Circulating tumour cells as a biomarker for diagnosis and staging in pancreatic cancer.

BACKGROUND: Current diagnosis and staging of pancreatic ductal adenocarcinoma (PDAC) has important limitations and better biomarkers are needed to guide initial therapy. We investigated the performance of circulating tumour cells (CTCs) as an adjunctive biomarker at the time of disease presentation. METHODS: Venous blood (VB) was collected prospectively from 100 consecutive, pre-treatment patients with PDAC. Utilising the microfluidic NanoVelcro CTC chip, samples were evaluated for the presence and number of CTCs. KRAS mutation analysis was used to compare the CTCs with primary tumour tissue. CTC enumeration data was then evaluated as a diagnostic and staging biomarker in the setting of PDAC. RESULTS: We found 100% concordance for KRAS mutation subtype between primary tumour and CTCs in all five patients tested. Evaluation of CTCs as a diagnostic revealed the presence of CTCs in 54/72 patients with confirmed PDAC (sensitivity=75.0%, specificity=96.4%, area under the curve (AUROC)=0.867, 95% CI=0.798-0.935, and P<0.001). Furthermore, a cut-off of ⩾3 CTCs in 4 ml VB was able to discriminate between local/regional and metastatic disease (AUROC=0.885; 95% CI=0.800-0.969; and P<0.001). CONCLUSION: CTCs appear to function well as a biomarker for diagnosis and staging in PDAC.

Bioimpedance phase angle indicates catabolism in Type 2 diabetes.

AIMS: Body cell mass is directly proportional to the bioimpedance phase-angle which is an indicator of the amount of electrical charge that cell membranes can hold and is an index of cellular health and function. To evaluate whether the bioimpedance phase angle is relevant for indicating catabolism in people with diabetes and whether it discriminates between people with diabetes receiving different types of therapy. METHODS: A cross-sectional study was performed in 182 people with Type 2 diabetes and 107 age- and BMI-matched control subjects. The phase angle was measured at 5, 50 and 100 kHz using multifrequency bioimpedance analysis. The phase angles were compared among different diabetes therapy groups (untreated patients with diabetes, patients receiving oral antidiabetic drugs and patients receiving insulin therapy). RESULTS: The phase angle at 100 kHz strongly correlated with total body potassium (r = 0.70, P = 0.001), and was therefore a good indicator of body cell mass. The phase angle at 100 kHz discriminated more strongly between patients with Type 2 diabetes and control subjects than did the phase angle at 50 kHz. Compared with control subjects, patients with Type 2 diabetes had a smaller phase angle at 100 kHz (men: 5.2° vs. 4.5°, P < 0.0001; women: 4.8° vs. 4.2°, P < 0.0001) and a smaller phase angle at 50 kHz (men: 5.9° vs. 5.3°, P < 0.0001; women: 5.4° vs. 4.8°, P = 0.0001), but a larger phase angle at 5 kHz (men: 2.0° vs. 2.6°, P = 0.0001; women: 2.3° vs. 3.0°, P = 0.00001). Phase angle ratios better discriminated between patients and control subjects than phase angles alone (phase angle at 5 kHz/ phase angle at 50 kHz ratio, P = 1.51 × 10(-16) ; phase angle at 5kHz/phase angle at 100 kHz ratio, P = 2.13 × 10(-15) ). No differences were found among phase angles in the different therapy groups. In patients with diabetes, the phase angle at 50 kHz and the phase angle at 100 kHz correlated inversely with duration of disease (men: P = 0.026, P = 0.016; women: only phase angle at 100 kHz, P = 0.003) and with HbA1c concentration (men: P = 0.010, P = 0.001; women: P = 0.007, P = 0.043). CONCLUSIONS: The phase angle at 100 kHz is a promising measurement for assessing catabolic state in people with diabetes.

PET lung ventilation/perfusion imaging using (68)Ga aerosol (Galligas) and (68)Ga-labeled macroaggregated albumin.

