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1.  Immunosuppression by Co-stimulatory Molecules: Inhibition of CD2-CD48/CD58 Interaction by Peptides from CD2 to Suppress Progression of Collagen-induced Arthritis in Mice 
Chemical biology & drug design  2013;82(1):106-118.
Targeting co-stimulatory molecules to modulate the immune response has been shown to have useful therapeutic effects for autoimmune diseases. Among the co-stimulatory molecules, CD2 and CD58 are very important in the early stages of generation of an immune response. Our goal was to utilize CD2-derived peptides to modulate protein-protein interactions between CD2 and CD58, thereby modulating the immune response. Several peptides were designed based on the structure of the CD58 binding domain of CD2 protein. Among the CD2-derived peptides, peptide 6 from the F and C β-strand region of CD2 protein exhibited inhibition of cell-cell adhesion in the nanomolar concentration range. Peptide 6 was evaluated for its ability to bind to CD58 in Caco-2 cells and to CD48 in T cells from rodents. A molecular model was proposed for binding a peptide to CD58 and CD48 using docking studies. Furthermore, in vivo studies were carried out to evaluate the therapeutic ability of the peptide to modulate the immune response in the collagen-induced arthritis (CIA) mouse model. In vivo studies indicated that peptide 6 was able to suppress the progression of CIA. Evaluation of the antigenicity of peptides in CIA and transgenic animal models indicated that this peptide is not immunogenic.
doi:10.1111/cbdd.12138
PMCID: PMC3707497  PMID: 23530775
CD2; CD58; collagen-induced arthritis; cyclic peptide; docking; immune response; protein-protein interaction
2.  SYNTHESIS, ANTIDEPRESSANT EVALUATION AND DOCKING STUDIES OF LONG-CHAIN ALKYLNITROQUIPAZINES AS SEROTONIN TRANSPORTER INHIBITORS 
Chemical biology & drug design  2013;81(6):695-706.
Twelve alkyl analogues (1 - 12) of the high-affinity serotonin transporter (SERT) inhibitor 6-nitroquipazine (6-NQ) were synthesised and studied using in vitro radioligand competition binding assays to determine their binding affinity (Ki). The putative antidepressant activity of five of the binders with the highest SERT binding affinities was studied by the forced swim and locomotor activity mouse tests. The three dimensional (3D) structures of 8 and 9 were determined using NOE NMR technique. Flexible docking of the compounds was undertaken to illustrate the binding of the compounds in the SERT model. Our results showed that several of the 6-NQ analogues are high-affinity SERT inhibitors and indicated that the octyl (8), decyl (10) and dodecyl (12) 6-NQ analogues exhibit moderate antidepressant activity.
doi:10.1111/cbdd.12116
PMCID: PMC3665732  PMID: 23574807
Serotonin transporter; 6-nitroquipazine alkyl analogues; Radioligand competition assay; NMR spectroscopy; Flexible docking; Porsolt forced swim test
3.  Semi-synthetic Mithramycin SA Derivatives with Improved Anti-Cancer Activity 
Chemical biology & drug design  2013;81(5):615-624.
doi:10.1111/cbdd.12107
PMCID: PMC3636171  PMID: 23331575
Mithramycin; Mithramycin SA; Semi-synthesis; Anti-cancer; Aureolic acid
4.  Investigating Mammalian Tyrosine Phosphatase Inhibitors as Potential ‘Piggyback’ Leads to Target Trypanosoma brucei Transmission 
Chemical biology & drug design  2013;81(2):291-301.
African trypanosomiasis is a neglected tropical disease affecting humans and animals across 36 sub-Saharan African countries. We have investigated the potential to exploit a ‘piggyback’ approach to inhibit Trypanosoma brucei transmission by targeting the key developmental regulator of transmission, T. brucei protein tyrosine phosphatase 1. This strategy took advantage of the extensive investment in inhibitors for human protein tyrosine phosphatase 1B, a key target for pharmaceutical companies for the treatment of obesity and diabetes. Structural predictions for human and trypanosome tyrosine phosphatases revealed the overall conservation of important functional motifs, validating the potential for exploiting cross specific compounds. Thereafter, nineteen inhibitors were evaluated; seventeen from a protein tyrosine phosphatase 1B-targeted inhibitor library and two from literature analysis – oleanolic acid and suramin, the latter of which is a front line drug against African trypanosomiasis. The compounds tested displayed similar inhibitory activities against the human and trypanosome enzymes, mostly behaving as noncompetitive inhibitors. However, their activity against T. brucei in culture was low, necessitating further chemical modification to improve their efficacy and specificity. Nonetheless, the results validate the potential to explore a ‘piggyback’ strategy targeting T. brucei protein tyrosine phosphatase 1 through exploiting the large pharmacological investment in therapies for obesity targeting protein tyrosine phosphatase 1B.
