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Eleftherios P. Diamandis - One of the best experts on this subject based on the ideXlab platform.
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Deciphering the ovarian cancer Ascites Fluid peptidome
Clinical proteomics, 2014Co-Authors: Anand Bery, Felix Leung, Christopher R. Smith, Eleftherios P. Diamandis, Vathany KulasingamAbstract:Background Conventional proteomic approaches have thus far been unable to identify novel serum biomarkers for ovarian cancer that are more sensitive and specific than the current clinically used marker, CA-125. Because endogenous peptides are smaller and may enter the circulation more easily than proteins, a focus on the low-molecular-weight region may reveal novel biomarkers with enhanced sensitivity and specificity. In this study, we deciphered the peptidome of Ascites Fluid from 3 ovarian cancer patients and 3 benign individuals (Ascites Fluid from patients with liver cirrhosis).
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abstract 4560 integrative proteomic profiling of pancreatic cancer cell line supernatants pancreatic juice and Ascites Fluid for the identification of novel pancreatic cancer biomarkers
Cancer Research, 2010Co-Authors: Shalini Makawita, Christopher R. Smith, Ihor Batruch, Felix Ruckert, Eleftherios P. DiamandisAbstract:Pancreatic cancer is a highly lethal malignancy for which circulating biomarkers with high sensitivity and specificity are urgently needed. Proteins secreted or shed from tumor cells and their microenvironment have the highest chance of reaching the circulation and serving as measurable indicators of disease status. In this respect, we extensively characterized the proteomes of pancreatic cancer cell culture supernatants, in conjunction with proximal biological Fluids, for the identification of novel candidate biomarkers. Specifically, we characterized the secretomes of the pancreatic cancer cell lines MIA-PaCa2, BxPc-3, Capan1, SU.86.86, CFPAC-1 and Panc1, along with the normal pancreatic ductal epithelial cell line HPDE, through multidimensional separation using strong-cation exchange chromatography, and reverse-phase chromatography coupled online to an LTQ-Orbitrap hybrid mass spectrometer. For each cell line, optimal growth conditions to achieve increased protein secretion with minimal cell death were first selected for, and each cell line was analyzed using three biological replicates. Generated spectra were searched against both MASCOT and X!Tandem using the human IPI 3.62 forward and reverse database and results were integrated using Scaffold 2.06 software. This resulted in a non-redundant list of between 2000 and 3500 proteins identified in the conditioned media from each of the cell lines, with a false positive rate of approximately 1%. Together, a total of 5009 non-redundant proteins with unique gene names were identified in the seven cell lines combined. Upon performing gene ontology annotations, approximately 27% of the proteins were found to localize to the extracellular or plasma membrane components. We subsequently performed proteomic analysis of pancreatic juice from cancer and chronic pancreatitis patients, as well as Ascites Fluid from pancreatic cancer patients. Through integration of the proteins identified in the pancreatic juice and Ascites Fluid analyses with that of the cell line conditioned media, we prioritized a list of candidate biomarkers for verification in serum from cancer versus controls through established quantitative methods. To aid in our candidate selection, we also performed multiple pathway and tissue specificity analyses using publically available databases, and made comparisons to proteins identified in relevant tissue proteomics and microarray studies found in the literature. Our analysis identified many previously studied pancreatic cancer biomarkers, which serves to validate our discovery approach. To our knowledge, this study comprises the most exhaustive proteome from pancreatic cancer cell lines, and the proteins identified through integration of the multiple biological sources can now serve as a mine for novel biomarkers and therapeutic targets. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4560.
