scholarly journals Defining the microenvironment landscape of bladder cancer using highly multiplexed spatial genomic and proteomic analysis

2019 ◽  
Author(s):  
Jason W Reeves ◽  
Zhaojie Zhang ◽  
Zachary K Norgaard ◽  
Denise M Zhou ◽  
JingJing Gong ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
T Cell ◽  
Immune Cell ◽  
Cell Types ◽  
Primary Resistance ◽  
Stromal Compartment ◽  
Spatial Profiling ◽  
Signaling Axis ◽  
Ffpe Tissues ◽  
Muscle Invasive

AbstractMuscle-invasive bladder cancer (MIBC) is an aggressive disease with limited therapeutic options. PD-1 pathway targeting immunotherapies have been approved to treat advanced bladder cancer, but most patients exhibit primary resistance, suggesting that immune evasion mechanisms exist. The PPARγ pathway has been identified as a potential therapeutic target in MIBC that is associated with reduced CD8+ T-cell infiltration and increased resistance to immunotherapies. We comprehensively profiled the tumor microenvironment (TME) in formalin-fixed, paraffin-embedded (FFPE) tissues from a cohort of PPARγhigh (n=13) and PPRARγlow (n=12) MIBC, integrating bulk gene expression, targeted mutation sequencing, immunohistochemistry and multiplex spatial profiling of RNA and protein expression on the GeoMx™ Digital Spatial Profiling (DSP) platform. Molecular subtyping was consistent between traditional methods and GeoMx profiling, and, in this cohort, we observed little evidence of spatial heterogeneity in tumor subtyping. The previously characterized T-cell exclusion phenotype of PPARγhigh MIBC was recapitulated on the GeoMx platform and was further extended to show that this is a general phenomenon across immune cell types, supporting potential combination of PPARγ inhibition with ICIs. Furthermore, we found that while immune cells were excluded from PPARγhigh tumors, the stromal compartment from these tumors was not significantly different than those PPARγlow tumors. By preserving spatial relationships during the GeoMx analysis, we also identify a novel association between lower immune cell expression in the tumors and higher expression of β-catenin in the stroma, and differential expression of other WNT pathway members associated with PPARγ activity.One Sentence SummaryA new method for capturing tumor-immune signaling in FFPE tissues explores how the PPARG signaling axis is associated with immune cell exclusion in bladder cancer.

Tumor Subtyping: Making Sense of Heterogeneity with a Goal Toward Treatment

2020 ◽  
pp. 1-11
Author(s):  
Joshua J. Meeks ◽  
Gottfrid Sjödahl ◽  
Seth P. Lerner ◽  
Arighno Das ◽  
David J. McConkey ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Immune Cell ◽  
Response To Therapy ◽  
Genomic Diversity ◽  
Intrinsic Resistance ◽  
Making Sense ◽  
Search Terms ◽  
Muscle Invasive ◽  
Next Generation Sequencing Ngs ◽  
Descriptive Feature

BACKGROUND: Bladder cancers have high total mutation burdens resulting in genomic diversity and intra- and inter-tumor heterogeneity that may impact the diversity of gene expression, biologic aggressiveness, and potentially response to therapy. To compare bladder cancers among patients, an organizational structure is necessary that describes the tumor at the histologic and molecular level. These “molecular subtypes”, or “expression subtypes” of bladder cancer were originally described in 2010 and continue to evolve secondary to next generation sequencing (NGS) and an increasing public repository of well-annotated cohorts. OBJECTIVE: To review the history and methodology of expression-based subtyping of non-muscle invasive (NMIBC) and muscle invasive bladder cancer (MIBC). METHODS: A literature review was performed of primary papers from PubMed that described subtyping methods and their descriptive feature including search terms of “subtype”, and “bladder cancer”. RESULTS: 21 papers were identified for review. Tumor subtyping developed from N = 2 to N = 6 subtyping schemes with most subtypes comprised of at least luminal and basal tumors. Most NMIBCs are luminal cancers and luminal MIBCs may be associated with less aggressive features, while one study of basal tumors identified a better clinical outcome with systemic chemotherapy. Tumors with a 53-like signature may have intrinsic resistance to chemotherapy. The heterogeneity of tumors, which is likely derived from stromal components and immune cell infiltration, affect subtype calls. CONCLUSION: Subtyping, while still evolving, is ready for testing in clinical trials. Improved patient selection with tumor subtyping may help with tumor classification and potentially match patient or tumor to therapy.

