Predictive scores in primary biliary cirrhosis: a retrospective single center analysis of 204 patients.

GOALS: The aim of this study was to assess the long-term outcome of primary biliary cirrhosis (PBC) patients and to test the clinical value of various outcome models, such as the Mayo Risk Score (MRS), in a large single-center cohort in Germany. BACKGROUND: PBC is a chronic autoimmune liver disease with a female gender predominance and a peak incidence in the fifth decade of life. PBC is characterized by portal inflammation and immune-mediated destruction of intrahepatic bile ducts in liver histology and the presence of antimitochondrial antibodies in the serum of nearly 95% of patients. In 5% to 20% of patients an overlap syndrome with autoimmune hepatitis (AIH) is diagnosed. Ursodeoxycholic acid is widely accepted as the standard medical treatment. STUDY: A total of 204 patients with PBC or PBC/AIH were retrospectively analyzed with regard to their clinical, biochemical, serological, and histologic features. PBC was diagnosed on the basis of the American Association for the Study of Liver Diseases criteria. Specific PBC scores, such as the MRS, the European and the Yale model, as well as nonspecific scores such as the Child-Pugh, the Model for End-stage Liver Disease, and Aspartate Aminotransferase to Platelet Ratio Index score were analyzed for their utility to predict the clinical outcome of patients. RESULTS: One hundred eighty-four patients with PBC alone and 20 with primary biliary cirrhosis/autoimmune hepatitis overlap were followed up for an average of 7.0 (range, 0.5 to 33.2) years. Importantly, baseline values of serum bilirubin, alkaline phosphatase, immunoglobulin M (IgM) and IgG, as well as antimitochondrial antibodies titers did not allow in properly predicting patient's outcome. The MRS proved clinical applicability. Patients with an R-value <6 did not develop liver-related complications. The Aspartate Aminotransferase to Platelet Ratio Index score had a significant correlation with the histologic degree of liver fibrosis, with limited value of scores between 1.0 and 1.5. Patients with a Model for End-stage Liver Disease score ≥8 (n=17) had a significantly higher risk to undergo liver transplantation or liver-related death. Outcome was less favorable than predicted by the European model. All scores showed low positive predictive values, limiting their applicability in clinical practice. CONCLUSIONS: Herein, we demonstrate that clinical risk scores in PBC should be interpreted with care. The MRS proved to be helpful to predict a favorable outcome. Novel approaches to predict outcome are needed to identify patients who may benefit from alternative, intensified treatment regimens.

Rectal cancer: dynamic contrast-enhanced MRI correlates with lymph node status and epidermal growth factor receptor expression.

PURPOSE: To evaluate correlations between dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) and clinicopathologic data as well as immunostaining of the markers of angiogenesis epidermal growth factor receptor (EGFR) and CXC-motif chemokine receptor 4 (CXCR4) in patients with rectal cancer. MATERIALS AND METHODS: Presurgical DCE-MRI was performed in 41 patients according to a standardized protocol. Two quantitative parameters (k21 , A) were derived from a pharmacokinetic two-compartment model, and one semiquantitative parameter (TTP) was assessed. Standardized surgery and histopathologic examinations were performed in all patients. Immunostaining for EGFR and CXCR4 was performed and evaluated with a standardized scoring system. RESULTS: DCE-MRI parameter A correlated significantly with the N category (P = 0.048) and k21 with the occurrence of synchronous and metachronous distant metastases (P = 0.029). A trend was shown toward a correlation between k21 and EGFR expression (P = 0.107). A significant correlation was found between DCE-MRI parameter TTP and the expression of EGFR (P = 0.044). DCE-MRI data did not correlate with CXCR4 expression. CONCLUSION: DCE-MRI is a noninvasive method which can characterize microcirculation in rectal cancer and correlates with EGFR expression. Given the relationship between the dynamic parameters and the clinicopathologic data, DCE-MRI data may constitute a prognostic indicator for lymph node and distant metastases in patients with rectal cancer.

The transcription factor IFN regulatory factor-4 controls experimental colitis in mice via T cell-derived IL-6.

