Ultrasound-Assisted Synthesis of 4-Alkoxy-2-methylquinolines: An Efficient Method toward Antitubercular Drug Candidates.

Tuberculosis (TB) has been described as a global health crisis since the second half of the 1990s. Mycobacterium tuberculosis (Mtb), the etiologic agent of TB in humans, is a very successful pathogen, being the main cause of death in the population among infectious agents. In 2019, it was estimated that around 10 million individuals were contaminated by this bacillus and about 1.2 million succumbed to the disease. In recent years, our research group has reported the design and synthesis of quinoline derivatives as drug candidates for the treatment of TB. These compounds have demonstrated potent and selective growth inhibition of drug-susceptible and drug-resistant Mtb strains. Herein, a new synthetic approach was established providing efficient and rapid access (15 min) to a series of 4-alkoxy-6-methoxy-2-methylquinolines using ultrasound energy. The new synthetic protocol provides a simple procedure utilizing an open vessel system that affords the target products at satisfactory yields (45-84%) and elevated purities (≥95%). The methodology allows the evaluation of a larger number of molecules in assays against the bacillus, facilitating the determination of the structure-activity relationship with a reduced environmental cost.

1,1-Diheterocyclic Ethylenes Derived from Quinaldine and Carbazole as New Tubulin-Polymerization Inhibitors: Synthesis, Metabolism, and Biological Evaluation.

We report the synthesis and metabolic and biological evaluation of a series of 17 novel heterocyclic derivatives of isocombretastatin-A4 (iso-CA-4) and their structure-activity relationships. Among these derivatives, the most active compound, 4f, inhibited the growth of a panel of seven cancer cell lines with an IC50 in the low nanomolar range. In addition, 4f showed interesting activity against CA-4-resistant colon-carcinoma cells and multidrug-resistant leukemia cells. It also induced G2/M cell-cycle arrest. Structural data indicated binding of 4f to the colchicine site of tubulin, likely preventing the curved-to-straight tubulin structural changes that occur during microtubule assembly. Also, 4f disrupted the blood-vessel-like assembly formed by human umbilical-vein endothelial cells in vitro, suggesting its function as a vascular-disrupting agent. An in vitro metabolism study of 4f showed its high human-microsomal stability in comparison with that of iso-CA-4. The physicochemical properties of 4f may be conducive to CNS permeability, suggesting that this compound may be a possible candidate for the treatment of glioblastoma.

In Silico Discovery of a Substituted 6-Methoxy-quinalidine with Leishmanicidal Activity in Leishmania infantum.

There is an urgent need for the discovery of new antileishmanial drugs with a new mechanism of action. Type 2 NADH dehydrogenase from Leishmania infantum (LiNDH2) is an enzyme of the parasite's respiratory system, which catalyzes the electron transfer from NADH to ubiquinone without coupled proton pumping. In previous studies of the related NADH: ubiquinone oxidoreductase crystal structure from Saccharomyces cerevisiae, two ubiquinone-binding sites (UQI and UQII) were identified and shown to play an important role in the NDH-2-catalyzed oxidoreduction reaction. Based on the available structural data, we developed a three-dimensional structural model of LiNDH2 using homology detection methods and performed an in silico virtual screening campaign to search for potential inhibitors targeting the LiNDH2 ubiquinone-binding site 1-UQI. Selected compounds displaying favorable properties in the computational screening experiments were assayed for inhibitory activity in the structurally similar recombinant NDH-2 from S. aureus and leishmanicidal activity was determined in the wild-type axenic amastigotes and promastigotes of L. infantum. The identified compound, a substituted 6-methoxy-quinalidine, showed promising nanomolar leishmanicidal activity on wild-type axenic promastigotes and amastigotes of L. infantum and the potential for further development.

Unconventional Coupling between Ligand Recognition and Allosteric Control in the Multidrug Resistance Gene Regulator, BmrR.

BmrR is a multidrug resistance (MDR) regulator that responds to diverse ligands. To obtain insight into signal recognition, allosteric control, and cooperativity, we used a quantitative in vitro transcription assay to determine the ligand-dependent activation profiles for a diverse set of cations, zwitterions, and uncharged ligands. As for many other biological switch systems, the data are well described by a modified Hill equation. Parameters extracted from curve fits to the data include L50 , RMAX and N. We found that L50 values correlate directly with ΔGBIND values, suggesting that the parameter reflects binding, whereas RMAX and N reflect allosteric control and cooperativity, respectively. Our results suggest unconventional coupling between ligand binding and allosteric control, with weakly interacting ligands exhibiting the highest levels of activation. Such properties are in stark contrast to those often exhibited by biological switch proteins, whereby ligand binding and allostery are tightly coupled, yielding both high selectivity and ultrasensitivity. We propose that weakened coupling, as observed for BmrR, may be important for providing robust activation responses to unrelated ligands. We also propose that other MDR proteins and other polyspecific switch systems will show similar features.

