Publications by authors named "Salman M Toor"

39 Publications

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

Sci Rep 2021 Jul 14;11(1):14456. Epub 2021 Jul 14.

College of Health and Life Sciences, Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

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.
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http://dx.doi.org/10.1038/s41598-021-93531-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8280114PMC
July 2021

Correction to: Metabolic reprogramming of T regulatory cells in the hypoxic tumor microenvironment.

Cancer Immunol Immunother 2021 Aug;70(8):2123

Biomedical Research Center, School of Science, Engineering and Environment, University of Salford, Manchester, M5 4WT, UK.

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http://dx.doi.org/10.1007/s00262-021-02999-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8289778PMC
August 2021

Transcriptome of Tumor-Infiltrating T Cells in Colorectal Cancer Patients Uncovered a Unique Gene Signature in CD4 T Cells Associated with Poor Disease-Specific Survival.

Vaccines (Basel) 2021 Apr 1;9(4). Epub 2021 Apr 1.

Biomedical Research Center, School of Science, Engineering and Environment, University of Salford, Manchester M5 4WT, UK.

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.
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http://dx.doi.org/10.3390/vaccines9040334DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8065799PMC
April 2021

Metabolic reprogramming of T regulatory cells in the hypoxic tumor microenvironment.

Cancer Immunol Immunother 2021 Aug 3;70(8):2103-2121. Epub 2021 Feb 3.

Biomedical Research Center, School of Science, Engineering and Environment, University of Salford, Manchester, M5 4WT, UK.

Metabolic dysregulation in the hypoxic tumor microenvironment (TME) is considered as a hallmark of solid tumors, leading to changes in biosynthetic pathways favoring onset, survival and proliferation of malignant cells. Within the TME, hypoxic milieu favors metabolic reprogramming of tumor cells, which subsequently affects biological properties of tumor-infiltrating immune cells. T regulatory cells (Tregs), including both circulating and tissue-resident cells, are particularly susceptible to hypoxic metabolic signaling that can reprogram their biological and physicochemical properties. Furthermore, metabolic reprogramming modifies Tregs to utilize alternative substrates and undergo a plethora of metabolic events to meet their energy demands. Major impact of this metabolic reprogramming can result in differentiation, survival, excessive secretion of immunosuppressive cytokines and proliferation of Tregs within the TME, which in turn dampen anti-tumor immune responses. Studies on fine-tuning of Treg metabolism are challenging due to heterogenicity of tissue-resident Tregs and their dynamic functions. In this review, we highlight tumor intrinsic and extrinsic factors, which can influence Treg metabolism in the hypoxic TME. Moreover, we focus on metabolic reprogramming of Tregs that could unveil potential regulatory networks favoring tumorigenesis/progression, and provide novel insights, including inhibitors against acetyl-coA carboxylase 1 and transforming growth factor beta into targeting Treg metabolism for therapeutic benefits.
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http://dx.doi.org/10.1007/s00262-020-02842-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8289790PMC
August 2021

Tumor-Infiltrating Lymphoid Cells in Colorectal Cancer Patients with Varying Disease Stages and Microsatellite Instability-High/Stable Tumors.

Vaccines (Basel) 2021 Jan 19;9(1). Epub 2021 Jan 19.

Biomedical Research Center, School of Science, Engineering and Environment, University of Salford, Manchester M5 4WT, UK.

Immune checkpoint inhibition is an effective anti-cancer therapeutic approach but has shown limited efficacy in treating colorectal cancer (CRC) patients. Importantly, immune constituents of the tumor microenvironment (TME) can influence therapy response and cancer progression. We investigated the expression of immune checkpoints (ICs) on lymphoid populations within the CRC TME and compared with cells from normal colon tissues using samples from 50 patients with varying disease stages. We found that the levels of B cells, T cells, and NK cells were similar, IC-expressing CD4 and CD4CD8 double positive T cells were higher, while CD8 T cells and CD4CD8 double negative T cells were significantly lower in CRC tumors. Notably, patients with mismatch-repair deficiency/microsatellite instability-high tumors had higher levels of IC-expressing CD4 and CD8 T cells than patients with proficient MMR and microsatellite stable tumors. Lastly, The Cancer Genome Atlas Colon Adenocarcinoma datasets showed associations between low expression of selective genes and poorer progression-free interval. Our findings highlight differential expression of ICs on lymphoid cells in CRC tumors in the era of cancer immunotherapy, which at present is solely approved for anti-PD-1 therapy in patients with dMMR/MSI-H tumors. Further investigations into their functionality have potentials for deciphering resistance mechanisms to IC inhibition.
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http://dx.doi.org/10.3390/vaccines9010064DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7832866PMC
January 2021

Epigenetic regulation of immune checkpoints and T cell exhaustion markers in tumor-infiltrating T cells of colorectal cancer patients.

