Design, synthesis and biological evaluation of diamino substituted cyclobut-3-ene-1,2-dione derivatives for the treatment of drug-resistant tuberculosis.

Mycobacterium tuberculosis (Mtb) ATP synthase is an important target for treating drug-resistant infections and sterilizing the bacteria, spurring intensive efforts to develop new TB therapeutics based on this target. In this work, four novel series including furan-2(5H)-ketone (3, 4), maleimide (5) and squaramide (6) derivatives were designed, respectively, through the strategy of scaffold morphing and hydrogen-bond introduction, using the selective Mtb ATP synthase inhibitor compound 2 as the lead compound. The result demonstrated that diamino substituted cyclobut-3-ene-1,2-dione compounds 6ab and 6ah displayed good to excellent in vitro anti-TB activities (MIC 0.452-0.963 µg/mL) with low cytotoxicity (IC50 > 64 µg/mL). In addition, not only did compound 6ab show effective activity against clinically isolated resistant strains, it also revealed good druggability profiles including improved metabolic stability, no hERG channel inhibition potential, and acceptable oral bioavailability. The preliminary result of docking study and in vitro anti-bedaquiline-resistant strain test compared to compound 2 suggested that Mtb ATP synthase is most likely the target of compound 6ab. The structure-activity relationship laid a good foundation for the identification of novel squaramides as a potential treatment of drug-resistant tuberculosis.

Pharmacological effects and mechanisms of muscone.

Musk, the dried secretion from the preputial follicles of the male musk deer (genus Moschus), possesses various pharmacological activities and has been used extensively in traditional Chinese medicine for thousands of years. Muscone is the main active ingredient of musk and exerts pharmacological effects similar to those of musk. Although muscone was notably used to treat various disorders and diseases, such as neurological disorders, chronic inflammation and ischemia-reperfusion injury, most of the mechanisms of the pharmacological action of muscone remain unclear because of slow progress in research before the 21st century. In recent years, the pharmacological activities and mechanisms of muscone have been clarified. The present article summarizes the pharmacological and biological studies on cerebrovascular disease, cardiovascular disease, neurological effects, cancer and others and the associated mechanisms of the action of muscone to date.

Muscone Ameliorates LPS-Induced Depressive-Like Behaviors and Inhibits Neuroinflammation in Prefrontal Cortex of Mice.

Depression is partially caused by inflammation in the central nervous system. Early study demonstrated that musk, glandular secretion from male musk deer, exerted an antidepressant-like effect. The aim of this study was to investigate if muscone, a bioactive ingredient in musk, could ameliorate neuroinflammation and depressive-like behaviors as well as explore the potential action mechanism. Mice were intraperitoneally (i.p.) injected with muscone for 2 weeks prior to administration of lipopolysaccharides (LPS, 1mg/kg, i.p.). Pre-treatment with muscone reversed the LPS-induced decrease in body weight within 24h and ameliorated depressive-like behaviors shown by sucrose preference, tail suspension test, and forced swimming test. LPS-induced activation of microglial cells and elevation in expression of inflammatory cytokines including IL-1ß, RANTES, and MCP-1 in the prefrontal cortex of mice were effectively abrogated by muscone, which significantly down-regulated expression of TLR4, MyD88, Caspase-1, NLRP3, renin, and Ang II. In addition, treatment of BV2 microglia cells with muscone markedly attenuated the LPS-induced rise in protein expression of TLR4, Ang II, and IL-1ß. This study revealed that muscone could ameliorate LPS-induced depressive-like behaviors by repressing neuroinflammation in the prefrontal cortex of mice caused by its suppression on microglia activation and production of inflammatory cytokines via acting on TLR4 pathway and RAS cascade.

Muscone/RI7217 co-modified upward messenger DTX liposomes enhanced permeability of blood-brain barrier and targeting glioma.

