An intracellular targeted antibody detects EGFR as an independent prognostic factor in ovarian carcinomas.

BACKGROUND: In ovarian cancer, the reported rate of EGFR expression varies between 4-70% depending on assessment method and data on patient outcome are conflicting. METHODS: In this study we investigated EGFR expression and its prognostic value in a cohort of 121 invasive ovarian carcinomas, using a novel antibody against the intracellular domain of the receptor. We further evaluated an association between EGFR, the nuclear transporter CRM1 as well as COX-2. Furthermore, we evaluated EGFR expression in ten ovarian cancer cell lines and incubated cancer cells with Leptomycin B, a CRM1 specific inhibitor. RESULTS: We observed a membranous and cytoplasmic EGFR expression in 36.4% and 64% of ovarian carcinomas, respectively. Membranous EGFR was an independent prognostic factor for poor overall survival in ovarian cancer patients (HR 2.7, CI 1.1-6.4, p = 0.02) which was also found in the serous subtype (HR 4.6, CI 1.6-13.4, p = 0.004). We further observed a significant association of EGFR with COX-2 and nuclear CRM1 expression (chi-square test for trends, p = 0.006 and p = 0.013, respectively). In addition, combined membranous EGFR/COX-2 expression was significantly related to unfavorable overall survival (HR 7.2, CI 2.3-22.1, p = 0.001).In cell culture, we observed a suppression of EGFR protein levels after exposure to Leptomycin B in OVCAR-3 and SKOV-3 cells. CONCLUSIONS: Our results suggest that the EGFR/COX-2/CRM1 interaction might be involved in progression of ovarian cancer and patient prognosis. Hence, it is an interesting anti-cancer target for a combination therapy. Further studies will also be needed to investigate whether EGFR is also predictive for benefit from EGFR targeted therapies.

Solid neuroendocrine carcinomas of the breast: metastases or primary tumors?

Neuroendocrine breast carcinomas are rare but may represent either metastatic or primary lesions. So far, clinical and preoperative histopathological examinations do not distinguish properly between a primary or metastatic breast tumor. Due to any possible consequences following an appropriate treatment, markers which may be helpful for such a distinguishment are needed. We addressed this study in order to evaluate the immunohistochemical expression of GCDFP-15 and mammaglobin in a subset of pure neuroendocrine breast carcinomas (n = 9) and compared the expression profile with a cohort of non-mammary neuroendocrine tumors (n = 99). We observed in our study that solid neuroendocrine breast carcinomas are characterized by the expression of estrogen and progesterone receptors as well as GCDFP-15 and/or mammaglobin. GCDFP-15 was expressed in 6 out of 9 cases, mammaglobin was positive in 4 out of 9 tumors. In contrast, neuroendocrine tumors of the non-mammary cohort expressed neither GCDFP-15 nor mammaglobin. We conclude that mammaglobin and GCDFP-15 as markers of epithelial breast origin may work as a new and reliable diagnostic tool to distinguish primary endocrine tumors of the breast from a metastatic neuroendocrine disease. This is of utmost importance, especially for surgical management.

Expression of classical NF-kappaB pathway effectors in human ovarian carcinoma.