doi:10.1111/cbdd.12079
PMCID: PMC4003393  PMID: 23066974
biological screening; drug discovery; kinase; phosphatase
5.  Synthesis and biological evaluation of tricyclic guanidine analogues of batzelladine K for antimalarial, antileishmanial, antibacterial, antifungal and anti-HIV activities 
Chemical biology & drug design  2013;81(4):10.1111/cbdd.1427.
Fifty analogues of batzelladine K were synthesized and evaluated for in vitro antimalarial (Plasmodium falciparum), antileishmanial (Leishmania donovani), antimicrobial (panel of bacteria and fungi), antiviral (HIV-1) activities. Analogues 14h and 20l exhibited potential antimalarial activity against chloroquine-sensitive D6 strain with IC50 1.25 and 0.88 μM and chloroquine-resistant W2 strain with IC50 1.64 and 1.07 μM, respectively. Analogues 12c and 14c having nonyl substitution showed the most potent antileishmanial activity with IC50 2.39 and 2.78 μM and IC90 11.27 and 12.76 μM respectively. Three analogues 12c, 14c and 14i were the most active against various pathogenic bacteria and fungi with IC50 <3.02 μM and MIC/MBC/MFC <6 μM. Analogue 20l having pentyl and methyl substituents on tricycle showed promising activities against all pathogens. However, none was found active against HIV-1. Our study demonstrated that the tricyclic guanidine compounds provide new structral class for broad spectrum activity.
doi:10.1111/cbdd.1427
PMCID: PMC3488358  PMID: 23534411
Batzelladine; tricyclic guanidine; antimalarial; antileishmanial; antimicrobial; anti-HIV
6.  Highly predictive ligand-based pharmacophore and homology models of ABHD6 
Chemical biology & drug design  2012;81(3):382-388.
α/β-hydrolase domain-containing 6 (ABHD6) represents a potentially attractive therapeutic target for indirectly potentiating 2-arachidonoylglycerol signaling, however, the enzyme is currently largely uncharacterized. Here we describe a five element, ligand-based pharmacophore model along with a refined homology model of ABHD6. Following a virtual screen of a modest database, both the pharmacophore and homology models were found to be highly predictive, preferentially identifying ABHD6 inhibitors over druglike noninhibitors. The models yield insight into the features required for optimal ligand binding to ABHD6 and the atomic structure of the binding site. In combination the two models should be very helpful not only in high throughput virtual screening, but also in lead optimization, and will facilitate the development of novel, selective ABHD6 inhibitors as potential drugs.
doi:10.1111/cbdd.12086
PMCID: PMC3573238  PMID: 23110439
7.  Biomolecular interactions of small-molecule inhibitors affecting the YopH protein tyrosine phosphatase 
Chemical biology & drug design  2013;81(3):323-333.
We have developed competitive and direct binding methods to examine small-molecule inhibitors of protein tyrosine phosphatase (PTPase) activity. Focusing on the Yersinia outer protein H (YopH), a potent bacterial PTPase, we describe how an understanding of the kinetic interactions involving YopH, peptide substrates, and small-molecule inhibitors of PTPase activity can be beneficial for inhibitor screening and we further translate these results into a microarray assay for high-throughput screening.
doi:10.1111/cbdd.12097
PMCID: PMC3573263  PMID: 23241354
Affinity screening; competitive binding assay; high-throughput screening; protein tyrosine phosphatase; peptide microarray; phosphopeptide; surface plasmon resonance (SPR); Yersinia pestis outer protein H (YopH)
8.  Structure-Based Design and Synthesis of Benzothiazole Phosphonate Analogues with Inhibitors of Human ABAD-Aβ for Treatment of Alzheimer’s disease 
Chemical biology & drug design  2012;81(2):238-249.