Rationale: The dual-targeted drug delivery system was designed for enhancing permeation of the blood-brain barrier (BBB) and providing an anti-glioma effect. As transferrin receptor (TfR) is over-expressed by the brain capillary endothelial (hCMEC/D3) and glioma cells, a mouse monoclonal antibody, RI7217, with high affinity and selectivity for TfR, was used to study the brain targeted drug delivery system. Muscone, an ingredient of traditional Chinese medicine (TCM) musk, was used as the "guide" drug to probe the permeability of the BBB for drug delivery into the cerebrospinal fluid. This study investigated the combined effects of TCM aromatic resuscitation and modern receptor-targeted technology by the use of muscone/RI7217 co-modified docetaxel (DTX) liposomes for enhanced drug delivery to the brain for anti-glioma effect. Methods: Cellular drug uptake from the formulations was determined using fluorescence microscopy and flow cytometry. The drug penetrating ability into tumor spheroids were visualized using confocal laser scanning microscopy (CLSM). In vivo glioma-targeting ability of formulations was evaluated using whole-body fluorescent imaging system. The survival curve study was performed to evaluate the anti-glioma effect of the formulations. Results: The results showed that muscone and RI7217 co-modified DTX liposomes enhanced uptake into both hCMEC/D3 and U87-MG cells, increased penetration to the deep region of U87-MG tumor spheroids, improved brain targeting in vivo and prolonged survival time of nude mice bearing tumor. Conclusion: Muscone and RI7217 co-modified DTX liposomes were found to show improved brain targeting and enhanced the efficacy of anti-glioma drug treatment in vivo.

Inhibitory Effects of Benzaldehyde, Vanillin, Muscone and Borneol on P-Glycoprotein in Caco-2 Cells and Everted Gut Sac.

AIMS: In clinical practice, herbal medicines have played an important role in the modulation of drug transporters through the combination of conventional prescription drugs, which necessitates the elucidation of herb-drug interactions. The present study was designed to investigate the inhibitory effects and mechanisms of benzaldehyde, vanillin, muscone, and borneol on P-glycoprotein (P-gp). METHODS: The effects of the 4 compounds on the intracellular accumulation of rhodamine-123 (Rho-123) in vinblastine-treated Caco-2 (VB-Caco-2) cells were studied by monitoring fluorescence intensity through a flow cytometry assay, and the effects of these compounds on Rho-123 transport through VB-Caco-2 monolayers and Rho-123 intestinal absorption in the rat everted gut sac were investigated by high-performance liquid chromatography. Moreover, P-gp expression in VB-Caco-2 cells was assessed using flow cytometry and Western blot analysis, and the relative ABCB1 mRNA level was determined by Real-time RT-PCR. KEY FINDINGS: The results showed that benzaldehyde, vanillin, muscone, and borneol significantly increased Rho-123 uptake in VB-Caco-2 cells, increased the absorption rate and apparent permeability coefficient of Rho-123 in rat jejunum and ileum, and decreased the efflux ratio of Rho-123 from 6.52 to less than 2 during transport across VB-Caco-2 cell monolayers. In addition, these compounds reduced the protein and ABCB1 mRNA levels of P-gp in VB-Caco-2 cells. CONCLUSIONS: These data indicate that benzaldehyde, vanillin, muscone and borneol could effectively reverse multidrug resistance via inhibiting the P-gp function and expression pathway. The data provide fodder for further investigation into the interaction between the 4 compounds and other drugs transported by P-gp.

[Brain Protection of Muscone in Rats with Brain Injury].

OBJECTIVE: To observe cerebral protective effect of muscone (nasal administration) on traumatic brain injury model rats. METHODS: SD rats were divided into the sham-operation group, the model group, and the treatment groups according to random digit table, 50 in each group. Traumatic brain injury model was established by controlled cortical strike. Rats in the sham-operation group received surgery and anesthesia procedures only, with no strike. Muscone (1.8 mg/kg) was delivered to rats in the treatment group using in situ nasal perfusion, 30 min each time, twice daily for 7 successive days. Water content of brain tissue was detected in each group before intervention (T1), at day 3 of intervention (T2), day 5 of intervention (T3), and after intervention (T4), respectively. Expression levels of brain derived neurotrophic factor (BDNF) and nerve growth factor (NGF) were detected using immunohistochemical analysis. RESULTS: Compared with the sham-operated group, water content of brain tissue increased (P < 0.05), and expression levels of NGF and BDNF decreased in the model group at T1, T2, T3, and T4 (P <0. 01). Compared with the model group, water content of brain tissue decreased (P < 0.05), and expression levels of NGF and BDNF increased (P < 0.01) in the treatment group at T1, T2, and T3. CONCLUSION: Nasal administration of muscone could reduce water content of brain tissue, alleviate cerebral edema, promote secretion of BDNF and NGF by olfactory ensheathing cells in traumatic brain injury rats.

