Nitro Analogue Research Articles

Influence of a Basic Side Chain on the Properties of Hypoxia-Selective Nitro Analogues of the Duocarmycins: Demonstration of Substantial Anticancer Activity in Combination with Irradiation or Chemotherapy.

A new series of nitro analogues of the duocarmycins was prepared and evaluated for hypoxia-selective anticancer activity. The compounds incorporate 13 different amine-containing side chains designed to bind in the minor groove of DNA while spanning a wide range of base strength from pKa 9.64 to 5.24. The most favorable in vitro properties were associated with strongly basic side chains, but the greatest in vivo antitumor activity was found for compounds containing a weakly basic morpholine. This applies to single-agent activity and for activity in combination with irradiation or chemotherapy (gemcitabine or docetaxel). In combination with a single dose of γ irradiation 50 at 42 μmol/kg eliminated detectable clonogens in some SiHa cervical carcinoma xenografts, and in combination with gemcitabine using a well-tolerated multidose schedule, the same compound caused regression of all treated A2780 ovarian tumor xenografts. In the latter experiment, three of seven animals receiving the combination treatment were completely tumor free at day 100.

Why does mutation of Gln61 in Ras by the nitro analog NGln maintain activity of Ras-GAP in hydrolysis of guanosine triphosphate?

Interpretation of the experiments showing that the Ras-GAP protein complex maintains activity in guanosine triphosphate (GTP) hydrolysis upon replacement of Glu61 in Ras with its unnatural nitro analog, NGln, is an important issue for understanding details of chemical transformations at the enzyme active site. By using molecular modeling we demonstrate that both glutamine and its nitro analog in the aci-nitro form participate in the reaction of GTP hydrolysis at the stages of proton transfer and formation of inorganic phosphate. The computed structures and the energy profiles for the complete pathway from the enzyme-substrate to enzyme-product complexes for the wild-type and mutated Ras suggest that the reaction mechanism is not affected by this mutation.

Mesophase stability in binary mixtures of monotropic nematogens

Binary phase diagrams were constructed from laterally substituted methyl azo/ester derivatives, namely 4-(4″-substituted phenylazo)-3-methyl phenyl-4″-alkoxy benzoates (Ina–d). In this group of compound the unsubstituted and chloro-substituted derivatives possess the nematic phase monotropically, while the nitro and methyl analogues are enantiotropically nematogenic. The binary phase diagrams constructed were made once from the monotropic nematogens with each other, and another with the enantiotropic nematogens. In both the cases enantiotropic nematic phase was observed that covers a wide range of composition. The mesophase behaviour of all binary mixtures was investigated by differential scanning calorimetry (DSC) and polarised light microscopy (PLM). The nematic phase was exhibited in all binary mixtures. Independent of the alkoxy chain length, the entropy change, ΔSN–I of the N–I transition of pure components was found to vary irregularly with the anisotropy of polarisability (∆αX) of the polar substituent, X.

Nitro and dinitroamino N-oxides of octaazaanthracene as high energy materials

The present study undertook the design of nitro and dinitroamino compounds from the skeleton of isomeric N-oxides of octaazanaphthalene, using computational methods to predict their degradation and explosive characteristics. The atom equivalent method was employed to evaluate the gas phase heats of formation of the designed species. Condensed phase heats of formation were also determined and found to be in the range of 220-286 kcal mol(-1). Crystal densities of all the designed molecules were calculated and found to be in the range of 1.91-1.98 g cm(-3). Detonation pressure (P) and detonation velocity (D) determined using the Kamlet-Jacobs equation showed that the performance of nitro-substituted compounds was comparable to that of RDX while that of dinitroamino compounds (P ≈ 43.4-43.7 GPa; D ≈ 9.6-9.7 km s(-1)) showed their superiority over HMX (P ≈ 39.3 GPa and D ≈ 9.10 km s(-1)). Impact sensitivity (h 50) of the designed molecules was compared with nitro- and nitramino-based commercial explosives on the basis of the available free space (∆V) per molecule in their crystal lattice estimated using wave function analysis. The study showed that dinitroamino compounds were more sensitive compared to their nitro analogs. Reactivity or chemical stability of the designed molecules were measured in terms of charge distribution, molecular electrostatic potential and frontier molecular orbital energy. The nitro compounds of N-oxides of octaazaanthracene were found to be more stable than their dinitroamino analogs.