Pulmonary imaging using ventilation/perfusion (V/P) single-photon emission tomography (V/P scan) with Tc-99m-labeled radiotracers is a well-established diagnostic tool for clinically suspected pulmonary embolism (PE). Ga-68 aerosol (Galligas) and Ga-68-labeled macroaggregated albumin (MAA) are potential tracers for positron emission tomography (PET) lung V/P imaging and could display an advantage over conventional V/P scans in terms of sensitivity and specificity. After radiochemical and animal studies, the clinical applicability of Ga-68 aerosol (Galligas) and Ga-68-labeled MAA was investigated in an exploratory study in patients with clinical suspicion of PE. PET scans were acquired using a 16-slice Gemini TF positron emission tomography/computed tomography (PET/CT) scanner. The acquisition protocol included low-dose computed tomography (CT) for attenuation correction (AC). Dosimetry calculations and continuative phantom measurements were performed. Structural analyses showed no modification of the particles due to the labeling process. In addition, in vitro experiments showed stability of Ga-68 MAA in various media. As expected, Ga-68-labeled human serum albumin microspheres (HSAM) were completely retained in the lung of the animals. In clinical use, PET lung ventilation and perfusion imaging using Ga-68 aerosol (Galligas) and Ga-68-labeled MAA was successful in all cases. In one case a clinically suspected PE could be detected and verified. The administered activity of Ga-68 aerosol (Galligas) and Ga-68-labeled MAA may be reduced by more than 50%, resulting in comparable radiation exposure to conventional V/P scans. In conclusion, Ga-68 aerosol (Galligas) and Ga-68-labeled MAA are efficient substitutes for clinical use and could be an interesting alternative with high accuracy for lung V/P imaging with Tc-99m-labeled radiotracers, especially in times of Mo-99 shortages and increasing use and spread of PET/CT scanners and Ga-68 generators, respectively.

Automatic brain response to facial emotion as a function of implicitly and explicitly measured extraversion.

Extraversion/introversion is a basic dimension of personality that describes individual differences in social behavior and sensory sensitivity. Previous neuroimaging research exclusively relied on self reports for assessing personality traits. In recent years, implicit measures of personality have been developed that aim at assessing the implicit self-concept of personality and complement self report instruments which are thought to measure aspects of the explicit self-concept of personality. In the present study functional magnetic resonance imaging was used to examine automatic brain reactivity to facial expression as a function of both implicitly and explicitly measured extraversion in 30 healthy women. Sad, happy, and neutral faces were presented for 33 ms masked by neutral faces beside a no face control condition. Subjects evaluated the briefly shown neutral mask faces. The Implicit Association Test (IAT) and the NEO Five-Factor Inventory (NEO-FFI) were applied as measures of extraversion which were not correlated in our sample. IAT extraversion was negatively correlated with automatic reactivity of the caudate head, thalamus, and inferior frontal cortex to sad faces. NEO-FFI extraversion was negatively correlated with response of the inferior frontal cortex and putamen to sad faces. For masked happy faces, an inverse correlation of the IAT effect for extraversion with activation of the caudate head and superior parietal lobule was observed. NEO-FFI extraversion was inversely correlated with the response of the thalamus to happy faces. Neither NEO-FFI extraversion nor IAT effect were significantly related to brain response to masked neutral faces (compared to the no face condition). Taken together, a specific heightened responsivity of the fronto-striatal-thalamic circuit to facial emotions which are arousing stimuli might underlie introverts' preference for avoiding social interactions. Research on the neurobiology of extraversion could benefit from the application of implicit in addition to explicit measurement instruments when automatic neural responses are investigated.

Laparoscopic radioisotope-guided sentinel lymph node mapping and excision of the rectum--an experimental study.

