Kazuhiro Yamanoi

Overexpression of leucine-rich repeat-containing G protein-coupled receptor 5 in gastric cancer

Gastric cancer is one of the most common malignancies worldwide and patients with advanced gastric cancer still have poor clinical outcomes. The overexpression of leucine‐rich repeat‐containing G protein‐coupled receptor 5 (LGR5) mRNA in colorectal cancer and its correlation with clinicopathological factors were recently reported by us. In this study, we show LGR5 mRNA overexpression in human gastric cancer specimens by quantitative RT‐PCR and in situ hybridization and assess a correlation with clinicopathological factors. The mean expression of LGR5 mRNA in cancerous tissues was five times higher than that in normal tissue (P = 0.0002). Furthermore, LGR5 mRNA expression show marked variation among cases and significantly increased in cases where lymphatic invasion was present compared with those where it was absent (P = 0.0056). Although the mean expression level of LGR5 was observed to be higher in nodal metastasis and venous invasion positive cases compared to negative cases, a significant difference was not observed. These results suggest that LGR5 can be a biomarker for malignancy in gastric cancer.

Epigenetic clustering of gastric carcinomas based on DNA methylation profiles at the precancerous stage: its correlation with tumor aggressiveness and patient outcome

Summary Single-CpG resolution genome-wide DNA methylation analysis indicated that distinct DNA methylation profiles are established during field cancerization in gastric mucosae, and such profiles at the precancerous stage are inherited by gastric cancers, thus determining tumor aggressiveness and patient outcome.

Multiple gastrointestinal stromal tumors with novel germline c-kit gene mutation, K642T, at exon 13

Multiple gastrointestinal stromal tumors (GISTs) caused by germline c-kit gene mutations are an extremely rare autosomal dominant disorder. A 57-year-old Japanese woman was referred to a hospital for appetite loss and severe weight loss. She had 2 large abdominal masses around the stomach, which were surgically resected. Histological examination revealed that these tumors were GISTs. Multiple microscopic GISTs and diffuse hyperplasia of the interstitial cells of Cajal were also seen in the background gastric and small intestinal walls. Characteristically, the GISTs showed severe hyalinization with calcification and partial heterotopic ossification, which may have caused the patients severe dysphagia. Mutational analysis of the c-kit gene revealed a substitution at codon 642 in exon 13 (K642T) in the tumor, normal ileal mucosa and peripheral blood leukocytes, indicating that the mutation is in the germline. This is the first case of multiple GISTs with novel germline c-kit gene mutation at exon 13.

Decreased expression of gastric gland mucin-specific glycan α1,4-linked N-acetylglucosamine on its scaffold mucin 6 is associated with malignant potential of pyloric gland adenoma of the stomach

Pyloric gland adenoma (PGA) is a unique gastric neoplasm expressing mucin 6 (MUC6), and is often associated with high‐grade dysplasia and/or adenocarcinoma. MUC6 secreted from the gastric gland mucous cells, such as pyloric gland cells, carries unique O‐glycans with terminal α1,4‐linked N‐acetylglucosamine (αGlcNAc) residues on its molecule. As we recently demonstrated that αGlcNAc serves as a tumour suppressor for gastric adenocarcinoma, this study aimed to investigate the significance of αGlcNAc expression in PGA.

Hepatoid adenocarcinoma of the stomach with multi-nucleated giant cell proliferation in a 100-year-old man

Few cases of gastric carcinoma with infiltrating multi‐nucleated giant cells (MGCs) have been reported, and giant cells infiltrating the gastric carcinoma were previously described as osteoclast‐like giant cells (OGCs). However, hepatoid adenocarcinomas have never been reported previously, and the present case is extremely rare. A 100‐year‐old Japanese man with gastralgia was found to have a mass in his gastric body. Histological examination showed a poorly differentiated adenocarcinoma with infiltration of MGCs. Vascular and lymphatic invasion were noted but there were no metastases. Almost all the tumor comprised cells with hepatoid‐like features. The MGCs proliferated, with infiltration of lymphoid cells. A few MGCs contained mucous material in their cytoplasm, indicating these were foreign‐body giant cells. Immunohistochemically, the hepatoid‐like components were positive for AFP, and staining with polyclonal antibodies against carcinoembryonic antigen (CEA) showed the canalicular pattern. We concluded that this component was hepatoid adenocarcinoma. The MGCs were positive for CD68, and surrounding infiltrating lymphoid cells were diffusely positive for CD3. Infiltration of MGCs in gastric cancer may represent a cellular immune response against gastric cancer invasion. Further studies are required to elucidate the etiology of MGCs in gastric cancer.

