The Experts below are selected from a list of 26913 Experts worldwide ranked by ideXlab platform
Craig W. Lindsley - One of the best experts on this subject based on the ideXlab platform.
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Targeting phospholipase D in cancer, infection and neurodegenerative disorders
Nature Reviews Drug Discovery, 2017Co-Authors: H. Alex Brown, Paul G. Thomas, Craig W. LindsleyAbstract:Lipid second messengers have essential roles in cellular function and contribute to the molecular mechanisms that underlie inflammation, malignant transformation, invasiveness, neurodegenerative disorders, and infectious and other pathophysiological processes. The phospholipase D (PLD) isoenzymes PLD1 and PLD2 are one of the major sources of signal-activated phosphatidic acid (PtdOH) generation downstream of a variety of cell-surface receptors, including G protein-coupled receptors (GPCRs), receptor tyrosine kinases (RTKs) and integrins. Recent advances in the development of isoenzyme-selective PLD inhibitors and in molecular genetics have suggested that PLD isoenzymes in mammalian cells and pathogenic organisms may be valuable targets for the treatment of several human diseases. Isoenzyme-selective inhibitors have revealed complex inter-relationships between PtdOH biosynthetic pathways and the role of PtdOH in pathophysiology. PLD enzymes were once thought to be undruggable owing to the ubiquitous nature of PtdOH in cell signalling and concerns that inhibitors would be too toxic for use in humans. However, recent promising discoveries suggest that small-molecule isoenzyme-selective inhibitors may provide novel compounds for a unique approach to the treatment of cancers, neurodegenerative disorders and other afflictions of the central nervous system, and potentially serve as broad-spectrum antiviral and antimicrobial therapeutics. Phospholipase D (PLD) enzymes are one source of receptor-generated phosphatidic acid (PtdOH),which may subsequently be metabolized to diacylglycerol (DAG) and lysophosphatidic acid. There are other pathways that lead to PtdOH generation, but differences in pathways and in the acyl composition of the products seem to provide some specificity. Both direct and indirect inhibitors of PLD activity have been identified despite a long-held suspicion that this pathway was undruggable. The identification of raloxifene and halopemide as direct inhibitors was followed by the systematic development of isoenzyme-preferring compounds that have been used to further differentiate the functions of PLD1 and PLD2. PLD2 in host cells has been associated with viral entry processes and innate immune response pathways such that inhibition blocks efficient infection. This PLD2 pathway has been linked to autophagy via AKT kinases. As a potential target in antiretroviral therapy, PLD1 works through the CAD enzyme (which contains carbamoyl aspartate synthase, aspartate transcarbamylase and dihydro-orotase domains) to modulate pyrimidine biosynthesis. PLD activity and expression have been shown to be upregulated in several types of human cancers, in which PLD enzymes function downstream of a variety of known oncogenes. Inhibition of PtdOH production has a marked effect on tumorigenesis and malignant invasion. PLD1, PLD2 and PLD3 have each been suggested to have a role in Alzheimer disease and other neurodegenerative conditions, but a mechanism has not yet emerged to explain the roles of these proteins in central nervous system pathophysiology. Lipid second messengers such as phosphatidic acid (PtdOH) have a role in a wide range of pathological processes, and phospholipase D (PLD) enzymes are one of the major sources of signal-activated PtdOH generation. In this Review, Brown, Thomas and Lindsley discuss the development of PLD inhibitors, with a focus on isoform-specific inhibitors, and their potential applications in the treatment of cancer, neurodegeneration and infection.
