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Background and Aim: Although rectal neuroendocrine tumor (NET-G1) have potential metastatic capability, even among small tumors, no predictive biomarker for invasion and metastasis has been reported. We analyzed microRNA (miRNA) expression profiles in rectal NET-G1 tissues with and without lymphovascular invasion (LVI). Moreover, we then investigated their target genes to clarify the mechanism of invasion/metastasis in NET-G1. Methods: miRNA array analysis was performed using 7 rectal NET-G1 tissues with LVI and 7 without LVI. miRNA expression was confirmed by quantitative real-time PCR. A NET cell line H727 was transfected with miRNA mimic or target gene small interfering RNA, and migration and invasion assays were performed. Results: The expression levels of miR-144-3p and miR-451a were significantly higher in NET-G1 with LVI versus without LVI, as determined by miRNA array analysis and RT-qPCR. A significant correlation was observed between miR-144-3p and miR-451a expression levels, strongly suggesting miR144/451 cluster overexpression in NET-G1 with LVI. Bioinformatic analysis of target genes revealed that miR-144-3p and miR-451a directly interact with PTEN and p19 mRNA, respectively. Immunohistochemistry revealed significantly lower expression of PTEN and p19 in NET-G1 tissues with LVI than in those without LVI. The miR-144-3p and miR-451a mimic significantly increased cell migration/invasion capability, respectively. Knockdown of PTEN and p19 induced significant augmentation of cell invasion and migration capability, respectively. Conclusions: Our data suggest that overexpression of miR-144/miR-451 cluster promotes LVI via repression of PTEN and p19 in rectal NET-G1 cells. miR-144/451 cluster may be a novel biomarker for predicting invasion/metastasis in rectal NET-G1.

Background: Although serrated polyposis syndrome (SPS) is associated with an increased colorectal cancer (CRC) risk, the carcinogenic mechanisms remain unknown. We investigated clinicopathological characteristics and genetic abnormalities in colorectal polyps and CRC to elucidate carcinogenic mechanisms in SPS. Methods: We retrospectively analyzed clinicopathological features of colorectal polyps in 44 SPS patients, and examined mutations of genes including APC, RAS, BRAF, and TP53, and microsatellite instability (MSI) in CRC tissues. Results: Of the 44 patients, 25 (56%) fulfilled WHO criterion 1, 11 (25%) fulfilled criterion 2, and 8 (18%) fulfilled both. A total of 956 polyps were observed; 642 (67%) hyperplastic polyps (HP), 204 (21%) sessile serrated lesions (SSL), 10 (1%) traditional serrated adenoma (TSA), and 100 (11%) adenomas. The median numbers of polyps (/patient) were 10.5 (IQR 2.75-23) HPs, 4.0 (2.0-6.0) SSLs, 0 (0-0) TSA, and 1 (0-3.3) adenoma. SSL and HP located preferentially in the proximal and distal colon respectively. Twenty-two CRCs were found in 18 patients. Based on the histological coexistence of SSL/TSA, BRAF mutation and MSI, 5 CRCs (26%) were classified as serrated-neoplasia pathway. Conversely, based on the coexistence of adenoma, APC/RAS and TP53 mutations, 11 CRCs (58%) were classified as adenoma-carcinoma pathway. The remaining 3 were unclassifiable. Most CRCs through adenoma-carcinoma pathway were located in the left-side colorectum and patients bearing those met criterion 2, characterized by many HP and advanced adenomas. Adenoma was a significant risk factor for CRC. Conclusions: Our results suggest that more than half of the CRCs, particularly those in the left-side colorectum, developed through the adenoma-carcinoma pathway in SPS patients. Adenoma was a risk factor for CRCs, suggesting its importance in colorectal carcinogenesis.

Background Cowden syndrome (CS) is an autosomal-dominant hereditary disorder caused by a germline PTEN variant and characterized by multiple hamartomas and a high risk of cancers. However, no detailed data on CS in Asian patients nor genotype-phenotype correlation have been reported. Methods We performed the first Japanese nationwide questionnaire survey on CS, and obtained questionnaire response data on 49 CS patients. Results Patients included 26 females (median age 48 y). The incidence of breast, thyroid, endometrium, and colorectal cancer was 32.7%, 12.2%, 19.2% (among females), and 6.1% respectively. The incidence of any cancers was relatively high among all patients (46.9%, 23/49), and particularly female patients (73.1%, 19/26), compared with previous reports from Western countries. Gastrointestinal (GI) polyps were more frequently found throughout the GI tract compared with previous studies. PTEN variants were detected in 95.6% (22/23) of patients; 12 in the N-terminal region (11 in phosphatase domain) and 10 in the C-terminal (C2 domain) region. The incidence of cancer in the C2 domain group was significantly higher than in the N-terminal region (phosphatase) group. All female patients with C2 domain variant had breast cancer. Conclusion Our data suggest that Japanese patients with CS, particularly female patients and patients with C2 domain variant may have a high risk of cancers.

