CD11c+CD163+ Cells and Signal Transducer and Activator of Transcription 3 (STAT3) Expression Are Common in Melanoma Leptomeningeal Disease

Leptomeningeal disease (LMD) in melanoma patients is associated with significant neurological sequela and has a dismal outcome, with survival measured typically in weeks. Despite the therapeutic benefit of targeted therapies and immunotherapies for Stage IV melanoma, patients with LMD do not typically benefit. A deeper understanding of the tumor microenvironment (TME) of LMD may provide more appropriate therapeutic selection. A retrospective analysis of subjects who underwent surgical resection with LMD (n=8) were profiled with seven color multiplex staining to evaluate the expression of the global immune suppressive hub - the signal transducer and activator of transcription 3 (STAT3) and for the presence of CD3+ T cells, CD68+ monocyte-derived cells, CD163+ immune suppressive macrophages, and CD11c+ cells [potential dendritic cells (DCs)] in association with the melanoma tumor marker S100B and DAPI for cellular nuclear identification. High-resolution cellular imaging and quantification was conducted using the Akoya Vectra Polaris. CD11c+ cells predominate in the TME (10% of total cells), along with immunosuppressive macrophages (2%). Another potential subset of DCs co-expressing CD11c+ and the CD163+ immunosuppressive marker is frequently present (8/8 of specimens, 8%). Occasional CD3+ T cells are identified, especially in the stroma of the tumor (p=0.039). pSTAT3 nuclear expression is heterogeneous in the various immune cell populations. Occasional immune cluster interactions can be seen in the stroma and on the edge. In conclusion, the TME of LMD is largely devoid of CD3+ T cells but is enriched in immune suppression and innate immunity.

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LMD-20. Immune Suppressive Macrophages and Signal Transducer and Activator of Transcription 3 (STAT3) Expression are common in Melanoma Leptomeningeal Disease

Neuro-Oncology Advances ◽

10.1093/noajnl/vdab071.045 ◽

2021 ◽

Vol 3 (Supplement_3) ◽

pp. iii11-iii12

Author(s):

Hinda Najem ◽

Anantha Marisetty ◽

Craig Horbinski ◽

Jared Burks ◽

Amy B Heimberger

Keyword(s):

T Cells ◽

Immune Cell ◽

Stage Iv ◽

Tumor Stroma ◽

Leptomeningeal Disease ◽

Signal Transducer ◽

Melanoma Tumor ◽

Immune Cell Population ◽

Melanoma Patients ◽

Transcription 3

Abstract Leptomeningeal disease (LMD) in melanoma patients is associated with significant neurological impairments and has a dismal outcome with a median survival of 1.8 months. Despite the therapeutic benefit of targeted therapies and immunotherapies for most kinds of Stage IV melanoma, patients with LMD do not typically benefit. A deeper understanding of the tumor microenvironment (TME) of LMD may provide more appropriate therapeutic selection. A retrospective analysis of subjects who underwent surgical resection with LMD (n=8) were profiled with seven color multiplex to evaluate the expression of the global immune suppressive hub - the signal transducer and activator of transcription 3 (STAT3) and for the presence of CD3 T cells, CD68+ monocytes, CD163 immune suppressive macrophages, CD11c+ antigen presenting cells (APCs) in association with the melanoma tumor marker S100B and DAPI for cellular nuclear identification. High-resolution cellular imaging and quantification was conducted using the Akoya Vectra Polaris. CD163+ macrophage is the most frequent immune cell population in the LMD TME. Occasional CD3+ T cells and CD11c+ APC are also identified, although the latter has concurrent expression of CD163. STAT3 nuclear localization is heterogeneously expressed in the various immune cell populations. Occasional immune cluster interactions can be seen in the tumor stroma and the tumor edge. In conclusion, the TME of LMD is largely devoid of CD3+ T cells, but is enriched for immune suppression and innate immunity.

