B cells are associated with survival and immunotherapy response in sarcoma.

SY16-02: Generation of B cell immunity in tertiary lymphoid structures supports response to immunotherapy

Esophageal squamous cell carcinoma (ESCC) is one of the most common upper gastrointestinal malignancies worldwide. Tertiary lymphoid structures (TLS) are tumor-infiltrating immune cells aggregates coupled with stromal cells which are similar to secondary lymphoid organs. The objective of this study is to explore the predictive effects of two common genes associated with TLS models on prognosis and immunotherapy effects in ESCC patients. Clinical information for ESCC patients in the TCGA(The Cancer Genome Altas) cohort and GSE 53625 were collected. All of the samples were classified as either high score group or low score group based on two TLS signatures, and the association between TLS signatures and survival, clinical indicators, genomic burden, stemness indices analysis, tumor microenvironment and immunotherapy response were performed. Furthermore, the mature TLS was also assessed in ESCC tissue microarray. In our study, we quantified the score of TLS_9 and TLS_12, respectively, reflecting the different statuses of TLS (TLS_9 = B and T cells in TLSs; TLS_12 = neogenesis of TLSs). Subsequently, we explored the effect of TLS score on ESCC tumor microenvironment quantified by multiple algorithms. We found that a correlation analysis indicated that TLS_9 and TLS_12 were all positively correlated with CD8+ T cell, NK cells, CD4+ T cells, M1 macrophages and so on. Meanwhile, some cells present a different correlation pattern of TLS_9 and TLS_12, including activated CD4+ memory T cells and Tgd cells. Immune-related analysis revealed that the TLS_12 and TLS_9 scores were all positively correlated with immune dysfunction, yet negatively correlated with immune exclusion. Following this, the biological roles of TLS_9 and TLS_12 scores were investigated. Also, we noticed that the TLS score could significantly affect the CAFs infiltration and be associated with the genomic burden and tumor stemness. In addition, we explored the prognostic value of mature TLS through tissue microarray (TMA). Our result displayed ESCC patients with the presence of mature TLS had a better prognosis than ESCC patients without it. Our study indicated that ESCC patients with the presence of TLS had better outcomes and an inflamed immune microenvironment. In addition, both TLS-9 and TLS-12 gene signatures could be used as potential biomarkers for the immunotherapy of ESCC patients.

Tertiary lymphoid structure, an ectopic lymphoid tissue induced under chronic inflammation, develops in various kidney diseases and is associated with poor prognosis. The immune system requires metabolic resources to support immune function and lymphocyte proliferation. Hence, dramatic metabolic alterations presumably occur during the formation of tertiary lymphoid structure. However, it remains unclear whether metabolic remodeling occurs during this formation and its underlying mechanism. In a murine model of renal tertiary lymphoid structures, we used imaging mass spectrometry and metabolome analysis to investigate the metabolic pathway that characterizes tertiary lymphoid structures. We also performed in situ hybridization with immunofluorescence and pharmacological inhibition to explore the expression and function of the key molecules governing the pivotal metabolic pathway. We analyzed urine samples from mice and humans to explore the metabolites estimating the presence of renal tertiary lymphoid structures. Significant glutathione accumulation and depletion of cysteine, which is essential for glutathione synthesis, was observed specifically within tertiary lymphoid structures. The kidneys with tertiary lymphoid structures exhibited higher glutathione concentrations than healthy kidneys. Tertiary lymphoid structures also showed significant accumulation of 4-HNE and 8-OHdG, markers of oxidative stress. Dendritic cells and fibroblasts within tertiary lymphoid structures expressed the cystine/glutamate transporter, that regulates glutathione synthesis, and supplied synthesized glutathione to lymphocytes, which lacked its expression. Pharmacological inhibition of the cystine/glutamate transporter prevented tertiary lymphoid structure formation in the kidney. Furthermore, enhanced glutathione synthesis within tertiary lymphoid structures was reflected in elevated urinary glutathione concentrations both in mice and humans, which effectively detected the presence of tertiary lymphoid structures in the kidney in IgA nephropathy patients. Glutathione significantly accumulated within tertiary lymphoid structures in the kidney. Inhibition of the cystine/glutamate transporter prevented the formation of tertiary lymphoid structures. Urinary glutathione served as a biomarker to detect tertiary lymphoid structures in the kidney.