An indolylquinoline derivative activates DNA damage response and apoptosis in human hepatocellular carcinoma cells.

Human liver cancer is one of the most frequently diagnosed cancers worldwide. The development of resistance to therapy limits the application against the disease. To improve treatment, new effective anticancer agents are constantly pursued. Previously, we reported that an indolylquinoline, 3-((7-ethyl-1H-indol-3-yl)-methyl)-2-methylquinoline (EMMQ), is effective in suppressing the growth of human lung cancer by impairing mitochondria functions. The present study revealed that EMMQ inhibited cell growth and induced apoptosis in liver cancer cells, but not in normal cells. This study demonstrated that EMMQ induced DNA damage by activating p53 and γ-H2AX and cell arrest by suppressing cyclin D1 and CDK2. Damaged DNA injured mitochondrial functions by lowering the membrane potential and producing reactive oxygen species. The subsequent mitochondrial cytochrome c release attenuated pro-survival signals and increased apoptotic characteristics. Introduction of p53 shRNA abrogated drug effects by reducing DNA damage while maintaining mitochondria integrity. In brief, the study demonstrates that the effectiveness of EMMQ accentuated apoptosis of hepatocarcinoma cells by activating p53. Based on these collective findings, the study offered a new perspective of EMMQ that was shown to be a promising candidate to treat liver cancer.

Cellular mechanisms of desynchronizing effects of hypothermia in an in vitro epilepsy model.

Hypothermia can terminate epileptiform discharges in vitro and in vivo epilepsy models. Hypothermia is becoming a standard treatment for brain injury in infants with perinatal hypoxic ischemic encephalopathy, and it is gaining ground as a potential treatment in patients with drug resistant epilepsy. However, the exact mechanism of action of cooling the brain tissue is unclear. We have studied the 4-aminopyridine model of epilepsy in mice using single- and dual-patch clamp and perforated multi-electrode array recordings from the hippocampus and cortex. Cooling consistently terminated 4-aminopyridine induced epileptiform-like discharges in hippocampal neurons and increased input resistance that was not mimicked by transient receptor potential channel antagonists. Dual-patch clamp recordings showed significant synchrony between distant CA1 and CA3 pyramidal neurons, but less so between the pyramidal neurons and interneurons. In CA1 and CA3 neurons, hypothermia blocked rhythmic action potential discharges and disrupted their synchrony; however, in interneurons, hypothermia blocked rhythmic discharges without abolishing action potentials. In parallel, multi-electrode array recordings showed that synchronized discharges were disrupted by hypothermia, whereas multi-unit activity was unaffected. The differential effect of cooling on transmitting or secreting γ-aminobutyric acid interneurons might disrupt normal network synchrony, aborting the epileptiform discharges. Moreover, the persistence of action potential firing in interneurons would have additional antiepileptic effects through tonic γ-aminobutyric acid release.

Glomerulus-specific, long-latency activity in the olfactory bulb granule cell network.

Reliable, stimulus-specific temporal patterns of action potentials have been proposed to encode information in many brain areas, perhaps most notably in the olfactory system. Analysis of such temporal coding has focused almost exclusively on excitatory neurons. Thus, the role of networks of inhibitory interneurons in establishing and maintaining this reliability is unclear. Here we use imaging of population activity in vitro to investigate the mechanisms of temporal pattern generation in mouse olfactory bulb inhibitory interneurons. We show that activity of these interneurons evolves slowly in time but that individual neurons fire at reliable times, with a timescale similar to the slow changes in the patterns of odor-evoked activity and to odor discrimination. Most strikingly, the latency of a single granule cell is highly reliable from trial to trial during repeated stimulation of the same glomerulus, whereas this same cell will have a markedly different latency when a different glomerulus is activated. These data suggest that the timing of granule cell-mediated inhibition in the olfactory bulb is tightly regulated by the source of input and that inhibition may contribute to the generation of reliable temporal patterns of mitral cell activity.

Synthesis and SAR study of acridine, 2-methylquinoline and 2-phenylquinazoline analogues as anti-prion agents.

