion of CXCL9 and CXCL10 by macrophages and led to an effective response to anti-PD1 therapy. Conclusions EMP3 is thus a critical immunosuppressive factor for recruiting TAMs in GBM and suppressing intratumoural T cell infiltration to facilitate tumour progression and is a potential therapeutic target.

scholarly journals In vivo imaging of cytotoxic T cell infiltration and elimination of a solid tumor

2007 ◽  
Vol 176 (4) ◽  
pp. i8-i8
Author(s):  
Alexandre Boissonnas ◽  
Luc Fetler ◽  
Ingrid S. Zeelenberg ◽  
Stéphanie Hugues ◽  
Sebastian Amigorena
Keyword(s):  
T Cell ◽  
Solid Tumor ◽  
In Vivo Imaging ◽  
Cell Infiltration ◽  
Cytotoxic T Cell ◽  
T Cell Infiltration

scholarly journals Male Bovine GH Transgenic Mice Have Decreased Adiposity With an Adipose Depot-Specific Increase in Immune Cell Populations

Endocrinology ◽  
2014 ◽  
Vol 156 (5) ◽  
pp. 1794-1803 ◽  
Author(s):  
Fabian Benencia ◽  
Stephanie Harshman ◽  
Silvana Duran-Ortiz ◽  
Ellen R. Lubbers ◽  
Edward O. List ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Cell ◽  
Stromal Vascular Fraction ◽  
Cell Infiltration ◽  
Cell Populations ◽  
Adipose Depot ◽  
T Cell Infiltration ◽  
Specific Manner ◽  
In Vivo Effects

White adipose tissue (WAT) is composed of mature adipocytes and a stromal vascular fraction (SVF), which contains a variety of cells, including immune cells that vary among the different WAT depots. Growth hormone (GH) impacts immune function and adiposity in an adipose depot-specific manner. However, its effects on WAT immune cell populations remain unstudied. Bovine GH transgenic (bGH) mice are commonly used to study the in vivo effects of GH. These giant mice have an excess of GH action, impaired glucose metabolism, decreased adiposity, increased lean mass, and a shortened lifespan. Therefore, the purpose of this study was to characterize the WAT depot-specific differences in immune cell populations in the presence of excess GH in vivo. Three WAT depots were assessed: inguinal (sc), epididymal (EPI), and mesenteric (MES). Subcutaneous and MES bGH WAT depots showed a significantly higher number of total SVF cells, yet only MES bGH WAT had higher leukocyte counts compared with control samples. By means of flow cytometry analysis of the SVF, we detected greater macrophage and regulatory T-cell infiltration in sc and MES bGH WAT depots compared with controls. However, no differences were observed in the EPI WAT depot. RNA-sequencing confirmed significant alterations in pathways related to T-cell infiltration and activation in the sc depot with fewer significant changes in the EPI bGH WAT depot. These findings collectively point to a previously unrecognized role for GH in influencing the distribution of WAT immune cell populations in a depot-specific manner.

88 Development of a 3D organoid autologous TIL co-culture platform for high throughput immuno-oncology studies

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A98-A98
Author(s):  
Garima Kaushik ◽  
Bhavna Verma ◽  
Amy Wesa
Keyword(s):  
T Cell ◽  
High Throughput ◽  
Fluorescent Dyes ◽  
Tumor Infiltrating Lymphocytes ◽  
Cell Infiltration ◽  
Patient Specific ◽  
T Cell Infiltration ◽  
Organoid Cultures ◽  
Infiltrating Lymphocytes

