| 1. |
- Bandaru, Sashidar, et al.
(författare)
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Lack of RAC1 in macrophages protects against atherosclerosis.
- 2020
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Ingår i: PLoS One. - : Public Library of Science (PLoS). - 1932-6203. ; 15:9
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Tidskriftsartikel (refereegranskat)abstract
- The Rho GTPase RAC1 is an important regulator of cytoskeletal dynamics, but the role of macrophage-specific RAC1 has not been explored during atherogenesis. We analyzed RAC1 expression in human carotid atherosclerotic plaques using immunofluorescence and found higher macrophage RAC1 expression in advanced plaques compared with intermediate human atherosclerotic plaques. We then produced mice with Rac1-deficient macrophages by breeding conditional floxed Rac1 mice (Rac1fl/fl) with mice expressing Cre from the macrophage-specific lysosome M promoter (LC). Atherosclerosis was studied in vivo by infecting Rac1fl/fl and Rac1fl/fl/LC mice with AdPCSK9 (adenoviral vector overexpressing proprotein convertase subtilisin/kexin type 9). Rac1fl/fl/LC macrophages secreted lower levels of IL-6 and TNF-α and exhibited reduced foam cell formation and lipid uptake. The deficiency of Rac1 in macrophages reduced the size of aortic atherosclerotic plaques in AdPCSK9-infected Rac1fl/fl/LC mice. Compare with controls, intima/media ratios, the size of necrotic cores, and numbers of CD68-positive macrophages in atherosclerotic plaques were reduced in Rac1-deficient mice. Moreover, we found that RAC1 interacts with actin-binding filamin A. Macrophages expressed increased RAC1 levels in advanced human atherosclerosis. Genetic inactivation of RAC1 impaired macrophage function and reduced atherosclerosis in mice, suggesting that drugs targeting RAC1 may be useful in the treatment of atherosclerosis.
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| 2. |
- Battisti, Umberto Maria, et al.
(författare)
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Exploration of Novel Urolithin C Derivatives as Non-Competitive Inhibitors of Liver Pyruvate Kinase
- 2023
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Ingår i: Pharmaceuticals. - : MDPI AG. - 1424-8247. ; 16:5
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Tidskriftsartikel (refereegranskat)abstract
- The inhibition of liver pyruvate kinase could be beneficial to halt or reverse non-alcoholic fatty liver disease (NAFLD), a progressive accumulation of fat in the liver that can lead eventually to cirrhosis. Recently, urolithin C has been reported as a new scaffold for the development of allosteric inhibitors of liver pyruvate kinase (PKL). In this work, a comprehensive structure-activity analysis of urolithin C was carried out. More than 50 analogues were synthesized and tested regarding the chemical features responsible for the desired activity. These data could pave the way to the development of more potent and selective PKL allosteric inhibitors.
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| 3. |
- Battisti, U. M., et al.
(författare)
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Serendipitous Identification of a Covalent Activator of Liver Pyruvate Kinase
- 2023
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Ingår i: ChemBioChem. - : Wiley. - 1439-4227 .- 1439-7633. ; 24:1
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Tidskriftsartikel (refereegranskat)abstract
- Enzymes are effective biological catalysts that accelerate almost all metabolic reactions in living organisms. Synthetic modulators of enzymes are useful tools for the study of enzymatic reactions and can provide starting points for the design of new drugs. Here, we report on the discovery of a class of biologically active compounds that covalently modifies lysine residues in human liver pyruvate kinase (PKL), leading to allosteric activation of the enzyme (EC50=0.29 μM). Surprisingly, the allosteric activation control point resides on the lysine residue K282 present in the catalytic site of PKL. These findings were confirmed by structural data, MS/MS experiments, and molecular modelling studies. Altogether, our study provides a molecular basis for the activation mechanism and establishes a framework for further development of human liver pyruvate kinase covalent activators.
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| 4. |
- Glise, Lars, 1988, et al.
(författare)
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pH-Dependent Protonation of Histidine Residues Is Critical for Electrostatic Binding of Low-Density Lipoproteins to Human Coronary Arteries
- 2022
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Ingår i: Arteriosclerosis Thrombosis and Vascular Biology. - : Ovid Technologies (Wolters Kluwer Health). - 1079-5642 .- 1524-4636. ; 42:8, s. 1037-1047
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Tidskriftsartikel (refereegranskat)abstract
- Background: The initiating step in atherogenesis is the electrostatic binding of LDL (low-density lipoprotein) to proteoglycan glycosaminoglycans in the arterial intima. However, although proteoglycans are widespread throughout the intima of most coronary artery segments, LDL is not evenly distributed, indicating that LDL retention is not merely dependent on the presence of proteoglycans. We aim to identify factors that promote the interaction between LDL and the vessel wall of human coronary arteries. Methods: We developed an ex vivo model to investigate binding of labeled human LDL to human coronary artery sections without the interference of cellular processes. Results: By staining consecutive sections of human coronary arteries, we found strong staining of sulfated glycosaminoglycans throughout the arterial intima, whereas endogenous LDL deposits were focally distributed. Ex vivo binding of LDL was uniform at all intimal areas with sulfated glycosaminoglycans. However, lowering the pH from 7.4 to 6.5 triggered a 35-fold increase in LDL binding. The pH-dependent binding was abolished by pretreating LDL with diethyl-pyrocarbonate, which blocks the protonation of histidine residues, or cyclohexanedione, which inhibits the positive charge of site B on LDL. Thus, both histidine protonation and site B are required for strong electrostatic LDL binding to the intima. Conclusions: This study identifies histidine protonation as an important component for electrostatic LDL binding to human coronary arteries. Our findings show that the local pH will have a profound impact on LDL's affinity for sulfated glycosaminoglycans, which may influence the retention and accumulation pattern of LDL in the arterial vasculature.