AIMS: Functional studies have demonstrated that nuclear factor (NF)-kappaB promotes tumour progression in ovarian cancer cells. However, surprisingly little is known of the expression of effectors of the NF-kappaB pathway in human ovarian cancer in vivo. METHODS AND RESULTS: Immunohistochemistry and in situ hybridization revealed that in a cohort of 85 primary ovarian carcinomas, total p65 expression was inversely correlated to nuclear and cytoplasmic phospho-IkappaBalpha (P = 0.002 and P = 0.05, respectively), and IkappaBalpha mRNA expression (P = 0.032). In contrast, phospho-p65 expression was paralleled by the expression of nuclear (P = 0.027) and cytoplasmic phospho-IkappaBalpha (P = 0.01). Total p65 expression was an adverse prognostic factor for overall survival (P = 0.018). In contrast, total IkappaBalpha and phosphorylated nuclear and cytoplasmic IkappaBalpha expression were favourable prognostic markers (P = 0.001, P = 0.031, P = 0.001, respectively). Cytoplasmic phospho-IkappaBalpha expression remained a significant prognostic factor on multivariate analysis (P = 0.010). In cultured, stimulated OVCAR-3 ovarian cancer cells the cytoplasmic retranslocation of p65 was delayed by inhibition of the nuclear membrane transporter chromosomal region maintenance/exportin1 protein (CRM1). A positive association of p65 and CRM1 expression was demonstrated in ovarian cancer tissue (P < 0.0001). CONCLUSIONS: Total and phosphorylated IkappaBalpha protein expression might serve as markers for NF-kappaB activation in human ovarian carcinoma. Cytoplasmic localization of p65 may be related to deregulated nucleocytoplasmic transport in carcinomas overexpressing CRM1.

A prognostic gene expression index in ovarian cancer - validation across different independent data sets.

Ovarian carcinoma has the highest mortality rate among gynaecological malignancies. In this project, we investigated the hypothesis that molecular markers are able to predict outcome of ovarian cancer independently of classical clinical predictors, and that these molecular markers can be validated using independent data sets. We applied a semi-supervised method for prediction of patient survival. Microarrays from a cohort of 80 ovarian carcinomas (TOC cohort) were used for the development of a predictive model, which was then evaluated in an entirely independent cohort of 118 carcinomas (Duke cohort). A 300-gene ovarian prognostic index (OPI) was generated and validated in a leave-one-out approach in the TOC cohort (Kaplan-Meier analysis, p = 0.0087). In a second validation step, the prognostic power of the OPI was confirmed in an independent data set (Duke cohort, p = 0.0063). In multivariate analysis, the OPI was independent of the post-operative residual tumour, the main clinico-pathological prognostic parameter with an adjusted hazard ratio of 6.4 (TOC cohort, CI 1.8-23.5, p = 0.0049) and 1.9 (Duke cohort, CI 1.2-3.0, p = 0.0068). We constructed a combined score of molecular data (OPI) and clinical parameters (residual tumour), which was able to define patient groups with highly significant differences in survival. The integrated analysis of gene expression data as well as residual tumour can be used for optimized assessment of the prognosis of platinum-taxol-treated ovarian cancer. As traditional treatment options are limited, this analysis may be able to optimize clinical management and to identify those patients who would be candidates for new therapeutic strategies.

Topoisomerase IIalpha mRNA and protein expression in ovarian carcinoma: correlation with clinicopathological factors and prognosis.

Topoisomerase IIalpha (Top IIalpha) is a nuclear enzyme that plays a central role in DNA metabolism, and is a molecular target for a variety of chemotherapeutic agents. Top IIalpha has recently gained attention as a biomarker for therapy response and patient survival. In this study, we attempted to assess the feasibility of measuring Top IIalpha gene expression in RNA, isolated from archival formalin-fixed paraffin-embedded tissue specimens, which are used routinely in pathology laboratories. We have employed a new technique on the basis of magnetic particles' separation and purification of nucleic acids, and evaluated both protein and mRNA expressions from the same routinely processed tissue blocks. We investigated the expression of Top IIalpha mRNA and protein by real-time reverse transcription polymerase chain reaction and immunohistochemistry, in a cohort of 133 primary ovarian carcinomas, and evaluated the association between Top IIalpha expression and clincopathological variables as well as patient outcome. Elevated Top IIalpha mRNA expression was observed in high-grade tumors (P=0.003) and advanced stage disease (P=0.011). In univariate Kaplan-Meier analysis, patients with higher expression of Top IIalpha nuclear protein had a significantly decreased overall survival (P=0.045). Interestingly, we detected cytoplasmic protein expression of Top IIalpha in a subset of samples. Cytoplasmic expression of Top IIalpha was associated with the expression of chromosomal region maintenance/exportin 1 (CRM1)-a nuclear export protein (P=0.008). Our study suggests that Top IIalpha overexpression is involved in the progression of ovarian cancer in a subset of the patients. Our results encourage the further evaluation of the prognostic and predictive values of Top IIalpha expression in ovarian carcinoma, which might help to assess the patients' risk profile, and the planning of an individualized therapy.

