Livia Theodor

Detection of Pancreatic Carcinoma (Diagnostic Value of K-ras Mutations in Circulating DNA from Serum)

Somatic activating mutations at codon 12 of theK-ras gene are present in the majority of exocrinepancreatic cancers and occur early in tumorgenesis. Theaim of this study was to test the feasibility of using a mutated K-ras gene from the serum as apotential tumor marker for detection of exocrinepancreatic carcinoma. Codon 12 K-ras mutations wereexamined in DNA extracted from the sera of 20 patients with pancreatic carcinomas, six patients withchronic pancreatitis, and five healthy individuals.K-ras gene mutations at codon 12 were detected in thesera of 14 of 20 patients with pancreatic carcinoma and in none of the six patients with chronicpancreatitis, or in the five healthy controls. Elevationof either CA19-9 or K-ras mutation was detected in 19/20patients. These results suggest that K-ras abnormalities in serum could be used as apotential tumor marker in patients with a pancreaticlesion. The absence of K-ras mutations in serum andpresence of CA19-9 in the normal range make thediagnosis of pancreatic cancer unlikely.

The I1307K APC polymorphism: prevalence in non-Ashkenazi Jews and evidence for a founder effect.

A missense mutation within the APC gene, I1307K, was described in Ashkenazi individuals at risk for colorectal cancer (CRC) and in the general population. The anecdotal reporting of the occurrence of this mutation in some non-Ashkenazi individuals led us to hypothesize that within the Jewish people, the I1307K polymorphism may reflect a founder mutation, and that the mutation is not restricted to ethnic Ashkenazis. To test that notion, and to establish the occurrence rate of the I1307K polymorphism in non-Ashkenazi Jewish populations, we screened Iraqi and Moroccan Jews and consecutive Jewish CRC patients and performed haplotype analysis with APC-linked markers in two I1307K carrier families. We analyzed Jewish individuals: 210 Moroccans, 160 Iraqis, 148 Ashkenazi, and 349 CRC patients (227 Ashkenazi and 122 non-Ashkenazi). The mutation detection scheme included PCR followed by denaturing gradient gel electrophoresis (DGGE) or modified restriction analysis (MRA). Haplotypes were assessed using three intragenic and three flanking markers. The I1307K polymorphism was detected in 29/227 Ashkenazi (12.8%), 2/122 (1.6%) non-Ashkenazi CRC patients, and in 2 individuals each (approximately 1%) within the Moroccan and Iraqi populations. Allelic pattern analysis in all our I1307K carriers, revealed a common haplotype for the three intragenic markers tested, in all mutation carriers, regardless of ethnic origin. The I1307K polymorphism, therefore, exists in all ethnic Jewish populations: Ashkenazi and non-Ashkenazi, with or without colon cancer. Jewish I1307K mutation carriers share a common allelic pattern with APC-linked markers. This strongly supports the notion of a founder mutation for I1307K.

The frequency of the predominant Jewish mutations in BRCA1 and BRCA2 in unselected Ashkenazi colorectal cancer patients

It is presently unclear whether carriers of BRCA1 mutations have an increased risk for colorectal cancer (CRC). To gain insight into this issue, 225 unselected Ashkenazi Jewish CRC patients were tested for the presence of the three common Jewish BRCA1/2 germline mutations: 185delAG and 5382insC (BRCA1) and 6174delT (BRCA2). A total of four carriers was found (4/225, 1.78%). This frequency is similar to the estimated normal Ashkenazi population frequency, thus suggesting that these specific mutations do not contribute to CRC predisposition.

Common origin of the I1307K APC polymorphism in Ashkenazi and non-Ashkenazi Jews.

A common germline missense mutation within the APC gene, I1307K, has recently been described in Ashkenazi Jews. We detected this polymorphism in two non-Ashkenazi Jewish women using denaturing gradient gel electrophoresis (DGGE), and hypothesized that in Jewish individuals it might not be restricted to Ashkenazim, and actually reflect a common ancestral polymorphism. To test this notion we performed allelic pattern determination using APC-linked markers in these two women and in nine Ashkenazi carrier controls. The pattern of the intragenic markers, as well as a single downstream marker 30–70 Kb from the APC gene was identical in all individuals, regardless of ethnic origin. We conclude that the I1307K polymorphism in Jewish individuals, is not restricted to Ashkenazim and probably reflects a founder mutation.

