A Simple Method to Improve the Odds in Finding 'Lead-Like' Compounds from Chemical Libraries

Horio, Kouhei; Muta, Hajime; Goto, Junki; Hirayama, Noriaki

doi:10.1248/cpb.55.980

Cited by 19 publications

(22 citation statements)

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“…Thrombosis is triggered by abnormal formation of blood clots in the blood stream. Fibrinolysis is a process characterized by the decomposition and liquidation of fibrin mesh formed from blood coagulation, and it involves two stages: activation of fibrinolytic enzyme and degradation of fibrin (Kivirikko et al, 1967; Morishima et al, 1997; Goto et al, 2004; Horio et al, 2007). First, prothrombin is converted to thrombin by prothrombin activator when the coagulation system is activated.…”

Section: Introductionmentioning

confidence: 99%

Inhibition of PAI-1 Activity by Toddalolactone as a Mechanism for Promoting Blood Circulation and Removing Stasis by Chinese Herb Zanthoxylum nitidum var. tomentosum

Zhang

Jin

et al. 2017

Front. Pharmacol.

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Traditional Chinese medicine has been used to treat a variety of human diseases for many centuries. Zanthoxylum nitidum var. tomentosum is used as an adjuvant to promote blood circulation and remove stasis. However, the mechanisms of improving circulation and other biological activities of Z. nitidum var. tomentosum are still unclear. Plasminogen activator inhibitor-1 (PAI-1) regulates the plasminogen activation system through inhibition of tissue-type and urokinase-type plasminogen activators (tPA and uPA). PAI-1 has been linked to fibrin deposition that evolves into organ fibrosis and atherosclerosis. In the present study, we showed that ethanol extract prepared from Z. nitidum var. tomentosum exhibited PAI-1 inhibitory activity, and identified toddalolactone as the main active component that inhibited the activity of recombinant human PAI-1 with IC50 value of 37.31 ± 3.23 μM, as determined by chromogenic assay, and the effect was further confirmed by clot lysis assay. In vitro study showed that toddalolactone inhibited the binding between PAI-1 and uPA, and therefore prevented the formation of the PAI-1/uPA complex. Intraperitoneal injection of toddalolactone in mice significantly prolonged tail bleeding and reduced arterial thrombus weight in a FeCl3-induced thrombosis model. In addition, the hydroxyproline level in the plasma and the degree of liver fibrosis in mice were decreased after intraperitoneal injection of toddalolactone in CCl4-induced mouse liver fibrosis model. Taken together, PAI-1 inhibition exerted by toddalolactone may represent a novel molecular mechanism by which Z. nitidum var. tomentosum manifests its effect in the treatment of thrombosis and fibrosis.

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Section: Introductionmentioning

confidence: 99%

Inhibition of PAI-1 Activity by Toddalolactone as a Mechanism for Promoting Blood Circulation and Removing Stasis by Chinese Herb Zanthoxylum nitidum var. tomentosum

Zhang

Jin

et al. 2017

Front. Pharmacol.

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“…Through the 2 filters we efficiently reduced the number of compounds to about 3000. 18 The second step involved docking simulation of the selected compounds to the PAI-1 molecule by the program Ph4Dock. 19 The crystal structure (1A7C) of the complex between PAI-1 and the inhibitory reactive-center loop peptide was obtained from the Protein Data Bank 20 and used as the target structure for the docking study.…”

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confidence: 99%

Inhibition of Plasminogen Activator Inhibitor-1

Izuhara

Takahashi

Nangaku

et al. 2008

ATVB

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Objective-Serine protease inhibitors (serpin) play a central role in various pathological processes including coagulation, fibrinolysis, malignancy, and inflammation. Inhibition of serpins may prove therapeutic. As yet, however, only very few small molecule serpin inhibitors have been reported. For the first time, we apply a new approach of virtual screening to discover novel, orally active, small molecule serpin inhibitors and report their effectiveness. Methods and Results-We focused on a clinically important serpin, plasminogen activator inhibitor-1 (PAI-1), whose crystal structure has been described. We identify novel, orally active molecules able to enter into the strand 4 position (s4A) of the A ␤-sheet of PAI-I as a mock compound. In vitro they specifically inhibit the PAI-1 activity and enhance fibrinolysis activity. In vivo the most effective molecule (TM5007) inhibits coagulation in 2 models: a rat arteriovenous (AV) shunt model and a mouse model of ferric chloride-induced testicular artery thrombosis. It also prevents the fibrotic process initiated by bleomycin in mouse lung. Conclusions-The present study demonstrates beneficial in vitro and in vivo effects of novel PAI-1 inhibitors. Our methodology proves to be a useful tool to obtain effective inhibitors of serpin activity. Serpins consist of a ␤-sheets-rich body. They contain an exposed mobile reactive center loop (RCL) 2 which, once cleaved by a target serine protease, inserts its N terminus part into the strand 4 position (s4A) of the A ␤-sheet, triggering its antiprotease activity. Small molecule compounds able to enter into the s4A position of the A ␤-sheet as a mock molecule may thus prevent the biological activity of the serpin.Up to now, however, only very few small molecule serpin inhibitors have been described, none of which are in clinical use. Most have been discovered by high-throughput random screening (HTS) of a large chemical library, 3-5 a rather inefficient strategy. In the present study, a new approach of virtual screening based on the 3-dimensional structure of a serpin, (PAI-1), was used to discover novel, orally active, small molecule compounds able to inhibit the target molecule.PAI-1 regulates the plasminogen activation system through inhibition of its target serine proteases, tissue-type and urokinase-type plasminogen activator (tPA and uPA). 1 Studies in humans and animals have demonstrated that PAI-1 expression is enhanced in various disorders such as thrombosis, fibrotic diseases, atherosclerosis, radiation damage, and cancer progression. 6 PAI-1 has been linked with fibrin deposition evolving into organ fibrosis and atherosclerosis, or with striking alterations of cell adhesion and migration mediating cancer progression. 7 The absence of PAI-1 in PAI-1 knockout mice markedly attenuates these pathological processes. 8 -12 Inhibition of PAI-1 by a neutralizing antibody 13 provides similar promising results in animal experiments. Small molecule PAI-1 inhibitors, active orally, should prove useful to treat not only thrombot...

