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J Trojan

Publications and source records attributed to J Trojan.

At least 37 records · Page 2Linked to original sources
PubMed2001

DNA methylation patterns in hereditary human cancers mimic sporadic tumorigenesis.

Cancer cells have aberrant patterns of DNA methylation including hypermethylation of gene promoter CpG islands and global demethylation of the genome. Genes that cause familial cancer, as well as other genes, can be silenced by promoter hypermethylation in sporadic tumors, but the methylation of these genes in tumors from kindreds with inherited cancer syndromes has not been well characterized. Here, we examine CpG island methylation of 10 genes (hMLH1, BRCA1, APC, LKB1, CDH1, p16(INK4a), p14(ARF), MGMT, GSTP1 and RARbeta2) and 5-methylcytosine DNA content, in inherited (n = 342) and non-inherited (n = 215) breast and colorectal cancers. Our results show that singly retained alleles of germline mutated genes are never hypermethylated in inherited tumors. However, this epigenetic change is a frequent second "hit", associated with the wild-type copy of these genes in inherited tumors where both alleles are retained. Global hypomethylation was similar between sporadic and hereditary cases, but distinct differences existed in patterns of methylation at non-familial genes. This study demonstrates that hereditary cancers "mimic" the DNA methylation patterns present in the sporadic tumors.

Breast Neoplasms↗
PubMed2001

Bethesda guidelines: relation to microsatellite instability and MLH1 promoter methylation in patients with colorectal cancer.

BACKGROUND: Microsatellite instability is a hallmark of mismatch repair deficiency in hereditary nonpolyposis colorectal cancer and results from mutations in the mismatch repair genes MLH1 or MSH2 or from gene inactivation associated with DNA methylation. The Bethesda guidelines were established to identify patients with colorectal cancer who should be tested for microsatellite instability. OBJECTIVE: To assess the Bethesda guidelines for detection of microsatellite instability and to determine the role of MLH1 promoter methylation in colorectal cancer. DESIGN: Prospective cohort study. SETTING: Tertiary care referral center in Frankfurt, Germany. PATIENTS: 125 consecutive patients with colorectal cancer. MEASUREMENTS: Patients were assessed according to the Bethesda guidelines, and tumor specimens were analyzed for microsatellite instability. Patients with microsatellite instability were tested for MLH1 promoter methylation and MLH1 and MSH2 germline mutations. RESULTS: Microsatellite instability was detected in 17 of 58 patients who fulfilled and 5 of 67 patients who did not fulfill criteria of the Bethesda guidelines. In 11 of 17 patients with microsatellite instability who fulfilled Bethesda guidelines, an MLH1 (n = 3), MSH2 (n = 7), or combined MLH1 and MSH2 (n = 1) mutation was found. Among the patients with microsatellite instability who did not fulfill Bethesda guidelines, no mutations were observed; MLH1 promoter methylation was observed in 6 of 11 patients with an MLH1 or MSH2 mutation and 5 of 11 patients without an MLH1 or MSH2 mutation. CONCLUSIONS: The Bethesda guidelines are useful for selecting patients for microsatellite instability testing. MLH1 and MSH2 testing should be recommended in all patients with colorectal cancer and microsatellite instability who fulfill at least one Bethesda criterion. MLH1 promoter methylation may accompany rather than initiate carcinogenesis in patients with colorectal cancer who have mismatch repair gene defects.

Adaptor Proteins, Signal Transducing↗
PubMed2001

[Event-related functional magnetic resonance imaging of cerebral pain processing].

Neurofunctional magnetic resonance imaging (fMRI) offers the possibility to map cerebral activity non-invasively. The development of event-related techniques during the past years allows to study brain processes with high spatial and temporal resolution. Based on these techniques, EPI- and FLASH sequences were developed in this study, to investigate cerebral processing of experimental thermal pain stimulation. Phasic and tonic stimulation paradigms were developed with an MR-compatible contact thermode. Functional mapping of pain-relevant areas was performed with these paradigms, as well as a specification of the temporal characteristics of the activation. Further, a randomized paradigm with several stimulus intensities could differentiate graded functional responses, dependent on stimulus intensity in specific "regions-of-interest". In this design, randomizing the stimulus order reduced habituation effects, while continuous subjective magnitude estimation of the stimuli kept attention of subjects maximal.