Background: STS are poorly understood cancers for which systemic therapy options are limited. Recent evidence suggests that some patients with two types of STS (undifferentiated pleomorphic sarcoma, UPS; dedifferentiated liposarcoma, DDLPS) benefit from ICB. Presence of tertiary lymphoid structures (TLS) and B cells within the sarcoma tumor immune microenvironment (TME) is associated with response to ICB and longer survival. It remains unclear how immune populations within this TME interact with each other and how they evolve upon ICB exposure. We characterized the TLS composition and spatial relationships between immune populations in untreated biopsies of UPS/DDLPS of responders (R) versus non-responders (NR). Methods: We completed a phase 2 trial of patients with resectable primary UPS (n=10) and DDLPS (n=17) who received ICB prior to surgery. Pre-ICB treatment biopsies were collected, and IMC was performed using a 34-plex antibody panel to characterize the TME of R vs NR. TLS composition, densities of immune cell subsets, and their spatial relationships were assessed across baseline biopsy specimens within 3 pre-defined regions-of-interest (ROIs): TLS, non-TLS, and TLS-non-TLS interface regions. Results: We identified distinct TME profiles in R compared to NR prior to ICB treatment. Intratumoral B cell density was higher in R compared to NR patients across pre-treatment biopsies (502 vs 277 cells/mm2, p=0.45) and within interface (1155 vs 710 cells/mm2, p=0.42) and non-TLS (458 vs 257 cells/mm2, p=0.17) regions but lower in TLS regions (506 vs 729 cells/mm2, p=0.52). NR patients tended to have higher densities of Tregs across pre-treatment samples (81 vs 37 cells/mm2, p=0.44) and in TLS (981 vs 824 cells/mm2, p=0.59), interface (119 vs 53 cells/mm2, p=0.14), and non-TLS (66 vs 15 cells/mm2, p<0.01) regions. When examining spatial distributions of TME immune populations, we noted a trend towards greater distances between B cells and T regs in the tumors of R compared to NR patients (78 vs 45 microns, p=0.13) which was also observed in TLS (116 vs 40 microns, p=0.05), non-TLS (76 vs 44 microns, p=0.14), and interface (179 vs 41 microns, p<0.01) regions. Conclusion: Although presence of intratumoral TLS is associated with ICB response and longer survival among patients with UPS/DDLPS, not everyone benefits. Response to ICB appears to be associated with the immune profile both within and beyond TLS regions of pre-treatment tumors. Lower Treg, higher B cell populations, and greater distances between Tregs and B cells within pre-treatment biopsies may help predict response to ICB for UPS and DDLPS. Studies are ongoing to assess the evolution of these spatial relationships after ICB treatment. Citation Format: Laurence P. Diggs, Elise Nassif Haddad, Susmita Kumari, Varshini Arunkumar, Sammy Ferri-Borgogno, Samuel Mok, Duncan Mak, Angelique Lin, Madeline McAllister, Akshay Basi, Manoj Chelvanambi, Javier Gomez, Jared Burks, Davis Ingram, Khalida Wani, Natalie Duarte, Trevor McKee, Alexander Lazar, Jennifer Wargo, Christina Roland, Emily Keung. Imaging mass cytometry (IMC) reveals distinct intratumoral B and T regulatory cell populations in responder versus non-responder soft tissue sarcoma (STS) patients before treatment with immune checkpoint blockade (ICB) [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 7124.

