The association between selected risk factors for pancreatic cancer and the expression of p53 and K-ras codon 12 mutations

Background Pancreatic cancer is a major contributor to cancer mortality. Studies suggest that a few risk factors, including cigarette smoking, body mass index, having a relative with pancreatic cancer, and diabetes may be related to pancreatic cancer risk. Aim of the Study We conducted a case–control study in southeastern Michigan to examine the relation between the abovementioned risk factors and mutations of the K-ras oncogene and p53 tumor suppressor gene. Methods Two hundred forty-five patients with newly diagnosed pancreatic cancer and 420 general population controls were enrolled in the study. For this analysis, all case subjects were restricted to the pancreatic cancer patients that had tissue blocks available for study (n = 51). In-person interviews were conducted to ascertain information on demographic and lifestyle factors. Adjusted logistic regression analyses were conducted to compare various subject characteristics of pancreatic cancer patients with K-ras and p53 mutations and their subtypes to the characteristics of the general population controls. Results Smoking (adjusted odds ratio [aOR] = 2.0; 95% confidence interval [95%CI] = 0.9–4.3) and diabetes diagnosed 5 or more years before interview (aOR = 3.4; 95%CI = 1.3–8.8) were associated with pancreatic cancer patients positive for K-ras codon 12 mutations, but not with pancreatic cancer patients negative for K-ras codon 12 mutations. On the other hand, none of the examined risk factors were meaningfully related to patients with p53 mutations. Conclusions This study suggests that some recognized risk factors for pancreatic cancer may also be associated with K-ras codon 12 mutations. However, further large-scale studies are warranted.     

Frequent K-ras mutations in small bowel adenocarcinomas

The reasons for the relatively rare occurrence of small bowel adenocarcinomas when compared to the high frequency of colonic adenocarcinomas are unknown. Activating mutations in the K-ras oncogene occur in about 40% of colonic adenocarcinomas, possibly reflecting the consequences of carcinogenic exposure. To study whether the low incidence of small bowel adenocarcinomas might be due to the absence of activation of cellular oncogenes in small bowel adenocarcinomas, we examined the frequency of K-ras mutations in small bowel adenocarcinomas. K-ras mutations were determined using a polymerase chain reaction (PCR)-based method to detect codon 12 mutations by restriction fragment length polymorphism. PCR amplification was successful in six of nine small bowel adenocarcinoma samples, and revealed point mutations of K-ras at codon 12 in five of these six cases. We conclude that the small bowel might be exposed to carcinogens similar to those responsible for colorectal cancer, but may have developed protective mechanisms against cancer formation.This research was supported by an American Cancer Society Grant (Sig 13), an award from the National Foundation for Cancer Research (to I.B.W.), by the Dr. Mildred Scheel-Stiftung fuer Krebsforschung (to T.S.), and by the National Dairy Council.     

K-ras mutations in sporadic colorectal tumors in Israel: unusual high frequency of codon 13 mutations and evidence for nonhomogeneous representation of mutation subtypes

Colorectal cancer is one of the most common malignancies in the western world, including Israel. An important step in progression includes induction of activating mutations in the protooncogene K-ras. This event is very frequent and is detected in about 40% of colorectal carcinomas. Previous studies of a variety of genetic disorders revealed unique gene mutation prevalence in Jewish populations, attributed both to differences in genetic background and to variability in environmental exposure. To determine the incidence and molecular subtypes of K-ras mutations in colorectal cancer in Israel, compared with other countries, DNA was isolated from a random collection of 105 colorectal carcinoma samples, and K-ras mutations were detected by an improved designed RFLP and direct sequencing. K-ras sporadic mutations in colorectal cancer in Israel are relatively frequent, with a higher fraction in codon 13 than reported thus far. Comparison with other countries shows a vast heterogeneity in terms of the relative abundance of the affected K-ras codon and in type and representation of specific mutations. The heterogeneous distribution found may be due to a variable genetic background and different environmental factors involved in the initiation and propagation of sporadic colorectal cancer.     

