Immune cell dynamics deconvoluted by single-cell RNA sequencing in normothermic machine perfusion of the liver.

Normothermic machine perfusion (NMP) has emerged as an innovative organ preservation technique. Developing an understanding for the donor organ immune cell composition and its dynamic changes during NMP is essential. We aimed for a comprehensive characterization of immune cell (sub)populations, cell trafficking and cytokine release during liver NMP. Single-cell transcriptome profiling of human donor livers prior to, during NMP and after transplantation shows an abundance of CXC chemokine receptor 1+/2+ (CXCR1+/CXCR2+) neutrophils, which significantly decreased during NMP. This is paralleled by a large efflux of passenger leukocytes with neutrophil predominance in the perfusate. During NMP, neutrophils shift from a pro-inflammatory state towards an aged/chronically activated/exhausted phenotype, while anti-inflammatory/tolerogenic monocytes/macrophages are increased. We herein describe the dynamics of the immune cell repertoire, phenotypic immune cell shifts and a dominance of neutrophils during liver NMP, which potentially contribute to the inflammatory response. Our findings may serve as resource to initiate future immune-interventional studies.

Treatment with tetrahydrobiopterin overcomes brain death-associated injury in a murine model of pancreas transplantation.

Brain death (BD) has been associated with an immunological priming of donor organs and is thought to exacerbate ischemia reperfusion injury (IRI). Recently, we showed that the essential nitric oxide synthase co-factor tetrahydrobiopterin (BH4) abrogates IRI following experimental pancreas transplantation. We therefore studied the effects of BD in a murine model of syngeneic pancreas transplantation and tested the therapeutic potential of BH4 treatment. Compared with sham-operated controls, donor BD resulted in intragraft inflammation reflected by induced IL-1ß, IL-6, VCAM-1, and P-selectin mRNA expression levels and impaired microcirculation after reperfusion (p < 0.05), whereas pretreatment of the BD donor with BH4 significantly improved microcirculation after reperfusion (p < 0.05). Moreover, BD had a devastating impact on cell viability, whereas BH4-treated grafts showed a significantly higher percentage of viable cells (p < 0.001). Early parenchymal damage in pancreatic grafts was significantly more pronounced in organs from BD donors than from sham or non-BD donors (p < 0.05), but BH4 pretreatment significantly ameliorated necrotic lesions in BD organs (p < 0.05). Pretreatment of the BD donor with BH4 resulted in significant recipient survival (p < 0.05). Our data provide novel insights into the impact of BD on pancreatic isografts, further demonstrating the potential of donor pretreatment strategies including BH4 for preventing BD-associated injury after transplantation.

Donor pretreatment with tetrahydrobiopterin saves pancreatic isografts from ischemia reperfusion injury in a mouse model.

Depletion of the nitric oxide synthase cofactor tetrahydrobiopterin (H4B) during ischemia and reperfusion is associated with severe graft pancreatitis. Since clinically feasible approaches to prevent ischemia reperfusion injury (IRI) by H4B-substitution are missing we investigated its therapeutic potential in a murine pancreas transplantation model using different treatment regimens. Grafts were subjected to 16 h cold ischemia time (CIT) and different treatment regimens: no treatment, 160 µM H4B to perfusion solution, H4B 50 mg/kg prior to reperfusion and H4B 50 mg/kg before recovery of organs. Nontransplanted animals served as controls. Recipient survival and endocrine graft function were assessed. Graft microcirculation was analyzed 2 h after reperfusion by intravital fluorescence microscopy. Parenchymal damage was assessed by histology and nitrotyrosine immunohistochemistry, H4B tissue levels by high pressure liquid chromatography (HPLC). Compared to nontransplanted controls prolonged CIT resulted in significant microcirculatory deterioration. Different efficacy according to route and timing of administration could be observed. Only donor pretreatment with H4B resulted in almost completely abrogated IRI-related damage showing graft microcirculation comparable to nontransplanted controls and restored intragraft H4B levels, resulting in significant reduction of parenchymal damage (p < 0.002) and improved survival and endocrine function (p = 0.0002 each). H4B donor pretreatment abrogates ischemia-induced parenchymal damage and represents a promising strategy to prevent IRI following pancreas transplantation.

Overexpression of p150, a part of the large subunit of the eukaryotic translation initiation factor 3, in colon cancer.