Epigenomics 2020 11 10;12(21):1871-1882. Epub 2020 Nov 10.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O.Box: 34110, Doha, Qatar.

 To elucidate the epigenetic alterations behind the upregulation of immune checkpoints and T cell exhaustion markers in colorectal cancer (CRC) patients. 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. 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 and upregulation in CRC tumors could be mediated by activating histone 3 lysine 4 trimethylation. 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.
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http://dx.doi.org/10.2217/epi-2020-0267DOI Listing
November 2020

Targeting TIM-3 in solid tumors: innovations in the preclinical and translational realm and therapeutic potential.

Expert Opin Ther Targets 2020 12 4;24(12):1251-1262. Epub 2020 Nov 4.

Biomedical Research Center, School of Science, Engineering and Environment, University of Salford , Manchester, United Kingdom.

Introduction: Immune checkpoint inhibitors (ICIs) have shown a great therapeutic efficacy in cancer patients. However, a significant proportion of cancer patients remain unresponsive or show limited response. T cell immunoglobulin and mucin-domain containing protein-3 (TIM-3) is a co-inhibitory receptor expressed on various cell types and is involved in the attenuation of immune responses. TIM-3 and its ligands are highly expressed in various solid malignancies and some studies have reported its association with worse disease outcomes. Thus, targeting TIM-3 could be a promising therapeutic approach to treat cancer patients.

Areas Covered: This review describes the role of TIM-3 and its ligands in regulating anti-tumor immunity and their contribution to cancer progression. Moreover, this review focuses on the preclinical models and translational data from important studies published in PubMed till October 2020, which demonstrate the therapeutic benefits of targeting TIM-3 signaling.

Expert Opinion: Despite the promising data obtained from targeting TIM-3 in preclinical models, precise mechanisms underlying the anti-tumor effects of TIM-3 inhibition are not fully elucidated. Therefore, mechanistic studies are required to provide better insights into the anti-tumor effects of targeting TIM-3, and clinical data are necessary to determine the safety profiles and therapeutic efficacy of TIM-3 inhibition in cancer patients.
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http://dx.doi.org/10.1080/14728222.2020.1841750DOI Listing
December 2020

Differential gene expression of tumor-infiltrating CD4 T cells in advanced versus early stage colorectal cancer and identification of a gene signature of poor prognosis.

Oncoimmunology 2020 09 30;9(1):1825178. Epub 2020 Sep 30.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

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, = 0.0001) and PFI (HR = 1.75, 95% CI 1.3-2.35, = 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.
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http://dx.doi.org/10.1080/2162402X.2020.1825178DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7553563PMC
September 2020

An evaluation of sorter induced cell stress (SICS) on peripheral blood mononuclear cells (PBMCs) after different sort conditions - Are your sorted cells getting SICS?

J Immunol Methods 2020 12 15;487:112902. Epub 2020 Oct 15.

Cancer Research Center, Qatar Biomedical Research Institute, Hamad Bin Khalifa University, Qatar Foundation, Doha, Qatar; Biomedical Research Center, School of Science, Engineering and Environment, University of Salford, Manchester, United Kingdom.

Flow cytometry and fluorescence-activated cell sorting have become invaluable tools to analyze and isolate specific cell populations in a wide range of biomedical research and clinical applications. In countless approaches worldwide, scientists are using single cell analyses to better understand the significance and variation within different cellular populations, and fluorescence-activated cell sorting has become a major technique for cell isolation in both basic and clinical research. However, majority of available cell sorters are pressurized, droplet-based systems, which apply significant environmental pressure and shear stress to cells during sorting. Recently, the flow cytometry community has become increasingly aware about the potential negative effects this could have on sorted cells and the term "sorter induced cell stress" (SICS) has been proposed. However, up to date only a limited number of studies have investigated the effects of cell sorting on cell viability and function. Therefore, solid data on the effects of sheath pressure and nozzle size on survival and function of sorted cells are surprisingly rare. With this in mind, we sorted "CD4" T-cells and "live" cells from human peripheral blood mononuclear cells (PBMCs) at different sort conditions and analyzed their quality before and after sorting in a series of assays. Here we present our findings in reference to cell viability and cell proliferation following sorting on different instruments (BD FACSAria III SORP and BD FACSJazz), utilizing different nozzle sizes (70 to 100 μm) and sheath pressure settings (20 to 70 psi). The results show no significant differences in cell viability and proliferation after the different tested sort conditions, but rather differences between individual experiments. These findings are evaluated and their potential significance in cell sorting experiments is discussed.
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http://dx.doi.org/10.1016/j.jim.2020.112902DOI Listing
December 2020

Differential gene expression of tumor-infiltrating CD33 myeloid cells in advanced- versus early-stage colorectal cancer.

Cancer Immunol Immunother 2021 Mar 30;70(3):803-815. Epub 2020 Sep 30.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O. Box: 34110, Doha, Qatar.