Brain↗
PubMed2001

Activation of a cryptic splice site of PTEN and loss of heterozygosity in benign skin lesions in Cowden disease.

Cowden disease is an autosomal dominant syndrome characterized by facial trichilemmomas, acral keratoses, papillomatous papules, mucosal lesions, and an increased risk for breast and nonmedullary thyroid cancer. Here, we describe a novel PTEN splicing site mutation in a family with classical Cowden disease and we studied benign skin lesions typical for Cowden disease for loss of heterozygosity. We found a PTEN IVS2 + 1G > Alpha 5'-splicing acceptor mutation resulting in activation of a cryptic splice site. Activation of this cryptic splice site is predicted to result in a frameshift with a premature stop codon, thus disrupting the phosphatase core motif of PTEN. Loss of heterozygosity analysis of two trichilemmomas, one fibroma, and three acanthomas of the index patient demonstrated loss of heterozygosity at the PTEN locus in four of these lesions. In conclusion, our data demonstrate that a PTEN splicing site mutation causes activation of a cryptic splice site, which results in aberrant transcripts.

Aged↗
PubMed2001

A de novo deletion of chromosome 5q causing familial adenomatous polyposis, dysmorphic features, and mild mental retardation.

Familial adenomatous polyposis (FAP) is an autosomal dominant disorder that typically presents with colorectal cancer secondary to extensive adenomatous polyps of the colon. The molecular basis and clinical phenotype of FAP are well known. Recurrent episodes of severe abdominal pain and a positive fecal occult blood test in an 18-yr-old boy with mild mental retardation and slight dysmorphic features of the face, head, and skeletal system led to the diagnosis of FAP. The clinical workup revealed the presence of over 100 sessile colonic polyps but no polyp formation in the upper GI tract, no cancer development, nor other FAP-associated lesions. To find out whether there is an association between mental retardation and FAP we performed a chromosome analysis including comparative genomic hybridization and an indirect genotype analysis with polymorphic markers from the APC gene region. Cytogenetic analysis showed an interstitial deletion of chromosomal region 5q that was confined to the region 5q21-q22 by comparative genomic hybridization. The deletion, spanning about 10 centimorgans, encompassed the complete APC gene and can be considered as causative for FAP. Moreover, molecular genetic analysis with polymorphic markers flanking the APC gene demonstrated a de novo deletion on the paternal chromosome. Cytogenetically detectable deletions on chromosome 5 including the APC gene generally lead to an associated gene deletion syndrome. Individuals who present with mild mental retardation and dysmorphic features should therefore be investigated for chromosomal deletions. If the deletion encompasses the APC gene, these patients are at high risk of developing FAP and associated complications.

Abnormalities, Multiple↗
PubMed2001

Human glioma cells transformed by IGF-I triple helix technology show immune and apoptotic characteristics determining cell selection for gene therapy of glioblastoma.

AIMS: Insulin-like growth factor type I (IGF-I) antisense cellular gene therapy of tumours is based on the following data: rat glioma or hepatoma cells transfected with the vector encoding IGF-I antisense cDNA lose their tumorigenicity and induce a tumour specific immune response involving CD8(+) T cells. Recently, using the IGF-I triple helix approach in studies of tumorigenicity, major histocompatibility complex class I (MHC-I) antigens were demonstrated in rat glioma transfected cells. This study used comparative IGF-I antisense and triple helix technologies in human primary glioma cells to determine the triple helix strategy that would be most appropriate for the treatment of glioblastoma. METHODS: The cells were transfected using the IGF-I triple helix expression vector, pMT-AG, derived from the pMT-EP vector. pMT-AG contains a cassette comprising a 23 bp DNA fragment transcribing a third RNA strand, which forms a triple helix structure within a target region of the human IGF-I gene. Using pMT-EP, vectors encoding MHC-I or B7 antisense cDNA were also constructed. RESULTS: IGF-I triple helix transfected glioma cells are characterised by immune and apoptotic phenomena that appear to be related. The expression of MHC-I and B7 in transfected cells (analysed by flow cytometry) was accompanied by programmed cell death (detected by dUTP fluorescein terminal transferase labelling of nicked DNA and electron microscopic techniques). Cotransfection of these cells with MHC-I and B7 antisense vectors suppressed the expression of MHC-I and B7, and was associated with a pronounced decrease in apoptosis. CONCLUSION: When designing an IGF-I triple helix strategy for the treatment of human glioblastoma, the transfected tumour cells should have the following characteristics: the absence of IGF-I, the presence of both MHC-I and B7 molecules, and signs of apoptosis.

