Rational design of specific ligands for human serum albumin separation and applications.

Human serum albumin is widely used in clinical practice, and the development of new ligands with high affinity is beneficial to improve its separation efficiency. The Site II of human serum albumin is an active binding site of various molecules such as l-tryptophan, which was studied with molecular simulation to obtain insights for the design of new ligands. The results showed that the carboxyl and indolyl groups of l-tryptophan were critical for the binding on Site II. Seven ligands containing carboxyl groups and indolyl groups were designed, and molecular simulation showed that indole-3-pentanoic acid was the best ligand. A new ligand combined indole-3-acetic acid and cysteine was designed for easier resin preparation, and molecular simulation also indicated that the new ligand bound strongly to Site II. Resins with the new ligand designed was prepared and static adsorption experiments indicated that the new resin had high adsorption capacity of human serum albumin and strong salt tolerance. Finally, recombinant human serum albumin was separated from yeast broth with high purity of 90.4% and recovery of 94.2%, which indicated that the new resin had good adsorption selectivity and strong potential for applications.

Molecular-docking-guided design and synthesis of new IAA-tacrine hybrids as multifunctional AChE/BChE inhibitors.

A series of new indole-3-acetic acid (IAA)-tacrine hybrids as dual acetylcholinesterase (AChE)/butyrylcholinesterase (BChE) inhibitors were designed and prepared based on the molecular docking mode of AChE with an IAA derivative (1a), a moderate AChE inhibitor identified by screening our compound library for anti-Alzheimer's disease (AD) drug leads. The enzyme assay results revealed that some hybrids, e.g. 5d and 5e, displayed potent dual in vitro inhibitory activities against AChE/BChE with IC50 values in low nanomolar range. Molecular modeling studies in tandem with kinetic analysis suggest that these hybrids target both catalytic active site and peripheral anionic site of cholinesterase (ChE). Molecular dynamic simulations and Molecular Mechanics/Poisson-Boltzmann Surface Area (MM-PBSA) calculations indicate that 5e has more potent binding affinity than hit 1a, which may explain the stronger inhibitory effect of 5e on AChE. Furthermore, their predicted pharmacokinetic properties and in vitro influences on mouse brain neural network electrical activity were discussed. Taken together, compound 5e can be highlighted as a lead compound worthy of further optimization for designing new anti-AD drugs.

New hybrid type strigolactone mimics derived from plant growth regulator auxin.

Strigolactones (SLs) constitute a new class of plant hormones of increasing importance in plant science. The structure of natural SLs is too complex for ready access by synthesis. Therefore, much attention is being given to design of SL analogues and mimics with a simpler structure but with retention of bioactivity. Here new hybrid type SL mimics have been designed derived from auxins, the common plant growth regulators. Auxins were simply coupled with the butenolide D-ring using bromo (or chloro) butenolide. D-rings having an extra methyl group at the vicinal C-3' carbon atom, or at the C-2' carbon atom, or at both have also been studied. The new hybrid type SL mimics were bioassayed for germination activity of seeds of the parasitic weeds S. hermonthica, O. minor and P. ramosa using the classical method of counting germinated seeds and a colorimetric method. For comparison SL mimics derived from phenyl acetic acid were also investigated. The bioassays revealed that mimics with a normal D-ring had appreciable to good activity, those with an extra methyl group at C-2' were also appreciably active, whereas those with a methyl group in the vicinal C-3' position were inactive (S. hermonthica) or only slightly active. The new hybrid type mimics may be attractive as potential suicidal germination agents in agronomic applications.

Syntheses of the plant auxin conjugate 2-O-(indole-3-acetyl)-myo-inositol IAInos.

The plant hormone conjugate 2-O-(indole-3-acetyl)-myo-inositol (IAInos) has been selectively prepared for the first time by two routes from myo-inositol. One of the syntheses depended upon the construction of the 3-indoleacetyl group by a Fischer indole synthesis on an unreactive axial hydroxyl group, while the other via a direct acylation of the equatorially orientated hydroxy group created by conformational constraint of the cyclohexane ring. The latter synthesis produced IAInos in 5 steps and 29% overall yield.