The proinflammatory cytokine IL-6 seems to have an important role in the intestinal inflammation that characterizes inflammatory bowel diseases (IBDs) such as Crohn disease and ulcerative colitis. However, little is known about the molecular mechanisms regulating IL-6 production in IBD. Here, we assessed the role of the transcriptional regulator IFN regulatory factor-4 (IRF4) in this process. Patients with either Crohn disease or ulcerative colitis exhibited increased IRF4 expression in lamina propria CD3+ T cells as compared with control patients. Consistent with IRF4 having a regulatory function in T cells, in a mouse model of IBD whereby colitis is induced in RAG-deficient mice by transplantation with CD4+CD45RB(hi) T cells, adoptive transfer of wild-type but not IRF4-deficient T cells resulted in severe colitis. Furthermore, IRF4-deficient mice were protected from T cell-dependent chronic intestinal inflammation in trinitrobenzene sulfonic acid- and oxazolone-induced colitis. In addition, IRF4-deficient mice with induced colitis had reduced mucosal IL-6 production, and IRF4 was required for IL-6 production by mucosal CD90+ T cells, which it protected from apoptosis. Finally, the protective effect of IRF4 deficiency could be abrogated by systemic administration of either recombinant IL-6 or a combination of soluble IL-6 receptor (sIL-6R) plus IL-6 (hyper-IL-6). Taken together, our data identify IRF4 as a key regulator of mucosal IL-6 production in T cell-dependent experimental colitis and suggest that IRF4 might provide a therapeutic target for IBDs.

Confocal laser endomicroscopy for the diagnosis of urothelial bladder neoplasia: a technology of the future?

OBJECTIVE: To evaluate bladder urothelium by confocal laser endomicroscopy (CLE) and correlate microscopic findings with standard histopathology. PATIENTS AND METHODS: Fresh bladder urothelium tissue specimens were topically stained with acriflavine for instantaneous microscopic imaging. A single-line laser in a handheld CLE probe delivered an excitation wavelength of 488 nm providing a high resolution of 0.7 µm and an adjustable imaging depth of 0-250 µm. Resection specimens of 18 patients were investigated with 1000-fold magnification and ex vivo findings were compared with targeted histopathology (haematoxylin and eosin staining). RESULTS: Typical tumour growth patterns such as altered nuclear to cytoplasmic ratio, pseudopapillar tissue stratification and neoangiogenesis were readily visible. Nuclear and subnuclear architecture of healthy bladder tissue could be discriminated against neoplastic tissue. CONCLUSIONS: In addition to white light cystoscopy, CLE is a promising novel tool for the in vivo microscopic visualization of bladder cancer; first results of the present study show its potential to define microscopic characteristics of bladder cancer tissue. Further in vivo studies are necessary to determine sensitivity and specificity of the technique.

The recipient CYP2D6 allele 4-associated poor metabolizer status correlates with an early fibrosis development after liver transplantation.

CYP2D6 is part of the cytochrome P450 system, which catalyzes biotransformation of endogenous substrates and xenobiotics. Approximately 10% of the Caucasian population has two null alleles, resulting in a poor metabolizer (PM) status. Mostly, allele four (CYP2D6*4) is responsible for the PM status, which is suspected to be associated with an accelerated fibrosis progression (FP). The aim of the present study was to analyze the role of the CYP2D6*4 genotype for FP after liver transplantation (LT). Genotypes were determined in liver biopsies (donor) and peripheral blood (recipient) by fluorescence resonance energy transfer. Data were correlated with clinical variables and risk factors for fibrosis. We analyzed 517 LTs performed between 1997 and 2009. Overall donor and recipient allele frequencies were comparable (18.0%, 20.5%; P = 0.43). The donor genotype did not correlate with FP. In contrast, recipients carrying CYP2D6*4, showed a significant higher risk for an accelerated FP (P = 0.011) in HCV positive (P = 0.038) and HCV negative patients (P = 0.033). Results were confirmed by multivariate analysis (Hazard ratio 1.65; P = 0.001). The CYP2D6*4-associated PM status of the donor liver seems to have no influence on FP after LT. Recipients, carrying the allele, have an elevated risk for an accelerated FP.

