Sang Geol Kim

Frequent CpG Island Methylation in Serrated Adenomas of the Colorectum

Serrated adenomas are characterized by a saw-toothed growth pattern with epithelial dysplasia (intraepithelial neoplasia). The CpG island methylator phenotype (CIMP) is a recently described mechanism for tumorigenesis in colorectal carcinomas and adenomas characterized by methylation of multiple CpG islands. The role of these epigenetic alterations in the pathogenesis of serrated adenomas is not clear. We therefore evaluated CIMP in 22 sporadic serrated adenomas and 6 serrated adenomas with multiple (6 to 10) hyperplastic polyps, including 5 with admixed hyperplastic glands and adenomatous glands, and compared the results with 34 conventional adenomas. Bisulfite methylation-specific polymerase chain reaction was used for the p16 and hMLH1 genes, and three MINT (methylated in tumor) loci (MINT1, MINT2, and MINT31). Patients with sporadic serrated adenomas had a higher frequency of hyperplastic polyps (1.3 ± 1.6) as compared to patients with tubular adenomas (0.4 ± 0.9, P = 0.02). Mean number of methylated sites was significantly higher in sporadic serrated adenomas (2.0 ± 1.7) than in tubular adenomas (0.8 ± 0.9, P = 0.00001). Sporadic serrated adenomas had significantly more frequent methylation of MINT1 (48%, 10 of 22) and MINT2 (71%, 15 of 21) than tubular adenomas (9%, 3 of 34, P = 0.001; and 18%, 6 of 34, P = 0.0001), respectively. Concordant methylation of two or more sites (CIMP-high) was also more frequent in sporadic serrated adenomas (68%, 15 of 22) than in tubular adenomas (18%, 6 of 34, P = 0.0005). All five serrated adenomas with admixed hyperplastic glands and adenomatous glands were CIMP-high. Our results indicate that CpG island methylation is common in sporadic serrated adenomas and may play an important role in their pathogenesis.

CpG island methylation in carcinoid and pancreatic endocrine tumors

Carcinoid tumors and pancreatic endocrine tumors (PETs) are uncommon neuroendocrine neoplasms and their genetic alterations are not well characterized. CpG island methylation is a mechanism of gene silencing, and concordant methylation of multiple CpG islands as CpG island methylator phenotype (CIMP) has been described in tumors. The aim of this study was to evaluate CIMP in carcinoid tumors and PETs. We studied 16 carcinoid tumors, 11 PETs, and 22 associated normal mucosa or pancreas. Methylation status of the p14, p16, cyclo-oxygenase 2 (COX2), O6-methyl-guanine methyltransferase (MGMT), estrogen receptor (ER), thrombospondin 1 (THBS1), retinoic acid receptor beta 2 (RARβ), T-type calcium channel (CACNA1G), and multiple endocrine neoplasia type-1 (MEN1) genes, and of MINT1, MINT2, MINT25, MINT27 and MINT31 loci was evaluated by methylation-specific-PCR (MSP) or combined bisulfite restriction analysis (COBRA). Carcinoid tumors were frequently methylated at RARβ, MGMT, p16, COX2, p14, THBS1, and ER ranging from 25 to 63% of tumors. Other CpG islands were infrequently methylated or unmethylated. The adjoining normal mucosa was also methylated for ER, COX2, and RARβ, but methylation at p14, p16, THBS1, and MGMT was tumor-specific. By contrast, PETs and normal pancreas were frequently methylated only at ER. Methylation was more frequent in carcinoid tumors than PETs at MGMT (25 versus 0%, p=0.03), THBS1 (44 versus 9%, p=0.04), p14 (44 versus 9%, p=0.04) and RARβ (25 versus 0%, p=0.03). Loss of p16 protein expression correlated with methylation of p16 gene in carcinoid tumors (p=0.006). Our study indicates that methylation profile of carcinoid tumors differs from PETs, reflecting different molecular pathogenesis.

Comparison of Epigenetic and Genetic Alterations in Mucinous Cystic Neoplasm and Serous Microcystic Adenoma of Pancreas