Colorectal cancer (CRC) has high mortality rates, especially in patients with advanced disease stages, who often do not respond to therapy. The cellular components of the tumor microenvironment are essentially responsible for dictating disease progression and response to therapy. Expansion of different myeloid cell subsets in CRC tumors has been reported previously. However, tumor-infiltrating myeloid cells have both pro- and anti-tumor roles in disease progression. In this study, we performed transcriptomic profiling of cells of myeloid lineage (CD33) from bulk CRC tumors at varying disease stages. We identified differentially expressed genes and pathways between CRC patients with advanced stage and early stages. We found that pro-angiogenic and hypoxia-related genes were upregulated, while genes related to immune and inflammatory responses were downregulated in CD33 myeloid cells from patients with advanced stages, implying that immune cell recruitment and activation could be compromised in advanced disease stages. Moreover, we identified a unique "poor prognosis CD33 gene signature" by aligning top upregulated and downregulated genes in tumor-infiltrating myeloid cells from our analyses with data from The Cancer Genome Atlas. Our results showed that this gene signature is an independent prognostic indicator for disease-specific survival in CRC patients, potentially reflecting its clinical importance.
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http://dx.doi.org/10.1007/s00262-020-02727-0DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7906947PMC
March 2021

Transcriptomic Analyses of Myeloid-Derived Suppressor Cell Subsets in the Circulation of Colorectal Cancer Patients.

Front Oncol 2020 2;10:1530. Epub 2020 Sep 2.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

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.
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http://dx.doi.org/10.3389/fonc.2020.01530DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7492613PMC
September 2020

Differential gene expression of tumor-infiltrating CD8 T cells in advanced versus early-stage colorectal cancer and identification of a gene signature of poor prognosis.

J Immunother Cancer 2020 09;8(2)

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar

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 CD3CD8 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 p<0.0001) and residual disease after primary therapy (χ 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.
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http://dx.doi.org/10.1136/jitc-2020-001294DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7511623PMC
September 2020

T-cell responses and therapies against SARS-CoV-2 infection.

Immunology 2021 01 27;162(1):30-43. Epub 2020 Oct 27.

Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O. Box: 34110, Doha, Qatar.

Coronavirus disease 2019 (COVID-19) is caused by SARS-CoV-2, a novel coronavirus strain. Some studies suggest that COVID-19 could be an immune-related disease, and failure of effective immune responses in initial stages of viral infection could contribute to systemic inflammation and tissue damage, leading to worse disease outcomes. T cells can act as a double-edge sword with both pro- and anti-roles in the progression of COVID-19. Thus, better understanding of their roles in immune responses to SARS-CoV-2 infection is crucial. T cells primarily react to the spike protein on the coronavirus to initiate antiviral immunity; however, T-cell responses can be suboptimal, impaired or excessive in severe COVID-19 patients. This review focuses on the multifaceted roles of T cells in COVID-19 pathogenesis and rationalizes their significance in eliciting appropriate antiviral immune responses in COVID-19 patients and unexposed individuals. In addition, we summarize the potential therapeutic approaches related to T cells to treat COVID-19 patients. These include adoptive T-cell therapies, vaccines activating T-cell responses, recombinant cytokines, Th1 activators and Th17 blockers, and potential utilization of immune checkpoint inhibitors alone or in combination with anti-inflammatory drugs to improve antiviral T-cell responses against SARS-CoV-2.
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http://dx.doi.org/10.1111/imm.13262DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7730020PMC
January 2021

Role of Epigenetic Modifications in Inhibitory Immune Checkpoints in Cancer Development and Progression.

Front Immunol 2020 14;11:1469. Epub 2020 Jul 14.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

A balance between co-inhibitory and co-stimulatory signals in the tumor microenvironment (TME) is critical to suppress tumor development and progression, primarily via maintaining effective immunosurveillance. Aberrant expression of immune checkpoints (ICs), including programmed cell death protein 1 (PD-1), cytotoxic T-lymphocyte-associated protein 4 (CTLA-4), T cell immunoglobulin and mucin-domain containing-3 (TIM-3), lymphocyte-activation gene 3 (LAG-3) and T cell immunoreceptor with Ig and ITIM domains (TIGIT), can create an immune-subversive environment, which helps tumor cells to evade immune destruction. Recent studies showed that epigenetic modifications play critical roles in regulating the expression of ICs and their ligands in the TME. Reports showed that the promoter regions of genes encoding ICs/IC ligands can undergo inherent epigenetic alterations, such as DNA methylation and histone modifications (acetylation and methylation). These epigenetic aberrations can significantly contribute to the transcriptomic upregulation of ICs and their ligands. Epigenetic therapeutics, including DNA methyltransferase and histone deacetylase inhibitors, can be used to revert these epigenetic anomalies acquired during the progression of disease. These discoveries have established a promising therapeutic modality utilizing the combination of epigenetic and immunotherapeutic agents to restore the physiological epigenetic profile and to re-establish potent host immunosurveillance mechanisms. In this review, we highlight the roles of epigenetic modifications on the upregulation of ICs, focusing on tumor development, and progression. We discuss therapeutic approaches of epigenetic modifiers, including clinical trials in various cancer settings and their impact on current and future anti-cancer therapies.
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http://dx.doi.org/10.3389/fimmu.2020.01469DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7371937PMC
April 2021

Transcriptomic Profiling of Circulating HLA-DR Myeloid Cells, Compared with HLA-DR Myeloid Antigen-presenting Cells.