Diagnostic efficiency of differentiating small cancerous and precancerous lesions using mucosal brush smears of the oral cavity--a prospective and blinded study.

The aim of this study was to evaluate the diagnostic accuracy of oral brush biopsy to identify early malignancy. One hundred and eighty-six brush biopsies of suspicious mucosal lesions were obtained, haematoxilin and eosin (H&E)-stained and compared with the histology of conventional excision biopsies of the same site performed concomitantly. The sensitivity for identifying squamous cell carcinoma (SCC) was 88.5%. High-risk lesions including squamous intraepithelial neoplasia (SIN II, SIN III) and SCC were identified with a sensitivity of 86.4%, using a pap-analogous classification, which is considered to be carcinomatous, as well as moderate and severe dysplastic cells positive. Depending on the cytopathologic definition for malignancy and the tumour size, the test accuracy varied: Extending the cytopathologic criteria for malignancy by defining all dysplastic or malignant cytopathologic findings as positive, the sensitivity was increased to 95.2% at the expense of the specificity, which was reduced from 94.9% to 82.3%. Separately analysing SCCs of less than 20 mm, the sensitivity was reduced by 9.5% to 78%. Although small malignant lesions seem to be less reliable by the conventional oral brush biopsy, it is a useful screening instrument for early diagnosis of suspicious, epithelial lesions and could therefore contribute to improved cancer prognosis.

Effectiveness of autofluorescence to identify suspicious oral lesions--a prospective, blinded clinical trial.

Regular screening through white light inspection of the entire oral mucosa is the most important examination method to identify precancerous lesions and early oral carcinoma. Additionally, the physiologic autofluorescence of the oral mucosa has been described as a novel screening method for the detection of mucosal lesions that are not visible by white light. This study aimed to evaluate the sensitivity and specificity of the autofluorescence examination. Seventy-eight patients were examined in this study. All of them suffered from suspicious oral mucosal lesions. Two different investigation methods were applied: the standard examination by white light and an examination by a novel light source of 400 nm that evoked a green light emission (>500 nm) in normal mucosa. It was proposed that malignant oral mucosal lesions show different autofluorescence characteristics than the green autofluorescence of healthy mucosa. Red autofluorescence indicated SCC with a sensitivity of 20% and a specificity of 98%. The results showed that dysplasia and carcinoma could be identified with a sensitivity of 96% and a specificity of 18% by using the autofluorescence method. The sensitivity decreased according to the grade of mucosal keratosis and was influenced by the localisation of the lesion. In conclusion, benign as well as malignant oral lesions could not be distinguished by a diminished autofluorescence signal. A red autofluorescence signal, however, could indicate cancerous processes of the oral mucosa.

Selective activation of trophoblast-specific PLAC1 in breast cancer by CCAAT/enhancer-binding protein beta (C/EBPbeta) isoform 2.

The trophoblast-specific gene PLAC1 (placenta-specific 1) is ectopically expressed in a wide range of human malignancies, most frequently in breast cancer, and is essentially involved in cancer cell proliferation, migration, and invasion. Here we show that basal activity of the PLAC1 promoter is selectively controlled by ubiquitous transcription factor SP1 and isoform 2 of CCAAT/enhancer-binding protein beta that we found to be selectively expressed in placental tissue and cancer cells. Binding of both factors to their respective elements within the PLAC1 promoter was essential to attain full promoter activity. Estrogen receptor alpha (ERalpha) signaling further augmented transcription and translation of PLAC1 and most likely accounts for the positive correlation between PLAC1 expression levels and the ERalpha status we observed in primary breast cancer specimens. DNA affinity precipitation and chromatin immunoprecipitation assays revealed that transactivation of the PLAC1 promoter by ligand-activated ERalpha is based on a nonclassical pathway independent of estrogen-response elements, by tethering of ERalpha to DNA-bound CCAAT/enhancer-binding protein beta-2, and SP1. Our findings provide first insight into a novel and hitherto unknown regulatory mechanism governing selective activation of trophoblast-specific gene expression in breast cancer.

Liver-specific Ldb1 deletion results in enhanced liver cancer development.