K-ras mutations in patients with early colorectal cancers

Background—Published data are contradictory aboutthe importance of K-ras mutations in advanced tumours andare not available for early cancers.
Aims—To establish whether specificK-ras mutations are prognostic markers in early stagecolorectal adenocarcinoma.
Methods—The presence of K-ras exon 1 mutations were correlated with tumour recurrence in two groups ofpatients: group 1 was a consecutive series of patients with resectedcolorectal adenocarcinoma at low risk of recurrence; group 2 werepatients referred for chemotherapy after relapse of previously resectedearly stage tumours. K-ras mutations were detected bydirect sequencing of whole tissue samples in all patients and in some,the leading edge and centre of the tumour were also microdissected outindividually and sequenced.
Results—Mutations were present in 26 (26.5%) of98 patients in group 1; 14 patients developed a recurrence, four(28.5%) of whom had a K-ras mutation. Seventy ninepatients have not developed tumour recurrence, 22 (28%) of whom had amutation (p=0.84). K-ras mutations were present in five of14 patients in group 2. Microdissection did not increase the number ofmutations detected.
Conclusions—Individual K-ras genotypesare distributed homogeneously throughout early stage colorectaladenocarcinomas, but detection of a mutation has no apparent prognostic value.

Keywords:Dukes' stage; colorectal cancer; K-ras; microdissection

     

Frequency of p16(INK4A) alterations and K-ras mutations in intrahepatic cholangiocarcinoma of the liver

BACKGROUND: Inactivation of the tumour suppressor gene p16 (CDKN2/MTS-1/INK4A) and K-ras mutations are among the most frequent genetic alterations in human malignancies. AIMS: To investigate the tumour suppressor gene p16 and its possible association with K-ras mutations in intrahepatic cholangiocarcinomas of the liver. METHODS: The status of p16 was evaluated in 41 cholangiocarcinomas by methylation specific polymerase chain reaction, microsatellite analysis, DNA sequencing, and immunohistochemical staining. K-ras mutations were determined by direct DNA sequencing analyses after microdissection. The results obtained were correlated with histopathological variables and patient survival. RESULTS: Hypermethylation of the 5' CpG island of the p16 gene was found in 34 of 41 (83%) carcinomas. Homozygous deletion at the p16 region was present in two (5%), and loss of heterozygosity (LOH) in eight cases (20%). We failed to detect p16 gene missense mutations. K-ras mutations were found in 22 of 41 (54%) cholangiocarcinomas and in two cases of tumour surrounding non-neoplastic liver tissue. All 22 cancers with K-ras mutations also exhibited methylated p16. We failed to observe a correlation between K-ras or p16 status and histopathological factors or prognosis of patients. CONCLUSION: These data suggest that inactivation of the p16 gene is a frequent event in cholangiocarcinoma. The most common somatic alteration is promotor methylation of the p16 gene which is closely associated with K-ras mutations. We failed to establish p16 or K-ras status as independent prognostic factors in these tumours.     

内镜下黏膜切除术治疗Peutz-Jeghers巨大十二指肠息肉71例临床研究北大核心CSCD

目的评估内镜下黏膜切除术(EMR)治疗Peutz-Jeghers综合征(PJS)巨大十二指肠息肉的安全性及有效性。方法收集2013年2月至2020年8月在空军特色医学中心确诊为PJS十二指肠息肉并经EMR治疗的病例资料,统计EMR治疗PJS十二指肠巨大息肉的完整切除率、并发症发生率。比较巨大息肉组(直径≥3 cm)与普通息肉组(直径<3 cm)患者EMR手术完整切除率和并发症发生情况,并分析EMR治疗PJS十二指肠息肉手术并发症发生的影响因素。结果共71例患者纳入研究,男44例,女27例,中位年龄为26岁(5~58岁)。内镜下切除息肉最大中位直径为2.0 cm(0.6~13.0 cm),所有患者均成功实施EMR手术,63例患者EMR治疗PJS十二指肠息肉实现完整切除(63/71,88.7%),巨大息肉组EMR手术完整切除率低于普通息肉组(77.4%比97.5%),差异有统计学意义(P=0.023)。EMR手术相关并发症总发生率5.6%(4/71),1例患者同时存在术中创面渗血和术后胰腺炎。巨大息肉组与普通息肉组间并发症发生率(9.7%比2.5%)差异无统计学意义(P>0.05)。内镜下EMR切除十二指肠息肉有无并发症发生在患者性别、年龄、有无PJS家族史、手术史、息肉数量、切除方式上的差异均无统计学意义(P值均>0.05),而息肉位于乳头部位者并发症发生率(50%,3/6)显著高于非乳头部位者(1.5%,1/65),差异有统计学意义(P=0.001)。结论EMR治疗PJS巨大十二指肠息肉总体安全有效的,可作为PJS十二指肠息肉的首选治疗方案。息肉部位是EMR手术相关并发症发生的重要影响因素。     

