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A cyclic peptide targeted to the immune checkpoint protein CTLA4 is designed using a hierarchy of computational methods, demonstrated to bind by a physical measurement, and shown to inhibit tumor growth in cell culture and in mice.

A high-dimensional neural network interatomic potential was developed and used in molecular dynamics simulations of condensed phase Ni and Ni systems with liquid–solid phase coexistence. The reference data set was generated by sampling the potential energy surface over a broad temperature-pressure domain using ab initio MD simulations to train a unified potential. Excellent agreement was achieved between bulk face-centred cubic nickel thermal expansion simulations and relevant experimental data. The same potential also yields accurate structures and diffusivities in the liquid state. The phase transition between liquid and solid phases was simulated using the two-phase interface method. The predicted melting point temperature is within a few kelvins of the literature value. The general methodology could be applied to describe crystals with much more complex phase behaviours.

Ordered nanoscale patterns have been observed by atomic force microscopy at graphene-water and graphite-water interfaces. The two dominant explanations for these patterns are that (i) they consist of self-assembled organic contaminants or (ii) they are dense layers formed from atmospheric gases (especially nitrogen). Here we apply molecular dynamics simulations to study the behavior of dinitrogen and possible organic contaminants at the graphene-water interface. Despite the high concentration of N

Molecular dynamics simulations show that thiol-containing capping agents for silver nanoparticles, such as mercaptohexanol and lipoic acid, spontaneously assemble into dense aggregates at the silver–water interface.

This data set for the manuscript entitled "Computational Design of a Cyclic Peptide that Inhibits the CTLA4 Immune Checkpoint Pathway" includes all files needed to run and analyze the simulations of a designed cyclic peptide (Peptide 16) bound to CTLA4 in the putative most stable binding configuration, which is detailed in Figure 6 of the paper. These files include molecular model structure files (NAMD psf), force field parameter files (in CHARMM format), initial atomic coordinates (pdb format), NAMD configuration files, NAMD output including restart files (in binary NAMD format) and trajectories in dcd format (downsampled to 10 ns per frame). Analysis is controlled by shell scripts (Bash-compatible) that call VMD Tcl scripts. These scripts and their output are also included. Version: 1.0 Conventions Used in These Files =============================== Structure Files ---------------- - ctla4_P16_wat.psf (original NAMD (XPLOR?) format psf file including atom details (type, charge, mass), as well as definitions of bonds, angles, dihedrals, and impropers for each dipeptide.) - ctla4_P16.pdb (initial coordinates before equilibration) - repart_*.psf (same as the above psf files, but the masses of non-water hydrogen atoms have been repartitioned by VMD script repartitionMass.tcl) - rest*.pdb (same as the above pdb files, but atoms have been marked for restraints in NAMD. These files are generated by doPrep.sh, with restraints applied to different atoms.) Force Field Parameters ---------------------- CHARMM format parameter files: - par_all36m_prot.prm (CHARMM36m FF for proteins) - toppar_water_ions_prot.str (CHARMM water and ions with NBFIX parameters needed for protein and others commented out) Template NAMD Configuration Files --------------------------------- These contain the most commonly used simulation parameters. They are called by the other NAMD configuration files (which are in the namd/ subdirectory): - template_min.namd (minimization) - template_rest.namd (NPT equilibration with different parts of the protein restrained) - template_prod.namd (for the long production simulations) Minimization ------------- - namd/min_*.0.namd Restraints ------------- - namd/rest_*.0.namd (both CTLA4 binding site and peptide atoms are restrained) - namd/rest_*.1.namd (CA atoms of CTLA4 and all atoms of the peptide are restrained) - namd/rest_*.2.namd (all atoms of only the peptide are restrained) - namd/rest_*.3.namd (only CA atoms of only the peptide are restrained) - namd/rest_*.4.namd (no atoms are restrained) Production ------------- - namd/pro_*.{D,E,F}.0.namd Analysis ------------- - interaction.sh (Shell script for running analysis with VMD) - calcSeparationNearestAtom.tcl (Calculate the separation between two selections, taking the shortest distance between any pair of atoms spanning the two selections. Accounts for (orthogonal) periodic boundary conditions.) - useful.tcl (VMD Tcl script with a library of useful procs, used by the script above) - sep_*.dat (Output of the above analysis containing rows with two columns: time in nanoseconds and minimum distance in Å) Scripts ------- Files with the .sh extension can be found throughout. These usually provide the highest level control for submission of simulations and analysis. Look to these as a guide to what is happening.&nbsp

Interfacial water participates in a wide range of phenomena involving graphite, graphite-like and 2D material interfaces. Recently, several high-spatial resolution experiments have questioned the existence of hydration layers on graphite, graphite-like and 2D material surfaces. Here, 3D AFM was applied to follow in real-time and with atomic-scale depth resolution the evolution of graphite-water interfaces. Pristine graphite surfaces upon immersion in water showed the presence of several hydration layers separated by a distance of 0.3 nm. Those layers were short-lived. After several minutes, the interlayer distance increased to 0.45 nm. At longer immersion times (∼50 min) we observed the formation of a third layer. An interlayer distance of 0.45 nm characterizes the layering of predominantly alkane-like hydrocarbons. Molecular dynamics calculations supported the experimental observations. The replacement of water molecules by hydrocarbons on graphite is spontaneous. It happens whenever the graphite-water volume is exposed to air.