BACKGROUND: P150, a 150 kDa protein, was isolated from virally and oncogene-transformed mouse cell lines, partially purified and cloned. P150 is part of the large subunit of the eukaryotic translation initiation factor 3 with sequence homology to centrosomin A. A significant correlation between p150 expression and malignancy in breast, cervical and esophageal cancer have recently been demonstrated. MATERIALS AND METHODS: Here, 110 colorectal carcinomas of different grades and stages, including lymph node and liver metastases were compared to adjacent normal mucosa by immunohistochemistry of P150. Western blot analysis of selected cases confirmed the expression levels determined by immunohistochemistry. Additionally, immuno-electron and laser scanning microscopy (LSM) was performed. RESULTS: All investigated carcinomas revealed high levels of p150 protein compared to normal adjacent mucosa. The staining intensity was slightly heterogeneous, and positivity was correlated to the tumor grade with statistically significant differences of p150 expression between normal and neoplastic mucosa (p<0.0001, Kruskal-Wallis test). Western blots confirmed higher expression levels of p150 in the tumor. Immunogold labelling and LSM investigation showed high expression levels of p150 on the rough endoplasmic reticulum and polyribosomes, indicating that p150 is translationally active in these tumors. CONCLUSION: Thus, we propose that p150 plays an important role in development and growth of colorectal carcinomas. Furthermore, p150 expression might provide us with reliable information on the biological behaviour of tumors and the clinical course of the disease.

An open-label, randomized phase II study of adecatumumab, a fully human anti-EpCAM antibody, as monotherapy in patients with metastatic breast cancer.

BACKGROUND: High-level expression of epithelial cell adhesion molecule (EpCAM) is associated with unfavorable prognosis in breast cancer. This study was designed to investigate two doses of the fully human IgG1 anti-EpCAM antibody adecatumumab (MT201) in patients with metastatic breast cancer (MBC). METHODS: A total of 109 patients were stratified into high- and low-level EpCAM expression by immunohistochemical staining of primary tumors and subsequently randomly assigned to receive monotherapy with either high- (6 mg/kg every two weeks (q2w)) or low-dose adecatumumab (2 mg/kg/ q2w) until disease progression. RESULTS: No complete or partial tumor responses could be confirmed by central RECIST assessment. The probability for tumor progression was significantly lower in patients receiving high-dose adecatumumab and expressing high levels of EpCAM (hazard ratio 0.43; P = 0.0057 versus low dose and low EpCAM). Three of 18 patients with highest EpCAM expression treated with adecatumumab developed new metastases up to week 6, compared with 14 of 29 patients with low EpCAM. Most frequent treatment-related adverse events (high dose/low dose) were chills (59%/20%), nausea (55%/18%), fatigue (39%/23%) and diarrhea (43%/7%). CONCLUSIONS: Single-agent adecatumumab shows dose- and target-dependent clinical activity in EpCAM-positive MBC, albeit no objective tumor regression. Further investigation of adecatumumab in patients with EpCAM-overexpressing tumors and lower tumor burden is warranted.

Practice of HER-2 immunohistochemistry in breast carcinoma in Austria.

Practice and accuracy of immunohistochemistry is known to vary highly. Reliability of HER-2 immunohistochemistry is critical because of its role in patient selection for therapeutical options in breast cancer. Therefore reliability of HER-2 immunohistochemistry in pathology laboratories in Austria was assessed. Ten tissue specimens of invasive ductal breast carcinomas and three cell line samples were tested. Presence/absence of gene amplification was determined by FISH to be used as a gold standard. Laboratories were asked to stain and assess slides using their routine immunohistochemical staining protocol. Overall the study consisted of 311 tests on tissue specimens and 142 on cell lines. In all cases manual scoring was performed. Participation was voluntary and was 94%. Overall sensitivity was 90.5% and specificity 99.2%. Overscoring including true false positive results were found in 6.7% and 6.3% in tissue specimens and cell lines, respectively. False negative determinations were obtained in 1.9% and 2.8% of tissue specimens and cell lines, respectively. HercepTest showed slightly higher reliability in comparison with individualized staining methods. By manual scoring inaccurate scoring affected 12.3% of test results and 62% of the laboratories. In conclusion participation rate and accuracy of HER-immunohistochemistry was high all over the country. Manually performed scoring demonstrated some limitations.

The effect of secretory leukocyte protease inhibitor (SLPI) on ischemia/reperfusion injury in cardiac transplantation.