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functional proteomics of kallikrein related peptidases in ovarian cancer Ascites Fluid
Biological Chemistry, 2010Co-Authors: Katerina Oikonomopoulou, Christopher R. Smith, Eleftherios P. Diamandis, Ihor Batruch, Antoninus Soosaipillai, Morley D HollenbergAbstract:Kallikrein-related peptidases (KLKs) are secreted serine proteinases with trypsin or chymotrypsin-like activity. Several family members, such as KLKs 6 and 10, are potential ovarian cancer biomarkers. Recently, using a newly developed assay for active KLK6, we found that only a very small proportion of immunoreactive KLK6 in tumor-derived clinical samples (malignant Ascites Fluid), in cerebrospinal Fluid, and in cancer cell line supernatants is enzymatically active. We therefore hypothesized that a proportion of other immunoreactive KLKs in such samples could be present, but might be partly complexed to endogenous serine proteinase inhibitors. Using a combination of immunological isolation of the enzymes, activity-based probe analysis and proteomics, we identified active KLK10 in ovarian cancer Ascites and we provide preliminary data that the activity of other KLKs present in these samples can be decreased by known proteinase inhibitors (e.g., a2-macroglobulin, a1-antitrypsin). Our data suggest that the enzymatic activity of ovarian cancer-released KLKs that are detected by regular immunoassays is low in vivo and very likely regulated by proteinase
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Retrieval of autoantibodies from ovarian cancer cells present in Ascites Fluid
Cancer Research, 2005Co-Authors: Geeth Gunawardana, Liu-ying Luo, Eleftherios P. DiamandisAbstract:1534 The immune system can differentiate self-antigens from non-self-antigens. Aberrantly expressed, mutated, or improperly processed self-proteins can also elicit immune responses, as is the case during cancer development. In general, the immune response to cancer can be either mediated by T cells (cellular immunity) or B cells (humoral immunity). In both cases, the immune response is against proteins known as “tumour associated antigens” (TAAs). Our previous studies and those of others have demonstrated the existence of humoral immune response in ovarian cancer. In this study, tumour-specific antibodies were isolated from tumour cells in the Ascites Fluid of patients with ovarian cancer. Ascites Fluid from ovarian cancer patients was centrifuged to pellet tumour cells. Cell pellets were washed under conditions that avoided cell lysis, yet ensured the removal of loosely bound antibodies, which were assumed to be non-specific. Bound antibodies were released from cells by a low pH wash under isotonic conditions. Figure 1 illustrates the efficiency of removing non-specifically bound IgG and of retrieving tumour specific IgG. Using ELISA, the IgGs isolated from the tumour cells were calculated to be 0.0002 % of the total IgG in the Ascites Fluid. We conclude that tumour-specific antibodies can be retrieved from the surface of tumour cells and these antibodies may be used to identify TAAs, which may then be used as targets for cancer vaccine therapy.
Harold F Dvorak - One of the best experts on this subject based on the ideXlab platform.
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pathogenesis of Ascites tumor growth vascular permeability factor vascular hyperpermeability and Ascites Fluid accumulation
Cancer Research, 1995Co-Authors: Janice A Nagy, Elizabeth M Masse, Kemp T Herzberg, Michelle S Meyers, Kiangteck J Yeo, Tetkin Yeo, Tracy M Sioussat, Harold F DvorakAbstract:Abstract Previous studies have shown that accumulation of tumor Ascites Fluid results in large part from increased permeability of peritoneal lining vessels (Nagy et al. , Cancer Res., 49: 5449–5458, 1989; Nagy et al. , Cancer Res., 53: 2631-2643, 1993). However, the specific microvessels rendered hyperpermeable have not been identified nor has the basis of peritoneal vascular hyperpermeability been established. To address these questions, TA3/St and MOT carcinomas, well-characterized transplantable murine tumors that grow in both solid and Ascites form, were studied as model systems. Ascites tumor cells of either type were injected i.p. into syngeneic A/Jax and C3Heb/FeJ mice, and Ascites Fluid and plasma were collected at intervals thereafter up to 8 and 28 days, respectively. Beginning several days after tumor cell injection, small blood vessels located in tissues lining the peritoneal cavity (mesentery, peritoneal wall, and diaphragm) became hyperpermeable to several macromolecular tracers ( 125 I-human serum albumin, FITC-dextran, colloidal carbon, and Monastral Blue B). Increased microvascular permeability correlated with the appearance in Ascites Fluid of vascular permeability factor (VPF), a tumor cell-secreted mediator that potently enhances vascular permeability to circulating macromolecules. VPF was measured in peritoneal Fluid by both a functional bioassay and a sensitive immunofluorometric assay. The VPF concentration, total peritoneal VPF, Ascites Fluid volume, tumor cell number, and hyperpermeability of peritoneal lining microvessels were found to increase in parallel over time. The close correlation of peritoneal Fluid VPF concentration with the development of hyperpermeable peritoneal microvessels in these two well-defined Ascites tumors suggests that VPF secretion by tumor cells is responsible, in whole or in part, for initiating and maintaining the Ascites pattern of tumor growth.