This study aimed to investigate the distribution, maturity, and prognostic significance of tertiary lymphoid structures (TLS) in primary breast cancer (BC), and to analyze the relationship between TLS clinicopathological features and the tumor immune microenvironment. Sixty postoperative tumor samples from patients with primary BC were retrospectively analyzed. The presence and maturity of TLS in tumor tissues were evaluated by immunohistochemical staining. The association of status and clinicopathological characteristics of TLS with immune cell infiltration was analyzed. Recurrence-free survival (RFS) and predictive factors were assessed using Kaplan-Meier survival analysis and univariate/multivariate logistic regression. TLS was detected in 35 (58.33%) patients, including 23 cases of secondary follicle-like TLS, 8 cases of early TLS, and 4 cases of primary follicle-like TLS. Compared with the non-TLS group, the TLS group showed significantly higher expression of estrogen receptor and progesterone receptor (p < 0.05). The presence of TLS was significantly associated with improved RFS (HR = 0.163, p = 0.035), with secondary follicle-like TLS showing the most favorable survival trend. TLS-positive tumors had significantly increased infiltration of CD4+ T cells (p = 0.004), while CD8+ T cell infiltration was slightly higher but not statistically significant. The presence of TLS, especially mature TLS, is associated with favorable clinicopathological features and improved prognosis in patients with primary BC, along with increased CD4+ T cell infiltration, suggesting an important role of TLS in regulating the tumor immune microenvironment. TLS may serve as a potential biomarker for prognosis evaluation and a target for immunomodulatory strategies in BC.

Tertiary lymphoid structures (TLSs) have emerged as critical regulators of antitumor immunity and prognostic indicators in various malignancies. However, the distribution patterns and prognostic significance of TLSs in hepatoblastoma (HB) remain poorly understood. This study aimed to investigate the presence, distribution, and prognostic value of TLSs in HB patients following neoadjuvant chemotherapy and to explore the underlying mechanisms linking TLSs to the tumor immune microenvironment. A total of 112 HB patients who underwent neoadjuvant chemotherapy and surgical resection at Shandong Provincial Hospital between 2015 and 2024 were retrospectively enrolled. The presence of TLSs was evaluated using hematoxylin and eosin (H&E) staining, and patients were classified into TLS-positive and TLS-negative groups. Univariate and multivariate Cox regression analyses were performed to identify independent prognostic factors for overall survival (OS). In addition, transcriptome data from the GEO database (GSE133039) were analyzed to construct a TLS gene signature score and explore immune-related mechanisms associated with TLS presence. TLSs were identified in 45 out of 112 hepatoblastoma patients (40.2%). Kaplan-Meier survival analysis demonstrated that TLS-positive patients had significantly longer overall survival (OS) compared to TLS-negative patients (p = 0.0017). Multivariate Cox regression analysis further confirmed the presence of TLSs as an independent favorable prognostic factor (HR = 0.061, p = 0.027). In contrast, advanced PRETEXT stage (III/IV), vascular invasion, and distant metastasis were identified as independent adverse prognostic factors, indicating that patients diagnosed at later stages tended to have a worse prognosis. Transcriptomic analysis revealed that TLS-positive tumors exhibited higher expression of antigen presentation and immune activation-related genes (e.g., HLA-DQA1, HLA-DQB1, SLAMF7), along with enriched infiltration of B cells, CD8+ T cells, and NK cells, suggesting a more active antitumor immune microenvironment. The presence of TLSs is significantly associated with favorable prognosis in HB patients and may contribute to enhanced antitumor immunity by recruiting and activating cytotoxic immune cells. TLSs represent a promising prognostic biomarker and potential immunotherapeutic target for HB patients. Tertiary lymphoid structures (TLSs) serve as a promising prognostic biomarker in hepatoblastoma (HB). Our study demonstrates that TLS-positive patients exhibit significantly prolonged overall survival. TLSs contribute to the tumor immune microenvironment by recruiting cytotoxic immune cells. These findings provide new insights into TLSs as a potential immunotherapeutic target for HB patients.

