A standardized analysis of tertiary lymphoid structures in human melanoma: disease progression- and tumor site-associated changes with germinal center alteration

2021 ◽  
Author(s):  
Franziska Werner
Keyword(s):  
Disease Progression ◽  
Germinal Center ◽  
Human Melanoma ◽  
Tumor Site ◽  
Tertiary Lymphoid Structures ◽  
Lymphoid Structures

scholarly journals A Standardized Analysis of Tertiary Lymphoid Structures in Human Melanoma: Disease Progression- and Tumor Site-Associated Changes With Germinal Center Alteration

2021 ◽  
Vol 12 ◽  
Author(s):  
Franziska Werner ◽  
Christine Wagner ◽  
Martin Simon ◽  
Katharina Glatz ◽  
Kirsten D. Mertz ◽  
...  
Keyword(s):  
Spatial Distribution ◽  
Disease Progression ◽  
Germinal Center ◽  
Human Melanoma ◽  
Melanoma Tumor ◽  
Invasive Tumor ◽  
Evaluation Approach ◽  
Tertiary Lymphoid Structures ◽  
Lymphoid Structures ◽  
And Function

There is increasing evidence that tertiary lymphoid structures (TLS) control not only local adaptive B cell responses at melanoma tumor sites but also the cellular composition and function of other immune cells. In human melanoma, however, a comprehensive analysis of TLS phenotypes, density and spatial distribution at different disease stages is lacking. Here we used 7-color multiplex immunostaining of whole tissue sections from 103 human melanoma samples to characterize TLS phenotypes along the expression of established TLS-defining molecular and cellular components. TLS density and spatial distribution were determined by referring TLS counts to the tissue area within defined intra- and extratumoral perimeters around the invasive tumor front. We show that only a subgroup of primary human melanomas contains TLS. These TLS rarely formed germinal centers and mostly located intratumorally within 1 mm distance to the invasive tumor front. In contrast, melanoma metastases had a significantly increased density of secondary follicular TLS. They appeared preferentially in stromal areas within an extratumoral 1 mm distance to the invasive tumor front and their density varied over time and site of metastasis. Interestingly, secondary follicular TLS in melanoma often lacked BCL6+ lymphatic cells and canonical germinal center polarity with the formation of dark and light zone areas. Our work provides an integrated qualitative, quantitative and spatial analysis of TLS in human melanoma and shows disease progression- and site-associated changes in TLS phenotypes, density and spatial distribution. The frequent absence of canonical germinal center polarity in melanoma TLS highlights the induction of TLS maturation as a potential additive to future immunotherapy studies. Given the variable evaluation strategies used in previous TLS studies of human tumors, an important asset of this study is the standardized quantitative evaluation approach that provides a high degree of reproducibility.

536 Divergent cancer etiologies drive distinct B cell signatures and tertiary lymphoid structures in head and neck cancer

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A572-A572
Author(s):  
Ayana Ruffin ◽  
Anthony Cillo ◽  
Tracy Tabib ◽  
Angen Liu ◽  
Sayali Onkar ◽  
...  
Keyword(s):  
T Cells ◽  
B Cells ◽  
B Cell ◽  
Head And Neck ◽  
Germinal Center ◽  
Spectral Unmixing ◽  
Cell Phenotype ◽  
Primary Tumors ◽  
Tertiary Lymphoid Structures ◽  
Lymphoid Structures

