Insights on cyclophosphamide metabolism and anticancer mechanism of action: A computational study.

The oxazaphosphorine cyclophosphamide (CP) is a DNA-alkylating agent commonly used in cancer chemotherapy. This anticancer agent is administered as a prodrug activated by a liver cytochrome P450-catalyzed 4-hydroxylation reaction that yields the active, cytotoxic metabolite. The primary metabolite, 4-hydroxycyclophosphamide, equilibrates with the ring-open aldophosphamide that undergoes ß-elimination to yield the therapeutically active DNA cross-linking phosphoramide mustard and the byproduct acrolein. The present paper presents a DFT investigation of the different metabolic phases and an insight into the mechanism by which CP exerts its cytotoxic action. A detailed computational analysis of the energy profiles describing all the involved transformations and the mechanism of DNA alkylation is given with the aim to contribute to an increase of knowledge that, after more than 60 years of unsuccessful attempts, can lead to the design and development of a new generation of oxazaphosphorines.

Ab initio molecular dynamics of solvation effects and reactivity at the interface between water and ascorbic acid covered anatase TiO2 (101).

In this work, we present a detailed study of the interaction between ascorbic acid (L-asc) and anatase TiO2 (101) surface both in gas phase and in contact with water by using density functional theory and ab initio molecular dynamics simulations. In gas phase, L-asc strongly binds the TiO2 (101) surface as a dianion (L-asc2- ), adopting a bridging bidentate coordination mode (BB), with the two acid protons transferred to two surface 2-fold bridging oxygens (O2c). AIMD simulations show that the interaction between the organic ligand and the anatase surface is stable and comparable to the vacuum one despite the possible solvent effects and/or possible structural distortions of the ligand. In addition, during the AIMD simulations hydroxylation phenomena occur forming transient H3 O+ ions at the solid-liquid interface. For the first time, our results provide insight into the role of the ascorbic acid on the electronic properties of the TiO2 (101), the influence of the water environment on the ligand-surface interaction and the nature of the solid-liquid interface.

Catalytic regeneration of metal-hydrides from their corresponding metal-alkoxides via the hydroboration of carbonates to obtain methanol and diols.

Thorium complexes decorated with 5-, 6-, and 7-membered N-heterocyclic iminato ligands containing mesityl wingtip substitutions have been synthesized and fully characterized. These complexes were found to be efficient in the hydroboration of cyclic and linear organic carbonates with HBpin or 9-BBN promoting their decarbonylation and producing the corresponding boronated diols and methanol. In addition, the hydroboration of CO2 breaks the molecule into "CO" and "O" forming boronated methanol and pinBOBpin. Moreover, the demanding depolymerization of polycarbonates to the corresponding boronated diols and methanol opens the possibility of recycling polymers for energy sources. Increasing the core ring size of the ligands allows a better performance of the complexes. The reaction proceeds with high yields under mild reaction conditions, with low catalyst loading, and short reaction times, and shows a broad applicability scope. The reaction is achieved via the recycling of a high-energy Th-H moiety from a stable Th-OR motif. Experimental evidence and DFT calculations corroborate the formation of the thorium hydride species and the reduction of the carbonate with HBpin to the corresponding Bpin-protected alcohols and H3COBpin through the formate and acetal intermediates.

Gold complexes with remote-substituted amino N-heterocyclic carbenes.

The 4-RN-1,3-Ar2-imidazolium salt, R = iPr, tBu, Ar = Mes, Dipp, Mes = mesityl, Dipp = 2,6-bis-diisopropyl-phenyl was metalated by AuI at the C2-, C5- and 4-RN positions depending on the reactants and conditions employed; a rare direct rearrangement of a AuI aminide to an abnormal imidazol-5-ylidene AuI complex was also observed and based on a DFT study it may involve TfO- facilitated H+ transfer.

Effect of Indole-2-carboxylic Acid on the Self-Corrosion and Discharge Activity of Aluminum Alloy Anode in Alkaline Al-Air Battery.