Amyloid binding alcohol dehydrogenase (ABAD), a mitochondrial protein, is a cofactor facilitating amyloid-β peptide (Aβ) induced cell stress. Antagonizing Aβ-ABAD interaction protects against aberrant mitochondrial and neuronal function and improves learning memory in the Alzheimer’s disease (AD) mouse model. Therefore, it offers a potential target for Alzheimer’s drug design, by identifying potential inhibitors of Aβ-ABAD interaction. 2D QSAR methods were applied to novel compounds with known IC50 values, which formed a training set. A correlation analysis was carried out comparing the statistics of the measured IC50 with predicted values. These selectivity-determining descriptors were interpreted graphically in terms of principle component analyses, which are highly informative for the lead optimization process with respect to activity enhancement. A 3D pharmacophore model also was created. The 2D QSAR and 3D pharmacophore models will assist in hi-throughput screening. In addition, ADME descriptors were also determined to study their pharmacokinetic properties. Finally, ABAD molecular docking study of these novel molecules was undertaken to determine whether these compounds exhibit significant binding affinity with the binding site. We have synthesized only the compounds that have shown the best drug like properties as candidates for further studies.
doi:10.1111/cbdd.12068
PMCID: PMC3640359  PMID: 23039767
Molecular Docking; Quantitative Structure Activity Relationship; ADME Prediction; ABAD Inhibitors; Benzothiazole Aminophosphonates
9.  Optimization of CD4/gp120 Inhibitors by Thermodynamic Guided Alanine Scanning Mutagenesis 
Chemical biology & drug design  2013;81(1):72-78.
Since protein/protein interactions usually trigger signaling processes, inhibitors of those interactions must preclude protein binding without eliciting the signaling process themselves. To accomplish those goals, small molecules need to target those protein residues that contribute the most to binding (binding hotspots) without disturbing those residues that initiate signaling processes (allosteric hotspots). The availability of a blueprint identifying binding and allosteric hotspots will significantly aid inhibitor design and optimization. In this paper, we show that in some situations the blueprint can be constructed by combining the standard technique of alanine scanning mutagenesis with isothermal titration calorimetry (ITC). We demonstrate the approach by developing the combined binding and allosteric hotspots blueprint for CD4/gp120, the initial interaction leading to HIV-1 cell infection. A major finding of these studies is that not all binding hotspots are allosteric hotspots opening the possibility for the rational design of inhibitors and antagonist or agonist modulators.
doi:10.1111/cbdd.12075
PMCID: PMC3527665  PMID: 23066870
Binding Affinity; Enthalpy; Entropy; Thermodynamic Optimization; Isothermal Titration Calorimetry; Alanine Scanning Mutagenesis
10.  Back to the Future: Can physical models of passive membrane permeability help reduce drug candidate attrition and move us beyond QSPR? 
Chemical biology & drug design  2013;81(1):61-71.
It is widely recognized that ADMET (Adsorption, Distribution, Metabolism, Excretion - Toxicology) liabilities kill the majority of drug candidates that progress to clinical trials. The development of computational models to predict small molecule membrane permeability is therefore of considerable scientific and public health interest. Empirical qualitative structure permeability relationship (QSPR) models of permeability have been a mainstay in industrial applications, but lack a deep understanding of the underlying biological physics. Others and we have shown that implicit solvent models to predict passive permeability for small molecules exhibit mediocre predictive performance when validated across experimental test sets. Given the vast increase in computer power, more efficient parallelization schemes, and extension of current atomistic simulation codes to general use graphical processing units (GPUs), the development and application of physical models based on all-atom simulations may now be feasible. Preliminary results from rigorous free energy calculations using all-atom simulations indicate that performance relative to implicit solvent models may be improved, but many outstanding questions remain. Here we review the current state of the art physical models for passive membrane permeability prediction, and present a prospective look at promising new directions for all-atom approaches.
doi:10.1111/cbdd.12074
PMCID: PMC3527668  PMID: 23066853
Passive membrane permeability; Implicit solvent; Molecular Dynamics; Homogenous solubility model
11.  Expanding the Number of “Druggable” Targets: Non-Enzymes and Protein-Protein Interactions 
Chemical biology & drug design  2013;81(1):22-32.