Immunol Invest 2020 Jul 29:1-12. Epub 2020 Jul 29.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population of cells with potent immunosuppressive functions, which can inhibit the activation of immune responses under a steady-state condition and pathological conditions. We performed transcriptomic profiling of circulating CD33HLA-DR myeloid antigen-presenting cells (APCs) and CD33HLA-DR myeloid cells (potentially MDSCs) in healthy individuals. We sorted both subpopulations from peripheral blood mononuclear cells (PBMCs) of 10 healthy donors and performed RNA sequencing (RNA-Seq). We found that several signaling pathways associated with the positive regulation of immune responses, such as antigen presentation/processing, FcγR-mediated phagocytosis and immune cell trafficking, phosphoinositide 3-kinase (PI3K)/Akt signaling, DC maturation, triggering receptor expressed on myeloid cells 1 (TREM1) signaling, nuclear factor of activated T cells (NFAT) and IL-8 signaling were downregulated in CD33HLA-DR myeloid cells. In contrast, pathways implicated in tumor suppression and anti-inflammation, including peroxisome proliferator-activated receptor (PPAR) and phosphatase and tensin homolog (PTEN), were upregulated in CD33HLA-DR myeloid cells. These data indicate that PPAR/PTEN axis could be upregulated in myeloid cells to keep the immune system in check in normal physiological conditions. Our data reveal some of the molecular and functional differences between CD33HLA-DR APCs and CD33HLA-DR myeloid cells in a steady-state condition, reflecting the potential suppressive function of CD33HLA-DR myeloid cells to maintain immune tolerance. For future studies, the same methodological approach could be applied to perform transcriptomic profiling of myeloid subsets in pathological conditions.
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http://dx.doi.org/10.1080/08820139.2020.1795875DOI Listing
July 2020

Editorial of Harnessing the Power of T Cells: The Promising Hope for a Universal Influenza Vaccine.

Vaccines (Basel) 2020 Jul 11;8(3). Epub 2020 Jul 11.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha 34110, Qatar.

The global burden of influenza-associated respiratory mortality is higher than previous estimates, with over 0 [...].
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http://dx.doi.org/10.3390/vaccines8030376DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7565606PMC
July 2020

Myeloid Cells in Circulation and Tumor Microenvironment of Colorectal Cancer Patients with Early and Advanced Disease Stages.

J Immunol Res 2020 12;2020:9678168. Epub 2020 Jun 12.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

Myeloid-derived suppressor cells (MDSCs) are a heterogenous population of cells that have been implicated in the development of an immunosuppressive environment, which promotes tumorigenesis and tumor progression. Numerous studies have reported expansion of MDSCs in circulation and the tumor microenvironment (TME) of cancer patients. However, due to the heterogenic nature of MDSCs and the different approaches for their identification, their detailed characterization and impact on disease progression in cancer patients are warranted. In this study, we investigated the levels of different myeloid cell subsets and antigen-presenting cells (APCs) using flow cytometry in unfractionated whole blood (WB), peripheral blood mononuclear cells (PBMCs), tumor tissue (TT), and adjacent normal tissue (NT) of colorectal cancer (CRC) patients. We found high levels of granulocytic myeloid cells (GMCs) in whole blood, but their levels were significantly lower in PBMCs. Importantly, we found significantly higher levels of GMCs in the TME compared to NT. In addition, monocytic myeloid cells (MMCs) showed significantly higher levels in PBMCs of CRC patients, compared to healthy donors (HDs). Notably, patients with advanced disease stages showed significantly higher levels of GMCs compared to early stages in whole blood, but PBMCs and tumor-infiltrating myeloid cells did not show any significant differences. Lastly, we found that levels of GMCs decreased, while IMCs increased in the TME with tumor budding. Our results highlight the importance of investigating the levels of different myeloid cell subsets in PBMCs versus whole blood of cancer patients and improve current knowledge on the potential prognostic significance of myeloid cells in CRC patients.
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http://dx.doi.org/10.1155/2020/9678168DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7306094PMC
May 2021

Blockade of PD-1, PD-L1, and TIM-3 Altered Distinct Immune- and Cancer-Related Signaling Pathways in the Transcriptome of Human Breast Cancer Explants.

Genes (Basel) 2020 06 25;11(6). Epub 2020 Jun 25.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha 34110, Qatar.