Al-air battery has been regarded as a promising new energy source. However, the self-corrosion of aluminum anode leads to a loss of battery capacity and a decrease in battery longevity, limiting its commercial applications. Herein, indole-2-carboxylic acid (ICA) has been added to 4 M NaOH as a corrosion inhibitor. Its impact on the self-corrosion of aluminum alloy and the enhancement of the functionality of Al-air batteries at various concentrations have been investigated. X-ray photoelectron spectroscopy (XPS), attenuated total reflection Fourier transform infrared (ATR-FTIR) spectroscopy, atomic force microscopy (AFM), and scanning electron microscopy (SEM) techniques have been used to examine the compositional and morphological alterations of aluminum alloy surfaces. Electrochemical and hydrogen evolution tests showed that indole-2-carboxylic acid is an efficient corrosion inhibitor in alkaline solutions, and its impact grows with concentration. Our findings demonstrated that when the inhibitor concentration is 0.07 M, the inhibition efficiency is 54.0%, the anode utilization rises from 40.2% to 79.9%, the capacity density increases from 1197.6 to 2380.9 mAh g-1, and the energy density increases from 1469.9 to 2951.8 Wh kg-1. In addition, theoretical calculations have been performed to support the experimental results.

Disrupting Cu trafficking as a potential therapy for cancer.

Copper ions play a crucial role in various cellular biological processes. However, these copper ions can also lead to toxicity when their concentration is not controlled by a sophisticated copper-trafficking system. Copper dys-homeostasis has been linked to a variety of diseases, including neurodegeneration and cancer. Therefore, manipulating Cu-trafficking to trigger selective cancer cell death may be a viable strategy with therapeutic benefit. By exploiting combined in silico and experimental strategies, we identified small peptides able to bind Atox1 and metal-binding domains 3-4 of ATP7B proteins. We found that these peptides reduced the proliferation of cancer cells owing to increased cellular copper ions concentration. These outcomes support the idea of harming copper trafficking as an opportunity for devising novel anti-cancer therapies.

Analysis of the Fragmentation Pathways for the Collision-Induced Dissociation of Protonated Cyclophosphamide: A Mass Spectrometry and Quantum Mechanical Study.

Cyclophosphamide is a well-known anticancer agent acting by means of DNA alkylation. Associated with its tumor selectivity, it also possesses a wide spectrum of toxicities. As the requirement of metabolic activation before cyclophosphamide exerts either its therapeutic or toxic effects is well recognized, research aiming at elucidating the pathways that lead to the activation of this drug is of key importance. This has created the necessity for developing an effective analytical method for detecting cyclophosphamide and its breakdown products. In this paper, an Acquity TQ tandem quadrupole mass spectrometer equipped with electrospray ionization in positive-ion mode was employed for detecting cyclophosphamide in its protonated form. The full-scan mass spectrum of cyclophosphamide shows two ion clusters displaying the characteristic isotopic pattern of two chlorine atoms and assigned as sodiated cyclophosphamide, [CP + Na]+, and protonated cyclophosphamide, [CP + H]+ or PCP. With the aid of quantum mechanical DFT calculation, free energy differences in the gas phase among PCP protomers were computed with respect to the most stable protomer being protonated on the 2-oxide oxygen of the 1,3,2-oxazaphosphorine-2-oxide ring. In addition, the interconversion mechanisms among the different protomers were also proposed by intercepting the corresponding transition states in the gas phase. Collision-induced dissociation (CID) of PCP generated six characteristic product ions. Fragmentation mechanisms were proposed and supported by computation. The calculated energy barriers for all of the located transition states were found to be accessible under the reported experimental conditions.

A green route to platinum N-heterocyclic carbene complexes: mechanism and expanded scope.

A sustainable and facile weak-base synthetic route to platinum N-heterocyclic carbene (NHC) complexes is disclosed. The mechanism of this reaction is also elucidated via experimental and computational investigations. This straightforward protocol is then used for the synthesis of novel Pt(II)-NHC complexes and its utility is further explored to access key Pt(0)-NHC precatalysts.