Following sequencing and assembly of the human genome, the preferred methods for identification of new drug targets have changed dramatically. Modern tactics such as genome-wide association studies (GWAS) and deep sequencing are fundamentally different from the pharmacology-guided approaches used previously, in which knowledge of small molecule ligands acting at their cellular targets was the primary discovery engine. A consequence of the “target-first, pharmacology-second” strategy is that many predicted drug targets are non-enzymes, such as scaffolding, regulatory or structural proteins, and their activities are often dependent on protein-protein interactions (PPIs). These types of targets create unique challenges to drug discovery efforts because enzymatic turnover cannot be used as a convenient surrogate for compound potency. Moreover, it is often challenging to predict how ligand binding to non-enzymes might affect changes in protein function and/or pathobiology. Thus, in the post-genomic era, targets might be strongly implicated by molecular biology-based methods, yet they often later earn the designation of “undruggable.” Can the scope of available targets be widened to include these promising, but challenging, non-enzymes? In this review, we discuss advances in high throughput screening technology and chemical library design that are emerging to deal with these challenges.
doi:10.1111/cbdd.12066
PMCID: PMC3531880  PMID: 23253128
12.  Discovery of potent, novel Nrf2 inducers via quantum modeling, virtual screening and in vitro experimental validation 
Chemical biology & drug design  2012;80(6):810-820.
Nuclear factor erythroid 2-related factor 2 (Nrf2) is the master transcription factor of the antioxidant response element (ARE) pathway, coordinating the induction of detoxifying and antioxidant enzymes. Nrf2 is normally sequestered in the cytoplasm by Kelch-like ECH associating protein 1 (Keap1). To identify novel small molecules that will disturb Nrf2:Keap1 binding and promote activation of the Nrf2-ARE pathway, we generated a quantum model based on the structures of known Nrf2-ARE activators. We used the quantum model to perform in silico screening on over 18 million commercially available chemicals to identify the structures predicted to activate the Nrf2-ARE pathway based on the quantum model. The top hits were tested in vitro and half of the predicted hits activated the Nrf2-ARE pathway significantly in primary cell culture. In addition, we identified a new family of Nrf2-ARE activating structures that all have comparable activity to tBHQ and protect against oxidative stress and dopaminergic toxins in vitro. The improved ability to identify potent activators of Nrf2 through the combination of in silico and in vitro screening described here improves the speed and cost associated with screening Nrf2-ARE activating compounds for drug development.
doi:10.1111/cbdd.12040
PMCID: PMC3484224  PMID: 22925725
13.  Farnesyl diphosphate synthase inhibitors from in silico screening 
Chemical biology & drug design  2013;81(6):742-748.
The relaxed complex scheme is an in silico drug screening method that accounts for receptor flexibility by using molecular dynamics simulations. Here, we used this approach combined with similarity searches and experimental inhibition assays to identify several low micro-molar, non-bisphosphonate inhibitors, bisamidines, of farnesyl diphosphate synthase (FPPS), an enzyme targeted by some anti-cancer and anti-microbial agents and for the treatment of bone resorption diseases. This novel class of FPPS inhibitors have more drug-like properties than existing bisphosphonate inhibitors, making them interesting pharmaceutical leads.
doi:10.1111/cbdd.12121
PMCID: PMC3671582  PMID: 23421555
14.  Structural Basis of the Allosteric Inhibitor Interaction on the HIV-1 Reverse Transcriptase RNase H domain 
Chemical biology & drug design  2012;80(5):706-716.
HIV-1 reverse transcriptase (RT) has been an attractive target for the development of antiretroviral agents. Although this enzyme is bi-functional, having both DNA polymerase and ribonuclease H (RNH) activities, there is no clinically approved inhibitor of the RNH activity. Here, we characterize the structural basis and molecular interaction of an allosteric site inhibitor, BHMP07, with the wild type (WT) RNH fragment. Solution NMR experiments for inhibitor titration on WT RNH showed relatively wide chemical shift perturbations, suggesting a long-range conformational effect on the inhibitor interaction. Comparisons of the inhibitor-induced NMR chemical-shift changes of RNH with those of RNH dimer, in the presence and absence of Mg2+, were performed to determine and verify the interaction site. The NMR results, with assistance of molecular docking, indicate that BHMP07 preferentially binds to a site that is located between the RNH active site and the region encompassing helices B and D (the “substrate-handle region”). The interaction site is consistent with the previous proposed site, identified using a chimeric RNH (p15-EC) [Gong, el (2011) Chem. Biol. Drug Des. 77, 39-47], but with slight differences that reflect the characteristics of the amino acid sequences in p15-EC compared to the WT RNH.