Expression of multidrug resistance-associated protein 1 in invasive ovarian carcinoma: implication for prognosis.

AIMS: Multidrug resistance is a major impediment in chemotherapeutic treatment of ovarian carcinoma patients. The aim of this study was to investigate the expression of multidrug resistance-associated protein 1 (MRP1) and to assess the possible associations with clinicopathological variables and patient outcome in primary ovarian carcinoma. METHODS AND RESULTS: Tumour specimens from 129 patients were obtained before chemotherapy and analysed by immunohistochemistry on tissue microarrays, and by real-time reverse transcriptase-polymerase chain reaction on RNA extracted from formalin-fixed paraffin-embedded tissue specimens using a new technique. Significantly increased MRP1 protein expression was observed in high-grade tumours (P = 0.005) and advanced International Federation of Gynaecology and Obstetrics stages (P = 0.036). On univariate Kaplan-Meier analysis, patients with higher expression of MRP1 protein had significantly decreased overall survival (P = 0.006). On multivariate Cox regression analysis, MRP1 protein expression retained its significance as an independent negative prognostic marker for overall survival (hazard ratio = 6.52, P = 0.003). Furthermore, MRP1 expression correlated with topoisomerase IIalpha expression both at mRNA and protein level (P < 0.001 and P = 0.023, respectively). CONCLUSION: In summary, in patients with primary ovarian cancer, overexpression of MRP1 is an adverse marker for patient outcome and cancer aggressiveness. Our data provide a translational basis for further clinical studies on the predictive value of MRP1 expression for response to chemotherapy.

High class I HDAC activity and expression are associated with RelA/p65 activation in pancreatic cancer in vitro and in vivo.

BACKGROUND: The strong association between aberrant HDAC activity and the occurrence of cancer has led to the development of a variety of HDAC inhibitors (HDIs), which emerge as promising new targeted anticancer therapeutics. METHODS: Due to the pivotal role of RelA/p65 in the tumorigenesis of pancreatic neoplasia we examined the expression of class I HDACs 1, 2 and 3 in a large cohort of human pancreatic carcinomas and correlated our findings with RelA/p65 expression status. Furthermore, we investigated the impact of the HDIs SAHA and VPA on RelA/p65 activity in pancreatic cancer cell culture models. RESULTS: Class I HDACs were strongly expressed in a subset of pancreatic adenocarcinomas and high expression was significantly correlated with increased nuclear translocation of RelA/p65 (p = 0.024). The link of HDAC activity and RelA/p65 in this tumor entity was confirmed in vitro, where RelA/p65 nuclear translocation as well as RelA/p65 DNA binding activity could be markedly diminished by HDI treatment. CONCLUSION: The RelA/p65 inhibitory effects of SAHA and VPA in vitro and the close relationship of class I HDACs and RelA/p65 in vivo suggest that treatment with HDIs could serve as a promising approach to suppress NF-kappaB activity which in turn may lead to enhanced apoptosis and chemosensitization of pancreatic cancers.

IMP3 expression in human ovarian cancer is associated with improved survival.