Immune checkpoint inhibitors (ICIs) are yet to have a major advantage over conventional therapies, as only a fraction of patients benefit from the currently approved ICIs and their response rates remain low. We investigated the effects of different ICIs-anti-programmed cell death protein 1 (PD-1), anti-programmed death ligand-1 (PD-L1), and anti-T cell immunoglobulin and mucin-domain containing-3 (TIM-3)-on human primary breast cancer explant cultures using RNA-Seq. Transcriptomic data revealed that PD-1, PD-L1, and TIM-3 blockade follow unique mechanisms by upregulating or downregulating distinct pathways, but they collectively enhance immune responses and suppress cancer-related pathways to exert anti-tumorigenic effects. We also found that these ICIs upregulated the expression of other IC genes, suggesting that blocking one IC can upregulate alternative ICs, potentially giving rise to compensatory mechanisms by which tumor cells evade anti-tumor immunity. Overall, the transcriptomic data revealed some unique mechanisms of the action of monoclonal antibodies (mAbs) targeting PD-1, PD-L1, and TIM-3 in human breast cancer explants. However, further investigations and functional studies are warranted to validate these findings.
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http://dx.doi.org/10.3390/genes11060703DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7349021PMC
June 2020

DNA methylation in the promoters of PD-L1, MMP9, ARG1, galectin-9, TIM-3, VISTA and TGF-β genes in HLA-DR myeloid cells, compared with HLA-DR antigen-presenting cells.

Epigenetics 2020 12 18;15(12):1275-1288. Epub 2020 May 18.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF) , Doha, Qatar.

Myeloid cells, including antigen-presenting cells (APCs) and myeloid-derived suppressor cells (MDSCs) play opposing roles to orchestrate innate and adaptive immune responses during physiological and pathological conditions. We investigated the role of DNA methylation in regulating the transcription of inhibitory/suppressive molecules in myeloid suppressive cells (identified as CD33HLA-DR) in comparison to APCs. We selected a number of immune checkpoints (ICs), IC ligands, and immunosuppressive molecules that have been implicated in MDSC function, including PD-L1, TIM-3, VISTA, galectin-9, TGF-β, ARG1 and MMP9. We examined their mRNA expression levels, and investigated whether DNA methylation regulates their transcription in sorted myeloid cell subpopulations. We found that mRNA levels of PD-L1, TIM-3, TGF-β, ARG1 and MMP9 in CD33HLA-DR cells were higher than APCs. However, VISTA and galectin-9 mRNA levels were relatively similar in both myeloid subpopulations. CpG islands in the promoter regions of TGF-β1, TIM-3 and ARG1 were highly unmethylated in CD33HLA-DRcells, compared with APCs, suggesting that DNA methylation is one of the key mechanisms, which regulate their expression. However, we did not find differences in the methylation status of PD-L1 and MMP9 between CD33HLA-DR and APCs, suggesting that their transcription could be regulated via other genetic and epigenetic mechanisms. The promoter methylation status of VISTA was relatively similar in both myeloid subpopulations. This study provides novel insights into the epigenetic mechanisms, which control the expression of inhibitory/suppressive molecules in circulating CD33HLA-DR cells in a steady-state condition, possibly to maintain immune tolerance and haemostasis.
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http://dx.doi.org/10.1080/15592294.2020.1767373DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7678924PMC
December 2020

Expression of immune checkpoints and T cell exhaustion markers in early and advanced stages of colorectal cancer.

Cancer Immunol Immunother 2020 Oct 11;69(10):1989-1999. Epub 2020 May 11.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O. Box: 34110, Doha, Qatar.

Despite recent advances in colorectal cancer (CRC) treatment, a large proportion of patients show limited responses to therapies, especially in advanced stages. There is an urgent need to identify prognostic biomarkers and/or therapeutic targets in advanced stages, aiming to improve the efficacy of current treatments. We aimed to determine prognostic biomarkers in tumor tissue and circulation of CRC patients, with a special focus on T cell exhaustion markers. We found that mRNA levels of PD-1, TIM-3, CTLA-4, TIGIT, CD160, CD244, KLRG1, TOX2, TOX3, Ki-67, and PRDM1 were elevated in CRC tumor tissues. We also investigated differences in gene expression between early and advanced disease stages. We found that TOX and potentially TIM-3, CTLA-4, VISTA, TIGIT, KLRG1, TOX2, SIRT1, Ki-67, and Helios mRNA levels in tumor tissue were elevated in advanced disease stages, suggesting their potential roles in CRC progression. In contrast, PD-1 and CD160 levels in tumor tissue were downregulated in advanced stages. In the circulation of CRC patients, mRNA levels of PD-1, VISTA and LAG-3 were higher than those of healthy individuals. Moreover, in circulation, PD-1, CTLA-4 and TIGIT mRNA levels were reduced in advanced stages. Interestingly, levels of PD-1 in both tumor tissue and circulation were reduced in advanced stages, suggesting that targeting PD-1 in patients with advanced stages could be less effective. Altogether, these findings suggest some potential T cell exhaustion markers that could be utilized as prognostic biomarkers and/or therapeutic targets for CRC. However, further investigations and validations in larger cohorts are required to confirm these findings.
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http://dx.doi.org/10.1007/s00262-020-02593-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7511277PMC
October 2020

Differential expression of TIM-3 in circulation and tumor microenvironment of colorectal cancer patients.