Transmissible spongiform encephalopathies (TSEs) are thought to arise from aggregation of a protease resistant protein denoted PrP(Sc), which is a misfolded isoform of the normal cellular prion protein PrP(C). Using virtual high-throughput screening we have selected structures analogous to acridine, 2-methyquinoline and 2-phenylquinazoline as potential therapeutic candidates for the treatment of TSEs. From the synthesis and screening of constructed libraries we have shown that an electron-rich aromatic ring attached through an amine linker to the position para to the ring nitrogen is beneficial to both binding to PrP(C) and the suppression of PrP(Sc) accumulation for acridine and 2-methylquinoline analogues. 2-Phenylquinazoline analogues appear to utilise a different mode of action by binding at a different location and/or pose. We report IC50s in the nanomolar range.

Dequalinium-induced protofibril formation of alpha-synuclein.

alpha-Synuclein is the major constituent of Lewy bodies, a pathological signature of Parkinson disease, found in the degenerating dopaminergic neurons of the substantia nigra pars compacta. Amyloidosis generating the insoluble fibrillar protein deposition has been considered to be responsible for the cell death observed in the neurodegenerative disorder. In order to develop a controlling strategy toward the amyloid formation, 1,1'-(1,10-decanediyl)-bis-[4-a-mino-2-methylquinolinium] (dequalinium), was selected and examined in terms of its specific molecular interaction with alpha-synuclein. The protein was self-oligomerized by dequalinium, which gave rise to the ladder formation on N-[2-hydroxy-1,1-bis(hydroxymethyl)ethyl]glycine/SDS-PAGE in the presence of a coupling reagent of N-(ethoxycarbonyl)-2-ethoxy-1,2-dihydroquinoline. The double-headed structure of dequalinium with the two cationic 4-aminoquinaldinium rings was demonstrated to be critical for the protein self-oligomerization. The dequalinium-binding site was located on the acidic C-terminal region of the protein with an approximate dissociation constant of 5.5 mum. The protein self-oligomerization induced by the compound has resulted in the protofibril formation of alpha-synuclein before it has developed into amyloids. The protofibrils were demonstrated to affect the membrane intactness of liposomes, and they have also been shown to influence cell viability of human neuroblastoma cells. In addition, dequalinium treatment of the alpha-synuclein-overexpressing cells exerted a significant cell death. Therefore, it is pertinent to consider that dequalinium could be used as a molecular probe to assess toxic mechanisms related to the amyloid formation of alpha-synuclein. Ultimately, the compound could be employed to develop therapeutic and preventive strategies toward alpha-synucleinopathies including Parkinson disease.

Modulation of Ca(2+)-dependent and Ca(2+)-independent miniature endplate potentials by phorbol ester and adenosine in frog.

Phorbol esters and adenosine modulate transmitter release from frog motor nerves through actions at separate sites downstream of calcium entry. However, it is not known whether these agents have calcium-independent sites of action. We therefore characterised calcium independent miniature endplate potentials (mepps) generated in response to 4-aminoquinaldine (4-AQ(A)) and then compared the modulation of these mepps by phorbol esters and adenosine with that of normal calcium dependent mepps. Application of 30 microM 4-AQ(A) resulted in the appearance of a population of mepps with amplitudes greater than twice the total population mode (mepp(>2M)). In the presence of 4-AQ(A), K(+) depolarisation or hypertonicity increased the numbers of normal amplitude mepps (mepp(N)) but had no effect on the frequency of mepp(>2M) events, suggesting that mepp(>2M) are not dependent on calcium. Treatment with the botulinum toxin (Botx) fractions C, D, or E (which selectively cleave syntaxin, synaptobrevin and SNAP-25, respectively) produced equivalent reductions in both normal and 4-AQ(A) induced mepps, suggesting that both mepp populations have equal dependence on the intact SNARE proteins. Phorbol dibutyrate (PDBu, 100 nM) increased the frequencies of both populations of mepps recorded in the presence of 4-AQ(A). Adenosine (25 microM) selectively reduced the numbers of mepp(N) with no effect on the frequency of mepp(>2M) events. These results suggest that mepp(>2M) events released in response to 4-AQ(A) are dependent on intact forms of syntaxin, synaptobrevin and SNAP-25, but unlike mepp(N) are independent of a functional calcium sensor. The selective action of adenosine, to reduce the numbers of normal amplitude mepps without effecting the frequency of mepp(>2M) events, suggests that adenosine normally inhibits transmitter release through a mechanism that is dependent on the presence of a functional calcium sensor.

Quinaldine derivatives: preparation and biological activity.