BackgroundThe preclinical screening of immune-modulatory therapies suffers from the absence of models that recapitulate in vivo heterogeneous tumor microenvironment (TME). 3D tumor organoid cultures provide a model that closely mimics in situ tumor architecture and is being aggressively used to evaluate therapeutic efficacy ex vivo. A vastly heterogenous TME impacts patient treatment response, and there is a dearth of human tumor models (2D or 3D), that mimic in vivo diversity of TME, including infiltrating immune populations. 3D organoid cultures typically contain neoplastic epithelium; however, they fall short in representing tumor to tumor-infiltrating lymphocytes (TILs) interactions, limiting their ability to generate a clinically relevant response to immunotherapeutics. Addition of immune cells from unrelated donors to organoids can simulate that microenvironment but is complicated by T cell alloreactivity. Here we describe 3D patient-derived xenograft organoid (PDXO) co-cultures with matching autologous human TILs to recapitulate the tumor-specific immune response, leveraging confocal high content analysis and luminex multiplex assays. This platform allows the evaluation and high throughput screening of novel immune targeting agents to determine impacts on patient-derived T cell function, T cell infiltration, and tumor cytotoxicity.MethodsSurgical resections from patients were used to generate patient-derived xenografts and tumor-infiltrating lymphocytes in parallel. PDX were resected and digested to establish PDXO. TILs and organoids from the same patient were fluorescent labeled and cultured together for four days to evaluate tumor infiltration and drug cytotoxicity in 3D cultures. CellInsight CX7 high content imaging platform was used to trace TILs and cancer cells and evaluate T cell infiltration and tumor cell killing in the presence and absence of immuno-modulatory therapies.ResultsPDXO were established to mimic in vivo tumor biology. Tumor-specific TILs were successfully expanded and characterized by flow cytometry. Co-culture resulted in TIL infiltration in organoids from day one in culture and increased over four days. Cytotoxicity and TIL infiltration were quantified using fluorescent dyes via high throughput imaging platform. Significantly enhanced TIL infiltration was observed in autologous co-cultures compared to non-autologous co-cultures. The established unique autologous PDXO immune organoid co-cultures could be used as an improved simulation of the modulatory activity of therapeutic agents in patient-specific T cells against their own tumors.ConclusionsPatient autologous TILs – PDXO co-culture platform is an advanced model for evaluating IO therapeutics with the tumor-specific immune microenvironment. The platform provides an opportunity for precision medicine and high throughput drug screening of immuno-modulatory therapies.Ethics ApprovalThe study was approved by Champions Oncology’s Institutional Animal Care and Use Committee (IACUC).

scholarly journals 404 Initial biomarker and clinical data of a phase 2a study of NT-I7, a long-acting interleukin-7, plus pembrolizumab: cohort of subjects with checkpoint inhibitor-naïve advanced MSS-colorectal cancer

2021 ◽  
Vol 9 (Suppl 3) ◽  
pp. A435-A435
Author(s):  
Richard Kim ◽  
Minal Barve ◽  
Hirva Mamdani ◽  
Melissa Johnson ◽  
Byung Ha Lee ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
T Cell ◽  
Checkpoint Inhibitor ◽  
Cd8 T Cell ◽  
T Cell Subsets ◽  
Cell Infiltration ◽  
T Cell Infiltration ◽  
Long Acting ◽  
Anti Tumor Activity