The insulin-like growth factor-II mRNA-binding protein IMP3 plays an important role in embryogenesis and recent reports suggest an involvement in tumorigenesis. Although IMP3 expression has been well studied in mouse and human fetal and adult gonads, its role in ovarian cancer is unknown. We investigated the expression of IMP3 at protein and mRNA levels in a cohort of primary ovarian carcinomas and in 11 ovarian cancer cell lines. Western blot analysis revealed an expression of IMP3 in all ovarian cancer cell lines and immunohistochemistry demonstrated a positive cytoplasmic staining in 32 of 68 carcinomas (47%). In contrast, epithelium of borderline tumors, as well as, benign ovarian lesions and normal ovaries exhibited only weak or no IMP3 expression. In univariate Kaplan-Meier analysis, IMP3 protein expression was significantly associated with better overall survival (P=0.048). To confirm these findings, we further determined IMP3 mRNA expression in 43 ovarian cancer specimens by real time quantitative reverse transcription-polymerase chain reaction. A significant correlation between protein and mRNA levels (r=0.414, P=0.006), as well as a correlation of IMP3 mRNA expression with patient overall survival (P=0.044), was observed. Our results demonstrate that IMP3 is expressed in a subpopulation of ovarian cancer and a marker of good prognosis.

Class I histone deacetylase expression has independent prognostic impact in human colorectal cancer: specific role of class I histone deacetylases in vitro and in vivo.

PURPOSE: Recently, several studies reported a strong functional link between histone deacetylases (HDAC) and the development of tumors of the large intestine. However, despite the importance of these molecules, comparably little is known on expression patterns and functions of specific HDAC isoforms in colorectal cancer. EXPERIMENTAL DESIGN: We characterized class I HDAC isoform expression patterns in a cohort of 140 colorectal carcinomas by immunohistochemistry. In addition, effects of HDAC inhibition by valproic acid and suberoylanilide hydroxamic acid, and specific HDAC isoform knockdown by short interfering RNA, were investigated in a cell culture model. RESULTS: We found class I HDACs highly expressed in a subset of colorectal carcinomas with positivity for HDAC1 in 36.4%, HDAC2 in 57.9%, and HDAC3 in 72.9% of cases. Expression was significantly enhanced in strongly proliferating (P = 0.002), dedifferentiated (P = 0.022) tumors. High HDAC expression levels implicated significantly reduced patient survival (P = 0.001), with HDAC2 expression being an independent survival prognosticator (hazard ratio, 2.6; P = 0.03). Short interfering RNA-based inhibition of HDAC1 and HDAC2 but not HDAC3 suppressed growth of colon cancer cells in vitro, although to a lesser extent than chemical HDAC inhibitors did. CONCLUSIONS: The strong prognostic impact of HDAC isoforms in colorectal cancer, the interactions of HDACs with tumor cell proliferation and differentiation in vivo, and our finding that HDACs are differentially expressed in colorectal tumors suggest that the evaluation of HDAC expression in clinical trials for HDAC inhibitors might help to identify a patient subgroup who will exceptionally profit from such a treatment.

Expression of the ELAV-like protein HuR in human prostate carcinoma is an indicator of disease relapse and linked to COX-2 expression.

The human ELAV-like protein HuR is involved in the stabilization of the mRNAs of a group of genes implicated in the regulation of cellular growth, angiogenesis and rapid inflammatory response. HuR is a nuclear shuttling protein, translocating bound mRNAs from the nucleus to the cytoplasm. We have previously observed an increased expression of cyclooxygenase-2 (COX-2) in prostate cancer while cell culture studies have shown that HuR stabilizes the mRNA of COX-2. Based on these mechanistic data, we aimed to investigate the role of HuR in prostate cancer by a tissue-based analysis combined with functional evaluation using a cell culture approach. Investigating 104 primary prostate carcinomas by immunohistochemistry, we found HuR expression to be shifted from a nuclear staining in normal prostate glands to a cytoplasmic staining in carcinoma tissue (p<0.0001). Cytoplasmic HuR expression was significantly correlated with COX-2 expression (p=0.005). Loss of nuclear HuR expression was an indicator of earlier PSA-relapse both in univariate (p=0.04) and multivariate survival analysis (p=0.04). HuR inhibition by Leptomycin B reduced the inducibility of COX-2 in PC-3 prostate cancer cells. We found that the subcellular localization of HuR is deregulated in a subset of prostate carcinomas, and that this deregulation is linked to an altered expression of the tumorigenic COX-2 protein as well as to an adverse patient prognosis. Our results point to a potential prognostic role of HuR expression in prostate cancer diagnostics and propose HuR as a future therapeutic target in prostate cancer therapy.

