Computer-aided Rational Design Research Articles

Nano-objects - sculpting and shape in molecular material design (The Pierre Gilles de Gennes ILCS prize lecture)

ABSTRACTEarly liquid crystal compounds often had relatively simple molecular structures, but with the invention of electro-optical devices, the design of materials became increasingly important. Property-structure correlations have promoted computer-aided rational design to the point where it is becoming a necessity in the selection of compounds for preparation. For simple molecular architectures, allowable electrostatic interactions in the face of steric repulsion were used in a phenomenological way to generate new mesophases, such as the hexatic phase, and to engineer materials with desirable physical properties. Materials chemists, in the creation of substances of greater complexity, have built upon these forms of research. Interestingly, the larger the system often the more simply it becomes as the molecular bulk and shape begin to dominate over the surface interactions. Nano-objects of unique shapes, such as ‘Janus’ grains, created in the process of design can kindle the formation of new mesophases, such as the twist-bend and splay phases generated through the crowding of molecular particles, and minimisation of the free volume. This article explores the development of the shapes of molecular grains and the mesophases they spawn.

Enhancing thermostability and removing hemin inhibition of Rhodopseudomonas palustris 5-aminolevulinic acid synthase by computer-aided rational design.

To enhance the thermostability and deregulate the hemin inhibition of 5-aminolevulinic acid (ALA) synthase from Rhodopseudomonas palustris (RP-ALAS) by a computer-aided rational design strategy. Eighteen RP-ALAS single variants were rationally designed and screened by measuring their residual activities upon heating. Among them, H29R and H15K exhibited a 2.3°C and 6.0°C higher melting temperature than wild-type, respectively. A 6.7-fold and 10.3-fold increase in specific activity after 1h incubation at 37°C was obtained for H29R (2.0U/mg) and H15K (3.1U/mg) compared to wild-type (0.3U/mg). Additionally, higher residual activities in the presence of hemin were obtained for H29R and H15K (e.g., 64% and 76% at 10μM hemin vs. 27% for wild-type). The ALA titer was increased by 6% and 22% in fermentation using Corynebacterium glutamicum ATCC 13032 expressing H29R and H15K, respectively. H29R and H15K showed high thermostability, reduced hemin inhibition and slightly high activity, indicating that these two variants are good candidates for bioproduction of ALA.

Tri-armed ligands of G-quadruplex on heteroarene-fused anthraquinone scaffolds: Design, synthesis and pre-screening of biological properties

Here, a combined molecular modelling methodology was used to identify the binding mode of 4,11-bis((2-guanidinoethyl)amino)anthra[2,3-b]thiophene-5,10-dione (1), a previously reported G4 ligand. After calculating the optimal interaction parameters 1 with the target, two series of tri-armed ligands based on furan- or thiophene-fused anthraquinone scaffolds were designed and synthesized. The new compounds bearing an additional side chain at the 2-position of the heterocycle and the 4,11-side chains with different spacer lengths and structures of terminal groups demonstrated much stronger affinity for telomeric G4 (4–15 times) versus the parental ligand. Moreover, the specificity to the quadruplex over duplex DNA was significantly improved (up to 75 times) when the 3-guanidinopropyl side chain was introduced at the 2-position of the heterocycle ring. All tri-armed ligands demonstrated modest antiproliferative potency, which is likely due to low intracellular penetration. Nevertheless, this work shows how computer-aided rational design of new potent compounds can be used for targeted anticancer therapy.

Identifying the Rate-Limiting Elementary Steps of Nitrogen Fixation with Single-Site Fe Model Complexes

Biomimetic nitrogen fixation provides an attractive alternative for the century-old Haber-Bosch process; however, the performance of the currently available molecular biomimetic catalysts is very limited. In this work, we are aiming to understand the catalytic cycle of one of the most promising biomimetic complex families that can be the cornerstone of future computer-aided rational design of biomimetic complexes. We calculate the Gibbs free energy of all elementary reaction steps of homogeneous dinitrogen reduction to NH3 on single-site iron complexes with EPPP tetradentate ligands (E = B, Si). We examine all possible mechanisms and identify the dominant pathways and the critical elementary steps that can be rate-determining in the catalytic cycle of nitrogen fixation. We find that the catalytic mechanism depends on the applied ligand and that the distal pathway observed with E = B is the most favorable route regarding the catalytic performance. Our calculations also reveal the lack of thermodynamic driving force in the last steps of the catalytic cycle that can be responsible for the low catalytic activity of the studied biomimetic catalysts. Our results can serve as a starting point for the rational design of biomimetic complexes, which should focus on establishing a steadily decreasing Gibbs free energy profile, as suggested by the Sabatier principle.