We investigated the role of secretory leukocyte protease inhibitor (SLPI) in ischemia/reperfusion injury in cardiac transplantation. SLPI-/- mouse hearts and wild-type (WT) controls were transplanted immediately or after 10 h of cold ischemia (CI). Recombinant SLPI (rSLPI) was added to the preservation solution or given systemically. After evaluation of myocardial performance, grafts were investigated for histology, SLPI, TNF-alpha, TGF-beta, NF-kappaB and protease expression at indicated time points. Early myocardial contraction was profoundly impaired in SLPI-/- hearts exposed to CI and associated with high intra-graft protease expression. Systemic administration of rSLPI had no effect, however, when SLPI was added to the preservation solution, myocardial contraction was restored to normal. At 10 days, inflammation, myocyte vacuolization and necrosis were significantly more severe in SLPI-/- hearts. SLPI gene expression was detected in WT mice at 12 and 24 h and was significantly higher after CI. SLPI protein was observed at 24 h and 10 days. High intra-graft concentrations of SLPI after administration of rSLPI were inversely correlated with protease levels early and TGF-beta expression late after reperfusion. SLPI plays a crucial role in early myocardial performance and postischemic inflammation after cardiac transplantation. A dual inhibitory effect on protease and TGF-beta expression might be the underlying mechanism.

Ep-CAM expression in pancreatic and ampullary carcinomas: frequency and prognostic relevance.

AIMS: Pancreatic adenocarcinoma is an aggressive gastrointestinal malignancy with only a few long-term survivors even after radical surgery. Patients with ampullary cancer have a better prognosis but adjuvant therapy needs further improvement. Epithelial cell adhesion molecule (Ep-CAM) is strongly expressed in a variety of epithelial cancers and represents a promising target for immunological tumour therapy. Thus, the aim of this study was to investigate Ep-CAM expression and its potential prognostic impact in pancreatic and ampullary carcinomas. METHODS: Ep-CAM expression was investigated retrospectively by immunohistochemistry in paraffin-embedded primary tumour tissue samples from a series of consecutive patients with pancreatic (n = 153) and ampullary cancer (n = 34). RESULTS: Ep-CAM overexpression was observed in 85 of 153 pancreatic cancer specimens (56%) and in 29 of 34 ampullary cancer samples (85%). Overall, Ep-CAM failed to be an independent prognostic marker. However, subgroup analyses showed that Ep-CAM overexpression correlated with shorter overall survival among patients with ampullary cancer and advanced stage pancreatic cancer. In the latter subgroup, survival gradually worsened with increasing Ep-CAM scores. Furthermore, in ampullary cancer, Ep-CAM overexpression was found to correlate with tumour stage. CONCLUSIONS: Ep-CAM overexpression was detectable in the majority of cases with pancreatic and ampullary cancer. Therefore, Ep-CAM represents an attractive target for immune-based therapeutic interventions in these tumour entities. However, the prognostic value of Ep-CAM overexpression remains undetermined.

Intraorbital plexiform neurofibroma in an NF-1-negative patient.

A 52-year-old patient presented with an orbital swelling and exophthalmos that enlarged over a period of about 40 years. The clinical examination showed massive exophthalmos and ptosis of the right eye without diplopia. The radiological investigation (MRI, CT and ultrasound) showed an unclear intraorbital mass with erosion of the orbital floor, infraorbital rim and orbital roof. The lesion was diagnosed histologically as a plexiform neurofibroma. The patient did not present any features of neurofibromatosis type 1 (NF-1) and molecular genetic analysis was unable to uncover a pathogenic sequence alteration in the NF-1 gene. Owing to the absence of clinical and ophthalmologic symptoms and the improbability of complete removal, the patient refused surgical intervention.

The transcription factor ZEB1 (deltaEF1) promotes tumour cell dedifferentiation by repressing master regulators of epithelial polarity.

Epithelial to mesenchymal transition (EMT) is implicated in the progression of primary tumours towards metastasis and is likely caused by a pathological activation of transcription factors regulating EMT in embryonic development. To analyse EMT-causing pathways in tumourigenesis, we identified transcriptional targets of the E-cadherin repressor ZEB1 in invasive human cancer cells. We show that ZEB1 repressed multiple key determinants of epithelial differentiation and cell-cell adhesion, including the cell polarity genes Crumbs3, HUGL2 and Pals1-associated tight junction protein. ZEB1 associated with their endogenous promoters in vivo, and strongly repressed promotor activities in reporter assays. ZEB1 downregulation in undifferentiated cancer cells by RNA interference was sufficient to upregulate expression of these cell polarity genes on the RNA and protein level, to re-establish epithelial features and to impair cell motility in vitro. In human colorectal cancer, ZEB1 expression was limited to the tumour-host interface and was accompanied by loss of intercellular adhesion and tumour cell invasion. In invasive ductal and lobular breast cancer, upregulation of ZEB1 was stringently coupled to cancer cell dedifferentiation. Our data show that ZEB1 represents a key player in pathologic EMTs associated with tumour progression.