Hydrogen bonding plays a critical role in the self-assembly of peptide amphiphiles (PAs). Herein, we studied the effect of replacing the amide linkage between the peptide and lipid portions of the PA with a urea group, which possesses an additional hydrogen bond donor. We prepared three PAs with the peptide sequence Phe-Phe-Glu-Glu (FFEE): two are amide-linked with hydrophobic tails of different lengths and the other possesses an alkylated urea group. The differences in the self-assembled structures formed by these PAs were assessed using diverse microscopies, nuclear magnetic resonance (NMR), and dichroism techniques. We found that the urea group influences the morphology and internal arrangement of the assemblies. Molecular dynamics simulations suggest that there are about 50% more hydrogen bonds in nanostructures assembled from the urea-PA than those assembled from the other PAs. Furthermore

Dopamine clearance in the brain is controlled by the dopamine transporter (DAT), a protein residing in the plasma membrane, which drives reuptake of extracellular dopamine into presynaptic neurons. Studies have revealed that the βγ subunits of heterotrimeric G proteins modulate DAT function through a physical association with the C-terminal region of the transporter. Regulation of neurotransmitter transporters by Gβγ subunits is unprecedented in the literature; therefore, it is interesting to investigate the structural details of this particular protein-protein interaction. Here, we refined the crystal structure of the Drosophila melanogaster DAT (dDAT), modeling de novo the N- and C-terminal domains; subsequently, we used the full-length dDAT structure to generate a comparative model of human DAT (hDAT). Both proteins were assembled with Gβ1γ2 subunits employing protein-protein docking, and subsequent molecular dynamics simulations were run to identify the specific interactions governing the formation of the hDAT:Gβγ and dDAT:Gβγ complexes. A [L/F]R[Q/E]R sequence motif containing the residues R588 in hDAT and R587 in dDAT was found as key to bind the Gβγ subunits through electrostatic interactions with a cluster of negatively charged residues located at the top face of the Gβ subunit. Alterations of DAT function have been associated with multiple devastating neuropathological conditions; therefore, this work represents a step toward better understanding DAT regulation by signaling proteins, allowing us to predict therapeutic target regions.

A water extract derived from the isolated cell walls of

The antitumor effects of a partially purified water extract from Euglena gracilis (EWE) and EWE treated by boiling (bEWE) were evaluated using orthotopic lung cancer syngeneic mouse models with Lewis lung carcinoma (LLC) cells. Daily oral administration of either EWE or bEWE started three weeks prior to the inoculation of LLC cells significantly attenuated tumor growth as compared to the phosphate buffered saline (PBS) control, and the attenuation was further enhanced by bEWE. The intestinal microbiota compositions in both extract-treated groups were more diverse than that in the PBS group. Particularly, a decrease in the ratio of Firmicutes to Bacteroidetes and significant increases in Akkermansia and Muribaculum were observed in two types of EWE-treated groups. Fecal microbiota transplantation (FMT) using bEWE-treated mouse feces attenuated tumor growth to an extent equivalent to bEWE treatment, while tumor growth attenuation by bEWE was abolished by treatment with an antibiotic cocktail. These studies strongly suggest that daily oral administration of partially purified water extracts from Euglena gracilis attenuates lung carcinoma growth via the alteration of the intestinal microbiota.

The antitumor effects of a partially purified water extract from

Metal oxide nanoparticles have great potential for selective adsorption and catalytic degradation of contaminants from aqueous solutions. In this study, we employ mass spectrometry and molecular dynamics simulations to better understand the chemical and physical mechanisms determining the affinity of chlorobenzenes and polychlorinated biphenyls (PCBs) for zinc oxide nanoparticles (ZnO NPs). The experiments and simulations both demonstrate that the adsorption coefficients for chlorobenzenes increase steadily with the number of chlorine atoms, while, for PCBs, the relation is more complex. The simulations link this complexity to chlorine atoms at ortho positions hindering coplanar conformations. For a given number of chlorine atoms, the simulations predict decreasing adsorption affinity with increasing numbers of ortho substitutions. Consequently, the simulations predict that some of the highest adsorption affinities for ZnO NPs are exhibited by dioxin-like PCBs, suggesting the possibility of selective sequestration of these most acutely toxic PCBs. Remarkably, the experiments show that the PCB adsorption coefficients of ZnO NPs with diameters ≤ 80 nm exceed those of a soil sample by 5–7 orders of magnitude, meaning that a single gram of ZnO NPs could sequester low levels of PCB contamination from as much as a ton of soil.

After opening, the Shaker voltage-gated potassium (KV) channel rapidly inactivates when one of its four N-termini enters and occludes the channel pore. Although it is known that the tip of the N-terminus reaches deep into the central cavity, the conformation adopted by this domain during inactivation and the nature of its interactions with the rest of the channel remain unclear. Here, we use molecular dynamics simulations coupled with electrophysiology experiments to reveal the atomic-scale mechanisms of inactivation. We find that the first six amino acids of the N-terminus spontaneously enter the central cavity in an extended conformation, establishing hydrophobic contacts with residues lining the pore. A second portion of the N-terminus, consisting of a long 24 amino acid α-helix, forms numerous polar contacts with residues in the intracellular entryway of the T1 domain. Double mutant cycle analysis revealed a strong relationship between predicted interatomic distances and empirically observed thermodynamic coupling, establishing a plausible model of the transition of KV channels to the inactivated state.