The series of quinaldine derivatives were prepared, some of them by means of novel synthetic methods. The synthetic approach, analytical and spectroscopic data of all newly synthesized compounds are presented. The prepared compounds were tested for their in vitro antifungal activity as well as for their photosynthesis-inhibiting activity (the inhibition of photosynthetic electron transport in spinach chloroplasts (Spinacia oleracea L.) and the reduction of chlorophyll content in Chlorella vulgaris Beij.). Structure-activity relationships among the chemical structure, the physical properties and the biological activities of the evaluated compounds are discussed in the article.

Effects of four anaesthetics on the innate immune response of gilthead seabream (Sparus aurata L.).

Anaesthesia may depress the immune system in mammals, but there is no available information on this topic in fish. In the present work, four anaesthetics that are used in aquaculture, MS222 (0 19 mM), benzocaine (0.21 mM), 2-phenoxyethanol (16 mM) and quinaldine sulphate (0.083 mM), were tested in order to observe their effects on the gilthead seabream (Sparus aurata L.) innate immune system. The results showed that the four anaesthetics produced increased blood glucose levels after an hour. In addition, benzocaine and 2-phenoxyethanol depressed complement activity and phagocytosis, while MS222 and quinaldine sulphate did not. Some anaesthesia is a common practice in aquaculture, the data obtained should be taken into account to avoid possible immunodepression in farmed fish.

Synthesis of some substituted quinazolinediones as potential inhibitors of smooth muscle contraction.

3-Substituted 2,4(1H,3H)-quinazolinediones were prepared from the corresponding N-substituted 2-aminobenzamides by treatment with ethyl chloroformate and KOH in ethanol. Also, a series of 3-substituted and 1-methyl-3-substituted 2,4(1H,3H)-quinazolinediones were synthesized by the reaction of 1-methyl-1,4-dihydro- and 1,4-dihydro-2,4-dioxo-3(2H)-quinazolineacetic acid with the corresponding N-substituted piperazines. The 13C NMR spectra and mass spectra of the compounds were measured and signals were assigned. Some of the compounds showed inhibitory action on contractile function of smooth muscle.

2,2-Dialkyl-1,2-dihydroquinolines: cytochrome P-450 catalyzed N-alkylporphyrin formation, ferrochelatase inhibition, and induction of 5-aminolevulinic acid synthase activity.

Incubation of 2,4-diethyl-1,2-dihydro-2-methylquinoline (DMDQ) with hepatic microsomes from rats pretreated with phenobarbital, 3-methylcholanthrene, pregnenolone-16 alpha-carbonitrile, or dexamethasone results in minor loss of the cytochrome P-450 chromophore and accumulation of a hepatic pigment. The hepatic pigment consists of the four regioisomers of N-ethylprotoporphyrin IX and minor amounts of the corresponding N-methyl regioisomers. Exposure of chick embryo liver cells to DMDQ results in inhibition of their ferrochelatase activity, induction of their 5-aminolevulinic acid synthase activity, and accumulation of protoporphyrin IX. 1,2-Dihydro-2,2,4-trimethylquinoline (TMDQ) causes negligible loss of cytochrome P-450 in rat liver microsomes but in vivo still produces the four N-methylprotoporphyrin IX regioisomers in low yield. Furthermore, it inhibits ferrochelatase activity, elevates 5-aminolevulinic acid synthase activity, and causes protoporphyrin IX accumulation in cultured chick embryo hepatocytes. One-electron oxidation of the 2,2-dialkyl-1,2-dihydroquinolines to radical cations is postulated to result in N-alkylation of the prosthetic heme group of cytochrome P-450. The N-alkylprotoporphyrins IX thus formed are potent inhibitors of ferrochelatase. Inhibition of ferrochelatase causes the induction of 5-aminolevulinic acid synthase and the accumulation of protoporphyrin IX. Heme alkylation and ferrochelatase inhibition may be generally associated with substrates that are subject to cytochrome P-450 mediated oxidative extrusion of alkyl radicals.

Quinoline and quninaldine as naturally occurring inhibitors specific for type A monoamine oxidase.

Type A monoamine oxidase (MAO-A) in human placental mitochondria was competitively inhibited by naturally occurring substances, quinoline and quinaldine, using kynuramine as substrate. Quinoline had a higher affinity for MAO than kynuramine. MAO-A in human brain synaptosomal mitochondria was also competitively inhibited by quinoline, while type B MAO (MAO-B) was reversibly and non-competitively inhibited by quinoline. Quinoline inhibited MAO-A much more potently than MAO-B. Of several compounds structurally similar to quinoline, isoquinoline noncompetitively inhibited MAO-A and -B activity.

