Genomic and transcriptomic correlates of immunotherapy response within the tumor microenvironment of leptomeningeal metastases

Abstract Leptomeningeal disease (LMD) is a devastating complication of solid tumor malignancies, with dire prognosis and no effective systemic treatment options. Over the past decade, the incidence of LMD has steadily increased due to therapeutics that have extended the survival of cancer patients, highlighting the need for new interventions. To examine the efficacy of immune checkpoint inhibitors (ICI) in patients with LMD, we completed two phase II clinical trials utilizing either Pembrolizumab alone or the combination of Ipilimumab and Nivolumab. We investigated the cellular and molecular features underpinning observed patient trajectories in these trials by applying single-cell RNA and cell-free DNA profiling to longitudinal cerebrospinal fluid (CSF) draws from enrolled patients. We isolated and sequenced 34,742 cells from both the malignant and immune compartment within CSF. Amongst the 19 patients included in the cohort, there were 13 pre-treatment and 24 post-treatment samples, and 9 patients were sampled across multiple timepoints. We detected dynamic changes in immune cell recruitment into the CSF and activation within 30 days of ICI, including increased effector T cell activation and IFN-gamma response pathways within T cells. Moreover, the overall level of IFN-gamma response and antigen processing within 30 days of ICI in malignant cells correlated with survival past clinical trial primary endpoint. Lastly, we observed evidence of longitudinal outgrowth of distinct immunogenic clones over the course of ICI. Overall, our study describes the liquid LMD tumor microenvironment prior to and following ICI treatment and provides unique insights into the compartmental and temporal variation during the course of ICI. Moreover, our findings demonstrate the clinical utility of cell- free and single-cell genomic measurements for LMD research.

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Patient-derived micro-organospheres recapitulate tumor microenvironment and heterogeneity for precision oncology.

Journal of Clinical Oncology ◽

10.1200/jco.2021.39.15_suppl.3076 ◽

2021 ◽

Vol 39 (15_suppl) ◽

pp. 3076-3076

Author(s):

Shengli Ding ◽

Zhaohui Wang ◽

Marcos Negrete Obando ◽

Grecia rivera Palomino ◽

Tomer Rotstein ◽

...

Keyword(s):

Drug Discovery ◽

Tumor Microenvironment ◽

Single Cell ◽

Stromal Cells ◽

Mononuclear Cells ◽

Immune Cell ◽

Tumor Biology ◽

Cell Types ◽

Precision Oncology ◽

Original Tumor

3076 Background: Preclinical models that can recapitulate patients’ intra-tumoral heterogeneity and microenvironment are crucial for tumor biology research and drug discovery. In particular, the ability to retain immune and other stromal cells in the microenvironment is vital for the development of immuno-oncology assays. However, current patient-derived organoid (PDO) models are largely devoid of immune components. Methods: We first developed an automated microfluidic and membrane platform that can generate tens of thousands of micro-organospheres from resected or biopsied clinical tumor specimens within an hour. We next characterized growth rate and drug response of micro-organospheres. Finally, extensive single-cell RNA-seq profiling were performed on both micro-organospheres and original tumor samples from lung, ovarian, kidney, and breast cancer patients. Results: Micro-organospheres derived from clinical tumor samples preserved all original tumor and stromal cells, including fibroblasts and all immune cell types. Single-cell analysis revealed that unsupervised clustering of tumor and non-tumor cells were identical between original tumors and the derived micro-organospheres. Quantification showed similar cell composition and percentages for all cell types and also preserved functional intra-tumoral heterogeneity.. An automated, end-to-end, high-throughput drug screening pipeline demonstrated that matched peripheral blood mononuclear cells (PBMCs) from the same patient added to micro-organospheres can be used to assess the efficacy of immunotherapy moieties. Conclusions: Micro-organospheres are a rapid and scalable platform to preserve patient tumor microenvironment and heterogeneity. This platform will be useful for precision oncology, drug discovery, and immunotherapy development. Funding sources: NIH U01 CA217514, U01 CA214300, Duke Woo Center for Big Data and Precision Health

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Applications of Single-Cell Omics to Dissect Tumor Microenvironment

Frontiers in Genetics ◽

10.3389/fgene.2020.548719 ◽

2020 ◽

Vol 11 ◽

Author(s):

Tingting Guo ◽

Weimin Li ◽

Xuyu Cai

Keyword(s):