Determination of membrane permeability to small molecules from first-principles represents a promising approach for screening lead compounds according to their permeation properties upstream in the drug discovery process and prior to their synthesis. Theoretical investigation of permeation events requires, at its core, a molecular model of the membrane, and the choice of this model impacts not only the predicted permeability but also its relation to the experimental measurements commonly performed in pharmaceutical settings with a variety of cell lines capable of mimicking intestinal passive permeation. Homogeneous single-lipid bilayers have traditionally been utilized in computer simulations of membrane permeability predictions due to the ease of sampling all the relevant configurations, as well as the availability of parameters for a range of components of the biological membrane. To assess the influence of the membrane heterogeneity on the permeability to small molecules, we have examined the permeation of ethanol in six different single-lipid bilayers and compared the computed free-energy and diffusivity profiles with those obtained using a mammalian cell membrane model consisting of 26 components. Our results suggest that the membrane permeability only mildly depends on the lipid composition, spanning only 1 order of magnitude between the small phosphoethanolamine and the large phosphocholine head groups, or the short, saturated lauryl and the long, unsaturated oleyl acyl chains, that is, nearly as close as current theoretical estimates can get to experiment. The staggering computer time required to obtain an accurate free-energy profile, devoid of hysteresis between the upper and the lower leaflets of the lipid bilayer, in excess of several microseconds, provides an impetus for the development of approximate routes for membrane permeability predictions. Here, we have modeled the free-energy profile underlying permeation by means of a series of free-energy perturbation calculations, whereby the substrate is reversibly coupled to its environment at fixed values in the direction normal to the lipid bilayer. The diffusivity profile is modeled based on the bulk self-diffusion of the permeant, and the membrane permeability is recovered without significant loss of accuracy. The proposed numerical approach can be seamlessly extended to the determination of the relative membrane permeability to alternate substrates, thereby allowing large sets of permeants to be screened at a fraction of the computational cost of a rigorous determination of their respective free-energy profile.

[EN] Hydration layers are formed on hydrophilic crystalline surfaces immersed in water. Their existence has also been predicted for hydrophobic surfaces, yet the experimental evidence is controversial. Using 3D-AFM imaging, we probed the interfacial water structure of hydrophobic and hydrophilic surfaces with atomic-scale spatial resolution. We demonstrate that the atomic-scale structure of interfacial water on crystalline surfaces presents two antagonistic arrangements. On mica, a common hydrophilic crystalline surface, the interface is characterized by the formation of 2 to 3 hydration layers separated by approximately 0.3 nm. On hydrophobic surfaces such as graphite or hexagonal boron nitride (h-BN), the interface is characterized by the formation of 2 to 4 layers separated by about 0.5 nm. The latter interlayer distance indicates that water molecules are expelled from the vicinity of the surface and replaced by hydrocarbon molecules. This creates a new 1.5-2 nm thick interface between the hydrophobic surface and the bulk water. Molecular dynamics simulations reproduced the experimental data and confirmed the above interfacial water structures. This journal is, European Research Council ERC-AdG-340177, the Ministerio de Ciencia, Innovación y Universidades (PID2019- 106801GB-I00; MAT2016-76507-R) and European Commission Marie Sklodowska-Curie grant agreement No. 721874. J.C. acknowledges financial support by the US National Science Foundation under Grant No. CHE-1726332 and DMR-1945589.

Lung cancer is the leading cause of cancer-related mortality in the world. Our previous studies have shown that the oral administration of Euglena water extract significantly inhibits the growth of grafted lung carcinoma tumors in mice. Euglena gracilis, a single-celled alga used as a nutritional dietary supplement, possesses a broad range of medicinal properties including anticancer activity against a few types of cancers. Most studies describing this anticancer activity have used xenograft cancer mouse models, but not carcinogen-induced cancer models. Since tobacco smoke carcinogen-induced lung carcinoma in mice mimics human lung cancer development, in this study, we investigated the anti-cancer properties and the underlying mechanism of the E. gracilis water extract against lung tumorigenesis induced by 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (NNK). A/J mice (n=10 per treatment group) were used and the treatment regimen consisted of: (i) PBS control, (ii) Euglena water extract 2 weeks before NNK injection (pre-NNK) and (iii) this extract 10 weeks after NNK injection (post-NNK). We examined the metabolite landscape of the gut microbiota by a high-throughput metabolomics approach. The Euglena water extract treatment greatly attenuated NNK-induced tumorigenesis in lungs of A/J mice. A Partial Least-Squares Discriminant Analysis demonstrated a separation of the fecal metabolites between the control and treatment groups. Using the untargeted metabolomics, we identified 20 potential metabolites that were differentially expressed in the Euglena extract (pre- and post-NNK) treatment groups as compared to the control group. Specifically, succinate, malate, triethanolamine, acetylserine were increased with the Euglena water extract treatment in comparison to the control. These metabolites are involved in the biosynthesis of short-chain fatty acids (SCFA). Furthermore, using the targeted analysis of SCFA, we observed a significant increase in acetic acid in the feces of mice treated with Euglena water extract both before and after NNK exposure; however, butyric and propionic acids were significantly higher in only post-treated mice. Moreover, in vitro treatment of both human and murine lung cancer cells with SCFA significantly suppressed their proliferation. The present study indicated that treating with the Euglena water extract both pre- and post-NNK exposure greatly inhibits NNK-induced lung tumorigenesis in mice even after early tumorigenesis has been established. This study also found that Euglena water extract increases the levels of SCFA and their precursors in the feces of the treated mice indicating a role of gut microbiota in suppressing the NNK-induced tumorigenesis. This study was supported by 2017 EUGLENA-RC2 (MT and JC), Kansas State University Johnson Cancer Research Center (MT and JC), and NIH grant P20 GM103418 (MT). Citation Format: Deepa Upreti, Susumu Ishiguro, Ayaka Nakashima, Kengo Suzuki, Jeffrey Comer, Masaaki Tamura. Euglena water extract attenuates lung tumorigenesis induced by tobacco-specific carcinogen through modulation of gut microbiota metabolites [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5944.

