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Humoral hypercalcemia of malignancy may reflect the synthesis and secretion of biologically active parathyroid hormone-related protein (PTHrP) by a given tumor. In the present study we investigated 25 human non-endocrine carcinomas which were clinically associated with hypercalcemia (Ca > 11 mg%). By applying PTHrP-specific immunocytochemistry, PTHrP could be detected in all tumors. The intra-tumorous distribution was heterogeneous with strong positivity in relatively few cells or weak positivity in the majority of cells. Surprisingly, in the PTHrP producing cells none of the marker proteins typical of endocrine cells (neuron-specific enolase, Leu-7 antigen, chromogranin, synaptophysin and endocrine granule constituent) was found. On the other hand, PTHrP producing cells of endocrine origin, such as medullary cancer, or normal and adenomatous parathyroid glands, all produce these endocrine markers. Thus for the first time, the existence of peptide hormone producing tumor cells is reported without expression of endocrine markers. This indicates a special mechanism of PTHrP secretion.

The people of Estonia, who until their independence had been systematically deprived of all forms of information, have shown a ready acceptance of the use of information technology in all areas of life. This environment and an excellent IT infrastructure have since the beginning of the new century provided favourable conditions for developing various individual projects in telemedicine. At the core of current telemedical applications in Estonia is the BITNET Project (in neurology, general medicine), built up with Swedish cooperation, and three German-Estonian projects (in telecardiology and telepathology). These projects are accompanied by studies of their cost-effectiveness. They constitute the basis for the plan that routine telemedical services be taken over by the Estonian health insurance. Differing from the situation in the Scandinavian countries, which have had an effective national telemedical service for over ten years, the special feature of telematrics in Estonia is its international networking with foreign centres of excellence. This has its origin in the aim of Estonian health policy to ensure medical services within its own country in all branches of medicine.

Decreased expression of p16 may result from hypermethylation of the promoter or from deletion of the gene. It can lead to intensified proliferation of neoplastic cells and to cytostatic drug resistance. The study was aimed at the examination of prognostic value of p16 expression in relation to Ki67 and caspase-3 in ovarian cancers using immunohistochemistry. The immunohistochemical studies were performed on 73 paraffinembedded samples of ovarian cancers from 43 patients and samples from 6 healthy ovaries. We have used monoclonal antibodies against p16. ABC method and DAB were used for antigens visualisation. The intensity of the immunohistochemical reactions was appraised using the semi-quantitative IRS scale. In healthy ovaries we have shown strong reaction in the nuclei of surface epithelium. In the case of studied ovarian cancers, the reaction of a nuclear and cytoplasmic localization was obtained. The mean overall immunoreactivity score of nuclear p16 expression amounted to 5.30±3.44 SD in primary laparotomy material and 6.61±4.34 SD in secondary cytoreduction material. Statistical analysis demonstrated that lower p16 expression was typical of the younger patients and the patients who died. Kaplan- Meier’s analysis proved that lower expression of p16 was characteristic of cases with shorter overall survival. In the present study we have demonstrated that lowered p16 expression represented an unfavourable prognostic index in ovarian cancer. Lowered p16 expression was also typical for chemotherapy-resistant ceases (cases of lower caspase-3 and higher Ki67 at secondary cytoreduction expression).

The new WHO series on Pathology and Genetics of Tumours of the Breast and Female Genital Organs (33) defined to name the group of epithelial ovarian tumors with a dignity between benign and malignant exclusively as Borderline Tumors of the Ovary (BOT) and to skip the term "... of low malignant potential". Further, the term "atypical proliferative tumour" was not recommended by the WHO. During a Consensus Meeting on Borderline Tumors of the Ovary held at Bethesda on August 27-28, 2003 (2) the expert panel also recommended to use the BOT terminology. However the term "tumour of low malignant potential" and--less favourable--"atypical proliferative tumour" may be used as synonym. Both groups agreed unanimously that the name carcinoma should not be included in the diagnosis. The group of borderline ovarian tumors is heterogeneous. 80 to 90% of the cases have a very favourable prognosis while 10-20% exhibit a recurrent clinical course with peritoneal implants and very rarely death from the tumor within 10 years. The morphological criteria and supporting methods for recognizing unfavorable BOT and for distinguishing them from highly differentiated ovarian carcinomas are summarized. The prognostic importance of peritoneal implants is described. The concept of micropapillary serous carcinomas (MPSC) and its implications on the diagnostic work of pathologists will be discussed. The presented data focus on serous tumors since this is by far the most common variant.