Effects of the lateral substituent on the cubic phase formation of two analogous compounds, 4ʹ-n-hexadecyloxy-3ʹ-cyanobiphenyl-4-carboxylic acid (ACBC-16) and its 3ʹ-nitro compound (ANBC-16)

Two cubic (Cub) phase forming compounds, 4ʹ-n-hexadecyloxy-3ʹ-cyanobiphenyl-4-carboxylic acid (ACBC-16) and its 3′-nitro analogue (ANBC-16) were studied by infrared (IR) spectroscopy. In the temperature region of the Cub phase, the molar fraction of hydrogen-bonded COOH groups estimated for ACBC-16 was by ≈0.05 at maximum larger than that for ANBC-16 and the aromatic ring C═C stretching (ν(C═C)ring) band frequency of ACBC-16 was by 3 cm−1 lower than that of ANBC-16. The quantum chemical calculation at DFT/B3LYP level, on the one hand, showed no meaningful difference in the stabilisation energy for dimerisation and the ν(C═C)ring band frequency between the respective model compounds. These results can be ascribed to the different steric effects of the side groups; the CN group would make possible the close contact of neighbouring phenyl rings while the bulky NO2 group would not, giving slightly more stabilised dimerisation of ACBC-16 in the Cub phase than in ANBC-16.

Preparation and properties of clickable amino analogues of the duocarmycins: factors that affect the efficiency of their fluorescent labelling of DNA.

Herein we report the synthesis of three DNA-alkylating amino analogues of the duocarmycins that carry an alkyne functional group suitable for copper-catalysed click chemistry. The alkyne-containing substituents are connected via a side chain position which projects away from the minor groove, and have only a small effect on DNA alkylation and cytotoxicity. The efficiency of click reactions with fluorophore azides was studied using alkylated ctDNA by analysing the adenine adducts produced after thermal depurination. Click reactions "on DNA" were sensitive to steric effects (tether length to the alkyne) and, surprisingly, to the nature of the fluorophore azide. With the best combination of click partners and reagents, adducts could be detected in the nuclei of treated cells by microscopy or flow cytometry, provided that an appropriate detergent (Triton X-100 and not Tween 20) was used for permeabilisation. The method is sensitive enough to detect adducts at physiologically relevant concentrations, and could have application in the development of nitro analogues of the duocarmycins as hypoxia-activated anticancer prodrugs.

Phenyl-imidazolo-cytidine analogues: structure-photophysical activity relationship and ability to detect single DNA mismatch.

To expand the arsenal of fluorescent cytidine analogues for the detection of genetic material, we synthesized para-substituted phenyl-imidazolo-cytidine ((Ph)ImC) analogues 5a-g and established a relationship between their structure and fluorescence properties. These analogues were more emissive than cytidine (λem 398-420 nm, Φ 0.009-0.687), and excellent correlation was found between Φ of 5a-g and σp(-) of the substituent on the phenyl-imidazolo moiety (R(2) = 0.94). Calculations suggested that the dominant tautomer of (Ph)ImC in methanol solution is identical to that of cytidine. DFT calculations of the stable tautomer of selected (Ph)ImC analogues suggested a relationship between the HOMO-LUMO gap and Φ and explained the loss of fluorescence in the nitro analogue. Incorporation of the CF3-(Ph)ImdC analogue into a DNA probe resulted in 6-fold fluorescence quenching of the former. A 17-fold reduction of fluorescence was observed for the G-matched duplex vs ODN(CF3-(Ph)ImdC), while for A-mismatched duplex, only a 2-fold decrease was observed. Furthermore, since the quantum yield of ODN(CF3-(Ph)ImdC):ODN(G) was reduced 17-fold vs that of a single strand, whereas that of ODN(CF3-(Ph)ImdC):ORN(G) was reduced only 3.8-fold, ODN(CF3-(Ph)ImdC) appears to be a DNA-selective probe. We conclude that the ODN(CF3-(Ph)ImdC) probe, exhibiting emission sensitivity upon single nucleotide replacement, may be potentially useful for DNA single nucleotide polymorphism (SNP) typing.