scholarly journals Low T-cell Receptor Diversity, High Somatic Mutation Burden, and High Neoantigen Load as Predictors of Clinical Outcome in Muscle-invasive Bladder Cancer

2016 ◽  
Vol 2 (4) ◽  
pp. 445-452 ◽  
Author(s):  
Noura J. Choudhury ◽  
Kazuma Kiyotani ◽  
Kai Lee Yap ◽  
Alexa Campanile ◽  
Tatjana Antic ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
T Cell ◽  
Clinical Outcome ◽  
T Cell Receptor ◽  
Somatic Mutation ◽  
Cell Receptor ◽  
Invasive Bladder Cancer ◽  
Muscle Invasive Bladder Cancer ◽  
Mutation Burden ◽  
Muscle Invasive

scholarly journals Intratumoral TIGIT+ CD8+ T-cell infiltration determines poor prognosis and immune evasion in patients with muscle-invasive bladder cancer

2020 ◽  
Vol 8 (2) ◽  
pp. e000978
Author(s):  
Zhaopei Liu ◽  
Quan Zhou ◽  
Zewei Wang ◽  
Hongyu Zhang ◽  
Han Zeng ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
T Cells ◽  
T Cell ◽  
Invasive Bladder Cancer ◽  
Th2 Cells ◽  
Cancer Center ◽  
Muscle Invasive Bladder Cancer ◽  
Cell Abundance ◽  
Immune Contexture ◽  
Muscle Invasive

BackgroundT-cell immunoglobulin and ITIM domain (TIGIT) is identified as a novel checkpoint receptor that can facilitate immune escape via mediating T-cell exhaustion in tumors. However, the clinical significance and immune contexture correlation of intratumoral TIGIT+ CD8+ T-cells remain to be further explored in muscle-invasive bladder cancer (MIBC).Methods259 patients with MIBC from two clinical centers (Zhongshan Hospital, n=141; Shanghai Cancer Center, n=118) were analyzed to evaluate the prognostic value and immune contexture association of TIGIT+ CD8+ T-cells through immunohistochemistry. Fresh tumor tissue samples from 26 patients with MIBC were examined to discover the phenotype of this CD8 subpopulation by flow cytometry.ResultsHigh infiltration of intratumoral TIGIT+ CD8+ T-cells predicted poor overall survival (OS) and recurrence-free survival (RFS) in MIBC. For patients with stage II MIBC with low infiltration of TIGIT+ CD8+ cells, adjuvant chemotherapy (ACT) could significantly prolong their OS and RFS. Intratumoral TIGIT+ CD8+ T-cell abundance was correlated with impaired CD8+ T-cell cytotoxicity and exhibited production of immunosuppressive cytokine IL-10. Further analysis of tumor-infiltrating immune cell landscape revealed TIGIT+ CD8+ T-cells were associated with suppressive immune contexture, including Th2 cells, regulatory T-cells, mast cells and neutrophils.ConclusionIntratumoral TIGIT+ CD8+ T-cell abundance could serve as an independent prognosticator for clinical outcome and a predictive biomarker for inferior ACT responsiveness. Intratumoral TIGIT+ CD8+ T-cell abundance correlated with dampened CD8+ T-cell antitumor immunity and immunosuppressive contexture abundance, highlighting a tumor-promoting role of TIGIT+ CD8+ T-cells.

MP98-14 CATHEPSIN S INHIBITION CHANGES REGULATORY T-CELL ACTIVITY IN REGULATING BLADDER CANCER AND IMMUNE CELL PROLIFERATION AND APOPTOSIS

2017 ◽  
Vol 197 (4S) ◽  
Author(s):  
Xiang Yan ◽  
Chun Wu ◽  
Tao Chen ◽  
Marcela M. Santos ◽  
Conglin Liu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Bladder Cancer ◽  
T Cell ◽  
Regulatory T Cell ◽  
Immune Cell ◽  
Cell Activity ◽  
Cathepsin S ◽  
Proliferation And Apoptosis

scholarly journals IMMU-08. MICROENVIRONMENT MODULATION BY TIM-3 BLOCKADE IMPROVES THE OUTCOME OF PRECLINICAL DIPG MODELS

