Cell phenotype determines PAI-1 antiproliferative effect - suppressed proliferation of the lung cancer but not prostate cancer cells

Wstęp: Inhibitor aktywatora plazminogenu typu 1 (PAI-1) jest ważnym regulatorem procesu wzrostu guza i tworzenia przerzutów nowotworowych, działając poprzez bezpośrednie hamowanie urokinazy (mechanizm antyfibrynolityczny) oraz niezależnie od kinaz poprzez powinowactwo z witronektyną. Autorzy pracy w poprzednim badaniu wykazali, że PAI-1 modyfikuje aktywność angiogenną komórek śródbłonka w stopniu zależnym od jego stężenia, jak również fenotypu komórek. Celem niniejszej pracy była ocena wpływu PAI-1 na aktywność proliferacyjną linii komórek nowotworowych - raka płuca (A549) i raka gruczołu krokowego (DU145), a także komórek strukturalnych - śródbłonka naczyniowego (HUVEC). Wyniki: Zmutowana postać PAI-1 (1, 10, 100 &#956;g/ml) charakteryzująca się znacząco przedłużoną aktywnością antyfibrynolityczną (T1/2 ~ 7000 godz.) w stopniu wyraźnie zależnym od dawki (p < 0,001) i czasu (p < 0,001) znamiennie hamowała aktywność proliferacyjną komórek raka płuca A549. Natomiast istotny wpływ PAI-1 na aktywność proliferacyjną komórek raka gruczołu krokowego DU145 wykazano tylko dla najwyższego użytego stężenia (100 &#956;g/ml) i tylko po 72 godzinach zahodowli (p < 0,001). Aktywność proliferacyjna komórek śródbłonka (HUVEC) była hamowana jedynie przez dawkę 100 &#956;g/ml PAI-1 po 24, 48 i 72 godzinach hodowli. Wniosek: Inhibitor aktywatora plazminogenu typu 1 moduluje aktywność proliferacyjną komórek w mechanizmie hamowania urokinazy w stopniu ściśle zależnym od fenotypu komórek, czasu działania i dawki. Pneumonol. Alergol. Pol. 2010; 78, 4: 279-283Introduction: Plasminogen inhibitor activator type 1 (PAI-1) is an important regulator of tumor growth and metastasis formation acting directly via specific urokinase complexing or indirectly due to its affinity to vitronectin. We have shown previously that PAI-1 modifies angiogenic activity of endothelial cells in a dose-dependent manner but also in close relationship to the cell phenotype. Present study aimed on evaluating the PAI-1 effect on the proliferative activity of lung cancer cells (A549), prostate cancer cells (DU145) as well as endothelial cells (HUVEC). Results: Mutated PAI-1 (1, 10, 100 &#956;g/mL) characterized by the prolonged antifibrinolytic activity (T1/2 ~ 7000 h) inhibited proliferation of lung cancer A549 cells in a dose-dependent (p < 0.001) and time-dependent (p < 0.001) manner. No significant effect on the DU145 prostate cancer cells has been observed except of the 72 h cultures with highest PAI-1 concentration (100 &#956;g/ml) (p < 0.001). Proliferative activity of endothelial cells (HUVEC) was affected by 100 &#956;g/ml PAI-1 only, and independent of the culture period (24, 48 and 72 h, p < 0.001). Conclusion: Plasminogen inhibitor activator type 1 modulates cell proliferation via antifibrynolitic mechanizm time- and dose-dependently, however final outcome is strongly affected by the cell phenotype. Pneumonol. Alergol. Pol. 2010; 78, 4: 279-28