Lipocalin-2 regulates the inflammatory response during ischemia and reperfusion of the transplanted heart.

Ischemia and reperfusion (IR) are known to negatively affect early allograft function following solid organ transplantation. Lipocalin-2 (Lcn-2) has been described as a marker and potential positive modulator of acute inflammation during these processes. Using a heterotopic murine heart transplant model we previously found that IR resulted in a pronounced upregulation of Lcn-2 mRNA in the heart at 12 (22.7-fold increase) and 24 h (9.8-fold increase) of reperfusion. We now confirm this increase at the protein level and provide evidence for infiltrating polymorphonuclear cells as the primary source of Lcn-2 protein. Lcn-2 levels are increased 6.6-fold at 12 h, 11.4-fold at 24 h and 6.4 fold at 48 h after reperfusion. In Lcn-2(-/-) grafts the number of infiltrating granulocytes is reduced by 54% (p < 0.05) at 2 h, 79% (p < 0.01) at 12 h, 72% (p < 0.01) at 24 h and 52% (p < 0.01) at 48 h after reperfusion compared to Lcn-2(+/+) grafts, without any differences in cardiomyocyte apoptosis. These data suggest a function of Lcn-2 in the initiation of the inflammatory response. Moreover, an increase in Lcn-2 is not only restricted to the transplanted heart, but is also observed in the kidney, hinting at a possible involvement of Lcn-2 in the systemic response to IR.

Non-invasive monitoring of kidney allograft rejection through IDO metabolism evaluation.

The immunomodulatory enzyme indoleamine 2,3-dioxygenase (IDO) is activated by interferon-gamma (IFN-gamma) and via tryptophan depletion, suppresses adaptive T cell-mediated immunity in inflammation, host immune defense, and maternal tolerance. Its role in solid organ transplantation is still unclear. Therefore, we investigated the usefulness of IDO-mediated tryptophan catabolism in the evaluation of kidney allograft rejection. Blood, urine, and tissue samples were collected from 34 renal transplant patients without rejection and from nine patients with biopsy-confirmed episodes of acute rejection (n=12). Concentrations of kynurenine and tryptophan in serum and urine were analyzed by high-pressure liquid chromatography. Kynurenine to tryptophan ratio (kyn/trp) was calculated to estimate IDO activity. Immunostaining for IDO was performed on renal biopsies. Neopterin was assessed using radioimmunoassay. Kyn/trp and neopterin were detectable at low levels in serum of healthy volunteers and were increased in non-rejecting allograft recipients. Serum levels of kyn/trp were higher in recipients with rejection compared to non-rejectors as early as by day 1 post-surgery. Rejection episodes occurring within 13+/-5.9 days after transplantation were accompanied by elevated kyn/trp in serum (114+/-44.5 micromol/mmol, P=0.001) and urine (126+/-65.9 micromol/mmol, P=0.02) compared to levels during stable graft function. Kyn/trp correlated significantly with neopterin suggesting an IFN-gamma-induced increase in IDO activity. Immunostaining showed upregulation of IDO in rejection biopsies, localized in tubular-epithelial cells. Non-rejected grafts displayed no IDO expression. Acute rejection is associated with simultaneously increased serum and urinary kyn/trp in patients after kidney transplantation. Thus, IDO activity might offer a novel non-invasive means of immunomonitoring of renal allografts.

STAT-1 expression in human glioblastoma and peritumoral tissue.