Aim: To investigate the therapeutic effect of lenvatinib (LEN) in liver disease etiology, especially nonviral hepatocellular carcinoma (HCC). Methods and Results: Sixty-seven patients with unresectable advanced HCC (u-HCC) treated with LEN and consisting of 26 hepatitis C virus (HCV), 19 hepatitis B virus (HBV), 11 alcohol, and 11 nonalcoholic steatohepatitis (NASH) cases were retrospectively recruited. Univariate and multivariate Cox proportional hazard models were used to determine predictive factors for survival. The objective response rate in the nonviral (alcohol and NASH) group was higher than that in the viral group (59.1% [13/22] vs. 46.7% [21/45]). Progression-free survival was significantly longer in the nonviral group than in the viral group (13.7 vs. 6.6 months; hazard ratio [HR] 0.324; 95% confidence interval [CI] 0.174–0.602; P

BACKGROUND AND AIMS: Small intestinal involvement in familial adenomatous polyposis (FAP) remains unclarified. We performed capsule endoscopy (CE) and balloon-assisted (BA) endoscopy to analyze small intestinal polyps and their genotype-phenotype correlations in a large cohort of FAP patients. METHODS: In this retrospective study, we performed CE in all 149 FAP patients and BA endoscopy in 74 patients with larger polyps. The prevalence of small intestinal adenoma with high-grade dysplasia and cancer was investigated. Correlation between APC variant and polyp phenotype was analyzed. RESULTS: Median patient age was 44 years and 69% were male. Pathogenic germline APC variants were found in 117/136 (86.1%). Overall prevalence and median number of small intestinal polyps were 85.2% (127/149) and 17 (IQR 6-36), respectively. A total of 5318 polyps were detected by CE and all 433 polyps histologically examined were confirmed as adenoma. The number of polyps significantly decreased from the proximal-to-distal tracts. Adenoma with high-grade dysplasia was detected in 26.2% (39/149), and its incidence among the polyps was approximately 1.4% (74/5318). Of these, 14 were intramucosal carcinomas with a prevalence of 5.4% (8/149). The germline APC variant in codon 1251-1580 was significantly correlated with a high number of small intestinal polyps. Multivariate analysis revealed APC variant in codon 1251-1580 and a high number of small intestinal polyps as independent risk factors for adenoma with high-grade dysplasia. Spigelman stage was significantly associated with number of jejunal/ileal polyps and their high-grade dysplasia. CONCLUSION: Among FAP patients, small intestinal adenoma with high-grade dysplasia was detected in 26.2% and cancer in 5.4%. FAP patients, particularly those with APC variant in codon 1251-1580 and/or high Spigelman stage may require surveillance for small intestinal polyps.

A primary splenic angiosarcoma is a rare type of soft tissue sarcoma and is associated with an extremely poor prognosis. In this study, we describe the case of a patient who was diagnosed with metastatic primary splenic angiosarcoma and survived for about 2 years. A 62-year-old female was referred to us for the treatment of splenic angiosarcoma with disseminated intravascular coagulation (DIC) and multiple liver and bone metastases. Paclitaxel therapy resulted in recovery from DIC and enabled her to continue sequential treatment through to sixth-line chemotherapy. We reviewed all splenic angiosarcoma case reports which were described as stage IV to date and compared with our case. From these data, we found that the median overall survival was 105 days, and the prognosis of splenic angiosarcoma of stage IV was worse than conventional case series. Splenectomy was performed in more patients than chemotherapy as a treatment. Moreover, various chemotherapeutic regimens were used. These data suggest that administering chemotherapy including paclitaxel to patients with splenic angiosarcoma might improve their prognosis.