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Discovery of a highly selective PLD2 inhibitor (ML395): a new probe with improved physiochemical properties and broad spectrum antiviral activity against influenza strains
ChemMedChem, 2014Co-Authors: Matthew C. O’reilly, H. Alex Brown, Paul G. Thomas, Sarah A Scott, Thomas Oguin, Ryan D Morrison, J. Scott Daniels, Charles W. Locuson, Craig W. LindsleyAbstract:Further chemical optimization of the halopemide-derived family of dual phospholipase D1/2 (PLD1/2) inhibitors afforded ML395 (VU0468809), a potent, >80-fold PLD2 selective allosteric inhibitor (cellular PLD1, IC50 >30,000 nM; cellular PLD2, IC50 =360 nM). Moreover, ML395 possesses an attractive in vitro DMPK profile, improved physiochemical properties, ancillary pharmacology (Eurofins Panel) cleaner than any other reported PLD inhibitor, and has been found to possess interesting activity as an antiviral agent in cellular assays against a range of influenza strains (H1, H3, H5 and H7).
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development of dual pld1 2 and PLD2 selective inhibitors from a common 1 3 8 triazaspiro 4 5 decane core discovery of ml298 and ml299 that decrease invasive migration in u87 mg glioblastoma cells
Journal of Medicinal Chemistry, 2013Co-Authors: Matthew C Oreilly, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Scott J Daniels, Ryan D Morrison, Alex H Brown, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 ...
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Development of dual PLD1/2 and PLD2 selective inhibitors from a common 1,3,8-Triazaspiro[4.5]decane Core: discovery of Ml298 and Ml299 that decrease invasive migration in U87-MG glioblastoma cells.
Journal of medicinal chemistry, 2013Co-Authors: Matthew C. O’reilly, H. Alex Brown, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Ryan D Morrison, J. Scott Daniels, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 ...
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development of dual pld1 2 and PLD2 selective inhibitors from a common 1 3 8 triazaspiro 4 5 decane core discovery of ml298 and ml299 that decrease invasive migration in u87 mg glioblastoma cells
Journal of Medicinal Chemistry, 2013Co-Authors: Matthew C Oreilly, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Scott J Daniels, Ryan D Morrison, Alex H Brown, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 nM). SAR studies revealed that a small structural change (incorporation of a methyl group) increased PLD1 activity within this classically PLD2-preferring core and that the effect was enantiospecific. Both probes decreased invasive migration in U87-MG glioblastoma cells.
Michael A Frohman - One of the best experts on this subject based on the ideXlab platform.
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oxidized ldl phagocytosis during foam cell formation in atherosclerotic plaques relies on a PLD2 cd36 functional interdependence
Journal of Leukocyte Biology, 2018Co-Authors: Ramya Ganesan, Michael A Frohman, Karen M. Henkels, Gilbert Di Paolo, Lucile E Wrenshall, Yasunori Kanaho, Julian GomezcambroneroAbstract:The uptake of cholesterol carried by low-density lipoprotein (LDL) is tightly controlled in the body. Macrophages are not well suited to counteract the cellular consequences of excess cholesterol leading to their transformation into "foam cells," an early step in vascular plaque formation. We have uncovered and characterized a novel mechanism involving phospholipase D (PLD) in foam cell formation. Utilizing bone marrow-derived macrophages from genetically PLD deficient mice, we demonstrate that PLD2 (but not PLD1)-null macrophages cannot fully phagocytose aggregated oxidized LDL (Agg-Ox-LDL), which was phenocopied with a PLD2-selective inhibitor. We also report a role for PLD2 in coupling Agg-oxLDL phagocytosis with WASP, Grb2, and Actin. Further, the clearance of LDL particles is mediated by both CD36 and PLD2, via mutual dependence on each other. In the absence of PLD2, CD36 does not engage in Agg-Ox-LDL removal and when CD36 is blocked, PLD2 cannot form protein-protein heterocomplexes with WASP or Actin. These results translated into humans using a GEO database of microarray expression data from atheroma plaques versus normal adjacent carotid tissue and observed higher values for NFkB, PLD2 (but not PLD1), WASP, and Grb2 in the atheroma plaques. Human atherectomy specimens confirmed high presence of PLD2 (mRNA and protein) as well as phospho-WASP in diseased arteries. Thus, PLD2 interacts in macrophages with Actin, Grb2, and WASP during phagocytosis of Agg-Ox-LDL in the presence of CD36 during their transformation into "foam cells." Thus, this study provides new molecular targets to counteract vascular plaque formation and atherogenesis.