[Sensitivity of fungi in the genus Candida to antimicrobial preparations]. / Issledovanie chuvstvitel'nosti gribov roda Candida k antimikrobnym preparatam.

Sensitivity of 50 Candida fungus strains isolated from patients with chronic intestine diseases was studied with respect to amphotericin B, levorin, enteroseptol, intestopan and decamethoxin. The rate of forming resistant variants of the strains with respect to decamethoxin and development of their cross resistance to levorin was estimated. It was shown that decamethoxin was the most active antifungal drug among the drugs tested. Estimation of sensitivity of the Candida strains to enteroseptol and intestopan revealed that the fungicidal concentration of enteroseptol for 78 per cent of the strains ranged within 3.9-7.8 micrograms/ml. It was demonstrated that development of resistance to decamethoxin in the strains of Candida albicans was slow and did not reach high levels. No cross resistance between decamethoxin and levorin was detected.

Effect of carcinogenic components of cigarette smoke on in vivo production of murine interferon.

Mice were treated with several different potentially carcinogenic components of tobacco smoke. The chemicals used were: 4-aminobiphenyl and aniline-HCl, which are found in high concentrations in sidestream tobacco smoke; hydrazine sulfate, which is found in high concentrations in mainstream tobacco smoke; and 2-methylquinoline, which is found in intermediate concentrations in both sidestream and mainstream smoke. The chemicals were injected i.p. into mice, and then alpha/beta interferon was induced in the mice by i.v. injection of polyriboinosinic-polyribocytidylic acid. The interferon was induced either 2, 24, or 48 hr after treatment with the tobacco smoke components. Mice treated with 4-aminobiphenyl showed some depression of interferon production 2 hr after treatment, maximum inhibition of interferon induction 24 hr after treatment, and a return to control levels of interferon 48 hr after treatment. Mice treated with hydrazine sulfate showed maximum inhibition of interferon induction 24 hr after treatment but no effects at any other treatment time. These components were the most carcinogenic chemicals of those utilized in this study. Treatment of mice with aniline-HCl, a chemical whose carcinogenic potential is still debated, resulted in marginal depression of interferon induction 24 hr after treatment. 2-Methylquinoline, the chemical with the lowest carcinogenic potential in this study, had no effect on interferon induction after administration to mice. In vivo interferon induction was, therefore, inhibited by treatment of mice with chemical carcinogens found in tobacco smoke. The efficacy of the chemical in inhibiting interferon induction was not influenced by the mainstream or sidestream smoke predominance of the chemical.

Effect of sidestream tobacco smoke components on alpha/beta interferon production.

Mouse embryo fibroblast cell cultures were treated with chemicals that are major components of sidestream (passive) cigarette smoke. These components were 4-aminobiphenyl and aniline-HCl. The cultures produced severely reduced levels of alpha/beta interferon after challenge with polyriboinosinic-polyribocytidylic acid when compared to control cultures. Treatment of additional cell cultures with 2-methylquinoline, and intermediate-level component of sidestream tobacco smoke, or hydrazine-sulfate, a minor component of sidestream tobacco smoke but a major component of mainstream (active) tobacco smoke, also resulted in inhibition of interferon induction with polyriboinosinic acid-polyribocytidylic acid. Therefore, treatment of the cell cultures with chemicals that are carcinogenic was equally effective in inhibiting alpha/beta interferon induction without regard to the sidestream or mainstream smoke origin of the chemical.

Hepatic microsomes from freshwater fish--II. Reduction of benzo(a)pyrene metabolism by the fish anesthetics quinaldine sulfate and tricaine.

1. A single in vivo exposure of brook trout (Salvelinus fontinalis) to a 30.0 mg/l solution of quinaldine sulfate or a 112.5 mg/l solution of tricaine for 5 min significantly reduced the in vitro hydroxylation of benzo(a)pyrene. 2. Since quinaldine sulfate and tricaine formed type I and II binding spectra, respectively, with brook trout hepatic cytochrome P-450, these chemicals probably reduced benzo(a)pyrene hydroxylase enzyme activity by altering the form(s) of cytochrome P-450 responsible for this activity. 3. Hepatic microsomal cytochrome P-450 from brook trout treated with tricaine for 5 min and then placed into fresh water for 24 hr had returned to control levels. 4. Caution should be exercised in the use of quinaldine sulfate or tricaine to anesthetize fish prior to analysis of hepatic microsomal mixed function oxidases.