doi:10.1111/cbdd.12010
PMCID: PMC3465473  PMID: 22846652
Reverse transcriptase; ribonuclease H; HIV; acylhydrazone; inhibitor; enzyme; protein; NMR; dimer; molecular docking
15.  Superior analgesic effect of H-Dmt-D-Arg-Phe-Lys-NH2 ([Dmt1]DALDA), a multifunctional opioid peptide, compared to morphine in a rat model of neuropathic pain 
Chemical biology & drug design  2012;80(5):771-774.
H-Dmt-D-Arg-Phe-Lys-NH2 ([Dmt1]DALDA), is a synthetic tetrapeptide with extraordinary selectivity for the mu-opioid receptor and is an extremely potent analgesic. [Dmt1]DALDA is unusual in the way that the greater part of its analgesic potency appears to be produced by its actions in the spinal cord. Furthermore, [Dmt1]DALDA inhibits norepinephrine re-uptake and is a mitochondria-targeted antioxidant. Such characteristics may make [Dmt1]DALDA particularly effective against neuropathic pain. The present study was designed to compare the effects of [Dmt1]DALDA and morphine on thermal hyperalgesia in an experimental neuropathic pain model. Neuropathic pain was induced in rats by surgical ligation of the L5 spinal nerve, and thermal pain thresholds were assessed by latencies of paw withdrawal to radiant heat. The increase in paw withdrawal latency was greater after the administration of [Dmt1]DALDA than that of morphine in neuropathic rats at doses that were equianalgesic in naïve animals. We conclude that [Dmt1]DALDA is more effective than morphine against thermal hyperalgesia in this experimental model of neuropathic pain.
doi:10.1111/cbdd.12003
PMCID: PMC3465489  PMID: 22834682
Opioid; neuropathic pain; hyperalgesia; [Dmt1]DALDA; morphine
16.  Modulation of Opioid Receptor Ligand Affinity and Efficacy Using Active and Inactive State Receptor Models 
Chemical biology & drug design  2012;80(5):763-770.
Mu opioid receptor (MOR) agonists are widely used for the treatment of pain; however chronic use results in the development of tolerance and dependence. It has been demonstrated that co-administration of a MOR agonist with a delta opioid receptor (DOR) antagonist maintains the analgesia associated with MOR agonists, but with reduced negative side effects. Using our newly refined opioid receptor models for structure-based ligand design, we have synthesized several pentapeptides with tailored affinity and efficacy profiles. In particular, we have obtained pentapeptides 8, Tyr-c(S-S)[DCys-1Nal-Nle-Cys]NH2, and 12, Tyr-c(S-S)[DCys-1Nal-Nle-Cys]OH, which demonstrates high affinity and full agonist behavior at MOR, high affinity but very low efficacy for DOR, and minimal affinity for the kappa opioid receptor (KOR). Functional properties of these peptides as MOR agonists/DOR antagonists lacking undesired KOR activity make them promising candidates for future in vivo studies of MOR/DOR interactions. Subtle structural variation of 12, by substituting D-Cys5 for L-Cys5, generated analog 13 which maintains low nanomolar MOR and DOR affinity, but which displays no efficacy at either receptor. These results demonstrate the power and utility of accurate receptor models for structure-based ligand design, as well as the profound sensitivity of ligand function on its structure.
doi:10.1111/cbdd.12014
PMCID: PMC3465523  PMID: 22882801
Delta opioid receptor; G protein-coupled receptors; mixed efficacy ligand; mu opioid receptor; opioid; peptide; structure-based design
17.  Neoflavonoids and tetrahydroquinolones as possible cancer chemopreventive agents 
Chemical biology & drug design  2012;80(4):616-624.
