Alternative chemical modifications reverse the binding orientation of a pharmacophore scaffold in the active site of macrophage migration inhibitory factor

Pharmacophores are chemical scaffolds upon which changes in chemical moieties (R-groups) at specific sites are made to identify a combination of R-groups that increases the therapeutic potency of a small molecule inhibitor while minimizing adverse effects. We developed a pharmacophore based on a carbonyloxime (OXIM) scaffold for macrophage migration inhibitory factor (MIF), a protein involved in the pathology of sepsis, to validate that inhibition of a catalytic site could produce therapeutic benefits. We studied the crystal structures of MIF.OXIM-based inhibitors and found two opposite orientations for binding to the active site that were dependent on the chemical structures of an R-group. One orientation was completely unexpected based on previous studies with hydroxyphenylpyruvate and (S,R)-3-(4-hydroxyphenyl)-4,5-dihydro-5-isoxazole acetic acid methyl ester (ISO-1). We further confirmed that the unexpected binding mode targets MIF in cellular studies by showing that one compound, OXIM-11, abolished the counter-regulatory activity of MIF on anti-inflammatory glucocorticoid action. OXIM-11 treatment of mice, initiated 24 h after the onset of cecal ligation and puncture-induced sepsis, significantly improved survival when compared with vehicle-treated controls, confirming that inhibition of the MIF catalytic site could produce therapeutic effects. The crystal structures of the MIF inhibitor complexes provide insight for further structure-based drug design efforts.     

Mutagenicity in a Molecule: Identification of Core Structural Features of Mutagenicity Using a Scaffold Analysis

With advances in the development and application of Ames mutagenicity in silico prediction tools, the International Conference on Harmonisation (ICH) has amended its M7 guideline to reflect the use of such prediction models for the detection of mutagenic activity in early drug safety evaluation processes. Since current Ames mutagenicity prediction tools only focus on functional group alerts or side chain modifications of an analog series, these tools are unable to identify mutagenicity derived from core structures or specific scaffolds of a compound. In this study, a large collection of 6512 compounds are used to perform scaffold tree analysis. By relating different scaffolds on constructed scaffold trees with Ames mutagenicity, four major and one minor novel mutagenic groups of scaffold are identified. The recognized mutagenic groups of scaffold can serve as a guide for medicinal chemists to prevent the development of potentially mutagenic therapeutic agents in early drug design or development phases, by modifying the core structures of mutagenic compounds to form non-mutagenic compounds. In addition, five series of substructures are provided as recommendations, for direct modification of potentially mutagenic scaffolds to decrease associated mutagenic activities.     

Novel analogues of Istaroxime,a potent inhibitor of Na+,K+-ATPase: Synthesis,structure–activity relationship and 3D-quantitative structure–activity relationship of derivatives at position 6 on the androstane scaffold

We report the synthesis and biological properties of novel analogues of Istaroxime acting as positive inotropic compounds through the inhibition of the Na⁺,K⁺-ATPase. We explored the chemical space around the position 6 of the steroidal scaffold by changing the functional groups at that position and maintaining a basic oximic chain in position 3. Some compounds showed inhibitory potencies of the Na⁺,K⁺-ATPase higher than Istaroxime and many of the compounds tested in vivo were safer than digoxin, the classic digitalis compound currently used for the treatment of congestive heart failure as inotropic agent. The 3D-QSAR analyses using comparative molecular field analysis (CoMFA) and comparative molecular similarity indices analysis (CoMSIA) methods have been successfully applied to a set of 63 androstane derivatives as Na⁺,K⁺-ATPase inhibitors. The contour plots provide many useful insights into relationships between structural features and inhibitory potency.     

