Transcriptomic analyses of myeloid-derived suppressor cell subsets in the circulation of colorectal cancer patients

Myeloid-derived suppressor cells (MDSCs) promote tumor immune evasion and favor tumorigenesis by activating various tumor-promoting downstream signals. MDSC expansion is evident in the circulation and tumor microenvironment of many solid tumors including colorectal cancer (CRC). We have recently reported the transcriptomic profiles of tumor-infiltrating MDSCs in CRC patients and uncovered pathways, which could potentially assist tumor progression. In this study, we sorted different subsets of circulating MDSCs in CRC patients and investigated their transcriptomic profiles in order to disclose pathways, which could potentially contribute to disease progression. The sorted subsets included polymorphonuclear/granulocytic MDSCs (PMN-MDSCs), immature MDSCs (I-MDSCs), and monocytic MDSCs (M-MDSCs). Our functional annotation analyses revealed that multiple pathways including DNA damage-, chemotaxis-, apoptosis-, mitogen-activated protein kinase-, transforming growth factor β-, and myeloid differentiation–related transcripts were higher in PMN-MDSCs, compared with monocytic antigen-presenting cells (APCs) or I-MDSCs. Furthermore, genes related to Janus kinase (JAK)–signal transducer and activator of transcription (STAT) were also elevated in PMN-MDSCs. These data suggest that upregulation of JAK-STAT pathway could trigger multiple downstream targets in PMN-MDSCs, which favor tumor progression. Additionally, we found that pathways including phosphatidyl inositol 3-kinase (PI3K), interleukin 6, and TGF-β in M-MDSCs and cell cycle–related pathways in I-MDSCs were upregulated, compared with monocytic APCs. Moreover, acetylation-related genes were upregulated in both PMN-MDSCs and M-MDSCs. This latter finding implicates that epigenetic modifications could also play a role in the regulation of multiple tumor-promoting genes in PMN-MDSCs and M-MDSCs. Taken together, this study reveals various signaling pathways, which regulate the function of MDSC subsets in the circulation of CRC patients. However, functional studies are warranted to support these findings

Transcriptomic profiling disclosed the role of DNA methylation and histone modifications in tumor-infiltrating myeloid-derived suppressor cell subsets in colorectal cancer

Increased numbers of myeloid-derived suppressor cells (MDSCs) are positively correlated with poor prognosis and reduced survivals of cancer patients. They play central roles in tumor immune evasion and tumor metastasis. However, limited data are available on phenotypic/transcriptomic characteristics of the different MDSCs subsets in cancer. These cells include immature (I-MDSCs), monocytic (M-MDSCs), and polymorphonuclear/granulocytic (PMN-MDSCs). Phenotypic characterization of myeloid subsets from 27 colorectal cancer (CRC) patients was assessed by flow cytometric analyses. RNA-sequencing of sorted I-MDSCs, PMN-MDSCs, and antigen-presenting cells (APCs) was also performed. We found that the levels of I-MDSCs and PMN-MDSCs were increased in tumor tissues (TT), compared with normal tissues (NT) in colorectal cancer. Our functional annotation analyses showed that genes associated with histone deacetylase (HDAC) activation- and DNA methylation-mediated transcriptional silencing were upregulated, and histone acetyl transferase (HAT)-related genes were downregulated in tumor-infiltrating I-MDSCs. Moreover, pathways implicated in cell trafficking and immune suppression, including Wnt, interleukin-6 (IL-6), and mitogen-activated protein kinase (MAPK) signaling, were upregulated in I-MDSCs. Notably, PMN-MDSCs showed downregulation in genes related to DNA methylation and HDAC binding. Using an ex vivo model, we found that inhibition of HDAC activation or neutralization of IL-6 in CRC tumor tissues downregulates the expression of genes associated with immunosuppression and myeloid cell chemotaxis, confirming the importance of HDAC activation and IL-6 signaling pathway in MDSC function and chemotaxis. This study provides novel insights into the epigenetic regulations and other molecular pathways in different myeloid cell subsets within the CRC tumor microenvironment (TME), giving opportunities to potential targets for therapeutic benefits

Transcriptomic profiling of tumor-infiltrating CD4 + TIM-3 + T Cells reveals their suppressive, exhausted, and metastatic characteristics in colorectal cancer patients