We report a concept that is not only applicable for telemicroscopy, but may be adapted for the remote control of any kind of scientific device, opening a new field of application for the Internet.

Fifty patients clinically suspected of having primary hyperparathyroidism were examined by scintigraphy (201Tl-99mTc subtraction) and sonography (5 MHz). In 28 cases, the results of subtraction scintigraphy, and in 26 cases, the results of sonography were checked by operation. In 25 patients there was a solitary parathyroid adenoma and in two, primary hyperplasia. In one patient all four parathyroid glands were normal. Subtraction scintigraphy had a sensitivity of 90% and a specificity of 98%, while sonography had a sensitivity of 82% and a specificity of 95%. The combination of 201Tl-99mTc subtraction scintigraphy as a functional investigation and sonography as a structural imaging method produced a sensitivity of 96% and is therefore superior to all other methods of localisation for normally situated glands. Diagnostic certainty is reduced by the presence of a nodular thyroid.

NF-kappaB has been demonstrated to activate proliferative, inflammatory, and angiogenic processes in ovarian cancer cells in vitro. To add translational information on the situation in vivo, we determined the expression pattern of p65, an important subunit of the classic NF-kappaB pathway, in ovarian carcinoma tissue, and investigated in vivo and in vitro whether this pathway is implicated in the known overexpression of cyclooxygenase-2 (COX-2).p65 siRNA, chemiluminescent NF-kappaB transcription factor assay, Taqman PCR, as well as immunoblotting were performed with OVCAR-3 ovarian cancer cells. 83 primary ovarian cancinomas as well as 17 cases of benign ovarian tissue were analyzed by p65 and COX-2 immunohistochemistry using a tissue microarray.DNA-binding avtivity as well as COX-2 mRNA and protein expression were strongly inducible by IL-1beta treatment in OVCAR-3 cells, while p65 siRNA inhibited IL-1beta-dependent p65 activity (p = 0.037) as well as COX-2 expression on the mRNA (p0.03) and on the protein level. In human tumor tissue, p65 protein expression was significantly associated with COX-2 expression (p = 0.002) as well as tumor grading (p = 0.005). Furthermore, p65 expression was a significant prognostic indicator of a reduced patient survival both in univariate (p = 0.038) and in multivariate analysis (p = 0.014).Our study indicates a deregulation of the classical NF-kappaB pathway in ovarian cancer, which results in the overexpression of the NF-kappaB target gene COX-2. Components of this pathway might constitute novel attractive targets for a specific therapy of advanced ovarian cancer.

While cigarette smoking and chronic alcohol consumption are the major risk factors for the development of head and neck cancer, it is assumed that genetic factors contribute to risk.We examined genotype frequencies from leukocyte DNA of 269 laryngeal cancer patients, 123 pharyngeal cancer patients and 216 controls. Polymorphisms at different glutathione-S-transferase (GST) gene loci were investigated. Losses of heterozygosity (LOH) at 12 different chromosomal gene loci were determined in 37 of the study patients by comparing blood and tumor cell DNA. The relationship between high risk genotypes and the occurrence of LOH was investigated.Glutathione-S-transferase high risk genotypes were identified at the first and third genes of the M family (GSTM1, GSTM3) and the first gene of the P family (GSTP1). These high risk genotypes are seen to have a statistically significant influence on the occurrence of LOH in the tumor tissue.There is evidence that the polymorphisms studied play a role in the carcinogenic process by influencing the chromosomal fragility which may lead to the inactivation of tumor suppressor genes or the activation of oncogenes.

