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There is an increased risk of colorectal cancer (CRC) development in patients with non-insulin-dependent type 2 diabetes. CD8+ T cells have been implicated in diabetes and are crucial for anti-tumor immunity. However, transcriptomic profiling for CD8+ T cells from CRC diabetic patients has not been explored. We performed RNA sequencing and compared transcriptomic profiles of CD8+ tumor-infiltrating T lymphocytes (CD8+ TILs) in CRC diabetic patients with CRC nondiabetic patients. We found that genes associated with ribogenesis, epigenetic regulations, oxidative phosphorylation and cell cycle arrest were upregulated in CD8+ TILs from diabetic patients, while genes associated with PI3K signaling pathway, cytokine response and response to lipids were downregulated. Among the significantly deregulated 1009 genes, 342 (186 upregulated and 156 downregulated) genes were selected based on their link to diabetes, and their associations with the presence of specific CRC pathological parameters were assessed using GDC TCGA colon database. The 186 upregulated genes were associated with the presence of colon polyps history (P = 0.0007) and lymphatic invasion (P = 0.0025). Moreover, CRC patients with high expression of the 186 genes were more likely to have poorer disease-specific survival (DSS) (Mantel–Cox log-rank P = 0.024) than those with low score. Our data provide novel insights into molecular pathways and biological functions, which could be altered in CD8+ TILs from CRC diabetic versus nondiabetic patients, and reveal candidate genes linked to diabetes, which could predict DSS and pathological parameters associated with CRC progression. However, further investigations using larger patient cohorts and functional studies are required to validate these findings. [ABSTRACT FROM AUTHOR]

Tumor-infiltrating lymphocytes (TILs) play indispensable roles in the progression and response to treatment of solid tumors. However, the prognostic significance of CD4+ TILs is not fully disclosed in cancers generally and in CRC in particular, mainly due to the existence of different functional subsets of CD4+ T cells. We performed transcriptomic profiling of CD4+ TILs isolated from CRC patients in order to identify differentially expressed genes and their functional pathways in early versus advanced disease stages. We found that in advanced stages, genes related to immune and inflammatory responses, in particular Th1-mediated immune response and cytotoxicity-mediated genes, were downregulated; while epigenetic-mediated silencing genes were upregulated. Interestingly, we identified genes, which were steadily upregulated or downregulated in CD4+ TILs with CRC progression from stage I to IV. Additionally, of the top 200 deregulated genes, 43 upregulated and 64 downregulated genes showed similar deregulation trends in the cancer genome atlas CRC dataset. From these 97 deregulated genes, we identified a "poor prognosis CD4 gene signature (ppCD4sig)". Patients with high ppCD4sig score showed shorter disease-specific survival (DSS) and progression-free interval (PFI). The ppCD4sig was an independent prognostic indicator for DSS (HR = 1.73, 95% CI 1.32–2.27, P = 0.0001) and PFI (HR = 1.75, 95% CI 1.3–2.35, P = 0.0016). Additionally, patients at advanced stages and at a younger age (<55 years) were more likely to have a high ppCD4sig score. Altogether, our data provide novel insights and a unique prognostic gene signature of CD4+ TILs in the CRC microenvironment. [ABSTRACT FROM AUTHOR]

Blockade of inhibitory immune checkpoints (ICs) is a promising therapeutic approach; however, it has shown limited success in some cancers including colorectal cancer (CRC). The tumor microenvironment (TME) is largely responsible for response to therapy, and its constituents may provide robust biomarkers for successful immunotherapeutic approaches. In this study, we performed phenotypical characterization and critical analyses of key inhibitory ICs and T regulatory cell (Treg)-related markers on CD4+ T cell subsets in CRC patients, and compared with normal colon tissues and peripheral blood from the same patients. We also investigated correlations between the levels of different CD4+ T cell subsets and the clinicopathologic features including disease stage and tumor budding. We found a significant increase in the levels of CD4+FoxP3+Helios+ T cells, which represent potentially highly immunosuppressive Tregs, in the CRC TME. Additionally, tumor-infiltrating CD4+ T cells upregulated programmed cell death protein-1 (PD-1), cytotoxic T-lymphocyte-associated protein-4 (CTLA-4), T cell immunoglobulin and mucin domain-3 (TIM-3) and lymphocyte-activation gene 3 (LAG-3). We also characterized the expression of PD-1, CTLA-4, TIM-3, and LAG-3 on different CD4+FoxP3−/+Helios−/+ T cell subsets. Interestingly, we found that CTLA-4, TIM-3, and LAG-3 were mainly co-expressed on FoxP3+Helios+ Tregs in the TME. Additionally, FoxP3high Tregs expressed higher levels of Helios, CTLA-4 and TIM-3 than FoxP3low T cells. These results highlight the significance of Tregs in the CRC TME and suggest that Tregs may hamper response to IC blockade in CRC patients, but effects of different IC inhibition regimes on Treg levels or activity warrants further investigations. We also found that CD4+CTLA-4+ T cells in circulation are increased in patients with advanced disease stage. This study simultaneously provides important insights into the differential levels of CD4+ T cell subpopulations and IC expression in CRC TME, compared to periphery and associations with clinicopathologic features, which could be used as potential biomarkers for CRC progression and response to therapy. [ABSTRACT FROM AUTHOR]