Muscone exerts neuroprotection in an experimental model of stroke via inhibition of the fas pathway.

Identifying small molecules that are neuroprotective against stroke injury will be highly beneficial for treatment therapies. A cell viability assay and gas chromatography-mass spectrometry were used to identify active small molecules in XingNaoJing, which is a well known Chinese medicine prescribed for the effective treatment of stroke. Studies have found that muscone is the active compound that prevents PC12 cell and cortical neuron damage following various injuries. Analysis of apoptosis indicated that muscone inhibited glutamate-induced apoptotic cell death of PC12 cells and cortical neurons. Fas and caspase-8 expression were upregulated following glutamate treatment in cortical neurons, and was markedly attenuated in the presence of muscone. Furthermore, muscone significantly reduced cerebral infarct volume, neurological dysfunction and inhibited cortical neuron apoptosis in middle cerebral artery occluded (MCAO) rats in a dose-dependent manner. Moreover, a significant decrease in Fas and caspase-8 expression in the rat cortex was observed in MCAO rats treated with muscone. Our results demonstrate that muscone may be a small active molecule with neuroprotective properties, and that inhibition of apoptosis and Fas is an important mechanism of neuroprotection by muscone. These findings suggest a potential therapeutic role for muscone in the treatment of stroke.

Immunoinhibitory effect of teuclatriol a guaiane sesquiterpene from Salvia mirzayanii.

BACKGROUND: Salvia mirzayanii, a native plant to Iran, is shown to have immunomodulatory effects on lymphocyte proliferation. OBJECTIVE: To identify the bioactive immunomodulatory compound(s) present in S. mirzayanii. METHODS: The crude extract was fractionated to five fractions in two steps using different solvents. The fractions were subjected to bioassay-guided fractionation. All the fractions were tested for bioactivity on human activated-peripheral blood lymphocytes (PBLs) using cell proliferation assay. RESULTS: The methanol fraction (Fr. M) showed the highest inhibitory effect on PBLs compared to other fractions. Fr. M was applied on a gravity column chromatography for further fractionation. Resultant fractions, demonstrated inhibitory effects at higher concentrations. Fr. 4 with an 18.9 ± 0.2% inhibitory activity at 200 µg/ml and with the highest quantity was applied on preparative TLC plates for further purification. The final purified compound was identified as teuclatriol, a guaiane sesquiterpene, by NMR analysis. This compound showed a significant anti-proliferative effect on human activated-peripheral blood lymphocytes (IC50, 72.8 ± 5.4 µg/ml). CONCLUSION: Teuclatriol was found to be one of the compounds responsible for the immunoinhibitory effect of Salvia mirzayanii. We suggest further studies on teuclatriol, exploring its mechanism of action as an immunomodulatory compound.

Protective effects of muscone on ischemia-reperfusion injury in cardiac myocytes.

ETHNOPHARMACOLOGICAL RELEVANCE: Musk has been traditionally used in Chinese medicine as the main ingredient of many formulations for the treatment of chest pain and angina pectoris. AIM OF THE STUDY: To investigate the protective effects of muscone (the active ingredient of musk) on ischemia-reperfusion (I/R) injury induced by hypoxia and low glucose in primary cultured rat cardiac myocytes. MATERIALS AND METHODS: Primary cultures of neonatal rat cardiac myocytes were subjected to ischemia-reperfusion in media, with or without muscone. Cell viability, release of lactic acid dehydrogenase (LDH), superoxide dismutase (SOD) activity, malondialdehyde (MDA) levels, creatine kinase (CK) and caspase-3 activities, as well as intracellular free Ca(2+) concentrations, were measured. Cellular apoptosis and mitochondrial membrane potential (MMP) were assessed by flow cytometry, and the expression of Bcl-2 and Bax proteins was assessed by Western blotting. RESULTS: Following the exposure of cardiac myocytes to ischemia-reperfusion, there was a marked decrease in pulsating frequency, cell viability, SOD activity, MMP, and the expression of Bcl-2 protein, accompanied by increased LDH release, MDA production, CK and caspase-3 activities, intracellular free Ca(2+) concentrations, rate of apoptosis, and expression of Bax protein. Pretreatment with muscone (0.215, 0.43, 0.86 µg/mL) prior to I/R injury significantly attenuated the above changes. CONCLUSION: Muscone has a protective effect against I/R injury in cardiac myocytes, indicating that muscone may potentially provide therapeutic benefit in I/R injury by inhibiting cellular oxidative stress and apoptosis.