The Tetrazole Analogue of the Auxin Indole-3-acetic Acid Binds Preferentially to TIR1 and Not AFB5.

Indole-3-acetic acid (auxin) is considered one of the cardinal hormones in plant growth and development. It regulates a wide range of processes throughout the plant. Synthetic auxins exploit the auxin-signaling pathway and are valuable as herbicidal agrochemicals. Currently, despite a diversity of chemical scaffolds all synthetic auxins have a carboxylic acid as the active core group. By applying bio-isosteric replacement we discovered that indole-3-tetrazole was active by surface plasmon resonance spectrometry, showing that the tetrazole could initiate assembly of the Transport Inhibitor Resistant 1 (TIR1) auxin coreceptor complex. We then tested the tetrazole's efficacy in a range of whole plant physiological assays and in protoplast reporter assays, which all confirmed auxin activity, albeit rather weak. We then tested indole-3-tetrazole against the AFB5 homologue of TIR1, finding that binding was selective against TIR1, absent with AFB5. The kinetics of binding to TIR1 are contrasted to those for the herbicide picloram, which shows the opposite receptor preference, as it binds to AFB5 with far greater affinity than to TIR1. The basis of the preference of indole-3-tetrazole for TIR1 was revealed to be a single residue substitution using molecular docking, and assays using tir1 and afb5 mutant lines confirmed selectivity in vivo. Given the potential that a TIR1-selective auxin might have for unmasking receptor-specific actions, we followed a rational design, lead optimization campaign, and a set of chlorinated indole-3-tetrazoles was synthesized. Improved affinity for TIR1 and the preference for binding to TIR1 was maintained for 4- and 6-chloroindole-3-tetrazoles, coupled with improved efficacy in vivo. This work expands the range of auxin chemistry for the design of receptor-selective synthetic auxins.

Weed resistance to synthetic auxin herbicides.

Herbicides classified as synthetic auxins have been most commonly used to control broadleaf weeds in a variety of crops and in non-cropland areas since the first synthetic auxin herbicide (SAH), 2,4-D, was introduced to the market in the mid-1940s. The incidence of weed species resistant to SAHs is relatively low considering their long-term global application with 30 broadleaf, 5 grass, and 1 grass-like weed species confirmed resistant to date. An understanding of the context and mechanisms of SAH resistance evolution can inform management practices to sustain the longevity and utility of this important class of herbicides. A symposium was convened during the 2nd Global Herbicide Resistance Challenge (May 2017; Denver, CO, USA) to provide an overview of the current state of knowledge of SAH resistance mechanisms including case studies of weed species resistant to SAHs and perspectives on mitigating resistance development in SAH-tolerant crops. © 2017 The Authors. Pest Management Science published by John Wiley & Sons Ltd on behalf of Society of Chemical Industry.

Design, Synthesis, and Evaluation of Novel Auxin Mimic Herbicides.

Due to the key roles of auxins as master regulators of plant growth, there is considerable interest in the development of compounds with auxin-like properties for growth management and weed control applications. Herein, we describe the design and multistep synthesis of ten compounds bearing combinations of functional groups commonly associated with auxin-type properties. Following synthesis, these compounds were tested against multiple weed species as well as sweet corn. In general, while these structures were not quite as active as commercial auxin mimic herbicides, multiple compounds exhibited broadleaf weed activity with concurrent selectivity in sweet corn (Zea mays L. var. saccharum). In addition, differential results were observed upon subtle changes to structure, providing insights into the structural properties required for activity.

The discovery of Arylex™ active and Rinskor™ active: Two novel auxin herbicides.