BACKGROUND & AIMS: LIM-domain-binding (Ldb) proteins have been demonstrated to be essential not only to key embryonic developmental processes but also to carcinogenesis. We have previously demonstrated Ldb1 to be of high biological and developmental relevance, as a targeted deletion of the Ldb1 gene in mice results in an embryonic lethal and pleiotropic phenotype. METHODS: We have now established a liver-specific Ldb1 knock out to investigate the role of Ldb1 in carcinogenesis, in particular in hepatocellular carcinoma (HCC) development, in vivo. RESULTS: These mice demonstrated a significantly enhanced growth of liver cancer by means of tumor size and number, advocating for an essential role of Ldb1 in HCC development. In addition, proliferation and resistance against apoptosis were increased. In order to identify the functional disturbances due to a lack of Ldb1, we performed a 15k mouse gene microarray expression analysis. We found the Myc oncogene to be regulated in the microarray analysis and were able to further confirm this regulation by demonstrating an over-expression of its downstream target Cyclin D1. Furthermore, we were able to demonstrate a down-regulation of the tumor suppressor p21. Finally, the liver stem cell marker EpCAM was also identified to be over expressed in Ldb1(-/-) knock out mice. CONCLUSIONS: We have established a significant role of Ldb1 in cancer development. Furthermore, we provided evidence for a myc/cyclin D1, p21, and EpCAM-dependent signalling to be key downstream regulators of this novel concept in HCC development.

Low expression of chemokine receptor CCR5 in human colorectal cancer correlates with lymphatic dissemination and reduced CD8+ T-cell infiltration.

BACKGROUND: Chemokines and their receptors have been proposed to distinctly contribute to tumor growth, dissemination, and local immune escape. The aim of this study was to evaluate the relevance of the chemokine receptor CCR5 expression for the progression of human colorectal cancer. METHODS: CCR5 expression was assessed by RT-PCR analysis in 103 colorectal cancer patients. Intensity of CCR5 expression was correlated with both tumor and patient characteristics. Infiltration of tumor margins with CD8(+) T cells in the context of CCR5 expression was analyzed by immunohistochemistry in additional 18 colorectal cancer specimens. RESULTS: Human colorectal cancer revealed variable intensities of CCR5 expression ranging from absent (48/103: 47%), weak (30/103: 29%), intermediate (13/103: 13%), to strong (12/103: 12%). Absent or weak CCR5 expression was significantly associated with advanced UICC stages (P=0.02) and lymphatic metastasis (P=0.05). In addition, CCR5 expression positively correlated with CD8(+) T-cell infiltration in tumor margins (P=0.001). CONCLUSION: In summary, intermediate and strong CCR5 expression was significantly associated with nonmetastatic colorectal cancer and increased CD8(+) T-cell infiltration.

Diagnostic accuracy of brush biopsy-based cytology for the early detection of oral cancer and precursors in Fanconi anemia.

BACKGROUND: Individuals with Fanconi anemia (FA) have a 500-fold to 700-fold elevated risk, much earlier onset, and limited therapeutic options for oral squamous cell carcinoma (SCC) compared with the general population. The early detection of SCC, or preferably its precursors, is mandatory to retain curative therapeutic options. Due to frequent synchronic and metachronic oral lesions, tissue biopsies, as usually recommended by guidelines, often are not feasible. In the current study, an alternative strategy for early detection using oral brush biopsy-based cytology was validated regarding its diagnostic accuracy. METHODS: Over a 12-year period, the oral cavities of a large cohort of 713 individuals with FA were inspected systematically and brush biopsy-based cytology of 1233 visible oral lesions was performed. In cases of inconclusive cytology, analysis of DNA ploidy was performed whenever possible. The results were correlated to a long-term clinicopathological follow-up reference standard. RESULTS: A total of 737 lesions were suitable for statistical analysis, including 86 lesions with at least high-grade oral epithelial dysplasia in 30 patients. For cytology, the sensitivity and specificity were 97.7% and 84.5%, respectively. Additional analysis of DNA ploidy increased the sensitivity and specificity to 100% and 92.2%, respectively. CONCLUSIONS: Careful inspection of the oral cavity of individuals with FA followed by brush biopsy-based cytology appears to identify visible oral, potentially malignant and malignant lesions that warrant treatment. Approximately 63% of SCC and precursor lesions are detected at a noninvasive or early stage. Negative cytology or a lack of DNA aneuploidy can exclude high-grade oral epithelial dysplasia or SCC with high accuracy and thus reduce the need for invasive diagnostic biopsies.