Clin Immunol 2020 06 19;215:108429. Epub 2020 Apr 19.

College of Health & Life Sciences, Hamad Bin Khalifa University, Qatar Foundation, Doha, Qatar; Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), PO Box 34110, Doha, Qatar. Electronic address:

T-cell immunoglobulin and mucin-domain containing-3 (TIM-3) is an inhibitory immune checkpoint, which suppresses anti-tumor immune responses. TIM-3 expression on different immune cells in periphery and tumor microenvironment (TME) of colorectal cancer (CRC) patients has not been fully investigated. We found that TIM-3 was mainly expressed on monocytic myeloid cells (MMCs) and antigen-presenting cells (APCs) in circulation but was mainly expressed on T cells and APCs in the TME. Additionally, TIM-3 T cells co-expressed higher levels of PD-1 than TIM-3 T cells in normal tissue. In contrast, TIM-3 T cells in the TME showed significantly higher PD-1 expression. Interestingly, there was a trend towards increased levels of TIM-3 APCs with disease stages; however, levels of TIM-3 T cells were decreased with disease stages in the TME. This study shows the differential expression of TIM-3 on different immune cells in circulation and TME of CRC patients, and their associations with disease stages.
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http://dx.doi.org/10.1016/j.clim.2020.108429DOI Listing
June 2020

Transcriptomic Profiling of Tumor-Infiltrating CD4TIM-3 T Cells Reveals Their Suppressive, Exhausted, and Metastatic Characteristics in Colorectal Cancer Patients.

Vaccines (Basel) 2020 02 6;8(1). Epub 2020 Feb 6.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O. Box 34110 Doha, Qatar.

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 CD3CD4 (CD8) TILs. CD4TIM-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 CD4TIM-3 TILs, which could favor proliferation/invasion and induce inflammatory and T-cell exhaustion pathways. In addition, we found that CD4TIM-3 TILs potentially support tumor invasion and metastasis, compared with conventional CD4CD25 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.
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http://dx.doi.org/10.3390/vaccines8010071DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7157206PMC
February 2020

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

Clin Epigenetics 2020 01 15;12(1):13. Epub 2020 Jan 15.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O. Box 34110, Doha, Qatar.

Background: 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).

Methods: 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.

Results: 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.

Conclusions: 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.
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http://dx.doi.org/10.1186/s13148-020-0808-9DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6964037PMC
January 2020

Identification of two HLA-A*0201 immunogenic epitopes of lactate dehydrogenase C (LDHC): potential novel targets for cancer immunotherapy.

Cancer Immunol Immunother 2020 Mar 13;69(3):449-463. Epub 2020 Jan 13.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

Lactate dehydrogenase C (LDHC) is an archetypical cancer testis antigen with limited expression in adult tissues and re-expression in tumors. This restricted expression pattern together with the important role of LDHC in cancer metabolism renders LDHC a potential target for immunotherapy. This study is the first to investigate the immunogenicity of LDHC using T cells from healthy individuals. LDHC-specific T cell responses were induced by in vitro stimulation with synthetic peptides, or by priming with autologous peptide-pulsed dendritic cells. We evaluated T cell activation by IFN-γ ELISpot and determined cytolytic activity of HLA-A*0201-restricted T cells in breast cancer cell co-cultures. In vitro T cell stimulation induced IFN-γ secretion in response to numerous LDHC-derived peptides. Analysis of HLA-A*0201 responses revealed a significant T cell activation after stimulation with peptide pools 2 (PP2) and 8 (PP8). The PP2- and PP8-specific T cells displayed cytolytic activity against breast cancer cells with endogenous LDHC expression within a HLA-A*0201 context. We identified peptides LDHC and LDHC from PP2 and PP8 to elicit a functional cellular immune response. More specifically, we found an increase in IFN-γ secretion by CD8 + T cells and cancer-cell-killing of HLA-A*0201/LDHC positive breast cancer cells by LDHC- and LDHC-induced T cells, albeit with a possible antigen recognition threshold. The majority of induced T cells displayed an effector memory phenotype. To conclude, our findings support the rationale to assess LDHC as a targetable cancer testis antigen for immunotherapy, and in particular the HLA-A*0201 restricted LDHC and LDHC peptides within LDHC.
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http://dx.doi.org/10.1007/s00262-020-02480-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7044258PMC
March 2020

Immune Checkpoints in Circulating and Tumor-Infiltrating CD4 T Cell Subsets in Colorectal Cancer Patients.