Cut-resistant protective gloves in pathology--effective and cost-effective.

Cutting injuries and needle-stitch injuries constitute a potentially fatal danger to both pathologists and autopsy personnel. We evaluated such injuries in a large German institute of pathology from 2002 to 2007 and analysed the effect of the introduction of cut-resistant gloves on the incidence of these injuries. In the observation period, 64 injuries (48 cutting injuries and 16 needle-stitch injuries) were noted in the injury report books. Most injuries were located at the non-dominant hand, preferentially at the index finger and the thumb. Around one fifths of the injuries were at the side of handedness. The average number of injuries per month was 1.22 for the 50 months prior to the introduction of cut-resistant gloves, more than seven times higher than after their introduction (0.158; 19 months; p < 0.001). Considering the medical and administrational costs of such injuries, cut-resistant protective gloves are an effective and cost-effective completion of personal occupational safety measures in surgical pathology and autopsy. We strongly recommend the use of such gloves, especially for autopsy personnel.

Activation of mTOR in a subgroup of ovarian carcinomas: correlation with p-eIF-4E and prognosis.

Ovarian carcinoma patients have an extremely poor prognosis; therefore, new molecular therapeutic approaches are urgently needed. The mTOR pathway, which may be targeted by substances such as Rapamycin or RAD001, is emerging as a promising target for anticancer therapy. So far, the expression and prognostic impact of mTOR signalling elements have not been completely studied in ovarian tumors. We analyzed p-mTOR, p-4E-BP1 and p-eIF-4E in 107 human ovarian lesions and observed an overexpression of p-mTOR (47%) and p-eIF-4E (56%) protein in primary ovarian carcinomas as compared to borderline tumors. Phospho-mTOR expression was significantly related to p-eIF-4E (p< or =0.001) and serous histological type (p=0.03). Increased p-4E-BP1 (31%) was associated with poor differentiation (p=0.04) and higher mitotic rate (p=0.004). In univariate analysis, increased expression of p-mTOR and p-eIF-4E was significantly associated with better overall survival (p=0.003, p=0.029). To connect the expression data with mechanistic studies, a set of 10 ovarian cancer cell lines was used. Expression of p-mTOR was increased in all cancer cell lines as compared to ovarian surface epithelial (HOSE) cells. Rapamycin treatment revealed a reduction of p-mTOR and p-4E-BP1 but increased p-AKT levels. We show for the first time an association of p-mTOR and p-eIF-4E with better overall survival for ovarian cancer patients. The combined results of our in vivo and cell culture studies suggest that a subpopulation of these patients may benefit from mTOR inhibition. The design of future clinical trials should incorporate biomarker testing to determine predictive markers for response to mTOR inhibitors.

Peroxisome proliferator-activated receptor gamma is highly expressed in pancreatic cancer and is associated with shorter overall survival times.