BackgroundCurrent FDA-approved immunotherapies aim to reinvigorate CD8+ T cells, but the contribution of the humoral arm of the immune response in human cancer remains poorly understood. B cells within tissues can mediate anti-tumor immunity and regulate immune responses by presenting antigen and producing tumor-specific antibodies and immunomodulatory cytokines. Head and neck squamous cell carcinoma (HNSCC) can be induced by human papillomavirus (HPV+) and carcinogens such as tobacco and alcohol (HPV-), and the immune infiltrate is quite distinct in the two etiologies, in particular, increased B cells in HPV+ HNSCC patients. Further, increased B cells in HNSCC patients correlate with improved patient survival. Our study seeks to differentiate B cell phenotype, function and location in HPV+ and HPV- HNSCC to identify putative B cell-centric immunotherapeutic targets.MethodsWe utilized a multi-level approach to clearly categorize B cells in HNSCC patients. Single cell RNA sequencing (scRNAseq) was performed on CD45+ tumor infiltrating lymphocytes (TIL) from HPV+ and HPV- HNSCC patients. HNSCC TIL and PBL were stained via spectral cytometry (Cytek Aurora,25 parameters) for unbiased analysis of B cell subsets via computational spectral unmixing. Paraffin embedded slides from HNSCC primary tumors were utilized for multispectral immunofluorescence (mIF) to identify tertiary lymphoid structures (TLS) and identify differences in HPV+ and HPV- disease.ResultsWe demonstrated distinct trajectories for B cells in HPV+ and HPV- disease. HPV- HNSCC tumors mainly contained memory B cells and plasma cells, while the B cells in HPV+ HNSCC were naïve and germinal center (GC). Further, we quantified B cells and CD4+ T cells in TLS, and germinal center-like TLS were associated with improved outcome in HPV+ disease. We also observed that transcriptional and protein expression of Semaphorin A (SEMA4a) was restricted to GC B cells and increased on GC B cells in HNSCC patients compared to healthy tonsils. Additionally, we identified distinct waves of gene expression in GC B cells in HNSCC tumors, ultimately revealing a novel transitional state for GC B cells in the tumor microenvironment (TME).ConclusionsUnderstanding B cell function in human cancers and how different TMEs influence B cells and TLS are important for devising novel therapeutic options for cancer patients. Ultimately, development of therapeutics to enhance B cell responses in the TME should be prioritized as a compliment to T-cell mediated therapies.

scholarly journals Germinal center reactions in tertiary lymphoid structures associate with neoantigen burden, humoral immunity and long-term survivorship in pancreatic cancer

2021 ◽  
Vol 10 (1) ◽  
pp. 1900635
Author(s):  
Andrew J. Gunderson ◽  
Venkatesh Rajamanickam ◽  
Cynthia Bui ◽  
Brady Bernard ◽  
Joanna Pucilowska ◽  
...  
Keyword(s):  
Pancreatic Cancer ◽  
Humoral Immunity ◽  
Germinal Center ◽  
Tertiary Lymphoid Structures ◽  
Lymphoid Structures

scholarly journals CXCL13 Neutralization Attenuates Neuropsychiatric Manifestations in Lupus-Prone Mice

2021 ◽  
Vol 12 ◽  
Author(s):  
Michelle W. Huang ◽  
Ariel D. Stock ◽  
Chaim Putterman
Keyword(s):  
Disease Progression ◽  
Lupus Erythematosus ◽  
Lymphoid Follicle ◽  
Beneficial Effects ◽  
Brain Gene Expression ◽  
Potential Contributor ◽  
Neuropsychiatric Manifestations ◽  
Tertiary Lymphoid Structures ◽  
Neurobehavioral Manifestations ◽  
Lymphoid Structures

Neuropsychiatric lupus (NPSLE), the nervous system presentation of systemic lupus erythematosus (SLE), remains challenging to treat due to its unclear pathogenesis and lack of available targeted therapies. A potential contributor to disease progression is brain tertiary lymphoid structures (TLS); these ectopic lymphoid follicles that can develop tissue-targeted antibodies have recently been described in the MRL/lpr lupus mouse strain, a classic model for studying NPSLE. The brains of MRL/lpr mice show a significant increase of CXCL13, an important chemokine in lymphoid follicle formation and retention that may also play a role in the disease progression of NPSLE. The aim of the present study was to inhibit CXCL13 and examine the effect of this intervention on lymphoid formation and the development of neurobehavioral manifestations in lupus mice. Female MRL/lpr mice were injected with an anti-CXCL13 antibody, an IgG1 isotype-matched antibody, or PBS either three times a week for 12 weeks intraperitoneally (IP) starting at 6-8 weeks of age, or continuously intracerebroventricularly (ICV) with an osmotic pump over a two-week period starting at 15 weeks of age. Cognitive dysfunction and depression-like behavior were assessed at the end of treatment. When treatment was delivered IP, anti-CXCL13 treated mice showed significant improvement in cognitive function when compared to control treated mice. Depression-like behavior was attenuated as well. Furthermore, mice that received anti-CXCL13 by the ICV route showed similar beneficial effects. However, the extent of lymphocyte infiltration into the brain and the general composition of the aggregates were not substantively changed by anti-CXCL13 irrespective of the mode of administration. Nevertheless, analysis of brain gene expression in anti-CXCL13 treated mice showed significant differences in key immunological and neuro-inflammatory pathways that most likely explained the improvement in the behavioral phenotype. Our results indicate that CXCL13 affects the behavioral manifestations in the MRL/lpr strain and is important to the pathogenesis of murine NPSLE, suggesting it as a potential therapeutic target.

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