Lipoxygenases - a Challenging Problem in Enzyme Inhibition and Drug Development

Lipoxygenases (LOXs), cytochromes P450 (CYPs) and cyclooxygenases (COXs) catalyze peroxidation of unsaturated fatty acids. In humans they convert arachidonic acid into a variety of eicosanoids, which play a role in all inflammatory responses, cardiovascular and kidney diseases, Alzheimer\u27s, cancer and other ailments. Blocking one pathway can prompt the body to switch to the available alternatives. In contrast to CYP and COX, LOX has a non-heme iron co-factor. Several LOXs are produced or stress-induced in the human body. They share the same mechanism, but differ in sequence causing catalysis on the same substrate to be regio- and stereospecific. The action of 15-LOXs could be pro- or anti-inflammatory, and pro- or anti-carcinogenic. Depending on the dose, LOXs inhibitors can induce or inhibit other oxygenases. Inhibition of these enzymes presents a great challenge in solving the problem of how to control their action and treat diseases, without causing severe side effects and maintaining/restoring a delicate equilibrium between them. Research on CYPs and COXs is more advanced, while studies of LOXs are lagging behind. This article presents a brief review about LOX structures and inhibition, their involvement in human diseases, and their interplay with other oxidoreductases

Evaluation of 12-Lipoxygenase (12-LOX) and Plasminogen Activator Inhibitor 1 (PAI-1) as Prognostic Markers in Prostate Cancer

In carcinoma of prostate, a causative role of platelet 12-lipoxygenase (12-LOX) and plasminogen activator inhibitor 1 (PAI-1) for tumor progression has been firmly established in tumor and/or adjacent tissue. Our goal was to investigate if 12-LOX and/or PAI-1 in patient’s plasma could be used to predict outcome of the disease. The study comprised 149 patients (age 70±9) divided into two groups: a study group with carcinoma confirmed by positive biopsy of prostate (n=116) and a reference group (n=33) with benign prostatic hyperplasia (BPH). The following parameters were determined by the laboratory test in plasma or platelet-rich plasma: protein level of 12-LOX, PAI-1, thromboglobulin (TGB), prostate specific antigen (PSA), C-reactive protein (CRP), hemoglobin (HGB, and hematocrit (HCT), as well as red (RBC) and white blood cells (WBC), number of platelets (PLT), international normalized ratio of blood clotting (INR), and activated partial thromboplastin time (APTT). The only difference of significance was noticed in the concentration of 12-LOX in platelet rich plasma, which was lower in cancer than in BPH group. Standardization to TGB and platelet count increases the sensitivity of the test that might be used as a biomarker to assess risk for prostate cancer in periodically monitored patients

Cell Phenotype Determines PAI1 Antiproliferative Effect—Suppressed Proliferation of the Lung Cancer but Not Prostate Cancer Cells

Introduction: Plasminogen inhibitor activator type 1 (PAI-1) is an important regulator of tumor growth and metastasis formation acting directly via specific urokinase complexing or indirectly due to its affinity to vitronectin. We have shown previously that PAI-1 modifies angiogenic activity of endothelial cells in a dose-dependent manner but also in close relation- ship to the cell phenotype. Present study aimed on evaluating the PAI-1 effect on the proliferative activity of lung cancer cells (A549), prostate cancer cells (DU145) as well as endothelial cells (HUVEC). Results: Mutated PAI-1 (1, 10, 100 μg/mL) characterized by the prolonged antifibrinolytic activity (T1/2 ~ 7000 h) inhibited proliferation of lung cancer A549 cells in a dose-dependent (p &lt; 0.001) and time-dependent (p &lt; 0.001) manner. No significant effect on the DU145 prostate cancer cells has been observed except of the 72 h cultures with highest PAI-1 concentration (100 μg/mL) (p &lt; 0.001). Proliferative activity of endothelial cells (HUVEC) was affected by 100 μg/mL PAI-1 only, and independent of the culture period (24, 48 and 72 h, p &lt; 0.001). Conclusion: Plasminogen inhibitor activator type 1 modulates cell proliferation via antifibrynolitic mechanizm time- and dose-dependently, however final outcome is strongly affected by the cell phenotype

Cell Phenotype Determines PAI-1 Antiproliferative Effect—Suppressed Proliferation of the Lung Cancer but Not Prostate Cancer Cells

