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Purpose: Circulating microRNAs (miRNA) are emerging as promising diagnostic biomarkers for colorectal cancer, but their usefulness for detecting early colorectal neoplasms remains unclear. This study aimed to identify serum miRNA biomarkers for the identification of patients with early colorectal neoplasms.Experimental Design: A cohort of 237 serum samples from 160 patients with early colorectal neoplasms (148 precancerous lesions and 12 cancers) and 77 healthy subjects was analyzed in a three-step approach that included a comprehensive literature review for published biomarkers, a screening phase, and a validation phase. RNA was extracted from sera, and levels of miRNAs were examined by real-time RT-PCR.Results: Nine miRNAs (miR-18a, miR-19a, miR-19b, miR-20a, miR-21, miR-24, miR-29a, miR-92, and miR-125b) were selected as candidate biomarkers for initial analysis. In the screening phase, serum levels of miR-21, miR-29a, and miR-125b were significantly higher in patients with early colorectal neoplasm than in healthy controls. Elevated levels of miR-21, miR-29a, and miR-125b were confirmed in the validation phase using an independent set of subjects. Area under the curve (AUC) values for serum miR-21, miR-29a, miR-125b, and their combined score in discriminating patients with early colorectal neoplasm from healthy controls were 0.706, 0.741, 0.806, and 0.827, respectively. Serum levels of miR-29a and miR-125b were significantly higher in patients who had only small colorectal neoplasms (≤5 mm) than in healthy subjects.Conclusions: Because serum levels of miR-21, miR-29a, and miR-125b discriminated patients with early colorectal neoplasm from healthy controls, our data highlight the potential clinical use of these molecular signatures for noninvasive screening of patients with colorectal neoplasia. Clin Cancer Res; 21(18); 4234–42. ©2015 AACR.

Supplementary figure 1: Expression levels of serum miR-21, miR-29a and miR-125b in healthy controls (HC) vs. patients with colorectal neoplasms (CRN), after exclusion of 8 patients with colorectal cancer.

Supplementary figure 2: Kaplan Meier survival curves based upon expression levels of miR-21, miR-29a, miR-125b and their combination, when comparing patients with colorectal neoplasms (CRN) with healthy controls (HC), and when comparing patients with advanced neoplasms (AN) with patients without advanced neoplasms (non-AN), after exclusion of all patients with colorectal cancer.

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Although concurrent occurrence of spondyloarthritis (SpA) and ulcerative colitis (UC) is sometimes seen, the profiles of cytokines have been poorly understood in UC-associated SpA. We herein report a case of UC-associated SpA successfully treated with infliximab. Profiles of cytokines in the serum and colonic mucosa were characterized by an enhanced expression of IL-6 but not TNF-α. Successful induction of remission by infliximab was associated with the downregulation of IL-6 expression but no significant alteration in TNF-α expression. These findings suggest that some cases of UC-associated SpA might be driven by IL-6, and infliximab might be effective in cases lacking enhanced TNF-α responses.

Polymorphisms in the autophagy-related protein 16 like 1 (ATG16L1) and nucleotide-binding oligomerization domain 2 (NOD2) genes are associated with Crohn’s disease (CD). Impaired interaction between ATG16L1 and NOD2 underlies CD immunopathogenesis. Although activation of the receptor-interacting serine–threonine kinase (RICK, also known as RIP2), a downstream signaling molecule for NOD2 and multiple toll-like receptors (TLRs), plays a pathogenic role in the development of inflammatory bowel disease, the molecular interaction between ATG16L1 and RICK/RIP2 remains poorly understood. In this study, we examined the physical interaction between ATG16L1 and RICK/RIP2 in human embryonic kidney 293 cells and human monocyte-derived dendritic cells (DCs) expressing excessive and endogenous levels of these proteins, respectively. We established that ATG16L1 binds to RICK/RIP2 kinase domain and negatively regulates TLR2-mediated nuclear factor-kappa B (NF-κB) activation and pro-inflammatory cytokine responses by inhibiting the interaction between TLR2 and RICK/RIP2. Binding of ATG16L1 to RICK/RIP2 suppressed NF-κB activation by down-regulating RICK/RIP2 polyubiquitination. Notably, the percentage of colonic DCs expressing ATG16L1 inversely correlated with IL-6 and TNF-α expression levels in the colon of CD patients. These data suggest that the interaction between ATG16L1 and RICK/RIP2 maintains intestinal homeostasis via the down-regulation of TLR-mediated pro-inflammatory cytokine responses.

