Neal K. Osborn

Prospective evaluation of methylated SEPT9 in plasma for detection of asymptomatic colorectal cancer

Background As screening methods for colorectal cancer (CRC) are limited by uptake and adherence, further options are sought. A blood test might increase both, but none has yet been tested in a screening setting. Objective We prospectively assessed the accuracy of circulating methylated SEPT9 DNA (mSEPT9) for detecting CRC in a screening population. Design Asymptomatic individuals ≥50 years old scheduled for screening colonoscopy at 32 US and German clinics voluntarily gave blood plasma samples before colon preparation. Using a commercially available assay, three independent blinded laboratories assayed plasma DNA of all CRC cases and a stratified random sample of other subjects in duplicate real time PCRs. The primary outcomes measures were standardised for overall sensitivity and specificity estimates. Results 7941 men (45%) and women (55%), mean age 60 years, enrolled. Results from 53 CRC cases and from 1457 subjects without CRC yielded a standardised sensitivity of 48.2% (95% CI 32.4% to 63.6%; crude rate 50.9%); for CRC stages I–IV, values were 35.0%, 63.0%, 46.0% and 77.4%, respectively. Specificity was 91.5% (95% CI 89.7% to 93.1%; crude rate 91.4%). Sensitivity for advanced adenomas was low (11.2%). Conclusions Our study using the blood based mSEPT9 test showed that CRC signal in blood can be detected in asymptomatic average risk individuals undergoing screening. However, the utility of the test for population screening for CRC will require improved sensitivity for detection of early cancers and advanced adenomas. Clinical Trial Registration Number: NCT00855348

Aberrant methylation of secreted frizzled-related protein genes in esophageal adenocarcinoma and Barrett's esophagus.

Hypermethylation of secreted frizzled‐related proteins (SFRP) genes frequently occurs with several cancers but has not been studied in esophageal adenocarcinoma or its precursor—Barretts esophagus. To explore the role of SFRP methylation in the neoplastic progression of Barretts esophagus and to evaluate methylated SFRP genes as biomarkers for Barretts esophagus and cancer, methylation of SFRP genes was determined in esophageal adenocarcinomas, Barretts esophagus and normal epithelia using methylation‐specific PCR. Protein expression of SFRP genes was then assessed in these tissues by immunohistochemistry. The mRNA expression of SFRP genes was quantified by real‐time reverse‐transcription PCR in esophageal adenocarcinoma cell lines with and without demethylation by 5‐aza‐2′deoxycytidine and inhibition of deacetylation by trichostatin A treatment. Hypermethylation of SFRP1, 2, 4 and 5 was detected in 93%, 83%, 73% and 85% of 40 cancers; 81%, 89%, 78% and 73% of 37 Barretts epithelia; 25%, 64%, 32% and 21% of 28 adjacent normal epithelia from Barretts patients; and 10%, 67%, 0% and 13% of 30 normal esophagogastric epithelia from healthy individuals, respectively (p < 0.001 for SFRP1, 4 and 5; p < 0.05 for SFRP2). Protein expression of SFRP1, 2 and 4 was downregulated in 87%, 67% and 90% of cancers, and expression correlated inversely with grade and stage of cancers and with grade of dysplasia. Expression of SFRP2 and SFRP4 proteins was lower in cancers with corresponding gene methylation (p < 0.05). Demethylation treatment effectively re‐expressed SFRP mRNA in cancer cell lines. Thus, hypermethylation of SFRP genes is a common early event in the evolution of esophageal adenocarcinoma, and methylation of SFRP1, 4 and 5 might serve as biomarkers for Barretts neoplasia. Aberrant promoter methylation appears to functionally silence SFRP gene expression in esophageal adenocarcinoma.

Frequent Methylation of Eyes Absent 4 Gene in Barrett's Esophagus and Esophageal Adenocarcinoma

Most esophageal adenocarcinomas arise within Barretts esophagus but the cause of this increasingly prevalent condition remains unknown. Early detection improves survival and discriminant screening markers for Barretts esophagus and cancer are needed. This study was designed to explore the natural history of eyes absent 4 (EYA4) gene methylation in the neoplastic progression of Barretts esophagus and to evaluate methylated EYA4 as a candidate marker. Aberrant promoter methylation of EYA4 was studied by methylation-specific PCR using bisulfite-treated DNA from esophageal adenocarcinomas, Barretts esophagus, and normal epithelia, and then confirmed by sequencing. Eight cancer cell lines were treated with the demethylation agent 5-aza-2′-deoxycytidine, and EYA4 mRNA expression with and without treatment was quantified by real-time reverse-transcription PCR. EYA4 hypermethylation was detected in 83% (33 of 40) of esophageal adenocarcinomas and 77% (27 of 35) of Barretts tissues, but only in 3% (2 of 58) of normal esophageal and gastric mucosa samples (P < 0.001). The unmethylated cancer cell lines had much higher EYA4 mRNA expression than the methylated cancer cell lines. Demethylation caused by 5-aza-2′-deoxycytidine increased the mRNA expression level by a median of 3.2-fold in methylated cells, but its effect on unmethylated cells was negligible. Results indicate that aberrant promoter methylation of EYA4 is very common during tumorigenesis in Barretts esophagus, occurs in early metaplasia, seems to be an important mechanism of down-regulating EYA4 expression, and represents an intriguing candidate marker for Barretts metaplasia and esophageal cancer.