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oxidized ldl phagocytosis during foam cell formation in atherosclerotic plaques relies on a PLD2 cd36 functional interdependence
Journal of Leukocyte Biology, 2018Co-Authors: Ramya Ganesan, Michael A Frohman, Karen M. Henkels, Gilbert Di Paolo, Lucile E Wrenshall, Yasunori Kanaho, Julian GomezcambroneroAbstract:The uptake of cholesterol carried by Low Density Lipoprotein (LDL) is tightly controlled in the body. Macrophages are not well suited to counteract the cellular consequences of excess cholesterol leading to their transformation into “foam cells”, an early step in vascular plaque formation. We have uncovered and characterized a novel mechanism involving phospholipase D (PLD) in foam cell formation. Utilizing bone marrow-derived macrophages from PLD genetically-deficient mice, we demonstrate that PLD2 (but not PLD1)-null macrophages cannot fully phagocytose aggregated oxidized LDL (Agg-Ox-LDL), which was phenocopied with with a PLD2-selective inhibitor. We also report a role for PLD2 in coupling Agg-oxLDL phagocytosis with WASP, Grb2 and Actin. Further, the clearance of LDL particles is mediated by both CD36 and PLD2, in a mutual dependence on each other. In the absence of PLD2, CD36 does not engage in Agg-ox-LDL removal and when CD36 is blocked, PLD2 cannot form protein-protein heterocomplexes with WASP or Actin. These result translated into humans using a GEO database of microarray expression data from atheroma plaques versus normal adjacent carotid tissue and observed higher values for NFkB, PLD2 (but not PLD1), WASP and Grb2 in the atheroma plaques. Human artherectomy specimens confirmed high presence of PLD2 (mRNA and protein) as well as phospho-WASP in diseased arteries. Thus, PLD2 interacts in macrophages with Actin, Grb2 and WASP during phagocytosis of Agg-ox-LDL in the presence of CD36 during their transformation into “foam cells”. This knowledge provides several new molecular targets to better understand the disease and counteract vascular plaque formation.
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the phospholipase d superfamily as therapeutic targets
Trends in Pharmacological Sciences, 2015Co-Authors: Michael A FrohmanAbstract:The phospholipase D (PLD) lipid-signaling enzyme superfamily has long been studied for its roles in cell communication and a wide range of cell biological processes. With the advent of loss-of-function genetic mouse models that have revealed that PLD1 and PLD2 ablation is overtly tolerable, small-molecule PLD1/2 inhibitors that do not cause unacceptable clinical toxicity, a PLD2 polymorphism that has been linked to altered physiology, and growing delineation of processes that are subtly altered in mice lacking PLD1/2 activity, the stage is being set for assessment of PLD1/2 inhibition for therapeutic purposes. Based on findings to date, PLD1/2 inhibition may be of more utility in acute rather than chronic settings, although this generalization will depend on the specific risks and benefits in each disease setting.