Introduction: Plasminogen inhibitor activator type 1 (PAI-1) is an important regulator of tumor growth and metastasis formation acting directly via specific urokinase complexing or indirectly due to its affinity to vitronectin. We have shown previously that PAI-1 modifies angiogenic activity of endothelial cells in a dose-dependent manner but also in close relationship to the cell phenotype. Present study aimed on evaluating the PAI-1 effect on the proliferative activity of lung cancer cells (A549), prostate cancer cells (DU145) as well as endothelial cells (HUVEC). Results: Mutated PAI-1 (1, 10, 100 μg/ml) characterized by the prolonged antifibrinolytic activity (T1/2~7000 h) inhibited proliferation of lung cancer A549 cells in a dose-dependent (p &lt; 0.001) and time-dependent (p &lt; 0.001) manner. No significant effect on the DU145 prostate cancer cells has been observed except of the 72 h cultures with highest PAI-1 concentration (100 μg/ml) (p &lt; 0.001). Proliferative activity of endothelial cells (HUVEC) was affected by 100 μg/ml PAI-1 only, and independent of the culture period (24, 48 and 72 h, p &lt; 0.001). Conclusion: Plasminogen inhibitor activator type 1 modulates cell proliferation via antifibrynolitic mechanizm time- and dose-dependently, however final outcome is strongly affected by the cell phenotype

Structure of bovine prothrombin fragment 1 refined at 2.25 Å resolution

The structure of bovine prothrombin fragment 1 has been refined at 2.25 Å resolution using high resolution measurements made with the synchrotron beam at CHESS. The synchrotron data were collected photographically by oscillation methods (R-merge = 0.08). These were combined with lower order diffractometer data for refinement purposes. The structure was refined using restrained least-squares methods with the program PROLSQ to a crystallographic R-value of 0.175. The structure includes 105 water molecules with occupancies of >0·6. The first 35 residues (Ala1-Leu35) of the N-terminal ?-carboxy glutamic acid-domain (Ala1-Cys48) of fragment 1 are disordered as are two carbohydrate chains of Mr ? 5000; the latter two combine to render 40% of the structure disordered. The folding of the kringle of fragment 1 is related to the close intramolecular contact between the inner loop disulfide groups. Half of the conserved sequence of the kringle forms an inner core surrounding these disulfide groups. The remainder of the sequence conservation is associated with the many turns of the main chain. The Pro95 residue of the kringle has a cis conformation and Tyr74 is ordered in fragment 1, although nuclear magnetic resonance studies indicate that the comparable residue of plasminogen kringle 4 has two positions. Surface accessibility calculations indicate that none of the disulfide groups of fragment 1 is accessible to solvent

Can EGCG Alleviate Symptoms of Down Syndrome by Altering Proteolytic Activity?

Down syndrome (DS), also known as “trisomy 21”, is a genetic disorder caused by the presence of all or part of a third copy of chromosome 21. Silencing these extra genes is beyond existing technology and seems to be impractical. A number of pharmacologic options have been proposed to change the quality of life and lifespan of individuals with DS. It was reported that treatment with epigallocatechin gallate (EGCG) improves cognitive performance in animal models and in humans, suggesting that EGCG may alleviate symptoms of DS. Traditionally, EGCG has been associated with the ability to reduce dual specificity tyrosine phosphorylation regulated kinase 1A activity, which is overexpressed in trisomy 21. Based on the data available in the literature, we propose an additional way in which EGCG might affect trisomy 21—namely by modifying the proteolytic activity of the enzymes involved. It is known that, in Down syndrome, the nerve growth factor (NGF) metabolic pathway is altered: first by downregulating tissue plasminogen activator (tPA) that activates plasminogen to plasmin, an enzyme converting proNGF to mature NGF; secondly, overexpression of metalloproteinase 9 (MMP-9) further degrades NGF, lowering the amount of mature NGF. EGCG inhibits MMP-9, thus protecting NGF. Urokinase (uPA) and tPA are activators of plasminogen, and uPA is inhibited by EGCG, but regardless of their structural similarity tPA is not inhibited. In this review, we describe mechanisms of proteolytic enzymes (MMP-9 and plasminogen activation system), their role in Down syndrome, their inhibition by EGCG, possible degradation of this polyphenol and the ability of EGCG and its degradation products to cross the blood–brain barrier. We conclude that known data accumulated so far provide promising evidence of MMP-9 inhibition by EGCG in the brain, which could slow down the abnormal degradation of NGF