Tumor Microenvironment ◽

Single Cell ◽

Immune Cells ◽

Immune Cell ◽

Cell Types ◽

Immune Checkpoint Blockade ◽

Lineage Tracing ◽

Great Success ◽

Novel Technologies ◽

Single Cell Sequencing

The recent technical and computational advances in single-cell sequencing technologies have significantly broaden our toolkit to study tumor microenvironment (TME) directly from human specimens. The TME is the complex and dynamic ecosystem composed of multiple cell types, including tumor cells, immune cells, stromal cells, endothelial cells, and other non-cellular components such as the extracellular matrix and secreted signaling molecules. The great success on immune checkpoint blockade therapy has highlighted the importance of TME on anti-tumor immunity and has made it a prime target for further immunotherapy strategies. Applications of single-cell transcriptomics on studying TME has yielded unprecedented resolution of the cellular and molecular complexity of the TME, accelerating our understanding of the heterogeneity, plasticity, and complex cross-interaction between different cell types within the TME. In this review, we discuss the recent advances by single-cell sequencing on understanding the diversity of TME and its functional impact on tumor progression and immunotherapy response driven by single-cell sequencing. We primarily focus on the major immune cell types infiltrated in the human TME, including T cells, dendritic cells, and macrophages. We further discuss the limitations of the existing methodologies and the prospects on future studies utilizing single-cell multi-omics technologies. Since immune cells undergo continuous activation and differentiation within the TME in response to various environmental cues, we highlight the importance of integrating multimodal datasets to enable retrospective lineage tracing and epigenetic profiling of the tumor infiltrating immune cells. These novel technologies enable better characterization of the developmental lineages and differentiation states that are critical for the understanding of the underlying mechanisms driving the functional diversity of immune cells within the TME. We envision that with the continued accumulation of single-cell omics datasets, single-cell sequencing will become an indispensable aspect of the immune-oncology experimental toolkit. It will continue to drive the scientific innovations in precision immunotherapy and will be ultimately adopted by routine clinical practice in the foreseeable future.

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SCISSOR™: a single-cell inferred site-specific omics resource for tumor microenvironment association study

NAR Cancer ◽

10.1093/narcan/zcab037 ◽

2021 ◽

Vol 3 (3) ◽

Author(s):

Xiang Cui ◽

Fei Qin ◽

Xuanxuan Yu ◽

Feifei Xiao ◽

Guoshuai Cai

Keyword(s):

Tumor Microenvironment ◽

Single Cell ◽

Clinical Outcomes ◽

Large Scale ◽

Cell Types ◽

Cell Interaction ◽

Specific Cell ◽

Dynamic Visualization ◽

Tissue Specific ◽

Cell Composition

Abstract Tumor tissues are heterogeneous with different cell types in tumor microenvironment, which play an important role in tumorigenesis and tumor progression. Several computational algorithms and tools have been developed to infer the cell composition from bulk transcriptome profiles. However, they ignore the tissue specificity and thus a new resource for tissue-specific cell transcriptomic reference is needed for inferring cell composition in tumor microenvironment and exploring their association with clinical outcomes and tumor omics. In this study, we developed SCISSOR™ (https://thecailab.com/scissor/), an online open resource to fulfill that demand by integrating five orthogonal omics data of >6031 large-scale bulk samples, patient clinical outcomes and 451 917 high-granularity tissue-specific single-cell transcriptomic profiles of 16 cancer types. SCISSOR™ provides five major analysis modules that enable flexible modeling with adjustable parameters and dynamic visualization approaches. SCISSOR™ is valuable as a new resource for promoting tumor heterogeneity and tumor–tumor microenvironment cell interaction research, by delineating cells in the tissue-specific tumor microenvironment and characterizing their associations with tumor omics and clinical outcomes.

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Roles of the CXCL8-CXCR1/2 Axis in the Tumor Microenvironment and Immunotherapy

Molecules ◽

10.3390/molecules27010137 ◽

2021 ◽

Vol 27 (1) ◽

pp. 137

Author(s):

Zhi-Jian Han ◽

Yang-Bing Li ◽

Lu-Xi Yang ◽

Hui-Juan Cheng ◽

Xin Liu ◽

...