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dependence of phospholipase d1 multi monoubiquitination on its enzymatic activity and palmitoylation
Journal of Biological Chemistry, 2010Co-Authors: Hao Yin, Michael A Frohman, Yu Gui, Xilong ZhengAbstract:Phospholipase D (PLD) is an important lipase in many cellular processes, including vesicular trafficking, cell survival, and cell migration. In the present study, we show that PLD1, but not PLD2, is posttranslationally modified by multi-monoubiquitination. Intriguingly, suppression of lipase activity either by mutation of the HKD motif (PLD1 H896R, K898R, or D903A) or the phosphatidylinositol 4,5-bisphosphate binding motif (PLD1 R691G,R695G) or through use of PLD-selective inhibitors impaired the ubiquitination of PLD1, although stimulation of lipase activity by phorbol 12-myristate 13-acetate did not enhance its ubiquitination. A palmitoylation-deficient mutant PLD1 allele, which exhibits altered patterns of vesicular trafficking, had significantly lower levels of monoubiquitination. In addition, the expression of ubiquitin-fused PLD1 induced aberrantly enlarged vesicles partially co-localized with the Golgi complex but not with early endosomes. The altered localization was reduced by the K898R mutation, suggesting a role of multi-monoubiquitination in PLD1 subcellular localization. Surprisingly, the degradation of PLD1, but not of PLD1 K898R or PLD2, was blocked by inhibitors of proteasomes but not by inhibitors of lysosomes or other proteases, suggesting a role of the ubiquitination in proteasomal degradation of PLD1. In summary, our studies show that PLD1, but not PLD2, is multi-monoubiquitinated. The ubiquitination modification might represent a novel regulatory mechanism in PLD1 functioning, particularly in the context of subcellular trafficking between different membrane compartments.
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dynamics and function of phospholipase d and phosphatidic acid during phagocytosis
Traffic, 2006Co-Authors: Matthias Corrotte, Guangwei Du, Mariefrance Bader, Sylvette Chasserotgolaz, Michael A Frohman, Nicolas Vitale, Nicholas T Ktistakis, Ping Huang, Nancy J. GrantAbstract:Phospholipase D (PLD) produces phosphatidic acid (PA), an established intracellular signalling lipid that has been also implicated in vesicular trafficking, and as such, PLD could play multiple roles during phagocytosis. Using an RNA interference strategy, we show that endogenous PLD1 and PLD2 are necessary for efficient phagocytosis in murine macrophages, in line with results obtained with wild-type constructs and catalytically inactive PLD mutants which, respectively, enhance and inhibit phagocytosis. Furthermore, we found that PA is transiently produced at sites of phagosome formation. Macrophage PLD1 and PLD2 differ in their subcellular distributions. PLD1 is associated with cytoplasmic vesicles, identified as a late endosomal/lysosomal compartment, whereas PLD2 localizes at the plasma membrane. In living cells undergoing phagocytosis, PLD1 vesicles are recruited to nascent and internalized phagosomes, whereas PLD2 is only observed on nascent phagosomes. These results provide evidence that both PLD isoforms are required for phagosome formation, but only PLD1 seems to be implicated in later stages of phagocytosis occurring after phagosomal internalization.
Sarah A Scott - One of the best experts on this subject based on the ideXlab platform.
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Discovery of a highly selective PLD2 inhibitor (ML395): a new probe with improved physiochemical properties and broad spectrum antiviral activity against influenza strains
ChemMedChem, 2014Co-Authors: Matthew C. O’reilly, H. Alex Brown, Paul G. Thomas, Sarah A Scott, Thomas Oguin, Ryan D Morrison, J. Scott Daniels, Charles W. Locuson, Craig W. LindsleyAbstract:Further chemical optimization of the halopemide-derived family of dual phospholipase D1/2 (PLD1/2) inhibitors afforded ML395 (VU0468809), a potent, >80-fold PLD2 selective allosteric inhibitor (cellular PLD1, IC50 >30,000 nM; cellular PLD2, IC50 =360 nM). Moreover, ML395 possesses an attractive in vitro DMPK profile, improved physiochemical properties, ancillary pharmacology (Eurofins Panel) cleaner than any other reported PLD inhibitor, and has been found to possess interesting activity as an antiviral agent in cellular assays against a range of influenza strains (H1, H3, H5 and H7).
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development of dual pld1 2 and PLD2 selective inhibitors from a common 1 3 8 triazaspiro 4 5 decane core discovery of ml298 and ml299 that decrease invasive migration in u87 mg glioblastoma cells
Journal of Medicinal Chemistry, 2013Co-Authors: Matthew C Oreilly, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Scott J Daniels, Ryan D Morrison, Alex H Brown, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 ...