Three new collagen mimetic peptides containing the CLK motif as anchoring arms were tested for silver nanoparticle surface stabilization. Our experimental and molecular dynamics data indicate that peptide length has an important effect not only in the resulting nanosilver's colloidal stability, but also in the biological performance of the composite.

The partially purified water extract from Euglena gracilis (EWE) was evaluated for its antitumor and immunomodulatory effects in cell cultures and in a mouse orthotopic lung carcinoma allograft model. In two-dimensional cell culture, the EWE treatment inhibited cell growth of both murine Lewis lung carcinoma (LLC) and human lung carcinoma cells (A549 and H1299) in a dose- and time-dependent manner. In contrast, the growth of mouse bone marrow cells (BMCs), but not mouse splenocytes (SPLs), was stimulated by the treatment with EWE. In three-dimensional spheroid culture, spheroid growth of LLC cells was significantly attenuated by EWE treatment. In a mouse LLC orthotopic allograft model, pretreatment with EWE (150–200 mg/kg/day, via drinking water) three weeks prior to the LLC cell inoculation, but not post-treatment after LLC cell inoculation, significantly attenuated the growth of LLC tumors in immunocompetent syngeneic mouse lung. This tumor growth attenuation coincided with a significant decrease in the population of myeloid-derived cells, primarily neutrophils. Flow cytometric analysis revealed that the EWE treatment significantly attenuated growth of granulocytic myeloid-derived suppressor cells (gMDSC) in BMCs and that this decrease was due to induction of gMDSC-specific apoptosis and differentiation of monocytic MDSCs (mMDSC) to macrophages. The present study provides evidence that EWE pretreatment inhibits lung carcinoma growth mainly by stimulating host antitumor immunity through attenuation of growth of gMDSCs and decreasing the number of peripheral granulocytes. This study suggests that the partially purified extract derived from Euglena gracilis contains significant bioactive materials that prevent lung carcinoma growth.

Prediction of membrane permeability to small molecules represents an important aspect of drug discovery. First-principles calculations of this quantity require an accurate description of both the thermodynamics and kinetics that underlie translocation of the permeant across the lipid bilayer. In this contribution, the membrane permeability to three drugs, or drug-like molecules, namely, 9-anthroic acid (ANA), 2',3'-dideoxyadenosine (DDA), and hydrocortisone (HYL), are estimated in a pure 1-palmitoyl-2-oleoylphosphatidylcholine (POPC) and in a POPC:cholesterol (2:1) mixture. On the basis of independent 2-5-μs free-energy calculations combined with a time-fractional Smoluchowski determination of the diffusivity, the estimated membrane permeabilities to these chemically diverse permeants fall within an order of magnitude from the experimental values obtained in egg-lecithin bilayers, with the exception of HYL in pure POPC. This exception is particularly interesting because the calculated permeability of the sterol-rich bilayer to HYL, in close agreement with the experimental value, is about 600 times lower than that of the pure POPC bilayer to HYL. In contrast, the permeabilities to ANA and DDA differ by less than a factor of 10 between the pure POPC and POPC:cholesterol bilayers. The unusual behavior of HYL, a large, amphiphilic compound, may be linked with the longer range perturbation of the lipid bilayer it induces, compared to ANA and DDA, suggestive of a possibly different translocation mechanism. We find that the tendency of lower permeabilities of the POPC:cholesterol bilayer relative to those of the pure POPC one is a consequence of increased free-energy barriers. Beyond reporting accurate estimates of the membrane permeability, the present contribution also demonstrates that rigorous free-energy calculations and a fractional-diffusion model are key in revealing the molecular phenomena linking the composition of a membrane to its permeability to drugs.

Predicting adsorption of organic pollutants onto graphene nanomaterials is not only useful for exploring their potential adsorbent applications, but also helpful for better understanding their fate and risks in aquatic environments. Herein molecular dynamics (MD) simulations and theoretical linear solvation energy relationships (TLSERs) were employed to construct prediction models for adsorption of neutral organic pollutants onto graphene and graphene oxides. The MD simulations for adsorption of 43 aromatic compounds onto graphene and diverse models of graphene oxides with various functional groups (hydroxyl, epoxy and carbonyl) demonstrate that graphene has a stronger affinity for the aromatic compounds than graphene oxides. The hydroxyl and carbonyl groups of graphene oxides were found to form hydrogen bonds with the aromatic adsorbates, while epoxy groups did not. TLSER models were developed for predicting the adsorption equilibrium coefficients (K) onto graphene and graphene oxide nanosheets. In the graphene prediction model, H-donating ability (eα) and dispersion/hydrophobic interactions (V) have significant effects on log K values, while in the graphene oxide model, eα is the most influential factor on log K values. The models provide in silico approaches for predicting adsorption affinities onto graphenic nanomaterials.