Multifunctional Effects of a Small-Molecule STAT3 Inhibitor on NASH and Hepatocellular Carcinoma in Mice.

Purpose: The incidence of hepatocellular carcinoma is increasing in the United States, and liver cancer is the second leading cause of cancer-related mortality worldwide. Nonalcoholic steatohepatitis (NASH) is becoming an important risk for hepatocellular carcinoma, and most patients with hepatocellular carcinoma have underlying liver cirrhosis and compromised liver function, which limit treatment options. Thus, novel therapeutic strategies to prevent or treat hepatocellular carcinoma in the context of NASH and cirrhosis are urgently needed.Experimental Design: Constitutive activation of STAT3 is frequently detected in hepatocellular carcinoma tumors. STAT3 signaling plays a pivotal role in hepatocellular carcinoma survival, growth, angiogenesis, and metastasis. We identified C188-9, a novel small-molecule STAT3 inhibitor using computer-aided rational drug design. In this study, we evaluated the therapeutic potential of C188-9 for hepatocellular carcinoma treatment and prevention.Results: C188-9 showed antitumor activity in vitro in three hepatocellular carcinoma cell lines. In mice with hepatocyte-specific deletion of Pten (HepPten- mice), C188-9 treatment blocked hepatocellular carcinoma tumor growth, reduced tumor development, and reduced liver steatosis, inflammation, and bile ductular reactions, resulting in improvement of the pathological lesions of NASH. Remarkably, C188-9 also greatly reduced liver injury in these mice as measured by serum aspartate aminotransferase and alanine transaminase levels. Analysis of gene expression showed that C188-9 treatment of HepPten- mice resulted in inhibition of signaling pathways downstream of STAT3, STAT1, TREM-1, and Toll-like receptors. In contrast, C188-9 treatment increased liver specification and differentiation gene pathways.Conclusions: Our results suggest that C188-9 should be evaluated further for the treatment and/or prevention of hepatocellular carcinoma. Clin Cancer Res; 23(18); 5537-46. ©2017 AACR.

Computer-aided rational design of Fe(iii)-catalysts for the selective formation of cyclic carbonates from CO2 and internal epoxides

The mechanism of Fe-catalyzed cyclic carbonate formation of CO2 and internal epoxides was studied to identify design strategies for more efficient catalysts.

Computer-aided rational design of novel EBF analogues with an aromatic ring.

Odorant binding proteins (OBPs) are important in insect olfactory recognition. These proteins bind specifically to insect semiochemicals and induce their seeking, mating, and alarm behaviors. Molecular docking and molecular dynamics simulations were performed to provide computational insight into the interaction mode between AgamOBP7 and novel (E)-β-farnesene (EBF) analogues with an aromatic ring. The ligand-binding cavity in OBP7 was found to be mostly hydrophobic due to the presence of several nonpolar residues. The interactions between the EBF analogues and the hydrophobic residues in the binding cavity increased in strength as the distance between them decreased. The EBF analogues with an N-methyl formamide or ester linkage had higher docking scores than those with an amide linkage. Moreover, delocalized π-π and electrostatic interactions were found to contribute significantly to the binding between the ligand benzene ring and nearby protein residues. To design new compounds with higher activity, four EBF analogues D1-D4 with a benzene ring were synthesized and evaluated based on their docking scores and binding affinities. D2, which had an N-methyl formamide group linkage, exhibited stronger binding than D1, which had an amide linkage. D4 exhibited particularly strong binding due to multiple hydrophobic interactions with the protein. This study provides crucial foundations for designing novel EBF analogues based on the OBP structure. Graphical abstract The design strategy of new EBF analogues based on the OBP7 structure.