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Development of dual PLD1/2 and PLD2 selective inhibitors from a common 1,3,8-Triazaspiro[4.5]decane Core: discovery of Ml298 and Ml299 that decrease invasive migration in U87-MG glioblastoma cells.
Journal of medicinal chemistry, 2013Co-Authors: Matthew C. O’reilly, H. Alex Brown, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Ryan D Morrison, J. Scott Daniels, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 ...
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development of dual pld1 2 and PLD2 selective inhibitors from a common 1 3 8 triazaspiro 4 5 decane core discovery of ml298 and ml299 that decrease invasive migration in u87 mg glioblastoma cells
Journal of Medicinal Chemistry, 2013Co-Authors: Matthew C Oreilly, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Scott J Daniels, Ryan D Morrison, Alex H Brown, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 nM). SAR studies revealed that a small structural change (incorporation of a methyl group) increased PLD1 activity within this classically PLD2-preferring core and that the effect was enantiospecific. Both probes decreased invasive migration in U87-MG glioblastoma cells.
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design synthesis and biological evaluation of halogenated n 2 4 oxo 1 phenyl 1 3 8 triazaspiro 4 5 decan 8 yl ethyl benzamides discovery of an isoform selective small molecule phospholipase d2 inhibitor
Journal of Medicinal Chemistry, 2010Co-Authors: Robert R Lavieri, Sarah A Scott, Scott J Daniels, Ryan D Morrison, Alex H Brown, Paige E Selvy, Satyawan Jadhav, Craig W. LindsleyAbstract:Phospholipase D (PLD) catalyzes the conversion of phosphatidylcholine to the lipid second messenger phosphatidic acid. Two mammalian isoforms of PLD have been identified, PLD1 and PLD2, which share 53% sequence identity and are subject to different regulatory mechanisms. Inhibition of PLD enzymatic activity leads to increased cancer cell apoptosis, decreased cancer cell invasion, and decreased metastasis of cancer cells; therefore, the development of isoform-specific, PLD inhibitors is a novel approach for the treatment of cancer. Previously, we developed potent dual PLD1/PLD2, PLD1-specific (>1700-fold selective), and moderately PLD2-preferring (>10-fold preferring) inhibitors. Here, we describe a matrix library strategy that afforded the most potent (PLD2 IC50 = 20 nM) and selective (75-fold selective versus PLD1) PLD2 inhibitor to date, N-(2-(1-(3-fluorophenyl)-4-oxo-1,3,8-triazaspiro[4.5]decan-8-yl)ethyl)-2-naphthamide (22a), with an acceptable DMPK profile. Thus, these new isoform-selective PLD inhib...
Wahn Soo Choi - One of the best experts on this subject based on the ideXlab platform.
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activation of rbl 2h3 mast cells is dependent on tyrosine phosphorylation of phospholipase d2 by fyn and fgr
Molecular and Cellular Biology, 2004Co-Authors: Ahmed Chahdi, Wahn Soo Choi, Young Mi Kim, Jun Ho Lee, Erk Her, Takaaki Hiragun, Hyoungpyo Kim, Jeung Whan HanAbstract:Phospholipase D (PLD) is activated via receptors in a wide variety of cells where it is thought to regulate intracellular signaling processes and functions such as membrane trafficking, cytoskeletal organization, and degranulation of mast cells (reviewed in references 15, 25, and 31). PLD catalyzes the hydrolysis of phosphatidylcholine to form phosphatidic acid, which is rapidly converted to other biologically active molecules, namely, lysophosphatidic acid and diacylglycerol. In the presence of relatively low concentrations of primary alcohols, the production of phosphatidic