BackgroundThe standard therapy for acute severe ulcerative colitis (ASUC) is intravenous corticosteroids; however, 30% of UC patients do not recover with corticosteroids alone. Infliximab or cyclosporin is indicated to decrease the rates of colectomy of hospitalized ASUC patients. Although a previous study showed that moderate to severe UC had a rapid response to tofacitinib, few studies have reported the efficacy and safety of tofacitinib for ASUC.We report a case series of successful first-line treatment consisting of high-dose tofacitinib (20 mg/day) administered to ASUC patients with steroid resistance. Case presentationPatients diagnosed with ASUC at our institution between October 2018 and February 2020 were retrospectively evaluated. They were administered high dose of tofacitinib (20 mg) after they had no response to steroid therapy comprising a dose of 1 to 1.5 mg/kg/day. This study included a total of eight patients with ASUC. There were four (50%) men and four (50%) women with a median age of 47.1 years (range, 19–65 years). All patients had a median UC duration of 4 years (range, 0–20 years). Four patients had no previous history of UC.A clinical response was observed in six of the eight patients before they experienced remission. Six patients were able to avoid colectomy. One patient (patient 2) had no response; however, remission was achieved after switching from tofacitinib to infliximab. One patient (patient 6) with no response to tofacitinib underwent total colectomy. When we used tofacitinib during induction and the follow-up phase, only one patient (patient 4) experienced a major adverse event, herpes zoster, which was treated with acyclovir without stopping tofacitinib. ConclusionClinical remission can be achieved with high probability and colectomy can be avoided by administering high-dose tofacitinib to steroid-resistant ASUC patients.

Collagenous colitis (CC), a prototypical microscopic colitis, is a chronic inflammatory disorder of the colon. The diagnosis of CC depends on the pathological examination. The colonic mucosa of patients with CC is characterized by the presence of a substantially thickened collagen band (>10μm) under the surface epithelium. In addition, intraepithelial and lamina propria lymphocytes are markedly increased in patients with CC. However, the roles played by the lymphocytes accumulating in the colonic mucosa of patients with CC are poorly defined. Recent studies indicate that T cells infiltrating the colonic mucosa of patients with CC are mainly represented by CD4+ T cells, CD8+ T cells, and forkhead box P3 (FOXP3)+ regulatory T cells (Tregs). Given that activation of CD4+/CD8+ T cells and FOXP3+ Tregs usually mediates pro-inflammatory and anti-inflammatory responses, respectively, alterations in the colonic numbers of these adaptive T cells might be related to the resolution of colitis in patients with CC. We determined alterations in the composition of colonic T cells by extensive immunohistochemical (IHC) analyses in a case of CC successfully treated with budesonide and metronidazole. Colonic lamina propria immune cells mainly comprised CD3+ T cells, CD4+ T cells, CD8+ T cells, CD68+ macrophages, and FOXP3+ Tregs, but not CD20+ B cells or myeloperoxidase (MPO)+ granulocytes in the active phase. During remission, the numbers of CD3+ T cells, CD4+ T cells, CD8+ T cells, and CD68+ macrophages did not change significantly in the colonic lamina propria, whereas FOXP3+ Tregs were markedly decreased, suggesting that induction of remission was achieved in a Treg-independent manner. Thus, our study indicates that accumulation of FOXP3+ Tregs in the colonic mucosa of patients with CC might be a counter-regulatory mechanism reflecting persistent inflammation and that induction of remission might be achieved without activation of Tregs.

