FLAP Inhibitor Research Articles

Identification of selective 5-LOX and FLAP inhibitors as novel anti-inflammatory agents by ligand-based virtual screening

Inflammation is a multifaceted biological process in which the conversion of arachidonic acid to eicosanoids, including prostaglandins and leukotrienes (LTs), plays a crucial role. 5-Lipoxygenase (5-LOX) is a key enzyme in cellular LT biosynthesis, and it is supported by the accessory protein 5-lipoxygenase-activating protein (FLAP). Pharmacological interventions to modulate LTs aim at either decreasing their biosynthesis or at mitigating their biological effects. Therefore, inhibiting 5-LOX or FLAP represents a useful strategy to reduce inflammation. Herein we present the identification and pharmacological evaluation of novel inhibitors targeting 5-LOX or FLAP. By means of a ligand-based virtual screening approach, we selected 38 compounds for in vitro assays. Among them, ALR-38 exhibits direct 5-LOX inhibition, while ALR-6 and ALR-27 showed potential as FLAP inhibitors. These latter not only reduced LT production but also promoted the generation of specialized pro-resolving mediators in specific human macrophage phenotypes. Interestingly, the identified compounds turned out to be selective for their respective targets, as none of them displayed activity towards microsomal prostaglandin E2 synthase-1 and soluble epoxide hydrolase, which are other proteins involved in eicosanoid biosynthesis. Thus, these compounds are endowed with potential therapeutic utility in mitigating inflammatory responses and might offer a venue for tackling inflammation-based disorders.

Identification of Novel 5-Lipoxygenase-Activating Protein (FLAP) Inhibitors by an Integrated Method of Pharmacophore Virtual Screening, Docking, QSAR and ADMET Analyses

This study explored a series of reported 5-lipoxygenase-activating protein (FLAP) inhibitors to understand their structural requirements and identify potential new inhibitor scaffolds through automated unbiased procedures. Docking studies have revealed that inhibitor binding affinity can be influenced by several key binding interactions with Phe114 and Lys116 from chain B and Val21, Phe25, His28 and Lys29 from chain C in the FLAP-binding site. A ligand-based alignment three-dimensional (3D)-quantitative structure–activity relationship (QSAR) was adopted, resulting in a robust model with a statistically significant noncross-validated coefficient ([Formula: see text]), a cross-validated correlation coefficient ([Formula: see text]) and a predictive squared correlation coefficient ([Formula: see text]). Overall, the analysis revealed the important electrostatic and steric attributes responsible for the FLAP inhibitory activity, which appeared to correlate well with the docking results. In addition, two statistically significant two-dimensional (2D)-QSAR models ([Formula: see text], [Formula: see text] and [Formula: see text], [Formula: see text]) were developed by a genetic function approximation (GFA). HypoGen 1, a proposed pharmacophore model, was used for database mining to identify potential new FLAP inhibitors. The bioactivity of the retrieved hits was then evaluated in silico based on the validated QSAR models, followed by pharmacokinetics and toxicity predictions.

Quinazoline-4(3H)-one-7-carboxamide Derivatives as Human Soluble Epoxide Hydrolase Inhibitors with Developable 5-Lipoxygenase Activating Protein Inhibition.

Soluble epoxide hydrolase (sEH) metabolizes epoxyeicosatrienoic acids (EETs), which are endowed with beneficial biological activities as they reduce inflammation, regulate endothelial tone, improve mitochondrial function, and decrease oxidative stress. Therefore, inhibition of sEH for maintaining high EET levels is implicated as a new therapeutic modality with broad clinical applications for metabolic, renal, and cardiovascular disorders. In our search for new sEH inhibitors, we designed and synthesized novel amide analogues of the quinazolinone-7-carboxylic acid derivative 5, a previously discovered 5-lipoxygenase-activating protein (FLAP) inhibitor, to evaluate their potential for inhibiting sEH. As a result, we identified new quinazolinone-7-carboxamides that demonstrated selective sEH inhibition with decreased FLAP inhibitor properties. The tractable SAR results indicated that the amide and thiobenzyl fragments flanking the quinazolinone nucleus are critical features governing the potent sEH inhibition, and compounds 34, 35, 37, and 43 inhibited the sEH activity with IC50 values of 0.30–0.66 μM. Compound 34 also inhibited the FLAP-mediated leukotriene biosynthesis (IC50 = 2.91 μM). In conclusion, quinazolinone-7-carboxamides can be regarded as novel lead structures, and newer analogues with improved efficiency against sEH along with or without FLAP inhibition can be generated.