Multiple classes of commercially important auxin herbicides have been discovered since the 1940s including the aryloxyacetates (2,4-D, MCPA, dichlorprop, mecoprop, triclopyr, and fluroxypyr), the benzoates (dicamba), the quinoline-2-carboxylates (quinclorac and quinmerac), the pyrimidine-4-carboxylates (aminocyclopyrachlor), and the pyridine-2-carboxylates (picloram, clopyralid, and aminopyralid). In the last 10 years, two novel pyridine-2-carboxylate (or picolinate) herbicides were discovered at Dow AgroSciences. This paper will describe the structure activity relationship study that led to the discovery of the 6-aryl-picolinate herbicides Arylex™ active (2005) and Rinskor™ active (2010). While Arylex was developed primarily for use in cereal crops and Rinskor is still in development primarily for use in rice crops, both herbicides will also be utilized in additional crops.

Synthesis, physicochemical characterization and pharmacological investigation of indolacin-5-fluorouracil-1-ylmethyl ester.

The objective of this work is to synthesize indolacin-5-fluorouracil-1-ylmethyl ester and the structure was confirmed by means of UV, IR, 1H-NMR, 13C-NMR and mass spectrometry. The physicochemical parameters of melting point, solubility, apparent partition coefficient were investigated. S180 sarcoma, H22 hapatitic cancer and Lewis-transplanted mice were used to evaluate the anti-tumor activity of indolacini-5-fluorouracil-1-ylmethyl ester compared with 5-fluorouracil in vivo. Anti-inflammatory and analgesic activities were evaluated in mice. The inhibitory ratio of indolacini- 5-fluorouracil-1-ylmethyl ester is comparative to that of 5-fluorouracil. This study indicates that 5-fluorouracil-1-ylmethyl ester may represent a new anticancer predrug of 5-fluorouracil to produce a combined effect of indolacin and 5-fluorouracil for cancer therapy.

Design, synthesis, and biological evaluation of a series of alkoxy-3-indolylacetic acids as peroxisome proliferator-activated receptor γ/δ agonists.

A series of alkoxy-3-indolylacetic acid analogs has been discovered as peroxisome proliferator-activated receptor (PPAR) agonists. Structure-activity relationship study indicated that PPARα/γ/δ activities were dependent on the nature of the hydrophobic group, the attachment position of the alkoxy linker to the indole ring, and N-alkylation of indole nitrogen. Some compounds presented significant PPARγ/δ activity and molecular modeling suggested their putative binding modes in the ligand binding domain of PPARγ. Of these, compound 51 was selected for in vivo study via an evaluation of microsomal stability in mouse and human liver. Compound 51 lowered the levels of fasting blood glucose, insulin, and HbA1c without gain in body weight in db/db mice. When compound 51 was treated, hepatic triglycerides level and the size of adipocytes in white adipose tissue of db/db mice were also reduced as opposed to treatment with rosiglitazone. Taken together, compound 51 shows high potential warranting further studies in models for diabetes and related metabolic disorders and may be in use as a chemical tool for the understanding of PPAR biology.

Synthesis of a high specific activity methyl sulfone tritium isotopologue of fevipiprant (NVP-QAW039).

The synthesis of a triple tritiated isotopologue of the CRTh2 antagonist NVP-QAW039 (fevipiprant) with a specific activity >3 TBq/mmol is described. Key to the high specific activity is the methylation of a bench-stable dimeric disulfide precursor that is in situ reduced to the corresponding thiol monomer and methylated with [(3)H3]MeONos having per se a high specific activity. The high specific activity of the tritiated active pharmaceutical ingredient obtained by a build-up approach is discussed in the light of the specific activity usually to be expected if hydrogen tritium exchange methods were applied.

Design, synthesis and biological evaluation of GY3-based derivatives for anti-type 2 diabetes activity.

A series of indole carboxylic acid derivatives were designed and synthesized. Their anti-type 2 diabetes activity was evaluated in HepG2 cell and db/db mice. The results showed that compounds 8c, 17a, 17b, 15a and 15b could significantly increase glucose consumption in HepG2 cell. Furthermore, compound 8c was able to lower the blood glucose level and induce less weight gain in db/db mice.