Apoptosis↗
PubMed2000

Alterations in tumorigenicity of embryonal carcinoma cells by IGF-I triple-helix induced changes in immunogenicity and apoptosis.

IGF-I antisense gene therapy has been applied successfully to animal models of glioma, hepatoma and teratocarcinoma. The antisense strategy has shown that tumor cells transfected with vectors encoding IGF-I antisense RNA lose tumorigenicity, become immunogenic and are associated with tumor specific immune response involving CD8+ lymphocytes. An IGF-I triple helix approach to gene therapy for glioma was recently described. The approach we have taken is to establish parameters of change using the IGF-I triple helix strategy. PCC-3 embryonal carcinoma cells derived from murine teratocarcinoma which express IGF-I were used as a model. The cells were transfected with vector which encodes an oligoribonucleotide that forms RNA-IGF-I DNA triple-helix structure. The triple-helix stops the production of IGF-I. Cells transfected in this manner underwent changes in phenotype and an increase in MHC-I and B-7 cell surface molecules. They also showed enhancement in the production of apoptotic cells (60-70%). The "triple helix" transfected cells lost the ability to induce tumor when injected subcutaneously in syngeneic 129 Sv mice. When co-transfected in vitro with expression vectors encoding both MHC-I and B-7 cDNA in antisense orientation, the "triple-helix" transfected cells were down-regulated in expression of MHC-I and B-7 and the number of apoptotic cells was significantly decreased. Injection of the doubly co-transfected cells into 129 Sv mice was associated with induction of teratocarcinoma. Comparison between antisense and triple-helix transfected cells strategies showed similar immunogenic and apoptotic changes. The findings suggest that triple-helix technology may offer a new clinical approach to treatement of tumors expressing IGF-I.

Animals↗
PubMed2000

Expression of insulin-like growth factor-I in rat glioma cells is associated with change in both immunogenicity and apoptosis.

Insulin-like growth factor I (IGF-I), has a role in cellular differentiation and is also expressed in neoplastic transformation of glioma cells. We recently demonstrated inhibition in expression of cellular IGF-I after transfection with vectors that incodes a segment of the human IGF-I RNA in antisense orientation. The transfected cells expressed increased levels of both MHC-I and B7 molecules. In this paper we show that IGF-I antisense transfected cells also become apoptotic. Moreover, the phenomenon of programmed cell death is related to the phenomenon that results in increased expression of MHC-I and B7 molecules. Co-transfection of rat glioma cells with the vector expressing IGF-I antisense RNA and with vectors encoding the expression of MHC-I and B7 antisense cDNA suppressed the expression of both of these molecules and was associated with a decrease in apoptosis.

Animals↗
PubMed2000

Extraintestinal polyps in Peutz-Jeghers syndrome: presentation of four cases and review of the literature. Deutsche Peutz-Jeghers-Studiengruppe.