Novel mutations in the STK11 gene in Thai patients with Peutz-Jeghers syndrome

Peutz-Jeghers syndrome (PJS), a rare autosomal dominant inherited disorder, is characterized by hamartomatous gastrointestinal polyps and mucocutaneous pigmentation. Patients with this syndrome have a predisposition to a variety of cancers in multiple organs. Mutations in the serine/threonine kinase 11 (STK11) gene have been identified as a major cause of PJS. Here we present the clinical and molecular findings of two unrelated Thai individuals with PJS. Mutation analysis by Polymerase Chain Reaction-sequencing of the entire coding region of STK11 revealed two potentially pathogenic mutations. One harbored a single nucleotide deletion (c.182delG) in exon 1 resulting in a frameshift leading to premature termination at codon 63 (p.Gly61AlafsX63). The other carried an in-frame 9-base-pair (bp) deletion in exon 7, c.907_915del9 (p.Ile303_Gln305del). Both deletions were de novo and have never been previously described. This study has expanded the genotypic spectrum of the STK11 gene.     

Carcinoma and polyps of the gallbladder associated with Peutz-Jeghers syndrome

Summary Peutz-Jeghers syndrome is a genetic condition characterized by mucocutaneous pigmentation and gastrointestinal polyposis. A variety of neoplasms have been found in the alimentary tract or else-where in patients with this entity. A 39-year-old female patient who had carcinoma and two polyps in the gallbladder in association with Peutz-Jeghers syndrome is described. Since the polyps consisted of the normal lining epithelium of the gallbladder and pseudopyloric gland-type metaplastic cells and obviously lacked cellular atypism, the authors would consider them hamartomatous. The carcinoma partly showing submucosal invasion existed in an area other than the polyps. This is the first documented case of the syndrome having gallbladder carcinoma.     

Novel serine/threonine kinase 11 gene mutations in Peutz-Jeghers syndrome patients and endoscopic management

AIM: To explore mutations in serine/threonine kinase 11 (STK11) gene in Peutz-Jeghers syndrome (PJS) with gastrointestinal (GI) hamartomatous polyps.METHODS: Six Japanese PJS patients in 3 families were enrolled in this study. Each of the cases had hamartomatous polyposis in the gastrointestinal tract, including the small intestine, along with mucocutaneous hyperpigmentation. Narrow-band imaging (NBI)-magnification endoscopy was employed to detect microvascular and microsurface irregularities in the GI lesions. NBI magnification findings could be classified into three groups (type A, type B, or type C). Endoscopic polypectomy was performed using double-balloon enteroscopy or colonoscopy. Genomic DNA was extracted from a whole blood sample from each subject. All of the coding exons of STK11 gene, its boundary regions, and the promoter region containing the polymorphic regions were amplified by polymerase chain reaction, and direct sequencing was performed to assess the germline mutations.RESULTS: NBI-magnification endoscopic observation could detect the abnormalities in microvessels and microsurface structures of GI polyps. Overall, we found 5 cases of type A and one case without the examination for the gastric polyps, while there were 4 cases of type B and 2 case of type A for the colorectal polyps. Seventy-nine small-bowel and 115 colorectal polyps over 27 sessions for each were resected endoscopically without significant complications. The only delayed complication included the occurrence of bleeding in a case, and this was successfully managed with hemoclips. Resected polyps contained no malignant components. Based on mutation analysis, all 3 cases in Family I exhibited the +658C>T nonsense mutation in exon 5, which resulted in the production of a truncated protein (Q220X). In Family II, a case had -252C>A and -193C>A in the promoter region. In Family III, a case was found to have the +1062C>G (F342L) mutation in exon 8.CONCLUSION: We found two novel mutations of STK11 in association with PJS. Endoscopic polypectomy of GI polyps in PJS patients appears to be useful to prevent emergency laparotomies and reduce the cancer risk.     

