Germline mutations in PTEN, encoding a dual-specificity phosphatase on 10q23.3 , cause Cowden syndrome (CS) , which is characterized by a high risk of breast and thyroid cancers. Loss of heterozygosity of 10q22-24 markers and somatic PTEN mutations have been found to a greater or lesser extent in a variety of sporadic component and noncomponent cancers of CS. Among several series of sporadic breast carcinomas , the frequency of loss of flanking markers around PTEN is approximately 30 to 40% , and the somatic intragenic PTEN mutation frequency is <5%. In this study , we analyzed PTEN expression in 33 sporadic primary breast carcinoma samples using immunohistochemistry and correlated this to structural studies at the molecular level. Normal mammary tissue had a distinctive pattern of expression: myoepithelial cells uniformly showed strong PTEN expression. The PTEN protein level in mammary epithelial cells was variable. Ductal hyperplasia with and without atypia exhibited higher PTEN protein levels than normal mammary epithelial cells. Among the 33 carcinoma samples , 5 (15%) were immunohistochemically PTEN-negative; 6 (18%) had reduced staining, and the rest were PTEN-positive. In the PTEN-positive tumors as well as in normal epithelium , the protein was localized in the cytoplasm and in the nucleus (or nuclear membrane). Among the immunostain negative group, all had hemizygous PTEN deletion but no structural alteration of the remaining allele. Thus, in these cases, an epigenetic phenomenon such as hypermethylation, decreased protein synthesis or increased protein degradation may be involved. In the cases with reduced staining, 5 of 6 had hemizygous PTEN deletion and 1 did not have any structural abnormality. Finally, clinicopathological features were analyzed against PTEN protein expression. Three of the 5 PTEN immunostain-negative carcinomas were also both estrogen and progesterone receptor-negative, whereas only 5 of 22 of the PTEN-positive group were double receptor-negative. The significance of this last observation requires further study. (Am J Pathol 1999, 155:1253-1260)

Summary PTEN is a putative tumour suppressor gene located on chromosome band 10q23. Mutations in PTEN have been identified in numerous human malignancies, including cancers of the brain, endometrium, ovary, and prostate. In this study, we screened 80 Barrett's oesophagus-associated adenocarcinomas (BOAd) for loss of heterozygosity (LOH) at 10q23, using the microsatellite markers D10S541, D10S219, and D10S551. Tumours demonstrating LOH were then screened for the presence or absence of PTEN mutations. LOH at one or more loci was identified in 17/80 (21%) cases. In none of these cases did we detect mutations in PTEN. The presence of LOH did not correlate with patient age, tumour stage, degree of differentiation, presence of perineural or vascular invasion, or overall survival. We conclude that LOH at chromosome 10q23 is uncommon in BOAd, is not associated with mutations in the PTEN tumour suppressor gene, and does not correlate with the clinical or pathologic features of these tumours. It is possible that PTEN is inactivated through other mechanisms in BOAd. MATERIALS AND METHODS Study group80 patients who had en bloc oesophageal resection at the Brigham and Women's Hospital and at the Beth Israel-Deaconess Hospital between 1973 and 1995 were identified. All patients had histologically confirmed BOAd, and none had received preoperative chemotherapy or radiation. All patients were treated surgically with an intent to cure.Selected clinical information (patient age, gender) and followup data were obtained from review of the patient's hospital charts and the hospital tumour registry, or from direct telephone interviews with the patient and/or his/her family when necessary. Follow-up time was calculated from the date of initial diagnosis to either the date of death or, for the patients who were still alive, to the date of the most recent clinical investigation. In the survival analysis, either death or tumour recurrence was considered a failure (event). Patients alive without disease at last follow-up were censored in the analysis. Pathologic analysisAll oesophageal resection specimens were received in the surgical pathology laboratory in the fresh state and fixed in 10% buffered formalin for subsequent tissue sectioning. Tissue sections were processed routinely, embedded in paraffin, and stained with haematoxylin and eosin (H & E).The following microscopic features were evaluated in all cases by one of the authors (RDO): 1) Pathologic stage according to the 1993 revised AJCC TNM classification (Fleming et al, 1997); 2) The presence or absence of lymphovascular invasion; 3) The presence or absence of perineural invasion; 4) Degree of tumour differentiation (well, > 95% of the tumour composed of glands; moderate, 50-95% of the tumour composed of glands; poor, < 50% of the tumour composed of glands). Molecular analysisSections from paraffin-embedded tumour specimens were cut. Tumour and normal tissue were identified and separated by microdissection. DNA extraction was performed using the QIAprep kit (Qiagen Inc, Chatswo...