Front Immunol 2019 17;10:2936. Epub 2019 Dec 17.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha, Qatar.

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 CD4FoxP3Helios 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 CD4FoxP3Helios T cell subsets. Interestingly, we found that CTLA-4, TIM-3, and LAG-3 were mainly co-expressed on FoxP3Helios Tregs in the TME. Additionally, FoxP3 Tregs expressed higher levels of Helios, CTLA-4 and TIM-3 than FoxP3 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 CD4CTLA-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.
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http://dx.doi.org/10.3389/fimmu.2019.02936DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6928042PMC
November 2020

Transcriptomic Analyses Revealed Systemic Alterations in Gene Expression in Circulation and Tumor Microenvironment of Colorectal Cancer Patients.

Cancers (Basel) 2019 Dec 11;11(12). Epub 2019 Dec 11.

College of Health & Life Sciences, Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), PO Box 34110, Doha, Qatar.

Colorectal cancer (CRC) is among the leading causes of cancer-related deaths worldwide, underscoring a need for better understanding of the disease and development of novel diagnostic biomarkers and therapeutic interventions. Herein, we performed transcriptome analyses on peripheral blood mononuclear cells (PBMCs), CRC tumor tissue and adjacent normal tissue from 10 CRC patients and PBMCs from 15 healthy controls. Up regulated transcripts from CRC PBMCs were associated with functions related to immune cell trafficking and cellular movement, while downregulated transcripts were enriched in cellular processes related to cell death. Most affected signaling networks were those involved in tumor necrosis factor (TNF) and interleukin signaling. The expression of selected immune-related genes from the RNA-Seq data were further validated using qRT-PCR. Transcriptome analysis of CRC tumors and ingenuity pathway analysis revealed enrichment in several functional categories related to cellular movement, cell growth and proliferation, DNA replication, recombination and repair, while functional categories related to cell death were suppressed. Upstream regulator analysis revealed activation of ERBB2 and FOXM1 networks. Interestingly, there were 18 common upregulated and 36 common downregulated genes when comparing PBMCs and tumor tissue, suggesting transcriptomic changes in the tumor microenvironment could be reflected, in part, in the periphery with potential utilization as disease biomarkers.
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http://dx.doi.org/10.3390/cancers11121994DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6966620PMC
December 2019

Pembrolizumab Interferes with the Differentiation of Human FOXP3-Induced T Regulatory Cells, but Not with FOXP3 Stability, through Activation of mTOR.

J Immunol 2020 01 4;204(1):199-211. Epub 2019 Dec 4.

Cancer Research Center, Qatar Biomedical Research Institute, Hamad Bin Khalifa University, Qatar Foundation, Doha, Qatar;

Programmed cell death 1 (PD-1) is critical for T regulatory cells (Tregs) to maintain peripheral tolerance to self-antigens. In the tumor microenvironment, interaction between PD-1 and its ligands supports tumor immune evasion. Pembrolizumab blocks interactions of PD-1 with its ligands, enhancing antitumor and clinical responses. We and others have reported that pembrolizumab does not affect function or phenotype of thymic-derived Tregs; however, little is known about its effect on extrathymic differentiation of peripheral Tregs. In this study, we investigated the effect of pembrolizumab on in vitro-induced Tregs (iTregs). Our work showed that PD-1 blockade interferes with iTreg differentiation and has no potential effect on the stability of FOXP3 after differentiation. Additionally, we found that both nontreated and pembrolizumab-treated iTregs were suppressive. However, pembrolizumab-treated iTregs were relatively less suppressive in higher Treg ratios and failed to produce IL-10 compared with their nontreated counterparts. Different methods including transcriptomic analyses confirmed that the downregulation of FOXP3 was mediated by activating mTOR and STAT1 and inhibiting MAPK pathways, shifting the iTreg polarization in favor of Th1 and Th17 subsets. To confirm the role of mTOR activation, we found that rapamycin diminished the effect of pembrolizumab-mediated downregulation of FOXP3. Ingenuity pathway analysis revealed that pembrolizumab-treated iTregs showed upregulation of genes promoting DNA repair and immune cell trafficking, in addition to downregulation of genes supporting cellular assembly and organization. To our knowledge, this is the first study to show that pembrolizumab interferes with differentiation of human FOXP3 iTregs and to disclose some of the molecular pathways involved.
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http://dx.doi.org/10.4049/jimmunol.1900575DOI Listing
January 2020

Breast Cancer Cells and PD-1/PD-L1 Blockade Upregulate the Expression of PD-1, CTLA-4, TIM-3 and LAG-3 Immune Checkpoints in CD4 T Cells.

Vaccines (Basel) 2019 Oct 12;7(4). Epub 2019 Oct 12.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), Doha 34110, Qatar.