K-ras codon 12 mutations in Barrett's oesophagus and adenocarcinomas of the oesophagus and oesophagogastric junction

BACKGROUND: Activation of the ras oncogene is commonly found in gastrointestinal tract cancers, but the role of ras in the development and progression of Barrett's oesophagus and associated cancers is uncertain. METHODS: The frequency of K-ras codon 12 point mutations was assessed in 52 paraffin-embedded tissues from 44 patients with oesophageal pathology. The specimens were classified pathologically as follows: adenocarcinoma of the oesophagus or oesophagogastric junction (n = 23), Barrett's high-grade dysplasia (n = 5), low-grade dysplasia (n = 14), intestinal metaplasia (n = 4), normal oesophagus (n = 5) or normal stomach (n = 1). DNA was extracted from three consecutive sections of each paraffin block and mutations at bases 1 and 2 of K-ras codon 12 were identified using a novel restriction endonuclease-mediated selective polymerase chain reaction method. RESULTS: Mutations were found in 7 of 23 (30.4%) adenocarcinomas and in 2 of 5 (40%) high-grade dysplasia specimens. No mutations were found in specimens of low-grade dysplasia, intestinal metaplasia without dysplasia, or normal oesophagus and stomach. There were no significant associations between the presence of mutations and clinicopathologic features in the patients with cancer. One patient who progressed from low-grade to high-grade dysplasia was found to have developed mutant K-ras in the course of this transformation. CONCLUSION: These results suggest that K-ras codon 12 mutations may occur frequently in patients with Barrett's oesophagus with high-grade dysplasia or adenocarcinoma of the oesophagus or oesophagogastric junction. K-ras mutation may be a late event in the Barrett's metaplasia-dysplasia-adenocarcinoma sequence.     

BRAF, K-ras and BAT26 mutations in colorectal polyps and stool

AIM: To assess the feasibility of using BRAF, K-ras and BAT26 genes as stool-based molecular markers for detection of colorectal adenomas and hyperplastic polyps (HPs). METHODS: We applied PCR-SSCP and direct sequencing to detect BRAF mutations of polyps and paired stool samples. Primer-mediated restriction fragment length polymorphism (RFLP) analysis and mutant-enriched PCR were used in detection of K-ras mutations of polyp tissues and paired stool samples respectively. BAT26, a microsatellite instability marker was examined by detection of small unstable alleles in a poly (A) repeat. RESULTS: No genetic alterations were detected in the 36 colonoscopically normal patients in either tissues or stools. BRAF, K-ras and BAT26 mutations were found in 4 (16%), 10 (40%) and 3 (12%) of 25 adenoma tissues and among them, 75%, 80% and 100% of patients were observed to contain the same mutations in their corresponding stool samples. In HPs, mutations of BRAF and K-ras were detected in the tumor DNA of 2 (11.1%) and 8 (33.3%) of 18 patients respectively, all of whom had identical alterations in their stools. Taken together, the three genetic markers detected 15 (60%) of 25 adenomas and 8 (44.4%) of 18 HPs. The sensitivity of stool detection was 80% for adenomas and 100% for HPs with an overall specificity of 92% for adenomas and 100% for HPs. CONCLUSION: BRAF, K-ras and BAT26 genes have the potential to be molecular markers for colorectal adenomas and HPs, and can be used as non-invasive screening markers for colorectal polyps.     

