Wolfgang Glienke

Curcumin Inhibits Constitutive STAT3 Phosphorylation in Human Pancreatic Cancer Cell lines and Downregulation of Survivin/BIRC5 Gene Expression

ABSTRACT The purpose of this study was to determine the effect of curcumin on Survivin/BIRC5 and on the role of signal transducer and activator of transcription 3 (STAT3) activation in Survivin/ BIRC5. We incubated two pancreatic cancer cell lines with different amounts of curcumin. This resulted in a downregulation of proliferation in all cell lines tested. The expression of Survivin/BIRC5 on mRNA and protein level was significantly downregulated and the phosphorylation of STAT3 was blocked. Treatment of pancreatic cancer cells with curcumin resulted in an induction of apoptosis. The results indicate that curcumin inhibits several key factors in cancer cellular pathways and may be of interest in pancreatic cancer.

UROKINASE RECEPTOR LOCALIZATION IN BREAST CANCER AND BENIGN LESIONS ASSESSED BY IN SITU HYBRIDIZATION AND IMMUNOHISTOCHEMISTRY

Abstract The serine protease urokinase-type plasminogen activator (uPA) mediates cancer invasion and metastasis by binding to a cell surface receptor (uPA-R, CD87) on both tumor and stromal cells. In the present study we assessed uPA-R distribution in formalin-fixed, paraffin-embedded breast cancer specimens (n=50) and benign lesions (n=10) by immunohistochemistry employing a newly developed polyclonal chicken antibody to uPA-R (pAb HU277) in parallel with established monoclonal antibody (mAb) 3936 to uPA-R. In addition, uPA-R mRNA synthesis was investigated by in situ hybridization. In all of the sections analyzed, macrophage-like cells reacted with either antibody type. In 22 of the 50 cancer specimens, tumor cells reacted with pAb HU277 in contrast to mAb 3936 which only stained 9 of the 22 positive cases. Nevertheless, in 49 of the 50 cases, uPA-R mRNA was detected in cancer and in stromal cells by in situ hybridization suggesting posttranscriptional regulation of uPA-R expression in breast cancer cells. In 18 of 50 cases, uPA-R mRNA was also visualized in blood vessel lining endothelial cells by in situ hybridization and applying pAb HU277 in 14 of these 18 cases by immunohistochemistry. mAb 3936 did not stain any endothelial cells. pAb HU277 reacted with the breast gland epithelial cells of benign lesions as well, in contrast to mAb 3936 which did not. As for the cancer tissue, in benign lesions, endothelial cells were sporadically stained by pAb HU277. This antibody, but not mAb 3936, also stained myoepithelial cells in intraductal areas of invasive breast carcinoma. The results presented demonstrate the usefulness of pAb HU277 in locating uPA-R in tumor and normal cells with high sensitivity in formalin-fixed, paraffin-embedded breast tissue.

Wilms' tumour gene 1 (WT1) as a target in curcumin treatment of pancreatic cancer cells.

The transcription factor WT1 plays an important role in cellular proliferation and survival of various cancer cells, and is frequently expressed in pancreatic cancer. Curcumin has been shown to be a potentially effective agent in pancreatic cancer. In this context, the purpose of this study was to determine the role of WT1 in a curcumin-treated pancreatic cancer cell line. To study the effect of curcumin on the expression of WT1, we incubated the pancreatic cancer cell line PANC-1 with different amounts of curcumin. The expression of WT1 on mRNA and protein level was measured with real-time RT-PCR and Western blot analysis. The incubation of the pancreatic cancer cell line PANC-1 with curcumin resulted in an inhibition of cellular proliferation as measured with MTT assay. The expression of WT1 on mRNA and protein level was significantly down-regulated in a concentration-dependent manner after treatment with curcumin. The WT1 mRNA levels were decreased by 20%, 25%, 40%, 78% and 88% in response to 10, 20, 30, 40 and 50 microM curcumin. The use of small inhibitory RNA (siRNA) targeting WT1 down-regulated the expression of WT1 about 90%. Combined treatment with curcumin and siRNA targeting WT1 resulted in a significant inhibition of cell proliferation compared to curcumin-treated cells alone. In conclusion, WT1 is involved in cellular proliferation of PANC-1 cells. Targeting WT1 gene expression with siRNA may enhance the efficacy of curcumin to inhibit cell proliferation.