BackgroundCheckpoint inhibitor (CPI) monotherapy is ineffective for microsatellite stable colorectal cancer (MSS-CRC). NT-I7 (efineptakin alfa) is the first-in-class long-acting IL-7 that can increase T-cell infiltration in the tumor microenvironment (TME). We hypothesize that NT-I7 may create a favorable immune-reactive TME to enhance the efficacy of CPI when combined with pembrolizumab (pembro).MethodsThis is an open-label, phase 2a study in subjects with relapsed/refractory (R/R) tumors, including CPI-naïve R/R MSS-CRC. Subjects received the recommended-phase-2-dose of NT-I7 intramuscularly at 1200 µg/kg every 6 weeks (Q6W) plus pembro 200 mg intravenously Q3W. Preliminary anti-tumor activity based on Overall Response Rate (ORR) was assessed by Response Evaluation Criteria in Solid Tumors (RECIST) v1.1 as a primary objective and by iRECIST as an exploratory objective. Biomarker analyses in peripheral blood and tumor biopsies were performed.ResultsAs of 15-July-2021, 19 subjects were enrolled in the CPI-naïve R/R MSS-CRC cohort. Six subjects are ongoing. Median age 58 years [37–81], ECOG PS 0 (26%), 1 (74%). Sixteen (84%) subjects received ≥ 2 prior therapies. All subjects had metastatic or locally advanced disease at enrollment. The median duration of follow-up was 4.64 months. Among 15 evaluable subjects, disease-control rate (DCR) based on RECIST1.1 was 47% and 1 subject achieved partial response per iRECIST (iPR) with 33% tumor reduction. Treatment-related adverse events (AEs) occurred in 14 (73.7%) subjects, 9 (47.4%) G1–2 events and 5 (26.3%) G3 events; no G4 or G5 AEs were reported. No subjects discontinued from the study due to AE. NT-I7 + pembro elicited a significant increase in the absolute lymphocyte count that peaked at week 3 (>3X from baseline, p<0.0001) and was sustained at least until week 18. CD4+/CD8+ T-cell subsets followed the same response pattern. Importantly, Stem-Cell Memory CD8+ T-cells (TSCM), the potential target for CPIs that differentiate into effectors, were remarkably increased post-study treatment (>25X from baseline, p<0.01). Plasmatic chemokines (CXCL9, CXCL10, CXCL11 and CCL9) were significantly increased after the first dose. The iPR subject had an enhanced T-cell infiltration in the TME at week 5. Subject’s follow-up continues and more updated data will be presented.ConclusionsThe chemo-free combination of NT-I7 + pembro was well tolerated and showed encouraging anti-tumor activity in subjects with CPI-naïve R/R MSS-CRC. Increased TSCM and CD8+ T-cell infiltration in TME may be the underlying mechanisms of action for the observed efficacy. These results support continued evaluation of NT-I7 + pembro in CPI-naïve subjects with R/R MSS-CRC.AcknowledgementsThe authors thank ICON for their partnership in conducting this trial.Trial RegistrationNCT04332653Ethics ApprovalThe trial was approved by MD Anderson IRB (#2020–0008_MOD001), Mary Crowley IRB (#20–13) and Advarra IRB (#Pro00042639)All participant gave informed consent prior to study enrollment.

scholarly journals B7 immune-checkpoints as targets for the treatment of neuroendocrine tumors

2021 ◽  
Author(s):  
Ziqiang Yuan ◽  
Juliet C Gardiner ◽  
Elaine C Maggi ◽  
Shuyu Huang ◽  
Asha Adem ◽  
...  
Keyword(s):  
T Cell ◽  
Neuroendocrine Tumors ◽  
T Cell Activation ◽  
Cell Infiltration ◽  
Immune Checkpoints ◽  
T Cell Infiltration ◽  
In Vivo Animal Model ◽  
B7 Family

The B7 family, and their receptors, the CD28 family, are major immune checkpoints that regulate T-cell activation and function. In the present study, we explore the role of two B7 immune-checkpoints: HERV-H LTR-Associating Protein 2 (HHLA2) and B7 Family Member, H4 (B7x), in the progression of gastrointestinal and pancreatic neuroendocrine tumors (GINETs and PNETs). We demonstrated that both HHLA2 and B7x were expressed to a high degree in human GINETs and PNETs. We determined that the expression of B7x and HHLA2 correlates with higher grade and higher incidence of nodal and distant spread. Furthermore, we confirmed that HIF-1 overexpression is associated with the upregulation of B7x both in our in vivo (animal model) and in vitro (cell culture) models. When grown in vitro, islet tumor β-cells lack B7x expression, unless cultured under hypoxic conditions, which results in both hypoxia inducible factor 1 subunit alpha (HIF-1α) and B7x upregulation. In vivo, we demonstrated that Men1/B7x double knockout (KO) mice (with loss of B7x expression) exhibited decreased islet β-cell proliferation and tumor transformation accompanied by increased T-cell infiltration compared with Men1 single knockout mice. We have also shown that systemic administration of a B7x mAb to our Men1 KO mice with PNETs promotes an antitumor response mediated by increased T-cell infiltration. These findings suggest that B7x may be a critical mediator of tumor immunity in the tumor microenvironment of NETs. Therefore, targeting B7x offers an attractive strategy for the immunotherapy of patients suffering from NETs.

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