Synthesis of Pro-XylaneTM: A new biologically active C-glycoside in aqueous media

The scope and limitation of Lubineau’s reaction were evaluated for the synthesis of C-glycosides (compounds 1–13). Further transformation of side chain carbonyl was also achieved (compounds 16–23). Optimization of these two steps was investigated in xylose case. Some of the compounds were shown to stimulate sulfated glycosaminoglycans (GAGs) synthesis. Compound 20 (called Pro-Xylane^TM) was identified as the best activator of GAGs biosynthesis. Pro-Xylane^TM was developed using environmentally friendly conditions relevant to ‘Green-Chemistry’ principles and launched on the market in September 2006. This compound is the first example of ‘Green’ chemical used in cosmetic.     

Bioactive Copper-Doped Glass Scaffolds Can Stimulate Endothelial Cells in Co-Culture in Combination with Mesenchymal Stem Cells

Bioactive glass (BG) scaffolds are being investigated for bone tissue engineering applications because of their osteoconductive and angiogenic nature. However, to increase the in vivo performance of the scaffold, including enhancing the angiogenetic growth into the scaffolds, some researchers use different modifications of the scaffold including addition of inorganic ionic components to the basic BG composition. In this study, we investigated the in vitro biocompatibility and bioactivity of Cu²⁺-doped BG derived scaffolds in either BMSC (bone-marrow derived mesenchymal stem cells)-only culture or co-culture of BMSC and human dermal microvascular endothelial cells (HDMEC). In BMSC-only culture, cells were seeded either directly on the scaffolds (3D or direct culture) or were exposed to ionic dissolution products of the BG scaffolds, kept in permeable cell culture inserts (2D or indirect culture). Though we did not observe any direct osteoinduction of BMSCs by alkaline phosphatase (ALP) assay or by PCR, there was increased vascular endothelial growth factor (VEGF) expression, observed by PCR and ELISA assays. Additionally, the scaffolds showed no toxicity to BMSCs and there were healthy live cells found throughout the scaffold. To analyze further the reasons behind the increased VEGF expression and to exploit the benefits of the finding, we used the indirect method with HDMECs in culture plastic and Cu²⁺-doped BG scaffolds with or without BMSCs in cell culture inserts. There was clear observation of increased endothelial markers by both FACS analysis and acetylated LDL (acLDL) uptake assay. Only in presence of Cu²⁺-doped BG scaffolds with BMSCs, a high VEGF secretion was demonstrated by ELISA; and typical tubular structures were observed in culture plastics. We conclude that Cu²⁺-doped BG scaffolds release Cu²⁺, which in turn act on BMSCs to secrete VEGF. This result is of significance for the application of BG scaffolds in bone tissue engineering approaches.     

Synthesis and characterization of new thiosemicarbazones,as potent urease inhibitors: In vitro and in silico studies

A new series of N-substituted thiosemicarbazones (3a-u) bearing 2-naphthyl and dihydrobenzofuranyl scaffolds were synthesized in good to excellent yields (78–95%). The synthesized compounds were characterized by advanced spectroscopic techniques, such as FTIR, ¹HNMR, ¹³CNMR and ESI-MS and evaluated as urease inhibitors. The structure of compound 3m was unambiguously confirmed by single crystal X-ray analysis. All compounds showed remarkable activities against urease enzyme with IC₅₀ values in range of 1.4–36.1 µM. The majority of the synthesized compounds showed higher activity than the standard compound thiourea. Molecular docking was performed to study the mode of interaction of these compounds and their structure-activity relationship. These studies revealed that the compounds bind at the active site and interacts with the nickel atom present in the binding site. The molecular docking demonstrated excellent co-relations with the experimental findings.     