BACKGROUND: Patients with a low-risk T1 rectal carcinoma can undergo the therapy of a local excision. In these patients the lymph node (LN) status remains unknown. There is a potential risk of up to 7% for nodal metastasis. To investigate the possibility of using the sentinel lymph node (SLN) concept, an experimental study on pigs was undertaken. The objective was to laparoscopically identify and extract SLNs from the rectum using a radioisotope (RI). METHODS: The experiment was conducted in 30 pigs, since the sample size calculation indicated that with 30 animals a two-sided 95% confidence interval for a single proportion using the large sample normal approximation would extend at most 0.107 from the observed proportion of 0.9. One milliliter of a mixture of the RI Technetium 99 m (Tc99 m) and patent blue V dye was administered in the rectum endoscopically and after the lapse of 1 h, we laparoscopically identified and excised all SLNs using a laparoscopic gamma camera probe. RESULTS: We found in all operated pigs (n = 30) at least one SLN (lymph node with highest measured counts per second (cps)). In mean we detected 1.6 SLN (range one to three SLNs). In 28 cases, the SLN concept was successful. Sensitivity for detecting SLNs was 93% (n = 28/30), the probe count rate ranged from 600-10,000 cps with a median of 3,800. CONCLUSION: Minimal invasive mapping and excision of SLN of the rectum using a RI is feasible. The sensitivity for detecting SLN was high (93%). The application of this procedure on humans seems to be possible.

Risk of malignancy in resected cystic tumors of the pancreas < or =3 cm in size: is it safe to observe asymptomatic patients? A multi-institutional report.

Recent international consensus guidelines propose that cystic pancreatic tumors less than 3 cm in size in asymptomatic patients with no radiographic features concerning for malignancy are safe to observe; however, there is little published data to support this recommendation. The purpose of this study was to determine the prevalence of malignancy in this group of patients using pancreatic resection databases from five high-volume pancreatic centers to assess the appropriateness of these guidelines. All pancreatic resections performed for cystic neoplasms < or =3 cm in size were evaluated over the time period of 1998-2006. One hundred sixty-six cases were identified, and the clinical, radiographic, and pathological data were reviewed. The correlation with age, gender, and symptoms (abdominal pain, nausea and vomiting, jaundice, presence of pancreatitis, unexplained weight loss, and anorexia), radiographic features suggestive of malignancy by either computed tomography, magnetic resonance imaging, or endoscopic ultrasound (presence of solid component, lymphadenopathy, or dilated main pancreatic duct or common bile duct), and the presence of malignancy was assessed using univariate and multivariate analysis. Among the 166 pancreatic resections for cystic pancreatic tumors < or =3 cm, 135 cases were benign [38 serous cystadenomas, 35 mucinous cystic neoplasms, 60 intraductal papillary mucinous neoplasms (IPMN), 1 cystic papillary tumor, and 1 cystic islet cell tumor], whereas 31 cases were malignant (14 mucinous cystic adenocarcinomas and 13 invasive carcinomas and 4 in situ carcinomas arising in the setting of IPMN). A greater incidence of cystic neoplasms was seen in female patients (99/166, 60%). Gender was a predictor of malignant pathology, with male patients having a higher incidence of malignancy (19/67, 28%) compared to female patients (12/99, 12%; p < 0.02). Older age was associated with malignancy (mean age 67 years in patients with malignant disease vs 62 years in patients with benign lesions (p < 0.05). A majority of the patients with malignancy were symptomatic (28/31, 90%). Symptoms that correlated with malignancy included jaundice (p < 0.001), weight loss (p < 0.003), and anorexia (p < 0.05). Radiographic features that correlated with malignancy were presence of a solid component (p < 0.0001), main pancreatic duct dilation (p = 0.002), common bile duct dilation (p < 0.001), and lymphadenopathy (p < 0.002). Twenty-seven of 31(87%) patients with malignant lesions had at least one radiographic feature concerning for malignancy. Forty-five patients (27%) were identified as having asymptomatic cystic neoplasms. All but three (6.6%) of the patients in this group had benign disease. Of the patients that had no symptoms and no radiographic features, 1 out of 30 (3.3%) had malignancy (carcinoma in situ arising in a side branch IPMN). Malignancy in cystic neoplasms < or =3 cm in size was associated with older age, male gender, presence of symptoms (jaundice, weight loss, and anorexia), and presence of concerning radiographic features (solid component, main pancreatic duct dilation, common bile duct dilation, and lymphadenopathy). Among asymptomatic patients that displayed no discernable radiographic features suggestive of malignancy who underwent resection, the incidence of occult malignancy was 3.3%. This study suggests that a group of patients with small cystic pancreatic neoplasms who have low risk of malignancy can be identified, and selective resection of these lesions may be appropriate.