Reduced αGlcNAc glycosylation on gastric gland mucin is a biomarker of malignant potential for gastric cancer, Barrett’s adenocarcinoma, and pancreatic cancer

Gastric gland mucin secreted from pyloric gland cells, mucous neck cells, and cardiac gland cells of the gastric mucosa harbors unique O-glycans carrying terminal α1,4-linked N-acetylglucosamine residues (αGlcNAc), which are primarily attached to the scaffold mucin core protein MUC6. αGlcNAc acts as an antibiotic against Helicobacter pylori (H. pylori), a microbe causing gastric cancer. In addition, mice deficient in A4gnt, which encodes the enzyme α1,4-N-acetylglucosaminyltransferase (α4GnT) that catalyzes αGlcNAc biosynthesis, spontaneously develop gastric differentiated-type adenocarcinoma, even if not infected by H. pylori. Thus, αGlcNAc prevents gastric cancer as both an antibiotic and a tumor suppressor (Nakayama in Acta Histochem Cytochem 47:1–9, 2014b). Indeed, in humans αGlcNAc loss on MUC6 in differentiated-type adenocarcinoma is closely associated with poor patient prognosis (Shiratsu et al. in Cancer Sci 105:126–133, 2014). Recently, we reported reduced αGlcNAc expression on MUC6 in both pyloric gland adenoma of the stomach and chronic atrophic gastritis, in Barrett’s esophagus, and in pancreatic intraductal papillary-mucinous neoplasm (IPMN)/pancreatic intraepithelial neoplasia (PanIN), all potentially premalignant conditions. This review discusses whether relatively reduced levels of αGlcNAc in these lesions could serve as a biomarker to predict malignant potential and cancer progression.

Gastric gland mucin-specific O -glycan expression decreases with tumor progression from precursor lesions to pancreatic cancer

Pancreatic cancer is lethal, as it is often detected late. Thus, novel biomarkers of precursor lesions are needed to devise timely therapies. Pancreatic intraepithelial neoplasia (PanIN) and intraductal papillary mucinous neoplasm (IPMN) are major precursors of pancreatic cancer. In normal gastric mucosa, gastric gland mucin‐specific O‐glycans are unique in having α1,4‐linked N‐acetylglucosamine (αGlcNAc) residues attached to MUC6. Recently we reported that αGlcNAc functions as a tumor suppressor for differentiated‐type gastric adenocarcinoma (Karasawa et al., J Clin Invest 122, 923, 2012). MUC6 is also expressed in pancreatic neoplasms, including PanIN and IPMN, but the role of αGlcNAc expression in pancreatic neoplasms remains unknown. Here, we analyze expression patterns of αGlcNAc, MUC6 and MUC5AC in pancreatic neoplasms and compare them with progression from PanIN to invasive ductal adenocarcinoma (IDAC) (the PanIN‐IDAC sequence; 20 cases) and from IPMN to IPMN with associated invasive carcinoma (IPMNAIC) (the IPMN‐IPMNAIC sequence; 20 cases). At both sequences, the frequency of MUC6‐positive and αGlcNAc‐positive lesions decreased with tumor progression. We then compared expression levels of αGlcNAc and MUC6 at each step of the progression. At the PanIN‐IDAC sequence, αGlcNAc expression significantly decreased relative to MUC6 in low‐grade PanIN (P = 0.021), high‐grade PanIN/intraductal spread of IDAC (P = 0.031) and IDAC (P = 0.013). At the IPMN‐IPMNAIC sequence, decreased αGlcNAc expression was also observed in low‐grade IPMN exhibiting gastric‐type morphology (P = 0.020). These results suggest that decreased expression of αGlcNAc relative to MUC6 occurs early and marks the initiation of tumor progression to pancreatic cancer.

Leucine-rich repeat-containing G-protein-coupled receptor 5 expression and clinicopathological features of colorectal neuroendocrine neoplasms: LGR5 in colorectal neuroendocrine neoplasms

LGR5 is expressed in various tumors and has been identified as a putative intestinal stem cell marker. Here we investigated LGR5 expression in colorectal neuroendocrine neoplasms and analyzed the correlation with pathological characteristics. We evaluated the clinicopathological features of 8 neuroendocrine tumor (NET) grade 1 (NET G1), 4 NET Grade 2 (NET G2), and 8 NET Grade 3 (NET G3; also termed neuroendocrine carcinoma, or NEC) cases. We examined LGR5 expression using an RNAscope, a newly developed RNA in situ hybridization technique, with a tissue microarray of the neuroendocrine neoplasm samples. LGR5 staining in individual tumor cells was semi‐quantitatively scored using an H‐score scale. We also performed a combination of LGR5 RNA in situ hybridization and synaptophysin immunohistochemistry. All cases contained tumor cells with some LGR5‐positive dots. For all cases, H‐scores showed a positive correlation with nuclear beta‐catenin expression. In the NEC group, there was a strong positive correlation between H‐score and beta‐catenin expression. Our findings suggest that LGR5 may serve as a stem cell marker in NEC, as is the case in colon adenocarcinoma. The positive correlation between H‐score and beta‐catenin expression suggests that LGR5 expression might be affected by beta‐catenin expression in neuroendocrine neoplasms and especially in NEC.