Patterns of allelic loss on chromosome 17 in sporadic breast carcinomas detected by fluorescent-labeled microsatellite analysis

Loss of heterozygosity (LOH) on chromosome 17 is a frequent genetic alteration in breast cancer. To assess whether the location of potential tumor suppressor genes is compatible with the LOH pattern in individual tumors, we analyzed allele loss on chromosome 17 in 121 invasive ductal breast carcinomas and 16 benign breast tumors with 14 polymorphic microsatellite markers (4 on 17p and 10 on 17q). Fluorescent polymerase chain reaction (PCR) for typing microsatellites coupled with DNA fragment analysis in an automated DNA sequencer was applied. Frequencies of LOH varied from 29.4% (D17S1322) to 57.4% (TP53-Alu). No LOH could be detected in benign breast tumors. In 54 tumors the deletion patterns were consistent with the complete loss of 17p (n = 28), 17q (n = 9), or the whole chromosome 17 (n = 17). Five smallest regions of overlap (SROs) were identified in tumors with interstitial deletion patterns. On 17p, two foci were detected affecting the TP53 locus and the hypermethylated in cancer I (HICI) region (17p13.3). On 17q, SROI was localized between markers THRAI and D17S855, centromeric to the breast/ovarian cancer gene BRCAI; SRO2 was flanked by markers AFM234 and NMEI, and SRO3 was centered between markers MPO and GH. Associations between LOH and histopathological characteristics were determined. Significant correlations were found between higher grade and loss of the TP53 gene (marker TP53, P = 0.019), loss of the BRCAI region (P < 0.009), LOH of marker AFM155 (P = 0.003) and marker NMEI (P = 0.026). For positive estrogen receptor status, only LOH of the THRAI marker correlated significantly, whereas highly significant correlations were determined between positive progesterone receptor and markers centromeric to the BRCAI region D17S250 (P = 0.00002), THRAI (P = 0.0006), and the intragenic BRCAI markers [D17S1322 (P = 0.021), D17S855 (P = 0.029)]. Results presented in this study identify five independent regions of chromosome 17 which are likely to contain potential tumor suppressor genes involved in the carcinogenesis of sporadic breast cancer. Genes Chromosom. Cancer 18:181–192, 1997. © 1997 Wiley-Liss, Inc.     

Isolation of DNA from the centromere of human chromosome 7 by microdissection

Centromeres remain the least characterized regions of human chromosomes because they have a very high content of repetitive DNA. Here, we describe a micro-dissection library from the centromeric region of human chromosome 7 and its use for generating sequence tagged sites (STSs). The library contains about 1500 clones with an average insert size of 150 bp and only about 15% of the clones harbour repetitive human DNA. Seven clones hybridizing to alphoid DNA were found to correspond to a fragment of the D7Z2 alphoid array on chromosome 7, thus confirming the origin of the library. A number of clones not containing known repetitive DNA were used to generate STSs that identified yeast artificial chromosomes (YACs) and in turn allowed the STSs to be placed on the physical map. One STS is located between the two Genethon genetic markers closest to the centromere on the q side. Another STS was located 3–4 cM away in 7q11.2, while a third identified YACs containing both low-copy and alphoid sequences that are not yet mapped but are clearly centromeric. The library therefore comprises a collection of sequences from the centromeric region of chromosome 7 that can be used to generate STSs and to map the entire centromeric region.This revised version was published online in November 2005 with corrections to the Cover Date.     

Proteomic analysis of field cancerization in pharynx and oesophagus: a prospective pilot study

‘Field cancerization’ in head and neck squamous cell carcinoma (HNSCC) is poorly understood and it may extend from the pharynx into the oesophagus. Both local recurrences and second primary carcinomas/second field tumours may originate from field cancerization. Our prospective pilot study aimed at the identification of patients suffering from field cancerization on the basis of mucosal protein profiles. Five mucosal biopsies from the oropharynx, hypopharynx and from three regions of the oesophagus were taken from 24 patients. Protein profiles were generated from the mucosal biopsies. After classifier learning, using the profiles of the patients without tumour diagnosis (n = 9), we were able to discriminate between the different mucosal sites and between healthy mucosa and HNSCC using tumour and healthy tissue samples. Mucosal biopsies of tumour patients (n = 15) revealed changes in the protein profiles similar to those in the tumours. During 42 months median follow‐up, six tumour patients experienced local recurrences and second field tumours, of which three occurred in the oesophagus. In all six cases, tumour relapse was correctly predicted by altered mucosal protein profiles (p = 0.007, Fisher's exact test, two‐tailed). Consequently, molecular field cancerization had a strong impact on progression‐free survival (p = 0.007, log‐rank test). Protein profiles of small diagnostic biopsies hold great promise to improve personalized risk assessment in HNSCC. Larger studies are needed to further substantiate these findings. Copyright © 2010 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.     

Multiparameter immunofluorescence on paraffin-embedded tissue sections.

Immunohistochemical techniques have gained increasing importance in diagnostics and research. While formalin-fixed, paraffin-embedded human tissue retains excellent morphology, the detection of antigens by immunofluorescence in its sections and especially the demonstration of multiple simultaneous antibodies have limitations. Double immunofluorescence labeling of routinely processed paraffin sections has been described previously. The signal intensity observed after triple labeling has been reported to be significantly inferior to that obtained by application of double fluorochromes. The authors show multicolor labeling of three and four primary antibodies in routinely processed paraffin-embedded tissue sections using a standardized immunofluorescence technique. In addition, procedures to reduce background staining and to avoid nonspecific double staining are described.     