Comparing pathologic outcomes for robotic versus laparoscopic surgery in rectal cancer resection: a propensity adjusted analysis of 7616 patients. Robotic surgery has gained popularity over the last decade with rapidly increasing usage of the robotic system [[1]]. In fact, recent trials suggest that for middle and low rectal cancer, robotic surgery by experienced surgeons resulted in better oncological quality of resection than conventional laparoscopic surgery, with less surgical trauma, and better postoperative recovery [[4]]. [Extracted from the article]

The article discusses a modified transanal minimally invasive surgery procedure for an ultralow rectal lesion. The management of rectal lesions has improved with advancements in surgical tools and techniques. Local excision of rectal lesions has traditionally been limited to lesions within 8 cm of the anal verge, but the transanal endoscopic microsurgery system has expanded the range of treatable lesions. Transanal minimally invasive surgery (TAMIS) has further revolutionized the treatment of mid and high rectal lesions, offering superior outcomes, a shallow learning curve, and lower costs compared to other methods. The article also provides guidelines for determining which rectal lesions are appropriate for local excision. [Extracted from the article]

Colorectal cancer (CRC) is influenced by infiltration of immune cell populations in the tumor microenvironment. While elevated levels of cytotoxic T cells are associated with improved prognosis, limited studies have reported associations between CD4+ T cells and disease outcomes. We recently performed transcriptomic profiling and comparative analyses of sorted CD4+ and CD8+ tumor-infiltrating lymphocytes (TILs) from bulk tumors of CRC patients with varying disease stages. In this study, we compared the transcriptomes of CD4+ with CD8+ TILs. Functional annotation pathway analyses revealed the downregulation of inflammatory response-related genes, while T cell activation and angiogenesis-related genes were upregulated in CD4+ TILs. The top 200 deregulated genes in CD4+ TILs were aligned with the cancer genome atlas (TCGA) CRC dataset to identify a unique gene signature associated with poor prognosis. Moreover, 69 upregulated and 20 downregulated genes showed similar trends of up/downregulation in the TCGA dataset and were used to calculate "poor prognosis score" (ppScore), which was significantly associated with disease-specific survival. High ppScore patients showed lower expression of Treg-, Th1-, and Th17-related genes, and higher expression of Th2-related genes. Our data highlight the significance of T cells within the TME and identify a unique candidate prognostic gene signature for CD4+ TILs in CRC patients. [ABSTRACT FROM AUTHOR]

Aim:  To elucidate the epigenetic alterations behind the upregulation of immune checkpoints and T cell exhaustion markers in colorectal cancer (CRC) patients. Materials & methods: mRNA expressions of different immune checkpoint/exhaustion markers were analyzed by quantitative real-time reverse transcriptase PCR and epigenetic investigations were performed using bisulfite sequencing and chromatin immunoprecipitation quantitative PCR. Results: mRNA expressions of PD-1, TIM-3, CTLA-4, PD-L1 and TOX2 were significantly upregulated in CD4 + and CD8 + tumor-infiltrating lymphocytes and bulk CRC tumor tissues. Histone 3 lysine 9 trimethylation was downregulated and histone 3 lysine 4 trimethylation was upregulated in PD-L1 and TOX2 promoters in tumor tissues, suggesting that PD-L1 and TOX2 upregulation in CRC tumors could be mediated by activating histone 3 lysine 4 trimethylation. Conclusion: Epigenetic modifications in promoters of immune checkpoint and T cell exhaustion genes could induce their upregulation, and potentially implicate the use of epigenetic modifiers to enhance antitumor immunity in CRC patients.

Tumor-infiltrating lymphocytes (TILs) play indispensable roles in the progression and response to treatment of solid tumors. However, the prognostic significance of CD4 + TILs is not fully disclosed in cancers generally and in CRC in particular, mainly due to the existence of different functional subsets of CD4 + T cells. We performed transcriptomic profiling of CD4 + TILs isolated from CRC patients in order to identify differentially expressed genes and their functional pathways in early versus advanced disease stages. We found that in advanced stages, genes related to immune and inflammatory responses, in particular Th1-mediated immune response and cytotoxicity-mediated genes, were downregulated; while epigenetic-mediated silencing genes were upregulated. Interestingly, we identified genes, which were steadily upregulated or downregulated in CD4 + TILs with CRC progression from stage I to IV. Additionally, of the top 200 deregulated genes, 43 upregulated and 64 downregulated genes showed similar deregulation trends in the cancer genome atlas CRC dataset. From these 97 deregulated genes, we identified a "poor prognosis CD4 gene signature (ppCD4sig)". Patients with high ppCD4sig score showed shorter disease-specific survival (DSS) and progression-free interval (PFI). The ppCD4sig was an independent prognostic indicator for DSS (HR = 1.73, 95% CI 1.32-2.27, P = 0.0001) and PFI (HR = 1.75, 95% CI 1.3-2.35, P = 0.0016). Additionally, patients at advanced stages and at a younger age (<55 years) were more likely to have a high ppCD4sig score. Altogether, our data provide novel insights and a unique prognostic gene signature of CD4 + TILs in the CRC microenvironment., (© 2020 The Author(s). Published with license by Taylor & Francis Group, LLC.)