Prognostic value of K-ras mutations and allelic imbalance on chromosome 18q in patients with resected colorectal cancer

PURPOSE: We designed this study to assess the frequency of K-ras mutations in patients with resected colorectal tumors and their association with survival. A second objective was to analyze the prognostic value of different K-ras genotypes. In a subgroup of patients we also investigated the presence of allelic imbalance on chromosome 18q and its relationship to clinical outcome. METHODS: One hundred fourteen colorectal tumors resected between 1983 and 1986 were analyzed to detect K-ras point mutations at codons 12, 13, and 61 by polymerase chain reaction followed by allele specific oligonucleotide hybridization. A subgroup of 77 tumors was further screened to detect loss of heterozygosity on chromosome 18q using three polymorphic microsatellite markers (D18S67, D18S474 andD18S58). RESULTS: K-ras mutations were detected in 29 percent (33/114) of patients. K-ras mutations correlated with age and preoperative carcinoembryonic antigen levels, and there was some indication that they may be linked to poor survival, especially in Stage II tumors, where a subgroup of patients with aspartic and serine mutations showed significantly reduced survival (P=0.03) compared with K-ras-negative patients. 18q loss of heterozygosity was present in 39 percent (25/63) of tumors. A multivariate analysis of Stage II tumors showed that 18q loss of heterozygosity was significantly associated with a worse prognosis (P=0.006). A significant decrease in survival was identified in ten patients harboring both genetic alterations (K-i mutations and 18q loss of heterozygosity;P=0.02). CONCLUSIONS: In colorectal tumors, K-ras mutations and 18q loss of heterozygosity are two genetic markers which may identify patients with more aggressive behavior, mainly in Stage II tumors. These findings warrant further research, because they can be useful in customizing adjuvant chemotherapy.     

Villous, hypermucinous mucosa in long standing ulcerative colitis shows high frequency of K-ras mutations.

BACKGROUND: K-ras mutation is one of the first genetic alterations in classical colorectal carcinogenesis. AIMS: To investigate the role of K-ras mutations in carcinogenesis, in long standing ulcerative colitis. METHODS: A total of 161 microdissected and 100 DNA samples from 13 patients were analysed for K-ras codons 12 and 13 mutations by means of a combination of enriched polymerase chain reaction amplification and temporal temperature gradient electrophoresis. RESULTS: K-ras mutations were found in 21/161 (13%) microdissected samples in 7/13 large bowels (16 and five in codons 12 and 13, respectively), and in 10/100 (10%) mucosal DNA samples (six and four, respectively). One of four patients with six adenocarcinomas had a K-ras mutation in a carcinoma, as well as one of two patients with large dysplasia associated lesion or mass (DALM). Eight of 13 (61%) areas with villous architecture and large, distended goblet cells, had a K-ras mutation, which was significantly more frequent than in low grade dysplasia (one of 23, 4%) but did not reach significance versus high grade dysplasia (four of 14, 28.5%). K-ras mutations were found in one of 20 (5%) flat lesions indefinite for dysplasia, two of 14 (14%) in non-villous, hypermucinous mucosa, and in one of 57 flat areas negative for dysplasia. CONCLUSION: The highest K-ras mutation frequency was found in villous, hypermucinous mucosa. We suggest that this entity should be investigated further as a potential risk lesion for cancer development. It may represent a pathway directly from non-classical dysplasia to cancer, not previously described.     

Detection of point mutations in K- ras gene at codon 12 in bile from percutaneous transhepatic choledochal drainage tubes for diagnosis of biliary strictures