Peutz-Jeghers syndrome (PJS) is a rare hereditary disorder characterized by hamartomatous polyps in the gastrointestinal tract and typical pigment lesions. Extraintestinal polyps have rarely been reported. Possible sites include the respiratory tract, urogenital tract, and gallbladder. We here describe four cases of extraintestinal polyps in PJS patients and review the literature on the need for operative therapy of extraintestinal polyps in PJS. Three nonrelated patients were examined who had PJS and polyps in the gallbladder; the fourth patient had PJS and recurrent choanal polyps. Surgery has so far been performed only for symptomatic polyps: one laparoscopic cholecystectomy and removal of the choanal polyps for recurrent infections of the respiratory tract. The remaining two patients reported no symptoms from the extraintestinal polyps. No malignant transformation was found in these patients, nor has such been reported in the literature on PJS. The frequent observation of this manifestation in our patients raises the question of clinical management: Is prophylactic surgery indicated? Since malignant transformation of PJS polyps in the intestine is extremely rare we see no reason for operative therapy as long as the polyps are small and asymptomatic. Regular sonographic controls are recommended since the risk of malignant transformation cannot be ruled out at present.

Adult↗
PubMed2000

Enhancement of immunogenicity of tumor cells by cotransfection with genes encoding antisense insulin-like growth factor-1 and B7.1 molecules.

Insulin-like growth factor-1 (IGF-1) is expressed in many tumor cell lines and has a role in both normal cell proliferation and in the growth of cancers. Tumor cells transfected with a vector encoding an IGF-1 antisense cDNA transcriptional cassette driven by the mouse metallothionein-1 promoter become immunogenic and lose their tumorigenicity in syngeneic animals. The enhanced immunogenicity is associated with an up-regulation in the expression of major histocompatibility complex class I molecule on cell surfaces. Blockade of the expression of IGF-1 in tumor cells by the IGF-1 antisense RNA approach is not uniformly effective in the induction of antitumoral protective immunity in low and nonimmunogenic tumor model systems. Here, we report that the immunogenicity of hepa 1-6 hepatoma and SMCC-1 colon carcinoma cells, which are poorly immunogenic and unresponsive to antisense IGF-1 gene transfer, can be induced by cotransfection with genes encoding antisense IGF-1 and mouse B7.1 molecules. The tumor cells modified in this manner become strongly immunogenic and can be used as a cellular vaccine to induce a protective immune response in vivo. Immunization with the transfected tumor cells also results in regression of the established hepa 1-6 hepatoma and SMCC-1 colon cancer. The immunity is tumor-specific and is mediated by CD3+ CD8+ T cells. Cytotoxic T lymphocytes generated in vitro by priming naive spleen cells and in vivo by immunizing mice with the double-transfected tumor cells specifically lysed autologous tumors cells and were effective in adoptive immunotherapy. The data suggest that modification of tumor cells in vitro by cotransfection with genes encoding antisense IGF-1 and B7.1 molecules may open a new avenue for cancer immunogene therapy.

Adjuvants, Immunologic↗
PubMed2000

5'-CpG island methylation of the LKB1/STK11 promoter and allelic loss at chromosome 19p13.3 in sporadic colorectal cancer.

BACKGROUND: In patients with Peutz-Jeghers syndrome (PJS), causative germline mutations in the LKB1/STK11 gene on chromosome 19p13.3 have been identified. Because of the loss of heterozygosity (LOH) at 19p13.3 in hamartomas and the cancer susceptibility of patients with PJS, LKB1/STK11 is suggested to act as a tumour suppressor. However, the frequency of genetic and epigenetic inactivation of LKB1/STK11 in sporadic tumours is unclear. AIMS: To investigate the LKB1/STK11 gene for promoter hypermethylation and allelic loss in tumour specimens of patients with sporadic colorectal cancer. METHODS: DNA from 50 consecutive paraffin embedded sporadic colorectal adenocarcinomas and corresponding normal epithelium was extracted. After bisulphite treatment, specimens were analysed for methylation of the LKB1/STK11 promoter 5'-CpG island by methylation specific polymerase chain reaction (MSP). In addition, tumours were analysed for LOH of chromosome 19p13.3. In tumours exhibiting LOH, LKB1/STK11 was sequenced. RESULTS: MSP was successful in 48 of 50 tumour specimens. Of those, four (8%) demonstrated hypermethylation of the LKB1/STK11 promoter 5'-CpG island. Moreover, LOH at either D19S886 or D19S878 was observed in five of 38 (13%) informative tumours. All five tumours showing LOH at 19p13.3 were advanced and four of five were located in the left sided colon. There was no correlation between LOH and LKB1/STK11 promoter hypermethylation or somatic mutation. CONCLUSIONS: In sporadic colorectal cancer, hypermethylation of the LKB1/STK11 promoter and allelic loss at the STK 11 gene locus are rare events. LOH at 19p13.3 was associated with advanced tumour stage and left sided location but not with LKB1/STK11 promoter hypermethylation or somatic mutation.