Unfavourable prognosis associated with K-ras gene mutation in pancreatic cancer surgical margins.

BACKGROUND: Despite intent to cure surgery with negative resection margins, locoregional recurrence is common in pancreatic cancer. AIMS: To determine whether detection of K-ras gene mutation in the histologically negative surgical margins of pancreatic cancer reflects unrecognised disease. PATIENTS: Seventy patients who underwent curative resection for pancreatic ductal adenocarcinoma were evaluated. METHODS: All patients had surgical resection margins (pancreatic transection and retroperitoneal) that were histologically free of invasive cancer. DNA was extracted from these paraffin embedded surgical margins and assessed by quantitative real time polymerase chain reaction to detect the K-ras gene mutation at codon 12. Detection of K-ras mutation was correlated with standard clinicopathological factors. RESULTS: K-ras mutation was detected in histologically negative surgical margins of 37 of 70 (53%) patients. A significant difference in overall survival was demonstrated between patients with margins that were K-ras mutation positive compared with negative (median 15 v 55 months, respectively; p = 0.0008). By univariate and multivariate analyses, detection of K-ras mutation in the margins was a significant prognostic factor for poor survival (hazard ratio (HR) 2.8 (95% confidence interval (CI) 1.5-5.3), p = 0.0009; and HR 2.8 (95% CI 1.4-5.5), p = 0.004, respectively). CONCLUSIONS: Detection of cells harbouring K-ras mutation in histologically negative surgical margins of pancreatic cancer may represent unrecognised disease and correlates with poor disease outcome. The study demonstrates that molecular-genetic evaluation of surgical resection margins can improve pathological staging and prognostic evaluation of patients with pancreatic ductal adenocarcinoma.

PET imaging with yttrium-86: comparison of phantom measurements acquired with different PET scanners before and after applying background subtraction.

Quantitative imaging with the positron emitter (86)Y is the method of choice to determine the uptake and dosimetry of (90)Y-labelled radiopharmaceuticals. To examine the quantitative accuracy of positron emission tomography findings with (86)Y, this non-pure positron emitter was evaluated in a cylindrical phantom with rods of Teflon, water and air and measured with three different scanners: ECAT EXACT (2D/3D), ECAT HR+ (2D/3D) and PC4096+ (2D). After standard reconstruction, (86)Y radioactivity measured with the ECAT EXACT and related to the true radioactivity varied between 0.84 and 0.99 in 2D and between 0.93 and 1.20 in 3D from the first to the last acquisition (eight half-life times later). The water and Teflon rods exhibited considerable amounts of reconstructed radioactivity-21% in 2D and 67% in 3D for water and 65% and 147%, respectively, for Teflon-compared with the actual (86)Y radioactivity of the phantom. For the ECAT HR+ similar results were obtained in 3D, but there were even greater overestimations in 2D. Measurements with the PC4096+ showed rather small errors, with 10% for water and 20% for Teflon. To correct for the background of gamma-coincidences, sinograms were analysed and an experimental percentage of the background was subtracted from the sinograms. In order to minimise the errors in reconstructed radioactivity, the subtraction value had to be different for the individual scanners and modes. Our results demonstrate that (90)Y/(86)Y-based dosimetry for bone and red marrow must be regarded with caution if it is derived from regions of interest over the bone, the density of which is similar to that of Teflon. To obtain more reliable estimates, an appropriate background correction must be applied and tailored individually with respect to the scanner and acquisition mode.