Radiosynthesis of 3-indolyl[1-¹¹C]acetic acid for phyto-PET-imaging: an improved production procedure and formulation method.

An improved production procedure and formulation method for the carbon-11 radiolabeled phytohormone, 3-indolyl-[l-(11)C]acetic acid ([(11)C]IAA), was developed by modifying selected original reaction parameters. This updated procedure both doubled the yield (from 25.9±6.7% (n=12) to 61.0±0.3% (n=10)) and increased the concentration (0.2-0.4 GBq/0.15-0.3 mL), enabling us to provide the radiotracer [(11)C]IAA suitable for in vivo phyto-PET-imaging studies. The specific activity was improved by more than a factor of three (26.7±5.6 GBq/µmol to 82.5±36.1 GBq/µmol). The total synthesis time for both production and formulation was 81.8±3.0 min (n=10). In addition, a streamlined semi-remote controlled production system, containing five processing modules, was designed and built for routine [(11)C]IAA production. This integrated system facilitated routine high radiation level production of [(11)C]IAA while minimizing radiation exposure to the production chemists.

Natural vs synthetic auxin: studies on the interactions between plant hormones and biological membrane lipids.

Analysis of the interactions between two representatives of plant hormones: synthetic (1-naphthaleneacetic acid, NAA) as well as natural (indole-3-acetic acid, IAA) and phospholipids occurring in biological membrane of both plant and animal cells was the subject of present studies. The aim of undertaken experiments was to elucidate the problem of direct influence of these plant growth regulators on phosphatidylcholines (PCs) and phosphatidylethanolamines (PEs) in monolayers at the air/water solution interface. The studied phospholipids differ not only as regards the structure of polar head-groups but also in the length of hydrophobic chains as well as their saturation degree. These differences result also in the main properties and functions of these phospholipids in biomembranes. The analysis of the results was based on the characteristics of the surface pressure (π)--area (A) isotherms registered for monolayers spread on the subphase containing plant hormone and as a reference on the surface of pure water. Moreover, as a complementary technique, Brewster angle microscopy was applied for the direct visualization of the investigated surface films. The obtained results revealed that auxins effectively influence phospholipids monolayers, regardless of the lipid structure, at the concentration of 10(-4)M. It was found that for this concentration, the influence of auxins was visibly larger in the case of PCs as compared to PEs. On the other hand, in the case of auxins solution of ≤ 10(-5)M, the observed trend was opposite. Generally, our studies showed that the natural plant hormone (IAA) interacts with the investigated lipid monolayers stronger than its synthetic derivative (NAA). The reason of these differences connects with the steric properties of both auxins; namely, the naphthalene ring of NAA molecule occupies larger space than the indole system of IAA. Therefore molecules of the latter compound penetrate easier into the region of phospholipids׳ polar head-groups. Moreover, the NH group of the indole moiety is capable of hydrogen bond formation with the acceptor groups in the polar fragment of lipid molecules. We proved also that among the investigated phospholipids, the highest susceptibility toward auxin influence show these lipids, for which during compression, surface film increases the degree of condensation.

Design, synthesis and structure-activity relationship studies of novel and diverse cyclooxygenase-2 inhibitors as anti-inflammatory drugs.

Because of the pivotal role of cyclooxygenase (COX) in the inflammatory processes, non-steroidal anti-inflammatory drugs (NSAIDs) that suppress COX activities have been used clinically for the treatment of inflammatory diseases/syndromes; however, traditional NSAIDs exhibit serious side-effects such as gastrointestinal damage and hyper sensitivity owing to their COX-1 inhibition. Also, COX-2 inhibition-derived suppressive or preventive effects against initiation/proliferation/invasion/motility/recurrence/metastasis of various cancers/tumours such as colon, gastric, skin, lung, liver, pancreas, breast, prostate, cervical and ovarian cancers are significant. In this study, design, synthesis and structure-activity relationship (SAR) of various novel {2-[(2-, 3- and/or 4-substituted)-benzoyl, (bicyclic heterocycloalkanophenyl)carbonyl or cycloalkanecarbonyl]-(5- or 6-substituted)-1H-indol-3-yl}acetic acid analogues were investigated to seek and identify various chemotypes of potent and selective COX-2 inhibitors for the treatment of inflammatory diseases, resulting in the discovery of orally potent agents in the peripheral-inflammation model rats. The SARs and physicochemical properties for the analogues are described as significant findings. For graphical abstract: see Supplementary Material. ( www.informahealthcare.com/enz ).