Identifying the novel inhibitors of lysine-specific demethylase 1 (LSD1) combining pharmacophore-based and structure-based virtual screening

Abstract

Lysine-specific demethylase 1 (LSD1) has been reported to connect with a range of solid tumors. Thus, the exploration of LSD1 inhibitors has emerged as an effective strategy for cancer treatment. In this study, we constructed a pharmacophore model based on a series of flavin adenine dinucleotide (FAD)-competing inhibitors bearing triazole???dithiocarbamate scaffold combining docking, structure–activity relationship (SAR) study, and molecular dynamic (MD) simulation. Meanwhile, another pharmacophore model was also constructed manually, relying on several speculated substrate-competing inhibitors and reported putative vital interactions with LSD1. On the basis of the two pharmacophore models, multi-step virtual screenings (VSs) were performed against substrate-binding pocket and FAD-binding pocket, respectively, combining pharmacophore-based and structure-based strategy to exploit novel LSD1 inhibitors. After bioassay evaluation, four compounds among 21 hits with diverse and novel scaffolds exhibited inhibition activity at the range of 3.63–101.43?μM. Furthermore, substructure-based enrichment was performed, and four compounds with a more potent activity were identified. After that, the time-dependent assay proved that the most potent compound with IC₅₀ 2.21?μM inhibits LSD1 activity in a manner of time-independent. In addition, the compound exhibited a cellular inhibitory effect against LSD1 in MGC-803 cells and may inhibit cell migration and invasion by reversing EMT in cultured gastric cancer cells. Considering the binding mode and SAR of the series of compounds, we could roughly deem that these compounds containing 3-methylxanthine scaffold act through occupying substrate-binding pocket competitively. This study presented a new starting point to develop novel LSD1 inhibitors.     

三种固相微萃取头对厚皮甜瓜果汁香气成分萃取效果分析

采用美国Supelco公司生产的Carboxen^TM/PDMS、DVB/CAR/PDMS和PDMS/DVB 3种固相微萃取头萃取厚皮甜瓜(Cucumis melo L.)果汁中的香气成分,并用气质联用仪对香气成分进行检测。结果表明, 3种萃取头检测到的香气成分均以酯类为主,醇类和醛类较少。Carboxen^TM/PDMS萃取的香气成分相对含量达91.55%,特征香气成分相对含量达到63.01%;DVB/CAR/PDMS也可以全面萃取甜瓜香气,但香气成分与特征香气的相对含量及总峰面积比Carboxen^TM/PDMS的低;PDMS/DVB萃取头吸附的香气成分种类最少,相对含量最低。可见,对厚皮甜瓜果汁香气成分的萃取以Carboxen^TM/PDMS萃取头较好。     

Chitin-based scaffolds are an integral part of the skeleton of the marine demosponge Ianthella basta

The skeletons of demosponges, such as Ianthella basta, are known to be a composite material containing organic constituents. Here, we show that a filigree chitin-based scaffold is an integral component of the I. basta skeleton. These chitin-based scaffolds can be isolated from the sponge skeletons using an isolation and purification technique based on treatment with alkaline solutions. Solid-state ¹³C NMR, Raman, and FT-IR spectroscopies, as well as chitinase digestion, reveal that the isolated material indeed consists of chitin. The morphology of the scaffolds has been determined by light and electron microscopy. It consists of cross-linked chitin fibers approximately 40–100 nm in diameter forming a micro-structured network. The overall shape of this network closely resembles the shape of the integer sponge skeleton. Solid-state ¹³C NMR spectroscopy was used to characterize the sponge skeleton on a molecular level. The ¹³C NMR signals of the chitin-based scaffolds are relatively broad, indicating a high amount of disordered chitin, possibly in the form of surface-exposed molecules. X-ray diffraction confirms that the scaffolds isolated from I. basta consist of partially disordered and loosely packed chitin with large surfaces. The spectroscopic signature of these chitin-based scaffolds is closer to that of α-chitin than β-chitin.     

