Antitumor activity of (R,R')-4-methoxy-1-naphthylfenoterol in a rat C6 glioma xenograft model in the mouse
Pharmacol Res Perspect. 2013 Dec;1(2):e00010. doi: 10.1002/prp2.10. Epub 2013 Dec 5
Bernier M, Paul RK, Dossou KS, Wnorowski A, Ramamoorthy A, Paris A, Moaddel R, Cloix JF, Wainer IW
(R,R')-4-methoxy-1-naphthylfenoterol (MNF) inhibits cancer cell proliferation in vitro through cell-type specific modulation of β2-adrenergic receptor and/or cannabinoid receptor function. Here, we report an investigation into antitumor activity of MNF in rat C6 glioma cells. The potent antiproliferative action of MNF in these cells (IC50 of ∼1 nmol/L) was refractory to pharmacological inhibition of β2-adrenergic receptor while a synthetic inverse agonist of cannabinoid receptor 1 significantly blocked MNF activity. The antitumor activity of MNF was then assessed in a C6 glioblastoma xenograft model in mice. Three days after subcutaneous implantation of C6 cells into the lower flank of nude mice, these animals were subjected to i.p. injections of saline or MNF (2 mg/kg) for 19 days and tumor volumes were measured over the course of the experiment. Gene expression analysis, quantitative RT-PCR and immunoblot assays were performed on the tumors after treatment. Significant reduction in mean tumor volumes was observed in mice receiving MNF when compared with the saline-treated group. We identified clusters in expression of genes involved in cellular proliferation, as well as molecular markers for glioblastoma that were significantly downregulated in tumors of MNF-treated mice as compared to saline-injected controls. The efficacy of MNF against C6 glioma cell proliferation in vivo and in vitro was accompanied by marked reduction in the expression of cell cycle regulator proteins. This study is the first demonstration of MNF-dependent chemoprevention of a glioblastoma xenograft model and may offer a potential mechanism for its anticancer action in vivo.
Sub-anesthetic concentrations of (R,S)-ketamine metabolites inhibit acetylcholine-evoked currents in alpha 7 nicotinic acetylcholine receptors
Eur J Pharmacol. 2013 Jan 5;698(1-3):228-34. doi: 10.1016/j.ejphar.2012.11.023. Epub 2012 Nov 23
Moaddel R, Abdrakhmanova G, Kozak J, Jozwiak K, Toll L, Jimenez L, Rosenberg A, Tran T, Xiao Y, Zarate CA, Wainer IW
The effect of the (R,S)-ketamine metabolites (R,S)-norketamine, (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine on the activity of α7 and α3β4 neuronal nicotinic acetylcholine receptors was investigated using patch-clamp techniques. The data indicated that (R,S)-dehydronorketamine inhibited acetylcholine-evoked currents in α7-nicotinic acetylcholine receptor, IC(50) = 55 ± 6 nM, and that (2S,6S)-hydroxynorketamine, (2R,6R)-hydroxynorketamine and (R,S)-norketamine also inhibited α7-nicotinic acetylcholine receptor function at concentrations ≤ 1 μM, while (R,S)-ketamine was inactive at these concentrations. The inhibitory effect of (R,S)-dehydronorketamine was voltage-independent and the compound did not competitively displace selective α7-nicotinic acetylcholine receptor ligands [(125)I]-α-bungarotoxin and [(3)H]-epibatidine indicating that (R,S)-dehydronorketamine is a negative allosteric modulator of the α7-nicotinic acetylcholine receptor. (R,S)-Ketamine and (R,S)-norketamine inhibited (S)-nicotine-induced whole-cell currents in cells expressing α3β4-nicotinic acetylcholine receptor, IC(50) 3.1 and 9.1 μM, respectively, while (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine were weak inhibitors, IC(50) >100 μM. The binding affinities of (R,S)-dehydronorketamine, (2S,6S)-hydroxynorketamine and (2R,6R)-hydroxynorketamine at the NMDA receptor were also determined using rat brain membranes and the selective NMDA receptor antagonist [(3)H]-MK-801. The calculated K(i) values were 38.95 μM for (S)-dehydronorketamine, 21.19 μM for (2S,6S)-hydroxynorketamine and>100 μM for (2R,6R)-hydroxynorketamine. The results suggest that the inhibitory activity of ketamine metabolites at the α7-nicotinic acetylcholine receptor may contribute to the clinical effect of the drug.