Serum Angiogenic Activity: Diagnostic Relevance in Renal Cell Carcinoma

OBJECTIVE Angiogenesis is essential for tumor growth and progression. However, reported data on angiogenic parameters in patients with renal cell carcinoma are contradictory. The objective of this study was to use serum to compare the systemic angiogenic activity in patients with renal cell carcinoma and to determine if pathologic stage and grade correlated to this angiogenesis parameter. METHODS Serum of 28 patients with a newly diagnosed renal cell carcinoma, 28 healthy volunteers and 9 patients with bladder carcinoma were used for this study. All sera were tested in a 72-hour endothelial cell proliferation assay. In addition the serum concentrations of the angiogenesis stimulators basic fibroblast growth factor (bFGF) and vascular endothelial growth factor (VEGF) were determined using standard ELISA assays. RESULTS The serum of renal cell carcinoma patients showed a median stimulation of human umbilical vein endothelial cells (HUVEC) of 89.79% (range 58.47-147.95%) and serum of healthy volunteers showed a median stimulation of 95.35% (range 74.64-141.77%) (p > 0.05). In contrast serum of patients with bladder carcinoma showed a median stimulation of 140.16% (range 64.82-200.16%) (p = 0.024). No correlations of the serum angiogenic activity and tumor stage or grade have been found in renal cell carcinoma patients. Furthermore, no correlations for serum bFGF and VEGF concentrations have been found. CONCLUSIONS Serum angiogenic activity of patients with renal cell carcinoma did not differ significantly from healthy controls, while serum of patients with bladder carcinoma showed a significant increase in endothelial cell stimulation. Furthermore, bFGF and VEGF serum concentrations did not correlate to serum angiogenic activity in patients with renal cell carcinoma. Therefore, the determination of systemic angiogenic parameters, in case of renal cell carcinoma, might not lead to adequate data concerning prognosis or therapeutic effects.

Targeting STAT3 signaling in pancreatic cancer promotes antiapoptotic gene expression.

Insulinomas are the most common type of functioning islet tumor of the pancreas and curable by surgical resection. The preoperative diagnosis of insulinomas has been challenging because of the small size of these tumors. There have been many reports of preoperative imaging examinations for localizing pancreatic insulinomas. Morganstein et al1 reported insulinoma detection rates for multiple imaging investigations, which included 43.5% of cases using computed tomography, 71% using magnetic resonance image, 86% using endoscopic ultrasound, 33% using octreotide scintigraphy, and 100% using arterial stimulation venous sampling (ASVS). In addition, Tseng et al stated that ASVS helps in the diagnosis of equivocal pancreatogenous hypoglycemia, indicating the pancreatic region of priority exploration and guiding a pancreatic resection. Selective ASVS was first introduced by Doppman et al in 1991. Positive results are derived from serum insulin gradient after stimulation with calcium injection into related arteries. Amore than 2-fold rise in the serum insulin concentration to the baseline concentration within 60 seconds after stimulation is considered to be positive for the pancreatic insulinoma. However, there is no case-control study in the series of reported literature. Therefore, we considered that a comparative study between patients with and without pancreatic insulinoma was necessary to establish adequate criteria for ASVS in patients with pancreatic insulinoma. In the present study, we retrospectively compared ASVS-derived indexes from patients with and without pancreatic insulinoma and determined the most reliable ASVS-derived indexes using statistical analyses and receiver operating characteristic curves (ROCs). MATERIALS AND METHODS

Laser microdissection and pressure catapulting (LMPC) in paraffin sections mounted on glass slides. A methodological report.

The technique of laser microdissection together with laser pressure catapulting (LMPC) is demonstrated in paraffin sections obtained from surgical specimens of brain tumors mounted on glass slides. A sufficient and precise application of microdissection techniques in tissue on glass slides is worthwhile, since it offers the possibility of a retrospective analysis of archived paraffin sections in histopathology. We could demonstrate a precise dissection of areas in tissues of different thicknesses (4 microm and 20 microm). Areas of tissue mounted directly on glass need to be dissected in a scanning mode in order to remove the total region in form of small tissue fragments row by row. This mode provided a precise microdissection of tissue areas of different sizes and shapes. A successful molecular biological analysis of the microdissected regions could be demonstrated. As an example for such an analysis, differential-PCR for detecting an amplification of the gene for the epidermal growth factor receptor (EGFR) was performed.