We investigated two critical aspects of rose Bengal (RB) photosensitized protein cross-linking that may underlie recently developed medical applications. Our studies focused on the binding of RB to collagen by physical interaction and the effect of this binding and certain amino acids on RB photochemistry. Molecular dynamics simulations and free-energy calculation techniques, complemented with isothermal titration calorimetry, provided insight into the binding between RB and a collagen-like peptide (CLP) at the atomic level. Electrostatic interactions dominated, which is consistent with the finding that RB bound equally well to triple helical and single chain collagen. The binding free energy ranged from −5.7 to −3 kcal/mol and was strongest near the positively charged amino groups at the N-terminus and on lysine side chains. At high RB concentration, a maximum of 16 ± 3 bound dye molecules per peptide was found, which is consistent with spectroscopic evidence for aggregated RB bound to collagen or the CLP. Within a tissue-mimetic collagen matrix, RB photobleached rapidly, probably due to electron transfer to certain protein amino acids, as was demonstrated in solutions of free RB and arginine. In the presence of arginine and low oxygen concentrations, a product absorbing at 510 nm formed, presumably due to dehalogenation after electron transfer to RB. In the collagen matrix without arginine, the dye generated singlet oxygen as well as the 510 nm product. These results provide the first evidence of the effects of a tissue-like environment on the photochemical mechanisms of rose Bengal.

Snake bite envenoming is a public health problem that was recently included in the list of neglected tropical diseases of the World Health Organization. In the search of new therapies for the treatment of local tissue damage induced by snake venom metalloproteinases (SVMPs), we tested the inhibitory activity of peptidomimetic compounds designed as inhibitors of matrix metalloproteinases on the activities of the SVMP Batx-I, from Bothrops atrox venom. The evaluated compounds show great potential for the inhibition of Batx-I proteolytic, hemorrhagic and edema-forming activities, especially the compound CP471474, a peptidomimetic including a hydroxamate zinc binding group. Molecular dynamics simulations suggest that binding of this compound to the enzyme is mediated by the electrostatic interaction between the hydroxamate group and the zinc cofactor, as well as contacts, mainly hydrophobic, between the side chain of the compound and amino acids located in the substrate binding subsites S1 and S1 &prime, These results show that CP471474 constitutes a promising compound for the development of co-adjuvants to neutralize local tissue damage induced by snake venom metalloproteinases.

Currently, there is a great interest in nanoparticle-based vaccine delivery. Recent studies suggest that nanoparticles when introduced into the biological milieu are not simply passive carriers but may also contribute immunological activity themselves or of their own accord. For example there is considerable interest in the biomedical applications of one of the physiologically-based inorganic metal oxide nanoparticle, zinc oxide (ZnO). Indeed zinc oxide (ZnO) NP are now recognized as a nanoscale chemotherapeutic or anticancer nanoparticle (ANP) and several recent reports suggest ZnO NP and/or its complexes with drug and RNA induce a potent antitumor response in immuno-competent mouse models. A variety of cell culture studies have shown that ZnO NP can induce cytokines such as IFN-γ, TNF-α, IL-2, and IL-12 which are known to regulate the tumor microenvironment. Much less work has been done on magnesium oxide (MgO), cobalt oxide (Co3O4), or nickel oxide (NiO); however, despite the fact that these physiologically-based metal oxide NP are reported to functionally load and assemble RNA and protein onto their surface and may thus also be of potential interest as nanovaccine platform. Here we initially compared in vitro immunogenicity of ZnO and Co3O4 NP and their effects on cancer-associated or tolerogenic cytokines. Based on these data we moved ZnO NP forward to testing in the ex vivo splenocyte assay relative to MgO and NiO NP and these data showed significant difference for flow cytometry sorted population for ZnO-NP, relative to NiO and MgO. These data suggesting both molecular and cellular immunogenic activity, a double-stranded anticancer RNA (ACR), polyinosinic:poly cytidylic acid (poly I:C) known to bind ZnO NP; when ZnO-poly I:C was injected into B16F10-BALB/C tumor significantly induced, IL-2 and IL-12 as shown by Cohen’s d test. LL37 is an anticancer peptide (ACP) currently in clinical trials as an intratumoral immuno-therapeutic agent against metastatic melanoma. LL37 is known to bind poly I:C where it is thought to compete for receptor binding on the surface of some immune cells, metastatic melanoma and lung cells. Molecular dynamic simulations revealed association of LL37 onto ZnO NP confirmed by gel shift assay. Thus using the well-characterized model human lung cancer model cell line (BEAS-2B), poly I:C RNA, LL37 peptide, or LL37-poly I:C complexes were loaded onto ZnO NP and delivered to BEAS-2B lung cells, and the effect on the main cancer regulating cytokine, IL-6 determined by ELISA. Surprisingly ZnO-LL37, but not ZnO-poly I:C or the more novel tricomplex (ZnO-LL37-poly I:C) significantly suppressed IL-6 by >98–99%. These data support the further evaluation of physiological metal oxide compositions, so-called physiometacomposite (PMC) materials and their formulation with anticancer peptide (ACP) and/or anticancer RNA (ACR) as a potential new class of immuno-therapeutic against melanoma and potentially lung carcinoma or other cancers.

An understanding of the nucleation and growth of hexagonal boron nitride (hBN) on nickel substrates is essential to its development as a functional material. In particular, fundamental insights into the formation of the hexagonal lattices with alternating boron (B) and nitrogen (N) atoms could be exploited to control hBN lattice morphologies for targeted applications. In this study, the preferred shapes and edge configurations of atomically smooth hBN on Ni(111) were investigated using molecular dynamics (MD) simulations, along with reactive force field (ReaxFF) developed to represent the Ni/B/N system and the lattice-building B–N bond formation. The obtained hBN lattices, from different B : N feed ratios, are able to confirm that hBN domain geometries can indeed be tuned by varying thermodynamic parameters (i.e., chemical potentials of N and B) – a finding that has only been predicted using quantum mechanical theories. Here, we also showed that the nitrogen fed to the system plays a more crucial role in dictating the size of hBN lattices. With an increase of the relative N content, the simulated hBN domain shapes also transition from equilateral triangles to hexagons, again, consistent with the anticipation based on Density Functional Theory (DFT) calculations. Hence, a plausible approach to acquire a desired hBN nanostructure depends on careful control over the synthesis conditions, which now can benefit from reliable molecular simulations.