Antibodies against PD-1 and PD-L1 can cause strong and durable anti-tumor immune responses in non-small cell lung cancer (NSCLC). Immunohistochemistry for PD-L1 (PD-L1 IHC) was tested as a predictive biomarker. Several IHC assays and interpretation criteria were developed in parallel.The clinical significance of PD-L1 IHC in NSCLC and the optimum method for staining and interpretation of the results are the subject of ongoing studies. The diagnostic application of immunotherapy in NSCLC necessitates harmonization of PD-L1 IHC to obtain evidence for guidelines; therefore, a consensus opinion on a well-founded diagnostic mode of testing should be defined based on published studies and the results of the first German PD-L1 IHC harmonization study.1. Summary of the current data situation. 2. Evaluation of the first German PD-L1 IHC harmonization study (centralized, staining with PD-L1 IHC analogous to studies, 15 cases of NSCLC, 4 IHC study assays [28‑8, 22C3, SP142 and SP263] and scoring by 9 pathologists).The use of PD-L1 IHC in NSCLC is suitable for identification of patients with an increased probability of a clinical benefit from immunotherapy. The various proportional cut-offs used to interpret the staining results can be summarized in a total score, which can be reproducibly assessed. The staining patterns of the four assays investigated were, however, not congruent in all situations.In principle, the use of PD-L1 IHC for assessment of the expression in tumor cells is a reliably determinable biomarker. Evaluation algorithms should be based on published clinical trials. For NSCLC approvals with obligatory PD-L1 IHC are to be expected but it remains to be seen to what extent PD-L1 IHC will be implemented in the clinical routine.

A 19-yr-old woman with previously diagnosed clear cell adenocarcinoma was referred to the Charité for further treatment. Biopsies were taken from the cervix, the endometrium, pseudomembranes in the peritoneum, and sentinel lymph nodes. The morphologic picture of pseudomembranes and inflammation together with the provided information about plasminogen deficiency of the patients led to the hypothesis of ligneous cervicitis. The previously taken biopsies of the adenocarcinoma were reevaluated and showed a clear cell lesion. Further immunohistochemical examination with antibodies against p16, Ki67, CEA, and p53 could not prove its malignant character. As a result we diagnosed an atypical form of microglandular hyperplasia in a patient with ligneous cervicitis. Ligneous cervicitis is a rare inflammatory condition that might affect all mucus membranes in patients with plasminogen deficiency. This case shows the importance of correlating pathologic and clinical findings in the diagnosis of ligneous cervicitis because of the rarity of the disease and the heterogeneity at presentation. © 2013 International Society of Gynecological Pathologists.

Oestrogen receptors (ESRs) regulate the growth and differentiation of normal ovarian epithelia. However, to date their role as biomarkers in the clinical setting of ovarian cancer remains unclear. In view of potential endocrine treatment options, we tested the role of ESR1 mRNA expression in ovarian cancer in the context of a neo-adjuvant chemotherapy trial. Study participants had epithelial ovarian or peritoneal carcinoma unsuitable for optimal upfront surgery and were treated with neo-adjuvant platinum-based chemotherapy before surgery. RNA was isolated from frozen tumour biopsies before treatment. RNA expression of ESR1 was determined by microarray and reverse transcriptase kinetic PCR technologies. The prognostic value of ESR1 was tested using univariate and multivariate Cox proportional hazards models, Kaplan–Meier survival statistics and the log-rank test. ESR1 positively correlates with proliferation markers and histopathological grading. ESR1 was a significant predictor of survival as a continuous variable in the univariate Cox regression analysis. In multivariate analysis, elevated baseline ESR1 mRNA levels predicted prolonged progression-free survival (P=0.041) and overall survival (P=0.01) after neo-adjuvant chemotherapy, independently of pathological grade and age. We conclude that pretreatment ESR1 mRNA is associated with tumour growth and is a strong prognostic factor in ovarian cancer, independent of the strongest clinical parameters used in clinical routine. We suggest that ESR1 mRNA status should be considered in order to minimize possible confounding effects in ovarian cancer clinical trials, and that early treatment with anti-hormonal agents based on reliable hormone receptor status determination is worth investigating.

