Zoning of mucosal phenotype, dysplasia, and telomerase activity measured by telomerase repeat assay protocol in Barrett's esophagus.

Glandular dysplasia in Barrett's esophagus may regress spontaneously but can also progress to cancer. The human telomerase RNA template and the human telomerase reverse transcriptase enzyme which do not, of themselves, correlate strongly with telomerase activity, are too often overexpressed in Barrett's dysplasia to predict individual cancer risk. This study relates telomerase activity, mucosal phenotype, and dysplasia in Barrett's esophagus. Biopsies (n = 256) from squamous esophagus, columnar-lined esophagus every 2 cm, esophago-gastric junction, gastric body, and antrum from 32 patients with long-segment Barrett's esophagus were evaluated by telomerase repeat assay protocol (TRAP). Three biopsies for histology (n = 794) were simultaneously taken at each anatomical level. These and all prior and subsequent biopsies (n = 1917) were reviewed for mucosal phenotypes and dysplasia severity. Intestinal-type Barrett's mucosa was present at all levels in Barrett's esophagus. At least one Barrett's biopsy was TRAP(+) in 22 of 32 patients. TRAP positivity of intestinal-type Barrett's mucosa increased distally, possibly as a consequence of mucosal exposure to acid or bile reflux. Native gastric mucosa was rarely TRAP(+) (1/31 corpus, 2/32 antrum), whereas native squamous mucosa usually was TRAP(+) (31/32). Dysplasia almost always involved intestinal-type Barrett's mucosa (85/87; P <.00001), without evidence of proximal-distal zoning. TRAP could be positive without dysplasia and negative in extensive, even high-grade, dysplasia. TRAP activity merits evaluation as a candidate biomarker for increased risk of persistent dysplasia and cancer progression in Barrett's esophagus.

Selective ablation of human cancer cells by telomerase-specific adenoviral suicide gene therapy vectors expressing bacterial nitroreductase.

Reactivation of telomerase maintains telomere function and is considered critical to immortalization in most human cancer cells. Elevation of telomerase expression in cancer cells is highly specific: transcription of both RNA (hTR) and protein (hTERT) components is strongly upregulated in cancer cells relative to normal cells. Therefore, telomerase promoters may be useful in cancer gene therapy by selectively expressing suicide genes in cancer cells and not normal cells. One example of suicide gene therapy is the bacterial nitroreductase (NTR) gene, which bioactivates the prodrug CB1954 into an active cytotoxic alkylating agent. We describe construction of adenovirus vectors harbouring the bacterial NTR gene under control of the hTR or hTERT promoters. Western blot analysis of NTR expression in normal and cancer cells infected with adenoviral vectors showed cancer cell-specific nitroreductase expression. Infection with adenoviral telomerase-NTR constructs in a panel of seven cancer cell lines resulted in up to 18-fold sensitization to the prodrug CB1954, an effect that was retained in two drug-resistant ovarian lines. Importantly, no sensitization was observed with either promoter in any of the four normal cell strains. Finally, an efficacious effect was observed in cervical and ovarian xenograft models following single intratumoural injection with low doses of vector, followed by injection with CB1954.

'Senescence-associated' beta-galactosidase activity in the upper gastrointestinal tract.

Beta-galactosidase activity at pH 6 is associated in vitro with senescence and cellular death, but in vivo data are sparse. This study undertook firstly to map 'senescence-associated' beta-galactosidase activity (SAbetaG) at pH 6 in normal epithelia and mucosae of the upper gastrointestinal tract. As escape from senescence confers a proliferative advantage, a reduction in SAbetaG activity might be predicted in neoplasia and their precursors in vivo. This prediction was tested in metaplastic, dysplastic, and neoplastic epithelium of the upper gastrointestinal tract. Histochemical staining for SAbetaG was performed at pH 6 on cryostat sections of 350 endoscopic biopsies from sites including oesophagus, stomach, and duodenum of 46 patients: 28 with Barrett's oesophagus (two with adenocarcinoma), 15 with gastric adenocarcinoma, and three with oesophageal squamous cancer. A staining score (range 0-6) was assigned to epithelial cells in all mucosae and scores were calculated for surface (luminal), intermediate, and deep (basal) layers. The strongest SAbetaG activity was in surface luminal cells of normal duodenal mucosa (mean score 3.6+/-0.5; n=19), 'specialized' Barrett's mucosa (mean 2.2+/-0.12; n=105), and intestinal metaplasia in the stomach (mean 2.4+/-0.40; n=16). Squamous epithelium was consistently negative for SAbetaG activity. Low- and high-grade Barrett's dysplasia showed no decrease in SAbetaG activity, but reduced activity was seen in gastric and oesophageal adenocarcinomas (mean 1.24+/-0.29; n=17; p=0.012). In six gastric adenocarcinomas, there was no detectable activity. Whether SAbetaG is truly a marker of cellular senescence in vivo remains to be determined. Activity is low in mucosal proliferation compartments and increases with cellular differentiation, especially in native or metaplastic intestinal mucosae. SAbetaG activity persists in dysplastic mucosae but may show some reduction or loss in adenocarcinomas (p=0.0012). Loss of SAbetaG activity is not, therefore, an early event in glandular dysplasia-neoplasia of the upper gastrointestinal tract.