Bioreductive Alkylation Research Articles

Study of the cytotoxic effects of 2,5-diaziridinyl-3,6-dimethyl-1,4-benzoquinone (MeDZQ) in mouse hepatoma cells.

A number of quinones have been shown to be efficient anticancer agents. However, some mechanisms of their action, in particular cell signaling are not well understood. The aim of this study was to partly fill this gap by characterizing the mode of cytotoxicity of 2,5-diaziridinyl-3,6-dimethyl-1,4-benzoquinone (MeDZQ) in malignant mouse hepatoma cells (MH-22A) with regard to the expression and activation of main molecules in MAPK cell signaling pathway. The study revealed unequal roles of MAP kinases in MeDZQ-induced cell death: the compound did not induce significant changes in ERK expression or its phosphorylation; JNK appeared to be responsible for cell survival, however, p38 kinase was shown to be involved in cell death. In order to assess the enzymatic activation mechanisms responsible for the action of MeDZQ, we have also found that the antioxidant N,N'-diphenyl-p-phenylene diamine, the iron-chelating agent desferrioxamine, and DT-diaphorase inhibitor, dicoumarol, partly protected the cells from MeDZQ cytotoxicity. It points to parallel oxidative stress and bioreductive alkylation modes of the cytotoxicity of MeDZQ.

Design and synthesis of biotinylated dimethylation of alkannin oxime derivatives

DMAKO-05, a novel dimethylation of alkannin oxime derivative, exhibits remarkable anticancer activity as well as excellent cellular selectivity and thus has been considered as a promising antineoplastic agent for colorectal carcinoma and melanoma. However, its potent cytotoxicity is not closely associated with reactive oxygen species (ROS) and bioreductive alkylation. Its specific antitumor target(s) has still remained elusive. To recognize the molecular target(s) of DMAKO-05 and its analogs, four biotinylated DMAKO derivatives were designed and prepared. The biotin moiety was successfully introduced in the molecule through a modified Mitsunobu reaction, which kept its anticancer activity. Moreover, the cell-based investigation demonstrated that replacement of the linker C4 chain with another alkyl chain (C6 or C8) gave rise to the enhancement of cytotoxicity. Among these biotinyl derivatives, both compound 16 and 8c exhibited more potent anticancer activity than DMAKO-05 against MCF-7 cells and were comparatively effective to alkannin toward HCT-15 cells. As expected, they might be thought as ideal chemical probes. Collectively, our present work could provide an available approach for the identification of the potential antineoplastic target(s) of DMAKO derivatives.

Single-electron reduction of quinone and nitroaromatic xenobiotics by recombinant rat neuronal nitric oxide synthase.

We examined the kinetics of single-electron reduction of a large number of structurally diverse quinones and nitroaromatic compounds, including a number of antitumour and antiparasitic drugs, and nitroaromatic explosives by recombinant rat neuronal nitric oxide synthase (nNOS, EC 1.14.13.39), aiming to characterize the role of nNOS in the oxidative stress-type cytotoxicity of the above compounds. The steady-state second-order rate constants (kcat/Km) of reduction of the quinones and nitroaromatics varied from 10² M⁻¹s⁻¹ to 10⁶ M⁻¹s⁻¹, and increased with an increase in their single-electron reduction potentials (E¹₇). The presence of Ca²⁺/calmodulin enhanced the reactivity of nNOS. These reactions were consistent with an 'outer sphere' electron-transfer mechanism, considering the FMNH∙/FMNH₂ couple of nNOS as the most reactive reduced enzyme form. An analysis of the reactions of nNOS within the 'outer sphere' electron-transfer mechanism gave the approximate values of the distance of electron transfer, 0.39-0.47 nm, which are consistent with the crystal structure of the reductase domain of nNOS. On the other hand, at low oxygen concentrations ([O₂] = 40-50 μM), nNOS performs a net two-electron reduction of quinones and nitroaromatics. This implies that NOS may in part be responsible for the bioreductive alkylation by two-electron reduced forms of antitumour aziridinyl-substituted quinones under a modest hypoxia.

