scholarly journals Characterization of Circulating Mucosal Associated Invariant T cells in Colorectal Cancer Patients

2020 ◽  
Author(s):  
Mohamed M Abu Hassan ◽  
Salah Elsafi ◽  
Suliman Y Al Omar ◽  
Hafez Halawaani ◽  
Lamjed Mansour ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Inverse Correlation ◽  
Stage Ii ◽  
Lower Percentage ◽  
Peripheral Blood Mononuclear ◽  
Mait Cells ◽  
Control Subjects ◽  
And Control

Abstract Background: Although MAIT cells regulate the pathogenesis of various inflammatory diseases, their roles in the development of colorectal cancer are still unclear. The objectives of this study is to investigate the level of circulating MAIT cells and the level of expression of the membranous KIRs receptors among CRC patients and control subjects. Peripheral blood mononuclear cells (PBMCs) were isolate and of MAIT cells were phenotypically identified by flow cytometry using various monoclonal antibodies. The presence of HLA-C1 and HLA-C2 groups were typed by PCR. Results: The percentage of MAIT cells were higher in CRC patients compared to control subjects. The percentage of MAIT cells was higher in patients with CRC in stage III and IV, but lower in stage II compared with control subjects. High frequency of HLA - C2 among patients with CRC (87%) compared with control subjects (74.4%). The protein expression of MAIT cells associated phenotyping antigens and some KIR receptors such as CD45RA, CD45RO, CD62L, CD11a, CD158a, CD158b, CD158e and CD158f were reported. A relatively lower percentage of CD45RA expression was seen in CRC patients compared with control subjects. There was a significant reduction in CD45RO, CD62L, CD158a, CD158e and CD158f expression in CRC patients. Stratification analysis of 46 CRC patients indicated that the percentages of circulating MAIT cells were lower in stage II and higher in stages III, and IV. The frequencies of circulating MAIT cells had not been reduced in patients with colorectal cancer.Conclusions: The number and level of circulating MAIT cells were evaluated in peripheral blood of CRC patients and control subjects. We compared the frequency of CD158a+MAIT cells in all stages of CRC and control subjects in the presence of its ligand HLA-C2 and found a significantly reduction in all stages of CRC. We stratified CRC patients and found that CD158b+ MAIT cells were decreased as the diseases progress to advancing cancer stages. When comparing the proportion of CD158a and b+ MAIT cells in all stages with control subjects, the results showed inverse correlation with cancer stages in CRC patients and the median gradually decrease in each stage.

scholarly journals Characterization of Circulating Mucosal Associated Invariant T cells in Colorectal Cancer Patients

2020 ◽  
Author(s):  
Mohamed M Abu Hassan ◽  
Salah Elsafi ◽  
Suliman Y Al Omar ◽  
Hafez Halawaani ◽  
Lamjed Mansour ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Inflammatory Diseases ◽  
Inverse Correlation ◽  
Peripheral Blood Mononuclear ◽  
Mait Cells ◽  
Control Subjects ◽  
Colorectal Cancer Patients ◽  
And Control

Abstract Background: Although MAIT cells regulate the pathogenesis of various inflammatory diseases, their roles in the development of colorectal cancer are still unclear. The objectives of this study is to investigate the level of circulating MAIT cells and the level of expression of the membranous KIRs receptors among CRC patients and control subjects. Peripheral blood mononuclear cells (PBMCs) were isolate and of MAIT cells were phenotypically identified by flow cytometry using various monoclonal antibodies. The presence of HLA-C1 and HLA-C2 groups were typed by PCR. Results: The result showed that the percentage of MAIT cells was slightly higher in the control subjects than in in all stages of CRC patients. Stratification analysis of 46 CRC patients indicated that the percentages of circulating MAIT cells were lower in stage II and higher in stages III, and IV. A higher frequency of HLA - C2 among patients with CRC (87%) compared with control subjects (74.4%) was detected. The protein expression of MAIT cells associated phenotyping antigens and some KIR receptors such as CD45RA, CD45RO, CD62L, CD11a, CD158a, CD158b, CD158e and CD158f were reported. There was a reduction in CD45RO, CD158a, CD158e and CD158f expression in CRC patients. The frequencies of circulating MAIT cells had not been reduced in patients with colorectal cancer.Conclusions: The number and level of circulating MAIT cells were evaluated in peripheral blood of CRC patients and control subjects. We compared the frequency of CD158a+MAIT cells in all stages of CRC and control subjects in the presence of its ligand HLA-C2 and found a reduction in all stages of CRC. We stratified CRC patients and found that CD158b+ MAIT cells were decreased as the diseases progress to advancing cancer stages. When comparing the proportion of CD158a and b+ MAIT cells in all stages with control subjects, the results showed inverse correlation with cancer stages in CRC patients and the median gradually decrease in each stage.