Impact of several histopathological prognosticators and local tumour extension on oncological outcome in pT3b/c N0M0 renal cell carcinoma.

OBJECTIVE: To investigate the prognostic relevance of different histopathological features and local tumour extension in patients with pT3b/c N0M0 renal cell carcinoma (RCC), as recently new proposals of reclassifying tumour fat invasion in pT3b/c RCC have been made but the effect of other histopathological tumour characteristics and combinations thereof with tumour invasion has yet to be determined in these patients. PATIENTS AND METHODS: Between 1990 and 2006, 1943 patients underwent surgical treatment for renal tumours in our institution, of which 175 patients (8.7%) had pT3b/c RCC. After exclusion of 57 patients (32.6%) with lymph node and/or distant metastases at the time of diagnosis, 118 (67.4%) remained for retrospective analysis. Different histopathological features and local tumour extension were studied for their association with cancer-specific-survival (CSS) and progression-free-survival (PFS) by univariate and multivariate analyses. Histopathology was reviewed and revised according to the 2002 Tumour-Nodes-Metastasis (TNM) classification system by one pathologist (S.B.). CSS and PFS were estimated by the Kaplan-Meier method. RESULTS: Follow-up data were obtained from 110 patients at a median (range) of 3.2 (0.3-16.1) years. In univariate analysis, microvascular invasion (MVI) and capsular invasion increased the risk of tumour progression by 2.05- and 2.72-times (P = 0.037 and P < 0.001). Overall, tumour fat invasion (TFI) and the presence of areas composed by cells with eosinophilic cytoplasm were associated with a higher risk of progression (P = 0.001 and P = 0.011) and reduced CSS (P = 0.037 and P = 0.017). In multivariate analysis, MVI and capsular invasion were associated with a two-fold increased risk of dying from cancer (hazard risk ratio, HR 2.22, P = 0.045 and HR 2.31, P = 0.011). TFI in general (P = 0.004) and specifically coexistent perirenal fat invasion (PFI) and renal sinus fat invasion (RSFI) were associated with a three-fold increased risk of developing tumour progression (HR 3.36, P = 0.001). The 10-year CSS and PFS rates were 39% and 36% for all patients, 47% and 45% for pT3b/c RCC with no PFI or RSFI, and 25% and 10% for PFI + RSFI. CONCLUSION: Patients with pT3b/c RCC with MVI, capsular invasion, TFI and especially PFI + RSFI, have a markedly reduced prognosis compared with patients with pT3b/c RCC without these features. When these results are corroborated by additional studies and external validation, modification of the TNM classification system would be a sensible consequence.

DNA image cytometry in the differential diagnosis of benign and malignant lesions of the bile duct, the pancreatic duct and the papilla of Vater.

UNLABELLED: The value of DNA image cytometry in the differential diagnosis of benign and malignant epithelial lesions was tested analyzing 174 specimens from the biliary tract, the pancreatic ducts and the papilla of Vater. PATIENTS AND METHODS: Cytologic specimens (n=45) or monolayer smears, prepared from three 50 microm thick sections by a cell separation technique (n=129), were stained according to Feulgen. The DNA content of 250 epithelial cells, chosen per random, was determined using a TV-image analysis system CM-1 (Hund, Wetzlar, Germany). The DNA content of 30 lymphocytes served as an internal standard for the normal diploid value in every individual case. Different DNA cytometric parameters and the mean nuclear area were calculated. RESULTS: In comparison with the benign cases (n=120), the mean values of all DNA cytometric variables were markedly increased in the malignant (n=54) (p<0.001). However, a certain overlay of the data distribution had to be considered. Requiring a specificity of >or=95%, sensitivity values up to 87% (mean ploidy) could be obtained. An aneuploid DNA stemline was present in 76% of the tumor cases. The occurrence of 9cEEs were completely restricted to malignant specimens (specificity: 100%), but only present in 50% of the tumor cases. As an additional finding in some benign cases, DNA polyploidy up to 8c could be demonstrated (11/120, 9.2%). CONCLUSION: DNA image cytometry represents a relevant tool in the objective identification of malignant changes of specimens of the biliary tract, the pancreatic duct or the papilla of Vater, if suitable variables are used for DNA data interpretation. However, mainly due to the fact that several cases reveal only slight ploidy changes, a well-preserved DNA ploidy profile does not exclude malignant changes.