Preclinical pharmacology of FL442, a novel nonsteroidal androgen receptor modulator

The preclinical profiles of two most potent compounds of our recently published cycloalkane[d]isoxazole pharmacophore-based androgen receptor (AR) modulators, FL442 (4-(3a,4,5,6,7,7a-hexahydro-benzo[d]isoxazol-3-yl)-2-(trifluoromethyl)benzonitrile) and its nitro analog FL425 (3-(4-nitro-3-(trifluoromethyl)phenyl)-3a,4,5,6,7,7a-hexahydrobenzo[d]isoxazole), were explored to evaluate their druggability for the treatment of AR dependent prostate cancer. The studies revealed that both compounds are selective to AR over other closely related steroid hormone receptors and that FL442 exhibits equal inhibition efficiency towards the androgen-responsive LNCaP prostate cancer cell line as the most widely used antiandrogen bicalutamide and the more recently discovered enzalutamide. Notably, FL442 maintains antiandrogenic activity with enzalutamide-activated AR mutant F876L. In contrast to bicalutamide, FL442 does not stimulate the VCaP prostate cancer cells which express elevated levels of the AR. Distribution analyses showed that [14CN]FL442 accumulates strongly in the mouse prostate. In spite of its low plasma concentration obtained by intraperitoneal administration, FL442 significantly inhibited LNCaP xenograft tumor growth. These findings provide a preclinical proof for FL442 as a promising AR targeted candidate for a further optimization.

Rules for the Design of Highly Fluorescent Nucleoside Probes: 8-(Substituted Cinnamyl)-Adenosine Analogues

Currently, there are no tools that can help the design of useful fluorescent analogues. Hence, we synthesized a series of 8-(substituted cinnamyl)-adenosine analogues, 5-17, and established a relationship between their structure and fluorescence properties. We attempted to find a correlation between maximum emission wavelengths (λ(em)) of 5-17 or their quantum yields (φ), and Hammett constants (σ(p) and σ(m)) of the substituent on the cinnamyl moiety. A linear correlation was observed at low-medium σ values, but not at high σ values (≥0.7). Next, we explored correlation between λ(em) and φ of 5-17 and computed HOMO and LUMO energy levels of fragments of 5-17, i.e., 8-vinyl 9-Me-adenine (fluorescent molecule), 18, and substituted toluene rings (fluoresence modulators), 19-30. High φ correlated with relatively close LUMO levels of 19-30 and 18 (-0.076 to -0.003 eV). The electron density of LUMO of nitro analogues 9 and 15 is localized on the aryl ring only, which explains their low φ. Calculation of HOMO-LUMO gap of 5-17 enables accurate prediction of the λ(abs) for a planned analogue, and LUMO levels of an aryl moiety vs 8-vinyl 9-Me-adenine, allows the prediction of high or low φ. These findings lay the ground for prediction of fluorescence properties of additional analogues having a similar structure.

