Addressing the metabolic activation potential of new leads in drug discovery: a case study using ion trap mass spectrometry and tritium labeling techniques

Metabolic activation of drug candidates to electrophilic reactive metabolites that can covalently modify cellular macromolecules may result in acute and/or idiosyncratic immune system-mediated toxicities in humans. This presents a significant potential liability for the future development of these compounds as safe therapeutic agents. We present here an example of an approach where sites of metabolic activation within a new drug candidate series were rapidly identified using online liquid chromatography/multi-stage mass spectrometry on an ion trap mass spectrometer. This was accomplished by trapping the reactive intermediates formed upon incubation of compounds with rat and human liver microsomes as their corresponding glutathione conjugates and mass spectral characterization of these thiol adducts. Based on the structures of the GSH adducts identified, potential sites and mechanisms of bioactivation within the chemical structure were proposed. These metabolism studies were interfaced with iterative structural modifications of the chemical series in order to block these bioactivation sites within the molecule. This strategy led to a significant reduction in the propensity of the compounds to undergo metabolic activation as evidenced by reductions in the irreversible binding of radioactivity to liver microsomal material upon incubation of tritium-labeled compounds with this in vitro system. With the efficiency and throughput achievable with such an approach, it appears feasible to identify and address the metabolic activation potential of new drug leads during routine metabolite identification studies in an early drug discovery setting.     

Reactions of metal carbonyl cluster complexes with multidentate phosphine ligands; reactions with bis(diphenylphosphino)methane

Reactions of Rh₆(CO)₁₆ with bis(diphenylphosphino)methane (dppm) gave Rh₆(CO)₁₄(dppm), Rh₆(CO)₁₂(dppm)₂, or Rh₆(CO)₁₀(dppm)₃, depending upon the reaction conditions. Rh₄(CO)₁₀(dppm) may be obtained from the reaction of Rh₄(CO)₁₂ with dppm, but this derivative rapidly decomposes in solution to give Rh₄(CO)₈(dppm)₂, Rh₆(CO)₁₄(dppm), and Rh₆(CO)₁₂(dppm)₂. Ir₄(CO)₁₀(dppm) and Ir₄(CO)₈(dppm)₂ have also been prepared, and their structures are discussed on the basis of infrared and ³¹P NMR spectroscopic data.     

Carbon nanohorns grown from ruthenium nanoparticles

A nanoscale ruthenium/gold bimetallic cluster of clusters has been used as a molecular precursor to produce pure ruthenium nanoparticles (seeds) as catalysts for the growth of carbon nanohorns (CNHs).     

Mirror image nanostructures

Chiral molecules that self-assemble to form chiral supramolecular structures exhibit interesting structural features reminiscent of tertiary and quaternary structures of proteins and have applications in catalysis and nonlinear optics. Often, these structures are hierarchical, with their chiral structure difficult to interpret on the molecular scale. In this communication, we observe chiral assembling molecules that form well-defined helices with a pitch of 28 nm. We observe the behavior in both R- and S-enantiomers of the molecule, forming mirror image nanostructures. The molecular chirality is determined by the dimethyloctyl alkyl coil of the molecule and is located more than 4 nm from the hydrogen-bonding segment. The nanostructures observed are not hierarchical, which could be a result of the significant separation between the stereocenter and hydrogen-bonding dendron. The subtle structural modification at the periphery of the molecule biases the supramolecular assembly, which is driven primarily by strong hydrogen-bonding and pi-pi stacking interactions.     

High-yield solvothermal formation of magnetic CoPt alloy nanowires

One-dimensional (1D) magnetic nanomaterials have attracted much attention recently because of their applications in magnetic recording and spintronics. Nevertheless, it remains a challenge to prepare free-standing magnetic nanowires in high yield. This Communication reports the successful high-yield synthesis of an interesting 1D ferromagnetic CoPt alloy by direct decomposition of platinum acetylacetonate and cobalt carbonyl compound in ethylenediamine solvent through a solvothermal reaction. The CoPt alloy nanowires obtained have a tunable diameter of 10-50 nm and a length along the longitudinal axis of up to several microns, depending on crystallization temperature and reaction time. A unique formation mechanism involving coarsening and ripening under solvothermal conditions was discovered. This research opens new opportunities in synthesizing nanomaterials through low-temperature solvothermal processes.     

THREE EXPONENTIAL FLUORESCENCE DECAY OF HORSE LIVER ALCOHOL DEHYDROGENASE REVEALED BY IODIDE QUENCHING

Abstract— The time resolved fluorescence decay of horse liver alcohol dehydrogenase was measured using a frequency doubled picosecond dye laser and time-correlated single-photon counting detection. A flow-cell technique is described which eliminates the photodegradation artifacts which commonly occur with laser excitation. A procedure is introduced which uses fluorescence quenching to reveal minor fluorescence lifetime components. The decay of the unquenched tryptophanyl fluorescence could be described by a double exponential decay law, but experiments conducted in the presence of iodide ion showed that the fluorescence decay must be more complex than this. A model is presented in which the fluorescence decay consists of three exponential components, only two of which are susceptible to quenching by iodide ion. Several possibilities are presented for the origin of this minor decay component, the most reasonable of which is that it arises from conformational heterogeneity in the solvent-exposed tryptophanyl residue.     

Multiple sprayer system for high-throughput electrospray ionization mass spectrometry

A multiple sprayer electrospray ion source for high-throughput analysis is described. The ion source is comprised of multiple electrospray capillaries, each with an ion lens located near the tip. The electric potentials applied to the ion lenses are used to control the sprayers. The use of ion lenses eliminates the need for mechanical blocking devices to selectively enable or disable the sprayers, and results in a less expensive and more reliable set-up. Sprayers can be enabled or disabled within approximately 50-250 ms when the lens potentials are controlled manually. For simultaneous operation of multiple electrospray capillaries, it is advantageous to orient the capillaries so that the spray from each passes directly in front of the entrance aperture of the mass spectrometer.     