General synthesis of ß-alanine-containing spider polyamine toxins and discovery of nephila polyamine toxins 1 and 8 as highly potent inhibitors of ionotropic glutamate receptors.

Certain spiders contain large pools of polyamine toxins, which are putative pharmacological tools awaiting further discovery. Here we present a general synthesis strategy for this class of toxins and prepare five structurally varied polyamine toxins. Electrophysiological testing at three ionotropic glutamate receptor subtypes reveals that two of these, Nephila polyamine toxins 1 (NPTX-1) and 8 (NPTX-8), comprise intriguing pharmacological activities by having subnanomolar IC(50) values at kainate receptors.

Diazine indole acetic acids as potent, selective, and orally bioavailable antagonists of chemoattractant receptor homologous molecule expressed on Th2 cells (CRTH2) for the treatment of allergic inflammatory diseases.

New classes of CRTH2 antagonists, the pyridazine linker containing indole acetic acids, are described. The initial hit 1 had good potency but poor permeability, metabolic stability, and PK. Initial optimization led to compounds of type 2 with low oxidative metabolism but poor oral bioavailability. Poor permeability was identified as a liability for these compounds. Addition of a linker between the indole and diazine moieties afforded a series with good potency, low rates of metabolism, moderate permeability, and good oral bioavailability in rodents. 32 was identified as the development track candidate. It was potent in cell based, binding, and whole blood assays and exhibited good PK profile. It was efficacious in mouse models of contact hypersensitivity (1 mg/kg b.i.d.) and house dust (20 mg/kg q.d.) when dosed orally. In sheep asthma, administration at 1 mg/kg iv completely blocked the LAR and AHR and attenuated the EAR phase.

Rational design of an auxin antagonist of the SCF(TIR1) auxin receptor complex.

The plant hormone auxin is a master regulator of plant growth and development. By regulating rates of cell division and elongation and triggering specific patterning events, indole 3-acetic acid (IAA) regulates almost every aspect of plant development. The perception of auxin involves the formation of a ternary complex consisting of an F-box protein of the TIR1/AFB family of auxin receptors, the auxin molecule, and a member the Aux/IAA family of co-repressor proteins. In this study, we identified a potent auxin antagonist, α-(phenylethyl-2-oxo)-IAA, as a lead compound for TIR1/AFB receptors by in silico virtual screening. This molecule was used as the basis for the development of a more potent TIR1 antagonist, auxinole (α-[2,4-dimethylphenylethyl-2-oxo]-IAA), using a structure-based drug design approach. Auxinole binds TIR1 to block the formation of the TIR1-IAA-Aux/IAA complex and so inhibits auxin-responsive gene expression. Molecular docking analysis indicates that the phenyl ring in auxinole would strongly interact with Phe82 of TIR1, a residue that is crucial for Aux/IAA recognition. Consistent with this predicted mode of action, auxinole competitively inhibits various auxin responses in planta. Additionally, auxinole blocks auxin responses of the moss Physcomitrella patens, suggesting activity over a broad range of species. Our works not only substantiates the utility of chemical tools for plant biology but also demonstrates a new class of small molecule inhibitor of protein-protein interactions common to mechanisms of perception of other plant hormones, such as jasmonate, gibberellin, and abscisic acid.

Structural requirements to obtain highly potent and selective 18 kDa Translocator Protein (TSPO) Ligands.