HER2 is an important tumour marker in breast cancer. However, there is controversy regarding which method reliably measures HER2 status. This study evaluates the concordance between HER2 gene amplification in invasive breast cancer determined by fluorescence in situ hybridisation (FISH) and a new silver enhanced in situ hybridisation (SISH) technique. Ninety-nine cases were analysed by direct-labelled manual FISH (PathVysion(R), Abbott/Vysis) and bright field automated SISH (INFORM(R), Ventana). For comparison, all specimens were stained by immunohistochemistry (Dako-HercepTesttrade mark and Ventana-PATHWAY(R)4B5). Evaluation was performed by five pathologists following the algorithms of the manufacturers and the American Society of Clinical Oncology/College of American Pathologists (ASCO/CAP) guidelines. Concordance was calculated and the value of kappa statistics estimated. Overall concordance between FISH and SISH was 96.0% (kappa = 0.754, 95%CI). Discrepancies were mostly seen in tumours with intra-tumoural heterogeneity of HER2 amplification. In conclusion, HER2 gene copy status can be reliably determined by SISH. The 96% concordance with FISH fulfils the ASCO/CAP requirement of greater than 95% concordance for amplified vs non-amplified cases. There was a low inter-observer variability in the interpretation of SISH, suggesting that SISH is equally reliable in determining HER2 amplification as FISH. Because SISH combines bright field microscopy with molecular analysis and full automation, it appears to be particularly suited for routine application in surgical pathology.