Combined Machine Learning and GRID-Independent Molecular Descriptor (GRIND) Models to Probe the Activity Profiles of 5-Lipoxygenase Activating Protein Inhibitors.

Leukotrienes (LTs) are pro-inflammatory lipid mediators derived from arachidonic acid (AA), and their high production has been reported in multiple allergic, autoimmune, and cardiovascular disorders. The biological synthesis of leukotrienes is instigated by transfer of AA to 5-lipoxygenase (5-LO) via the 5-lipoxygenase-activating protein (FLAP). Suppression of FLAP can inhibit LT production at the earliest level, providing relief to patients requiring anti-leukotriene therapy. Over the last 3 decades, several FLAP modulators have been synthesized and pharmacologically tested, but none of them could be able to reach the market. Therefore, it is highly desirable to unveil the structural requirement of FLAP modulators. Here, in this study, supervised machine learning techniques and molecular modeling strategies are adapted to vaticinate the important 2D and 3D anti-inflammatory properties of structurally diverse FLAP inhibitors, respectively. For this purpose, multiple machine learning classification models have been developed to reveal the most relevant 2D features. Furthermore, to probe the 3D molecular basis of interaction of diverse anti-inflammatory compounds with FLAP, molecular docking studies were executed. By using the most probable binding poses from docking studies, the GRIND model was developed, which indicated the positive contribution of four hydrophobic, two hydrogen bond acceptor, and two shape-based features at certain distances from each other towards the inhibitory potency of FLAP modulators. Collectively, this study sheds light on important two-dimensional and three-dimensional structural requirements of FLAP modulators that can potentially guide the development of more potent chemotypes for the treatment of inflammatory disorders.

Intrinsic 5-lipoxygenase activity regulates migration and adherence of mantle cell lymphoma cells

Human B-lymphocytes express 5-lipoxygenase (5-LOX) and 5-LOX activating protein (FLAP) and can convert arachidonic acid to leukotriene B4. Mantle cell lymphoma (MCL) cells contain similar amounts of 5-LOX as human neutrophils but the function and mechanism of activation of 5-LOX in MCL cells, and in normal B-lymphocytes, are unclear. Here we show that the intrinsic 5-LOX pathway in the MCL cell line JeKo-1 has an essential role in migration and adherence of the cells, which are important pathophysiological characteristics of B-cell lymphoma. Incubation of JeKo-1 with the FLAP inhibitor GSK2190915 or the 5-LOX inhibitor zileuton, at a concentration below 1 μM, prior to stimulation with the chemotactic agent CXCL12, led to a significant reduction of migration. CRISPR/Cas9 mediated deletion of ALOX5 gene in JeKo-1 cells also led to a significantly decreased migration of the cells. Furthermore, 5-LOX and FLAP inhibitors markedly decreased the adherence of JeKo-1 cells to stromal cells. In comparison, these drugs had a similar effect on adherence of JeKo-1 cells as the Bruton tyrosine kinase inhibitor ibrutinib, which has a proven anti-tumour effect. These results indicate that inhibition of 5-LOX may be a novel treatment for MCL and certain other B-cell lymphomas.

In Vitro Effects of 5-Lipoxygenase Pathway Inhibition on Rhinovirus-Associated Bronchial Epithelial Inflammation.

IntroductionThe leukotriene pathway may be implicated in the induction of virus-induced inflammation. Respiratory epithelial cells may express low levels of 5-lipoxygenase (5-LO) and release leukotrienes (LTs) C4, D4, and E4, upon exposure to viruses or other stimuli. Enhanced expression of 5-LO pathway proteins after rhinovirus (RV) infection has previously been described. We hypothesized that anti-leukotriene treatment of epithelial cells, with or without exposure to RV-infected peripheral blood mononuclear cells (PBMCs)-conditioned media, may inhibit RV-induced up-regulation of inflammatory cytokines.MethodsPBMCs from a healthy donor were exposed to RV1B and supernatants were harvested at 48 h post infection. BEAS-2B cells were infected with RV, with or without conditioning with the PBMC supernatant. Treatment with anti-LT agents was performed either on both PBMCs and BEAS-2B or at the bronchial epithelial level only, with varying concentrations of montelukast (CysLT receptor antagonist) or MK-886 [FLAP(5-lipoxygenase-activating-protein) inhibitor]. Evaluation of the inflammatory cytokines IL-8, RANTES, IL-11, IL-6, and IP-10 was performed using ELISA.ResultsOur results show that anti-LT treatment of RV-infected bronchial epithelial cells suppresses epithelial RV-mediated cytokine production, independent of conditioning.ConclusionsThis observation may represent an indirect mode of action of the anti-leukotrienes in virus-induced asthma.Supplementary InformationThe online version contains supplementary material available at 10.1007/s41030-021-00152-x.