2021 ◽  
Vol 23 (Supplement_1) ◽  
pp. i28-i28
Author(s):  
Iker Ausejo-Mauleon ◽  
Sara Labiano ◽  
Virginia Laspidea ◽  
Marc Garcia-Moure ◽  
Daniel de la Nava ◽  
...  
Keyword(s):  
T Cell ◽  
Tumor Microenvironment ◽  
Nk Cells ◽  
Immune Cell ◽  
Cell Types ◽  
Pediatric Brain Tumors ◽  
Immune Memory ◽  
Cd8 Cells ◽  
Functional Studies

Abstract Diffuse Midline Gliomas (DMGs), encompassing Diffuse Intrinsic Pontine Gliomas (DIPGs), are the most aggressive pediatric brain tumors. Their meagre survival has not changed despite the combination of radiotherapy with targeted therapies emphasizing the urgent need for effective treatments. Recent research suggested that the DIPG tumor microenvironment is neither highly immunosuppressive nor inflammatory. These analyses showed the lack of infiltrating lymphocytes and the abundance of CD11b+ cells. TIM-3 (HAVCR2) is a member of the T-cell immunoglobulin and mucin domain protein family which is expressed on multiple immune cell types including T cells, Tregs, NK cells, monocytes, dendritic cells and microglia, where it potently regulates not only adaptive immunity but also innate immunity. Therefore, the central hypothesis of this study is that TIM-3 inhibitors could stimulate a cytotoxic immune effect and challenge several components in the tumor microenvironment including microglia, thereby providing a potential effective treatment for DMGs. In silico assessment of TIM-3 expression in a DIPG datasets showed a robust expression of this gene. Moreover, single-cell sequencing analyses of DIPG biopsies uncover its expression on tumor cells, especially in the OPCs compartment. In vivo efficacy studies showed that treatment with anti-TIM-3 antibody significantly increase the overall survival in two DIPG immunocompetent orthotopic animal models (doubling the median), lead to long-term survivors (50%) and showed immune memory. Analyses of CD45+ populations in the tumor microenvironment showed a significant increase in B, NK and CD8+ cells corresponding with a T-cell activate phenotype in treated-mice. The potential therapeutic involvement of NK cells was certified using nude mice and functional studies. Involvement of microglia in currently being analysed. In summary, these data underscore TIM-3 as a potential target DIPGs and uncover the potential involvement of NKs and other immune mechanisms in the efficacy of anti-TIM-3 therapy.

scholarly journals Higher cytolytic score correlates with an immunosuppressive tumor microenvironment and reduced survival in glioblastoma

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Alexander F. Haddad ◽  
Jia-Shu Chen ◽  
Taemin Oh ◽  
Matheus P. Pereira ◽  
Rushikesh S. Joshi ◽  
...  
Keyword(s):  
T Cell ◽  
Immune Cell ◽  
Cell Types ◽  
Cd8 T Cell ◽  
The Cancer Genome Atlas ◽  
Cell Infiltration ◽  
Sequencing Data ◽  
Gene Score ◽  
T Cell Infiltration ◽  
The Relationship

Abstract Cytolytic score (CYT), calculated from mRNA expression levels of granzyme and perforin, positively correlates with CD8+ T cell infiltration/activity in a variety of cancers. Unlike other cancers, higher CYT has been associated with worse prognosis in glioblastoma (GBM). To address this discrepancy, we sought to investigate the relationship between CYT and immune checkpoint gene score (ICGscore), as well as their correlation with patient survival and tumor immune cell infiltration. Clinical and RNA-sequencing data for patients with newly diagnosed GBM were obtained from The Cancer Genome Atlas. Maximally-selected rank statistics was used to dichotomize subgroups. CIBERSORT was used to estimate abudence of immune cell-types. Spearman correlation was used to characterize the relationship between CYT and ICGscore. Kaplan–Meier curves were generated for survival analysis. Overall, 28/151 patients had high CYT. High CYT was associated with a mesenchymal subtype (p < 0.001) and worse survival (7.45 vs. 12.2 months, p < 0.001). There were no differences in patient demographics, IDH/MGMT mutation status, or treatment. On subgroup analysis, patients with high CYT/ICGscore had significantly increased CD8+ infiltration (p < 0.001), as expected, and worse survival (HR 0.445, p < 0.01). Furthermore, CYT strongly correlated with ICGscore (RS = 0.675, p < 0.001). The high CYT/ICGscore subgroup was associated with greater infiltration of M2 macrophages (p = 0.011) and neutrophils (p = 0.055). Our study highlights a multidimensional immunosuppressive GBM microenvironment in patients with higher CYT and potentially identifies patients with high CYT/ICGscore as a subgroup that may particularly benefit from multi-faceted immunotherapies, given their already elevated tumor CD8+ T cell levels.