In this chapter, we provide a review of the use of molecular modeling and simulation to reveal the atomic interactions and thermodynamics underlying biomolecule adsorption on inorganic nanoparticles. We describe how these methods have helped to understand the effects of the interface structure on adsorption processes, highlighting the recent advances and novel applications. Also, we will discuss the importance of computational techniques to permit rational design of new functionalized nanomaterials. The challenges and future opportunities of the use of molecular simulation in studying adsorption phenomena is also addressed, with special emphasis on the enhancing sampling methods used to estimate the adsorption free energy, which in many cases provide a realistic kinetic model of the interactions of organic compounds or biomolecules with metal nanomaterials. Finally, we describe recent advances in the use of hybrid (QM/MM) methods and reactive force fields (ReaxFF) to characterize bionanomolecule complexes.

Beta glucans are known to have immunomodulatory effects that mediated by a variety of mechanisms. In this article, we describe experiments and simulations suggesting that beta-1,3 glucans may promote activation of T cells by a previously unknown mechanism. First, we find that treatment of a T lymphoblast cell line with beta-1,3 oligoglucan significantly increases mRNA levels of T cell activation-associated cytokines, especially in the presence of the agonistic anti-CD3 antibody. This immunostimulatory activity was observed in the absence of dectin-1, a known receptor for beta-1,3 glucans. To clarify the molecular mechanism underlying this activity, we performed a series of molecular dynamics simulations and free-energy calculations to explore the interaction of beta-1,3 oligoglucans with potential immune receptors. While the simulations reveal little association between beta-1,3 oligoglucan and the immune receptor CD3, we find that beta-1,3 oligoglucans bind to CD28 near the region identified as the binding site for its natural ligands CD80 and CD86. Using a rigorous absolute binding free-energy technique, we calculate a dissociation constant in the low millimolar range for binding of 8-mer beta-1,3 oligoglucan to this site on CD28. The simulations show this binding to be specific, as no such association is computed for alpha-1,4 oligoglucan. This study suggests that beta-1,3 glucans bind to CD28 and may stimulate T cell activation collaboratively with T cell receptor activation, thereby stimulating immune function.

Measurements of ionic currents through nanopores partially blocked by DNA have emerged as a powerful method for characterization of the DNA nucleotide sequence. Although the effect of the nucleotide sequence on the nanopore blockade current has been experimentally demonstrated, prediction and interpretation of such measurements remain a formidable challenge. Using atomic resolution computational approaches, here we show how the sequence, molecular conformation, and pore geometry affect the blockade ionic current in model solid-state nanopores. We demonstrate that the blockade current from a DNA molecule is determined by the chemical identities and conformations of at least three consecutive nucleotides. We find the blockade currents produced by the nucleotide triplets to vary considerably with their nucleotide sequences despite having nearly identical molecular conformations. Encouragingly, we find blockade current differences as large as 25% for single-base substitutions in ultra small (1.6 nm × 1.1 nm cross section; 2 nm length) solid-state nanopores. Despite the complex dependence of the blockade current on the sequence and conformation of the DNA triplets, we find that, under many conditions, the number of thymine bases is positively correlated with the current, whereas the number of purine bases and the presence of both purines and pyrimidines in the triplet are negatively correlated with the current. Based on these observations, we construct a simple theoretical model that relates the ion current to the base content of a solid-state nanopore. Furthermore, we show that compact conformations of DNA in narrow pores provide the greatest signal-to-noise ratio for single base detection, whereas reduction of the nanopore length increases the ionic current noise. Thus, the sequence dependence of the nanopore blockade current can be theoretically rationalized, although the predictions will likely need to be customized for each nanopore type.

Phytochemicals have been developed into treatments against many diseases, including various cancers. Euglena is a genus of single-cell flagellate eukaryotes having features of both animals and plants. These organisms, which are found in both fresh and salt water, have been used as a dietary supplement for their rich nutrient content. Although recent studies suggest that Euglena extracts have anti-tumor properties, the mechanism is yet to be clarified. Accordingly, the anti-tumor and immunomodulatory effects of partially purified water extract from Euglena gracilis (EWE) was evaluated in cell culture and a mouse orthotopic lung carcinoma allograft model. The EWE was prepared by a PBS extraction at 37°C for 30 min, centrifugation at 2,000 g for 20 min, and filtration through a membrane filter (0.22 μm pore size). Primary components of this extract are water soluble materials and exclude mature paramylons, intact chloroplasts, and oil droplets. In two-dimensional cell culture, the EWE treatment inhibited cell growth of both murine Lewis lung carcinoma (LLC) and human lung carcinoma cells (A549 and H1299) in a dose (1-100 μg/ml) and time (24-72 h)-dependent manner. In contrast, the growth of mouse bone marrow cells (BMCs), but not mouse splenocytes (SPLs) were dose (1-100 μg/ml) and time (24-72 h)-dependently stimulated by the treatment with EWE. In three-dimensional spheroid culture, spheroid growth of LLC cells was significantly attenuated by EWE treatment. Flow cytometry revealed that the EWE treatment attenuated granulocytic myeloid-derived suppressor cells in BMCs. In a mouse LLC orthotopic allograft model, pretreatment with EWE (100-200 mg/kg/day, via drinking water) three weeks prior to LLC cell inoculation significantly attenuated the growth of LLC tumors in the lungs of immunocompetent syngeneic mice. However, this attenuation was not seen for EWE treatment initiated after LLC cell inoculation. The tumor growth attenuation by EWE pretreatment coincided with a significant decrease of myeloid-derived cells, primarily neutrophils. EWE pretreatment appears to inhibit lung carcinoma growth mainly by stimulating host anti-tumor immunity via an attenuation of the host myeloid-derived immune suppressor cells. Hence, the present study suggests that the partially purified extract derived from Euglena gracilis contains bioactive materials that prevent lung carcinoma growth. This study was supported by 2016EUGLENA-RC1 (MT), 2017EUGLENA-RC2 (MT and JC), a Kansas State University College of Veterinary Medicine SMILE award (MT and JC), and NIH grant P20 RR017686 (MT). Citation Format: Susumu Ishiguro, Deepa Upreti, Nicole Robben, Page Cote, Riley Burghart, Damilola Ogun, Tran Le, Jennifer Delzeit, Jayson Carswell, Arashi Nakashima, Ayaka Nakashima, Kengo Suzuki, Jeffrey Comer, Masaaki Tamura. Water extract from Euglena gracilis prevents lung carcinoma growth in mice via attenuation of myeloid-derived suppressor cells and granulocytes [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 3459.