Triple negative breast cancer (TNBC) is the most aggressive breast cancer subtype, and it exhibits resistance to common breast cancer therapies. Immune checkpoint inhibitors (ICIs) targeting programmed cell death 1 (PD-1) and its ligand, PD-L1, have been approved to treat various cancers. However, the therapeutic efficacy of targeting PD-1/PD-L1 axis in breast cancer is under clinical investigation. In addition, the mechanisms of action of drugs targeting PD-1 and PD-L1 have not been fully elucidated. In this study, we investigated the effect of human TNBC cell lines, MDA-MB-231 and MDA-MB-468, and the non-TNBC cell line, MCF-7, on the expression of immune checkpoints (ICs) on CD4 T cell subsets, including regulatory T cells (Tregs), using a co-culture system. We also examined the effect of blocking PD-1 or PD-L1 separately and in combination on IC expression by CD4 T cell subsets. We found that breast cancer cells upregulate the expression of ICs including PD-1, cytotoxic T lymphocyte-associated antigen-4 (CTLA-4), T cell immunoglobulin and mucin domain-containing protein 3 (TIM-3) and lymphocyte activation gene-3 (LAG-3) in CD4 T cell subsets. We also found that the co-blockade of PD-1 and PD-L1 further upregulates the co-expression of TIM-3 and LAG-3 on CD4CD25 T cells and CD4CD25FoxP3Helios Tregs in the presence of TNBC cells, but not in non-TNBC cells. Our results indicate the emergence of compensatory inhibitory mechanisms, most likely mediated by Tregs and activated non-Tregs, which could lead to the development of TNBC resistance against PD-1/PD-L1 blockade.
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http://dx.doi.org/10.3390/vaccines7040149DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6963740PMC
October 2019

Investigation of the Effect of PD-L1 Blockade on Triple Negative Breast Cancer Cells Using Fourier Transform Infrared Spectroscopy.

Vaccines (Basel) 2019 Sep 9;7(3). Epub 2019 Sep 9.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), P.O. Box 34110 Doha, Qatar.

Interactions between programmed death-1 (PD-1) with its ligand PD-L1 on tumor cells can antagonize T cell responses. Inhibiting these interactions using immune checkpoint inhibitors has shown promise in cancer immunotherapy. MDA-MB-231 is a triple negative breast cancer cell line that expresses PD-L1. In this study, we investigated the biochemical changes in MDA-MB-231 cells following treatment with atezolizumab, a specific PD-L1 blocker. Our readouts were Fourier Transform Infrared (FTIR) spectroscopy and flow cytometric analyses. Chemometrical analysis, such as principal component analysis (PCA), was applied to delineate the spectral differences. We were able to identify the chemical alterations in both protein and lipid structure of the treated cells. We found that there was a shift from random coil and α-helical structure to β-sheet conformation of PD-L1 on tumor cells due to atezolizumab treatment, which could hinder binding with its receptors on immune cells, ensuring sustained T cell activation for potent immune responses. This work provides novel information about the effects of atezolizumab at molecular and cellular levels. FTIR bio-spectroscopy, in combination with chemometric analyses, may expedite research and offer new approaches for cancer immunology.
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http://dx.doi.org/10.3390/vaccines7030109DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6789440PMC
September 2019

Immune checkpoints in the tumor microenvironment.

Semin Cancer Biol 2020 10 29;65:1-12. Epub 2019 Jun 29.

Cancer Research Center, Qatar Biomedical Research Institute (QBRI), Hamad Bin Khalifa University (HBKU), Qatar Foundation (QF), PO Box 34110, Doha, Qatar; Institute of Cancer Sciences, University of Manchester, Manchester, United Kingdom. Electronic address:

Interactions between immune checkpoints (ICs) and their ligands negatively regulate T cell activation pathways involved in physiological immune responses against specific antigens. ICs and their ligands are frequently upregulated in the tumor microenvironment (TME) of various malignancies, and they represent significant barriers for induction of effective anti-tumor immune responses. Several IC inhibitors (ICIs) have been developed, with some currently in clinical trials and others have been approved for the treatment of different cancers. However, tumor cells are able to counteract the activity of ICIs and can commission additional inhibitory pathways via expression of other ICs/ligands within the TME. This review discusses the expression of various ICs/ligands in the TME and their impact on tumor immune evasion. Additionally, we discuss various regulatory mechanisms, including genetic and epigenetic, and other modulatory factors including hypoxia and the presence of immunosuppressive populations in the TME, which result in upregulation of ICs in various cancers. Moreover, we discuss the prognostic significance of ICs and their ligands, and the potential strategies to enhance treatment responses to ICIs. This review aims to advance our current knowledge on the role of ICs in the TME and the clinical benefits of targeting them.
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http://dx.doi.org/10.1016/j.semcancer.2019.06.021DOI Listing
October 2020
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