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| 5. |
- Nain-Perez, Amalyn, et al.
(författare)
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Anthraquinone derivatives as ADP-competitive inhibitors of liver pyruvate kinase
- 2022
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Ingår i: European Journal of Medicinal Chemistry. - : Elsevier BV. - 0223-5234 .- 1768-3254. ; 234
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Tidskriftsartikel (refereegranskat)abstract
- Liver pyruvate kinase (PKL) is a major regulator of metabolic flux and ATP production during liver cell glycolysis and is considered a potential drug target for the treatment of non-alcoholic fatty liver disease (NAFLD). In this study, we report the first ADP-competitive PKL inhibitors and identify several starting points for the further optimization of these inhibitors. Modeling and structural biology guided the optimization of a PKL-specific anthraquinone-based compound. A structure-activity relationship study of 47 novel synthetic derivatives revealed PKL inhibitors with half-maximal inhibitory concentration (IC50) values in the 200 nM range. Despite the difficulty involved in studying a binding site as exposed as the ADP site, these derivatives feature expanded structural diversity and chemical spaces that may be used to improve their inhibitory activities against PKL. The obtained results expand the knowledge of the structural requirements for interactions with the ADP-binding site of PKL.(C) 2022 The Authors. Published by Elsevier Masson SAS.& nbsp;
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| 6. |
- Nain-Perez, Amalyn, et al.
(författare)
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Tuning liver pyruvate kinase activity up or down with a new class of allosteric modulators
- 2023
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Ingår i: European Journal of Medicinal Chemistry. - : Elsevier BV. - 0223-5234. ; 250
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Tidskriftsartikel (refereegranskat)abstract
- The liver isoform of pyruvate kinase (PKL) has gained interest due to its potential capacity to regulate fatty acid synthesis involved in the progression of non-alcoholic fatty liver disease (NAFLD). Here we describe a novel series of PKL modulators that can either activate or inhibit the enzyme allosterically, from a cryptic site at the interface of two protomers in the tetrameric enzyme. Starting from urolithin D, we designed and synthesised 42 new compounds. The effect of these compounds on PKL enzymatic activity was assessed after incubation with cell lysates obtained from a liver cell line. Pronounced activation of PKL activity, up to 3.8-fold, was observed for several compounds at 10 mu M, while other compounds were prominent PKL inhibitors reducing its activity to 81% at best. A structure-activity relationship identified linear-shaped sulfone-sulfonamides as activators and nonlinear compounds as inhibitors. Crystal structures revealed the conformations of these modulators, which were used as a reference for designing new modulators.
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| 7. |
- Xu, Yongjin, et al.
(författare)
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Design and development of a photoswitchable DFG-out kinase inhibitor
- 2021
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Ingår i: Chemical Communications. - : Royal Society of Chemistry (RSC). - 1359-7345 .- 1364-548X. ; 57, s. 10043-10046
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Tidskriftsartikel (refereegranskat)abstract
- We report the synthesis and characterisation of a photoswitchable DFG-out kinase inhibitor. Photocontrol of the target kinase in both enzymatic and living cell assays is demonstrated.
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| 8. |
- Xu, Yongjin, et al.
(författare)
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Design and development of photoswitchable DFG-Out RET kinase inhibitors
- 2022
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Ingår i: European Journal of Medicinal Chemistry. - : Elsevier BV. - 0223-5234 .- 1768-3254. ; 234
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Tidskriftsartikel (refereegranskat)abstract
- REarranged during Transfection (RET) is a transmembrane receptor tyrosine kinase that is required for development of multiple human tissues, but which is also an important contributor to human cancers. RET activation through rearrangement or point mutations occurs in thyroid and lung cancers. Further-more, activation of wild type RET is an increasingly recognized mechanism promoting tumor growth and dissemination of a much broader group of cancers. RET is therefore an attractive therapeutic target for small-molecule kinase inhibitors. Non-invasive control of RET signaling with light offers the promise of unveiling its complex spatiotemporal dynamics in vivo. In this work, photoswitchable DFG-out RET kinase inhibitors based on heterocycle-derived azobenzenes were developed, enabling photonic control of RET activity. Based on the binding mode of DFG-out kinase inhibitors and using RET kinase as the test model, we developed a photoswitchable inhibitor with a quinoline "head " constituting the azoheter-oarene. This azo compound was further modified by three different strategies to increase the difference in biological activity between the E-isomer and the light enriched Z-isomer. Stilbene-based derivatives were used as model compounds to guide in the selection of substituents that could eventually be introduced to the corresponding azo compounds. The most promising quinoline-based compound showed more than a 15-fold difference in bioactivity between the two isomers in a biochemical assay. However, the same compound showed a decreased Z/E (IC50) ratio in the cellular assay, tentatively assigned to stability issues. The corresponding stilbene compound gave a Z/E (IC50) ratio well above 100, consistent with that measured in the biochemical assay. Ultimately, a 7-azaindole based photoswitchable DFG-out kinase inhibitor was shown to display more than a 10-fold difference in bioactivity between the two isomers, in both a biochemical and a cell-based assay, as well as excellent stability even under reducing conditions.(C) 2022 The Authors. Published by Elsevier Masson SAS.
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