Identification of a novel selective small-molecule inhibitor of protein arginine methyltransferase 5 (PRMT5) by virtual screening,resynthesis and biological evaluations

As one of the most promising anticancer target in protein arginine methyltransferase (PRMT) family, PRMT5 has been drawing more and more attentions, and many efforts have been devoted to develop its inhibitors. In this study, three PRMT5 inhibitors (9, 16, and 23) with novel scaffolds were identified by performing pharmacophore- and docking-based virtual screening combined with in vitro radiometric-based scintillation proximity assay (SPA). Substructure search based on the scaffold of the most active 9 afforded 26 additional analogues, and SPA results indicated that two analogues (9–1 and 9–2) showed increased PRMT5 inhibitory activity compared with the parental compound. Resynthesis of 9, 9–1, and 9–2 confirmed their PRMT5 enzymatic inhibition activity. In addition, compound 9–1 displayed selectivity against PRMT5 over other key homological members (PRMT1 and CARM1 (PRMT4)). While the structure–activity relationship (SAR) of this series of compounds was discussed to provide clues for further structure optimization, the probable binding modes of active compounds were also probed by molecular docking and molecular dynamics simulations. Finally, the antiproliferative effect of 9–1 on MV4-11 leukemia cell line was confirmed and its impact on regulating the target gene of PRMT5 was also validated. The hit compounds identified in this work have provided more novel scaffolds for future hit-to-lead optimization of small-molecule PRMT5 inhibitors.     

The Boar-Operated-System: a Novel Method to Deliver Baits to Wild Pigs

ABSTRACT Bait-delivered pharmaceuticals, increasingly used to manage populations of wild boar (Sus scrofa) and feral pigs, may be ingested by nontarget species. Species-specificity could be achieved through a delivery system. We designed the BOS^TM (Boar-Operated-System) as a device to deliver baits to wild pigs. The BOS^TM consists of a metal pole onto which a round perforated base is attached. A metal cone with a wide rim slides up and down the pole and fully encloses the base onto which the baits are placed. We conducted a pilot, captive trial and found that captive wild boar fed from the BOS^TM either directly, by lifting the cone, or indirectly, by feeding once another animal had lifted the cone. Thus, we tested whether free-living wild boar fed from the BOS^TM and whether the BOS^TM could prevent bait uptake by nontarget species. We observed that free-living wild boar fed regularly from the BOS^TM and that the device successfully prevented bait uptake by nontarget species. The BOS^TM should be trialed more extensively to confirm its effectiveness and species-specificity to distribute pharmaceuticals to wild suids. If successful, the BOS^TM could be used to deliver vaccines in disease control programs as well as contraceptives to manage overabundant populations of wild suids.     

Sulfonamides containing curcumin scaffold: Synthesis,characterization, carbonic anhydrase inhibition and molecular docking studies

Curcumin is a multi-functional pharmacologically safe natural agent with proven cytoprotective effects to healthy human cells. In this study, a new series of sulfonamides with curcumin scaffold were synthesized, characterized and investigated for their carbonic anhydrase isoenzyme I (human) and II (bovine) isoforms. The structures of newly synthesized compounds were described by IR, ¹H NMR and ¹³C NMR spectral data. Compound 14 showed the K_i value of 0.99 µM with highest inhibitory activity among all other synthesized compounds against hCA-I enzyme. Similarly enzyme kinetic studies of compound 14, 16 and 30 against bCAII enzyme showed Ki values of 0.71, 0.67 and 0.71 µM respectively. Our biological assays results showed that most of active compounds have similar inhibitory activities compared to standard acetazolamide drug. The molecular docking predicted binding modes showed that these compounds bind with hCA-1 enzyme in similar fashion.     

Bioavailability, absorption mechanism, and toxicity of microencapsulated iron (I) sulfate

