Decreased expression of fecal miR-4478 and miR-1295b-3p in early-stage colorectal cancer

BACKGROUND: Colorectal cancer (CRC) is a major cause of cancer-related deaths world-wide. Detection of molecular markers in stool samples is a promising strategy for CRC screening. MicroRNAs (miRNAs) are short, non-coding RNA molecules that are commonly dysregulated in neoplasia. OBJECTIVE: The objective of this study was to evaluate the fecal miRNAs differentiation between early-stage CRC patients and healthy subjects. METHODS: Stool samples were collected from 40 patients with early stage (I, II) CRC and 16 healthy controls. RNA was extracted from all samples using miRNAeasy Mini Kits. MiRNA microarray expression profiling was performed with Agilent's miRNA Microarray system on 12 CRC and 8 normal stool samples. The expression levels of miR-4478 and miR-1295b-3p were determined by the SYBR Green miScript PCR system. RESULTS: In profiling study, we found 215 down-regulated miRNAs in CRC group. Furthermore, in validation study we found that the expression levels of fecal miR-4487 and miR-1295b-3p were significantly decreased in CRC patients compared to healthy controls. CONCLUSIONS: The expression of miR-4478 and miR-1295b-3p were significantly diminished in stool samples of CRC patients with early stage (I, II) in comparison with normal group. These miRNAs maybe use as potential non-invasive molecular markers for CRC diagnosis, but further studies are needed. © 2015 - IOS Press and the authors. All rights reserved

Downregulation of plasma MiR-142-3p and MiR-26a-5p in patients with colorectal carcinoma

Background: Colorectal cancer is one of the most commonly diagnosed cancers and cancer- related death worldwide. Identification of new specific biomarkers could be helpful to detection of this malignancy. Altered plasma microRNA expression has been identified in many cancers, including colorectal cancer. Objectives: The main objective of this study was to identify the circulating microRNAs with the most expression changes in colorectal cancer patients compared with neoplasm free healthy individuals. Materials and Methods: MicroRNA expression profiling was performed on plasma samples of 37 colorectal cancer patients and 8 normal subjects using microRNA microarray. Quantitative real-time reverse transcription polymerase chain reaction was used to validate the two selected altered microR NAs. Plasma samples from 61 colorectal cancer patients and 24 normal subjects were used in our validation study. Results: In profiling study we found a panel of six plasma microRNAs with significant downregulation. MicroRNA-142-3p and microRNA-26a-5p were selected and validated by polymerase chain reaction. Our results demonstrated that expression levels of plasma microRNA-142-3p and microRNA-26a-5p were significantly downregulated in patients with colorectal cancer when compared to control group. Conclusions: Our findings suggest that downregulation of plasma microRNA-142-3p and microRNA-26a-5p might serve as novel noninvasive biomarkers in the diagnosis of colorectal cancer, although more studies are needed to highlight the theoretical strengths. © 2015, Iranian Journal of Cancer Prevention

Expression, Tissue Distribution and Function of miR-21 in Esophageal Squamous Cell Carcinoma

Objective:MiR-21 is an oncomir expressed by malignant cells and/or tumor microenvironment components. In this study we focused on understanding the effects of stromal miR-21 on esophageal malignant cells.Design:MiR-21 expression was evaluated in formalin-fixed paraffin-embedded samples from patients with esophageal squamous-cell carcinoma (SCC) by quantitative RT-PCR. MiR-21 tissue distribution was visualized with in situ hybridization. A co-culture system of normal fibroblasts and esophageal cancer cells was used to determine the effects of fibroblasts on miR-21 expression levels, and on SCC cell migration and invasion.Results:MiR-21 was overexpressed in SCCs, when compared to the adjacent non-tumor tissues (P = 0.0007), and was mainly localized in the cytoplasm of stromal cells adjacent to malignant cells. Accordingly, miR-21 expression was increased in tumors with high versus low stromal content (P = 0.04). When co-cultured with normal fibroblasts, miR-21 expression was elevated in SCC cells (KYSE-30), while its expression was restricted to fibroblasts when co-cultured with adenocarcinoma cells (OE-33 and FLO-1). MiR-21 was detected in conditioned media of cancer cell lines, illustrating the release of this miRNA into the environment. Co-culturing with normal fibroblasts or addition of fibroblast conditioned media caused a significant increase in cell migration and invasion potency of KYSE-30 cells (P<0.0001). In addition, co-culturing cancer cells with fibroblasts and expression of miR-21 induced the expression of the cancer associated fibroblast (CAF) marker S100A4.Conclusions:MiR-21 expression is mostly confined to the SCC stroma and its release from fibroblasts influences the migration and invasion capacity of SCC cells. Moreover, miR-21 may be an important factor in "activating" fibroblasts to CAFs. These findings provide new insights into the role of CAFs and the extracellular matrix in tumor microenvironment formation and in tumor cell maintenance, and suggest miR-21 may contribute to cellular crosstalk in the tumor microenvironment. © 2013 Nouraee et al

A novel signaling role for miR-451 in esophageal tumor microenvironment and its contribution to tumor progression