scholarly journals Stem-Like Signature Predicting Disease Progression in Early Stage Bladder Cancer. The Role of E2F3 and SOX4

Biomedicines ◽  
2018 ◽  
Vol 6 (3) ◽  
pp. 85 ◽  
Author(s):  
Joaquim Bellmunt
Keyword(s):  
Stem Cells ◽  
Bladder Cancer ◽  
Cancer Stem Cells ◽  
Tumor Progression ◽  
Cell Types ◽  
Invasive Bladder Cancer ◽  
Low Grade ◽  
High Grade ◽  
Muscle Invasive Bladder Cancer ◽  
Muscle Invasive

The rapid development of the cancer stem cells (CSC) field, together with powerful genome-wide screening techniques, have provided the basis for the development of future alternative and reliable therapies aimed at targeting tumor-initiating cell populations. Urothelial bladder cancer stem cells (BCSCs) that were identified for the first time in 2009 are heterogenous and originate from multiple cell types; including urothelial stem cells and differentiated cell types—basal, intermediate stratum and umbrella cells Some studies hypothesize that BCSCs do not necessarily arise from normal stem cells but might derive from differentiated progenies following mutational insults and acquisition of tumorigenic properties. Conversely, there is data that normal bladder tissues can generate CSCs through mutations. Prognostic risk stratification by identification of predictive markers is of major importance in the management of urothelial cell carcinoma (UCC) patients. Several stem cell markers have been linked to recurrence or progression. The CD44v8-10 to standard CD44-ratio (total ratio of all CD44 alternative splicing isoforms) in urothelial cancer has been shown to be closely associated with tumor progression and aggressiveness. ALDH1, has also been reported to be associated with BCSCs and a worse prognosis in a large number of studies. UCC include low-grade and high-grade non-muscle invasive bladder cancer (NMIBC) and high-grade muscle invasive bladder cancer (MIBC). Important genetic defects characterize the distinct pathways in each one of the stages and probably grades. As an example, amplification of chromosome 6p22 is one of the most frequent changes seen in MIBC and might act as an early event in tumor progression. Interestingly, among NMIBC there is a much higher rate of amplification in high-grade NMIBC compared to low grade NMIBC. CDKAL1, E2F3 and SOX4 are highly expressed in patients with the chromosomal 6p22 amplification aside from other six well known genes (ID4, MBOAT1, LINC00340, PRL, and HDGFL1). Based on that, SOX4, E2F3 or 6q22.3 amplifications might represent potential targets in this tumor type. Focusing more in SOX4, it seems to exert its critical regulatory functions upstream of the Snail, Zeb, and Twist family of transcriptional inducers of EMT (epithelial–mesenchymal transition), but without directly affecting their expression as seen in several cell lines of the Cancer Cell Line Encyclopedia (CCLE) project. SOX4 gene expression correlates with advanced cancer stages and poor survival rate in bladder cancer, supporting a potential role as a regulator of the bladder CSC properties. SOX4 might serve as a biomarker of the aggressive phenotype, also underlying progression from NMIBC to MIBC. The amplicon in chromosome 6 contains SOX4 and E2F3 and is frequently found amplified in bladder cancer. These genes/amplicons might be a potential target for therapy. As an existing hypothesis is that chromatin deregulation through enhancers or super-enhancers might be the underlying mechanism responsible of this deregulation, a potential way to target these transcription factors could be through epigenetic modifiers.