The iron compounds used for food fortification have to meet certain requisites related to their bioavailability, absorption mechanism, and toxicity, since they will be consumed by a massive population group. With these purposes, we evaluated a new product used for the iron fortification of milk and lacteous derivatives, called SFE-171^TM, which is a ferrous sulfate, microencapsulated with phospholipids. The bioavailability studies were carried out using four groups of 30 female mice each. In two groups, we studied the absorption of ferrous ascorbate and ferrous sulfate, both in water as reference standards, which show absorptions of 13.1±4.9% and 13.2±4.3%, respectively. With the third group, we studied the absorption of ferrous sulfate in milk; its value, 7.9±3.2%, is significantly lower than that of the remaining groups, with ap<0.01. The studies with SFE-171^TM in milk, were performed on the fourth group, with a result of 11.6±4.5%, demonstrating that its absorption does not differ significantly from that of the reference standards. The absorption mechanism was determined by means of in vivo self-displacement studies of the ferrous ion and the SFE-171^TM, taking ferrous sulfate as the reference compound. For this study, 210 female mice were used, and no significant difference between the absorption mechanism of both products could be observed. Toxicity studies of the new product with regard to ferrous sulfate were carried out with two groups of 70 female mice each and two groups of 70 male mice each. The lethal dose 50% LD₅₀ for SFE-171^TM and for ferrous sulfate was 1200 and 680 mg/kg for female mice and 1230 and 670 mg/kg for male mice, respectively, demonstrating that the toxicity of the first product is substantially lower than that of the reference standard. We conclude that the iron product under study has a high bioavailability, an absorption mechanism equal to that of nonhemic iron, and lower toxicity than ferrous sulfate.     

Discovery of new inhibitors against both NF-κB and osteoclastogenesis from in-house library with α, β-unsaturated-enone fragment

The α,β-unsaturated-enone contained natural products have been reported showing NF-κB inhibition effect. It is well known that NF-κB inhibitors can also be used to inhibit osteoclastogenesis. In a continual discovery new agents for anti-osteoclastogenesis, 8 different type compounds with α,β-unsaturated-enone fragments from our in-house library were evaluated for NF-κB inhibition and anti-osteoclastogenesis. Experimental results indicated five compounds exhibited inhibition of NF-κB signal pathway. Among them, one compound ((E)-2-(4-fluorobenzylidene)-3,4-dihydronaphthalen-1(2H)-one, 6a) simultaneously inhibits both osteoclastogenesis and NF-κB signal pathway. Furthermore, 12 compounds with similar scaffold with 6a were tested for anti-osteoclastogenesis. As a result, 9 compounds inhibited both NF-κB and osteoclastogenesis. Among them, compound 6b is the most potent inhibitor against NF-κB (IC₅₀ = 2.09 μM) and osteoclast differentiation (IC₅₀ = 0.86 μM). Further studies show that compound 6b blocks the phosphorylation of both p65 and IκBα, and suppresses NF-κB targeted gene expression without interfering MAPKs and PI3K/Akt signal transduction pathways. This study demonstrates that we can identify promising synthesized compounds with new scaffolds as therapeutic solutions against osteoclastogenesis inspired by the privileged fragment derived from natural leads.     

Biological designer self-assembling peptide nanofiber scaffolds significantly enhance osteoblast proliferation, differentiation and 3-D migration

A class of self-assembling peptide nanofiber scaffolds has been shown to be an excellent biological material for 3-dimension cell culture and stimulating cell migration into the scaffold, as well as for repairing tissue defects in animals. We report here the development of several peptide nanofiber scaffolds designed specifically for osteoblasts. We designed one of the pure self-assembling peptide scaffolds RADA16-I through direct coupling to short biologically active motifs. The motifs included osteogenic growth peptide ALK (ALKRQGRTLYGF) bone-cell secreted-signal peptide, osteopontin cell adhesion motif DGR (DGRGDSVAYG) and 2-unit RGD binding sequence PGR (PRGDSGYRGDS). We made the new peptide scaffolds by mixing the pure RAD16 and designer-peptide solutions, and we examined the molecular integration of the mixed nanofiber scaffolds using AFM. Compared to pure RAD16 scaffold, we found that these designer peptide scaffolds significantly promoted mouse pre-osteoblast MC3T3-E1 cell proliferation. Moreover, alkaline phosphatase (ALP) activity and osteocalcin secretion, which are early and late markers for osteoblastic differentiation, were also significantly increased. We demonstrated that the designer, self-assembling peptide scaffolds promoted the proliferation and osteogenic differentiation of MC3T3-E1. Under the identical culture medium condition, confocal images unequivocally demonstrated that the designer PRG peptide scaffold stimulated cell migration into the 3-D scaffold. Our results suggest that these designer peptide scaffolds may be very useful for promoting bone tissue regeneration.     