acid is diverted to more metabolically inert phosphatidylalcohols by transphosphatidylation, a reaction that is unique to PLD and one that is utilized in the assay of PLD in vivo (39) and to unmask the physiologic roles of phosphatidic acid (62). Two isoforms of PLD have been cloned, PLD1 and PLD2, with PLD1 existing as two variants, PLD1a and PLD1b (11, 21). PLD1 is activated in vitro by small GTPases such as ARF and Rho and protein kinase C (PKC) α in the presence of phosphatidylinositol 1,4-bisphosphate (PIP2) (4, 21, 37, 43, 55). There is also evidence that PLD1 can be regulated in vivo by Rho kinase (48), Ca2+/calmodulin-dependent kinase II (35), and PKC in a catalytically dependent or independent manner (21, 26, 63). PLD2, in contrast, is activated in vitro by PIP2 alone, and this activity is minimally affected by the small GTPases or PKCα (11, 32, 54). However, the mechanisms regulating PLD2 activity in vivo are unclear. There are reports of tyrosine phosphorylation of PLD1 (33, 36) and PLD2 (1, 44, 51) and indications from pharmacological studies that tyrosine phosphorylation may regulate PLD activity (6, 27, 36, 44). In addition, PLD2 was shown to associate with, and be phosphorylated by, the tyrosine kinase receptor for epidermal growth factor (EGF) (51) and by Src kinase (1, 42). Nevertheless, the role of such phosphorylation is uncertain. Although tyrosine-11 was identified as the specific residue phosphorylated in PLD2, mutation of this site enhanced basal PLD2 activity but had no effect on the magnitude of the PLD2 response to EGF (51). Mast cells and blood basophils are responsible for a variety of allergic disorders (5, 59). These cells respond to immunoglobulin E (IgE)-directed antigens via the high-affinity receptor for IgE, namely, FcɛRI, by release of granules that contain preformed inflammatory mediators and the generation of inflammatory lipids and cytokines. PLD is thought to play an essential role in mast cell degranulation (7, 10, 58). PLD is activated in isolated mast cells (12) and cultured mast cell lines (10, 28, 30) by a variety of stimulants, including antigen. Cross-linking of the IgE/FcɛRI complex with antigen results in the recruitment and activation of Src kinases and subsequently other tyrosine kinases. The function of the individual PLD isoforms in mast cells has been studied in the RBL-2H3 cell line, which is now known to be an analog of rat mucosal mast cells (49). Studies with transiently expressed forms of both PLDs in RBL-2H3 cells indicate that PLD1b and PLD2 associate with granule membranes and the plasma membrane, respectively (7, 9), and that both isoforms are activated upon antigen stimulation (8, 40). The mechanisms of activation of these PLDs by antigen are unknown. However, the location of PLD2 at the plasma membrane makes this isoform particularly accessible to FcɛRI-associated tyrosine kinases. As reported here, activation of PLD and degranulation in antigen-stimulated RBL-2H3 cells is inhibited by low concentrations of the Src kinase inhibitor PP2. We investigated whether Src kinases regulate PLD directly by tyrosine phosphorylation and, if so, whether this phosphorylation is essential for degranulation. We show by coexpression studies, site-directed mutagenesis, and the use of small interfering RNAs (siRNAs) directed against Src kinases that Fyn and Fgr phosphorylate PLD2 but not PLD1b in vitro and in vivo and that this phosphorylation is required for the activation of PLD2 in vivo. Furthermore, suppression of this phosphorylation or the activation of PLD2 itself by various strategies also results in suppression of degranulation in stimulated RBL-2H3 cells.