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LKB1 deficiency in T cells promotes the development of gastrointestinal polyposis

Science ◽

10.1126/science.aan3975 ◽

2018 ◽

Vol 361 (6400) ◽

pp. 406-411 ◽

Cited By ~ 10

Author(s):

M. C. Poffenberger ◽

A. Metcalfe-Roach ◽

E. Aguilar ◽

J. Chen ◽

B. E. Hsu ◽

...

Keyword(s):

T Cells ◽

Cancer Progression ◽

Immune Cell ◽

Inflammatory Factors ◽

Gastrointestinal Polyposis ◽

Cancer Predisposition Syndrome ◽

Stat3 Signaling ◽

Liver Kinase B1 ◽

Peutz Jeghers Syndrome ◽

Transcription 3

Germline mutations in STK11, which encodes the tumor suppressor liver kinase B1 (LKB1), promote Peutz–Jeghers syndrome (PJS), a cancer predisposition syndrome characterized by the development of gastrointestinal (GI) polyps. Here, we report that heterozygous deletion of Stk11 in T cells (LThet mice) is sufficient to promote GI polyposis. Polyps from LThet mice, Stk11+/− mice, and human PJS patients display hallmarks of chronic inflammation, marked by inflammatory immune-cell infiltration, signal transducer and activator of transcription 3 (STAT3) activation, and increased expression of inflammatory factors associated with cancer progression [interleukin 6 (IL-6), IL-11, and CXCL2]. Targeting either T cells, IL-6, or STAT3 signaling reduced polyp growth in Stk11+/− animals. Our results identify LKB1-mediated inflammation as a tissue-extrinsic regulator of intestinal polyposis in PJS, suggesting possible therapeutic approaches by targeting deregulated inflammation in this disease.

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Exploiting the STAT3 Nexus in Cancer-Associated Fibroblasts to Improve Cancer Therapy

Frontiers in Immunology ◽

10.3389/fimmu.2021.767939 ◽

2021 ◽

Vol 12 ◽

Author(s):

Amr Allam ◽

Marina Yakou ◽

Lokman Pang ◽

Matthias Ernst ◽

Jennifer Huynh

Keyword(s):

Tumor Cells ◽

Immune Suppression ◽

Immune Cells ◽

Therapeutic Benefit ◽

Cancer Associated Fibroblasts ◽

Signal Transducer ◽

Growing Body ◽

Heterogenous Population ◽

Transcription 3 ◽

Soluble Components

The tumor microenvironment (TME) is composed of a heterogenous population of cells that exist alongside the extracellular matrix and soluble components. These components can shape an environment that is conducive to tumor growth and metastatic spread. It is well-established that stromal cancer-associated fibroblasts (CAFs) in the TME play a pivotal role in creating and maintaining a growth-permissive environment for tumor cells. A growing body of work has uncovered that tumor cells recruit and educate CAFs to remodel the TME, however, the mechanisms by which this occurs remain incompletely understood. Recent studies suggest that the signal transducer and activator of transcription 3 (STAT3) is a key transcription factor that regulates the function of CAFs, and their crosstalk with tumor and immune cells within the TME. CAF-intrinsic STAT3 activity within the TME correlates with tumor progression, immune suppression and eventually the establishment of metastases. In this review, we will focus on the roles of STAT3 in regulating CAF function and their crosstalk with other cells constituting the TME and discuss the utility of targeting STAT3 within the TME for therapeutic benefit.

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Deciphering Treg accumulation in melanoma.