A colon cancer growth inhibitor partially purified from the isolated cell wall membrane fraction of Chlorella sorokiniana, here referred to as Chlorella membrane factor (CMF), was evaluated for its antitumor and immunomodulatory effects in cell culture and in a colon carcinoma mouse model. The CMF treatment dose- and time-dependently inhibited colon carcinoma cell growth in 2-dimensional cultures. Treatment with CMF also significantly inhibited the growth of colon carcinoma spheroids in 3-dimensional cell culture in coculture with T lymphocytes. In a mouse CT26 colon carcinoma peritoneal dissemination model, intraperitoneal injection of CMF (10 or 30 mg dry weight/kg body weight, every other day) dose-dependently and significantly attenuated the growth of tumor nodules via induction of tumor cell apoptosis. Evaluation of immune cell populations in ascites showed that CMF treatment tended to increase T lymphocytes but lower granulocyte populations. The present study suggests that the cell wall membrane fraction of Chlorella sorokiniana contains a bioactive material that inhibits colon carcinoma growth via direct cell growth inhibition and stimulation of host antitumor immunity. Hence, it is suggested that the Chlorella cell wall membrane extract or a bioactive substance in the extract is an attractive complementary medicine for cancer therapy.

Adsorption of organic molecules from aqueous solution to the surface of carbon nanotubes or graphene is an important process in many applications of these materials. Here we use molecular dynamics simulation, supplemented by analytical chemistry, to explore in detail the adsorption thermodynamics of a diverse set of aromatic compounds on graphenic materials, elucidating the effects of the solvent, surface coverage, surface curvature, defects, and functionalization by hydroxy groups. We decompose the adsorption free energies into entropic and enthalpic components and find that different classes of compounds-such as phenols, benzoates, and alkylbenzenes-can easily be distinguished by the relative contributions of entropy and enthalpy to their adsorption free energies. Overall, entropy dominates for the more hydrophobic compounds, while enthalpy plays the greatest role for more hydrophilic compounds. Experiments and independent simulations using two different force field frameworks (CHARMM and Amber) support the robustness of these conclusions. We determine that concave curvature is generally associated with greater adsorption affinity, more favorable enthalpy, and greater contact area, while convex curvature reduces both adsorption enthalpy and contact area. Defects on the graphene surfaces can create concave curvature, resulting in localized binding sites. As the graphene surface becomes covered with aromatic solutes, the affinity for adsorbing an additional solute increases until a complete monolayer is formed, driven by more favorable enthalpy and partially canceled by less favorable entropy. Similarly, hydroxylation of the surface leads to preferential adsorption of the aromatic solutes to remaining regions of bare graphene, resulting in less favorable adsorption entropy, but compensated by an increase in favorable enthalpic interactions.

Most of the snakebite envenomations in Central and South America are caused by species belonging to Bothrops genus. Their venom is composed mainly by zinc-dependent metalloproteinases, responsible of the hemorrhage characteristic of these envenomations. The aim of this study was to determine the inhibitory ability of ten flavonoids on the in-vitro proteolytic activity of Bothrops atrox venom and on the hemorrhagic, edema-forming and myonecrotic activities of Batx-I, the most abundant metalloproteinase isolated from this venom. Myricetin was the most active compound, exhibiting an IC 50 value of 150 μ M and 1021 μ M for the inhibition of proteolytic and hemorrhagic activity, respectively. Independent injection experiments, with a concentration of 1600 μ M of myricetin administered locally, immediately after toxin injection, demonstrated a reduction of 28 ± 6 % in the hemorrhagic lesion. Additionally, myricetin at concentrations 800, 1200 and 1600 μ M promoted a reduction in plasma creatine kinase activity induced by Batx-I of 21 ± 2 % , 60 ± 5 % and 63 ± 2 % , respectively. Molecular dynamics simulations coupled with the adaptive biasing method suggest that myricetin can bind to the metalloproteinase active site via formation of hydrogen bonds between the hydroxyl groups 3’, 4’ and 5’ of the benzyl moiety and amino acid Glu143 of the metalloproteinase. The hydroxyl substitution pattern of myricetin appears to be essential for its inhibitory activity. Based on this evidence, myricetin constitutes a candidate for the development of inhibitors to reduce local tissue damage in snakebite envenomations.