Enhanced Thermostability of Lipoxygenase from Anabaena sp. PCC 7120 by Site-Directed Mutagenesis Based on Computer-Aided Rational Design.

Lipoxygenase from Anabaena sp. PCC 7120 (Ana-LOX) was thermally unstable. So, improving the thermostability of the enzyme was quite essential. The target site of Ana-LOX selected for site-directed mutagenesis was based on computer-aided rational design. The thermostability and specific activity of Ana-LOX were improved with replacing valine with alanine at the target site 421 and the site 40. Compared to the wild-type enzyme which has a half-life (T 1/2) of inactivation of 3.8min at 50°C, the T 1/2 of mutant enzymes with V421A and V40A substitution increased to 4.4 and 7.0min, respectively. The double mutant V421A/V40A showed a synergistic effect with a T 1/2 value of 8.3min, resulting in a 1.18-fold improvement compared to the original Ana-LOX. V421A, V40A, and V421A/V40A also obtained 4.83, 41.58, and 80.07% increase in specific activity, respectively. This study provides useful theoretical reference for enzyme molecular modification and computer-aided rational design.

LLL12, a novel small inhibitor targeting STAT3 for hepatocellular carcinoma therapy.

The constitutive activation of signal transducer and activator of transcription 3 (STAT3) is frequently detected in clinical incidences of hepatocellular carcinoma (HCC) but not in normal human hepatocytes. STAT3 signaling plays pivotal roles in angiogenesis, survival, metastasis, and growth of HCC. Recent evidence suggests that the blockade of aberrant STAT3 pathways can be exploited as a therapeutic strategy for HCC. We have developed the novel small molecular STAT3 inhibitor LLL12 on the basis of curcumin structure using computer-aided rational design. LLL12 has shown antitumor activity in various solid tumors including breast, brain, pancreatic cancer, and glioblastoma in vitro and in vivo. In this study, we hypothesized LLL12 inhibits STAT3 phosphorylation at tyrosine 705 (Y705) in HCC and show antitumor activity in HCC in vitro and in vivo. Our results show that LLL12 selectively inhibited HCC cell proliferation and induced apoptosis in SNU387, SNU398, SNU449, and Hep3B HCC cells in vitro. Furthermore, LLL12 at 5 mg/kg/day significantly inhibited the growth of SNU398 xenografts in nude mice. Collectively, our results indicate that LLL12 could be used to target STAT3 for the effective prevention or treatment of HCC.

QM/MM Calculations Reveal the Different Nature of the Interaction of Two Carborane-Based Sulfamide Inhibitors of Human Carbonic Anhydrase II

The crystal structures of two novel carborane-sulfamide inhibitors in the complex with human carbonic anhydrase II (hCAII) have been studied using QM/MM calculations. Even though both complexes possess the strongly interacting sulfamide···zinc ion motif, the calculations have revealed the different nature of binding of the carborane parts of the inhibitors. The neutral closo-carborane cage was bound to hCAII mainly via dispersion interactions and formed only very weak dihydrogen bonds. On the contrary, the monoanionic nido cage interacted with the protein mainly via electrostatic interactions. It formed short and strong dihydrogen bonds (stabilization of up to 4.2 kcal/mol; H···H distances of 1.7 Å) with the polar hydrogen of protein NH2 groups. This type of binding is unique among all of the classical organic and inorganic inhibitors of hCAII. Virtual glycine scanning allowed us to identify the amino-acid side chains, which made important contributions to ligand-binding energies. In summary, using QM/MM calculations, we have provided a detailed understanding of the differences between the interactions of two carborane sulfamides, identified the amino acids of hCAII with which they interact, and thus paved the way for the computer-aided rational design of selective boron-cluster-containing hCAII inhibitors.