Tissue microarrays are reliable tools for the clinicopathological characterization of lung cancer tissue.

BACKGROUND: The advantage of tissue microarray (TMA) is its ability to efficiently analyze large numbers of tissue specimens in a methodologically uniform way. The reliability of TMAs, especially with regard to clinicopathological characterizations, when compared to conventional immunohistochemistry (IHC) was evaluated. MATERIALS AND METHODS: Seventy-two embedded tissue sections from lung cancer specimens were stained with monoclonal antibodies against the tumor-associated markers TA-MUC1 and Lewis Y. Three representative cores of every tumor were embedded in a paraffin array multiblock. The IHC was evaluated by the immunoreactive score (IRS). RESULTS: The data for the TMA IHC and the conventional IHC were concordant (kappa > or = 80%) for both markers. Likewise, discordance (McNemar's test) was low, and sensitivity and specificity were above 80% for both markers. In the samples with high positive expression, the concordance increased (kappa > or = 90%), discordance disappeared (McNemar p = 1.0), and sensitivity and specificity increased above 90% for both markers. Using Cox regression models, all the clinicopathological dependencies were equivalent for both techniques and both markers. CONCLUSION: Immunohistochemistry with tissue microarrays is valid and provides results equivalent to conventional immunohistochemistry with respect to expression patterns and clinicopathological characterizations.

Automated detection of cancer cells in effusion specimens by DNA karyometry.

BACKGROUND: The average sensitivity of conventional cytology for the identification of cancer cells in effusion specimens is only approximately 58%. DNA image cytometry (DNA-ICM), which exploits the DNA content of morphologically suspicious nuclei measured on digital images, has a sensitivity of up to 91% for the detection of cancer cells. However, when performed manually, to our knowledge to date, an expert needs approximately 60 minutes for the analysis of a single slide. METHODS: In the current study, the authors present a novel method of supervised machine learning for the automated identification of morphologically suspicious mesothelial and epithelial nuclei in Feulgen-stained effusion specimens. The authors compared this with manual DNA-ICM and a gold standard cytological diagnosis for 121 cases. Furthermore, the authors retrospectively analyzed whether the amount of morphometrically abnormal mesothelial or epithelial nuclei detected by the digital classifier could be used as an additional diagnostic marker. RESULTS: The presented semiautomated DNA karyometric solution identified more diagnostically relevant abnormal nuclei compared with manual DNA-ICM, which led to a higher sensitivity (76.4% vs 68.5%) at a specificity of 100%. The ratio between digitally abnormal and all mesothelial nuclei was found to identify cancer cell-positive slides at 100% sensitivity and 70% specificity. The time effort for an expert therefore is reduced to the verification of a few nuclei with exceeding DNA content, which to our knowledge can be accomplished within 5 minutes. CONCLUSIONS: The authors have created and validated a computer-assisted bimodal karyometric approach for which both nuclear morphology and DNA are quantified from a Feulgen-stained slide. DNA karyometry thus increases the diagnostic accuracy and reduces the workload of an expert when compared with manual DNA-ICM.

Loss of LLGL1 Expression Correlates with Diffuse Gastric Cancer and Distant Peritoneal Metastases.