PURPOSE: Peroxisome proliferator-activated receptor gamma (PPARgamma) is a ligand-activated transcription factor that has been implicated in carcinogenesis and progression of various solid tumors, including pancreatic carcinoma. We aimed to clarify the expression patterns of PPARgamma in pancreatic ductal carcinomas and to correlate these to clinicopathologic variables, including patient survival. EXPERIMENTAL DESIGN: Array-based expression profiling of 19 microdissected carcinomas and 14 normal ductal epithelia was conducted. Additionally, Western blots of pancreatic cancer cell lines and paraffinized tissue of 129 pancreatic carcinomas were immunostained for PPARgamma. For statistical analysis, Fisher's exact test, chi2 test for trends, correlation analysis, Kaplan-Meier analysis, and Cox's regression were applied. RESULTS: Expression profiles showed a strong overexpression of PPARgamma mRNA (change fold, 6.9; P=0.04). Immunohistochemically, PPARgamma expression was seen in 71.3% of pancreatic cancer cases. PPARgamma expression correlated positively to higher pT stages and higher tumor grade. Survival analysis showed a significant prognostic value for PPARgamma, which was found to be independent in the clinically important subgroup of node-negative tumors. CONCLUSIONS: PPARgamma is commonly up-regulated in pancreatic ductal adenocarcinoma and might be a prognostic marker in this disease. Both findings corroborate the importance of PPARgamma in tumor progression of pancreatic cancer.

Acute myocardial infarction caused by coronary tumor thromboembolism: a rare primary manifestation of malignant tumor disease.

Myocardial infarction in the context of malignant tumor disease is rare but well documented and can occur through various pathophysiological mechanisms. We report an unusual case of a patient with recurrent myocardial infarction due to coronary tumor thromboembolism as the first clinical manifestation of a previously unknown squamous cell carcinoma of the lung. The possible pitfalls leading to a wrong preliminary diagnosis in our case as well as the literature on myocardial infarction in patients with cancer were discussed.

Erlotinib in patients with advanced non-small-cell lung cancer: impact of dose reductions and a novel surrogate marker.

Erlotinib is a relatively well-tolerated treatment option for patients with advanced non-small-cell lung cancer (NSCLC). Some patients suffer from severe skin toxicity or diarrhea, making dose reductions or even treatment cessation necessary. Recent clinical trials usually defined a 100 mg daily dose as the lowest acceptable dose, whereas little is known about the efficacy with lower doses. We retrospectively reviewed the files of all patients with advanced non-small-cell lung cancer (NSCLC) treated with erlotinib. We assessed demographic, disease- and treatment-related information. We tried to correlate tolerability with clinical efficacy. EGF receptor exon 18/19/21 mutations were analyzed in selected patients. Fifty-three patients with advanced non-small-cell lung cancer were treated with erlotinib. In nine patients (17%), the doses had to be reduced to 75 or 50 mg daily due to toxicity. We observed several sustained disease stabilizations in this subgroup. Patients suffering from paronychia with erlotinib had a significantly longer time to progression than did subjects without nail toxicity (P = 0.04). If patients were free from any toxicity, they were at high risk for early tumor progression (P = 0.001) and death. In patients with disease stabilization for 6 months or longer, we observed EGFR 18/19/21 wild type, exon 19 and exon 21 mutations. In conclusion, several patients required dose reductions during treatment with erlotinib. However, in tumors with sensitivity to erlotinib, even daily doses of 50-75 mg can result in sustained disease control. Paronychia represents a favorable surrogate marker for efficacy.

Prognostic impact of AMP-activated protein kinase expression in ovarian carcinoma: correlation of protein expression and GC/TOF-MS-based metabolomics.