Click chemistry for drug development and diverse chemical-biology applications
Chem Rev. 2013 Jul 10;113(7):4905-79. doi: 10.1021/cr200409f. Epub 2013 Mar 27
Thirumurugan P, Matosiuk D, Jozwiak K
Chemistry, traditionally being the science of synthesis and structural manipulations of molecules, has gradually undertaken the more challenging task of biology-oriented synthesis. The generation of molecules/molecular assemblies possessing well-defined biological functions remains an extremely challenging task; immediate refinements in conventional synthetic tactics are necessary. New and more efficient chemical reactions and methodologies, which may override the laborious protection/deprotection and purification steps in conventional total synthesis, could revolutionize the next-generation chemical and biological research. A set of chemical reactions, known as bioorthogonal reactions, that are orthogonal to most functional groups in biological systems has so far shown promising applications in biological research. Of these reactions, the Cu(I)-catalyzed version of the Huisen 1,3-dipolar cycloaddition reaction between azides and terminal alkynes for the construction of trizoles, referred to as a “click chemistry reaction”, was defined by nobel laureate KB Sharpless and associates in 2001. Click chemistry has recently emerged to become one of the most powerful tools in drug discovery, chemical biology, and proteomic applications. In recent years, the design and synthesis of pharmacologically relevant heterocyclic molecules by combinatorial techniques have proven to be a promising strategy in the search for new pharmaceutical lead structures. Click chemistry is one of the powerful reactions for making carbon–heteroatom–carbon bonds in aqueous environment with a wide variety of chemical and biological applications in various fields.
Influence of acute or chronic calcium channel antagonists on the acquisition and consolidation of memory and nicotine-induced cognitive effects in mice
Naunyn Schmiedebergs Arch Pharmacol. 2013 Jul;386(7):651-64. doi: 10.1007/s00210-013-0866-z. Epub 2013 Apr 12
Biala G, Kruk-Slomka M, Jozwiak K
Nicotinic cholinergic receptors (nAChRs) form a heterogeneous family of ligand-gated ion channels found in the nervous system. The main objective of our research was to investigate the interaction between cholinergic nicotinic system and calcium homeostasis in cognitive processes using the modified elevated plus maze memory model in mice. The time each mouse took to move from the open arm to either of the enclosed arms on the retention trial (transfer latency, TL2) was used as an index of memory. Our results showed that a single injection of nicotine (0.035 and 0.175 mg/kg) shortened TL2 values, improving memory-related processes. Similarly, L-type calcium channel antagonists (CCAs), i.e., flunarizine, verapamil, amlodipine, nimodipine, nifedipine, and nicardipine (at the range of dose 5-20 mg/kg) administered before or after training, decreased TL2 value improving memory acquisition and/or consolidation. Interestingly, at the subthresold doses, flunarizine, nicardipine, amlodipine, verapamil, and bupropion, a nAChR antagonist, significantly reversed the nicotine improvement of memory acquisition, while flunarizine, verapamil, and bupropion attenuated the improvement of memory consolidation provoked by an acute injection of nicotine (0.035 mg/kg, s.c.). After subchronic administration (14 days, i.p.) of verapamil and amlodipine, two CCAs with the highest affinity for nAChRs, only verapamil (5 mg/kg) impaired memory acquisition and consolidation while both verapamil and amlodipine, at the subthresold, ineffective dose (2.5 mg/kg), significantly reversed the improvement of memory provoked by an acute injection of nicotine (0.035 mg/kg, s.c.). Our findings can be useful to better understand the interaction between cholinergic nicotinic receptors and calcium-related mechanisms in memory-related processes.
Lipophilicity - methods of determination and its role in medicinal chemistry.
Acta Pol Pharm. 2013 Jan-Feb;70(1):3-18
Rutkowska E, Pajak K, Jóźwiak K
Lipophilicity is a physicochemical property of crucial importance in medicinal chemistry. On the molecular level it encodes information on the network of inter- and intramolecular forces affecting drug transport through lipid structures as well as drug's interactions with the target protein. In result, on the organism level, lipophilicity is an important factor defining pharmacokinetics and pharmacodynamics of a drug substance. Thus, it is a meaningful parameter that found innumerable applications in drug development, Quantitative Structure-Activity Relationships (QSARs) and Quantitative Structure-Pharmacokinetic Relationships (QSPkRs) analyses. This report reviews the importance of lipophilicity on each step of the presence of a medicinal substance in the organism and describes progress in experimental methods of its determination. It has been documented that the retention of a compound in reversed-phase liquid chromatography is governed by its lipophilicity and shows significant correlation with n-octanol/water partition coefficient. Hence, reversed phase chromatography may provide relevant information about the compounds' property. Elaboration of biomimetic stationary phases provides better insight into biological partition processes. Nowadays, there is an urgent need for both precise and quick procedures for quantification of molecular lipophilicity.