Quantum Mechanical Scoring: Structural and Energetic Insights into Cyclin-Dependent Kinase 2 Inhibition by Pyrazolo[1,5-a]pyrimidines

A quantum mechanics (QM)-based scoring function has been applied to complexes of cyclin-dependent kinase 2 (CDK2) and thirty-one pyrazolo[1,5-a]pyrimidine-based inhibitors and their bioisosteres. A hybrid three-layer QM/MM setup (DFT-D/PM6-D3H4X/AMBER in generalized Born solvent) was used here for the first time as an extension of our previous full QM and SQM/MM (SQM means semiempirical QM) approaches. Two approaches to obtain the structures of the CDK2/inhibitor complexes were examined: i) building the modifications from one X-ray structure available coupled with a conformational search and ii) docking the compounds into CDK2. The QM-based scoring entailed a QM/SQM/MM optimization followed by calculations of the binding scores which were subsequently correlated with the experimental binding free energies. The correlation for the building protocol was good (r(2) = 0.64, predictive index = 0.81), whereas the docking approach failed. A decomposition of the interaction energies to ligand fragments enabled us to rationalize the differences in the binding affinities. In conclusion, we have developed and refined a QM-based scoring protocol and successfully applied it to reproduce the binding affinities in congeneric series of CDK2 inhibitors and to rationalize their potency. We thus propose that such a tool can be used in computer-aided rational drug design.

Rational Design of Cyclic Peptide Modulators of the Transcriptional Coactivator CBP: A New Class of p53 Inhibitors

The CREB binding protein (CBP) is a human transcriptional coactivator consisting of several conserved functional modules, which interacts with distinct transcription factors including nuclear receptors, CREB, and STAT proteins. Despite the importance of CBP in transcriptional regulation, many questions regarding the role of its particular domains in CBP functions remain unanswered. Therefore, developing small molecules capable of selectively modulating a single domain of CBP is of invaluable aid at unraveling its prominent activities. Here we report the design, synthesis, and biological evaluation of conformationally restricted peptides as novel modulators for the acetyl-lysine binding bromodomain (BRD) of CBP. Utilizing a target structure-guided and computer-aided rational design approach, we developed a series of cyclic peptides with affinity for CBP BRD significantly greater than those of its biological ligands, including lysine-acetylated histones and tumor suppressor p53. The best cyclopeptide of the series exhibited a K(d) of 8.0 μM, representing a 24-fold improvement in affinity over that of the linear lysine 382-acetylated p53 peptide. This lead peptide is highly selective for CBP BRD over BRDs from other transcriptional proteins. Cell-based functional assays carried out in colorectal carcinoma HCT116 cells further demonstrated the efficacy of this compound to modulate p53 stability and function in response to DNA damage. Our results strongly argue that these CBP modulators can effectively inhibit p53 transcriptional activity by blocking p53K382ac binding to CBP BRD and promoting p53 instability by changes of its post-translational modification states, a different mechanism than that of the p53 inhibitors reported to date.

Ligand-Based Computer-Aided Discovery of Tyrosinase Inhibitors. Applications of the TOMOCOMD-CARDD Method to the Elucidation of New Compounds

In this review an overview of the application of computational approaches is given. Specifically, the uses of Quantitative Structure-Activity Relationship (QSAR) methods for in silico identification of new families of compounds as novel tyrosinase inhibitors are revised. Assembling, validation of models through prediction series, and virtual screening of external data sets are also shown, to prove the accuracy of the QSAR models obtained with the TOMOCOMD-CARDD (TOpological MOlecular COMputational Design-Computer-Aided Rational Drug Design) software and Linear Discriminant Analysis (LDA) as statistical technique. Together with this, a database is collected for these QSAR studies, and could be considered a useful tool in future QSAR modeling of tyrosinase activity and for scientists that work in the field of this enzyme and its inhibitors. Finally, a translation to real world applications is shown by the use of QSAR models in the identification and posterior in-vitro evaluation of different families of compounds. Several different classes of compounds from various sources (natural and synthetic) were identified. Between them, we can find tetraketones, cycloartanes, ethylsteroids, lignans, dicoumarins and vanilloid derivatives. Finally, some considerations are discussed in order to improve the identification of novel drug-like compounds based on the use of QSAR-Ligand-Based Virtual Screening (LBVS).