scholarly journals IMMU-18. IMMUNOGENOMIC RESPONDER PHENOTYPE FROM A PHASE I TRIAL OF ANTI-LAG3 OR ANTI-CD137 ALONE AND IN COMBINATION WITH ANTI-PD-1 IN PATIENTS WITH RECURRENT GBM

2019 ◽  
Vol 21 (Supplement_6) ◽  
pp. vi122-vi123
Author(s):  
Christina Jackson ◽  
John Choi ◽  
JiaJia Zhang ◽  
Anna Piotrowski ◽  
Tobias Walbert ◽  
...  
Keyword(s):  
T Cells ◽  
T Cell ◽  
Cd8 T Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Checkpoint Inhibitors ◽  
Lower Percentage ◽  
Radiographic Evaluation ◽  
Peripheral Blood Mononuclear ◽  
Initial Resection

Abstract BACKGROUND Immune checkpoint inhibitors (ICIs) are not uniformly effective in glioblastoma treatment. Immunogenomic determinants may identify patients who are most likely to benefit from these therapies. Therefore, we compared the immunogenomic phenotype of a responder to combination anti-LAG-3 and anti-PD-1 therapy to non-responders. METHODS We performed T cell receptor (TCR) sequencing and gene expression analysis on pre-treatment, post-chemoradiation, and post-immunotherapy tumor specimens of glioblastoma patients treated with anti-LAG3 in combination with anti-PD-1 after first recurrence (NCT02658981, ongoing). We evaluated T cell clonotypes and immunophenotype of serially collected peripheral blood mononuclear cells (PBMCs) during treatment using multi-parametric flow cytometry. RESULTS To date, six patients have been enrolled in the initial anti-LAG-3 and anti-PD-1 cohort. One patient demonstrated complete response, one had stable disease, and four had progressive disease by radiographic evaluation. The responder demonstrated substantially higher TCR clonality in the resected tumor at initial diagnosis compared to non-responders (mean 0.028 vs. 0.005). Shared tumor infiltrating clonotypes with pre-immunotherapy PBMCs exhibited an increase in frequency from initial resection (6.8%) to resection at recurrence (20%). The responder’s tumor at initial resection exhibited increased gene signatures of PD1low CD8+ T cells, chemokine signaling, and interferon gamma pathways. On PBMC phenotypic analysis, the responder demonstrated significantly higher percentages of CD137+ CD8+T cells (median 8.38% vs 3.24%, p=0.02) and lower percentages of Foxp3+CD137+ CD4+T cells compared to non-responders (median 18.5% vs. 38.5%, p=0.006). Interestingly, dynamic analysis of PBMCs showed that the responder demonstrated a lower percentage of PD1+ CD8+ T cells pre-immunotherapy (median 2.5% vs.12.4%, p=0.002), with persistent decrease over the course of treatment while non-responders showed no consistent pattern. CONCLUSION Our preliminary results demonstrate significant differences in tumor and peripheral blood immunogenomic characteristics between responder and non-responders to anti-LAG3 and anti-PD-1 therapy. These immunogenomic characteristics may help stratify patients’ response to combination ICIs.

scholarly journals Immune Characterization Of Metastatic Colorectal Cancer Patients Post Reovirus Administration

2020 ◽  
Author(s):  
Ruwan Parakrama ◽  
Elisha Fogel ◽  
Carol Chandy ◽  
Titto Augustine ◽  
Matt Coffey ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Flow Cytometry ◽  
Cancer Patients ◽  
Tumor Cells ◽  
Peripheral Blood Mononuclear Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Sequencing Analysis ◽  
Peripheral Blood Mononuclear ◽  
Blood Mononuclear Cells