Synthesis and antibacterial activities of N-substituted-glycinyl 1H-1,2,3-triazolyl oxazolidinones

A series of 1H-1,2,3-triazolyl piperazino oxazolidinone analogs with optionally varied glycinyl substitutions were synthesized and their antibacterial activity assessed against a panel of susceptible and resistant Gram-positive and selected Gram-negative bacteria including clinical isolates. The N-aroyl- and N-heteroaroyl-glycinyl (MIC: 0.06–4 μg/ml) derivatives were more potent than the N-acylglycinyl (2–8 μg/ml) derivatives against all Gram-positive bacteria tested. Nitro substitution on aryl and heteroaryl rings significantly enhanced activity against Gram-positive bacteria, as noted with the 3,5-dinitrobenzoyl (6m and 6n) and 5-nitro-2-furoyl (6u and 6v) derivatives with MIC ranges of and 0.25–0.5 and 0.06–0.5 μg/ml, respectively. These nitro analogs also showed more potent extended activity against Moraxella catarrhalis, with MICs ranges of 0.25–1 μg/ml, compared to linezolid (MIC: 8 μg/ml). Hence, the presence of the N-aroyl and/or N-heteroaroyl glycinyl structural motifs as spacer group could significantly enhance the antibacterial activities of 1H-1,2,3-triazolyl oxazolidinone class of compounds.

Evaluation of genotoxic effects of new molecules with possible trypanocidal activity for Chagas disease treatment.

Chagas disease is responsible for a large number of human infections and many are also at risk of infection. There is no effective drug for Chagas disease treatment. The Institute of Pharmaceutical Technology at Fiocruz, Brazil, has designed three nitro analogs of the nitroimidazole-thiadiazole, megazol: two triazole analogs PTAL 05-02 and PAMT 09 and a pyrazole analog PTAL 04-09. A set of Salmonella enterica serovar Typhimurium strains were used in the bacterial reverse mutation test (Ames test) to determine the mutagenicity and cytotoxicity of megazol and its nitro analogs. Megazol presented positive mutagenic activity at very low concentration, either with or without metabolic activation S9 mix. The mutagenic response of the analogs was detected at higher concentration than the lowest megazol concentration to yield mutagenic activity showing that new advances can be made to develop new analogs. The micronucleus test with rat macrophage cells was used in the genotoxic evaluation. The analogs were capable of inducing micronucleus formation and showed cytotoxic effects. PTAL 04-09 structural modifications might be better suitable for the design of promising new drugs candidate for Chagas' disease treatment.

P55 Selective turn-on fluorescent probes for detection of hydrogen sulfide in living cells

Background Hydrogen sulfide (H2S) is an important gasotransmitter that mediates a wide range of physiological effects. Although H2S is known for its toxicity and rotten egg smell, it has recently been established to play a role in neurological degradation and is correlated with central nervous system diseases, including Alzheimer’s disease, Down syndrome, and multiple sclerosis. H2S is produced endogenously by three enzymes: cystathionine-β-synthase (CBS), cystathionine-γ-lyase (CSE), and 3-mercaptopyruvate sulfur transferase (3MST). Although its biological genesis is well understood, the lack of techniques and methods for selectively monitoring H2S has led to a poor understanding of its multifaceted roles in biology. The development and use of selective fluorescent probes offers a technique to monitor endogenous small molecules. Our interest in developing chemical tools to study the biological roles of H2S has led us to synthesize two new compounds for H2S detection based on the 4-amino-1,8-naphthalimide fluorophore. Methods Hydrogen sulfide can be used to reduce nitro (RNO2) and azido (RN3) functional groups to the parent amine. Based on these chemical properties, we sought to design H2S-selective fluorescent probes in which H2S selectively reduces a nitro (HSN1) or azide (HSN2) analog of the parent naphthalimide probe. In addition to testing the response to H2S, the developed probes were tested under anaerobic conditions with various reactive species including cysteine, glutathione, α-lipoic acid, nitric oxide, hydrogen peroxide, sulfite, and thiosulfate. Results Treatment of a 5 μM solution of HSN1 or HSN2 in aqueous buffer (50 mM PIPES, 100 mM KCl, pH 7.4) with 100 equivalents of H2S resulted in a 15 and 60-fold fluorescence turn-on, respectively. Reduction of a nitro or azido analog of the parent naphthalimide probe generates the parent fluorescent amine, resulting in a large fluorescence turn-on. The developed fluorescent probes are able to selectively detect H2S in the presence of reactive sulfur, oxygen, nitrogen, species (RSONs) including cysteine, glutathione, α-lipoic acid, nitric oxide, hydrogen peroxide, sulfite, and thiosulfate. Furthermore, HSN2 remains highly selective for H2S even in the presence of 2000 equivalents (10 mM) of cysteine or glutathione. HSN1 and HSN2 were also used to detect exogenous H2S in live HeLa cells. After incubating HeLa cells with 5 μM HSN1 or HSN2, addition of 250 μM H2S resulted in a significant fluorescence enhancement by comparison to the control cells. Summary HSN1 and HSN2 are highly-selective fluorescent probes for H2S. Reactions of these probes with H2S resulted in a 15–60-fold fluorescence turn-on and were demonstrated to be viable H2S-detection methods in live HeLa cells.