Keyword(s):

Tumor Microenvironment ◽

Tumor Progression ◽

Immune Cells ◽

Epithelial To Mesenchymal Transition ◽

Checkpoint Inhibitors ◽

Lymphatic Vessels ◽

Cell Types ◽

Mesenchymal Transition ◽

Development Of Resistance ◽

Immunosuppressive Cells

In humans, Interleukin-8 (IL-8 or CXCL8) is a granulocytic chemokine with multiple roles within the tumor microenvironment (TME), such as recruiting immunosuppressive cells to the tumor, increasing tumor angiogenesis, and promoting epithelial-to-mesenchymal transition (EMT). All of these effects of CXCL8 on individual cell types can result in cascading alterations to the TME. The changes in the TME components such as the cancer-associated fibroblasts (CAFs), the immune cells, the extracellular matrix, the blood vessels, or the lymphatic vessels further influence tumor progression and therapeutic resistance. Emerging roles of the microbiome in tumorigenesis or tumor progression revealed the intricate interactions between inflammatory response, dysbiosis, metabolites, CXCL8, immune cells, and the TME. Studies have shown that CXCL8 directly contributes to TME remodeling, cancer plasticity, and the development of resistance to both chemotherapy and immunotherapy. Further, clinical data demonstrate that CXCL8 could be an easily measurable prognostic biomarker in patients receiving immune checkpoint inhibitors. The blockade of the CXCL8-CXCR1/2 axis alone or in combination with other immunotherapy will be a promising strategy to improve antitumor efficacy. Herein, we review recent advances focusing on identifying the mechanisms between TME components and the CXCL8-CXCR1/2 axis for novel immunotherapy strategies.

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Effects of sample size on plant single-cell RNA profiling

10.21203/rs.3.rs-549533/v1 ◽

2021 ◽

Author(s):

Longbiao Guo ◽

Hongyu Chen ◽

Xinxin Yin ◽

Xi Chen ◽

Qinjie Chu ◽

...

Keyword(s):

Sample Size ◽

Single Cell ◽

High Reliability ◽

Cell Types ◽

Cell Number ◽

Cellular Heterogeneity ◽

Root Cells ◽

Rna Profiling ◽

Molecular Features ◽

Cell Clustering

Abstract Background: Single-cell RNA (scRNA) profiling or scRNA-sequencing (scRNA-seq) is a rapidly developing technology and an important frontier of molecular biology science. scRNA profiling makes it possible to parallelly investigate diverse molecular features of multiple types of cells in a given plant tissue, and promotes elucidation of cellular heterogeneity and discovery of developmental processes underpinning cell differentiation. While it is assumed that the power of scRNA profiling in uncovering cellular heterogeneity largely depends on the depth of scRNA-seq, no study about the effect of the sequenced cell numbers on the power of plant scRNA-seq has ever been reported. Results: In this study, on the basis of analyzing the sample coverage of 1,244 available scRNA-seq studies (including 30 in plants) and the effect of sample coverage on cell clustering and identification of cell types, we evaluated the effects of sample size (i.e., cell number) on the outcome of single cell transcriptome analysis by sampling different number of cells from a pool of ~57,000 Arabidopsis thaliana root cells integrated from five published studies. Our results indicated that the most significant principle components could be achieved when 20,000-30,000 cells were sampled, a relatively high reliability of cell clustering could be achieved by using ~20,000 cells with little further improvement by using more cells, 96% of the differentially expressed genes could be successfully identified with no more than 20,000 cells, and a relatively stable pseudotime could be estimated in the sub-sample with 5,000 cells. Conclusions: Our results imply that ~20,000 (or 10,000 - 30,000[1] ) cells are enough for profiling Arabidopsis root cells using scRNA-seq, although the applicability of this number to other Arabidopsis tissues and other plants is yet to be further determined by analyzing scRNA-seq data generated from diverse tissues of different plant species. Nevertheless, our results provide a general guide for optimizing sample size to be used in plant scRNA-seq studies. Change to “or up to 300000”?

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Blood–Brain Barrier, Cell Junctions, and Tumor Microenvironment in Brain Metastases, the Biological Prospects and Dilemma in Therapies

Frontiers in Cell and Developmental Biology ◽

10.3389/fcell.2021.722917 ◽

2021 ◽

Vol 9 ◽

Author(s):

Zhiyuan Guan ◽

Hongyu Lan ◽

Xin Cai ◽

Yichi Zhang ◽

Annan Liang ◽

...