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Mastoparan Selectively Activates Phospholipase D2 in Cell Membranes
The Journal of biological chemistry, 2003Co-Authors: Ahmed Chahdi, Wahn Soo Choi, Young Mi Kim, Michael A. BeavenAbstract:Abstract Both known isoforms of phospholipase (PL) D, PLD1 and PLD2, require phosphatidylinositol 4,5-bisphosphate for activity. However, PLD2 is fully active in the presence of this phospholipid, whereas PLD1 activation is dependent on additional factors such as ADP-ribosylation factor-1 (ARF-1) and protein kinase Cα. We find that mastoparan, an activator of Gi and mast cells, stimulates an intrinsic PLD activity, most likely PLD2, in fractions enriched in plasma membranes from rat basophilic leukemia 2H3 mast cells. Overexpression of PLD2, but not of PLD1, results in a large increase in the mastoparan-inducible PLD activity in membrane fractions, particularly those enriched in plasma membranes. As in previous studies, expressed PLD2 is localized primarily in the plasma membrane and PLD1 in granule membranes. Studies with pertussis toxin and other agents indicate that mastoparan stimulates PLD2 independently of Gi, ARF-1, protein kinase C, and calcium. Kinetic studies indicate that mastoparan interacts synergistically with phosphatidylinositol 4,5-bisphosphate and that oleate, itself a weak stimulant of PLD2 at low concentrations, is a competitive inhibitor of mastoparan stimulation of PLD2. Therefore, mastoparan may be useful for investigating the regulation of PLD2, particularly in view of the well studied molecular interactions of mastoparan with certain other strategic signaling proteins.
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phospholipases d1 and d2 regulate different phases of exocytosis in mast cells
Journal of Immunology, 2002Co-Authors: Wahn Soo Choi, Michael A Frohman, Young Mi Kim, Christian A Combs, Michael A. BeavenAbstract:The rat mast cell line RBL-2H3 contains both phospholipase D (PLD)1 and PLD2. Previous studies with this cell line indicated that expressed PLD1 and PLD2 are both strongly activated by stimulants of secretion. We now show by use of PLDs tagged with enhanced green fluorescent protein that PLD1, which is largely associated with secretory granules, redistributes to the plasma membrane in stimulated cells by processes reminiscent of exocytosis and fusion of granules with the plasma membrane. These processes and secretion of granules are suppressed by expression of a catalytically inactive mutant of PLD1 or by the presence of 50 mM 1-butanol but not tert-butanol, an indication that these events are dependent on the catalytic activity of PLD1. Of note, cholera toxin induces translocation of PLD1-labeled granules to the plasma membrane but not fusion of granules with plasma membrane or secretion. Subsequent stimulation of calcium influx with Ag or thapsigargin leads to rapid redistribution of PLD1 to the plasma membrane and accelerated secretion. Also of note, PLD1 is recycled from plasma membrane back to granules within 4 h of stimulation. PLD2, in contrast, is largely confined to the plasma membrane, but it too participates in the secretory process, because expression of catalytically inactive PLD2 also blocks secretion. These data indicate a two-step process: translocation of granules to the cell periphery, regulated by granule-associated PLD1, and a calcium-dependent fusion of granules with the plasma membrane, regulated by plasma membrane-associated PLD2 and possibly PLD1.
Alex H Brown - One of the best experts on this subject based on the ideXlab platform.
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development of dual pld1 2 and PLD2 selective inhibitors from a common 1 3 8 triazaspiro 4 5 decane core discovery of ml298 and ml299 that decrease invasive migration in u87 mg glioblastoma cells
Journal of Medicinal Chemistry, 2013Co-Authors: Matthew C Oreilly, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Scott J Daniels, Ryan D Morrison, Alex H Brown, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 ...
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development of dual pld1 2 and PLD2 selective inhibitors from a common 1 3 8 triazaspiro 4 5 decane core discovery of ml298 and ml299 that decrease invasive migration in u87 mg glioblastoma cells
Journal of Medicinal Chemistry, 2013Co-Authors: Matthew C Oreilly, Paul G. Thomas, Sarah A Scott, Kyle A Brown, Thomas Oguin, Scott J Daniels, Ryan D Morrison, Alex H Brown, Craig W. LindsleyAbstract:An iterative parallel synthesis effort identified a PLD2 selective inhibitor, ML298 (PLD1 IC50 > 20 000 nM, PLD2 IC50 = 355 nM) and a dual PLD1/2 inhibitor, ML299 (PLD1 IC50 = 6 nM, PLD2 IC50 = 20 nM). SAR studies revealed that a small structural change (incorporation of a methyl group) increased PLD1 activity within this classically PLD2-preferring core and that the effect was enantiospecific. Both probes decreased invasive migration in U87-MG glioblastoma cells.