Identification of natural compounds as potent inhibitors of SARS-CoV-2 main protease using combined docking and molecular dynamics simulations

Coronavirus disease 2019 (COVID-19) has emerged from China and globally affected the entire population through the human-to-human transmission of a newly emerged virus called severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). The genome of SARS-CoV-2 encodes several proteins that are essential for multiplication and pathogenesis. The main protease (M^pro or 3CL^pro) of SARS-CoV-2 plays a central role in its pathogenesis and thus is considered as an attractive drug target for the drug design and development of small-molecule inhibitors. We have employed an extensive structure-based high-throughput virtual screening to discover potential natural compounds from the ZINC database which could inhibit the M^pro of SARS-CoV-2. Initially, the hits were selected on the basis of their physicochemical and drug-like properties. Subsequently, the PAINS filter, estimation of binding affinities using molecular docking, and interaction analyses were performed to find safe and potential inhibitors of SARS-CoV-2 M^pro. We have identified ZINC02123811 (1-(3-(2,5,9-trimethyl-7-oxo-3-phenyl-7H-furo[3,2-g]chromen-6-yl)propanoyl)piperidine-4-carboxamide), a natural compound bearing appreciable affinity, efficiency, and specificity towards the binding pocket of SARS-CoV-2 M^pro. The identified compound showed a set of drug-like properties and preferentially binds to the active site of SARS-CoV-2 M^pro. All-atom molecular dynamics (MD) simulations were performed to evaluate the conformational dynamics, stability and interaction mechanism of M^pro with ZINC02123811. MD simulation results indicated that M^pro with ZINC02123811 forms a stable complex throughout the trajectory of 100 ns. These findings suggest that ZINC02123811 may be further exploited as a promising scaffold for the development of potential inhibitors of SARS-CoV-2 M^pro to address COVID-19.     

Discovery of novel PTP1B inhibitors via pharmacophore-oriented scaffold hopping from Ertiprotafib

An integrated molecular design strategy combining pharmacophore recognition and scaffold hopping was exploited to discover novel PTP1B inhibitors based on the known PTP1B inhibitor Ertiprotafib. A composite pharmacophore model was proposed from the interaction mode of Ertiprotafib, and 21 diverse molecules from five distinct structural classes were designed and synthesized accordingly. New compounds with considerable inhibition against PTP1B were identified from each series, and the most active compound 3a showed IC₅₀ value of 1.3 μmol L^?1 against human recombinant PTP1B. Docking study indicated that the new inhibitors assumed binding modes similar to that of Ertiprotafib.     

Development of a highly sensitive in vitro phototoxicity assay using the SkinEthicTM reconstructed human epidermis

The reconstituted human epidermis model SkinEthic^TM was used to evaluate the phototoxicity of topically applied chemicals. For comparison with published data, we first tested a library of 13 nonphototoxic (NPT) and phototoxic (PT) compounds, applied onto SkinEthic^TM reconstituted human epidermal tissues, in a protocol as close as possible to the one described by Liebsch using another skin tissue model. The results showed that, under these nonoptimized conditions, the SkinEthic^TM model was already able to fully discriminate between known NPT and PT compounds. Furthermore, these epidermal tissues being highly resistant to UVA irradiation, it was possible to increase irradiation by (at least) 3-fold without decrease in tissue viability. In such conditions, the phototoxicity assay is much more sensitive, so that the model is expected to be of great interest for the detection not only of strong but also of weak phototoxic compounds. This revised version was published online in July 2006 with corrections to the Cover Date.