Journal of Clinical Oncology ◽

10.1200/jco.2013.31.15_suppl.e22170 ◽

2013 ◽

Vol 31 (15_suppl) ◽

pp. e22170-e22170

Author(s):

Kristen E. Tobin Vealey ◽

I. Caroline Le Poole ◽

Jared S. Klarquist ◽

Jonathan M. Eby

Keyword(s):

T Cells ◽

Metastatic Melanoma ◽

Chemokine Receptor ◽

Chemokine Receptors ◽

Intervention Strategy ◽

Host Immune System ◽

Healthy Controls ◽

Receptor Signaling ◽

Melanoma Tumor ◽

Melanoma Patients

e22170 Background: A high density of regulatory T-cells (Tregs) at melanoma tumor sites correlates with poor prognosis due to immunosuppressive effects on the host immune system. While the concentration of circulating Tregs in patients with melanoma is not significantly different from that of healthy controls, Tregs tend to accumulate at tumor sites. We hypothesized that the influx of Tregs observed in melanoma tumors may be explained by enhanced tumor homing. Methods: To quantify the relative expression of chemokine receptors CCL4 and CCL8, and skin homing receptor CLA, blood was drawn from five metastatic melanoma patients and five healthy controls. Lymphocytes were ficolled and negatively sorted for CD3 expression. T-cells were subjected to 7-color FACS staining where Tregs were defined as CD4+CD25+CD127lowFoxP3+ and were simultaneously assessed for expression of CCR4, CCR8, and CLA. To further understand chemokine-receptor signaling involved in Treg homing in melanoma we analyzed the number of CCL22 producing cells in four human metastatic melanoma tissues and four healthy skin tissues obtained during surgery. Dermal CCL22+ cells were detected by indirect immunohistochemistry and quantified to a depth of three times the thickness of the epidermis. Results: Lymphocytes of melanoma patients were found to exhibit enhanced expression of CCR4 (mean of 65.9 versus 51.2%, p<0.01) and CCR8 (3.0 versus 1.5%, p<0.05), but not CLA (21.9 versus 23.9%) as compared with healthy controls. The mean fluorescence intensities of CCR4 and CCR8 were likewise elevated by 63 and 49% over controls (p<0.05 and p<0.01), respectively. A markedly increased number of CCL22 secreting cells were found in tumor samples (mean of 1063.1 versus 207.4, p= 0.02) as compared with healthy controls. Conclusions: These data strongly suggest that enhanced homing of CCR4+ Tregs towards the ligand CCL22 is likely involved in accumulating Tregs at melanoma tumor sites, ultimately impeding effective anti-tumor immune responses. Targeting chemokine-receptor signaling may thus be an effective intervention strategy in melanoma.

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

Journal of Clinical Oncology ◽

10.1200/jco.2021.39.15_suppl.e14532 ◽

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.

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Prolonged Survival of Dendritic Cell–Vaccinated Melanoma Patients Correlates With Tumor-Specific Delayed Type IV Hypersensitivity Response and Reduction of Tumor Growth Factor β-Expressing T Cells

Journal of Clinical Oncology ◽

10.1200/jco.2008.18.0794 ◽

2009 ◽

Vol 27 (6) ◽

pp. 945-952 ◽

Cited By ~ 97

Author(s):

Mercedes N. López ◽

Cristian Pereda ◽

Gabriela Segal ◽

Leonel Muñoz ◽

Raquel Aguilera ◽

...

Keyword(s):