Radiosynthesis and evaluation of an (18)F-labeled positron emission tomography (PET) radioligand for brain histamine subtype-3 receptors based on a nonimidazole 2-aminoethylbenzofuran chemotype.

A known chemotype of H(3) receptor ligand was explored for development of a radioligand for imaging brain histamine subtype 3 (H(3)) receptors in vivo with positron emission tomography (PET), namely nonimidazole 2-aminoethylbenzofurans, represented by the compound (R)-(2-(2-(2-methylpyrrolidin-1-yl)ethyl)benzofuran-5-yl)(4-fluorophenyl)methanone (9). Compound 9 was labeled with fluorine-18 (t(1/2) = 109.7 min) in high specific activity by treating the prepared nitro analogue (12) with cyclotron-produced [(18)F]fluoride ion. [(18)F]9 was studied with PET in mouse and in monkey after intravenous injection. [(18)F]9 showed favorable properties as a candidate PET radioligand, including moderately high brain uptake with a high proportion of H(3) receptor-specific signal in the absence of radiodefluorination. The nitro compound 12 was found to have even higher H(3) receptor affinity, indicating the potential of this chemotype for the development of further promising PET radioligands.

En route towards α-benzotriazoyl nitroso derivatives

A new class of geminally-substituted nitroso compounds, i.e. α-benzotriazoyl nitroso derivatives, is presented. These compounds display a rather different behavior than other related nitroso compounds bearing a geminal electron-withdrawing group. An unexpected and spontaneous oxidation to the nitro analog is observed in solution.

The effect of a bromide leaving group on the properties of nitro analogs of the duocarmycins as hypoxia-activated prodrugs and phosphate pre-prodrugs for antitumor therapy

Nitro seco analogs (nitroCBIs) of the antitumor antibiotic duocarmycins are a new class of hypoxia activated prodrugs. These compounds undergo hypoxia-selective metabolism to form potent DNA alkylating agents. A series of four nitroCBI alcohol prodrugs containing a bromide rather than chloride or sulfonate leaving group was synthesized. In assays for in vitro hypoxia-selective cytotoxicity against human tumor cell lines the two bromides with DNA minor groove binding basic side chains displayed hypoxic cytotoxicity ratios (HCRs) of 52–286 in HT29 cells and 41–43 in SiHa cells. These values compare well with a related previously reported chloride analog. The corresponding more water soluble phosphate pre-prodrugs of the bromides were synthesized and evaluated for in vivo antitumor activity against SiHa human tumor xenografts. All four phosphates, with both neutral and basic side chains, demonstrated activity providing statistically significant hypoxic log 10 cell kills of 0.87–2.80 at non-toxic doses, matching or proving superior to those of their chloride analogs.