Mucinous cystic neoplasms and serous microcystic adenomas account for the majority of cystic tumors of pancreas. Mucinous cystic neoplasms and serous microcystic adenomas have different frequencies of progression to malignancy. The genetic and epigenetic alterations of these tumors have not been studied in detail. In this study, we compared methylation status of p16, p14, VHL, and ppENK genes by methylation-specific PCR (MSP), and genetic alterations including K-ras and β-catenin gene mutations, chromosome 3p loss, and microsatellite instability in 15 mucinous cystic neoplasms (10 benign and 5 borderline) and 16 serous microcystic adenomas. There were no significant differences between mucinous cystic neoplasms and serous microcystic adenomas in methylation of p16 (14%, 2/14 and 12%, 2/16), p14 (15%, 2/13 and 37%, 6/16), VHL (0/14 and 7%, 1/14), and ppENK (0/14 and 0/13), respectively. K-ras mutation was present only in mucinous cystic neoplasms but not in serous microcystic adenomas (33%, 5/15 versus 0/16; P = .004). In addition, LOH at 3p25, the chromosomal location of VHL gene, was present in 57% (8/14) of serous microcystic adenomas compared with in 17% (2/12) of mucinous cystic neoplasms (P = .03). No β-catenin mutation, microsatellite instability, or mutation of transforming growth factor β type II receptor was present in either type of tumors. In conclusion, K-ras mutations and allelic loss of VHL locus at 3p25, but not methylation, distinguished mucinous cystic neoplasms and serous microcystic adenomas. The differences in genetic alterations but not epigenetic alterations may explain the pathogenesis and progression to malignancy of these cystic tumors of pancreas.

Epigenetic and Genetic Alterations in Duodenal Carcinomas Are Distinct From Biliary and Ampullary Carcinomas

BACKGROUND & AIMS Carcinomas of the extrahepatic bile ducts, ampulla of Vater, and duodenum are uncommon, and their epigenetic and genetic alterations are not well characterized. METHODS We therefore compared the methylation profile and genetic alterations in 18 extrahepatic biliary, 9 ampullary, and 12 duodenal carcinomas. We evaluated methylation at p16, p14, and human Mut L homologue (hMLH1) by methylation- specific PCR (MSP), and at cyclooxygenase 2 (COX2), O(6)-methyl-guanine methyltransferase (MGMT), estrogen receptor (ER), retinoic acid receptor beta 2 (RAR beta), and T-type calcium channel (CACNA1G) genes, and methylated in tumor 1 (MINT1), MINT2, MINT25, MINT27, and MINT31 loci by combined bisulfite restriction analysis (COBRA); mutation of K-ras, p53, p16, and p14 genes by sequencing; loss of heterozygosity of chromosome 9p; and microsatellite instability (MSI). RESULTS Duodenal carcinomas were methylated more frequently or had increased methylation densities than biliary carcinomas at p14 (P = 0.04), hMLH1 (P = 0.04), MGMT (P = 0.01), MINT1 (P = 0.01), MINT25 (P = 0.01), MINT27 (P = 0.001), RAR beta (P = 0.03), and ER (P = 0.001), and than ampullary carcinomas at RAR beta (P = 0.02) and ER (P = 0.03). In contrast, the methylation profiles of biliary and ampullary carcinomas were not statistically different. Simultaneous methylation of 3 or more CpG islands (CpG island methylator phenotype-high) was more common in duodenal cancers (P = 0.004). MGMT methylation was associated with G-to-A mutation in K-ras (P = 0.006), and hMLH1 methylation was associated with MSI-high (P = 0.01). CONCLUSIONS Our findings indicate that the methylation profile and genetic alterations of duodenal carcinomas are distinct from biliary and ampullary carcinomas, and that tumor-specific methylation is associated with gene mutation and MSI.

Totally laparoscopic associating liver partition and portal vein ligation for staged hepatectomy using anterior approach in HCC patient with Type II portal vein anomaly

Associating liver partition and portal vein ligation for staged hepatectomy (ALPPS) has gradually developed because of rapid hypertrophy of the future liver remnant volume (FLR) in spite of high morbidity. To minimize the patients postoperative pain and morbidity including wound complication caused by two consecutive major abdominal operations, we adopted a totally laparoscopic approach and used a composite mesh graft. Also, to maximize the oncologic efficacy, we adopted the “anterior approach” technique. A 44-year-old woman with large hepatitis B-related hepatocellular carcinoma (HCC) in her right lobe was transferred to our hospital for surgical treatment. Preoperatively predicted FLR by a CT scan was 21% and type II portal vein anomaly was detected. A totally laparoscopic approach was planned. During the first stage operation, right anterior and posterior portal veins were meticulously dissected and tied. After parenchymal transection by the “anterior approach” technique, two glissonian pedicles of the right liver were individually isolated. A composite mesh graft was used to prevent severe adhesion on both liver partition surfaces. During the second-stage operation, 9 days after the first stage operation, the two isolated glissonian pedicles were initially transected. After full mobilization of the right lobe, the right hepatic vein was also transected. The right lobe was removed through the Pfannenstiel incision. She was discharged 7 days after the second stage operation. Her postoperative course was uneventful and there was no HCC recurrence for 15 months after hepatectomy. A totally laparoscopic ALPPS procedure can be a feasible technique that ensures patient safety and oncologic superiority, even in patients with complicated anatomical variation.