Keyword(s):

Tumor Microenvironment ◽

Brain Metastasis ◽

Checkpoint Inhibitors ◽

Target Therapy ◽

Tumor Hypoxia ◽

Cell Types ◽

Cell Junctions ◽

Primary Tumors ◽

Tumor Microenvironments ◽

Brain Radiotherapy

Brain metastasis is the most commonly seen brain malignancy, frequently originating from lung cancer, breast cancer, and melanoma. Brain tumor has its unique cell types, anatomical structures, metabolic constraints, and immune environment, which namely the tumor microenvironment (TME). It has been discovered that the tumor microenvironment can regulate the progression, metastasis of primary tumors, and response to the treatment through the particular cellular and non-cellular components. Brain metastasis tumor cells that penetrate the brain–blood barrier and blood–cerebrospinal fluid barrier to alter the function of cell junctions would lead to different tumor microenvironments. Emerging evidence implies that these tumor microenvironment components would be involved in mechanisms of immune activation, tumor hypoxia, antiangiogenesis, etc. Researchers have applied various therapeutic strategies to inhibit brain metastasis, such as the combination of brain radiotherapy, immune checkpoint inhibitors, and monoclonal antibodies. Unfortunately, they hardly access effective treatment. Meanwhile, most clinical trials of target therapy patients with brain metastasis are always excluded. In this review, we summarized the clinical treatment of brain metastasis in recent years, as well as their influence and mechanisms underlying the differences between the composition of tumor microenvironments in the primary tumor and brain metastasis. We also look forward into the feasibility and superiority of tumor microenvironment-targeted therapies in the future, which may help to improve the strategy of brain metastasis treatment.

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Proteasome Inhibition: Thinking outside the Box

Clinical Medicine Therapeutics ◽

10.4137/cmt.s3072 ◽

2009 ◽

Vol 1 ◽

pp. CMT.S3072

Author(s):

Michael B. Armstrong

Keyword(s):

Multiple Myeloma ◽

Cancer Therapy ◽

Treatment Options ◽

Proteasome Inhibition ◽

Cell Types ◽

Malignant Cell ◽

26S Proteasome ◽

Malignant Neoplasms ◽

New Era

A new era in cancer therapy is emerging with the development of tumor-specific agents exhibiting less toxicity. Since the advent of imatinib, several tumor-directed treatment options have been developed. However, therapies not directed specifically at a tumor target also have potential benefits. The 26S proteasome is a critical regulator of cell homeostasis through the degradation of key signaling molecules including p21, p27, and p53. Additionally, the proteasome degrades I-κB which inhibits the activity of NF-κB, an important promoter of cell proliferation. Blocking function of the proteasome disrupts tumor growth by shifting the balance of the cell from proliferation to apoptosis. In vitro, the proteasome inhibitor, bortezomib, inhibits NF-κB activity and prevents growth of several malignant cell types including multiple myeloma. Given the central role of NF-κB in the pathogenesis of multiple myeloma, bortezomib was a good candidate for use in therapy. Treatment of heavily pre-treated patients with bortezomib led to response rates of 30%-40%. More importantly, bortezomib led to improvements in bone metabolism, a major cause of morbidity in multiple myeloma. This effect was seen independent of the response of the myeloma. This finding correlates with in vitro studies which demonstrate increased BMP2 expression and osteoblast number after exposure to bortezomib. Moreover, bortezomib blocks NF-κB-mediated angiogenesis and tumor cell metastasis. While tumor-targeted treatments have an important role in the future of cancer therapy, these examples show that it is important not to lose sight of the benefits of less-specific agents in the treatment of malignant neoplasms.

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A Systematic Review on the Therapeutic Potentiality of PD-L1-Inhibiting MicroRNAs for Triple-Negative Breast Cancer: Toward Single-Cell Sequencing-Guided Biomimetic Delivery

Genes ◽

10.3390/genes12081206 ◽

2021 ◽

Vol 12 (8) ◽

pp. 1206

Author(s):

Mahdi Abdoli Shadbad ◽

Sahar Safaei ◽

Oronzo Brunetti ◽

Afshin Derakhshani ◽

Parisa Lotfinejad ◽

...