Preparation and Antitumour Properties of the Enantiomers of a Hypoxia‐Selective Nitro Analogue of the Duocarmycins

Racemic 2-{[1-(chloromethyl)-5-nitro-3-{5-[2-(dimethylamino)ethoxy]indol-2-carbonyl}-1,2-dihydro-3H-benzo[e]indol-7-yl]sulfonyl}aminoethyl dihydrogen phosphate, a synthetic nitro derivative of the duocarmycins, is a hypoxia-selective prodrug active against radiation-resistant tumour cells at nontoxic doses in mice. An intermediate in the synthesis of this prodrug was resolved by chiral HPLC and the absolute configuration assigned by X-ray crystallography. The intermediate was used to prepare the prodrug's enantiomers, and also the enantiomers of the active nitro and amino metabolites. In vitro analysis in the human cervical carcinoma cell line SiHa showed that both nitro enantiomers are hypoxia-selective cytotoxins, but the "natural" S enantiomer is at least 20-fold more potent. Examination of extracellular amino metabolite concentrations demonstrated no enantioselectivity in the hypoxia-selective reduction of nitro to amino. Low levels of amino derivative were also found in aerobic cell suspensions, sufficient to account for the observed oxic toxicity of the nitro form. At an equimolar dose in SiHa-tumour bearing animals, the (-)-R enantiomer of the prodrug was inactive, while the (+)-S enantiomer caused significantly more hypoxic tumour cell kill than the racemate. At this dose, the combination of (+)-S-prodrug and radiation eliminated detectable colony-forming cells in four out of five treated tumour-bearing animals.

The effect of sulfonate leaving groups on the hypoxia-selective toxicity of nitro analogs of the duocarmycins

A series of 3-substituted (5-nitro-2,3-dihydro-1H-benzo[e]indol-1-yl)methyl sulfonate (nitroCBI) prodrugs containing sulfonate leaving groups undergo hypoxia-selective metabolism to form potent DNA minor groove alkylating agents. They were evaluated (along with chloride leaving group analogs for comparison) for their cytotoxicity against cultures of SKOV3 and HT29 human tumor cell lines under both aerobic and hypoxic conditions. Sulfonates with neutral side chains (e.g., 5,6,7-trimethoxyindole; TMI) show consistently higher hypoxic cytotoxicity ratios (HCRs) (34–246) than the corresponding chloro analogs (2.8–3.1) in SKOV3 cells, but these trends do not hold for compounds with cationic or polar neutral side chains.

Selective Treatment of Hypoxic Tumor Cells In Vivo: Phosphate Pre‐Prodrugs of Nitro Analogues of the Duocarmycins

Selective Treatment of Hypoxic Tumor Cells In Vivo: Phosphate Pre‐Prodrugs of Nitro Analogues of the Duocarmycins

Selective Treatment of Hypoxic Tumor Cells In Vivo: Phosphate Pre‐Prodrugs of Nitro Analogues of the Duocarmycins

Selective Treatment of Hypoxic Tumor Cells In Vivo: Phosphate Pre‐Prodrugs of Nitro Analogues of the Duocarmycins

ChemInform Abstract: Substituted Isoquinolines by Noyori Transfer Hydrogenation: Enantioselective Synthesis of Chiral Diamines Containing an Aniline Subunit.

Transfer hydrogenation using the Noyori catalyst 5-Ts is effective for the enantioselective hydrogenation of imines containing fully substituted nitrogen groups (12 or 13). Analogues such as 11c could not be reduced in practical yield, apparently due to product inhibition of the catalyst. Asymmetric transfer hydrogenation of the aniline imine 8a was possible, but required impractical purity levels for the substrate, and the nitro analogue 7 could not be reduced efficiently. The best results were obtained with the bromophenyl imine 20. In the case of 20b, the product 21b was formed with 98.7% ee, and the material could be upgraded to >99% ee by crystallization of the hydrochloride salt. Reaction of 21b with NH3 or MeNH2 in the presence of Cu/CuCl gave the chiral anilines 10b or 23b. The latter substance is comparable to the commercially available 1 as a chiral proton donor for amide enolates and provides access to the hitherto unavailable enantiomeric series.