The Bioreductive Alkylation of DNA by Kalafungin: A Theoretical Investigation

The thermochemical cascades for the bioreductive alkylation of DNA by kalafungin were calculated using density functional theory (DFT). Guanine (G) was used as a model nucleotide. According to the calculations both one- and two-electron reduction of kalafungin is possible in vivo. Furthermore, a clear pathway was found for both mono- and bis-alkylations of G with the former favoured. Alkylation at C-8 position of G is considerably more exothermic than on the N2-exocyclic amine. In the absence of experimentally identified adduct structures of kalafungin, the results presented here support the idea that this compound readily forms covalent bonds with DNA resulting in pro-mutagenic lesions.

A few considerations on quinones as antitumor agents

Although a number of quinones represent important antitumor drugs, the role of the quinone ring in this activity does not seem to be established. Active quinones are relatively large molecules which either possess a large quinoid system, in particular that of anthraquinone, or contain the benzoquinone ring imbedded in a larger molecular skeleton. Simple small quinones have little if any clinical utility. The cellular target of quinones is DNA. Drugs having a large anthraquinone ring interact with DNA essentially by intercalation, small quinones imbedded in a larger molecular framework react with DNA essentially by covalent bond formation which, however, does not involve the quinone ring itself. The establishment of the adducts seems to necessitate the activation of the drugs, which may be produced by protonation or, better, by bioreductive alkylation. There does not seem to be any straightforward correlation between the antitumor potency and the one-electron reduction potential of the quinone–semiquinone couples. The generation of free radicals upon the reoxidation of the hydroquinone moiety of DNA-bound quinones may cause single-strand breaks in DNA, but these are also produced by intercalating drugs which cannot undergo bioreductive alkylation. Analogs of active quinones which lack the quinone system may also manifest antitumor activity. The determination of the specific significance of the quinone system in antitumor activity needs thus further investigation.

Pyranonaphthoquinone Lactones: A New Class of AKT Selective Kinase Inhibitors Alkylate a Regulatory Loop Cysteine

The naturally occurring pyranonaphthoquinone (PNQ) antibiotic lactoquinomycin and related aglycones were found to be selective inhibitors of the serine-threonine kinase AKT. A set of synthetic PNQs were prepared and a minimum active feature set and preliminary SAR were determined. PNQ lactones inhibit the proliferation of human tumor cell lines containing constitutively activated AKT and show expected effects on cellular biomarkers. Biochemical data are presented supporting a proposed bioreductive alkylation mechanism of action.

The molecular mechanisms involved in the cytotoxicity of alkannin derivatives

In order to better understand the molecular aspects of the cytotoxic action mechanisms, the cytotoxicity of alkannin derivatives, 1– 10, on five human tumor cell lines were examined and their standard redox potentials in aprotic medium were tested by means of cyclic voltammetry. It was suggested that the oxidative potential is closely related to the cytotoxicity. The more negative the oxidative potential of the hydroquinones, the higher cytotoxicity of these derivatives. The results of the compounds 5, 7, 9 and 10 with bad leaving groups, have higher cytotoxic action is not agreed with the bioreductive alkylation mechanism of quinones. It indicates that the molecular mechanism involving cytotoxicity of alkannin derivatives may favor the mechanism of production of reactive oxygen.

Some Applications of Electrochemistry in Biomedical Chemistry. Emphasis on the Correlation of Electrochemical and Bioactive Properties

This review summarises some of the more relevant achievements in the correlation between electrochemical processes and parameters and bioactive properties, mainly related to cancer and tropical diseases. Despite the broad range of possibilities and the complexity of cell/tissue/extracell chemistry, it is possible to rationalise the role of electrochemistry in few basic theoretical frameworks, mainly, the one based on electron transfer-oxidative stress and in situ generation of toxic species, other than the reactive oxygen species; interaction with endobiotics, with emphasis on bioreductive alkylation and replacement of endobiotics with function in biological redox reactions. The use of electrochemical methods to obtain relevant informations about drugs' mechanism of action and analysis of cellular events is also presented. Electrochemical methods and/or parameters play essential but not absolute roles.