Adenocarcinoma↗
PubMed2000

Thyrotropin-releasing hormone time-dependently influences thyrotropin microheterogeneity--an in vivo study in euthyroidism.

Thyrotropin (TSH) is secreted not as one distinct hormone, but rather as a group of isohormones which differ in their oligosaccharide composition. Although the mechanisms regulating TSH glycosylation are not fully understood, there is strong evidence that TRH plays an important role. The aim of our study was to determine the dynamic influence of TRH on TSH microheterogeneity. Sera were obtained from euthyroid volunteers (n=20) before and 30, 60, 120, 180 and 240 min after intravenous, nasal and oral administration of TRH in three independent runs (randomized order, at a time-interval of 3 weeks between each run). TSH was immuno-concentrated and analysed by isoelectric focusing (IEF) and lentil lectin affinity chromatography. TSH immunoreactivity was measured by an automated second-generation TSH immunoassay. Overall, serum TSH concentrations reached maximal values 30 min after intravenous, 60 min after nasal and 180 min after oral TRH stimulation. IEF analysis revealed 63.3+/-3.3% of pituitary standard TSH (IRP 80/558) in the neutral pH range (8>pH>6). In contrast, 30 min after TRH stimulation 80.8+/-3.7% (P<0.001) and 60 min after TRH stimulation 44.9+/-2.2% (P<0.001) of the TSH of euthyroid probands were found in this pH range, whereas 180 min after TRH stimulation 58.4+/-2.3% (P<0.001) were detected in the acidic pH range (pH<6). This shift of TSH composition in euthyroidism after TRH stimulation was confirmed by lentil lectin analysis of TSH: core-fucose content of euthyroid TSH was 73.4+/-3.8% 30 min and 22.9+/-3.2% 120 min after TRH stimulation in contrast to basal (53.3+/-1.8%; P<0.001) and pituitary standard (IRP 80/558) TSH (63.0+/-0.9%; P<0.001). In conclusion, in euthyroidism, TRH stimulation time-dependently changes the distribution pattern of the TSH isoforms from an alkaline and neutral to a more acidic one. This corresponds to the secretion of isohormones with altered bioactivity which could influence the fine-tuning of thyroid function.

Administration, Intranasal↗
PubMed1999

Peutz-Jeghers syndrome: molecular analysis of a three-generation kindred with a novel defect in the serine threonine kinase gene STK11.

The Peutz-Jeghers syndrome, phenotypically characterized by mucocutaneous pigmentation and hamartomatous polyposis, is an autosomal dominant disease with variable expression and incomplete penetrance. Moreover, affected patients are at increased risk for gastrointestinal and other malignancies. Recently, a mutated gene encoding abnormal forms of the novel serine threonine kinase STK11 has been identified as a genetic cause of Peutz-Jeghers syndrome. Here, we report the molecular analysis of the STK11 gene in a patient with Peutz-Jeghers syndrome, which in exon 1 revealed a guanine (G) insertion in the 5 G repeat of codons 51-53. The insertion leads to a frameshift with a premature TGA stop codon 324 bp downstream in codon 162, predicting the expression of a truncated protein without kinase activity. This heterozygous germline mutation was also found in the affected father and in one affected sister of the index patient, but not in any phenotypically unaffected family member or in unrelated control subjects. In DNA isolated from microdissected hamartomatous polyps of the index patient, exon 1 of the STK11 gene could not be amplified suggesting that both alleles of STK11 exon 1 were lost in the hamartomatous polyps. Identification of a STK11 gene mutation in an index patient offers the possibility of a predictive diagnosis, and initiation of specific screening programs in the genetically affected kindred.