Objective: We evaluated miR-451 expression in serum and tissue samples of esophageal squamous cell carcinoma (ESCC) patients. Then, we examined a secretory role of miR-451 in esophageal tumor microenvironment. Methods: miR-451 expression was evaluated in 39 serum samples from esophageal SCC patients compared to 39 normal individuals as well as 26 pairs of fresh-frozen tumor and adjacent normal tissues from patients with ESCC, using qRT-PCR. In a co-culture system of human normal fibroblasts (HFSF-PI3) and esophageal cancer cell line (KYSE-30), we evaluated exosomal miR-451 secretion into the conditioned medium (CM) of both cell lines. Then, we analyzed the effect of primiR-451-transfected fibroblasts on the migration potency of their neighboring KYSE-30 cells. Results: We detected miR-451 over-expression in serum samples of esophageal cancer patients compared to the normal group (P = 0.005). Interestingly, fresh-frozen tumor tissues from the same patients showed miR-451 down-regulation compared to their adjacent normal counterparts (P = 0.043). Co-culturing the KYSE-30 cell line with normal fibroblasts significantly induced miR-451 exosomal secretion into the CM. Moreover, co-culture of KYSE-30 cell line with miR-451-over-expressing fibroblasts significantly induced migration tendency in KYSE-30 cell line compared to the mock-transfected fibroblasts (P < 0.0001). In this system, MIF expression (a validated target of miR-451) in the KYSE-30 cell line was increased although this alteration was not statistically significant (fold change = 4.44). Conclusions: Our data suggest that cancer-associated fibroblasts use exosomal miR-451 as a signaling molecule to provide a favorable niche for tumor cell migration and cancer progression. Our findings provide new insights into the stromal role of miR-451 in the esophageal tumor microenvironment as a communicatory molecule and suggest a signaling role for miR-451 in extracellular matrix cross-talks. © 2016, Federación de Sociedades Españolas de Oncología (FESEO)

Single-cell transcriptomics identifies multiple pathways underlying antitumor function of TCR- and CD8αβ-engineered human CD4⁺ T cells.

Transgenic coexpression of a class I-restricted tumor antigen-specific T cell receptor (TCR) and CD8αβ (TCR8) redirects antigen specificity of CD4 &lt;sup&gt;+&lt;/sup&gt; T cells. Reinforcement of biophysical properties and early TCR signaling explain how redirected CD4 &lt;sup&gt;+&lt;/sup&gt; T cells recognize target cells, but the transcriptional basis for their acquired antitumor function remains elusive. We, therefore, interrogated redirected human CD4 &lt;sup&gt;+&lt;/sup&gt; and CD8 &lt;sup&gt;+&lt;/sup&gt; T cells by single-cell RNA sequencing and characterized them experimentally in bulk and single-cell assays and a mouse xenograft model. TCR8 expression enhanced CD8 &lt;sup&gt;+&lt;/sup&gt; T cell function and preserved less differentiated CD4 &lt;sup&gt;+&lt;/sup&gt; and CD8 &lt;sup&gt;+&lt;/sup&gt; T cells after tumor challenge. TCR8 &lt;sup&gt;+&lt;/sup&gt; CD4 &lt;sup&gt;+&lt;/sup&gt; T cells were most potent by activating multiple transcriptional programs associated with enhanced antitumor function. We found sustained activation of cytotoxicity, costimulation, oxidative phosphorylation- and proliferation-related genes, and simultaneously reduced differentiation and exhaustion. Our study identifies molecular features of TCR8 expression that can guide the development of enhanced immunotherapies

A Costimulatory CAR Improves TCR-based Cancer Immunotherapy.

T-cell receptors (TCR) recognize intracellular and extracellular cancer antigens, allowing T cells to target many tumor antigens. To sustain proliferation and persistence, T cells require not only signaling through the TCR (signal 1), but also costimulatory (signal 2) and cytokine (signal 3) signaling. Because most cancer cells lack costimulatory molecules, TCR engagement at the tumor site results in incomplete T-cell activation and transient antitumor effects. To overcome this lack of signal 2, we genetically modified tumor-specific T cells with a costimulatory chimeric antigen receptor (CoCAR). Like classical CARs, CoCARs combine the antigen-binding domain of an antibody with costimulatory endodomains to trigger T-cell proliferation, but CoCARs lack the cytotoxic CD3ζ chain to avoid toxicity to normal tissues. We first tested a CD19-targeting CoCAR in combination with an HLA-A*02:01-restricted, survivin-specific transgenic TCR (sTCR) in serial cocultures with leukemia cells coexpressing the cognate peptide-HLA complex (signal 1) and CD19 (signal 2). The CoCAR enabled sTCR+ T cells to kill tumors over a median of four additional tumor challenges. CoCAR activity depended on CD19 but was maintained in tumors with heterogeneous CD19 expression. In a murine tumor model, sTCR+CoCAR+ T cells improved tumor control and prolonged survival compared with sTCR+ T cells. We further evaluated the CoCAR in Epstein-Barr virus-specific T cells (EBVST). CoCAR-expressing EBVSTs expanded more rapidly than nontransduced EBVSTs and delayed tumor progression in an EBV+ murine lymphoma model. Overall, we demonstrated that the CoCAR can increase the activity of T cells expressing both native and transgenic TCRs and enhance antitumor responses