Background: Cytotoxic CD8 + T cell-mediated response is the most important arm of adaptive immunity, which dictates the capacity of the host immune response in eradicating tumor cells. Due to tumor intrinsic and/or extrinsic factors, the density and function of CD8 + tumor-infiltrating lymphocytes (TILs) could be compromised, leading to poor prognosis and survival., Methods: Using RNA-Seq, transcriptomes of sorted CD3 + CD8 + TILs from treatment-naïve colorectal cancer (CRC) patients at advanced stages (III and IV) were compared with those from patients with early stages (I and II). A signature referred to as 'poor prognosis CD8 gene signature (ppCD8sig)' was identified and analyzed in The Cancer Genome Atlas CRC dataset. Scores for the ppCD8sig were calculated and classified as high, intermediate and low, and its prognostic significance was assessed using multivariate analysis and Cox proportional hazard model. Densities of CD3 + and CD8 + T cell infiltration in tumors from patients with high and low ppCD8sig scores were assessed by flow cytometry and immunostaining., Results: Genes related to epigenetic regulation and response to hypoxia were upregulated in CD8 + TILs from patients with advanced stages, while genes related to T cell activation, cell proliferation and cell cycle were downregulated. Patients with high ppCD8sig score had poorer disease-specific survival (DSS) and shorter progression-free interval (PFI). The ppCD8sig was an independent prognostic indicator for DSS (HR 1.83, 95% CI 1.40 to 2.38, p<0.0001) and PFI (HR 1.42, 95% CI 1.04 to 1.93, p=0.026). Additionally, patients with high ppCD8sig score were more likely to have advanced stages (χ 2 p<0.0001) and residual disease after primary therapy (χ 2 p=0.046). Patients with high ppCD8sig score had reduced levels of CD3 + and CD8 + TILs and low Immunoscores (IS), compared to patients with low ppCD8sig score., Conclusions: Our data provided insights into the altered regulation of biological mechanisms and signaling pathways in CD8 + TILs during CRC progression, and revealed a gene signature as an independent prognostic indicator. Patients with high ppCD8sig score had lower levels of TILs and low IS. These data further confirm the prognostic value of the identified ppCD8sig and potentially highlight its clinical relevance., Competing Interests: Competing interests: None declared., (© Author(s) (or their employer(s)) 2020. Re-use permitted under CC BY-NC. No commercial re-use. See rights and permissions. Published by BMJ.)

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., Competing Interests: The authors declare no conflicts of interest.

Blockade of inhibitory immune checkpoints (ICs) is a promising therapeutic approach; however, it has shown limited success in some cancers including colorectal cancer (CRC). The tumor microenvironment (TME) is largely responsible for response to therapy, and its constituents may provide robust biomarkers for successful immunotherapeutic approaches. In this study, we performed phenotypical characterization and critical analyses of key inhibitory ICs and T regulatory cell (Treg)-related markers on CD4 + T cell subsets in CRC patients, and compared with normal colon tissues and peripheral blood from the same patients. We also investigated correlations between the levels of different CD4 + T cell subsets and the clinicopathologic features including disease stage and tumor budding. We found a significant increase in the levels of CD4 + FoxP3 + Helios + T cells, which represent potentially highly immunosuppressive Tregs, in the CRC TME. Additionally, tumor-infiltrating CD4 + T cells upregulated programmed cell death protein-1 (PD-1), cytotoxic T-lymphocyte-associated protein-4 (CTLA-4), T cell immunoglobulin and mucin domain-3 (TIM-3) and lymphocyte-activation gene 3 (LAG-3). We also characterized the expression of PD-1, CTLA-4, TIM-3, and LAG-3 on different CD4 + FoxP3 -/+ Helios -/+ T cell subsets. Interestingly, we found that CTLA-4, TIM-3, and LAG-3 were mainly co-expressed on FoxP3 + Helios + Tregs in the TME. Additionally, FoxP3 high Tregs expressed higher levels of Helios, CTLA-4 and TIM-3 than FoxP3 low T cells. These results highlight the significance of Tregs in the CRC TME and suggest that Tregs may hamper response to IC blockade in CRC patients, but effects of different IC inhibition regimes on Treg levels or activity warrants further investigations. We also found that CD4 + CTLA-4 + T cells in circulation are increased in patients with advanced disease stage. This study simultaneously provides important insights into the differential levels of CD4 + T cell subpopulations and IC expression in CRC TME, compared to periphery and associations with clinicopathologic features, which could be used as potential biomarkers for CRC progression and response to therapy., (Copyright © 2019 Toor, Murshed, Al-Dhaheri, Khawar, Abu Nada and Elkord.)