Mark W.c. Hatton - One of the best experts on this subject based on the ideXlab platform.
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human ovarian cancer Ascites Fluid contains a mixture of incompletely degraded soluble products of fibrin that collectively possess an antiangiogenic property
International Journal of Gynecological Cancer, 2006Co-Authors: N Jandu, M Richardson, Gurmit Singh, H Hirte, Mark W.c. HattonAbstract:Ovarian cancer Ascites Fluid (OCAF) displayed an antiangiogenic property in a chick chorioallantoic membrane (CAM) assay. This property was attributed in part to angiostatin although angiostatin-free OCAF retained a net antiangiogenic property. Recently, immunopurified fibrin(ogen) degradation products (FDPs) from malignant effusions of VX2 tumor–burdened rabbits exhibited antiangiogenic activity on the CAM. We questioned whether the FDPs of OCAF were also antiangiogenic. FDPs were immunopurified from individual OCAF samples, characterized by sodium dodecyl sulfate–polyacrylamide gel electrophoresis /western blots, enzyme-linked immunosorbent assays, and CAM assays. FDPs of OCAFs consisted of soluble high molecular weight (MW) fragments (>200 kd; ∼40% of total FDPs), D-dimer (∼180 kd; ∼37%), fragment D (∼90 kd; ∼15%), and fragment E (∼50 kd; ∼8%); intact fibrinogen was absent. When applied to CAM surfaces (0.5–1.6 mg/10 mL), purified FDPs significantly reduced the area of chorionic capillaries from 90% (in controls) to 47% over a 48-h period; from CAM sections, capillary density was reduced from 60% (controls) to 26%. FDPs prepared from fibrinogen displayed a similar antiangiogenic effect. Further digestion of OCAF FDPs by human plasmin caused degradation of high MW fragments, releasing additional D-dimer, fragment D, and fragment E. Of the fibrinogen-related components, OCAF contained only soluble FDPs (including incompletely digested fibrin fragments). Collectively, these FDPs contributed to the net antiangiogenic property of Ascites Fluid.
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A tumor model of Ascites Fluid production: Intrahepatic injection of the rabbit with rabbit VX2 tumor cells
Cancer Research, 2005Co-Authors: Bonnie Ross, Suzanne M.r. Southward, Mary Richardson, Mark W.c. HattonAbstract:1049 />From our previous work, approximately 15% of New Zealand White rabbits (NZW; weight range: 2.5 - 3.5 kg), that were injected intravenously with VX2 tumor cells and yielded lung tumors at 28d, accumulated an effusion in the interpleural space. This effusion contained the products and catabolic fragments of hemostasis (e.g. angiostatin, fibrin degradation products (FDPs)) that, when isolated and purified, contributed towards the net anti-angiogenic activity of VX2 effusions as shown by a chick embryo chorioallantoic membrane (CAM) assay. To study the dynamics of tumor effusion formation in vivo, a tumor model has been developed in which an increased proportion of tumor-bearing rabbits accumulate an effusion. Anesthetized NZW rabbits were subjected to a small incision into the abdominal cavity below the xiphoid cartilage. Live tumor cells, from a freshly-harvested lung or liver tumor from a donor rabbit, were injected (2-3 x 105/0.5ml saline) directly into the liver at two sites and the wound closed. After 28d, the rabbit was anesthetized; the abdominal cavity contained an extensive tumor growth associated with the liver, the omentum and the site of incision. Ascites Fluid was present in 88% of female rabbits (32 ± 25 ml/rabbit; n = 8) and in 36% of male rabbits (18 ± 45 ml; n = 11). Analyses of total protein content, fibrinolytic factors and FDPs, and anti-angiogenic properties (by the CAM method) indicated that Ascites Fluid and pleural effusion from VX2-tumor rabbits were similar in many respects. The volume of recovered Ascites Fluid varied positively with the tumor burden of the liver, whereas the concentration of live VX2 cells/ml of Ascites Fluid was independent of tumor burden. This rabbit VX2-tumor liver model will be used to study the dynamics of Ascites Fluid production in the tumor-burdened rabbit, and to investigate more about the changing contents of malignant effusions as tumor growth progresses in vivo.