Preliminary data on biodistribution and dosimetry for therapy planning of somatostatin receptor positive tumours: comparison of (86)Y-DOTATOC and (111)In-DTPA-octreotide.

The somatostatin analogue (90)Y-DOTATOC (yttrium-90 DOTA- D-Phe(1)-Tyr(3)-octreotide) is used for treatment of patients with neuroendocrine tumours. Accurate pretherapeutic dosimetry would allow for individual planning of the optimal therapeutic strategy. In this study, the biodistribution and resulting dosimetric calculation for therapeutic exposure of critical organs and tumour masses based on the positron emission tomography (PET) tracer (86)Y-DOTATOC, which is chemically identical to the therapeutic agent, were compared with results based on the tracer commonly used for somatostatin receptor scintigraphy, (111)In-DTPA-octreotide (indium-111 DTPA- D-Phe(1)-octreotide, OctreoScan). Three patients with metastatic carcinoid tumours were investigated. Dynamic and static PET studies with 77-186 MBq (86)Y-DOTATOC were performed up to 48 h after injection. Serum and urinary activity were measured simultaneously. Within 1 week, but not sooner than 5 days, patients were re-investigated by conventional scintigraphy with (111)In-DTPA-octreotide (110-187 MBq) using an equivalent protocol. Based on the regional tissue uptake kinetics, residence times were calculated and doses for potential therapy with (90)Y-DOTATOC were estimated. Serum kinetics and urinary excretion of both tracers showed no relevant differences. Estimated liver doses were similar for both tracers. Dose estimation for organs with the highest level of radiation exposure, the kidneys and spleen, showed differences of 10.5%-20.1% depending on the tracer. The largest discrepancies in dose estimation, ranging from 23.1% to 85.9%, were found in tumour masses. Furthermore, there was a wide inter-subject variability in the organ kinetics. Residence times (tau(organs)) for (90)Y-DOTATOC therapy were: tau(liver) 1.59-2.79 h; tau(spleen) 0.07-1.68 h; and tau(kidneys) 0.55-2.46 h (based on (86)Y-DOTATOC). These data suggest that dosimetry based on (86)Y-DOTATOC and (111)In-DTPA-octreotide yields similar organ doses, whereas there are relevant differences in estimated tumour doses. Individual pretherapeutic dosimetry for (90)Y-DOTATOC therapy appears necessary considering the large differences in organ doses between individual patients. If possible, the dosimetry should be performed with the chemically identical tracer (86)Y-DOTATOC.

Identification of genes differentially induced by hypoxia in pancreatic cancer cells.

A hypoxic microenvironment is characteristic of many solid tumors, including pancreatic cancer, the fifth leading cause of cancer death in the United States. Hypoxia causes the stabilization of the HIF-1 (hypoxia-inducible factor-1) transcription factor and the induction of many genes that promote angiogenesis, tumor growth, and metastasis. We performed representational difference analysis (RDA) using mRNA extracted from hypoxic and normoxic Capan-2, a human pancreatic cancer cell line. cDNAs corresponding to hypoxia-inducible genes were cloned and sequenced. We identified GPI/NLK/AMF (glucose phosphate isomerase/neuroleukin/autocrine motility factor) as a hypoxic inducible gene. In addition, hexokinase II and DEC1/Stra13, genes known to be hypoxia inducible in other systems, were found to be hypoxia inducible in our pancreatic cancer system. We thus identified three genes that are induced by hypoxia in a human pancreatic cancer, including GPI/NLK/AMF, which was not previously known to be hypoxia inducible in any other system. These genes may provide new targets for diagnosis and treatment of pancreatic cancer.

