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Multiregion single-cell sequencing reveals the transcriptional landscape of the immune microenvironment of colorectal cancer.

ABSTRACT: The tumor microenvironment is a complex ecosystem formed by distinct and interacting cell populations, and its composition is related to cancer prognosis and response to clinical treatment. In this study, we have taken the advantage of two single-cell RNA sequencing technologies (Smart-seq2 and DNBelab C4) to generate an atlas of 15,115 immune and nonimmune cells from primary tumors and hepatic metastases of 18 colorectal cancer (CRC) patients. We observed extensive changes in the proportions and functional states of T cells and B cells in tumor tissues, compared to those of paired non-tumor tissues. Importantly, we found that B cells from early CRC tumor were identified to be pre-B like expressing tumor suppressors, whereas B cells from advanced CRC tumors tended to be developed into plasma cells. We also identified the association of IgA⁺ IGLC2⁺ plasma cells with poor CRC prognosis, and demonstrated a significant interaction between B-cell and myeloid-cell signaling, and found CCL8⁺ cycling B cells/CCR5⁺ T-cell interactions as a potential antitumoral mechanism in advanced CRC tumors. Our results provide deeper insights into the immune infiltration within CRC, and a new perspective for the future research in immunotherapies for CRC.

SUBMITTER: Wang W

PROVIDER: S-EPMC7775989 | biostudies-literature | 2021 Jan

REPOSITORIES: biostudies-literature

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Multiregion single-cell sequencing reveals the transcriptional landscape of the immune microenvironment of colorectal cancer.

Wang Wei W Zhong Yu Y Zhuang Zhenkun Z Xie Jiarui J Lu Yueer Y Huang Chengzhi C Sun Yan Y Wu Liang L Yin Jianhua J Yu Hang H Jiang Zhiqiang Z Wang Shanshan S Wang Chunqing C Zhang Yuanhang Y Huang Yilin Y Han Chongyin C Zhong Zhenggang Z Hu Jialin J Ouyang Ying Y Liu Huisheng H Yu Mengya M Wei Xiaochan X Chen Dandan D Huang Lizhen L Hou Yong Y Lin Zhanglin Z Liu Shiping S Ling Fei F Yao Xueqing X

Clinical and translational medicine 20210101 1

The tumor microenvironment is a complex ecosystem formed by distinct and interacting cell populations, and its composition is related to cancer prognosis and response to clinical treatment. In this study, we have taken the advantage of two single-cell RNA sequencing technologies (Smart-seq2 and DNBelab C4) to generate an atlas of 15,115 immune and nonimmune cells from primary tumors and hepatic metastases of 18 colorectal cancer (CRC) patients. We observed extensive changes in the proportions an ...[more]

PMID: 33463049

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Project description:BackgroundThe objective response rate of microsatellite instability-high (MSI-H) metastatic colorectal cancer (mCRC) patients with first-line anti-programmed cell death protein-1 (PD-1) monotherapy is only 40-45%. Single-cell RNA sequencing (scRNA-seq) enables unbiased analysis of the full variety of cells comprising the tumor microenvironment. Thus, we used scRNA-seq to assess differences among microenvironment components between therapy-resistant and therapy-sensitive groups in MSI-H/mismatch repair-deficient (dMMR) mCRC. Resistance-related cell types and genes identified by this analysis were subsequently verified in clinical samples and mouse models to further reveal the molecular mechanism of anti-PD-1 resistance in MSI-H or dMMR mCRC.MethodsThe response of primary and metastatic lesions to first-line anti-PD-1 monotherapy was evaluated by radiology. Cells from primary lesions of patients with MSI-H/dMMR mCRC were analyzed using scRNA-seq. To identify the marker genes in each cluster, distinct cell clusters were identified and subjected to subcluster analysis. Then, a protein‒protein interaction network was constructed to identify key genes. Immunohistochemistry and immunofluorescence were applied to verify key genes and cell marker molecules in clinical samples. Immunohistochemistry, quantitative real-time PCR, and western blotting were performed to examine the expression of IL-1β and MMP9. Moreover, quantitative analysis and sorting of myeloid-derived suppressor cells (MDSCs) and CD8+ T cells were performed using flow cytometry.ResultsTumor responses in 23 patients with MSI-H/dMMR mCRC were evaluated by radiology. The objective response rate was 43.48%, and the disease control rate was 69.57%. ScRNA-seq analysis showed that, compared with the treatment-resistant group, the treatment-sensitive group accumulated more CD8+ T cells. Experiments with both clinical samples and mice indicated that infiltration of IL-1β-driven MDSCs and inactivation of CD8+ T cells contribute to anti-PD-1 resistance in MSI-H/dMMR CRC.ConclusionsCD8+ T cells and IL-1β were identified as the cell type and gene, respectively, with the highest correlation with anti-PD-1 resistance. Infiltration of IL-1β-driven MDSCs was a significant factor in anti-PD-1 resistance in CRC. IL-1β antagonists are expected to be developed as a new treatment for anti-PD-1 inhibitor resistance.