Malignant and benign breast masses on 3D US volumetric images: effect of computer-aided diagnosis on radiologist accuracy

PURPOSE: To retrospectively investigate the effect of using a custom-designed computer classifier on radiologists' sensitivity and specificity for discriminating malignant masses from benign masses on three-dimensional (3D) volumetric ultrasonographic (US) images, with histologic analysis serving as the reference standard. MATERIALS AND METHODS: Informed consent and institutional review board approval were obtained. Our data set contained 3D US volumetric images obtained in 101 women (average age, 51 years; age range, 25-86 years) with 101 biopsy-proved breast masses (45 benign, 56 malignant). A computer algorithm was designed to automatically delineate mass boundaries and extract features on the basis of segmented mass shapes and margins. A computer classifier was used to merge features into a malignancy score. Five experienced radiologists participated as readers. Each radiologist read cases first without computer-aided diagnosis (CAD) and immediately thereafter with CAD. Observers' malignancy rating data were analyzed with the receiver operating characteristic (ROC) curve. RESULTS: Without CAD, the five radiologists had an average area under the ROC curve (A(z)) of 0.83 (range, 0.81-0.87). With CAD, the average A(z) increased significantly (P = .006) to 0.90 (range, 0.86-0.93). When a 2% likelihood of malignancy was used as the threshold for biopsy recommendation, the average sensitivity of radiologists increased from 96% to 98% with CAD, while the average specificity for this data set decreased from 22% to 19%. If a biopsy recommendation threshold could be chosen such that sensitivity would be maintained at 96%, specificity would increase to 45% with CAD. CONCLUSION: Use of a computer algorithm may improve radiologists' accuracy in distinguishing malignant from benign breast masses on 3D US volumetric images.     

Strategies for the reduction of cerebral microembolism during transmyocardial laser revascularization

BACKGROUND AND OBJECTIVES: During transmyocardial laser revascularization (TMLR), multiple microembolic signals (MES) can be detected in cerebral arteries. We sought to characterize composition and clinical relevance of these MES and to evaluate strategies to reduce cerebral microembolization during TMLR. STUDY DESIGN/MATERIALS AND METHODS: TMLR was performed in pigs. Laser energy was set to 4-10 J (group A) or 80 J (group B). Oxygen concentration was varied between 21 and 100%. MES were recorded in the ophthalmic artery. Brain and spinal cord were investigated histologically after 10 days. RESULTS: More MES could be detected during high- compared to low-energy laser procedures. Ventilation with 100% oxygen reduced the number of MES. No lesions were found on histology. CONCLUSIONS: The number of MES depends on the laser energy. Laser-induces cavitation-effects lead to an additional release of nitrogen bubbles. Thus, the microembolic load can be reduced by ventilation with 100% oxygen and by decreasing the laser energy.     

Perioral reflex myoclonias: a controlled study in patients with JME and focal epilepsies

PURPOSE: Perioral reflex myoclonias (PORM) are obvious, frequent, but often unobserved focal seizures in different epileptic syndromes and the leading seizure type in reading epilepsy. PORMs remain often undiagnosed because the patients are not aware that these are epileptic seizures and fail to report them. Their semiology is not fundamentally different in various epileptic syndromes. METHODS: We studied the frequency of PORM in patients with juvenile myoclonic epilepsy (JME) compared with patients with focal epilepsies. Twenty-five patients with JME were investigated with a standardized neuropsychological test program and compared with 25 matched patients with focal epilepsies. Statistical significance was calculated by using Fisher's exact test. RESULTS: We found significant differences between the groups regarding both frequency of PORM and activation of epileptic discharges. These observations seem to indicate that PORM, like praxis-induced seizures, are typical traits in JME. CONCLUSIONS: PORM are more frequent in JME compared with focal epilepsies. The distinction between focal and generalized epileptic ictogenesis may be less clear than is traditionally believed.     

Antiepileptic drug use in nursing home residents: a cross-sectional, regional study.

The use of antiepileptic drugs (AED), their primary indication, comorbid conditions, and concomitant medications were collected from 565 nursing homes (NH) residents of six NH located around the city of Mainz, Germany representing 5.05% of all NH residents in the area. Data were collected from the electronic pharmacy files and by reviewing all available medical records. Average age was 82.2 +/- 2.4 years, 85.5% were women. Of 565 NH residents 28 (4.96%) received AED therapy, of which in 17 (63%) AED were prescribed for a seizure-related diagnosis. In 76.5% seizure types were unspecified and a distinction in focal and generalized epilepsy was made in only 23.5% of patients. Three patients never had epileptic seizures and in four residents the reason for AED use was unclear. AEDs most frequently prescribed were carbamazepine (37.1%), valproic acid (25.9%), and phenytoin (14.8%). Five patients received benzodiazepines (18.5%), newer generation AED were used in only four (14.8%) cases. Residents on average took n = 5.6 +/- 3.3 other drugs. Opportunities exist for health care professionals to improve the medical management of nursing facility residents receiving AEDs.