Abstract Background KRAS mutations are prevalent in 40-45% of patients with colorectal cancer (CRC) and targeting this gene has remained elusive. Viruses are well known immune sensitizing agents. The therapeutic efficacy of oncolytic reovirus in combination with chemotherapy is underway in a phase 1 study of metastatic CRC. This study evaluates the nature of immune response by determining the cytokine expression pattern in peripheral circulation along with the distribution of antigen presenting cells (APCs) and activated T lymphocytes. Further the study evaluates the alterations in exosomal and cellular microRNA levels along with the effect of reovirus on leukocyte transcriptome.Methods Reovirus was administered as a 60-minute intravenous infusion for 5 consecutive days every 28 days, at a tissue culture infective dose (TCID50) of 3x1010. Peripheral blood mononuclear cells (PBMC) were isolated from whole blood prior to reovirus administration and post-reovirus on days 2, 8, and 15. The expression profile of 25 cytokines in plasma was assessed (post PBMC isolation) on an EMD Millipore multiplex Luminex platform. Exosome and cellular levels of miR-29a-3p was determined in pre and post reovirus treated samples. Peripheral blood mononuclear cells were stained with fluorophore labelled antibodies against CD4, CD8, CD56, CD70, and CD123, fixed and evaluated by flow cytometry. The expression of granzyme B was determined on core biopsy of one patient. Finally, Clariom D Assay, was used to determine the expression of 847 immune-related genes when compared to pre reovirus treatment by RNA sequencing analysis. A change was considered if the expression level either doubled or halved and the significance was determined at a p value of 0.001. Results Cytokine assay indicated upregulation at day 8 for IL-12p40 (2.95; p=0.05); day 15 for GM-CSF (3.56; p=0.009), IFN-Ƴ (1.86; p=0.0004) and IL-12p70 (2.42; p=0.02). An overall reduction in IL-8, VEGF and RANTES/CCL5 was observed over the 15-day period. Statistically significant reductions were observed at Day 15 for IL-8 (0.457-fold, 53.3% reduction; p=0.03) and RANTES/CC5 (0.524-fold, 47.6% reduction; p=0.003). An overall increase in IL-6 was observed, with statistical significance at day 8 (1.98-fold; 98% increase, p=0.00007). APCs were stimulated within 48 hours and activated (CD8+ CD70+) T cells within 168 hours as determine by flow cytometry. Sustained reductions in exosomal and cellular levels of miR-29a-3p (a microRNA upregulated in CRC and associated with decreased expression of the tumor suppressor WWOX gene) was documented. Reovirus administration further resulted in increases in KRAS (33x) , IFNAR1 (20x), STAT3(5x), and TAP1 (4x) genes after 2 days; FGCR2A (23x) and CD244 (3x) after 8 days; KLRD1 (14x), TAP1 (2x) and CD244(2x) after 15 days. Reductions (>0.5x) were observed in VEGFA (2x) after 2 days; CXCR2 (2x), ITGAM (3x) after 15 days.Conclusions Reovirus has profound immunomodulatory properties that span the genomic, protein and immune cell distribution levels. This is the first study with reovirus in cancer patients that demonstrates these multi-layered effects, demonstrating how reovirus can function as an immune stimulant (augmenting the efficacy of immuno-chemo-therapeutic drugs), and an oncolytic agent. Reovirus thus functions bimodally as an oncolytic agent causing lysis of tumor cells, and facilitator of immune-mediated recognition and destruction of tumor cells.

scholarly journals MicroRNA signatures in peripheral blood mononuclear cells of chronic heart failure patients

2010 ◽  
Vol 42 (3) ◽  
pp. 420-426 ◽  
Author(s):  
Christine Voellenkle ◽  
Jeroen van Rooij ◽  
Claudia Cappuzzello ◽  
Simona Greco ◽  
Diego Arcelli ◽  
...  
Keyword(s):  
Gene Expression ◽  
Heart Failure ◽  
Chronic Heart Failure ◽  
Peripheral Blood Mononuclear Cells ◽  
Mirna Expression ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Peripheral Blood Mononuclear ◽  
Control Subjects ◽  
Blood Mononuclear Cells