Towards a systematic analysis of structure-activity relationships of 5-LOX inhibitors through activity landscape and chemotype enrichment

Pharmacological inhibition of the 5-lipoxygenase (5-LOX) pathway used to inhibit leukotrienes’ production (LT) has been shown to have therapeutic properties in many respiratory and allergic diseases. Because of the toxicity effect of FDA approved orally active 5-LOX inhibitor, zileuton, the scientific community is seeking novel 5-LOX inhibitors. As a result, the significant and relevant amount of structure-activity information of 5-LOX inhibitors and its activating protein inhibitors, i.e., FLAP inhibitors, has been released and stored in public databases. To the best of our knowledge, there are no detailed analyzes of their structure-activity relationship (SAR) so far. This study has tried to locate, characterize, and extract SAR from the ChEMBL database and present them intuitively using activity landscape analysis and chemotype enrichment study. We have found eight important activity cliff generators and more than six cyclic systems with a large proportion of active molecules for each protein. These molecules can provide key information on the pharmacophoric regions that are significant to determine their potency. Therefore, they can be used for lead optimization. Besides, this study found that the smooth SAR region in the 5-LOX chemical space’s activity landscape opens up the possibility of developing robust and reliable QSAR models.

Fingerprint-based computational models of 5-lipo-oxygenase activating protein inhibitors: Activity prediction and structure clustering.

Inflammatory diseases can be treated by inhibiting 5-lipo-oxygenase activating protein (FLAP). In this study, a data set containing 2,112 FLAP inhibitors was collected. A total of 25 classification models were built by five machine learning algorithms with five different types of fingerprints. The best model, which was built by support vector machine algorithm with ECFP_4 fingerprint had an accuracy and a Matthews correlation coefficient of 0.862 and 0.722 on the test set, respectively. The predicted results were further evaluated by the application domain dSTD-PRO (a distance between one compound to models). Each compound had a dSTD-PRO value, which was calculated by the predicted probabilities obtained from all 25 models. The application domain results suggested that the reliability of predicted results depended mainly on the compounds themselves rather than algorithms or fingerprints. A group of customized 10-bit fingerprint was manually defined for clustering the molecular structures of 2,112 FLAP inhibitors into eight subsets by K-Means. According to the clustering results, most of inhibitors in two subsets (subsets 2 and 4) were highly active inhibitors. We found that aryl oxadiazole/oxazole alkanes, biaryl amino-heteroarenes, two aromatic rings (often N-containing) linked by a cyclobutene group, and 1,2,4-triazole group were typical fragments in highly active inhibitors.

A cheminformatic study on chemical space characterization and diversity analysis of 5-LOX inhibitors

The process of blocking 5-lipoxygenase (5-LOX) catalyzed leukotriene biosynthesis has been recognized for the past few decades as a promising therapeutic strategy for acute inflammatory, allergic, and respiratory diseases. Due to the toxicity effect of FDA approved 5-LOX inhibitor zileuton, novel 5-LOX inhibitors have been sought by the scientific community. As a result, a significant and relevant amount of information on the structure-activity of 5-LOX inhibitors has been released and stored in public databases. In this study, we aimed at the comprehensive cheminformatic characterization of the diversity and complexity of the chemical space of 5-LOX inhibitors and its activating protein FLAP inhibitors by comparing it with the Approved drug space and virtual LOX library. The visual representation of the property space indicates some compounds in the 5-LOX inhibitors space broaden the traditional medicinal space. The structural diversity of the databases is computed using complementary approaches, including Physicochemical Property (PCP) descriptors, molecular fingerprints, and molecular scaffold. With the apparent exception of approved drugs, the 5-LOX dataset shows more diversity compared to FLAP and LOX virtual library set. This study was able to identify the underlying patterns in the chemical and pharmacological properties space that were decisive for the drug discovery and development of 5-LOX inhibitors.

