Teppei Tagawa

Distinct molecular features of different macroscopic subtypes of colorectal neoplasms.

BACKGROUND Colorectal adenoma develops into cancer with the accumulation of genetic and epigenetic changes. We studied the underlying molecular and clinicopathological features to better understand the heterogeneity of colorectal neoplasms (CRNs). METHODS We evaluated both genetic (mutations of KRAS, BRAF, TP53, and PIK3CA, and microsatellite instability [MSI]) and epigenetic (methylation status of nine genes or sequences, including the CpG island methylator phenotype [CIMP] markers) alterations in 158 CRNs including 56 polypoid neoplasms (PNs), 25 granular type laterally spreading tumors (LST-Gs), 48 non-granular type LSTs (LST-NGs), 19 depressed neoplasms (DNs) and 10 small flat-elevated neoplasms (S-FNs) on the basis of macroscopic appearance. RESULTS S-FNs showed few molecular changes except SFRP1 methylation. Significant differences in the frequency of KRAS mutations were observed among subtypes (68% for LST-Gs, 36% for PNs, 16% for DNs and 6% for LST-NGs) (P<0.001). By contrast, the frequency of TP53 mutation was higher in DNs than PNs or LST-Gs (32% vs. 5% or 0%, respectively) (P<0.007). We also observed significant differences in the frequency of CIMP between LST-Gs and LST-NGs or PNs (32% vs. 6% or 5%, respectively) (P<0.005). Moreover, the methylation level of LINE-1 was significantly lower in DNs or LST-Gs than in PNs (58.3% or 60.5% vs. 63.2%, P<0.05). PIK3CA mutations were detected only in LSTs. Finally, multivariate analyses showed that macroscopic morphologies were significantly associated with an increased risk of molecular changes (PN or LST-G for KRAS mutation, odds ratio [OR] 9.11; LST-NG or DN for TP53 mutation, OR 5.30; LST-G for PIK3CA mutation, OR 26.53; LST-G or DN for LINE-1 hypomethylation, OR 3.41). CONCLUSION We demonstrated that CRNs could be classified into five macroscopic subtypes according to clinicopathological and molecular differences, suggesting that different mechanisms are involved in the pathogenesis of colorectal tumorigenesis.

Molecular features of colorectal polyps presenting Kudo's type II mucosal crypt pattern: are they based on the same mechanism of tumorigenesis?

Background and study aims: The molecular features of serrated polyps (SPs) with hyperplastic crypt pattern, also called Kudo’s type II observed by chromoendoscopy, were evaluated. Methods: The clinicopathological and molecular features of 114 SPs with a hyperplastic pit pattern detected under chromoendoscopy (five dysplastic SPs, 63 sessile serrated adenoma/polyps (SSA/Ps), 36 microvesicular hyperplastic polyps (MVHPs), and 10 goblet cell-rich hyperplastic polyps (GCHPs)) were examined. The frequency of KRAS and BRAF mutations and CpG island methylator phenotype (CIMP) were investigated. Results: Dysplastic SPs and SSA/Ps were frequently located in the proximal colon compared to others (SSA/Ps vs. MVHPs or GCHPs, P < 0.0001). No significant difference was found in the frequency of BRAF mutation among SPs apart from GCHP (60 % for dysplastic SPs, 44 % for SSA/Ps, 47 % for MVHPs, and 0 % for GCHPs). The frequency of CIMP was higher in dysplastic SPs or SSA/Ps than in MVHPs or GCHPs (60 % for dysplastic SPs, 56 % for SSA/Ps, 32 % for MVHPs, and 10 % for GCHPs) (SSA/Ps vs. GCHP, P = 0.0068). When serrated neoplasias (SNs) and MVHPs were classified into proximal and distal lesions, the frequency of CIMP was significantly higher in the proximal compared to the distal SNs (64 % vs. 11 %, P = 0.0032). Finally, multivariate analysis showed that proximal location and BRAF mutation were significantly associated with an increased risk of CIMP. Conclusions: Distinct molecular features were observed between proximal and distal SPs with hyperplastic crypt pattern. Proximal MVHPs may develop more frequently through SSA/Ps to CIMP cancers than distal MVHPs.