Diagnostic Value of K-ras Mutations in Serum of Pancreatic Cancer Patients

The diagnosis of pancreatic carcinoma is sometimes difficult. Distinguishing pancreatic carcinoma from chronic pancreatitis is important and of obvious clinical implication. However, in some patients, neither the vague clinical presentation nor the current imaging modalities have clear-cut distinctive features.1,2 A variety of tumor markers have been tested in the diagnostic workup of a pancreatic lesion, yet none has been proven as the ultimate one.3,4 Therefore, an accurate and readily accessible test to differentiate pancreatic cancer from chronic pancreatitis is highly warranted. K-ras oncogene is frequently mutated in pancreatic carcinomas, and several studies have reported its presence in the pancreatic tissue of 75–95% of human pancreatic adenocarcinoma. The mutation is at codon 12 of the K-ras oncogene and is considered as a critical and early event in pancreatic oncogenesis.5–9 High rates of codon 12 K-ras mutations have been detected in DNA obtained by fine-needle aspiration from pancreatic masses,10–14 as well as from duodenal or pure pancreatic juice15–19 collected from patients with pancreatic cancer. However, K-ras mutations were also detected in a significant proportion of patients with chronic pancreatitis,20–24 markedly reducing the specificity of mutated K-ras gene as a potential tumor marker. A potential noninvasive source of DNA in pancreatic cancer patients is the serum. Circulating serum DNA levels were found to be elevated in patients with gastrointestinal malignancies and, in particular, in patients with pancreatic carcinoma.25–27 Indeed, K-ras mutations were demonstrated in plasma from 3/3 patients with pancreatic cancer.28 The aim of this study was to test the feasibility of using mutated K-ras gene from the serum as a potential tumor marker for detection of exocrine pancreatic carcinoma. In addition, the diagnostic value of both CA19-9 and K-ras mutations in serum was further investigated.

Serum CA 19-9 levels as a diagnostic marker in cystic fibrosis patients with borderline sweat tests

Abstract.Patients with normal or borderline sweat tests present a diagnostic challenge. In spite of the availability of genetic analysis and measurement of nasal potential difference, there is still uncertainty in diagnosing cystic fibrosis in some patients. CA 19–9 is a tumor-associated antigen whose levels were previously found to be elevated in some cystic fibrosis patients. We investigated whether serum CA 19–9 levels can contribute to establishing the diagnosis of cystic fibrosis in patients with a borderline sweat test, and evaluated the influence of different clinical variables on CA 19–9 levels. Serum CA 19–9 levels were measured in 82 cystic fibrosis patients grouped according to their genotype and in 38 healthy individuals. Group A included 50 patients who carried two mutations previously found to be associated with a pathological sweat test and pancreatic insufficiency (ΔF508, W1282X, G542X, N1303K, and S549R). Group B included 13 compound heterozygote cystic fibrosis patients who carried one mutation known to cause mild disease with a borderline or normal sweat test and pancreatic sufficiency (3849+10kb C→T, 5T). Group C included 38 normal controls. Nineteen cystic fibrosis patients carried at least one unidentified mutation. An association between CA 19–9 levels and age, pulmonary function, pancreatic status, sweat chloride, previous pancreatitis, serum lipase, meconium ileus, distal intestinal obstruction, liver disease, and diabetes was investigated. The distribution of CA 19–9 levels was significantly different between the three groups (p<0.01); high CA 19–9 levels were found in 60% (30/50) of group Apatients and in 46.6% (6/13) of group B patients, but in only 5.2% (2/38) of the controls. CA 19–9 levels were inversely related to forced expiratory volume in 1 s, while no association was found with the other clinical parameters examined. Our findings suggest that the serum CA 19–9 in cystic fibrosis patients originates in the respiratory system, and has a useful ancillary role, particularly when diagnostic uncertainty exists. Hence, the diagnosis of cystic fibrosis should be considered in patients with borderline sweat tests and high CA 19–9 levels, but normal levels do not exclude cystic fibrosis.

Immunohistochemical analyses of colon cancer in I1307K APC mutation carriers compared with noncarriers

The I1307K APC germline mutation is associated with an increased risk to colorectal cancer (CRC). Whether and to what extent the somatic features and the molecular pathways of cancer development in mutation carriers differ from colorectal cancer in noncarriers remains unknown. To gain insight into this issue, 52 Israeli patients with CRC, 24 of whom were I1307K APC mutation carriers, were analyzed. The expression pattern of genes known to be involved in the pathogenesis of sporadic CRC was assessed immunohistochemically: E-cadherin, ß-catenin, deleted in colon cancer (DCC), and p53. In addition, tumors were genotyped for somatic activating mutations in Ki-rasoncogene. Mutation carriers and noncarriers were comparable in age at diagnosis (64.3 ± 10.1 years for carriers and 60.8 ± 14.1 years for noncarriers), tumor location in the colon, and disease stages. Tumors of I1307K mutation carriers displayed positive p53 immunostaining and loss of ß-catenin, E-cadherin, and DCC expression more often compared with noncarriers, although none of these differences reached statistical significance. Mutation frequencies in the Ki-ras gene were similar in both groups. In conclusion, the molecular pathways in CRC in I1307K APC mutation carriers are seemingly similar to those of sporadic cases, but a larger study is clearly needed.