Development of a highly sensitive liquid chromatography–mass spectrometry method to quantify plasma leukotriene E4 and demonstrate pharmacological suppression of endogenous 5-LO pathway activity in man

Low basal endogenous concentrations (<20 pg/mL) of the 5-lipoxygenase (5-LO) pathway biomarker leukotriene E4 (LTE4) in human plasma present a significant analytical challenge. Analytical methods including liquid chromatography–mass spectrometry and enzyme linked immunosorbent assays have been used to quantify plasma LTE4 in the past but have not provided consistent data in the lower pg/mL-range. With our new method, a detection limit (<1 pg/mL plasma) significantly below basal levels of LTE4 was achieved by combining large volume sample purification and enrichment by anion-exchange mixed mode solid phase extraction (SPE) with large volume injection followed by chromatographic separation by ultra performance liquid chromatography (UPLC) and quantification by highly sensitive negative-ion electrospray tandem mass spectrometry (MS/MS). The method was reproducible, accurate and linear between 1 and 120 pg/mL plasma LTE4. The method was used to perform an analysis of plasma samples collected from healthy volunteers in a Phase 1 study with the FLAP (5-lipoxygenase activating protein) inhibitor AZD5718. Basal endogenous LTE4 levels of 5.1 ± 2.7 pg/mL were observed in healthy volunteers (n = 34). In subjects that had been administered a single oral dose of AZD5718, significant suppression (>80%) of plasma LTE4 level was observed, providing pharmacological evidence that endogenous 5-LO pathway activity could be assessed.

Discovery of Novel 5-Lipoxygenase-Activating Protein (FLAP) Inhibitors by Exploiting a Multistep Virtual Screening Protocol.

Leukotrienes (LTs) are proinflammatory mediators derived from arachidonic acid (AA), which play significant roles in inflammatory diseases. The 5-lipoxygenase-activating protein (FLAP) is an integral membrane protein, which is essential for the initial step in LT biosynthesis. The aim of this study was to discover novel and chemically diverse FLAP inhibitors for treatment of inflammatory diseases requiring anti-LT therapy. Both ligand- and structure-based approaches were applied to explain the activities of known FLAP inhibitors in relation to their predicted binding modes. We gained valuable insights into the binding modes of the inhibitors by molecular modeling and generated a multistep virtual screening (VS) workflow in which 6.2 million compounds were virtually screened, and the molecular hypotheses were validated by testing VS-hit compounds biologically. The most potent hit compounds showed significant inhibition of FLAP-dependent cellular LT biosynthesis with IC50 values in the range from 0.13 to 0.87 μM. Collectively, this study provided novel bioactive chemotypes with potential for further development as effective anti-inflammatory drugs.

Concise Synthesis of a Potential 5-Lipoxygenase Activating Protein (FLAP) Inhibitor and Its Analogs through Late-Stage Alkene Dicarbofunctionalization

We report a five-step synthesis of the biologically important 1,1-diarylalkane 1, a potential 5-lipoxygenase activating protein (FLAP) inhibitor that was synthesized previously in 12 steps. In this synthesis, we apply a three-component alkene dicarbofunctionalization reaction as a key final step to assemble the potential FLAP inhibitor 1 from commercially available starting materials. In addition, we also report the synthesis of a variety of new analogs of the inhibitor 1 and its regioisomer.

Novel Chemical Series of 5-Lipoxygenase-Activating Protein Inhibitors for Treatment of Coronary Artery Disease.

5-Lipoxygenase (5-LO)-activating protein (FLAP) inhibitors have proven to attenuate 5-LO pathway activity and leukotriene production in human clinical trials. However, previous clinical candidates have been discontinued and the link between FLAP inhibition and outcome in inflammatory diseases remains to be established. We here describe a novel series of FLAP inhibitors identified from a screen of 10k compounds and the medicinal chemistry strategies undertaken to progress this series. Compound 4i showed good overall properties and a pIC50 hWBfree of 8.1 and an lipophilic ligand efficiency of 5.2. Target engagement for 4i was established in dogs using ex vivo measurement of leukotriene B4 (LTB4) levels in blood with good correlation to in vitro potency. A predicted human dose of 280 mg b.i.d. suggests a wide margin to any identified in vitro off-target effects and sufficient exposure to achieve an 80% reduction of LTB4 levels in humans. Compound 4i is progressed to preclinical in vivo safety studies.