Summary Detection of K-ras mutations at codon 12 constitutes one modality for diagnosis of pancreatic tumors. We attempted to detect K-ras mutations in DNA from bile collected through percutaneous transhepatic choledochal drainage (PTCD) tubes as a diagnostic approach to biliary strictures. Since bile salts induce cell damage, we first investigated the degeneration of cells according to bile exposure time using cell lines. High-mol-wt DNA could be extracted from cells exposed to bile for 6 h, but not from those exposed for 12 h. However, DNA exposed to bile for up 12 h could be amplified by the polymerase chain reaction (PCR) method. Therefore, K-ras mutations in fresh bile specimens collected from 15 patients through PTCD tubes were examined using PCR with restriction enzyme digestion. K-ras mutations were found in five out of five (100%) pancreatic cancers, all of which were negative according to cytodiagnosis of the same bile. On the other hand, K-ras mutations were not detected in bile from biliary tract cancers or metastatic neoplasms, except for one bile duct carcinoma and one metastatic case. Thus, although K-ras mutation alone is not an absolute marker for cancer, detection of K-ras mutations in fresh bile from PTCD tubes is a useful adjunct for diagnosis of pancreatic carcinomas in cases of biliary tract strictures.     

Clinical features,endoscopic polypectomy and STK11 gene mutation in a nine-month-old Peutz-Jeghers syndrome Chinese infant

AIM: To investigate multiple polyps in a Chinese PeutzJeghers syndrome(PJS) infant. METHODS: A nine-month-old PJS infant was admitted to our hospital for recurrent prolapsed rectal polyps for one month. The clinical characteristics, a colonoscopic image, the pathological characteristics of the polyps and X-ray images of the intestinal perforation were obtained. Serine threonine-protein kinase 11(STK11) gene analysis was also performed using a DNA sample from this infant.RESULTS: Here we describe the youngest known Chinese infant with PJS. Five polyps, including a giant polyp of approximately 4 cm × 2 cm in size, were removed from the infant's intestine. Laparotomy was performed to repair a perforation caused by pneumoperitoneum. The pathological results showed that this child had PJS. Molecular analysis of the STK11 gene further revealed a novel frameshift mutation(c.64_65het_del AT) in exon 1 in this PJS infant.CONCLUSION: The appropriate treatment method for multiple polyps in an infant must be carefully considered. Our results also show that the STK11 gene mutation is the primary cause of PJS.     

Prognostic significance of K-ras andTP53 mutations in the role of adjuvant chemotherapy on survival in patients with dukes C colon cancer

PURPOSE: Mutations in K-ras andTP53 genes are common in colorectal cancer. They affect biologic behavior and might influence chemotherapy susceptibility in these tumors. We investigated whether the survival of patients with Dukes C colon cancer treated with adjuvant chemotherapy is influenced by K-ras andTP53 mutations. METHODS: Mutation screening of the hot spots of the K-ras gene and of the evolutionarily conserved regions of theTP53 gene was performed by denaturing gradient gel electrophoresis technique in formalin-fixed paraffin-embedded specimens of 55 consecutive patients with Dukes C colon cancer treated with adjuvant 5-fluorouracil-based chemotherapy. The median follow-up was 47 (range, 32–66) months. RESULTS: Alterations in the mutation hot spots of K-ras were found at codon 12 (n=11) and 13 (n=4) in 15 of the 55 carcinomas (27 percent). No mutation was found at codon 61. Mutations of a probably causative nature in the evolutionarily conserved regions (exons 5–8) of theTP53 gene were found in 24 tumors (44 percent). K-ras andTP53 mutations were found equally in the group with recurrent disease (7/26 (26 percent) and 12/27 (44 percent), respectively) and in the group without recurrences (8/28 (24 percent) and 12/28 (43 percent), respectively). Cancer-specific survival did not differ significantly between patients with K-ras orTP53 or both mutated and nonmutated tumors, respectively (log-rank test: K-ras, P=0.72 andTP53, P=0.77; K-ras andTP53, P=0.8). Also, potentially aggressive K-ras codon 12 and 13 mutations had the same survival as tumors without these mutations (log-rank test;P=0.73). CONCLUSIONS: Patients with K-ras orTP53 or both mutated Dukes C colon tumors have the same survival as nonmutated tumors when treated with adjuvant chemotherapy. These data suggest that mutations in K-ras orTP53 alone are not prognostic indicators in patients with Dukes C colon cancer receiving adjuvant 5-Fluorouracil-based therapy.Supported by a grant form the Netherlands Cancer Foundation (NKB/KWF).Presented at the European Cancer Conference, Vienna, Austria, Sept 13 to 16, 1999.