BACKGROUND: Glioblastoma is a very aggressive brain tumour with poor prognosis despite radical surgery or radiotherapy. Signal transducers and activators of transcription (STAT) proteins are important elements in intracellular signalling and part of the JAK-STAT pathway. They are activated by growth factors and cytokines and translocate into the nucleus upon activation to exert their function as transcription factors. STAT-1 can be induced by interferons and has also been found to be important in sensitizing tumours to chemotherapeutic drugs. MATERIALS AND METHODS: Forty-six glioblastoma samples have been analysed for the expression of STAT-1 by immunohistochemistry. RESULTS: In our study performed by immunohistochemistry, 22 out of 46 glioblastomas (48%) were strongly positive for staining with a STAT-1 antibody, 9 (20%) showed an intermediate reactivity, 8 (17%) low immunoreactivity, and 7 (15%) were completely negative. In the tumour tissue, STAT-1 expression was mostly localized in the cytoplasm. This location of STAT-1 suggests the predominant presence of an inactive form of STAT-1. Tumour giant cells were frequently strongly stained. Part of the peritumoral brain tissue showed strongly positively reactive glial cells. Interestingly, within the infiltration area strong STAT-1 expression was found in reactive astrocytes, glia, and particularly in microglial components. CONCLUSION: The expression of STAT-1 in the majority of glioblastomas, together with its documented role in apoptosis and in the action of chemotherapeutic drugs on tumour cell lines point to a possible function of this protein in the response of glioblastomas to chemotherapy.

Tetrahydrobiopterin attenuates microvascular reperfusion injury following murine pancreas transplantation.

In this study we investigated the effect of tetrahydrobiopterin (BH4), an essential cofactor for nitric oxide synthases, on ischemia-reperfusion injury (IRI) following murine pancreas transplantation. Pancreatic grafts were exposed to prolonged cold ischemia times (CIT) and different treatment regimens: normal saline (S), S + 16 h CIT, BH4 50 mg/kg + 16 h CIT. Nontransplanted animals served as controls. Graft microcirculation was analyzed by means of functional capillary density (FCD) and capillary diameters (CD) after 2 h reperfusion using intravital microscopy. Quantification of inflammatory responses (mononuclear infiltration) and endothelial disintegration (edema formation) was done by histology (hematoxylin and eosin), and peroxynitrite formation assessed by nitrotyrosine immunostaining. FCD was significantly reduced after prolonged CIT, paralleled by increased peroxynitrite formation as compared with controls (all p < 0.05). Microcirculatory changes correlated significantly with intragraft peroxynitrite generation (Spearman: r = -0.56; p < 0.01). Pancreatic grafts treated with BH4 displayed markedly higher FCD values (p < 0.01) and abrogated nitrotyrosine staining (p = 0.03). CD were not significantly different in any group. Histology showed increased inflammation, interstitial edema, hemorrhage, acinar vacuolization and focal areas of necrosis after 16 h CIT, which was diminished by BH4 administration (p < 0.01). BH4 treatment significantly reduces post-ischemic deterioration of microcirculation as well as histologic damage and might be a promising novel strategy in attenuating IRI following pancreas transplantation.

The RANKL/OPG system is activated in inflammatory bowel disease and relates to the state of bone loss.

BACKGROUND AND AIMS: A substantial proportion of patients with inflammatory bowel disease (IBD) develops osteopenia and osteoporosis in the course of disease. Recent data from a mouse model of colitis suggest that the receptor activator of nuclear factor kappa B (RANKL)/osteoprotegerin (OPG) system may be responsible for bone loss. METHODS: We investigated the activation state of the RANKL/OPG system and its association with bone loss in human IBD. Plasma levels of OPG and RANKL were correlated with bone mineral density and current IBD therapy. Colonic secretion of OPG and RANKL and cell types responsible for such secretion were determined. RESULTS: OPG plasma levels were elevated 2.4-fold in Crohn's disease (CD) and 1.9-fold in ulcerative colitis (UC) whereas soluble RANKL (sRANKL) levels were not significantly different in IBD patients compared with healthy controls. High levels of OPG were released from colonic explant cultures (CEC) derived from inflamed IBD specimens, and colonic macrophages and dendritic cells costained for OPG. sRANKL levels from CEC were low both in IBD patients and healthy controls. Interestingly, increased expression of RANKL was mainly confined to cells in the lamina muscularis. A significant negative correlation was found between OPG plasma levels and femoral neck/lumbar spine bone mineral density. CONCLUSIONS: We have demonstrated that IBD is associated with alterations in the RANKL/OPG system. Applying results from a murine model of colitis associated bone loss, the constellation of OPG and sRANKL regulation observed in our study raises the possibility that RANKL/OPG may contribute to the development of bone loss in IBD.

Aberrant tetranectin expression in human breast carcinomas as a predictor of survival.