MicroRNAs (miRNAs) are noncoding RNAs that act as negative regulators of gene expression. Interestingly, specific alterations of miRNA expression have been found in failing hearts of different etiologies. The aim of this study was to identify the miRNA expression pattern of peripheral blood mononuclear cells (PBMCs) derived from chronic heart failure (CHF) patients affected by ischemic (ICM) and nonischemic dilated (NIDCM) cardiomyopathy. The expression profile of 257 miRNAs was assessed in 7 NIDCM patients, 8 ICM patients, and 9 control subjects by quantitative real-time PCR. Significantly modulated miRNAs were validated by using an independent set of 34 CHF patients (NIDCM = 19, ICM = 15) and 19 control subjects. Three miRNAs (miR-107, -139, and -142-5p) were downmodulated in both NIDCM and ICM patients versus control subjects. Other miRNAs were deregulated in only one of the CHF classes analyzed compared with control subjects: miR-142-3p and -29b were increased in NIDCM patients, while miR-125b and -497 were decreased in ICM patients. Bioinformatic analysis of miRNA predicted targets and of gene expression modifications associated with CHF in PBMCs indicated a significant impact of the miRNA signature on the transcriptome. Furthermore, miRNAs of both the NIDCM and the ICM signature shared predicted targets among CHF-modulated genes, suggesting potential additive or synergistic effects. The present study identified miRNAs specifically modulated in the PBMCs of NIDCM and ICM patients. Intriguingly, most of these miRNAs were previously reported as deregulated in human and/or mouse failing hearts. The identified miRNAs might have a potential diagnostic and/or prognostic use in CHF.

Ectopic Expression of Guanylyl Cyclase C in CD34+Progenitor Cells in Peripheral Blood

2001 ◽  
Vol 19 (19) ◽  
pp. 3951-3959 ◽  
Author(s):  
Tracy A. Fava ◽  
Rodwige Desnoyers ◽  
Stephanie Schulz ◽  
Jason Park ◽  
David Weinberg ◽  
...  
Keyword(s):  
Colorectal Cancer ◽  
Epithelial Cell ◽  
Healthy Volunteers ◽  
Progenitor Cells ◽  
Peripheral Blood ◽  
Mononuclear Cells ◽  
Rt Pcr ◽  
Peripheral Blood Mononuclear ◽  
Total Rna ◽  
Blood Mononuclear Cells

PURPOSE: To examine the utility of guanylyl cyclase C (GC-C)–specific nested reverse transcriptase polymerase chain reaction (RT-PCR) to detect circulating tumor cells in patients with colorectal cancer.PATIENTS AND METHODS: Peripheral-blood mononuclear cells from 24 patients with Dukes’ stage D colorectal cancer were analyzed by GC-C-specific nested RT-PCR using 1 μg of total RNA. Peripheral-blood mononuclear cells from 20 healthy volunteers served as controls. Additionally, peripheral-blood CD34+progenitor cells were assayed for the expression of both GC-C and other epithelial cell–specific markers.RESULTS: GC-C mRNA was detected in blood mononuclear cells from all 24 patients with colorectal cancer and all healthy volunteers. These unexpected positive results reflected low-level ectopic transcription of GC-C in CD34+progenitor cells. Moreover, CD34+progenitor cells expressed other epithelial cell–specific markers, including prostate-specific antigen, prostate-specific membrane antigen, carcinoembryonic antigen, CK-19, CK-20, mucin 1, and GA733.2. Limiting the quantity of mononuclear cell total RNA analyzed to ≤ 0.8 μg eliminated detection of GC-C and other tissue-specific transcripts in blood of healthy volunteers. However, under the same conditions, GC-C mRNA was detected in mononuclear cells from all 24 patients with metastatic colorectal cancer. Using 0.5 μg of total RNA and GC-C–specific primers, nested RT-PCR detected a single human colon carcinoma cell (approximately 20 to 200 GC-C transcripts/cell) in 106to 107mononuclear blood cells.CONCLUSION: These data suggest that GC-C may be useful for detecting circulating colorectal cancer cells. They also demonstrate that CD34+cells are a source of ectopically expressed epithelial cell–specific markers and that CD34+cells may contribute to the high false-positive rate generally observed when those markers are used to detect rare circulating metastatic cancer cells by RT-PCR.

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