Several lactone and lactam based neoflavonoids and tetrahydroquinolones were synthesized and evaluated for cancer chemopreventive studies using cell and molecular target based in vitro bioassays, namely NFκB, aromatase, and quinone reductase 1 (QR1). These analogues blocked TNF-α-induced NFκB activation in a dose-dependent manner with IC50 values in the range of 0.11–3.2 μM. In addition, compound 8 inhibited aromatase activity with an IC50 value of 12.12 μM and compound 10 affected QR1 induction (IR: 3.6, CD: 19.57 μM). Neoflavonoids 8 and 10 exhibiting good results, can further be optimized for improved therapeutic profiles. However, investigations into the actions of neoflavonoids and tetrahydroquinolones, especially those related to the NFκB signaling pathway, aromatase inhibition, induction of QR1 expression and in vivo studies could provide new insights into the cancer chemopreventive ability of these molecules.
doi:10.1111/j.1747-0285.2012.01439.x
PMCID: PMC3448855  PMID: 22726671
Neoflavonoids; tetrahydroquinolone; cancer chemoprevention; NFκB; aromatase; quinone reductase 1
18.  Identification of FDA-approved drugs that computationally bind to MDM2 
Chemical biology & drug design  2012;80(4):631-637.
The integrity of the p53 tumor suppressor pathway is compromised in the majority of cancers. In 7% of cancers, p53 is inactivated by abnormally high levels of MDM2—an E3 ubiquitin ligase that polyubiquitinates p53, marking it for degradation. MDM2 engages p53 through its hydrophobic cleft and blockage of that cleft by small molecules can re-establish p53 activity. Small molecule MDM2 inhibitors have been developed, but there is likely to be a high cost and long time period before effective drugs reach the market. An alternative is to repurpose FDA-approved drugs. This report describes a new approach, called Computational Conformer Selection, to screen for compounds that potentially inhibit MDM2. This screen was used to computationally generate up to 600 conformers of 3,244 FDA-approved drugs. Drug conformer similarities to 41 computationally-generated conformers of MDM2 inhibitor nutlin 3a were ranked by shape and charge distribution. Quantification of similarities by Tanimoto combo scoring resulted in scores that ranged from 0.142 to 0.802. In silico docking of drugs to MDM2 was used to calculate binding energies and to visualize contacts between the top-ranking drugs and the MDM2 hydrophobic cleft. We present 15 FDA-approved drugs predicted to inhibit p53/MDM2 interaction.
doi:10.1111/j.1747-0285.2012.01428.x
PMCID: PMC3448857  PMID: 22703617
MDM2; cancer; nutlin; FDA; repurposing; repositioning; p53; drugs
19.  Enhancing and limiting endothelin-1 signaling with a cell penetrating peptide mimicking the third intracellular loop of the ETB receptor 
Chemical biology & drug design  2012;80(3):374-381.
TAT (a 13-mer derived from the HIV-1 Tat protein) linked cell-permeable peptides deliver plasma membrane impermeable cargos into the cell. We investigated the effect of a TAT linked intracellular third loop (IC3) of the endothelin-1 (ET-1) type B receptor on ET-1 activation of ERK. The effect of this peptide on ERK activation was determined in ETB receptor cDNA transfected CHO cells and in ETA and ETB expressing human pulmonary artery smooth muscle cells (hPASMC) obtained from a normal and a bone morphogenetic protein-2 receptor (BMPR2), exon 1–8 deletion subject, with pulmonary hypertension. In the CHO cells the peptide, at optimum 10uM concentration, suppressed ET-1 activation. In the normal hPASMC, the peptide marginally enhanced ET-1 activation of ERK. However, it markedly enhanced the ET-1 activation of ERK in the BMP2R hPASMC. While the effective concentration for ET-1 activation of ERK remained unchanged in the BMP2R hPASMC, the number of ETB receptors declined by 2/3. These data point to the IC3 peptide as having variable receptor interactive effects with both signal repressive and enhancing capabilities. Peptides that can alter ET-1 signal capabilities are potentially important in the study and treatment of pulmonary hypertension.
doi:10.1111/j.1747-0285.2012.01405.x
PMCID: PMC3496424  PMID: 22553998
drug delivery; G-protein coupled receptor; peptide; signal transduction and modulators (activation / inhibition); endothelin; endothelin receptors
20.  Comparison of Dynamics of Extracellular Accesses to the β1 and β2 Adrenoceptors Binding Sites Uncovers the Potential of Kinetic Basis of Antagonist Selectivity 
Chemical biology & drug design  2012;80(2):215-226.