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malignant Ascites Fluid maf including ovarian cancer associated maf contains angiostatin and other factor s which inhibit angiogenesis
Gynecologic Oncology, 2002Co-Authors: Mark W.c. Hatton, M Richardson, Gurmit Singh, J Gunawan, Eric Seidlitz, Hal W HirteAbstract:Abstract Objective. The aim was to determine whether human malignant Ascites Fluid (MAF) associated with abdominal cancer, including ovarian cancer, contained factors which inhibit angiogenesis as well as others which stimulate this process. Methods. MAF was collected from six patients, four with ovarian cancer, one with gastric cancer, and one with liver metastases. Using the chick chorioallantoic membrane (CAM) the effect of MAF on 7-day-old CAM capillaries was examined for 48 h. Vascular endothelial growth factor (VEGF) was evaluated by ELISA. Five samples of MAF were fractionated by lysine–Sepharose chromatography and the lysine-bound and -unbound fractions were eluted by ϵ-amino- n -hexanoic acid. Whole MAF, the lysine-bound and -unbound fractions, and human angiostatin were subjected to SDS–PAGE/Western blot analysis and immunostained after exposure to anti-human plasminogen. Human plasminogen was exposed to conditioned medium from ovarian epithelial cancer (HEY) cells and subjected to similar Western blot analysis. Results. Despite containing VEGF, each MAF sample examined caused a loss of capillaries from the CAM; a similar response was seen using purified human angiostatin. Whole MAF and the lysine-bound fraction contained plasminogen (90 kDa) and a 55-kDa protein which migrated in a similar manner to human angiostatin on Western blot. Both the lysine-bound and -unbound fractions caused a loss of capillaries in the CAM. Human plasminogen exposed to conditioned medium from HEY cells yielded a fragment which was similar in size to angiostatin. Conclusions. MAF from patients with various clinical presentations contains angiostatin and VEGF as well as other factors which are capable of inhibiting angiogenesis.
Janice A Nagy - One of the best experts on this subject based on the ideXlab platform.
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pathogenesis of Ascites tumor growth vascular permeability factor vascular hyperpermeability and Ascites Fluid accumulation
Cancer Research, 1995Co-Authors: Janice A Nagy, Elizabeth M Masse, Kemp T Herzberg, Michelle S Meyers, Kiangteck J Yeo, Tetkin Yeo, Tracy M Sioussat, Harold F DvorakAbstract:Abstract Previous studies have shown that accumulation of tumor Ascites Fluid results in large part from increased permeability of peritoneal lining vessels (Nagy et al. , Cancer Res., 49: 5449–5458, 1989; Nagy et al. , Cancer Res., 53: 2631-2643, 1993). However, the specific microvessels rendered hyperpermeable have not been identified nor has the basis of peritoneal vascular hyperpermeability been established. To address these questions, TA3/St and MOT carcinomas, well-characterized transplantable murine tumors that grow in both solid and Ascites form, were studied as model systems. Ascites tumor cells of either type were injected i.p. into syngeneic A/Jax and C3Heb/FeJ mice, and Ascites Fluid and plasma were collected at intervals thereafter up to 8 and 28 days, respectively. Beginning several days after tumor cell injection, small blood vessels located in tissues lining the peritoneal cavity (mesentery, peritoneal wall, and diaphragm) became hyperpermeable to several macromolecular tracers ( 125 I-human serum albumin, FITC-dextran, colloidal carbon, and Monastral Blue B). Increased microvascular permeability correlated with the appearance in Ascites Fluid of vascular permeability factor (VPF), a tumor cell-secreted mediator that potently enhances vascular permeability to circulating macromolecules. VPF was measured in peritoneal Fluid by both a functional bioassay and a sensitive immunofluorometric assay. The VPF concentration, total peritoneal VPF, Ascites Fluid volume, tumor cell number, and hyperpermeability of peritoneal lining microvessels were found to increase in parallel over time. The close correlation of peritoneal Fluid VPF concentration with the development of hyperpermeable peritoneal microvessels in these two well-defined Ascites tumors suggests that VPF secretion by tumor cells is responsible, in whole or in part, for initiating and maintaining the Ascites pattern of tumor growth.