Adult↗
PubMed1999

Fluorine-18 FDG positron emission tomography for imaging of hepatocellular carcinoma.

OBJECTIVE: The detection of increased fluorine-18 fluorodeoxyglucose (18F-FDG) uptake by positron emission tomography (PET) is based on the enhanced glucose metabolism of tumor cells. Because the detection and staging of hepatocellular carcinoma (HCC) in patients with liver cirrhosis can be difficult, we prospectively evaluated the sensitivity of 18F-FDG PET in 14 consecutive patients with HCC. METHODS: Whole body and regional 18F-FDG PET of the liver were obtained. The results were compared with ultrasonography, contrast-enhanced, helical CT, histological grading, p53 protein expression of HCC, and serum alpha-fetoprotein (AFP) level. RESULTS: In 7 patients PET demonstrated increased tumor 18F-FDG uptake, whereas HCC was not distinguishable from nonmalignant liver tissue in 7 other patients. Hepatic lesions were detected by ultrasonography in all patients, whereas only 11 of 14 HCCs could be identified by CT. In 3 patients extrahepatic spread was demonstrated by 18F-FDG PET. Patients with increased tumor 18F-FDG uptake had significantly larger hepatic lesions and higher serum AFP levels than those with normal 18F-FDG uptake. Lesions could be visualized by 18F-FDG PET in 7 of 8 patients with moderately or poorly differentiated HCC, whereas none of the six well-differentiated tumors was detected. Two patients with strong p53 expression demonstrated increased tumor 18F-FDG uptake and extrahepatic metastases. CONCLUSIONS: The sensitivity of 18F-FDG PET for the imaging of HCC is low. Nevertheless, in patients with moderately or poorly differentiated HCC, tumors >5 cm, or with markedly elevated AFP levels 18F-FDG PET may contribute to an effective noninvasive staging.

Adult↗
PubMed1999

Evaluation of rapid microsatellite analysis of paraffin-embedded specimens in screening for hereditary nonpolyposis colorectal cancer.

Length alterations in short repetitive DNA sequences, termed microsatellite instability (MSI), are used as a diagnostic criterion of replication errors caused by various mutations in at least five mismatch repair genes. Therefore, MSI analysis is useful in clinical practice to identify patients with hereditary nonpolyposis colorectal cancer (HNPCC). MSI can be detected by amplification of microsatellite loci in DNA extracted from paraffin-embedded tumor and corresponding peritumoral specimens after numerous time consuming steps limiting the clinical utilities. Rapid microsatellite analysis, a efficient and rapid DNA extraction technique based on Triton X-100 preincubation, was compared with the conventional DNA extraction for HNPCC screening in colorectal tumor specimens from 12 patients. Five complex and two noncomplex (CA)n microsatellite loci were tested, with use of multicolor fluorescent analysis. MSI and loss of heterozygosity in colorectal tumor samples could equally be assessed with the two DNA preparation methods, whereas the number of initially unsuccessful DNA extractions from paraffin-embedded tissue specimens and overall duration for MSI analysis were significantly reduced when rapid microsatellite analysis was used. A replication error-positive phenotype was detected in 2 of 10 patients with a positive family history for colorectal cancer, and diagnosis of HNPCC was finally confirmed by detection of a specific germline mutation. The described rapid microsatellite analysis is less time consuming and more efficient, and, in general, it reduces the risk of contamination by limiting the number of steps required. Therefore, it might replace current DNA extraction procedures. Furthermore, techniques using fluorescent polymerase chain reaction and semiautomated DNA sequencer allow for precise, observer-independent, and rapid scoring in MSI and loss of heterozygosity assessment. A combination of our rapid DNA extraction method and the use of a highly specific microsatellite marker might improve replication error analysis in HNPCC screening.

Adult↗