From the molecular mechanism of antagonist unbinding in the β1 and β2 adrenoceptors investigated by steered molecular dynamics, we attempt to provide further possibilities of ligand subtype and subspecies selectivity. We have simulated unbinding of β1-selective Esmolol and β2-selective ICI-118551 from both receptors to the extracellular environment and found distinct molecular features of unbinding. By calculating work profiles, we show different preference in antagonist unbinding pathways between the receptors, in particular, perpendicular to the membrane pathway is favourable in the β1 adrenoceptor, whereas the lateral pathway involving helices 5, 6 and 7 is preferable in the β2 adrenoceptor. The estimated free energy change of unbinding based on the preferable pathway correlates with the experimental ligand selectivity. We then show that the non-conserved K347 (6.58) appears to facilitate in guiding Esmolol to the extracellular surface via hydrogen bonds in the β1 adrenoceptor. In contrast, hydrophobic and aromatic interactions dominate in driving ICI-118551 through the easiest pathway in the β2 adrenoceptor. We show how our study can stimulate design of selective antagonists and discuss other possible molecular reasons of ligand selectivity, involving sequential binding of agonists and glycosylation of the receptor extracellular surface.
doi:10.1111/j.1747-0285.2012.01390.x
PMCID: PMC3399986  PMID: 22530954
adrenergic receptors; drug design; G protein-coupled receptors; molecular dynamics; selectivity
21.  A strategy for selective O6-alkylguanine-DNA alkyltransferase depletion under hypoxic conditions 
Chemical biology & drug design  2012;80(2):279-290.
Cellular resistance to chemotherapeutics that alkylate the O-6 position of guanine residues in DNA correlates with their O6-alkylguanine-DNA alkyltransferase (AGT) activity. In normal cells high [AGT] is benefical, sparing the host from toxicity, whereas in tumor cells high [AGT] prevents chemotherapeutic response. Therefore, it is necessary to selectively inactivate AGT in tumors. The oxygen deficient compartment unique to solid tumors is conducive to reduction, and could be utilized to provide this selectivity. Therefore, we synthesized 2-nitro-6-benzyloxypurine (2-NBP), an analog of O6-benzylguanine (O6-BG) in which the essential 2-amino group is replaced by a nitro moiety, 2-NBP is >2000-fold weaker than O6-BG as an AGT inhibitor. We demonstrate oxygen concentration sensitive net reduction of 2-NBP by cytochrome P450 reductase, xanthine oxidase and EMT6, DU145 and HL-60 cells to yield O6-BG. We show that 2-NBP treatment depletes AGT in intact cells under oxygen deficient conditions and selectively sensitizes cells to laromustine (an agent that chloroethylates the O-6 position of guanine) under oxygen deficient but not normoxic conditions. 2-NBP represents a proof of concept lead compound, however, its facile reduction (E1/2 – 177 mV vs. Ag/AgCl) may result in excessive oxidative stress and/or the generation of AGT inhibitors in normoxic regions in vivo.
doi:10.1111/j.1747-0285.2012.01401.x
PMCID: PMC3399964  PMID: 22553921
2-Nitro-6-benzyloxypurine; O6-benzylguanine; prodrug; AGT; hypoxia; targeting; sensitization; chemotherapy
22.  MCR Synthesis of Praziquantel Derivatives 
Chemical biology & drug design  2012;79(4):470-477.
Schistosomiasis, a high volume neglected tropical disease affecting more than 200 million people worldwide, can only be effectively treated by the tetrahydroisoquinoline drug praziquantel (PZQ). Herein, we describe an efficient approach to access PZQ derivatives by the Ugi 4-component reaction followed by the Pictet-Spengler reaction in a two-step, one-pot procedure. 30 Novel PZQ derivatives are described based on the Ugi 4-component reaction and an X-ray structure of a novel derivative revealing different conformation compared with PZQ is discussed. Several analogues comparable in activity to the drug PZQ have been identified based on an in vitro Schistosoma mansoni worm viability assay.
doi:10.1111/j.1747-0285.2011.01288.x
PMCID: PMC3712825  PMID: 22151001
multicomponent reaction; Ugi; Pictet-Spengler; isocyanide; schistosomiasis; neglected tropical disease
23.  Structure-activity relationship study of collagen derived anti-angiogenic biomimetic peptides 
Chemical biology & drug design  2012;80(1):27-37.