Discovery and Early Clinical Development of an Inhibitor of 5-Lipoxygenase Activating Protein (AZD5718) for Treatment of Coronary Artery Disease.

5-Lipoxygenase activating protein (FLAP) inhibitors attenuate 5-lipoxygenase pathway activity and reduce the production of proinflammatory and vasoactive leukotrienes. As such, they are hypothesized to have therapeutic benefit for the treatment of diseases that involve chronic inflammation including coronary artery disease. Herein, we disclose the medicinal chemistry discovery and the early clinical development of the FLAP inhibitor AZD5718 (12). Multiparameter optimization included securing adequate potency in human whole blood, navigation away from Ames mutagenic amine fragments while balancing metabolic stability and PK properties allowing for clinically relevant exposures after oral dosing. The superior safety profile of AZD5718 compared to earlier frontrunner compounds allowed us to perform a phase 1 clinical study in which AZD5718 demonstrated a dose dependent and greater than 90% suppression of leukotriene production over 24 h. Currently, AZD5718 is evaluated in a phase 2a study for treatment of coronary artery disease.

Targeting biosynthetic networks of the proinflammatory and proresolving lipid metabolome.

Nonsteroidal anti-inflammatory drugs interfere with the metabolism of arachidonic acid to proinflammatory prostaglandins and leukotrienes by targeting cyclooxygenases (COXs), 5-lipoxygenase (LOX), or the 5-LOX-activating protein (FLAP). These and related enzymes act in conjunction with marked crosstalk within a complex lipid mediator (LM) network where also specialized proresolving LMs (SPMs) are formed. Here, we present how prominent LM pathways can be differentially modulated in human proinflammatory M1 and proresolving M2 macrophage phenotypes that, upon exposure to Escherichia coli, produce either abundant prostaglandins and leukotrienes (M1) or SPMs (M2). Targeted liquid chromatography-tandem mass spectrometry-based metabololipidomics was applied to analyze and quantify the specific LM profiles. Besides expected on-target actions, we found that: 1) COX or 15-LOX-1 inhibitors elevate inflammatory leukotriene levels, 2) FLAP and 5-LOX inhibitors reduce leukotrienes in M1 but less so in M2 macrophages, 3) zileuton blocks resolution-initiating SPM biosynthesis, whereas FLAP inhibition increases SPM levels, and 4) that the 15-LOX-1 inhibitor 3887 suppresses SPM formation in M2 macrophages. Conclusively, interference with discrete LM biosynthetic enzymes in different macrophage phenotypes considerably affects the LM metabolomes with potential consequences for inflammation-resolution pharmacotherapy. Our data may allow better appraisal of the therapeutic potential of these drugs to intervene with inflammatory disorders.-Werner, M., Jordan, P. M., Romp, E., Czapka, A., Rao, Z., Kretzer, C., Koeberle, A., Garscha, U., Pace, S., Claesson, H.-E., Serhan, C. N., Werz, O., Gerstmeier, J. Targeting biosynthetic networks of the proinflammatory and proresolving lipid metabolome.

Identification of multi-target inhibitors of leukotriene and prostaglandin E2 biosynthesis by structural tuning of the FLAP inhibitor BRP-7

Leukotrienes (LTs) and prostaglandin (PG)E2 are enzymatically produced from arachidonic acid and represent highly bioactive lipid mediators with pro-inflammatory functions. Here, we report on novel multi-target inhibitors that potently and dually interfere with 5-lipoxygenase-activating protein (FLAP) and microsomal prostaglandin E2 synthase (mPGES)-1 in LT and PGE2 biosynthesis, based on the previously identified selective FLAP inhibitor BRP-7 (8, IC50 = 0.31 μM). C (5)-substitution of the benzimidazole ring of BRP-7 by carboxylic acid and its bioisosteres provided compounds, exemplified by 57 that potently suppress LT formation (IC50 = 0.05 μM) by targeting FLAP along with inhibition of mPGES-1 (IC50 = 0.42 μM). Besides FLAP, also 5-lipoxygenase (5-LO) and LTC4 synthase activities were inhibited by 57, albeit with lower potency (IC50 = 0.6 and 6.2 μM) than FLAP. Docking studies and molecular dynamic simulations with FLAP, mPGES-1 and 5-LO provide valuable insights into potential binding interactions of the inhibitors with their targets. Together, these novel benzimidazole derivatives may possess potential as leads for development of effective anti-inflammatory drugs with multi-target properties for dually inhibiting LT and PGE2 production.