New equations for estimating body cell mass from bioimpedance parallel models in healthy older Germans.

The objectives of this study were to assess for elderly Germans the validity of existing equations for predicting body cell mass (BCM) and to develop from single- and multifrequency bioimpedance (SFBIA, MFBIA) models new prediction equations. In a data-splitting approach, validation and cross-validation were performed in 160 healthy elderly (60- to 90-yr) subjects. BCM was determined using a tetrapolar bioimpedance analyzer (800 microA; 4 fixed frequencies: 1, 5, 50, and 100 kHz; electrodes placed to hand, wrist, ankle, and foot) and whole body (40)K counting as a reference method. New prediction equations were derived by multiple stepwise regression analysis. The Bland-Altman procedure was used for methods comparison. Relative to whole body counting, the manufacturer's equation overestimated BCM by 9% in men (P < 0.0001, paired t-test) and 4% in women (P = 0.002). Compared with the manufacturer's equation, the newly derived equations (r = 0.92, RMSE = 6-9%) improved accuracy (pure error = 13 vs. 7-8%) and reduced bias and limits of agreement. SFBIA and MFBIA equations did not differ in precision or accuracy. We conclude that the newly derived equations improved BCM estimates in the elderly compared with existing equations. There was no advantage of MFBIA over SFBIA equations.

[Effect of the lavage of the digestive tract on microflora in patients with polyps in the large intestine]. / Vliianie lavazha pishchevaritel'nogo trakta na mikrofloru u bol'nykh s polipami v tolstoi kishke.

The microbial status of the intestine and the influence of lavage with polyethylene glycol and balanced electrolyte solution (PEG + E), used in the process of the preparation of patients to polypectomy, on this status were evaluated. The study of microflora was made before oral lavage after, and 48-72 hours later its completion. For control, a group of healthy volunteers, also subjected to oral lavage with PEG + E, was used. The lavage of the digestive tract with PEG + E led to a sharp change in the microbial status in both groups. Some microorganisms, previously absent in the intestine, were found after lavage. The processes of the restoration of intestinal microflora after lavage in healthy volunteers and in patients with polyps had certain differences. In healthy volunteers intestinal microflora was completely restored, and even improved, 48-72 hours after lavage with PEG + E, while at the expiration of this time intestinal microflora in the patients with polyps could be characterized as dysbiotic.

Peroxisome proliferator-activated receptor gamma induces pancreatic cancer cell apoptosis.

Peroxisome proliferator-activated receptor gamma (PPAR-gamma) decreases the growth of certain cancer cells. In the present study, we found that six different human pancreatic cancer cell lines (AsPC-1, BxPC-3, Capan-2, HPAF-II, MIA PaCa-2, and PANC-1) expressed PPAR-gamma m-RNA and synthesized the protein. The endogenous and exogenous PPAR-gamma ligands 15-deoxy-d12,14-prostaglandin J(2) (15-PGJ(2)) and ciglitazone decreased cell number, cell viability, and increased floating/attached ratio, in a time- and dose-dependent fashion. 15-PGJ(2) increased intracellular nucleosome concentration after 6 h, but did not increase caspase-3 activity even after 96 h. Combined treatment with both 15-PGJ(2) and the caspase-3 inhibitor DEVD-CHO had no effect on cell viability, but the general caspase inhibitor ZVAD-FMK reduced 15-PGJ(2)-induced apoptosis. We concluded that the six human pancreatic cancer cells tested all expressed PPAR-gamma receptor, and treatment with PPAR-gamma agonists decreased cell viability and growth in a time- and dose-dependent manner. These effects were partially mediated by induction of caspase-3 independent apoptosis.

[Intestinal microflora in children from Mongolia, Russia, and Switzerland]. / Mikroflora kishechnika u detei Mongolii, Rossii i Shveitsarii.