DFT Investigation of Substitutional and Interstitial Nitrogen-Doping Effects on a ZnO(100)-TiO2(101) Heterojunction.

Density Functional Theory (DFT) calculations have been performed to investigate the structural and electronic properties of the ZnO(wurtzite)-ATiO2(anatase) heterojunction in the absence and presence of substitutional, interstitial nitrogen (N) doping and oxygen vacancies (OV). We report a detailed study of the interactions between the two nonpolar ZnO and TiO2 surfaces and on the role of N-doping and oxygen vacancies, which are decisive for improving the photocatalytic activity of the heterojunction. Our calculations show that substitutional N-doping is favored in the ATiO2 portion, whereas the interstitial one is favored in the ZnO region of the interface. Both substitutional and interstitial N-doped sites (i) induce gap states that act as deep electronic traps improving the charge separation and delaying electron-hole recombination, (ii) facilitate the OV formation causing a decrease in the formation energy (E FORM), and (iii) do not affect the band alignment when compared to the undoped analogue system. The presented results shed light on the N-doping effect on the electronic structure of the ZnO(100)-TiO2(101) heterojunction and how N-doping improves its photocatalytic properties.

Straightforward synthesis of [Cu(NHC)(alkynyl)] and [Cu(NHC)(thiolato)] complexes (NHC = N-heterocyclic carbene).

Synthetic access to monomeric copper-alkynyl and copper-thiolato complexes of the type [(NHC)Cu(R)] (R = alkynyl or thiolato) using a weak base approach is reported. All reported reactions proceed under mild conditions in air and in environmentally acceptable solvents. The novel complexes are fully characterized and single crystal X-ray analyses unambiguously establish the atom connectivity in these mononuclear complexes. The importance of the supporting NHC ligand's steric properties in stabilizing mononuclear complexes is discussed.

A Boron-Containing Compound Acting on Multiple Targets Against Alzheimer's Disease. Insights from Ab Initio and Molecular Dynamics Simulations.

Given the multifactorial nature and pathogenesis of Alzheimer's disease, therapeutic strategies are addressed to combine the benefits of every single-target drug into a sole molecule. Quantum mechanics and molecular dynamics (MD) methods were employed here to investigate the multitarget action of a boron-containing compound against Alzheimer's disease. The antioxidant activity as a radical scavenger and metal chelator was explored by means of density functional theory. The most plausible radical scavenger mechanisms, which are hydrogen transfer, radical adduct formation, and single-electron transfer in aqueous and lipid environments, were fully examined. Metal chelation ability was investigated by considering the complexation of Cu(II) ion, one of the metals that in excess can even catalyze the ß-amyloid (Aß) aggregation. The most probable complexes in the physiological environment were identified by considering both the stabilization energy and the shift of the λmax induced by the complexation. The excellent capability to counteract Aß aggregation was explored by performing MD simulations on protein-ligand adducts, and the activity was compared with that of curcumin, chosen as a reference.

All-Atom Simulations Disclose How Cytochrome Reductase Reshapes the Substrate Access/Egress Routes of Its Partner CYP450s.

Cytochromes P450 enzymes (CYP450s) promote the oxidative metabolism of a variety of substrates via the electrons supplied by the cytochrome P450 reductase (CPR) and upon formation of a CPR/CYP450 adduct. In spite of the pivotal regulatory importance of this process, the impact of CPR binding on the functional properties of its partner CYP450 remains elusive. By performing multiple microsecond-long all-atom molecular dynamics simulations of a 520 000-atom model of a CPR/CYP450 adduct embedded in a membrane mimic, we disclose the molecular terms for their interactions, considering the aromatase (HA) enzyme as a proxy of the CYP450 family. Our study strikingly unveils that CPR binding alters HA's functional motions, bolstering a change in the shape and type of the channels traveled by substrates/products during their access/egress to/from the enzyme's active site. Our outcomes unprecedentedly contribute to extricate the many entangled facets of the CYP450 metabolon, redrafting its intricate panorama from an atomic-level perspective.