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“…Similarity-based virtual screening (Horio et al, 2007) of a chemical database containing approximately five million entities was performed using two reference structures (staurosporine and phorbol ester). Seventy of the top-ranking compounds were purchased from chemical suppliers through Namiki Shoji Co. Ltd. (Tokyo, Japan).…”

Section: Methodsmentioning

confidence: 99%

Na⁺/H⁺Exchanger 1 Is Regulated via Its Lipid-Interacting Domain, Which Functions as a Molecular Switch: A Pharmacological Approach Using Indolocarbazole Compounds

et al. 2013

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The plasma membrane Na 1 /H 1 exchanger 1 (NHE1) is rapidly activated in response to various stimuli. The membraneproximal cytoplasmic region (∼60 residues), termed the lipidinteracting domain (LID), is an important regulatory domain of NHE1. Here, we used a pharmacological approach to further characterize the role of LID in the regulation of NHE1. Pharmacological analysis using staurosporine-like indolocarbazole and bisindolylmaleimide compounds suggested that the phorbol ester-and receptor agonist-induced activation of NHE1 occurs through a protein kinase C-independent mechanism. In particular, only indolocarbazole compounds that inhibited NHE1 activation were able to interact with the LID, suggesting that the inhibition of NHE1 activation is achieved through the direct action of these compounds on the LID. Furthermore, in addition to phorbol esters and a receptor agonist, okadaic acid and hyperosmotic stress, which are known to activate NHE1 through unknown mechanisms, were found to promote membrane association of the LID concomitant with NHE1 activation; these effects were inhibited by staurosporine, as well as by a mutation in the LID. Binding experiments using the fluorescent ATP analog trinitrophenyl ATP revealed that ATP and the NHE1 activator phosphatidylinositol 4,5-bisphosphate bind competitively to the LID. These findings suggest that modulation of NHE1 activity by various activators and inhibitors occurs through the direct binding of these molecules to the LID, which alters the association of the LID with the plasma membrane. IntroductionThe ubiquitous Na 1 /H 1 exchanger isoform 1 (NHE1) catalyzes acid extrusion across the cell membrane by electroneutral ion exchange coupled to Na 1 influx and serves as a key regulator of intracellular pH (pH i ), Na 1 concentration, and cell volume in virtually all tissues (Wakabayashi et al., 1997;Orlowski and Grinstein, 2004). NHE1 elicits relatively high exchange activity under unstimulated physiologic conditions (basal state) and is further activated in response to various stimuli, including hormones, growth factors, and mechanical stress (activated state). This activation is attributable to a change in its affinity for intracellular H 1 ; thus, it can be easily detected in the neutral pH i range as a stimuli-induced cytoplasmic alkalinization in the absence of bicarbonate (Wakabayashi et al., 1997). Although NHE1 activation plays a physiologically important role in optimizing the intracellular ionic environment, it is also thought to promote the pathogenesis of disease, such as heart failure and cancer (Cardone et al., 2005;Karmazyn et al., 2008;Wakabayashi et al., 2013). In fact, treatment with NHE1 inhibitors reduced the pathologic phenotypes of animal models with acute and chronic heart diseases (Karmazyn et al., 2008).The molecular mechanism of the hormonal activation of NHE1 has been the focus of many investigations. The stimulation of Gq-coupled receptors induces the hydrolysis of phosphatidylinositol 4,5-bisphosphate [PtdIns(4,5)P 2 ] via the a...

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A Simple Method to Improve the Odds in Finding 'Lead-Like' Compounds from Chemical Libraries

Cited by 19 publications

References 4 publications

Inhibition of PAI-1 Activity by Toddalolactone as a Mechanism for Promoting Blood Circulation and Removing Stasis by Chinese Herb Zanthoxylum nitidum var. tomentosum

Inhibition of PAI-1 Activity by Toddalolactone as a Mechanism for Promoting Blood Circulation and Removing Stasis by Chinese Herb Zanthoxylum nitidum var. tomentosum

Inhibition of Plasminogen Activator Inhibitor-1

Na⁺/H⁺Exchanger 1 Is Regulated via Its Lipid-Interacting Domain, Which Functions as a Molecular Switch: A Pharmacological Approach Using Indolocarbazole Compounds

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A Simple Method to Improve the Odds in Finding 'Lead-Like' Compounds from Chemical Libraries

Cited by 19 publications

References 4 publications

Inhibition of PAI-1 Activity by Toddalolactone as a Mechanism for Promoting Blood Circulation and Removing Stasis by Chinese Herb Zanthoxylum nitidum var. tomentosum

Inhibition of PAI-1 Activity by Toddalolactone as a Mechanism for Promoting Blood Circulation and Removing Stasis by Chinese Herb Zanthoxylum nitidum var. tomentosum

Inhibition of Plasminogen Activator Inhibitor-1

Na+/H+Exchanger 1 Is Regulated via Its Lipid-Interacting Domain, Which Functions as a Molecular Switch: A Pharmacological Approach Using Indolocarbazole Compounds

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Na⁺/H⁺Exchanger 1 Is Regulated via Its Lipid-Interacting Domain, Which Functions as a Molecular Switch: A Pharmacological Approach Using Indolocarbazole Compounds