Although right-sided colorectal cancer (CRC) shows a worse prognosis than left-sided CRC, the underlying mechanism remains unclear. We established patient-derived organoids (PDOs) from left- and right-sided CRCs and directly compared cell proliferation and invasion capability between them. We then analyzed the expression of numerous genes in signal transduction pathways to clarify the mechanism of the differential prognosis. Cell proliferation activity and invasion capability in right-sided cancer PDOs were significantly higher than in left-sided cancer PDOs and normal PDOs, as revealed by Cell Titer Glo and transwell assays, respectively. We then used quantitative RT-PCR to compare 184 genes in 30 pathways among right-sided and left-sided cancer and normal PDOs and found that the TIMP1 mRNA level was highest in right-sided PDOs. TIMP1 protein levels were upregulated in right-sided PDOs compared with normal PDOs but was downregulated in left-sided PDOs. TIMP1 knockdown with shRNA significantly decreased cell proliferation activity and invasion capability in right-sided PDOs but not in left-sided PDOs. Moreover, TIMP1 knockdown significantly decreased pFAK and pAkt expression levels in right-sided PDOs but not in left-sided PDOs. A database analysis of The Cancer Genome Atlas revealed that TIMP1 expression in right-sided CRCs was significantly higher than in left-sided CRCs. Kaplan–Meier survival analysis showed significantly shorter overall survival in high-TIMP1 patients versus low-TIMP1 patients with right-sided CRCs but not left-sided CRCs. Our data suggest that TIMP1 is overexpressed in right-sided CRCs and promotes cell proliferation and invasion capability through the TIMP1/FAK/Akt pathway, leading to a poor prognosis. The TIMP1/FAK/Akt pathway can be a target for therapeutic agents in right-sided CRCs.

N-acetylglucosamine (GlcNAc) is the fundamental amino sugar moiety that is essential for protein glycosylation. UDP-GlcNAc, an active form of GlcNAc, is synthesized through the hexosamine biosynthetic pathway (HBP). Arabidopsis N-acetylglucosamine-1-P uridylyltransferases (GlcNAc1pUTs), encoded by GlcNA.UTs, catalyze the last step in the HBP pathway, but their biochemical and molecular functions are less clear. In this study, the GlcNA.UT1 expression was knocked down by the double-stranded RNA interference (dsRNAi) in the glcna.ut2 null mutant background. The RNAi transgenic plants, which are referred to as iU1, displayed the reduced UDP-GlcNAc biosynthesis, altered protein N-glycosylation and induced an unfolded protein response under salt-stressed conditions. Moreover, the iU1 transgenic plants displayed sterility and salt hypersensitivity, including delay of both seed germination and early seedling establishment, which is associated with the induction of ABA biosynthesis and signaling. These salt hypersensitive phenotypes can be rescued by exogenous fluridone, an inhibitor of ABA biosynthesis, and by introducing an ABA-deficient mutant allele nced3 into iU1 transgenic plants. Transcriptomic analyses further supported the upregulated genes that were involved in ABA biosynthesis and signaling networks, and response to salt stress in iU1 plants. Collectively, these data indicated that GlcNAc1pUTs are essential for UDP-GlcNAc biosynthesis, protein N-glycosylation, fertility, and the response of plants to salt stress through ABA signaling pathways during seed germination and early seedling development.

Background/Aims: Sessile serrated adenomas/polyps (SSA/Ps) are a putative precursor lesion of colon cancer. Although the relevance of DNA hypermethylation in the SSA/P-cancer sequence is well documented, the role of DNA hypomethylation is unknown. We investigated the biological relevance of DNA hypomethylation in the SSA/P-cancer sequence by using 3-dimensional organoids of SSA/P. Methods: We first analyzed hypomethylated genes using datasets from our previous DNA methylation array analysis on 7 SSA/P and 2 cancer in SSA/P specimens. Expression levels of hypomethylated genes in SSA/P specimens were determined by RT-PCR and immunohistochemistry. We established 3-dimensional SSA/P organoids and performed knockdown experiments using a lentiviral shRNA vector. DNA hypomethylation at CpG sites of the gene was quantitated by MassARRAY analysis. Results: The mean number of hypomethylated genes in SSA/P and cancer in SSA/P was 41.6 ± 27.5 and 214 ± 19.8, respectively, showing a stepwise increment in hypomethylation during the SSA/P-cancer sequence. S100P, S100α2, PKP3, and MUC2 were most commonly hypomethylated in SSA/P specimens. The mRNA and protein expression levels of S100P, S100α2, and MUC2 were significantly elevated in SSA/P compared with normal colon tissues, as revealed by RT-PCR and immunohistochemistry, respectively. Among these, mRNA and protein levels were highest for S100P. Knockdown of the S100P gene using a lentiviral shRNA vector in 3-dimensional SSA/P organoids inhibited cell growth by >50% (p < 0.01). The mean diameter of SSA/P organoids with S100P gene knockdown was significantly smaller compared with control organoids. MassARRAY analysis of DNA hypomethylation in the S100P gene revealed significant hypomethylation at specific CpG sites in intron 1, exon 1, and the 5′-flanking promoter region. Conclusion: These results suggest that DNA hypomethylation, including S100P hypomethylation, is supposedly associated with the SSA/P-cancer sequence. S100P overexpression via DNA hypomethylation plays an important role in promoting cell growth in the SSA/P-cancer sequence.