Immune Response ◽

Overall Survival ◽

T Cells ◽

Growth Factor ◽

Regulatory T Cells ◽

Disease Progression ◽

Stage Iv ◽

Stage Iii ◽

Type Iv ◽

Melanoma Patients

PurposeThe aim of this work was to assess immunologic response, disease progression, and post-treatment survival of melanoma patients vaccinated with autologous dendritic cells (DCs) pulsed with a novel allogeneic cell lysate (TRIMEL) derived from three melanoma cell lines.Patients and MethodsForty-three stage IV and seven stage III patients were vaccinated four times with TRIMEL/DC vaccine. Specific delayed type IV hypersensitivity (DTH) reaction, ex vivo cytokine production, and regulatory T-cell populations were determined. Overall survival and disease progression rates were analyzed using Kaplan-Meier curves and compared with historical records.ResultsThe overall survival for stage IV patients was 15 months. More than 60% of patients showed DTH-positive reaction against the TRIMEL. Stage IV/DTH-positive patients displayed a median survival of 33 months compared with 11 months observed for DTH-negative patients (P = .0014). All stage III treated patients were DTH positive and remained alive and tumor free for a median follow-up period of 48 months (range, 33 to 64 months). DTH-positive patients showed a marked reduction in the proportion of CD4+ transforming growth factor (TGF) β+ regulatory T cells compared to DTH-negative patients (1.54% v 5.78%; P < .0001).ConclusionOur findings strongly suggest that TRIMEL-pulsed DCs provide a standardized and widely applicable source of melanoma antigens, very effective in evoking antimelanoma immune response. To our knowledge, this is the first report describing a correlation between vaccine-induced reduction of CD4+TGFβ+ regulatory T cells and in vivo antimelanoma immune response associated to improved patient survival and disease stability.

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Hydroxychloroquine (HCQ) reverses anti-PD-1 immune murine checkpoint blockade: TCF1 as a marker in humans for COVID-19 and HCQ therapy

10.1101/2020.09.29.20193110 ◽

2020 ◽

Author(s):

Janna Kreuger ◽

Francois Santinon ◽

Alexandra Kazanova ◽

Mark Issa ◽

Bruno Larrivee ◽

...

Keyword(s):

Public Health ◽

T Cells ◽

Cd8 T Cells ◽

Deleterious Effect ◽

Therapeutic Benefit ◽

Tumor Rejection ◽

Checkpoint Blockade ◽

Global Public Health ◽

Melanoma Tumor ◽

First Time

Coronavirus disease 2019 (COVID-19) pandemic caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a serious threat to global public health. Hydroxychloroquine (HCQ) and the antibiotic azithromycin (AZ) are still being used by thousands and numerous hospitals to treat COVID-19. In a related context, immunotherapy using checkpoint blockade (ICB) with antibodies such as anti-PD-1 has revolutionised cancer therapy. Given that cancer patients on ICB continue to be infected with SARS-CoV-2, an understanding of the effects of HCQ and AZ on the elimination of tumors by anti-PD-1 ICB is urgently needed. In this study, we report that HCQ alone, or in combination with AZ, at doses used to treat COVID-19 patients, reverses the therapeutic benefit of anti-PD-1 in controlling B16 melanoma tumor growth in mice. No deleterious effect was seen on untreated tumors, or in using AZ alone in anti-PD-1 immunotherapy. Mechanistically, HCQ and HCQ/AZ inhibited PD-L1 expression on tumor cells, while specifically targeting the anti-PD-1 induced increase in progenitor CD8+CD44+PD-1+TCF1+ tumor-infiltrating T-cells (TILs) and the generation of CD8+CD44+PD-1+ effectors. Surprisingly, it also blocked the appearance of a subset of terminally exhausted CD8+ TILs. No effect was seen on the presence of CD4+ T-cells, FoxP3+ Tregs, thymic subsets, B-cells, antibody production, myeloid cells, or the vasculature of mice. Lastly, we identified TCF-1 expression in peripheral CD8+ T-cells from cancer or non-cancer human patients infected with SARs CoV2 as a marker for the effects of COVID-19 and HCQ on the immune system. This study indicates for the first time that HCQ and HCQ/AZ negatively impact the ability of anti-PD-1 checkpoint blockade to promote tumor rejection.

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Evaluation of the Melanoma Tumor Burden Score (MTBS) in a real-world setting.

Journal of Clinical Oncology ◽

10.1200/jco.2017.35.15_suppl.9565 ◽

2017 ◽

Vol 35 (15_suppl) ◽

pp. 9565-9565 ◽

Cited By ~ 2

Author(s):

Michael Weichenthal ◽

Gamze Senel ◽

Marcus Both ◽

Ilka Senkpiehl ◽

Dragana Lekic ◽

...