Identification and Validation of Colorectal Neoplasia–Specific Methylation Markers for Accurate Classification of Disease

Aberrant DNA methylation occurs early in oncogenesis, is stable, and can be assayed in tissues and body fluids. Therefore, genes with aberrant methylation can provide clues for understanding tumor pathways and are attractive candidates for detection of early neoplastic events. Identification of sequences that optimally discriminate cancer from other diseased and healthy tissues is needed to advance both approaches. Using well-characterized specimens, genome-wide methylation techniques were used to identify candidate markers specific for colorectal neoplasia. To further validate 30 of these candidates from genome-wide analysis and 13 literature-derived genes, including genes involved in cancer and others with unknown functions, a high-throughput methylation-specific oligonucleotide microarray was used. The arrays were probed with bisulfite-converted DNA from 89 colorectal adenocarcinomas, 55 colorectal polyps, 31 inflammatory bowel disease, 115 extracolonic cancers, and 67 healthy tissues. The 20 most discriminating markers were highly methylated in colorectal neoplasia (area under the receiver operating characteristic curve > 0.8; P < 0.0001). Normal epithelium and extracolonic cancers revealed significantly lower methylation. Real-time PCR assays developed for 11 markers were tested on an independent set of 149 samples from colorectal adenocarcinomas, other diseases, and healthy tissues. Microarray results could be reproduced for 10 of 11 marker assays, including eight of the most discriminating markers (area under the receiver operating characteristic curve > 0.72; P < 0.009). The markers with high specificity for colorectal cancer have potential as blood-based screening markers whereas markers that are specific for multiple cancers could potentially be used as prognostic indicators, as biomarkers for therapeutic response monitoring or other diagnostic applications, compelling further investigation into their use in clinical testing and overall roles in tumorigenesis. (Mol Cancer Res 2007;5(2):153–63)

Cronkhite-Canada Syndrome Hamartomatous Polyps Are Infiltrated with IgG4 Plasma Cells

(CCS) is a rare autoimmune disorder characterized by hamartomatous intestinal polyps, protein-losing enteropathy and ectodermal changes [2] . To investigate if CCS is a manifestation of IRAD we examined hamartomatous CCS intestinal polyps for IgG4 plasma cell infiltration using Dear Sir, IgG4-related autoimmune disease (IRAD) is a recently described multisystem disorder characterized by IgG4 plasma cell infiltration with manifestations including autoimmune pancreatitis, sclerosing cholangitis and retroperitoneal fibrosis [1] . Cronkhite-Canada syndrome Published online: May 18, 2007

Verrucous carcinoma of the esophagus: clinicopathophysiologic features and treatment of a rare entity.

In 1948, Ackerman (1), first described verrucous squamous cell carcinoma (VSC) as a variant of oral squamous cell carcinoma. Fewer than 20 cases have been reported. These tumors are slow growing, well-differentiated, irregular wartlike masses with a propensity for local invasion into surrounding tissue. They have been described in the mouth, nasal cavity, larynx, glans penis, scrotum, vulva, vagina, cervix, endometrium, urinary bladder, and anorectal region. VSCs are typically associated with chronic mucosal irritation or a long-term local disease process. Symptoms are insidious and are usually present for a long time before the diagnosis is made. Dysphagia and weight loss are typical at presentation, and usually there is a long delay between the onset of symptoms and detection of the lesion. These tumors uniformly respond well to surgical excision if they are diagnosed before invasion of local tissue. Endoscopically, VSC of the esophagus has a shaggy, white, exophytic, wartlike appearance (hence the name “verrucous”). A conclusive diagnosis of VSC is difficult to make because superficial biopsies tend to show only nonspecific acanthosis, parakeratosis, or hyperkeratosis, with associated acute and chronic inflammation. Deeper, full-thickness biopsies or a fully resected specimen is often needed to detect invasion through the superficial epithelium which is the distinguishing diagnostic feature of VSC. Morbidity and mortality are usually due to complications from local invasion. Metastasis is considered to be exceptionally rare (2). Without esophageal resection, mortality is very high, and patients often die within days

Diagnosis and management of early onset colorectal cancer with non-germline microsatellite instability: what is the algorithm?

Diagnosis and management of early onset colorectal cancer with non-germline microsatellite instability: what is the algorithm?

Sporadic duodenal gastrinoma presenting as Zollinger-Ellison syndrome requiring surgical treatment

Sporadic duodenal gastrinoma presenting as Zollinger-Ellison syndrome requiring surgical treatment