Keyword(s):

Tumor Microenvironment ◽

Single Cell ◽

Immune Checkpoint ◽

Triple Negative ◽

De Novo ◽

Checkpoint Inhibitors ◽

Oncogenic Signaling ◽

Single Cell Sequencing ◽

Oncogenic Signaling Pathways ◽

Tumoral Cells

The programmed death-ligand 1 (PD-L1)/programmed cell death protein 1 (PD-1) is a well-established inhibitory immune checkpoint axis in triple-negative breast cancer (TNBC). Growing evidence indicates that tumoral PD-L1 can lead to TNBC development. Although conventional immune checkpoint inhibitors have improved TNBC patients’ prognosis, their effect is mainly focused on improving anti-tumoral immune responses without substantially regulating oncogenic signaling pathways in tumoral cells. Moreover, the conventional immune checkpoint inhibitors cannot impede the de novo expression of oncoproteins, like PD-L1, in tumoral cells. Accumulating evidence has indicated that the restoration of specific microRNAs (miRs) can downregulate tumoral PD-L1 and inhibit TNBC development. Since miRs can target multiple mRNAs, miR-based gene therapy can be an appealing approach to inhibit the de novo expression of oncoproteins, like PD-L1, restore anti-tumoral immune responses, and regulate various intracellular singling pathways in TNBC. Therefore, we conducted the current systematic review based on the preferred reporting items for systematic reviews and meta-analyses (PRISMA) to provide a comprehensive and unbiased synthesis of currently available evidence regarding the effect of PD-L1-inhibiting miRs restoration on TNBC development and tumor microenvironment. For this purpose, we systematically searched the Cochrane Library, Embase, Scopus, PubMed, ProQuest, Web of Science, Ovid, and IranDoc databases to obtain the relevant peer-reviewed studies published before 25 May 2021. Based on the current evidence, the restoration of miR-424-5p, miR-138-5p, miR-570-3p, miR-200c-3p, miR-383-5p, miR-34a-5p, miR-3609, miR-195-5p, and miR-497-5p can inhibit tumoral PD-L1 expression, transform immunosuppressive tumor microenvironment into the pro-inflammatory tumor microenvironment, inhibit tumor proliferation, suppress tumor migration, enhance chemosensitivity of tumoral cells, stimulate tumor apoptosis, arrest cell cycle, repress the clonogenicity of tumoral cells, and regulate various oncogenic signaling pathways in TNBC cells. Concerning the biocompatibility of biomimetic carriers and the valuable insights provided by the single-cell sequencing technologies, single-cell sequencing-guided biomimetic delivery of these PD-L1-inhibiting miRs can decrease the toxicity of traditional approaches, increase the specificity of miR-delivery, enhance the efficacy of miR delivery, and provide the affected patients with personalized cancer therapy.

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Pan-cancer analysis identified inflamed microenvironment associated multi-omics signatures

10.1101/2020.03.17.996199 ◽

2020 ◽

Author(s):

Ben Wang ◽

Mengmeng Liu ◽

Zhujie Ran ◽

Xin Li ◽

Jie Li ◽

...

Keyword(s):

Tumor Microenvironment ◽

Single Cell ◽

Histone Deacetylase Inhibitors ◽

Immune Cell ◽

Cell Interaction ◽

Molecular Characteristics ◽

Molecular Features ◽

Phenotype Transformation ◽

Multiple Drugs ◽

Pan Cancer

AbstractBackgroundImmunotherapy has revolutionized cancer therapy. However, responses are not universal. The inflamed tumor microenvironment has been reported to correlate with response in tumor patients. However, how different tumors shape their tumor microenvironment remains a critical unsolved problem. A deeper insight into the molecular characteristics of inflamed tumor microenvironment may be needed.Materials and methodsHere, based on single-cell RNA sequencing technology and TCGA pan-cancer cohort, we investigated multi-omics molecular features of tumor microenvironment phenotypes. Based on single-cell RNA-seq analysis, we classified pan-cancer tumor samples into inflamed or non-inflamed tumor and identified molecular features of these tumors. Analysis of integrating identified gene signatures with a drug-genomic perturbation database identified multiple drugs which may be helpful for converting non-inflamed tumors to inflamed tumors.ResultsOur results revealed several inflamed/non-inflamed tumor microenvironments-specific molecular characteristics. For example, inflamed tumors highly expressed miR-650 and lncRNA including MIR155HG and LINC00426, these tumors showed activated cytokines-related signaling pathways. Interestingly, non-inflamed tumors tended to express several genes related to neurogenesis. Multi-omics analysis demonstrated the neuro phenotype transformation may be induced by hypomethylated promoters of these genes and down-regulated miR-650. Drug discovery analysis revealed histone deacetylase inhibitors may be a potential choice for helping favorable tumor microenvironment phenotype transformation and aiding current immunotherapy.ConclusionOur results provide a comprehensive molecular-level understanding of tumor cell-immune cell interaction and may have profound clinical implications.

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