PHOTODYNAMIC THERAPY INDUCED ULTRASTRUCTURAL ALTERATIONS IN MICROVASCULATURE OF THE RAT CREMASTER MUSCLE

Abstract— Photodynamic therapy disrupts blood flow to tumors and produces tumor necrosis. These effects may be due to a localized generation of singlet oxygen. The current studies used direct observations of the rat cremaster microvasculature to examine the vascular effects of PDT. The objective of the morphological examination was to delineate the structural basis for the altered blood flow in photodynamic therapy. Dihematoporphyrin ether given 30 min or 48 h prior to the experiment was activated with green light (wavelength530–560 nm, 120 J/cm²). After the in vivo activation the tissues were prepared for electron microscopy. Light alone induced little or no change in the luminal content or vessel wall. On exposure to activating light both acute (30 min) and long term (48 h) dihematoporphyrin ether pretreated samples displayed formation of luminal aggregates, granulocyte margination and migration, and endothelial cell and smooth muscle cell damage. The latter was more pronounced in the arterioles than the venules. Perivascular changes included interstitial edema and damage to striated myocytes. Some of the alterations such as interstitial edema may be transient; however, smooth and skeletal muscle cell injury are important in normal and tumor tissue necrosis after photodynamic therapy.     

The structural characteristics of organozinc complexes incorporating N,N'-bidentate ligands

Dimethylzinc reacts with an excess of N-2-pyridylaniline 6 to give the homoleptic species, Zn[PhN(2-C(5)H(4)N)](2) 8. Single crystal X-ray diffraction reveals a solid-state dimer based on an 8-membered (NCNZn)(2) core motif. Zn[CyN(2-C(5)H(4)N)]Me (Cy =c-C(6)H(11)) 10, prepared by the combination of ZnMe(2) with the corresponding cyclohexyl-substituted pyridylamine, is also dimeric in the solid state but reveals a central (ZnN)(2) metallacycle. Employment of (p-Tol)NH(2-C(5)H(4)N)(p-Tol = 4-MeC(6)H(4)) 11 yielded the tris(zinc) adduct Zn(3)[(p-Tol)N(2-C(5)H(4)N)](4)Me(2) 12, which incorporates a central chiral molecule of 'Zn[(p-Tol)N(2-C(5)H(4)N)](2)' 12a, that bridges two 'Zn[(p-Tol)N(2-C(5)H(4)N)]Me' 12b units. A similar trimetallic structure is noted when the pyridylaniline substrate 11 is replaced with the bicyclic guanidine 1,3,4,6,7,8-hexahydro-2H-pyrimido[1,2-a]pyrimidine (hppH), affording Zn(3)(hpp)(4)Me(2) 13. Spectroscopic studies point to retention of the solid-state structure of in hydrocarbon solution. Reaction of 13 with dimesityl borinic acid, Mes(2)BOH (Mes = mesityl), affords Zn(3)(hpp)(4)(OBMes(2))(2) 14 in which the trimetallic core is retained. This reactivity is in contrast to the closely related reaction of dimeric Zn[Me(2)NC[N(i)Pr](2)]Me 15 with Mes(2)BOH, which yielded Zn[Me(2)NC[N(i)Pr](2)][OBMes(2)].Me(2)NC[N(i)Pr][NH(i)Pr] 16 as a result of protonation at the guanidine ligand in addition to the Zn-Me bond.     

BonDNet: a graph neural network for the prediction of bond dissociation energies for charged molecules

A broad collection of technologies, including e.g. drug metabolism, biofuel combustion, photochemical decontamination of water, and interfacial passivation in energy production/storage systems rely on chemical processes that involve bond-breaking molecular reactions. In this context, a fundamental thermodynamic property of interest is the bond dissociation energy (BDE) which measures the strength of a chemical bond. Fast and accurate prediction of BDEs for arbitrary molecules would lay the groundwork for data-driven projections of complex reaction cascades and hence a deeper understanding of these critical chemical processes and, ultimately, how to reverse design them. In this paper, we propose a chemically inspired graph neural network machine learning model, BonDNet, for the rapid and accurate prediction of BDEs. BonDNet maps the difference between the molecular representations of the reactants and products to the reaction BDE. Because of the use of this difference representation and the introduction of global features, including molecular charge, it is the first machine learning model capable of predicting both homolytic and heterolytic BDEs for molecules of any charge. To test the model, we have constructed a dataset of both homolytic and heterolytic BDEs for neutral and charged (−1 and +1) molecules. BonDNet achieves a mean absolute error (MAE) of 0.022 eV for unseen test data, significantly below chemical accuracy (0.043 eV). Besides the ability to handle complex bond dissociation reactions that no previous model could consider, BonDNet distinguishes itself even in only predicting homolytic BDEs for neutral molecules; it achieves an MAE of 0.020 eV on the PubChem BDE dataset, a 20% improvement over the previous best performing model. We gain additional insight into the model''s predictions by analyzing the patterns in the features representing the molecules and the bond dissociation reactions, which are qualitatively consistent with chemical rules and intuition. BonDNet is just one application of our general approach to representing and learning chemical reactivity, and it could be easily extended to the prediction of other reaction properties in the future.

Prediction of bond dissociation energies for charged molecules with a graph neural network enabled by global molecular features and reaction difference features between products and reactants.