AMP-activated protein kinase (AMPK) plays a central role in regulating energy metabolism in cells. AMPK activation results in down-regulation of anabolic pathways (e.g., fatty acid biosynthesis) and switches on catabolic processes such as glucose uptake, glycolysis or fatty acid oxidation. Recent studies in cell culture models have shown that the growth of tumor cell lines was inhibited by AMPK activation, but the expression of AMPK in human ovarian tumors has not been reported so far. In this study we investigated AMPK expression in a cohort of 70 ovarian carcinomas, 14 borderline tumors and 5 normal ovaries and linked the protein expression data to Gas chromatography/ time of flight mass spectrometry (GC/TOF-MS) based metabolomics. We observed a significantly higher expression in ovarian carcinomas compared to borderline tumors and normal ovaries (p=0.038). Decreased AMPK expression correlated significantly with higher tumor grade (p=0.009) and was of adverse prognosis in patients with advanced tumor stages (p=0.016) as well as in patients with serous ovarian carcinomas (p=0.037). GC/TOF-MS based metabolomics revealed a significantly higher concentration of glucose in AMPK-negative carcinomas (p=0.022) as well as overexpression of other metabolites from carbohydrate metabolism. Our results indicate a role for AMPK in progression of ovarian tumors and point towards a prognostic impact of AMPK expression for patient overall survival. Furthermore, our data suggest a deregulation of the AMPK-dependent energy metabolism in human ovarian carcinomas. In future clinical studies, activation of AMPK in ovarian carcinoma patients with advanced tumor stages might be an interesting therapeutic approach.

Flat epithelial atypia is a common subtype of B3 breast lesions and is associated with noninvasive cancer but not with invasive cancer in final excision histology.

The biological behavior and the optimal management of benign breast lesions with uncertain malignant potential, the so-called B3 lesions, found in breast needle core biopsies is still under debate. We addressed this study to compare histologic findings in B3 needle core biopsies with final excision specimens to determine associated rates of malignancy. Consecutive needle core biopsies were performed in a 3-year period (January 1, 2006-December 31, 2008). Biopsies were image-guided (31 by ultrasound, 85 stereotactic vacuum-assisted, 6 unknown) for evaluation of breast abnormalities. We reviewed 122 needle core biopsies with B3 lesions of 91 symptomatic patients and 31 screen-detected women and compared the B3 histologic subtypes with the final excision histology. A total of 1845 needle core biopsies were performed and B3 lesions comprised 6.6% of all B categories. The most common histologic subtype in biopsies was flat epithelia atypia in 35.2%, followed by papillary lesions in 21% and atypical ductal hyperplasia in 20%. Reports on excision specimens were available in 66% (81 patients). Final excision histology was benign in 73 (90.2%) and malignant in 8 (9.8%) patients (2 invasive cancer, 6 ductal carcinoma in situ). Of all B3 subtypes, atypical ductal hyperplasia and flat epithelial atypia were associated with malignancy, whereas only atypical ductal hyperplasia was accompanied by invasive cancer. Of all lesions, flat epithelial atypia was most frequently found in excision specimens (18%). In our study, flat epithelial atypia and atypical ductal hyperplasia are common lesions of the B3 category in needle core biopsies of the breast. Both lesions are associated with malignancy, whereas only atypical ductal hyperplasia was related to invasive cancer. We conclude that an excision biopsy after diagnosis of flat epithelial atypia is recommended depending on clinical and radiologic findings.

Down-regulation of the antigen processing machinery is linked to a loss of inflammatory response in colorectal cancer.

Antitumor inflammatory response is known to inhibit tumor growth in colorectal carcinoma. The density and functionality of tumor-infiltrating lymphocytes (TIL) is regulated by the antigen processing machinery through regulator proteins such as transporters associated with antigen processing (TAP) and major histocompatibility complex (MHC) class I antigen. We aimed to investigate the in vivo association of those factors and their impact on prognosis in colorectal cancer. TAP1, TAP2 and MHC class I antigen expression, inflammatory infiltrate and TIL (CD4(+), CD8(+), and CD20(+)) were assessed by immunohistochemistry in 336 sporadic colorectal carcinomas. The factors were correlated with each other and with clinic-pathological parameters and patient outcome. We found TAP1 and TAP2 expression to be significantly associated with MHC class I antigen expression (TAP1: r = 0.363, P < .001; TAP2: r = 0.393, P < .001). Increased density of CD8(+) TIL was predominantly found in TAP1, TAP2 and MHC class I antigen-positive cases. Increased density of CD4(+) TIL was linked with TAP1 and TAP2, but not with MHC class I antigen. High CD4(+) and CD8(+) cell count but not TAP1, TAP2 and MHC class I antigen expression had favorable prognostic impact in colorectal cancer (P = .003 and P = .003, respectively). In conclusion, our data show that the expression of key components of the antigen processing machinery is tightly linked to the density of TIL, which are positive prognostic factors in colorectal cancer in vivo. This implies that modulation of these factors may help to enhance antitumor inflammatory response which in turn may improve patient prognosis.