455 Impact of EphB4 and PD-1 treatment on immune infiltrate in advanced bladder cancer

2020 ◽  
Vol 8 (Suppl 3) ◽  
pp. A482-A482
Author(s):  
Sarmad Sadeghi ◽  
Tyler Hether ◽  
Jason Yeon ◽  
Richard Mangio ◽  
Jason Reeves ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
T Cell ◽  
Tumor Microenvironment ◽  
Combination Therapy ◽  
Mouse Models ◽  
Immune Cell ◽  
Clinical Investigation ◽  
Post Treatment ◽  
Rna Expression ◽  
Preclinical Models

BackgroundBladder cancer is the fourth most common cancer in American men with chances of 1 in 27 developing this form of cancer. Despite the progress in treating these patients with immunomodulatory agents, the vast majority of patients remain refractory to therapeutic intervention. EphB4 and EphrinB2 are induced in the tumor vasculature and modulate immune response within the tumor microenvironment. Intervention blocking Ephrin and PD-1/PD-L1 pathway has shown promising data in preclinical models. These data form the basis of clinical investigation of combined therapy in bladder cancer and other tumor types.MethodsPreclinical mouse models were treated with decoy soluble EphB4 and tumor infiltrating immune cells were profiled by RNA expression analysis post-treatment and compared to control treated mice. Next, patients were treated with soluble Eph4B in combination with anti-PD1 therapy, biopsies were obtained prior to and during the course of treatment. Biopsies were used for analysis of localized protein and RNA expression by GeoMx Digital Spatial Profiling (DSP). DSP analysis focused on tumor rich regions of interest (ROIs), adjacent stromal immune populations and microniches around vascular sites, with emphasis on sites where CD45+ T-cells were observed to be surrounding capillaries within and surrounding the tumor, presumably from extravasation.ResultsIn preclinical mouse models, EphB4 was found to induce several inflammatory pathways as a monotherapy including key immunomodulatory checkpoints such as PD1, PDL1, PDL2. Similarly, patients enrolled in this study were observed to have elevated T-cell infiltration in primary and secondary tumor sites, resulting in tumor mass reduction in post-treatment observations. DSP between matched samples discovered interesting differences in T-cell populations between both protein and mRNA expression. We observed evidence of tumor-debulking by decreased expression of epithelial markers such as Pan-cytokeratin and S100B within tumor ROIs, and increased infiltration within these ROIs measured by immune cell markers such as CD3 and CD163. Additionally, we observed increased GZMA expression post-treatment in perivascular regions suggestive of higher ongoing response by cells entering the tumor microenvironment. Additional analysis of localized RNA expression provided further support for activation of inflammatory cascades in post-treatment samples.ConclusionsThese discoveries provide insights into the mechanism of action of EphB4 combination therapy in bladder cancer, providing support for a role of EphB4 acting as an adjuvant for PD1 therapy. Our results highlight the ability of EphB4 to activate the immune system both in preclinical models and in key structures within the tumor microenvironment during combination therapy.Trial RegistrationNAEthics ApprovalThe studies were approved by USC IRB Protocol 4B 15-11 and IACUC Protocol 20570.

scholarly journals An integrated multi-omics analysis identifies prognostic molecular subtypes of non-muscle-invasive bladder cancer

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Sia Viborg Lindskrog ◽  
Frederik Prip ◽  
Philippe Lamy ◽  
Ann Taber ◽  
Clarice S. Groeneveld ◽  
...  
Keyword(s):  
Bladder Cancer ◽  
Chromosomal Instability ◽  
Prognostic Value ◽  
Immune Cell ◽  
Cell Infiltration ◽  
Invasive Bladder Cancer ◽  
Immune Cell Infiltration ◽  
Muscle Invasive Bladder Cancer ◽  
Omics Analysis ◽  
Muscle Invasive

AbstractThe molecular landscape in non-muscle-invasive bladder cancer (NMIBC) is characterized by large biological heterogeneity with variable clinical outcomes. Here, we perform an integrative multi-omics analysis of patients diagnosed with NMIBC (n = 834). Transcriptomic analysis identifies four classes (1, 2a, 2b and 3) reflecting tumor biology and disease aggressiveness. Both transcriptome-based subtyping and the level of chromosomal instability provide independent prognostic value beyond established prognostic clinicopathological parameters. High chromosomal instability, p53-pathway disruption and APOBEC-related mutations are significantly associated with transcriptomic class 2a and poor outcome. RNA-derived immune cell infiltration is associated with chromosomally unstable tumors and enriched in class 2b. Spatial proteomics analysis confirms the higher infiltration of class 2b tumors and demonstrates an association between higher immune cell infiltration and lower recurrence rates. Finally, the independent prognostic value of the transcriptomic classes is documented in 1228 validation samples using a single sample classification tool. The classifier provides a framework for biomarker discovery and for optimizing treatment and surveillance in next-generation clinical trials.

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