Structure-activity relationship (SAR) studies are essential in the generation of peptides with enhanced activity and efficacy as therapeutic agents. In this study we report a SAR study for a family of mimetic peptides derived from type IV collagen with potent anti-angiogenic properties. The SAR study was conducted using a number of validated in vitro assays including cell proliferation, adhesion, migration and tubule formation. We report a critical sequence (NINNV) within this peptide series which is required for the potent anti-angiogenic activity. Detailed amino acid substitutions resulted in peptides with superior efficacy. Specifically, substitutions with Isoleucine at positions twelve and eighteen along with the substitution of the Methionine at position ten with the non-natural amino acid d-Alanine led to an increase in potency by two orders of magnitude over the parent peptide. Several mimetic peptides in this series exhibit a significant improvement of activity over the parent peptide. This improved in vitro activity is expected to correlate with an increase in in vivo activity leading to effective peptides for anti-angiogenic therapy for different disease applications including cancer and age-related macular degeneration.
doi:10.1111/j.1747-0285.2012.01376.x
PMCID: PMC3371172  PMID: 22405100
angiogenesis; peptidomimetics; endothelial cell; cancer; age-related macular degeneration; lymphangiogenesis
24.  Accounting for receptor flexibility and enhanced sampling methods in computer aided drug design 
Chemical biology & drug design  2013;81(1):10.1111/cbdd.12051.
Protein flexibility plays a major role in biomolecular recognition. In many cases it is not obvious how molecular structure will change upon association with other molecules. In proteins these changes can be major, with large deviations in overall backbone structure, or they can be more subtle as in a side chain rotation. Either way the algorithms that predict the favorability of biomolecular association require relatively accurate predictions of the bound structure to give an accurate assessment of the energy involved in association. Here we review a number of techniques that have been proposed to accommodate receptor flexibility in the simulation of small molecules binding to protein receptors. We investigate modifications to standard rigid receptor docking algorithms, and also explore enhanced sampling techniques, and the combination of free energy calculations and enhanced sampling techniques. The understanding and allowance for receptor flexibility are helping to make computer simulations of ligand protein binding more accurate. These developments may help improve the efficiency of drug discovery and development. Efficiency will be essential as we begin to see personalized medicine tailored to individual patients, which means specific drugs are needed for each patient’s genetic makeup.
doi:10.1111/cbdd.12051
PMCID: PMC3540989  PMID: 23253130
Computer aided drug design; structure based drug design; receptor flexibility; ensemble docking; relaxed complex scheme; molecular dynamics; accelerated molecular dynamics; free energy calculation
25.  Inhibitors of Tubulin Assembly Identified through Screening a Compound Library 
Chemical biology & drug design  2008;72(6):513-524.
Tubulin is the proposed target for drugs against cancer and helminths and is also a validated target in kinetoplastid parasites. With the aim of identifying new lead compounds against Leishmania sp., tubulin isolated from L. tarentolae was used to screen a 10 000 compound library. One compound, Chembridge No. 7992831 (5), displayed an IC50 of 13 μm against Leishmania tubulin in an in vitro assembly assay and showed a greater than threefold selectivity over mammalian tubulin. Another compound, Chembridge No. 9067250 (8), exhibited good activity against mammalian tubulin (IC50 = 5.0 μm). This compound was also toxic to several cancer cell lines with IC50 values in the region of 1 μM. Subsequent testing of analogues of 8 contained within the library identified two compounds with greater potency against mammalian tubulin (IC50 values of 1.1 and 2.8 μM). The more potent antitubulin agent also showed promising activity against cancer cell lines in vitro, with IC50 values ranging from 0.18 to 0.73 μM.
doi:10.1111/j.1747-0285.2008.00729.x
PMCID: PMC3677961  PMID: 19090918
Tubulin; screen; library; Leishmania

Results 1-25 (145)