Naphthazarin derivatives: synthesis, cytotoxic mechanism and evaluation of antitumor activity.

The rate of the GSH conjugate formation, the inhibition of DNA topoisomerase-I and the cytotoxic activity against L1210 cells of the naphthoquinones showed the same order; 5,8-dimethoxy-1,4-naphthoquinone (DMNQ) > 6-(1-hydroxyethyl)-DMNQ > 2-(1-hydroxyethyl)-DMNQ; the steric hindrance of the substituents, particularly 2-substutuent, in reacting with cellular nucleophiles must be the main cause for lowering the bioactivities. Acetylation of 2-(1-hydroxyethyl)-DMNQ producing 2-(acetyloxyethyl)-DMNQ potentiated the bioactivities; 2-(1-hydroxyethyl)-DMNQ did not react with GSH and the enzyme, and showed ED50 of 0.680 microgram/ml, whereas the values of 2-(1-acetyloxyethyl)-DMNQ were the conjugate formation of 0.14 microM, IC50 value of 81 microM for the enzyme inhibition and ED50 of 0.146 microgram/ml for the cytotoxcity. Furthermore, the acetylation 2-(1-hydroxyethyl)-DMNQ (T/C, 119%) enhanced the T/C values for the mice bearing S-180 tumor [T/C of 2-(1-acetyloxyethyl)-DMNQ, 276%]. It was assumed that the difference in bioactivities ensued by acetylation was based on the mechanism of the so-called bioreductive alkylation.

Electrochemistry of potentially bioreductive alkylating quinones. Part 4. Qualitative and quantitative structure‐activity relationships of aziridinylquinones

AbstractThe concept of bioreductive alkylation as a mechanism of action of aziridinylquinoid anticancer agents has been investigated. The influence of quinone substituents on quinone reduction, on protonation of the aziridines prior to and following quinone reduction and on partitioning properties of the compound was examined. Parameters obtained from a combined electrochemical, chemical‐stability and lipophilicity study describing these processes were determined and correlated quantitatively in a Hansch‐type QSAR study with biological data obtained from three experimental tumor models. Poor quantitative correlations between cytotoxicity in a L1210 clonogenic assay and the parameters were obtained. Good linear relationships, however, between antitumor activity in vivo (vs. L1210 leukemic mice and vs. B16 melanoma‐bearing mice) and the lipophilic properties of the quinone were found. These relationships, showing a negative correlation between antitumor activity and lipophilicity, can be used to predict the activity of new, unknown compounds. No trend was evident between antitumor activity and other parameters, although some indications for potential importance of electronic and steric properties of the substituents and of their ability to form hydrogen bonds were found.

Electrochemistry of potentially bioreductive alkylating quinones: Part 3. Quantitative structure-electrochemistry relationships of aziridinylquinones

The concept of bioreductive alkylation as a mechanism of action of aziridinylquinoid anticancer agents has been investigated by the use of electrochemical techniques. Properly substituted aziridinylquinones are activated by an electrochemical step (reduction of the quinone function), followed by protonation of the aziridinyl moiety to the alkylating species. The influence of substitution on quinone reduction, on protonation and on subsequent opening of the aziridines (prior to and after quinone reduction) has been examined. A series of mono- and poly(1-aziridinyl)-quinones has been synthesized and analyzed by direct current (d.c.) polarography. The half-wave potential ( E 1 2 value of the quinone reduction and the p K red and p K red2 (reflecting the ease of protonation at the mercury electrode of one and two aziridinyl rings, respectively) were used in a Hammett type QSAR analysis. A linear relationship between E 1 2 and the electronic substituent constant σ p was obtained for simple quinones. Deviations from linearity were observed, due to steric and/or resonance interactions which influence quinone reduction, with amino- and halogen-substituted quinones. Unknown σ p values could be calculated. Relationship between p K red2 and some physical-chemical parameters show that electronic and steric properties of quinone substituents affect p K red2. In addition, the formation of a hydrogen bond between the quinone substituent and the adjacent aziridinyl ring (which may thwart aziridine protonation) and the presence of a methyl substituent at position 2 of the aziridine (which facilitates protonation) are of importance. Results of this study have lead to a better knowledge of the individual substituents with respect to their electronic and steric influences on quinone reduction and aziridine protonation, which may be of importance if these processes play a decisive role in cytostatic activity as well as toxicity.