Project description:Brain metastases is the most common intracranial tumor and account for approximately 20% of all systematic cancer cases. It is a leading cause of death in advanced-stage cancer, resulting in a five-year overall survival rate below 10%. Therefore, there is a critical need to identify effective biomarkers that can support frequent surveillance and promote efficient drug guidance in brain metastasis. Recently, the remarkable breakthroughs in single-cell RNA-sequencing (scRNA-seq) technology have advanced our insights into the tumor microenvironment (TME) at single-cell resolution, which offers the potential to unravel the metastasis-related cellular crosstalk and provides the potential for improving therapeutic effects mediated by multifaceted cellular interactions within TME. In this study, we have applied scRNA-seq and profiled 10,896 cells collected from five brain tumor tissue samples originating from breast and lung cancers. Our analysis reveals the presence of various intratumoral components, including tumor cells, fibroblasts, myeloid cells, stromal cells expressing neural stem cell markers, as well as minor populations of oligodendrocytes and T cells. Interestingly, distinct cellular compositions are observed across different samples, indicating the influence of diverse cellular interactions on the infiltration patterns within the TME. Importantly, we identify tumor-associated fibroblasts in both our in-house dataset and external scRNA-seq datasets. These fibroblasts exhibit high expression of type I collagen genes, dominate cell-cell interactions within the TME via the type I collagen signaling axis, and facilitate the remodeling of the TME to a collagen-I-rich extracellular matrix similar to the original TME at primary sites. Additionally, we observe M1 activation in native microglial cells and infiltrated macrophages, which may contribute to a proinflammatory TME and the upregulation of collagen type I expression in fibroblasts. Furthermore, tumor cell-specific receptors exhibit a significant association with patient survival in both brain metastasis and native glioblastoma cases. Taken together, our comprehensive analyses identify type I collagen-secreting tumor-associated fibroblasts as key mediators in metastatic brain tumors and uncover tumor receptors that are potentially associated with patient survival. These discoveries provide potential biomarkers for effective therapeutic targets and intervention strategies.

Project description:IntroductionColorectal cancer (CRC) is a highly heterogeneous cancer. The molecular and cellular characteristics differ between the colon and rectal cancer type due to the differences in their anatomical location and pathological properties. With the advent of single-cell sequencing, it has become possible to analyze inter- and intra-tumoral tissue heterogeneities.MethodsA comprehensive CRC immune atlas, comprising 62,398 immune cells, was re-structured into 33 immune cell clusters at the single-cell level. Further, the immune cell lineage heterogeneity of colon, rectal, and paracancerous tissues was explored. Simultaneously, we characterized the TAM phenotypes and analyzed the transcriptomic factor regulatory network of each macrophage subset using SCENIC. In addition, monocle2 was used to elucidate the B cell developmental trajectory. The crosstalk between immune cells was explored using CellChat and the patterns of incoming and outgoing signals within the overall immune cell population were identified. Afterwards, the bulk RNA-sequencing data from The Cancer Genome Atlas (TCGA) were combined and the relative infiltration abundance of the identified subpopulations was analyzed using CIBERSORT. Moreover, cell composition patterns could be classified into five tumor microenvironment (TME) subtypes by employing a consistent non-negative matrix algorithm. Finally, the co-expression and interaction between SPP1+TAMs and Treg cells in the tumor microenvironment were analyzed by multiplex immunohistochemistry.ResultsIn the T cell lineage, we found that CXCL13+T cells were more widely distributed in colorectal cancer tissues, and the proportion of infiltration was increased. In addition, Th17 was found accounted for the highest proportion in CD39+CD101+PD1+T cells. Mover, Ma1-SPP1 showed the characteristics of M2 phenotypes and displayed an increased proportion in tumor tissues, which may promote angiogenesis. Plasma cells (PCs) displayed a significantly heterogeneous distribution in tumor as well as normal tissues. Specifically, the IgA+ PC population could be shown to be decreased in colorectal tumor tissues whereas the IgG+ PC one was enriched. In addition, information flow mediated by SPP1 and CD44, regulate signaling pathways of tumor progression. Among the five TME subtypes, the TME-1 subtype displayed a markedly reduced proportion of T-cell infiltration with the highest proportion of macrophages which was correlated to the worst prognosis. Finally, the co-expression and interaction between SPP1+TAMs and Treg cells were observed in the CD44 enriched region.DiscussionThe heterogeneity distribution and phenotype of immune cells were analyzed in colon cancer and rectal cancer at the single-cell level. Further, the prognostic role of major tumor-infiltrating lymphocytes and TME subtypes in CRC was evaluated by integrating bulk RNA. These findings provide novel insight into the immunotherapy of CRC.

Dataset Information

Multiregion single-cell sequencing reveals the transcriptional landscape of the immune microenvironment of colorectal cancer.

Publications

Multiregion single-cell sequencing reveals the transcriptional landscape of the immune microenvironment of colorectal cancer.

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