Background: Loss of LLGL1 has been associated with loss of cellular adhesion and dissemination of cells from colorectal cancer and malignant melanoma. Regulation and relevance of LLGL1 were analyzed in gastric cancer patients with lymphatic and distant dissemination. Furthermore, LLGL1 expression was analyzed in relation to the cellular adhesion protein E-cadherin. Methods: LLGL1 and E-cadherin transcription levels were evaluated in 56 gastric cancer patients and five gastric cancer cell lines. IHC staining for LLGL1 was performed on 39 gastric cancer specimens. LLGL1 was stably transfected into LLGL1 negative gastric cancer cell line SNU16 (del(17) (p11.2)) for functional in vitro assays and a xenograft bioassay. Results: Gastric cancer specimens and cell lines displayed LLGL1 and E-cadherin expression levels with variable intensity. In gastric mucosa, LLGL1 exhibited weak cytoplasmic and strong cortical staining. Loss of LLGL1 expression occurred in 65% of gastric cancers and significantly correlated with loss of E-cadherin expression (P=0.00009). Loss of LLGL1 expression was associated with the diffuse type of gastric cancer (P=0.029) with peritoneal carcinomatosis (M1; P=0.006) and with female gender (P=0.017). Stable reexpression of LLGL1 in SNU16 cells significantly increased both plastic surface adhesion and extracellular matrix proteins laminin and fibronectin, but had no impact on in vitro proliferation, apoptosis, or invasion or on in vivo proliferation or differentiation in our xenograft bioassay. Conclusion: LLGL1 is coexpressed with E-cadherin. Loss of expression of either protein is associated with diffuse gastric cancer and peritoneal metastases. LLGL1 does not impact on proliferation or epithelial-mesenchymal transition (EMT) rather increasing cellular adhesion.

Pseudodominant inheritance of goitrous congenital hypothyroidism caused by TPO mutations: molecular and in silico studies.

CONTEXT AND OBJECTIVE: Most cases of goitrous congenital hypothyroidism (CH) from thyroid dyshormonogenesis 1) follow a recessive mode of inheritance and 2) are due to mutations in the thyroid peroxidase gene (TPO). We report the genetic mechanism underlying the apparently dominant inheritance of goitrous CH in a nonconsanguineous family of French Canadian origin. DESIGN, SETTING, AND PARTICIPANTS: Two brothers identified by newborn TSH screening had severe hypothyroidism and a goiter with increased (99m)Tc uptake. The mother was euthyroid, but the father and two paternal uncles had also been diagnosed with goitrous CH. After having excluded PAX8 gene mutations, we hypothesized that the underlying defect could be TPO mutations. RESULTS: Both compound heterozygous siblings had inherited a mutant TPO allele carried by their mother (c.1496delC; p.Pro499Argfs2X), and from their father, one brother had inherited a missense mutation (c.1978C-->G; p.Gln660Glu) and the other an insertion (c.1955insT; p.Phe653Valfs15X). The thyroid gland of one uncle who is a compound heterozygote for TPO mutations (p.Phe653Valfs15X/p.Gln660Glu) was removed because of concurrent multiple endocrine neoplasia type 2A. Immunohistochemistry revealed normal TPO staining, implying that Gln660Glu TPO is expressed properly. Modeling of this mutant in silico suggests that its three-dimensional structure is conserved, whereas the electrostatic binding energy between the Gln660Glu TPO and its heme group becomes repulsive. CONCLUSION: We report a pedigree presenting with pseudodominant goitrous CH due to segregation of three different TPO mutations. Although goitrous CH generally follows a recessive mode of inheritance, the high frequency of TPO mutations carriers may lead to pseudodominant inheritance.

The host defence peptide LL-37/hCAP-18 is a growth factor for lung cancer cells.

Cancer development can be viewed as dysregulated repair. Antimicrobial peptides (AMPs) are effector molecules of the innate immune system with direct antimicrobial activity. Beside this host defence function several AMPs play a role in the regulation of inflammation and tissue repair. The aim of the present study was to investigate whether the human cathelicidin AMP LL-37/hCAP-18 is involved in the biology of lung cancer. Human cancer cell lines were found to express the human cathelicidin LL-37/hCAP-18 mRNA and peptide at different levels. Immunohistochemistry of human lung cancers showed that the peptide is expressed mostly in adenocarcinoma and squamous cell carcinoma. Application of exogenous LL-37 at low concentrations of 5ng/ml to cancer cell lines increased proliferation and growth of anchorage-independent colonies. At the molecular level, LL-37 induced phosphorylation of the epidermal growth factor receptor (EGFR) and activation of downstream MAP kinase signalling pathways. Lung cancer cell lines that stably overexpress the peptide by means of a doxycycline-regulated promoter system also showed a faster growth. When these cell lines were injected subcutaneously into nude mice, cathelicidin overexpression resulted in increased tumourigenicity and the formation of significantly larger tumours. In conclusion, cathelicidin is expressed in human lung cancers. The peptide activates tumour cells resulting in increased cell growth in vitro and in an animal model. The host defence peptide cathelicidin LL-37/hCAP-18 acts as growth factor for human lung cancer.