Phospho-mTOR and phospho-4EBP1 in endometrial adenocarcinoma: association with stage and grade in vivo and link with response to rapamycin treatment in vitro.

PURPOSE: Endometrial adenocarcinoma, due to a frequent activation of PI3 K/AKT has been proposed as a candidate neoplasm for the treatment with mTOR inhibitors. Yet, data on the expression of mTOR cascade components in endometrial cancer are lacking. METHODS: To provide a basis for futher studies with mTOR inhibitors, we used immunohistochemistry to evaluate the expression of activated mTOR pathway components in 57 endometrial cancer surgical specimens in vivo, and investigated in vitro the relation between the activation of AKT/mTOR and the response to rapamycin. RESULTS: p-mTOR expression was associated with nuclear p-4EBP1 expression (P = 0.02), and was more frequent in tumors extending ouside the uterine corpus (P = 0.011). Nuclear p-4EBP1 expression was increased in carcinomas of poor differentiation (P = 0.012). In cultivated PTEN-deficient Ishikawa cells, in addition to an activation of AKT, a phosphorylation of mTOR and 4EBP1 was evident, while PTEN-wild type HEC-1A cells lacked AKT activation but revealed a reduced expression of p-mTOR and p-4EBP1. Rapamycin induced a growth reduction, which was clearly more pronounced in Ishikawa cells than in HEC-1A cells (P < 0.03) and could be observed for up to 6 days. CONCLUSISONS: Expression of mTOR and 4EBP1 characterize high-grade, high-stage endometrial adenocarcinomas and might be predictive markers of a response to rapamycin. Based on our results, we suggest that the expression of elements of the mTOR pathway in human tumor tissue should be further evaluated as a possible predictive marker in large-scale clinical studies as well as translational research protocols in clinical studies with mTOR inhibitors.

Vascular endothelial growth factor C mRNA expression is a prognostic factor in epithelial ovarian cancer as detected by kinetic RT-PCR in formalin-fixed paraffin-embedded tissue.

Vascular endothelial growth factor C (VEGF-C) is a well described chemotactic and growth factor for lymphatic endothelial cells. Its inhibition leads to suppression of lymphatic and distant metastases in mouse models. In ovarian cancer, the relationship between VEGF-C expression and tumor behavior has not yet been determined by a quantitative method in vivo. Therefore, we used a new technique of RNA extraction from formalin-fixed paraffin-embedded tissue samples and determined the expression levels of VEGF-C mRNA in a study group of 97 ovarian cancer patients. Expression levels were correlated with clinicopathological features and patient survival. High VEGF-C expression was associated with worse overall (p = 0.0393) and progression-free (p = 0.0155) patient survival. In the subgroups of serous tumors and high-grade tumors, VEGF-C mRNA was still a negative indicator for patient survival (p = 0.0190 and 0.0311, respectively). A trend was observed among patients with high clinical stage (p = 0.0634). In multivariate survival analysis VEGF-C mRNA retained its prognostic influence on progression-free survival (p = 0.006, HR = 0.319 with a 95% confidence interval of 0.142-0.720). High VEGF-C expression was further associated with an increased residual tumor mass after primary cytoreductive surgery. We found no correlation of VEGF-C expression with tumor grade, FIGO stage, lymph node, or distant metastases. Our study demonstrates that high VEGF-C expression is associated with aggressive tumor behavior in ovarian cancer. mRNA extracted from paraffin-embedded tumor samples is suitable for VEGF-C gene expression studies.