Synthesis of a 5-Lipoxygenase Activating Protein (FLAP) Inhibitor

Key words BI 665915 - FLAP inhibitor - classical resolution - 1,2,4-oxadiazole ring formation - Suzuki–Miyaura cross-coupling

Ligand-based Modeling for the Prediction of Pharmacophore Features for Multi-targeted Inhibition of the Arachidonic Acid Cascade.

The single-target drugs against the arachidonic acid inflammatory pathway are associated with serious side effects, hence, as a first step towards multi-target drugs, we have studied the pharmacophoric features common to the inhibitors of 5-lipoxygenase-activating protein (FLAP), microsomal prostaglandin E-synthase 1 (mPGES-1) and leukotriene A4 hydrolase (LTA4H). FLAP and mPGES-1 shared subfamily-specific positions (SSPs) and four mPGES-1 inhibitors binding to them mapped onto the pharmacophore derived from FLAP inhibitors (Ph-FLAP). The reactions of mPGES-1 and LTA4H had high structural similarity. The pharmacophore derived from two substrate mimic inhibitors of LTA4H (Ph-LTA4H) also mapped onto three mPGES-1 inhibitors. Screening of in-house database for Ph-FLAP and Ph-LTA4H identified one compound, C1. It inhibited the production of the mPGES-1 product, prostaglandin E2 (PGE2) by 97.8±1.6 % at 50 μM in HeLa cells and can be a starting point for designing molecules inhibiting all three targets simultaneously.

Discovery and optimization of oxadiazole-based FLAP inhibitors

Structure activity relationship (SAR) investigation of an oxadiazole based series led to the discovery of several potent FLAP inhibitors. Lead optimization focused on achieving functional activity while improving physiochemical properties and reducing hERG inhibition. Several compounds with favorable in vitro and in vivo properties were identified that were suitable for advanced profiling.

Pharmacological profile and efficiency in vivo of diflapolin, the first dual inhibitor of 5-lipoxygenase-activating protein and soluble epoxide hydrolase

Arachidonic acid (AA) is metabolized to diverse bioactive lipid mediators. Whereas the 5-lipoxygenase-activating protein (FLAP) facilitates AA conversion by 5-lipoxygenase (5-LOX) to pro-inflammatory leukotrienes (LTs), the soluble epoxide hydrolase (sEH) degrades anti-inflammatory epoxyeicosatrienoic acids (EETs). Accordingly, dual FLAP/sEH inhibition might be advantageous drugs for intervention of inflammation. We present the in vivo pharmacological profile and efficiency of N-[4-(benzothiazol-2-ylmethoxy)-2-methylphenyl]-N′-(3,4-dichlorophenyl)urea (diflapolin) that dually targets FLAP and sEH. Diflapolin inhibited 5-LOX product formation in intact human monocytes and neutrophils with IC50 = 30 and 170 nM, respectively, and suppressed the activity of isolated sEH (IC50 = 20 nM). Characteristic for FLAP inhibitors, diflapolin (I) failed to inhibit isolated 5-LOX, (II) blocked 5-LOX product formation in HEK cells only when 5-LOX/FLAP was co-expressed, (III) lost potency in intact cells when exogenous AA was supplied, and (IV) prevented 5-LOX/FLAP complex assembly in leukocytes. Diflapolin showed target specificity, as other enzymes related to AA metabolism (i.e., COX1/2, 12/15-LOX, LTA4H, LTC4S, mPGES1, and cPLA2) were not inhibited. In the zymosan-induced mouse peritonitis model, diflapolin impaired vascular permeability, inhibited cysteinyl-LTs and LTB4 formation, and suppressed neutrophil infiltration. Diflapolin is a highly active dual FLAP/sEH inhibitor in vitro and in vivo with target specificity to treat inflammation-related diseases.