Unraveling the Impact of Cysteine-to-Serine Mutations on the Structural and Functional Properties of Cu(I)-Binding Proteins.

Appropriate maintenance of Cu(I) homeostasis is an essential requirement for proper cell function because its misregulation induces the onset of major human diseases and mortality. For this reason, several research efforts have been devoted to dissecting the inner working mechanism of Cu(I)-binding proteins and transporters. A commonly adopted strategy relies on mutations of cysteine residues, for which Cu(I) has an exquisite complementarity, to serines. Nevertheless, in spite of the similarity between these two amino acids, the structural and functional impact of serine mutations on Cu(I)-binding biomolecules remains unclear. Here, we applied various biochemical and biophysical methods, together with all-atom simulations, to investigate the effect of these mutations on the stability, structure, and aggregation propensity of Cu(I)-binding proteins, as well as their interaction with specific partner proteins. Among Cu(I)-binding biomolecules, we focused on the eukaryotic Atox1-ATP7B system, and the prokaryotic CueR metalloregulator. Our results reveal that proteins containing cysteine-to-serine mutations can still bind Cu(I) ions; however, this alters their stability and aggregation propensity. These results contribute to deciphering the critical biological principles underlying the regulatory mechanism of the in-cell Cu(I) concentration, and provide a basis for interpreting future studies that will take advantage of cysteine-to-serine mutations in Cu(I)-binding systems.

Recent advances in computational design of potent aromatase inhibitors: open-eye on endocrine-resistant breast cancers.

Introduction: The vast majority of breast cancers (BC) are estrogen receptor positive (ER+). The most effective treatments to fight this BC type rely on estrogen deprivation therapy, by inhibiting the aromatase enzyme, which performs estrogen biosynthesis, or on blocking the estrogens signaling path via modulating/degrading the estrogen's specific nuclear receptor (estrogen receptor-α, ERα). While being effective at early disease stage, patients treated with aromatase inhibitors (AIs) may acquire resistance and often relapse after prolonged therapies. Areas covered: In this compendium, after an overview of the historical development of the AIs currently in clinical use, and of the computational tools which were used to identify them, the authors focus on current advances in obtaining innovative inhibitors via molecular simulations. These inhibitors may help prevent or delay relapse to AIs. Expert opinion: BC remains the most diagnosed and the leading cause of death in women. In spite of the success of the adjuvant endocrine therapy, which has enormously prolonged woman's survival rate, the increasing emergence of the resistance phenomena calls for the development of novel approaches and drugs to fight it. The discovery of the last generation of AIs dates back to two decades ago, underlying a paucity of research efforts.

The pivotal role of MBD4-ATP7B in the human Cu(i) excretion path as revealed by EPR experiments and all-atom simulations.

Copper's essentiality and toxicity require a meticulous mechanism for its acquisition, cellular distribution and excretion, which remains hitherto elusive. Herein, we jointly employed electron paramagnetic resonance spectroscopy and all-atom simulations to resolve the copper trafficking mechanism in humans considering the route travelled by Cu(i) from the metallochaperone Atox1 to the metal binding domains 3 and 4 of ATP7B. Our study shows that Cu(i) in the final part of its extraction pathway is most likely mediated by binding of Atox1 monomer to MBD4 of ATP7B. This interaction takes place through weak metal-stabilized protein-protein interactions.

Post-Translational Regulation of CYP450s Metabolism As Revealed by All-Atoms Simulations of the Aromatase Enzyme.