X-ray crystallographic structures as a source of ligand alignment in 3D-QSAR
J Chem Inf Model. 2013 Jun 24;53(6):1406-14. doi: 10.1021/ci400004e. Epub 2013 Jun 5
Urniaż RD, Jóźwiak K
Three-dimensional quantitative structure-activity relationships (3D-QSAR) analyses are methods correlating a pharmacological property with a mathematical representation of a molecular property distribution around three-dimensional molecular models for a set of congeners. 3D-QSAR methods are known to be highly sensitive to ligand conformation and alignment method. The current study collects 32 unique positions of congeneric ligands co-crystallized with the binding domain of AMPA receptors and aligns them using protein coordinates. Thus, it allows for a unique opportunity to consider a ligands' orientation aligned by their mode of binding in a native molecular target. Comparative molecular field analysis (CoMFA) models were generated for this alignment and compared with the results of analogous modeling using standard structure-based alignment or obtained in docking simulations of the ligands' molecules. In comparison with classically derived models, the model based on X-ray crystallographic studies showed much better performance and statistical significance. Although the 3D-QSAR methods are mainly employed when crystallographic information is limited, the current study underscores the importance that the selection of inappropriate molecular conformations and alignment methods can lead to generation of erroneous models and false conclusions.
Role of the nicotinic receptor beta4 subunit in the antidepressant activity of novel N,6-dimethyltricyclo[188.8.131.52²,⁶]decan-2-amine enantiomers
Neurosci Lett. 2013 Oct 11;553:186-90. doi: 10.1016/j.neulet.2013.08.036. Epub 2013 Aug 27
Targowska-Duda KM, Jozwiak K, Arias HR
The role of the nicotinic receptor β4 subunit in the antidepressant activity of N,6-dimethyltricyclo[184.108.40.206(2,6)]decan-2-amine enantiomers was investigated using wild-type (β4+/+) and knockout (β4-/-) mice. Mice were injected (i.p.) with saline (control) or with either enantiomer (1.0mg/kg base drug) daily for the first two weeks. Forced swim tests (FST) were performed on Day 1 to determine the acute effect of each enantiomer, and on Day 7 and 14, to determine the chronic activity. To examine the remnant effects after drug treatment, a withdrawal period of two more weeks was continued with FSTs performed on Day 21 and 28. Our results indicate that: (1) the acute antidepressant effect elicited by the (S,S)-enantiomer is observed in β4+/+ mice from both sexes, whereas the effect elicited by the (R,R)-enantiomer is only observed in male β4+/+ mice. There is no antidepressant effect for both novel compounds on male and female β4-/- mice, (2) the chronic antidepressant effect elicited by both enantiomers is observed in β4+/+, but not in β4-/-, mice from both sexes, and (3) the residual antidepressant effect mediated by each enantiomer after withdrawal was observed only in female β4+/+ mice. Our results clearly indicate that β4-containing AChRs are targets for the antidepressant activity of these compounds, and that this activity is gender-dependent.
Molecular interactions between fenoterol stereoisomers and derivatives and the beta2-adrenergic receptor binding site studied by docking and molecular dynamics simulations
J Mol Model. 2013 Nov;19(11):4919-30. doi: 10.1007/s00894-013-1981-y. Epub 2013 Sep 17
Plazinska A, Kolinski M, Wainer IW, Jozwiak K
The β2 adrenergic receptor (β2-AR) has become a model system for studying the ligand recognition process and mechanism of the G protein coupled receptors activation. In the present study stereoisomers of fenoterol and some of its derivatives (N = 94 molecules) were used as molecular probes to identify differences in stereo-recognition interactions between β2-AR and structurally similar agonists. The present study aimed at determining the 3D molecular models of the fenoterol derivative-β2-AR complexes. Molecular models of β2-AR have been developed by using the crystal structure of the human β2-AR T4 lysozyme fusion protein with bound (S)-carazolol (PDB ID: 2RH1) and more recently reported structure of a nanobody-stabilized active state of the β2-AR with the bound full agonist BI-167107 (PDB ID: 3P0G). The docking procedure allowed us to study the similarities and differences in the recognition binding site(s) for tested ligands. The agonist molecules occupied the same binding region, between TM III, TM V, TM VI and TM VII. The residues identified by us during docking procedure (Ser203, Ser207, Asp113, Lys305, Asn312, Tyr308, Asp192) were experimentally indicated in functional and biophysical studies as being very important for the agonist-receptor interactions. Moreover, the additional space, an extension of the orthosteric pocket, was identified and described. Furthermore, the molecular dynamics simulations were used to study the molecular mechanism of interaction between ligands ((R,R')- and (S,S')-fenoterol) and β2-AR. Our research offers new insights into the ligand stereoselective interaction with one of the most important GPCR member. This study may also facilitate the design of improved selective medications, which can be used to treat, prevent and control heart failure symptoms.