Small molecule inhibitors of snake venom metalloproteinases (SVMPs) could provide a means to rapidly halt the progression of local tissue damage following viperid snake envenomations. In this study, we examine the ability of candidate compounds based on a pentacyclic triterpene skeleton to inhibit SVMPs. We leverage molecular dynamics simulations to estimate the free energies of the candidate compounds for binding to BaP1, a P-I type SVMP, and compare these results with experimental assays of proteolytic activity inhibition in a homologous enzyme (Batx-I). Both simulation and experiment suggest that betulinic acid is the most active candidate, with the simulations predicting a standard binding free energy of &Delta, G ∘ = &minus, 11.0 &plusmn, 1.4 kcal/mol. The simulations also reveal the atomic interactions that underlie binding between the triterpenic acids and BaP1, most notably the electrostatic interaction between carboxylate groups of the compounds and the zinc cofactor of BaP1. Together, our simulations and experiments suggest that occlusion of the S1 &prime, subsite is essential for inhibition of proteolytic activity. While all active compounds make hydrophobic contacts in the S1 &prime, site, &beta, boswellic acid, with its distinct carboxylate position, does not occlude the S1 &prime, site in simulation and exhibits negligible activity in experiment.

Introduction Biomedical applications of nanoparticles (NPs) as enzyme inhibitors have recently come to light. Oxides of metals native to the physiological environment (eg, Fe, Zn, Mg, etc.) are of particular interest—especially the functional consequences of their enzyme interaction. Materials and methods Here, Fe2O3, zinc oxide (ZnO), magnesium oxide (MgO) and nickel oxide (NiO) NPs are compared to copper (Cu) and boron carbide (B4C) NPs. The functional impact of NP interaction to the model enzyme luciferase is determined by 2-dimensional fluorescence difference spectroscopy (2-D FDS) and 2-dimensional photoluminescence difference spectroscopy (2-D PLDS). By 2-D FDS analysis, the change in maximal intensity and in 2-D FDS area under the curve (AUC) is in the order Cu~B4C>ZnO>NiO>>Fe2O3>MgO. The induced changes in protein conformation are confirmed by tryptic digests and gel electrophoresis. Results Analysis of possible trypsin cleavage sites suggest that cleavage mostly occurs in the range of residues 112–155 and 372–439, giving a major 45 kDa band. By 2-D PLDS, it is found that B4C NPs completely ablate bioluminescence, while Cu and Fe2O3 NPs yield a unique bimodal negative decay rate, −7.67×103 and −3.50×101 relative light units respectively. Cu NPs, in particular, give a remarkable 271% change in enzyme activity. Molecular dynamics simulations in water predicted that the surfaces of metal oxide NPs become capped with metal hydroxide groups under physiological conditions, while the surface of B4C becomes populated with boronic acid or borinic acid groups. These predictions are supported by the experimentally determined zeta potential. Thin layer chromatography patterns further support this conception of the NP surfaces, where stabilizing interactions were in the order ionic>polar>non-polar for the series tested. Conclusion Overall the results suggest that B4C and Cu NP functional dynamics on enzyme biochemistry are unique and should be examined further for potential ramifications on other model, physiological or disease-relevant enzymes., Video abstract

Dendrimers provide a means to control the synthesis of gold nanoparticles and stabilize their suspensions. However, design of improved dendrimers for this application is hindered by a lack of understanding how the dendrimers and synthesis conditions determine nanoparticle morphology and suspension stability. In the present work, we evaluate the effect of polyamidoamine (PAMAM) dendrimers terminated with different functional groups (-OH or -NH

Gold nanorods (AuNRs) possess unique photothermal properties due to their strong plasmonic absorption in the near-infrared region of the electromagnetic spectrum. They have been explored widely as an alternative or a complement to chemotherapy in cancer treatment. However, the use of AuNRs as an injectable medicine is greatly hindered by their stability in biological media. Therefore, studies have been focused on improving the stability of AuNRs by introducing biocompatible surface functionalizations such as polyethylene glycol (PEG) coatings. However, these coatings can affect heat conduction and alter their photothermal behavior. Herein, we studied how functionalization of AuNRs with PEG chains of different molecular weights determined the temperature distribution of suspensions under near-infrared irradiation, cell uptake in vitro, and hyperthermia-induced cytotoxicity. Thermogravimetric analysis of the PEG-conjugated AuNRs exhibited slightly different PEG mass fractions of 12.0%, 12.7%, and 18.5% for PEG chains with molecular weights of 2, 5, and 10 kDa, respectively, implying distinct structures for PEG brushes. When exposed to near-infrared radiation, we found greater temperatures and temperature gradients for longer PEG chains, while rapid aggregation was observed in unmodified (raw) AuNRs. The effect of the PEG coating on heat transport was investigated using molecular dynamics simulations, which revealed the atomic scale structure of the PEG brushes and demonstrated lower thermal conductivity for PEG-coated AuNRs than for unmodified AuNRs. We also characterized the uptake of the AuNRs into mouse melanoma cells in vitro and determined their ability to kill these cells when subjected to near-infrared radiation. For all PEG-coated AuNRs, exposure to 10 s of near-infrared radiation significantly reduced cell viability relative to unirradiated controls, with this viability further decreasing with increasing AuNR doses, indicating potential phototherapeutic effects. The 5 kDa PEG coating appeared to yield the best performance, yielding significant phototoxicity at even the lowest dose considered (0.5 μg mL-1), while also exhibiting high colloidal stability, which could help in rational design consideration of AuNRs for NIR induced photothermal therapy.