The presence of tertiary lymphoid structures (TLSs) and a high intratumoral density of B lymphocytes have been associated with improved survival in several tumor types. We investigated the impact of TLS on responses to immunotherapy and, analyzing a large-scale retrospective analysis of three independent cohorts of patients with cancer, including sarcomas, who were treated with anti-PD-1 or anti-PD-L1 antibodies, we showed that the presence of mature TLSs was associated with improved objective response rates, progression-free survival and overall survival, independent of PD-L1 expression status and CD8+T cell density. To decipher the role of B cells in such responses, we performed spatial transcriptomics and examined the nature of B cell responses within TLS structure. B cells were enriched in TLS, and therein we could identify all B cell maturation stages toward plasma cell (PC) formation. B cell repertoire analysis revealed clonal diversification, selection, expansion in TLS and the presence of full mature clonotypes at distance. In TLS+ tumors, IgG producing PCs disseminated into the tumor beds along fibroblastic tracks. TLS+ tumors exhibited high frequencies of IgG-producing PCs and IgG-stained and apoptic malignant cells, suggestive of anti-tumor effector activity. We validated these findings in a prospective trial in metastatic Soft Tissue Sarcoma (STS) bearing patients. PEMBROSARC is a multi-cohort phase II study of pembrolizumab combined with low-dose cyclophosphamide in patients with advanced STS. The 6-months non-progressive rate (NPR) and the objective response rates previously reported in cohorts including all comers were 4.9% and 2.4%, respectively. In the cohort enrolling patients selected based on the presence of TLS, the 6-month NPR was 40% and the objective response rate was 30%. Exploratory analyses reveled that infiltration by IgG antibody-producing plasma cells was associated with improved clinical outcome as well as the presence of IgG antibodies on tumor cells. Altogether, these results establish that maturation, selection and amplification of B cells in intratumoral TLSs results in Plasma Cells generation which may produce anti-tumor antibodies. The latter may amplify in situ antitumor immune responses leading to better therapeutic responses to immunotherapy. Citation Format: Wolf H. Fridman. B cell immunity generation in tertiary lymphoid structures, a major determinant of sarcoma immune response to immunotherapy [abstract]. In: Proceedings of the AACR Special Conference: Sarcomas; 2022 May 9-12; Montreal, QC, Canada. Philadelphia (PA): AACR; Clin Cancer Res 2022;28(18_Suppl):Abstract nr IA007.

Background Tertiary lymphoid structures (TLS) in inflammatory bowel disease (IBD) have traditionally been regarded as secondary structures accompanying chronic inflammation. We hypothesize that TLS are crucial components in the pathological environment that promote ulcer formation and that biological agents function by disrupting this TLS-dependent niche. Our goal is to examine the relationship between inflammatory cytokines—targets for biological agents—and TLS; assess the effect of anti-TNF therapy on TLS; and evaluate the potential to predict anti-TNF therapy effectiveness using biopsy TLS indicators. Methods To address the limitation of mucosal biopsies that do not cover the entire ulcerative niche, Xenium analysis was performed on full-thickness intestinal segments from surgical resection specimens, including four untreated cases and four cases treated with TNF inhibitors. Additionally, from 849 surgical cases collected between 2014 and 2019, 105 cases were selected—82 untreated and 23 treated with anti-TNF monotherapy. In a propensity-score-matched immunohistochemistry cohort (1:3 ratio, n = 60), the presence of TLS, follicular dendritic cells, and high endothelial venules were quantitatively assessed. Furthermore, in public biopsy data (CD and ulcerative colitis; UC), prediction of TNF inhibitor responsiveness was performed using pre-treatment TLS indicators. Results (1) The number of TLS was linked to clinical and histological activity indices, showing TLS burden as a measurable marker of the ulcerative niche. (2) In untreated samples, TNF-positive cells targeted by TNF inhibitors were sparse within the mucosal lamina propria and were mostly concentrated within the TLS. (3) In cases treated with anti-TNF therapy, there was a notable reduction in TNF-positive cells and CXCL13-positive follicular dendritic cells (FDCs) within the TLS, along with changes such as an increase in M2 macrophages inside the TLS. (4) In cases that did not respond to TNF inhibitors, TLS persisted despite FDC depletion, with CXCL13-positive Tfh cells still present, indicating a Tfh-dependent compensatory niche. (5) TLS status at pre-treatment biopsy was a reliable predictor of response to TNF inhibitors in both CD and UC (CD: AUC 0.762; UC: AUC 0.867). Conclusion TLS is more than just an indicator; it is an ulcerative niche. By interfering with CXCL13⁺ follicular dendritic cell networks, anti-TNF therapy disrupts TLS-supporting niches and reduces TLS burden. Conversely, Tfh-derived CXCL13 may play a distinct role in promoting local tolerance. The TLS index in pre-treatment biopsy specimens proved useful for anti-TNF stratification in CD/UC. Conflict of interest: Dr. Kodama, Makoto: No conflict of interest Nagase, Yoshihiro: No conflict of interest Okano, Soh: No conflict of interest Yamamoto, Kouhei: No conflict of interest Furukawa, Asuka: No conflict of interest Yamamoto, Kurara: No conflict of interest Miyata, Ryota: No conflict of interest Abe, Keiko: No conflict of interest Ohteki, Toshiaki: No conflict of interest Makishima, Makoto: No conflict of interest