The (18 kDa) Translocator Protein (TSPO), was initially identified in 1977 as peripheral binding site for the benzodiazepine diazepam and named "Peripheral-type benzodiazepine receptor (PBR)". It is an evolutionarily well-conserved protein particularly located at the outer/inner mitochondrial membrane contact sites, in closely association with the 32 kDa voltage-dependent anion channel (VDAC) and the 30 kDa adenine nucleotide translocase (ANT), thus forming the mitochondrial permeability transition pore (MPTP). TSPO is ubiquitary expressed in peripheral tissues (steroid producing tissues, liver, heart, kidney, lung, immune system) and in lower levels in the central nervous system, where it is mainly located in glial cells, and in neurons. TSPO is involved in a variety of biological processes such as cholesterol transport, steroidogenesis, calcium homeostasis, lipid metabolism, mitochondrial oxidation, cell growth and differentiation, apoptosis induction, and regulation of immune functions. In the last decade, many studies have reported that TSPO basal expression is altered in a number of human pathologies, such as cancer and neurodegenerative disorders (Huntington's and Alzheimer's diseases), as well as in various forms of brain injury and inflammation and anxiety. Consequently, TSPO has not only been suggested as a promising drug target for a number of therapeutic applications (anticonvulsant, anxiolytic, immunomodulating, etc.), but also as valid diagnostic marker for related-disease state and progression, prompting the development of specific labelled ligands as powerful tools for imaging techniques. A number of structurally different classes of ligands have been reported, showing high affinity and selectivity towards TSPO. Indeed, most of these ligands have been designed starting from selective CBR ligands which were structurally modified in order to shift their affinity towards TSPO. Extensive structure-activity relationship studies were performed allowing to hypothesize various TSPO pharmacophore models. The purpose of this paper is to highlight the structural requirements needed to obtain TSPO ligands with high affinity and selectivity.

Preparation of magnetic indole-3-acetic acid imprinted polymer beads with 4-vinylpyridine and ß-cyclodextrin as binary monomer via microwave heating initiated polymerization and their application to trace analysis of auxins in plant tissues.

Auxin is a crucial phytohormone for precise control of growth and development of plants. Due to its low concentration in plant tissues which are rich in interfering substances, the accurate determination of auxins remains a challenge. In this paper, a new strategy for isolation and enrichment of auxins from plant tissues was obtained by the magnetic molecularly imprinted polymer (mag-MIP) beads, which were prepared by microwave heating initiated suspension polymerization using indole-3-acetic acid (IAA) as template. In order to obtain higher selective recognition cavities, an enhanced imprinting method based on binary functional monomers, 4-vinylpyridine (4-VP) and ß-cyclodextrin (ß-CD), was adopted for IAA imprinting. The morphological and magnetic characteristics of the mag-MIP beads were characterized by scanning electron microscopy, Fourier-transform infrared spectroscopy and vibrating sample magnetometry. A majority of resultant beads were within the size range of 80-150µm. Porous surface morphology and good magnetic property were observed. Furthermore, the mag-MIP beads fabricated with 4-VP and ß-CD as binary functional monomers exhibited improved recognition ability to IAA, as compared with the mag-MIP beads prepared with the individual monomer separately. Competitive rebinding experiment results revealed that the mag-MIP beads exhibited a higher specific recognition for the template than the non-imprinted polymer (mag-NIP) beads. An extraction method by mag-MIP beads coupled with high performance liquid chromatography (HPLC) was developed for determination of IAA and indole-3-butyric acid (IBA) in plant tissues. Linear ranges for IAA and IBA were in the range of 7.00-100.0µgL(-1) and 10.0-100.0µgL(-1), and the detection limits were 3.9 and 7.4µgL(-1), respectively. The analytical performance was also estimated by seedlings or immature embryos samples from three different plant tissues, pea, rice and wheat. Recoveries were in the range of 70.1-93.5%. The results show that the present imprinting method is a promising approach for preparation of selective adsorbents for sample preparation of auxin analysis in plant tissues.