T cell immunoglobulin mucin-3 (TIM-3) is an immune checkpoint identified as one of the key players in regulating T-cell responses. Studies have shown that TIM-3 is upregulated in the tumor microenvironment (TME). However, the precise role of TIM-3 in colorectal cancer (CRC) TME is yet to be elucidated. We performed phenotypic and molecular characterization of TIM-3+ T cells in the TME and circulation of CRC patients by analyzing tumor tissues (TT, TILs), normal tissues (NT, NILs), and peripheral blood mononuclear cells (PBMC). TIM-3 was upregulated on both CD4+ and CD3+CD4− (CD8+) TILs. CD4+TIM-3+ TILs expressed higher levels of T regulatory cell (Tregs)-signature genes, including FoxP3 and Helios, compared with their TIM-3− counterparts. Transcriptomic and ingenuity pathway analyses showed that TIM-3 potentially activates inflammatory and tumor metastatic pathways. Moreover, NF-κB-mediated transcription factors were upregulated in CD4+TIM-3+ TILs, which could favor proliferation/invasion and induce inflammatory and T-cell exhaustion pathways. In addition, we found that CD4+TIM-3+ TILs potentially support tumor invasion and metastasis, compared with conventional CD4+CD25+ Tregs in the CRC TME. However, functional studies are warranted to support these findings. In conclusion, this study discloses some of the functional pathways of TIM-3+ TILs, which could improve their targeting in more specific therapeutic approaches in CRC patients

Integrated whole transcriptome and small RNA analysis revealed multiple regulatory networks in colorectal cancer

Colorectal cancer (CRC) remains a global disease burden and a leading cause of cancer related deaths worldwide. The identification of aberrantly expressed messenger RNA (mRNA), long non-coding RNA (lncRNA), and microRNA (miRNA), and the resulting molecular interactions and signaling networks is essential for better understanding of CRC, identification of novel diagnostic biomarkers and potential development of therapeutic interventions. Herein, we performed microRNA (miRNA) sequencing on fifteen CRC and their non-tumor adjacent tissues and whole transcriptome RNA-Seq on six paired samples from the same cohort and identified alterations in miRNA, mRNA, and lncRNA expression. Computational analyses using Ingenuity Pathway Analysis (IPA) identified multiple activated signaling networks in CRC, including ERBB2, RABL6, FOXM1, and NFKB networks, while functional annotation highlighted activation of cell proliferation and migration as the hallmark of CRC. IPA in combination with in silico prediction algorithms and experimentally validated databases gave insight into the complex associations and interactions between downregulated miRNAs and upregulated mRNAs in CRC and vice versa. Additionally, potential interaction between differentially expressed lncRNAs such as H19, SNHG5, and GATA2-AS1 with multiple miRNAs has been revealed. Taken together, our data provides thorough analysis of dysregulated protein-coding and non-coding RNAs in CRC highlighting numerous associations and regulatory networks thus providing better understanding of CRC

RNA-seq analysis of colorectal tumor-infiltrating myeloid-derived suppressor cell subsets revealed gene signatures of poor prognosis

Elevated levels of myeloid-derived suppressor cells (MDSCs), including polymorphonuclear MDSCs (PMN-MDSCs) and immature MDSCs (I-MDSCs), are usually associated with disease progression in cancer patients, including colorectal cancer (CRC). However, biological mechanisms and molecular pathways regulated by MDSC subpopulations in the CRC tumor microenvironment (TME) have not been fully investigated. In this study, we performed transcriptomic analysis of tumor-infiltrating I-MDSCs and PMN-MDSCs isolated from tumor tissues of six CRC patients, compared to antigen-presenting cells (APCs). We also compared the transcriptomic profiles of tumor-infiltrating PMN-MDSCs to I-MDSCs. Our results showed different molecular pathways regulated by each MDSC subset, potentially reflecting their phenotypical/molecular/functional characteristics in the CRC TME. Moreover, we identified gene signatures in PMN-MDSC and I-MDSC of poor overall survival (OS) and disease-free survival (DFS) using the Cancer Genome Atlas (TCGA) dataset from patients with colon adenocarcinoma (COAD). However, functional studies are required to validate these findings