BackgroundOral squamous cell carcinomas are often heavily infiltrated by immune cells. The organization of B-cells, follicular dendritic cells, T-cells and high-endothelial venules into structures termed tertiary lymphoid structures have been detected in various types of cancer, where their presence is found to predict favourable outcome. The purpose of the present study was to evaluate the incidence of tertiary lymphoid structures in oral squamous cell carcinomas, and if present, analyse whether they were associated with clinical outcome.MethodsTumour samples from 80 patients with oral squamous cell carcinoma were immunohistochemically stained for B-cells, follicular dendritic cells, T-cells, germinal centre B-cells and high-endothelial venules. Some samples were sectioned at multiple levels to assess whether the presence of tertiary lymphoid structures varied within the tumour.ResultsTumour-associated tertiary lymphoid structures were detected in 21 % of the tumours and were associated with lower disease-specific death. The presence of tertiary lymphoid structures varied within different levels of a tissue block.ConclusionsTertiary lymphoid structure formation was found to be a positive prognostic factor for patients with oral squamous cell carcinoma. Increased knowledge about tertiary lymphoid structure formation in oral squamous cell carcinoma might help to develop and guide immune-modulatory cancer treatments.

Blockade of the inhibitory PD1 pathway on CD8+ and CD4+ tumor infiltrating lymphocytes (TILs) has revolutionized standard of care for cancer patients. However, these approaches only benefit approximately 20% of cancer patients, thus a more comprehensive understanding of the immune response is paramount for the development of new therapeutic approaches and enhancement of the anti-tumor immune response. The presence of tertiary lymphoid structures (TLS) correlates with enhanced anti-tumor immunity and improved prognosis in several solid tumors. These ectopic lymphoid aggregates can exhibit features like secondary lymphoid organs (SLOs), including high endothelial venules, a T cell zone with mature dendritic cells (DCs), and a germinal center (GC) with follicular DCs and B cells, that facilitate the induction of immune responses in situ. Furthermore, the presence of TLS correlate with superior responses to immunotherapy and mature, GC-containing TLS correlate with increased T cell function in human tumors. However, current immunotherapies do not target TLS despite their predominance in the tumor microenvironment (TME) and key role in the adaptive immune response. Developing therapies that adequately induce TLS is limited by a lack of objective, bioinformatic analyses to elucidate the cellular and locational heterogeneity of TLS. TLS differences in composition are thought to define maturation levels i.e. early TLS have B and T cell aggregates but lack follicular DC and GC appear as the TLS mature. However, cellular composition is just one aspect of TLS heterogeneity. To successfully increase mature TLS in cancer patients for maximal humoral immunity, we must first understand the complete transcriptomic and proteomic profile of TLS in cancer patients. We have utilized the innovative Nanostring GeoMx platform to spatially interrogate the transcriptomics and proteomics associated with TLS. By pairing this technology with high level multispectral immunofluorescence via the Vectra Polaris system, we can link TLS heterogeneity to TLS position within the tumor. Further, we can analyze the complete activation profile of B and T cells to understand how TLS heterogeneity affects lymphocyte function. Our data have indicated that TLS can be linked to increased CD8+ T cell infiltrate and function. In fact, when mature TLS are not present, CD8+ T cell and Treg interactions increase, ultimately creating a more immunosuppressive TME. Lastly, utilizing a novel, robust bioinformatic pipeline, we have identified new genes that will aid in an objective definition of TLS. Specifically, CD27 is increased within TLS that are proximal to the tumor compared to those in the normal adjacent tissue. These studies will revolutionize the way in which TLS are defined within the TME, ultimately offering comprehensive gene and protein analysis of TLS that includes spatial geography of the structures and an unbiased, objective definition of TLS for future clinical utility and immunotherapeutic targeting. Citation Format: Dongyan Liu, Xiang Li, Rajesh Acharya, Ernest M. Meyer, Shelley Reynolds, Ayana Ruffin, Robert L. Ferris, Dario A.A. Vignali, Riyue Bao, Tullia C. Bruno. Utilizing spatial transcriptomics to elucidate tertiary lymphoid structure heterogeneity in human cancer [abstract]. In: Proceedings of the AACR Virtual Special Conference on Tumor Heterogeneity: From Single Cells to Clinical Impact; 2020 Sep 17-18. Philadelphia (PA): AACR; Cancer Res 2020;80(21 Suppl):Abstract nr PO-083.

1452P - Prognostic significance of elements of the adaptive immunity in the microenvironment of early-stage non-small cell lung cancer

BackgroundRecent evidence suggests that dietary fiber intake is associated with improved clinical and pathologic response to anti-PD-1 therapy,1 though this relationship is unclear in soft-tissue sarcoma. We evaluated the impact...