AIMS: Tetranectin (TN), a plasminogen kringle 4 binding protein, is thought to play a prominent role in the regulation of proteolytic processes via binding to plasminogen. The aim of this study was to evaluate the expression of TN in human breast cancer and adjacent normal breast tissue and to determine the impact of this expression on survival. METHODS: A retrospective analysis was performed on 189 patients with breast cancer, with a median follow up time of 10.6 years. The expression of TN was assessed in tumour tissue and adjacent normal breast tissue by immunohistochemistry, and the prognostic relevance of its expression in tumour cells was evaluated. RESULTS: TN was highly expressed in connective tissue fibres surrounding normal breast epithelium, but not in normal epithelial cells. High expression of TN in tumour cells was found in 131 (69%) of the tumour samples. By western blot analysis, no significant difference in the amount and molecular weight of TN was seen between tumour tissue and normal tissue. Strong TN immunoreactivity in tumour tissue was predictive of poor disease free and tumour specific overall survival. By multivariate analysis, high TN expression in cancer cells was an independent prognostic factor for disease free and tumour specific overall survival. CONCLUSIONS: Our results demonstrate differential TN expression in normal and malignant breast tissue and a prognostic impact of TN protein expression in breast carcinoma tissue. These data suggest a possible role of TN in invasiveness and the metastatic spread of human breast cancer.

Skin implant metastasis after percutaneous radio-frequency therapy of liver metastasis of a colorectal carcinoma.

Percutaneous radio-frequency ablation (RFA) of liver tumors has been reported to be an effective approach. Skin implant metastases have been described after RFA of hepatocellular carcinoma. A 56-year-old man underwent resection of the transverse colon for an adenocarcinoma (pT3N2M0) following by adjuvant chemotherapy. He developed multiple liver metastases and underwent RFA. Six weeks after RFA, the patient noticed a painful skin lesion at the entrance side of the probe in the right upper abdominal quadrant. Ultrasound examination and computed tomography scan revealed an intracutaneous tumor of 2-cm diameter. The tumor was excised and revealed a metastasis of the previously described adenocarcinoma. CPT-11 monotherapy was started; however, due to tumor progression, the patient died 35 months after colonic resection and 10 months after RFA. This is the first case of an implant skin metastasis after RFA of secondary liver tumors. Although RFA is a promising option in the palliation of such tumors, such rare complications have to be considered.

Distribution of DMT 1 within the human glandular system.

BACKGROUND & AIMS: DMT1 is a transmembrane protein which transports the divalent metal ions Fe2+, Zn2+, Cu2+ and Mn2+. Although DMT1 has been functionally linked to duodenal absorption and cellular utilisation of iron hardly anything is known about its distribution and potential role within the human glandular system. METHODS: Two polyclonal antibodies were raised to study the expression of DMT1 in tissues obtained from human corpus by the means of immunocytochemistry and Western blotting. RESULTS: All antibodies specifically detected a 60 kD protein band referring to human DMT1. Significant amounts of DMT1 expression were detected on the luminal site of organs, which are involved in excretion/re-absorption processes, such as salivary glands, pancreas, biliary tract and gallbladder. CONCLUSIONS: Our results suggest that DMT1 may be of pivotal importance for the regulation of metal ion homeostasis within organs involved in absorption and excretion of ions.

Her2/neu overexpression in differentiated thyroid carcinomas predicts metastatic disease.

To investigate the prognostic value of Her2/neu expression in differentiated thyroid carcinomas 103 patients were retrospectively investigated. All of them received surgical and an identical follow-up treatment. The patients with papillary and follicular thyroid cancer were further separated into two groups concerning their clinical development, including one group without distant metastasis (follow-up of minimum 8 years). The second group presented with distant metastases as a sign of an aggressive behaviour. Her2/neu was immunohistochemically detected on sections from formalin-fixed, paraffin-embedded tissues using c-erbB-2/Her-2/neu oncoprotein Ab-17 monoclonal antibody (mAb). In statistical analysis using the Mann-Whitney U-test and chi(2) test, Her2/neu protein overexpression was significantly correlated with prognosis. Both tumour entities without distant metastases showed significantly less cytoplasmic immunostaining than patients with development of metastases. Concerning the clinical outcome, Her2/neu overexpression may be regarded as a prognostic factor in differentiated thyroid carcinomas. Moreover, in addition to standard radio-iodine elimination therapy, application of Herceptin could lead to new successful therapeutic concepts for a number of patients with progressive thyroid cancer.