Intestinal microflora was studied in 3 groups of children; 55--living in Mongolia, 18--in Switzerland and 28--in Russia. Age of children of both sexual groups was 1.5-3 years. This study revealed that in none of these groups of clinically healthy children living in different regions and having different diet normal intestinal microflora corresponded to the standard considered to be the norm. The revision of norm criteria for normal intestinal microflora in children is recommended.

Age-related decline in body cell mass in elderly men and women, determined by a noninvasive nuclear technique: effects of physical activity and dietary potassium intake.

The body cell mass (BCM) represents the actively metabolizing cellular components of the human body. In this study, the hypothesis was tested that physical activity and a sufficient dietary intake of potassium attenuate the age-related decline in BCM in the elderly. Cross-sectional data were collected in 82 male and 79 female non-institutionalized elderly (60-90 years) from Mainz, Germany, and were analyzed by age groups. BCM was calculated from total body potassium, measured by whole-body counting of naturally occurring 40K. Physical activity level (PAL) was assessed by a standardized questionnaire. Dietary intake of potassium (DIP) was estimated from a 7-day food diary. The results showed the following trends: (1) BCM decreased continuously from age 60 to 90 years by 11.2% (men) and 7.0% (women). BCM was inversely correlated with age in both sexes (men, P < 0.001; women, P < 0.05), but significant age group differences only existed for men (P < 0.01, one-way analysis-of-variance). Women had less BCM than men in all age groups (P < 0.001, t tests). (2) Correlation analyses demonstrated that in both sexes PAL declined with advancing age (P < 0.05), whereas DIP did not change significantly with age. (3) In both sexes, BCM showed a strong positive correlation with PAL (P < 0.001), but did not correlate significantly with DIP. Multiple linear regression analysis (independent variables were PAL, DIP, age, height, weight) demonstrated that PAL, age, and height explained 45% of the variability in BCM in men. In women, PAL was the only significant predictor of BCM, explaining 23% of the variance. The study supports the hypothesis that the level of physical activity is associated with the decline of BCM in the elderly.

Platelet activating factor antagonism reduces the systemic inflammatory response in a murine model of acute pancreatitis.

BACKGROUND: The platelet activating factor (PAF) antagonist, Lexipafant, has been used in experimental models and clinical trials to treat severe acute pancreatitis (AP). The purpose of this study was to determine whether Lexipafant reduces the local and systemic components of AP in a murine model of mild, edematous AP. MATERIALS AND METHODS: Forty-eight female Swiss-Webster mice were divided into four groups. Group 1 received 50 microl of saline ip every hour for 6 h (sham). Group 2 received saline treatment, plus Lexipafant (25 mg/kg dose ip, every 3 h starting 1 h after the first saline injection) (sham/Lex). Group 3 received cerulein (50 microg/kg dose ip, every hour for 6 h) (AP). Group 4 received AP, plus therapeutic treatment with Lexipafant (AP/Lex). Animals were sacrificed 3 h after the last injection. Serum cytokine levels were determined by ELISA. Standard assays were performed for serum amylase activity and lung myeloperoxidase activity (MPO). Histology was scored by two blinded investigators. RESULTS: Serum cytokines (TNFalpha, IL-1beta), lung MPO, and serum amylase activity were reduced by PAF antagonism. Histology showed a trend toward improvement with Lexipafant, but did not reach statistical significance. CONCLUSION: The PAF antagonism reduces the severity of systemic inflammation when given after the induction of mild AP in mice. These results suggest that Lexipafant may be useful in the treatment of mild pancreatitis after its clinical onset.

Real-time quantitative PCR of telomerase mRNA is useful for the differentiation of benign and malignant pancreatic disorders.