Early tumor shrinkage as a predictor of favorable outcomes in patients (pts) with advanced pancreatic cancer treated with FOLFIRINOX.

237 Background: Results from the phase III PRODIGE 4/ACCORD 11 trial provided one of current standard regimens for advanced pancreatic cancer (PC), consisting of oxaliplatin, irinotecan, fluorouracil, and leucovorin (FOLFIRINOX), which has superior response rate (RR) and survival benefit even with severe toxicity (Thierry C, et al. N Engl J Med 2011;364:1817-1825). There are several reports on the correlation between early tumor shrinkage (ETS) or depth of response (DpR) and survival in chemotherapies for colorectal cancer; however, few studies have investigated it in PC. We therefore analyzed retrospectively whether the ETS will predict outcomes in pts with PC treated with FOLFIRINOX therapy. Methods: Advanced PC pts with ECOG PS of 0 or 1, who received FOLFIRINOX as 1st- or 2nd-line treatment between November 2012 and July 2015 in 3 institutes of Showa University were included in this analysis. ETS was defined as a reduction ≥ 20% of target lesions’ diameters measured at 8 weeks from treatment start. We...

Sa1683 Five Different Subclasses of Colorectal Neoplasias Identified by Integrated Genetic and Epigenetic Analysis: Association With Polypoid and Nonpolypoid Carcinogenesis

Background: Tumor necrosis factor-α (TNF-α) is a pro-inflammatory cytokine that is highly up-regulated in inflammatory bowel disease and reduces the expression of the intestinal specific homeodomain transcription factor, CDX2. CDX2 is a master regulator of the intestinal homeostasis and is considered to be a tumor suppressor. WNT/ β-catenin signaling is critical for intestinal cell proliferation, but decreased CDX2 expression contributes to an abnormal increased activation of WNT signaling and progression of cancer. Furthermore, low CDX2 expression is associated with enhanced epithelial-to-mesenchymal transition (EMT) in colorectal cancer (CRC). Inflammation is often observed at the invasive front of the colon cancer, and it has been suggested that the inflammatory microenvironment around a tumor also induces EMT. Hence, it is likely that down-regulation of CDX2 expression in the invasive front in CRC contributes to enhanced tumorigenicity by regulating genes associated with the WNT/β-catenin pathway. Aim: To determine the role of TNFα in the regulation of CDX2 expression in CRC and its influence onWNT/β-catenin signaling. Methods: Carcinoma specimens were obtained from patients undergoing surgical resection of the rectum. Ten cases with tumor cell buddings were selected, and the expression patterns of TNFα and CDX2 at the invasive front were evaluated on immunostained slides. The mechanisms behind TNF-α-mediated down-regulation of CDX2 were investigated in vitro by selective inhibitors, and the impact of TNF-α on APC, AXIN2 and GSK3β expression were analyzed by quantitative real-time PCR (qPCR) in Caco-2 cells. Subsequently, in vivo CDX2-DNA interactions to APC, AXIN2 and GSK3β gene regulatory elements were investigated by chromatin immunoprecipitation in relation to the TNF-α status (presence vs. absence). Results: Immunohistochemical staining showed reduced CDX2 positive-cells in tumor buddings in areas with TNF-α expression in the surrounding inflammatory cells. TNF-α treatment showed a dosedependent decrease of CDX2 mRNA and protein expression dependent on p38 and NFκB, but not on JNK and ERK signaling pathways. Down-regulation of CDX2 leads to significantly decreased levels of APC (p,0.05), AXIN2 (p,0.01), and GSK3β (p,0.05) mRNA. In line with these results, TNF-α treatment impaired the ability of CDX2 to interact and activate the expression of APC (p,0.05), AXIN2 (p,0.01), and GSK3β (p,0.05). Conclusions: This study suggests that TNF-α has a tumor-promoting effect on CRC by influencing the activity of WNT signaling through down-regulation of CDX2 expression. These findings provide a novel insight into the molecular regulation of genes in the β-catenin degradation complex in response to the pro-inflammatory cytokine TNFα in cancer cells.