Keyword(s):

Metastatic Melanoma ◽

Stage Iv ◽

Tumor Burden ◽

Sensitivity Analyses ◽

Melanoma Tumor ◽

Clinical Cancer ◽

Metastatic Lesions ◽

Tumor Load ◽

Melanoma Patients ◽

Stage Iv Melanoma

9565 Background: Clinical cancer registration is increasingly important for healthcare delivery and outcome research in oncology. As compared to clinical trial data, information from clinical routine is often limited regarding the granularity and quality of measures for individual tumor load and distribution. Methods: In an effort to implement a robust and useful measure of tumor burden for patients with metastatic melanoma in a German national skin cancer registry (ADOReg) we evaluated the melanoma tumor burden score (MTBS), originally developed for analyzing chemotherapy data in melanoma patients. The MTBS contains a simple categorization of size, number an distribution of metastatic lesions in individual patients. It is aimed at being used on routine radiologic report allowing for a certain level of uncertainty and imprecise quantification of metastatic lesions. Basically, the lesions are categorized per affected organ with respect to number (solitary, few, multiple) and size (≤1cm, >1- 5cm, >5cm). For evaluation of prognostic significance the summary score was calculated and included in univariate and multivariate survival analysis. We performed extensive sensitivity analyses for a variety of different model settings. Results: In the primary analysis set we re-evaluated 898 radiologic reports in a total of 235 various chemotherapies in n=128 stage IV melanoma patients. The confirmatory data sets consisted of n=384 stage IV melanoma patients with various treatments including chemotherapy, BRAF inhibitor treatment, and immune checkpoint blockade. MTBS categorization could be applied on routine radiologic reports in the majority of cases (95.7 %). In a multivariate model MTBS remained significantly correlated with outcome when adjusted for age, sex, LDH, and number of metastatic sites. Moreover, change in MTBS correlated to a formal response evaluation according to RECIST. Conclusions: The MTBS appears to be a promising tool for meaninful quantification of metastatic tumor load in metastatic melanoma for real life data collection like in clinical cancer registries. [Table: see text]

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Peripheral CD8 effector-memory type 1 T-cells correlate with outcome in ipilimumab-treated stage IV melanoma patients

European Journal of Cancer ◽

10.1016/j.ejca.2016.12.011 ◽

2017 ◽

Vol 73 ◽

pp. 61-70 ◽

Cited By ~ 33

Author(s):

Kilian Wistuba-Hamprecht ◽

Alexander Martens ◽

Florian Heubach ◽

Emanuela Romano ◽

Marnix Geukes Foppen ◽

...

Keyword(s):

T Cells ◽

Stage Iv ◽

Effector Memory ◽

Melanoma Patients ◽

Memory Type ◽

Stage Iv Melanoma

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STAT3, a Master Regulator of Anti-Tumor Immune Response

Cancers ◽

10.3390/cancers11091280 ◽

2019 ◽

Vol 11 (9) ◽

pp. 1280 ◽

Cited By ~ 19

Author(s):

Rébé ◽

Ghiringhelli

Keyword(s):

Cancer Progression ◽

Immune Cells ◽

Cell Function ◽

Immune Cell ◽

Checkpoint Inhibitors ◽

Cancer Growth ◽

Signal Transducer ◽

Immune Cell Function ◽

Transcription 3

Immune cells in the tumor microenvironment regulate cancer growth. Thus cancer progression is dependent on the activation or repression of transcription programs involved in the proliferation/activation of lymphoid and myeloid cells. One of the main transcription factors involved in many of these pathways is the signal transducer and activator of transcription 3 (STAT3). In this review we will focus on the role of STAT3 and its regulation, e.g. by phosphorylation or acetylation in immune cells and how it might impact immune cell function and tumor progression. Moreover, we will review the ability of STAT3 to regulate checkpoint inhibitors.

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