Introduction Tertiary lymphoid structures (TLSs) and tumor-infiltrating lymphocytes (TILs) in breast carcinomas are prognostic for survival and predictive of certain therapy responses. The presence of TLSs and TILs are identified by manual pathological examination; however, this method often lacks reproducibility, limiting its use in routine clinical practice. Here, we demonstrate that morphological evaluation of whole slide images (WSIs) using an artificial intelligence (AI)-based analytic workflow comprised of convolutional neural network (CNN) deep learning models that accurately and reproducibly characterizes TILs, measured as the lymphocyte immune-infiltrated area (LIIA), and TLSs in the tumor microenvironment (TME) of breast carcinomas. Methods We collected a cohort of 445 TCGA breast cancer H&amp;E WSIs, including clinical and sequencing data, and divided this cohort into luminal invasive lobular carcinoma (ILC) (n = 192), HER2-enriched (n = 110), and basal-like (n = 143) molecular subtypes. After 55 samples were excluded due to artifacts or incomplete clinical annotation, a total of 390 samples were analyzed. A combination of CNN-based deep learning models was used to detect and classify the tumor area, TLSs present in the TME, TLS density (number of TLS per mm2 of tumor), and lymphocyte-rich regions. The LIIA was calculated as the area of the stromal and TIL components of the TME. Validation was performed by manually annotating 10 random WSIs from the dataset. Spatial model predictions of the tumor and TLSs were combined to identify TLS locations. Each model’s predictions were verified by univariate (Kaplan-Meier) and multivariate (Cox regression) survival analyses, and the log-rank test was used to calculate overall survival. Additionally, the relationship between TLSs and LIIAs with CD274 expression (PD-L1) and a high tumor mutational burden (TMB &amp;gt; 10) was analyzed. Statistical analyses included Spearman’s rank correlation and Mann-Whitney tests. Results TLS were detected in 53% (n = 207) of the samples, with a mean density of 26.02 TLS/mm2 (Q3 = 5.53 TLS/mm2). TLS density was higher in basal-like subtype samples compared to luminal and HER2-enriched subtypes. While LIIA and TMB-high samples exhibited a significant relationship (p = 0.00001), no significant association was found between TME and TLS quantities or density. PD-L1 gene expression exhibited weak to moderate correlations with predicted LIIA in basal-like (r = 0.38, p = 0.00001) and HER2-enriched subtypes (r = 0.38, p = 0.0001). The luminal subtype had no significant correlation between PD-L1 expression and predicted LIIA. As a result, LIIA and TLS were characterized as positive prognostic factors for the basal-like subtype. After adjusting for age, stage, and grade, the LIIA and TLS density were found to be significant independent positive prognostic overall survival factors for the basal-like subtype (LIIA HR: 0.02, p = 0.003; TLS-high group HR: 0.09, p = 