Background Despite the high incidence of spondyloarthritis (SpA) as an extra-intestinal manifestation of Crohn's disease (CD), the immunopathogenesis of CD-associated SpA remains largely unknown. Objective We tried to explore molecular mechanisms accounting for the development of CD-associated SpA in a patient successfully treated with infliximab. Methods Peripheral blood mononuclear cells (PBMCs) before infliximab treatment were stimulated with Toll-like receptor (TLR) ligands to measure pro-inflammatory cytokine responses. Endoscopic biopsy samples before and after infliximab treatment were subjected to quantitative polymerase chain reaction. Results PBMCs from this CD-associated SpA patient exhibited higher production of pro-inflammatory cytokines upon stimulation with TLR ligands than PBMCs from healthy controls. Induction of remission by infliximab was associated with the downregulation of pro-inflammatory cytokine responses in the small intestinal mucosa, which is continually exposed to TLR ligands. Conclusions Excessive pro-inflammatory cytokine responses to TLR ligands might underlie the immunopathogenesis of CD-associated SpA.

Inflammatory bowel diseases (IBDs) are becoming more frequent worldwide. A significant fraction of patients with IBD are refractory to various types of therapeutic biologics and small molecules. Therefore, identification of novel therapeutic targets in IBD is required. Receptor-interacting serine/threonine kinase 2 (RIPK2), also known as receptor-interacting protein 2 (RIP2), is a downstream signaling molecule for nucleotide-binding oligomerization domain 1 (NOD1), NOD2, and Toll-like receptors (TLRs). RIPK2 is expressed in antigen-presenting cells, such as dendritic cells and macrophages. Recognition of microbe-associated molecular patterns by NOD1, NOD2, and TLRs leads to the interaction between RIPK2 and these innate immune receptors, followed by the release of pro-inflammatory cytokines such as TNF-α, IL-6, and IL-12/23p40 through the activation of nuclear factor kappa B and mitogen-activated protein kinases. Thus, activation of RIPK2 plays a critical role in host defense against microbial infections. Recent experimental and clinical studies have provided evidence that activation of RIPK2 is involved in the development of autoimmune diseases, especially IBDs. In addition, the colonic mucosa of patients with IBD exhibits enhanced expression of RIPK2 and associated signaling molecules. Furthermore, the blockage of RIPK2 activation ameliorates the development of experimental murine colitis. Thus, activation of RIPK2 underlies IBD immunopathogenesis. In this review, we attempt to clarify the roles played by RIPK2 in the development of IBD by focusing on its associated signaling pathways. We also discuss the possibility of using RIPK2 as a new therapeutic target in IBD.

Due to the tendency of gastric linitis plastica (GLP) to cause extensive submucosal infiltration, a superficial endoscopic biopsy sometimes yields no evidence of malignancy, hindering definite diagnosis. The present study was a single-center retrospective analysis of 54 consecutive patients diagnosed with GLP between 2016 and 2020 to evaluate EUS-guided fine-needle aspiration (EUS-FNA) biopsy outcomes in patients with negative endoscopic biopsy findings. A pathological GLP diagnosis was achieved by endoscopic biopsy in 40 patients (74.1%). EUS-FNA biopsy with a 22-gauge needle was performed in 13 of the remaining 14 patients, and GLP diagnosis was confirmed in 10 patients, with a median of three needle passes. The remaining four patients were laparoscopically diagnosed with GLP. The diagnostic ability of EUS-FNA biopsy for GLP was 76.9%, and EUS-FNA biopsy contributed to GLP diagnosis in 18.5% (10/54) of all cases. None of the 13 patients exhibited EUS-FNA biopsy-related adverse events. Univariable and multivariable analyses revealed an absence of superficial ulcerations as a predictor of false-negative endoscopic biopsy findings in patients with GLP. These results suggest EUS-FNA biopsy as a minimally invasive and safe alternative diagnostic modality for GLP in cases where conventional endoscopic biopsy fails to verify malignancy, although prospective studies with larger cohorts are warranted to confirm these findings.