ChemInform Abstract: Pulse Radiolytic Kinetic Study of the Decay of α‐Methyl‐ Substituted Benzoquinone Radical Anions: A Possible Mechanistic Model for Bioreductive Alkylation.

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Pulse radiolytic kinetic study of the decay of .alpha.-methyl-substituted benzoquinone radical anions: a possible mechanistic model for bioreductive alkylation

Pulse radiolysis and transient absorption spectroscopy were used to kinetically characterize the decay of several α-methyl-substituted 1,4-benzoquinone radical anions. Electron attachment rates, absorption spectra, extinction coefficients, and pK a ' s for the reduced quinones are very similar regardless of substituent, but the decay kinetics are dependent on both substituent and pH. The semiquinone radical anions disproportionate at pH 7 regardless of substituent. At pH 3, the radical anion is readily protonated, and in the phenoxy- and methoxy-substituted models reductive elimination becomes competitive with disproportionation. The involvement of an intramolecular H-bonded intermediate is proposed in the formation of the quinone methide. The existence of such an intermediate is supported by theoretical results (AM1 calculations)

Mitomycin C

The mitomycins are antitumor antibiotics that are under investigation now for more than 30 years. Mitomycin C (MMC) is the best investigated subtype. It serves as a prototype for drugs with bioreductive alkylation, which is a unique feature of this class. MMC is mainly active under anaerobic circumstances. The pharmacokinetics are linear in a two-compartment model. The main toxicities of MMC are thrombocytopenia and leucocytopenia. Rare but severe side effects are a hemolytic uremic syndrome, pneumonitis and cardiac failure. MMC has a wide clinical antitumor spectrum with efficacy in various tumor types such as gastric cancer, pancreatic cancer, breast cancer, non-small cell lung cancer, cervical cancer, prostate cancer and bladder cancer. Still, the above mentioned side effects prevent a more widespread use. The most important features of the drug will be reviewed.

Bioreductive alkylation as a trigger for toxic effects of dynemicin

The endiyne derivative, dynemicin, is postulated to undergo bioreductive cis ring opening of a benzylic epoxide and activate the endiyne unit for Bergman rearrangement to an arene-1,4-diyl.

Mechanism(s) of bioreductive activation: The example of diaziquinone (AZQ)

Bioreduction in the activation of diaziquone (2,5-diaziridinyl-3,6-bis (carboethoxyamino)-1,4-benzoquinone) has been investigated by exploring its reduction by whole cells, rat liver microsomes and purified enzymes. The mechanism of bioreduction was further investigated by exploring the chemical and electrochemical reduction of diaziquione as well as its photochemistry. Reduced diaziquone (be several means) was then tested for activity against parent compound. It appears that reduced diaziquone in most cases is more active than the oxidized form. Diaziquone redox cycles, but it is easily reduced to the hydroquinone which oxidizes to the semiquinone yielding free radicals under aerobiosis. The most probable mechanism of action is that of bioreductive alkylation where the alkylating aziridines are protonated after reduction facilitating the opening of the aziridine rings and thus alkylation.

53 Bioreductive alkylation

53 Bioreductive alkylation

Bioactivation as a model for drug design bioreductive alkylation.

Bioactivation as a model for drug design bioreductive alkylation.