Phosphorylation by kinases enzymes is a widespread regulatory mechanism able of rapidly altering the function of target proteins. Among these are cytochrome P450s (CYP450), a superfamily of enzymes performing the oxidation of endogenous and exogenous substrates thanks to the electron supply of a redox partner. In spite of its pivotal role, the molecular mechanism by which phosphorylation modulates CYP450s metabolism remains elusive. Here by performing microsecond-long all-atom molecular dynamics simulations, we disclose how phosphorylation regulates estrogen biosynthesis, catalyzed by the Human Aromatase (HA) enzyme. Namely, we unprecedentedly propose that HA phosphorylation at Y361 markedly stabilizes its adduct with the flavin mononucleotide domain of CYP450s reductase (CPR), the redox partner of microsomal CYP450s, and a variety of other proteins. With CPR present at physiological conditions in a limiting ratio with respect to its multiple oxidative partners, the enhanced stability of the CPR/HA adduct may favor HA in the competition with the other proteins requiring CPR's electron supply, ultimately facilitating the electron transfer and estrogen biosynthesis. As a result, our work elucidates at atomic-level the post-translational regulation of CYP450s catalysis. Given the potential for rational clinical management of diseases associated with steroid metabolism disorders, unraveling this mechanism is of utmost importance, and raises the intriguing perspective of exploiting this knowledge to devise novel therapies.

Hydrolysis in Acidic Environment and Degradation of Satraplatin: A Joint Experimental and Theoretical Investigation.

For the synthesis and selection of active platinum-based anticancer drugs that perform better than cisplatin and its analogues, six-coordinate octahedral Pt(IV) complexes appear to be promising candidates as, being kinetically more inert and more resistant to ligand substitution than four-coordinate Pt(II) centers, they are able to minimize unwanted side reactions with biomolecules prior to DNA binding. Due to their kinetic inertness, Pt(IV) complexes have also been exploited to bypass inconvenient intravenous administration. The most prominent example is satraplatin (Sat.) which is the first platinum antineoplastic agent reported to have oral activity. The present paper deals with a theoretical DFT investigation of the influence that the acidity of the biological environment can have on the activity of satraplatin and analogous octahedral Pt(IV) complexes having two carboxylates as axial ligands. Moreover, here the outcomes of a joint electrospray ionization mass spectrometry and DFT investigation of the fragmentation pathways of the protonated satraplatin are reported. Calculations show that the simulated acidic environment has an important impact on the satraplatin reactivity causing a significant lowering of the barrier that is necessary to overcome for the hydrolysis of the first acetate ligand to occur. Data from electrospray ionization mass spectrometry, 1H NMR, and potentiometric experiments strongly suggest that the loss of CH3COOH from the protonated satraplatin ion [Sat. + H]+ takes place almost immediately upon dissolution of satraplatin in methanol-water, D2O, and water solutions, respectively, at room temperature.

Investigation of the Inertness to Hydrolysis of Platinum(IV) Prodrugs.

Platinum(IV) complexes are an important class of compounds that can act as prodrugs, and due to their inertness, if correctly designed, they could have low toxicity outside the cancer cell and improve the pharmacological properties of the platinum(II) anticancer agents that are currently used in the clinic. Because of the efforts that are concentrated on the use of axial ligands able to control the reduction potentials, lipophilicity, charge, selectivity, targeting, and cell uptake of the Pt(IV) complexes, we considered to be of interest to probe the inertness of such complexes that is assumed to be a fulfilled prerequisite. To this aim, a density functional theory computational analysis of the hydrolysis mechanism and the corresponding energy profiles for a series of Pt(IV) derivatives of cisplatin, carboplatin, and oxaliplatin with acetato, haloacetato, and chlorido ligands was performed to probe their stability in biological fluids. The heights of the barriers calculated along the hydrolysis pathways for the associative displacement of ligands both in axial and equatorial positions confirm that Pt(IV) complexes are, in general, more inert than the corresponding Pt(II) drugs even if inertness is lower than expected. Some exceptions exist, such as derivatives of oxaliplatin for the hydrolysis in equatorial position. The nature of the axial ligands influences the course of the hydrolysis reaction even if a decisive role is played by the ligands in equatorial positions. The mechanism of the aquation in axial position of cisplatin Pt(IV) derivative with two chlorido axial ligands assisted by Pt(II) cisplatin was elucidated, and the calculated activation energy confirms the catalytic role played by the Pt(II) complex.