Mit der Entdeckung der katalytischen Aktivität von Ribonukleinsäuren wurde begonnen, darauf basierende therapeutische Strategien zu entwickeln, die auf genetischer Ebene den Stoffwechsel oder virale Infektionen von Zellen beeinflussen. Entsprechende Oligoribonukleotide waren befähigt, spezifisch RNA-Stränge zu erkennen und zu spalten. In Analogie zu Enzymen wurden sie als Ribozyme bezeichnet. Aufgabe der vorliegenden Arbeit war die Entwicklung eines metabolisch stabilen Hammerhead-Ribozyms. Dieses sollte lipidvermittelt in die Zellen des Nierenzellkarzinoms RCC 95/96 transfiziert werden und dort die Genexpression des dem Parathormon verwandten Proteins (PTHrP) durch die Spaltung der PTHrP-mRNA unterdrücken. Ausgangspunkt der Entwicklung war das Hammerhead-Ribozym RbO. Es wies in zellfreien Versuchen bei der Spaltung einer 232 Basen langen Substrat-RNA mit einem k(obs)-Wert von 47,42 E-3 /min eine zur Literatur vergleichbar hohe Aktivität auf, jedoch zeigte es sich als reiner RNA-Strang gegenüber Nukleasen sehr fragil. Das Endprodukt der in mehreren Schritten abgelaufenen Weiterentwicklung des Ribozyms RbO stellte das Ribozym RbS dar. Im Vergleich zum Ausgangsribozym bestand das in der Stammschleife verkürzte Ribozym zu 71 % aus DNA. Es besaß Phosphorothioatmodifi-kationen in den Flanken und in den konservierten Sequenzbereichen. Zudem war durch einen Basenwechsel im katalytischen Teil der Stammschleife eine Pyrimidinbase durch eine Purinbase ausgetauscht worden. Zu-sammen bedingten die genannten Veränderungen eine Steigerung der Ribozymstabilität um mehr als das Zehnfache. Dabei war die katalytische Aktivität des Ribozyms RbS mit einem k(obs)-Wert von 45,76 E-3 /min gegenüber RbO annähernd identisch. Das schrittweise Vorgehen bei der Ribozymentwicklung mit dem Erstellen von 20 unterschiedlich modifi-zierten Ribozymen ermöglichte es, den Einfluss einzelner Modifikationen auf die Ribozymaktivität zu prüfen. Mehrfach konnten dabei die Ergebnisse anderer Forschungsgruppen bestätigt werden. So wurde erkannt, dass für viele Modifikationen, wie Ribozymverkleinerung, RNA-DNA-Basenaustausch und Phosphorothioa-teinbindung, die Auswirkung auf die katalytische Aktivität nur begrenzt vorherzusagen ist und optimale Er-gebnisse nur durch Testreihen zu erlangen sind. Bei der Behandlung der Tumorzellen in Monolayer-Zellkultur ließ sich für das Ribozym RbS keine signifi-kante Wirkung nachweisen. Als Ursache dafür ist am ehesten die auf Grund von Sekundärstrukturen fehlende Erkennung der Zielsequenz innerhalb der PTHrP-mRNA anzunehmen., The development of therapeutic strategies affecting cellular metabolism and viral infections of cells was in-troduced after the discovery of the catalytic activity of ribonucleic acid. Appropriate oligoribonucletides were able to specificly recognize and cleave RNA strands. They were called ribozymes by analogy with enzymes. The main task of our research was the development of a metabolic stable hammerhead ribozym. The ribozym should be transfected by lipid mediated transport into renal carcinoma cells RCC 95/96 where it should suppress the gene expression of parathormone-related peptide (PTHrP) by means of cleavage of the PTHrP mRNA. The hammerhead ribozyme RbO was the starting point of the development. Its activity in cleavage of 232 base long subsrats in cell-free tests was comparable to the literature (k(obs) value 47,42 E-3 /min). However it was very fragile regarding nucleases. The end product of the gradual further develop-ment of the ribozyme RbO was the ribozyme RbS. This ribozyme which was shortened in helix 2 consisted of 71 percent DNA in comparison to the original one. It was modified with phosphorothioates in helix 1 and 3 and in the conserved sequence regions. Furthermore a pyrimidine base was exchanged for a purine base in the catalytic part of helix 2. Altogether the named alterations increased the stability of the ribozyme more than 10 times. The catalytic activity of the ribozyme RbS compared to RbO was approximately identical (k(obs) value 45,76 E-3 /min). The step-by-step development of the ribozymes with the creation of 20 different modified ribozyms made it possible to study the impact of individual modifications on the ribozyme activity. Often the results of other research groups were confirmed. Thus it was detected that the effects of some modifications like ribozyme reduction, RNA-DNA base exchange and phosphorothioates integration are only partly predictable. Therefo-re optimal results are only obtained by a number of tests. Finally we could not demonstrate a significant effect of the ribozyme RbS in the treatment of tumor cells in monolayer. The most likely explanation for this seems to be that the target sequence inside of the PTHrP-mRNA wasn't recognized due to secondary structures.