[Protective effects and machanism of muskone on pheochromocytoma cell injure induced by glutamate].

OBJECTIVE: To explore the mechanism of glutamate (Glu)-induced PC12 cell apoptosis and the protective effect of muscone. METHOD: PC12 cells were randomly divided into six groups: normal group (Normal), model group injured by glutamate (Glu), nimodipine group (Nim) and muskone groups (Mus) of high, middle and low doses. The PC12 cells were pretreated with or without different concentrations of muskone for 30 min and then exposed to glutamate. MTT assay for cell survival, flow cytometric detection of apoptotic cells, DCF assay for reactive oxygen species (ROS) and flow cytometric assay was performed to determine the mitochondrial membrane potential in PC12 cell. RESULT: PC12 cell damage, the concentrations of [Ca2+], and apoptosis induced by Glu were decreased after being administrated with paeoniflorin. CONCLUSION: Muskone inhibited Glu-induced apoptosis in PC12 cells. The mechanism is related to inhibiting intracellular Ca2+ overload and maintaining mitochondral membrance potential.

[Effects of series of Muskone on heart hemodynamics and myocardial consumption of oxygen in experimental dogs].

OBJECTIVE: To observe the effects of series of Muskone (the muskone includes Slender Dutchmanspipe Root, Inula Root and neither kind of Common Aucklandia Root) on the heart hemodynamics and myocardial consumption of oxygen in experimental dogs, and to explain its pharmacological action on cardiovascular system. METHOD: Arterial blood pressure, coronary blood flow, resistance in coronary artery, total peripheral resistance, work of left artrium and oxygen consumption index of the cardiac muscles were observed in anaesthetic dogs. RESULT: The series of Muskone decreased arterial blood pressure significantly, dilated coronary artery and peripheral arteries significantly, increased coronary blood flow, decreased resistance in coronary artery, improved the work of left artrium, the oxygen availability of cardiac muscles and the complaisance of arteries in cardiac muscles.

[Review on investigations related to chemical constituents and biological activities of Periplaneta americana].

With the fast development of medicinal animals as new drugs, research on Periplaneta americana become hot recently. Several drugs which mainly consisted of P. americana were approved for clinical applications. The chemical constituents and pharmacological bioactivities of this insect were summarized herein, which provides informativon for further researches on this medicinal animal.

[Study on comparative pharmacology of series of Muskone].

OBJECTIVE: To study the therapeutic effects of the series of Muskone (the Muskone includes Slender Dutchmanspipe Root, Tumuxiang, and not Slender Dutchmanspipe Root) on experimental myocardial infarct and pain in rats. METHOD: Coronary artery ligation was applied for the model of myocardial infarct. Therapeutic effects were evaluated by measuring parameters of histomorphometry, blood plasm of ET, 6- keto-PGF1alpha and TXB2. Intraperitoneal injection acetic was applied for the model of ache, the frequency and eclipse period of writhing were evaluated its effect of resisting pain. RESULT: The Muskone including Radix Aristolociae, the Muskone including Radix Inulae and the Muskone without Radix Aucklandiae all can decrease the area of myocardial infarction in rats, the level of TXB2, ET, and the frequency of writhing significantly. Also it can increase the level 6-keto-PGF1alpha, the ratio of 6-keto-PGF1alpha and TXB2. Single Radix Aristolociae or Radix Inulae only relieved pain. CONCLUSION: The Muskone including Radix Aristolociae, the Muskone including Radix Inulae and the Muskone without Radix Aucklandiae all have significant therapeutic effect on both myocardial infarction and pain, while single Radix Aristolociae or Radix Inulae only can relieve pain.