Live attenuated influenza virus vaccines by computer-aided rational design

Influenza claims 250,000 - 500,000 lives annually worldwide. Despite existing vaccines and enormous efforts in biomedical research, these staggering numbers have not changed significantly over the last two decades1, motivating the search for new, more effective, vaccines that can be rapidly designed and easily produced. Using influenza virus strain A/PR/8/34, we describe a systematic, rational approach, termed Synthetic Attenuated Virus Engineering (SAVE), to develop new, efficacious live attenuated influenza virus vaccine candidates through genome-scale changes in codon pair bias. Attenuation is based on many hundreds of nucleotide changes across the viral genome, offering high genetic stability and a wide margin of safety. The method can be applied rapidly to any emerging influenza virus in its entirety, an advantage that is significant for dealing with seasonal epidemics and pandemic threats, such as H5N1- or 2009-H1N1 influenza.

Computer-aided design and synthesis of tetra-aryl-substituted alkenes and their bioevaluation as a selective modulator of estrogen-related receptor γ

This study reports on a translational exercise in computer-aided rational drug design, chemical synthesis, and bioevaluation using a cell-based reporter gene assay, pursued exclusively for the development of specific estrogen-related receptor γ (ERR γ) inverse agonists with selectivity over estrogen receptor α (ER α). We designed and synthesized a 9-membered small-molecule collection, which has, as the key molecular framework, tetra-aryl-substituted alkene derived from 4-hydroxytamoxifen (4-OHT), a known ERR γ inverse agonist and antagonist of ER α. Although we could not achieve a more potent inverse agonist than GSK5182 from our compound collection, we demonstrated a reasonable correlation between the in silico docking simulation and biological data of transcriptional regulation on nuclear receptors. Therefore, we suggest that structural information regarding proteins-of-interest provides a novel insight into the rational design of new therapeutic agents and that the utilization of docking simulation as a preliminary filtering tool might be a useful option for medicinal chemists or chemical biologists.

Abstract B95: A novel small molecule, LLL12 inhibits STAT3 phosphorylation and activities and exhibits potent growth suppressive activity in human hepatocellular carcinoma cells

Abstract Hepatocellular carcinoma is a major worldwide public health problem, and the third most frequent cause of cancer deaths after lung and stomach cancers. Hepatocellular carcinoma is a highly malignant tumor type with average survival rates that are currently less than 1 year following diagnosis. The etiology of hepatocellular carcinoma is still not well understood but involves a multi-step process of signaling protein dysregulation, which includes STAT3. The constitutive activation of STAT3 is frequently detected in clinical incidences of hepatocellular carcinoma and in more than 50% of human hepatocellular carcinoma cell lines but not in normal human hepatocytes. Constitutive STAT3 signaling contributes to hepatocellular carcinoma progression by promoting angiogenesis, survival, metastasis, and growth of hepatocellular carcinoma cells. Recent evidence suggests that the blockade of aberrant STAT3 signal pathway can be exploited as a therapeutic strategy for hepatocellular carcinoma. We have developed a novel small molecular STAT3 inhibitor LLL12 based on computer-aided rational design, which inhibits STAT3 activity by binding to its Src homology 2 (SH2) domain. LLL12 can inhibit STAT3 tyrosine 705 phosphroylation, STAT3 DNA binding activitivies. and prevent STAT translocation into the nucleus, thus decrease the transcription of target genes. And LLL12 also can inhibit IL-6 induced STAT3 phorsphorylation in Hep3B hepatocellular carcinoma cell. Our previous data show it can inhibit breast cancer (MDA-MB231 cells), brain tumor (U87 cells), and pancreatic cancer (PANC-1 cells) in vitro and in vivo. In this study, we report that LLL12 can selectively inhibit proliferation and induce apoptosis in SNU398, SNU387, Hep3B, and SNU449 hepatocellular carcinoma cells in vitro. Furthermore, the LLL12 can significantly inhibit tumor growth of SNU398 hepatocellular carcinoma cell line xenograft in nude mice at the dose of 5mg/kg. Collectively, our results show LLL12 could be used as a targeted drug candidate to target STAT3 for chemoprevention or treatment of Hepatocellular carcinoma in human. Citation Information: Cancer Prev Res 2010;3(1 Suppl):B95.