Purpose: Circulating microRNAs (miRNA) are emerging as promising diagnostic biomarkers for colorectal cancer, but their usefulness for detecting early colorectal neoplasms remains unclear. This study aimed to identify serum miRNA biomarkers for the identification of patients with early colorectal neoplasms. Experimental Design: A cohort of 237 serum samples from 160 patients with early colorectal neoplasms (148 precancerous lesions and 12 cancers) and 77 healthy subjects was analyzed in a three-step approach that included a comprehensive literature review for published biomarkers, a screening phase, and a validation phase. RNA was extracted from sera, and levels of miRNAs were examined by real-time RT-PCR. Results: Nine miRNAs (miR-18a, miR-19a, miR-19b, miR-20a, miR-21, miR-24, miR-29a, miR-92, and miR-125b) were selected as candidate biomarkers for initial analysis. In the screening phase, serum levels of miR-21, miR-29a, and miR-125b were significantly higher in patients with early colorectal neoplasm than in healthy controls. Elevated levels of miR-21, miR-29a, and miR-125b were confirmed in the validation phase using an independent set of subjects. Area under the curve (AUC) values for serum miR-21, miR-29a, miR-125b, and their combined score in discriminating patients with early colorectal neoplasm from healthy controls were 0.706, 0.741, 0.806, and 0.827, respectively. Serum levels of miR-29a and miR-125b were significantly higher in patients who had only small colorectal neoplasms (≤5 mm) than in healthy subjects. Conclusions: Because serum levels of miR-21, miR-29a, and miR-125b discriminated patients with early colorectal neoplasm from healthy controls, our data highlight the potential clinical use of these molecular signatures for noninvasive screening of patients with colorectal neoplasia. Clin Cancer Res; 21(18); 4234–42. ©2015 AACR.

Objectives: Early detection of precancerous lesions and early-stage cancers are essential to reduce mortality from colorectal cancer (CRC). Serum/plasma microRNAs (miRs) have been reported as diagnostic and prognostic biomarkers of CRC, but their utility in detecting patients with early lesions has not been fully explored. The aim of this study is to identify serum miRs which are potentially useful as diagnostic biomarkers for early colorectal neoplasms, and potential confounders in a blood test. Methods: Candidate biomarker miRs were selected from previously published studies. Serum samples were collected from patients with low-grade intraepithelial neoplasias (LGINs) including tubular adenomas and tubulovillous adenomas, high-grade intraepithelial neoplasias (HGINs), early cancers, and healthy volunteers, and divided into discovery and validation sets. Total RNA was extracted from serum samples and levels of candidate miRs were measured by real-time RT-PCR. To evaluate the impact of hemolysis on serum miR levels, serially diluted hemolysed controls were prepared from stock solution made by hemolysing the red blood cells in distilled water. Levels of candidate miRs in hemolysed controls were also examined by real-time RT-PCR. Absorbance of serum samples at 560 nm, 576 nm, and 592 nm was measured by spectrophotometry and hemoglobin levels were estimated from a standard curve. Results: Fourteen miRs were selected as candidate diagnostic biomarkers of early colorectal neoplasms from a review of the literature. Serum levels of five miRs were too low to be accurately quantified by real-time RT-PCR, and excluded from further evaluation. Sera from 12 patients with LGINs, 8 patients with HGINs, 4 patients with cancers, and 25 healthy volunteers were included in the discovery phase. Of 9 miRs examined, miR21 (p = 0.0007), miR29a (p < 0.0001) and miR125b (p = 0.0198) showed significantly higher levels in sera from early colorectal neoplasms compared to those from healthy volunteers. Higher levels of miR29a and miR125b in sera from early colorectal neoplasms were confirmed in the validation phase using independent set of samples. Levels of miR21, 29a, and miR125b in sera significantly correlated with the degree of hemolysis. Conclusions: Serum miRs may be useful biomarkers to detect patients with early colorectal neoplasia, including adenomas. Hemolysis affects serum miR levels, and will confound measurement. Since hemolysis occurs frequently during blood collection, its impact on serum miRs should be further investigated. Citation Format: Atsushi Yamada, Takahiro Horimatsu, Yoshinaga Okugawa, Naoshi Nishida, Tadayuki Kou, Toshihiro Kusaka, Hajime Honjo, Yusuke Amanuma, Osamu Kikuchi, Manabu Muto, Ajay Goel, C. Richard Boland. Serum microRNAs as diagnostic biomarkers for early colorectal neoplasms. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 867. doi:10.1158/1538-7445.AM2014-867