[Study on therapeutic effects of series of muskone on myocardial infarction canines].

OBJECTIVE: To observe the effects of the series of Muskone (the Muskone includes Slender Dutchmanspipe Root, Tumuxiang, and not Slender Dutchmanspipe Root) on myocardial ischemia, myocardial infarction and hematological index in experimental canines, and to explain the pharmacological action and characteristic of its therapeutic effect on ischemic heart disease. METHOD: The range and degree of myocardial ischemia was evaluated by epicardial electrogram mapping, and the range extent of myocardial infarction was determined by quantitate histology (N-BT staining method). Meanwhile, the changes of ET, TXB2, 6-Keto-PGF1alpha were determined to study the effects of the series of Muskone on myocardial ischemia, myocardial infarction and hematological index in experimental canines. RESULT: The series of Muskone can improve myocardial ischemia and infarction in experimental canines, and relieve significantly the degree of myocardial ischemia (Sigma-ST) determined by epicardial electrogram mapping, decrease the range of myocardial ischemia (N-ST) determined by epicardial electrogram mapping and decrease infarction zone determined by N-BT staining method. And it has a significant inhibition on activity of ET induced by myocardial ischemia and infarction, and increases 6-Keto-PGF1alpha and 6-Keto-PGF1alpha/TXB2 induced by myocardial ischemia. CONCLUSION: The series of Muskone has significant therapeutic effect on myocardial infarction.

Polyhalogenated and perfluorinated compounds that disobey the Meyer-Overton hypothesis.

Fourteen polyhalogenated, completely halogenated (perhalogenated), or perfluorinated compounds were examined for their anesthetic effects in rats. Anesthetic potency or minimum alveolar anesthetic concentration (MAC) was quantified using response/nonresponse to electrical stimulation of the tail as the end-point. For compounds that produced excitable behavior, and/or did not produce anesthesia when given alone, we determined MAC by additivity studies with desflurane. Nine of 14 compounds had measurable MAC values with products of MAC x oil/gas partition coefficient ranging from 3.7 to 24.8 atm. Because these products exceed that for conventional inhaled anesthetics (1.8 atm), they demonstrate a deviation from the Meyer-Overton hypothesis. Five compounds (CF3CCIFCF3, CF3CCIFCCIFCF3, perfluorocyclobutane, 1,2-dichloroperfluorocyclobutane, and 1,2-dimethylperfluorocyclobutane) had no anesthetic effect when given alone, had excitatory effects when given alone, and tended to increase the MAC for desflurane. These five compounds had no anesthetic properties in spite of their abilities to dissolve in lipids and tissues, to penetrate into the central nervous system, and to be administered at high enough partial pressures so that they should have an anesthetic effect as predicted by the Meyer-Overton hypothesis. Such compounds will be useful in identifying and differentiating anesthetic sites and mechanisms of action. Any physiologic or biophysical/biochemical change produced by conventional anesthetics and deemed important for the anesthetic state should not be produced by nonanesthetics.

Repeated intrathecal injections of dezocine produce antinociception without evidence for neurotoxicity in the rat: a study of morphometric evaluation of spinal cord histology.

This study was designed to determine the antinociceptive and spinal cord histologic effects of a new agonist/antagonist opioid drug dezocine. This drug was injected intrathecally in rats at a dose of 50 or 125 micrograms twice daily for 14 days. The tail-flick test showed that the antinociceptive effect declined gradually, with no detectable effects by day 14. Quantitative histologic techniques and light and electron microscopy showed that neither dose, compared with vehicle, created any morphologic changes in the spinal cord that could be attributed to a neurotoxic or otherwise degenerative effect of the drug. In conclusion, dezocine is a drug that gives rise to sustained antinociceptive effects when administered intrathecally and causes no morphologic changes in the rat spinal cord that could be indicative of neurotoxic potential.