0.002). For the HER2-enriched subtype, TLS density was also a significant predictor (HR: 0.05, p = 0.035), while LIIA was not a statistically significant prognostic factor (HR: 0.0002, p = 0.08). Associations were not observed between the TLSs and LIIA between the ILC subtypes and survival outcomes. The same result was observed for univariate analyses. Conclusion The developed analytic pipeline accurately identified the presence of LIIA and TLS on H&amp;E slides, demonstrating the potential of CNN for automated characterization of the breast cancer TME. AI-based TLS and LIIA quantification can be a robust tool for pathology processes, offering additional information to help in clinical decision-making. This approach can be used to detect features of immune morphology biomarkers in other cancer types. Citation Format: Vladimir Kushnarev, Daniil Dymov, Nadezhda Lukashevich, Lev Popyvanov, Anna Belozerova, Diana Shamsutdinova, Aida Akaeva, Yury Popov, Svetlana Khorkova, Ivan Valiev, Anastasia Zotova, Jessica H. Brown, Anna Love, Alexander Bagaev, Ekaterina Postovalova, Nathan Fowler. AI-based prediction of tertiary lymphoid structures and lymphocyte immune infiltration in breast carcinomas [abstract]. In: Proceedings of the 2022 San Antonio Breast Cancer Symposium; 2022 Dec 6-10; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2023;83(5 Suppl):Abstract nr P6-04-15.

Lymphocyte Function in Tertiary Lymphoid Structures Predicts Hepatocellular Carcinoma Outcome

BACKGROUND Recently, the presence of tertiary lymphoid structure (TLS), composed mainly of intratumoral B cells, has been reported as a favorable prognostic factor in various types of cancer. However, TLS has not been well studied in glioblastoma. In this study, we performed spatial and genomic analyses in human glioblastoma, focusing on TLS. METHODS We evaluated lymphocytic infiltration by immunostaining in 50 cases, which were diagnosed with glioblastoma from April 2018 to March 2023 in Okayama University Hospital. RNA-sequencing (RNA-seq) was performed in 39 cases to compare those with and without TLS by Differential Gene Expression (DGE) analysis and Gene Set Enrichment Analysis (GSEA). In addition, we performed spatial transcriptomic analysis using the tissue sections with TLS. RESULTS TLS was detected in 4 (8.0%) of the 50 cases. DGE analysis showed increased gene expression related to the T cell activation marker (TNFRSF9), chemokine (CCL5), etc., in patients with TLS. GSEA also confirmed the activation of immune responses such as Adaptive immune response, Cytokine production, and Lymphocyte activation etc. Spatial transcriptomic analysis revealed the local upregulation of genes detected by RNA-seq analysis in and around TLS region. Moreover, it seemed to be the secondary lymphoid follicles and follicular helper T cells in TLS, which suspected the presence of mature TLS. CONCLUSION We analyzed a total of 50 cases of glioblastoma. Glioblastoma with TLS were immune activated.