DFT Investigation of the Mechanism of Action of Organoiridium(III) Complexes As Anticancer Agents.

The potential use of synthetic metal complexes able to catalyze chemical transformations in living organisms is currently attracting a great deal of attention. Recently, organometallic ruthenium and iridium complexes have revealed an unexpected ability to modulate the redox status of cancer cells. In particular, half-sandwich organoiridium(III) cyclopentadienyl complexes of general formula [(η(5)-Cp(x))Ir(III)(X(∧)Y)Z](0/+), where Cp(x) = Cp*, Cp(xph) (tetramethyl-(phenyl)cyclopentadienyl) or Cp(xbiph) (tetramethyl(biphenyl)-cyclopentadienyl), X(∧)Y = bidentate ligand with nitrogen, oxygen, and/or carbon donor atoms, and Z = Cl, H2O, or pyridine (py) have shown promising antiproliferative activity toward cancer cells, higher potency than cisplatin, and a different mechanism of action due to the increase of the oxidative stress in cells. As such, complexes can belong to the class of DNA interacting compounds and attack on DNA can represent a secondary mechanism of action. We have explored here by means of density functional calculations (M06-L) and with the support of experimental observations for both [(η(5)-Cp(xbiph))Ir-(phpy) (Cl)], 1-Cl, and [(η(5)-Cp(xbiph))Ir-(phpy) (py)], 1-py, complexes the mechanistic aspects of the hydrolysis reaction, H2O2 ROS production by assisted hydride transfer from NADH to molecular oxygen, interaction with purine nucleobases adenine and guanine as well as gluthatione, that is highly abundant in cells, alongside the reaction mechanism for the oxidation of the formed sulfur-coordinated thiolate to the corresponding sulfenato complex. The comparison between kinetic and thermodynamic parameters calculated for all the involved processes shows that, according to the hypothesis based on experimental findings, the interaction with the tripeptide glutathione causes deactivation of 1-Cl, whereas 1-py, in both its aquated and nonaquated form, can induce cell apoptosis in a dual manner: DNA damage and H2O2 ROS production to increase oxidative stress.

Mass Spectrometric and Computational Investigation of the Protonated Carnosine-Carboplatin Complex Fragmentation.

Platinum(II)-based anticancer drugs are square-planar d(8) complexes that, activated by hydrolysis, cause cancer cell death by binding to nuclear DNA and distorting its structure. For that reason, interactions of platinum anticancer drugs with DNA have been extensively investigated, aiming at disentangling the mechanism of action and toxicity. Less attention, however, has been devoted to the formation of adducts between platinum drugs with biological ligands other than DNA. These adducts can cause the loss and deactivation of the drug before it arrives at the ultimate target and are also thought to contribute to the drug's toxicity. Here are reported the outcomes of electrospray ionization mass spectrometry experiments and density functional theory (DFT) computations carried out to investigate the fragmentation pathways of the protonated carnosine-carboplatin complex, [Carnosine + CarbPt + H](+). DFT calculations at the B3LYP/LANL2DZ level employed to probe fragmentation mechanisms account for all experimental data. Because of the relative rigidity of the structure of the most stable 1A conformer, stabilized by three strong hydrogen bonds, the first step of all of the examined fragmentation pathways is the interconversion of the 1A conformer into the less stable structure 1B. Formation of the [Carnosine + H](+) fragment from the precursor ion, [Carnosine + CarbPt + H](+), is calculated to be the lowest-energy process. At slightly higher energies, the loss of two amino groups is observed to produce the [Carnosine + (CarbPt - NH3) + H](+) and [Carnosine + (CarbPt - 2NH3) + H](+) ions. At significantly higher energies, the loss of CO2 occurs, yielding the final [Carnosine + (CarbPt - NH3) - CO2 + H](+) and [Carnosine + (CarbPt - 2NH3) - CO2 + H](+) products. Formation of the [CarbPt + H](+) fragment from [Carnosine + CarbPt + H](+), even if not hampered by a high activation barrier, is calculated to be very unfavorable from a thermodynamic point of view.