Recent development of two chitinase inhibitors, Argifin and Argadin, produced by soil microorganisms

Chitin, the second most abundant polysaccharide in nature, occurs in fungi, some algae and many invertebrates, including insects. Thus, chitin synthesis and degradation could represent specific targets for fungicides and insecticides. Chitinases hydrolyze chitin into oligomers of N-acetyl-d-glucosamine at key points in the life cycles of organisms, consequently, chitinase inhibitors have become subject of increasing interest. This review covers the development of two chitinase inhibitors of natural origin, Argifin and Argadin, isolated from the cultured broth of microorganisms in our laboratory. In particular, the practical total synthesis of these natural products, the synthesis of lead compounds via computer-aided rational molecular design, and discovery methods that generate only highly-active compounds using a kinetic target(chitinase)-guided synthesis approach (termed in situ click chemistry) are described.

Computer-aided rational molecular design of argifin-derivatives with increased inhibitory activity against chitinase B from Serratia marcescens

Argifin, a novel pentapeptide chitinase inhibitor isolated from Gliocladium fungal culture, is a promising candidate for the development of new fungicides, insecticides, and anti-asthma medications. In this study, we undertook rational molecular design of argifin-derivatives and tested them against chitinase B from Serratia marcescens ( SmChiB). The work involved molecular dynamics simulation with explicit water molecules, the molecular docking calculation, and free-energy analysis using the molecular mechanics Poisson–Boltzmann surface area method. The custom-designed derivatives were synthesized via effective solid phase synthesis, developed recently in our laboratory, and their inhibitory activities were measured against SmChiB. Finally, we identified and obtained a derivative which exhibited 28-fold more inhibition than argifin itself, a compound in which the d-Ala(5) of argifin was replaced with d-Leu and the 4-benzylpiperdine was attached to l-Asp(4).

Computer-aided rational design of the phosphotransferase system for enhanced glucose uptake in Escherichia coli

The phosphotransferase system (PTS) is the sugar transportation machinery that is widely distributed in prokaryotes and is critical for enhanced production of useful metabolites. To increase the glucose uptake rate, we propose a rational strategy for designing the molecular architecture of the Escherichia coli glucose PTS by using a computer-aided design (CAD) system and verified the simulated results with biological experiments. CAD supports construction of a biochemical map, mathematical modeling, simulation, and system analysis. Assuming that the PTS aims at controlling the glucose uptake rate, the PTS was decomposed into hierarchical modules, functional and flux modules, and the effect of changes in gene expression on the glucose uptake rate was simulated to make a rational strategy of how the gene regulatory network is engineered. Such design and analysis predicted that the mlc knockout mutant with ptsI gene overexpression would greatly increase the specific glucose uptake rate. By using biological experiments, we validated the prediction and the presented strategy, thereby enhancing the specific glucose uptake rate.

Novel Pharmacological Targets for Controlling Dendrite Branching and Growth During Neuronal Development

Processing of information by the central nervous system (CNS) depends on the dendritic morphology of postsynaptic neurons. The patterning of dendrites is determined by extrinsic and intrinsic factors that promote the activation of cellular signaling pathways. These factors and signaling cascades may lead to the transcriptional activation of regulators of neuronal morphology. Interestingly, when there is an abnormal decrease in the number of dendrite branches and disruption of proper networks, neurodegenerative diseases, including Rett Syndrome, autism, and mental retardation, may result. In this review, we evaluate the potential of regulators of dendrite patterning as targets for drug design for the treatment of neurodegenerative diseases and altered neuronal growth in the CNS. Particular attention is directed towards a specific regulator of dendrite branching reported by our group, cypin (cytosolic PSD-95 interactor). We discuss this novel intrinsic regulator of dendrite branching as an innovative pharmacological target for the use of computer-aided rational drug design to control guanine levels, microtubule assembly, and neuronal differentiation during CNS development and in disease states. Keywords: Dendrite branching, dendrite development, signaling pathways, transcriptional regulation, neurotrophins, cypin, guanine deaminase, neurological disorders