Lei Zhu - One of the best experts on this subject based on the ideXlab platform.
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detection of soluble apo 1 fas in plasma pleural and Ascites Fluid of malignant tumor patients and its clinical significance
Journal of Zhejiang University (Medical Sciences), 2003Co-Authors: Luhong Luo, Lei ZhuAbstract:OBJECTIVE To study the changes of soluble Apo-1/Fas levels in plasma, pleural and Ascites Fluid of malignant tumor patients and to evaluate their clinical significance. METHODS The soluble Apo-1/Fas levels were measured by enzyme-linked immunosorbent assay (ELISA) in the plasma of 157 malignant tumor patients and 25 normal controls as well as in the pleural and ascite Fluids of 129 patients with various diseases. RESULT The plasma soluble Apo-1/Fas levels in acute and chronic leukemia and multiple myeloma were significantly higher than those in normal controls (P <0.05). The plasma soluble Apo-1/Fas levels in chronic myeloid leukemia and chronic lymphocytic leukemia were significantly higher than those in acute myeloid leukemia and acute lymphocytic leukemia, respectively (P <0.05). After chemotherapy, the plasma soluble Apo-1/Fas levels in complete remission group were distinctly decreased(P <0.05),whereas the levels in no remission and recurrence groups remained high. Compared with normal controls, the plasma soluble Apo-1/Fas levels in solid tumors were significantly increased (P <0.01), and the levels in metastasis cancers were significantly higher than those in non-metastasis cancer (P <0.0 1). Simultaneously the levels in remission cancer patients after operation and radiotherapy were distinctly lower than those before treatment(P <0.01), but were significantly increased in recurrence cancer patients (P <0.01). The soluble Apo-1/Fas levels in pleural and Ascites Fluid of malignant tumors were significantly higher than those in tuberculous effusions and transudates. CONCLUSION The soluble Apo-1/Fas levels in plasma, pleural and Ascites Fluid of malignant tumor patients are markedly increased, which might be associated with the progress of cancers. The changes of soluble Apo-1/Fas levels may be useful for understanding the pathologic process of cancers and to differential diagnosis of various pleural and Ascites Fluids.
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Detection of soluble Apo-1/Fas in plasma, pleural and Ascites Fluid of malignant tumor patients and its clinical significance
Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences, 2003Co-Authors: Luhong Luo, Lei ZhuAbstract:OBJECTIVE To study the changes of soluble Apo-1/Fas levels in plasma, pleural and Ascites Fluid of malignant tumor patients and to evaluate their clinical significance. METHODS The soluble Apo-1/Fas levels were measured by enzyme-linked immunosorbent assay (ELISA) in the plasma of 157 malignant tumor patients and 25 normal controls as well as in the pleural and ascite Fluids of 129 patients with various diseases. RESULT The plasma soluble Apo-1/Fas levels in acute and chronic leukemia and multiple myeloma were significantly higher than those in normal controls (P