Das Ziel der vorliegenden Arbeit bestand in der molekularzytogenetischen Charakterisierung von invasiv duktalen und invasiv lobulären Mammakarzinomen sowie der Identifizierung unterschiedlicher genetischer Alterationsmuster in Tumorsubgruppen der invasiv duktalen Mammakarzinome. Mit Hilfe der komparativen genomischen Hybridisierung (CGH) wurden 30 invasiv duktale und 20 invasiv lobuläre Mammakarzinome auf genetische Veränderungen untersucht. In beiden Tumorarten überwiegen DNA-Deletionen gegenüber DNA-Gewinnen, daher scheint die Inaktivierung tumorsupprimierender Gene einen größeren Einfluß auf das maligne Wachstumsverhalten auszuüben. Invasiv duktale Mammakarzinome zeichnen sich durch eine im Mittel größere Anzahl an Alterationen pro Tumorfall aus als invasiv lobuläre Karzinome (14,9 vs. 8,9). Neben einer größeren Anzahl an DNA-Verlusten ist auch die Anzahl der DNA-Überrepräsentierungen deutlich höher verglichen mit invasiv lobulären Mammakarzinomen (6,2 pro Tumorfall vs. 2,7 pro Tumorfall). DNA-Gewinne auf Chromosom 1q und DNA-Verluste auf den Chromosomen 6q, 11q22- qter und 13q fanden sich in >=40% der Fälle in beiden histologischen Tumortypen. In invasiv duktalen Mammakarzinomen traten DNA-Gewinne mit größerer Frequenz auf den Chromosomen 6p, 8q, 11q13, 16p, 17q, 19p/q und 20q auf. Ebenfalls häufiger waren DNA-Deletionen auf den Chromosomen 2q, 3p, 4p/q, 5q, 7p, 8p, 9q, 10q und 15q zu finden. DNA-Verluste auf den Chromosomen 16q, 17p, 18q und 22q wurden dagegen vermehrt in invasiv lobulären Karzinomen detektiert. Gut (G1) und schlecht (G3) differenzierte invasiv duktale Mammakarzinome zeichnen sich durch ein unterschiedliches genetisches Muster aus. Während gut differenzierte Tumore durch DNA-Gewinne auf 1q, 11q11-13, 16p und 20q gekennzeichnet sind, weisen die schlecht differenzierten Tumore zusätzlich DNA-Deletionen im Bereich der Chromosomen 5q, 18q und 21q21 auf. Außerdem läßt sich ein unterschiedliches genetisches Muster bei Östrogenrezeptor- positiven und Östrogenrezeptor-negativen invasiv duktalen Mammakarzinomen feststellen. Die Östrogenrezeptor-negativen Tumore zeigen eine größere Anzahl an Alterationen, dazu gehören zusätzliche DNA-Überrepräsentierungen auf 1p, 6p und 22q und DNA-Verluste der Chromosomen 5q, 7p, 8p und 12q. Somit läßt sich feststellen, daß sich invasiv duktale und invasiv lobuläre Mammakarzinome durch ein wiederkehrendes Muster chromosomaler Veränderungen charakterisieren lassen., The aim of the study was to analyze invasive ductal and invasive lobular breast carcinomas regarding to molecular DNA imbalances as well as different DNA imbalances in tumor subgroups of the invasive ductal carcinomas. Comparative genomic hybridization (CGH) was applied to analyze 30 invasive ductal and 20 invasive lobular breast carcinomas to carry out genetic alterations. In both tumor subgroups, DNA losses showed a higher incidence compared to DNA gains, suggesting a higher influence of inactivation of tumor suppressor genes in tumor progression. Invasive ductal carcinomas showed a higher incidence of alterations per case compared to invasive lobular carcinomas (14,9 vs. 8,9). Besides a higher incidence of DNA losses, particularly DNA gains were statistically significant in invasive ductal carcinomas (6,2 per tumorcase vs. 2,7 per tumorcase). DNA gains on chromosome 1q and DNA losses on chromosomes 6q, 11q22-qter and 13q were investigated in >=40% of all tumor cases. DNA gains were observed with a higher frequency in invasive ductal carcinomas on 6p, 8q, 11q13, 16p, 17q, 19p/q and 20q. Additionally, DNA losses showed a higher incidence on 2q, 3p, 4p/q, 5q, 7p, 8p, 9q, 10q and 15q compared to invasive lobular carcinomas. In contrast, DNA losses on 16q, 17p, 18q and 22q reached statistical significance in invasive lubular carcinomas. Well-differentiated (G1) and poorly differentiated (G3) invasive ductal carcinomas reflected different genetic imbalances. Well-differentiated carcinomas are associated with DNA gains on 1q, 11q11-13, 16p and 20q, whereas poorly differentiated tumors showed additional DNA losses on 5q, 18q and 21q21. Furthermore, the investigation indicated that the average number of alterations is correlated also to the estrogen receptor content of the invasive ductal carcinomas. Tumors with estrogen receptor negative content showed a higher incidence of alterations on the following regions, i.e., DNA gains on 1p, 6p and 22q and DNA losses on 5q, 7p, 8p and 12q. Consequently, invasive ductal and invasive lobular carcinomas are characterized by distinct patterns of chromosomal alterations.