PDGFRalpha/beta expression correlates with the metastatic behavior of human colorectal cancer: a possible rationale for a molecular targeting strategy.

As new multi-target tyrosine kinase inhibitors are emerging in the therapy of various malignancies, our aim was to define the co-expression pattern of receptor-tyrosine-kinase platelet-derived growth factor receptors alpha and beta (PDGFRalpha/beta) in human colorectal cancer. The co-expression pattern of PDGFRalpha/beta was analyzed by RT-PCR in 99 histologically confirmed human colorectal carcinomas and five colorectal cancer cell lines. In addition, immunohistochemical (IHC) staining was applied for confirmation of expression and analysis of receptor tyrosine kinase (RTK) localisation. The colorectal cancer cell lines that were analysed revealed varying expression intensities of PDGFRalpha and PDGFRbeta. The majority of human colorectal cancer specimens revealed a PDGFRalpha (83%) or PDGFRbeta (60%) expression. While PDGFRalpha showed a predominantly cytoplasmic staining in tumor cells as well as in stromal pericytes, PDGFRbeta was restricted to stromal pericytes only. Furthermore, PDGFRalpha expression significantly correlated with lymph node metastasis (P=0.0082) and advanced UICC stages III/IV (P=0.018) in older patients (P=0.043). PDGFRbeta expression only revealed a trend towards lymphatic dissemination (P=0.099). Co-expression of PDGFRalpha/beta occurred in 57% of the colorectal cancer samples, whereas another 29% of the samples depicted mono-expression of PDGFRalpha or PDGFRbeta. Notably, PDGFRalpha/beta expression significantly correlated with lymphatic metastasis (P=0.007) and advanced UICC stages III/IV (P=0.017) in older patients (P=0.03). In summary, our results revealed that PDGFRalpha/beta expression significantly correlates with lymphatic dissemination and therefore encourages application of PDGFRalpha/beta RTK-inhibitors within a combination therapy.

Co-expression of receptor tyrosine kinases in esophageal adenocarcinoma and squamous cell cancer.

This study aimed to define the co-expression pattern of target receptor tyrosine kinases (RTKs) in human esophageal adenocarcinoma and squamous cell cancer. The co-expression pattern of vascular endothelial growth factor receptor (VEGFR)1-3, platelet-derived growth factor receptor (PDGFR)alpha/beta and epidermal growth factor receptor 1 (EGFR1) was analyzed by RT-PCR in 50 human esophageal cancers (35 adenocarcinomas and 15 squamous cell cancers). In addition, IHC staining was applied for the confirmation of the expression and analysis of RTK localisation. The adenocarcinoma samples revealed VEGFR1 (97%), VEGFR2 (94%), VEGFR3 (77%), PDGFRalpha (91%), PDGFRbeta (85%) and EGFR1 (97%) expression at different intensities. Ninety-four percent of the esophageal adenocarcinomas expressed at least four out of six RTKs. Similarly, squamous cell cancers revealed VEGFR1 (100%), VEGFR2 (100%), VEGFR3 (53%), PDGFRalpha (100%), PDGFRbeta (87%) and EGFR1 (100%) expression at different intensities. All esophageal squamous cell carcinomas expressed at least four out of six RTKs. While VEGFR1-3 and PDGFRalpha and EGFR1 was expressed by tumor cells, PDGFRbeta was restricted to stromal cells, which also depicted a PDGFRalpha expression. Our results revealed a high rate of RTK co-expression in esophageal adenocarcinoma and squamous cell cancer and may encourage application of multi-target RTK inhibitors within a multimodal concept as a promising novel approach for innovative treatment strategies.