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autoregulation of phospholipase d activity is coupled to selective induction of phospholipase d1 expression to promote invasion of breast cancer cells
International Journal of Cancer, 2011Co-Authors: Dong Woo Kang, Craig W. Lindsley, Mi Hee Park, Alex H Brown, Young Jun Lee, Hyung Sik Kim, Do Sik MinAbstract:Phospholipase D (PLD) is an important signaling enzyme implicated in the control of many biological processes, including cell proliferation and survival. Despite the importance of the duration and amplitude of PLD signaling in carcinogenesis, mechanisms that regulate PLD expression remain poorly understood. In our study, we define the regulatory components of the machinery that specifies selective PLD1 induction via signals propagated through PLD activity. We demonstrate for the first time that establishment of a positive feedback loop that is dependent on enzymatic activity originating from both PLD1 and PLD2 isozymes enhances selective expression of PLD1, but not PLD2. Phosphatidic acid, the product of PLD activity, leads to an increase in the Ras-ERK/PI3K-NFκB signaling cascade and enhances binding of NFκB to the PLD1 promoter, consequently inducing selective PLD1 expression in SK-BR3 breast cancer cells. Moreover, selective PLD inhibitor suppressed epidermal growth factor-induced matrix metalloproteinase upregulation and invasion by inhibiting PLD1 expression. In conclusion, we propose that autoregulation of PLD activity might be coupled to induction of PLD1 expression, and thereby play a role in carcinogenesis.
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design synthesis and biological evaluation of halogenated n 2 4 oxo 1 phenyl 1 3 8 triazaspiro 4 5 decan 8 yl ethyl benzamides discovery of an isoform selective small molecule phospholipase d2 inhibitor
Journal of Medicinal Chemistry, 2010Co-Authors: Robert R Lavieri, Sarah A Scott, Scott J Daniels, Ryan D Morrison, Alex H Brown, Paige E Selvy, Satyawan Jadhav, Craig W. LindsleyAbstract:Phospholipase D (PLD) catalyzes the conversion of phosphatidylcholine to the lipid second messenger phosphatidic acid. Two mammalian isoforms of PLD have been identified, PLD1 and PLD2, which share 53% sequence identity and are subject to different regulatory mechanisms. Inhibition of PLD enzymatic activity leads to increased cancer cell apoptosis, decreased cancer cell invasion, and decreased metastasis of cancer cells; therefore, the development of isoform-specific, PLD inhibitors is a novel approach for the treatment of cancer. Previously, we developed potent dual PLD1/PLD2, PLD1-specific (>1700-fold selective), and moderately PLD2-preferring (>10-fold preferring) inhibitors. Here, we describe a matrix library strategy that afforded the most potent (PLD2 IC50 = 20 nM) and selective (75-fold selective versus PLD1) PLD2 inhibitor to date, N-(2-(1-(3-fluorophenyl)-4-oxo-1,3,8-triazaspiro[4.5]decan-8-yl)ethyl)-2-naphthamide (22a), with an acceptable DMPK profile. Thus, these new isoform-selective PLD inhib...
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design and synthesis of isoform selective phospholipase d pld inhibitors part i impact of alternative halogenated privileged structures for pld1 specificity
Bioorganic & Medicinal Chemistry Letters, 2009Co-Authors: Jana A. Lewis, Sarah A Scott, Alex H Brown, Robert R Lavieri, Paige E Selvy, Michelle D. Armstrong, Jason R Buck, Sydney L Stoops, Craig W. LindsleyAbstract:This Letter describes the synthesis and structure–activity-relationships (SAR) of isoform-selective PLD inhibitors. By virtue of the installation of alternative halogenated piperidinyl benzimidazolone privileged structures, in combination with a key (S)-methyl group, novel PLD inhibitors with low nM potency and unprecedented levels of PLD1 isoform selectivity (1700-fold) over PLD2 were developed.