The presence of telomerase activity has been proposed as a specific and sensitive marker for malignant tissue, and positivity rates of up to 95% have been reported in pancreatic cancer. In the present study telomerase activity analysis was reevaluated in 29 pancreatic cancer tissues compared with 36 chronic pancreatitis tissues and 21 normal controls, and a study was made of whether malignant and benign pancreatic disorders can be better differentiated using a novel technique real-time quantitative PCR analysis-analyzing telomerase mRNA expression. Telomerase activity was present in 35% (10 of 29) of pancreatic cancer samples, 3% (one of 36) of chronic pancreatitis samples, and none of the normal pancreatic tissue samples in the TRAP assay. Real-time quantitative PCR analysis revealed the presence of telomerase mRNA expression in 50% (10 of 20) of normal, 86% (31 of 36) of chronic pancreatitis, and 90% (26 of 29) of pancreatic cancer samples. However, quantification of the expression data revealed that the relative increase above normal was 5.5 (range, 3.5-8.6) for chronic pancreatitis and 23.9 (range, 18.6-30.7) for pancreatic cancer samples (p < 0.01). No relationship was found between telomerase activity and the fold increase of telomerase mRNA above normal and gender, patient age, tumor stage, or tumor grade. These data indicate that detection of telomerase activity using the TRAP assay has limitations in differentiating benign and malignant pancreatic disorders. However, telomerase mRNA analysis by real-time quantitative PCR analysis allows a highly sensitive detection and differentiation of pancreatic cancer from normal pancreas and chronic pancreatitis and thereby may serve as a new reliable, easy, and effective diagnostic tool for cancer diagnosis.

Angiogenesis inhibitor TNP-470 reduces human pancreatic cancer growth.

In this study we investigated the effects of the angiogenesis inhibitor TNP-470 on human pancreatic cancer cells in vitro and in vivo. The action of TNP-470 on vascular endothelial growth factor (VEGF) was also assessed. In vitro human pancreatic cancer cells (MIAPaCa-2, AsPC-1, and Capan-1), and human umbilical vein endothelial cells (HUVEC) were exposed to increasing concentrations (1 pg/ml to 100 microg/ml) of TNP-470. Cell proliferation was assessed after 3 days by cell count and MTT assay. In vivo, 5 x 10(6) pancreatic cancer cells were injected subcutaneously into nude mice. Four weeks later, 1 mm3 fragments of the resulting tumors were implanted into the pancreas of other mice. Animals received either TNP-470 (30 mg/kg every other day) or vehicle subcutaneously for 14 weeks. The volume of the primary tumor and metastatic spread were determined at autopsy. Concentrations of VEGF were determined in serum (VEGF(S)) and ascites (VEGF(A)) by enzyme-linked immunosorbent assay. Microvessel density was analyzed by immunohistochemistry in CD31-stained tumor sections. In vitro, proliferation and viability of the human pancreatic cancer cell lines were significantly inhibited at high concentrations of TNP-470 (> 1 microg/ml). In contrast, TNP-470 effectively decreased the growth of HUVEC at 100 pg/ml. In vivo, tumor volume and dissemination scores were significantly lower in all three pancreatic cancer cell lines. VEGF(S) and VEGF(A) were not different between treated groups. Treatment with TNP-470 significantly reduced neoangiogenesis in tumors of all three human pancreatic cancer cell lines: MIAPaCa-2 = 74.8 +/- 7.8/0.74 mm2 vs. 24.8 +/- 3.7/0.74 mm2; AsPC-1 = 65.3 +/- 5.0/0.74 mm2 vs. 26.0 +/- 3.4/0.74 mm2; and Capan-1 = 82.2 +/- 5.8/0.74 mm2 vs. 26.9 +/- 2.5/0.74 mm2 (P < 0.001). However, survival was not statistically different between groups. TNP-470 reduced tumor growth and metastatic spread of pancreatic cancer in vivo. This was probably due to the antiproliferative effect of the agent on endothelial cells rather than to the direct inhibition of pancreatic cancer cell growth. TNP-470 activity was not associated with alteration of VEGF secretion.