N,6-dimethyltricyclo[,6]decan-2-amine Enantiomers Interact with Luminal and Non-luminal Domains at the Human 34 Nicotinic Acetylcholine Receptor



Hugo R. Arias, Katarzyna M. Targowska-Duda, Krzysztof Jozwiak



Hugo R. Arias, Department of Medical Education, College of Medicine, California Northstate University, Elk Grove, CA 95757, USA

Katarzyna M. Targowska-Duda, Krzysztof Jozwiak, Department of Chemistry, Laboratory of Medicinal Chemistry and Neuroengineering, Medical University of Lublin, 20-093 Lublin, Poland

Correspondence to: Hugo R. Arias, Department of Medical Education, California Northstate University College of Medicine, 9700 W. Taron Dr., Elk Grove, CA 95757, USA.

Email: hugo.arias@cnsu.edu

Telephone: +1-916-647-0458           Fax: +1-916-686-7310       

Received: November 22, 2014        Revised: January 20, 2015

Accepted: January 25, 2015

Published online: March 30, 2015



The objective of current study is to characterize the interaction of (S,S)- and (R,R)-N,6-dimethyltricyclo[,6]decan-2-amine enantiomers with human (h) 34 nicotinic acetylcholine receptors (AChRs) in different conformational states by using pharmacological and structural approaches. The pharmacological results established that both enantiomers bind with very low affinity to the [3H]imipramine binding sites at h34 AChRs in the resting and desensitized states (apparent Ki~0.5-0.8 mM). The obtained nH values are lower than unity (~0.5) indicating negative cooperative interactions between imipramine and each enantiomer. Based on these results, we suggest that these interactions are allosteric in nature, and consequently, imipramine and each enantiomer bind to non-overlapping sites. This possibility is supported by molecular docking and dynamics results, indicating that the enantiomers, in the protonated and neutral states, interact with mixed luminal/non-luminal and intersubunit (i.e., non-luminal) domains, both formed between the 3 and 4 transmembrane segments. Based on our finding, we suggest that the previously observed antidepressant-like activity elicited by these enantiomers (Targowska-Duda et al, 2013; Neurosci. Lett. 553, 186-190) may be mediated by a novel negative allosteric modulatory mechanism at the h34 AChR.

© 2015 ACT. All rights reserved.


Key words: Nicotinic acetylcholine receptors; Antidepressants; Allosteric modulation; Conformational states; Molecular modeling; Molecular dynamics


Arias HR, Targowska-Duda KM, Jozwiak K. N,6-dimethyltricyclo [,6]decan-2-amine Enantiomers Interact with Luminal and Non-luminal Domains at the Human 34 Nicotinic Acetylcholine Receptor. Journal of Biochemistry and Molecular Biology Research 2015; 1(1): 19-24 Available from: URL: http://www.ghrnet.org/index.php/jbmbr/article/view/929



An increasing amount of data supports the concept that nicotinic acetylcholine receptors (AChRs) are targets for the clinical activity of different antidepressants[1,2]. Structurally and functionally different antidepressants inhibit various neuronal AChRs by noncompetitive mechanisms[1-5]. In particular, 34 AChRs can be inhibited by different antidepressant-like compounds, including tricyclic antidepressants (TCAs) and mecamylamine[6], serotonin selective reuptake inhibitors (SSRIs)[7], and bupropion[8].

    Recent studies demonstrated that the N,6-dimethyltricyclo [,6]decan-2-amine enantiomers show antidepressant-like activity[9], and interact with luminal and non-luminal binding sites at the human (h) 42 AChR[10]. These novel compounds inhibit 42 and 34 AChRs in the submicromolar concentration range (unpublished data, Targacept, Inc., Winston-Salem, NC, USA). However, we do not have a clear picture of how these compounds interact with 34 AChRs and what are the structural components of its binding site(s).

    AChRs containing the 4-subunit (e.g., 34 and 334) are expressed in the habenulo-interpeduncular pathway in high amounts[11]. These AChR subtypes are involved in drug addiction and mood disorders, among other pathological states[2]. Interestingly, these receptors modulate the antidepressant-like activity of bupropion[12] and nicotine[13]. Thus, to characterize the interaction of N,6-dimethyltricyclo[,6]decan-2-amine enantiomers (Figure 1) with h34 AChRs in different conformational states, structural and functional approaches are applied, including radioligand competition binding assays using [3H]imipramine as a probe for the antidepressant sites[6], as well as molecular docking and molecular dynamics (MD) studies.



Materials and Methods


[3H]Imipramine (47.5 Ci/mmol) was obtained from PerkinElmer Life Sciences Products, Inc. (Boston, MA, USA) and stored in ethanol at -20. The N,6-dimethyltricyclo[,6]decan-2-amine enantiomers were obtained from Targacept, Inc. (Winston-Salem, NC, USA). Imipramine hydrochloride, polyethylenimine, and trypsin/EDTA were purchased from Sigma Chemical Co. (St. Louis, MO, USA). ()-Epibatidine hydrochloride was obtained from Tocris Bioscience (Ellisville, MO, USA). -Bungarotoxin (-BTx) was obtained from Biotoxins Incorporated (St. Cloud, FL, USA). Fetal bovine serum (FBS) was obtained from Aleken Biologicals (Nash, TX, USA). Salts were of analytical grade.


[3H]Imipramine competition binding experiments using h34 AChRs in different conformational states

The effect of (S,S)- and (R,R)-N,6-dimethyltricyclo[,6]decan-2-amine on [3H]imipramine binding to h34 AChRs in different conformational states was determined using membranes prepared from HEK293-h34 cells as described previously[6]. In this regard, HEK293-h34 cells, cultured as described previously[6], express predominantly AChRs with the (3)3(4)2 stoichiometry[14]. AChR membranes (1.5 mg/mL), prepared from these cells, were suspended in binding saline buffer (50 mM Tris-HCl, 120 mM NaCl, 5 mM KCl, 2 mM CaCl2, 1 mM MgCl2, pH 7.4) with 20 nM [3H]imipramine, in the presence of 0.1 M -BTx (resting/-BTx-bound state) or 0.1 M ()-epibatidine (desensitized/epibatidine-bound state), and preincubated for 30 min at room temperature (RT). Bungarotoxins such as -BTx are competitive antagonists that maintain the AChRs in the resting (closed) state[15]. The nonspecific binding was determined in the presence of 100 µM imipramine. The total volume was divided into aliquots, and increasing concentrations of each enantiomer were added to each tube and incubated for 90 min at RT. AChR-bound radioligand was then separated from free ligand by a filtration assay using a 48-sample harvester system with GF/B Whatman filters (Brandel Inc., Gaithersburg, MD, USA), previously soaked with 0.5% polyethylenimine for 30 min. The membrane-containing filters were transferred to scintillation vials with 3 mL of Bio-Safe II (Research Product International Corp, Mount Prospect, IL, USA), and the radioactivity was determined using a Beckman LS6500 scintillation counter (Beckman Coulter, Inc., Fullerton, CA, USA). 

    The concentrationCresponse data were curve-fitted by nonlinear least squares analysis using the Prism software (GraphPad Software, San Diego, CA). The calculated IC50 and Hill coefficient (nH) values were calculated using the following equation. The observed IC50 values from the competition experiments described above were transformed into inhibition constant (Ki) values using the Cheng-Prusoff relationship[16]: Ki = IC50/{1+([[3H]imipramine]/Kdimipramine)} (1)

where [[3H]imipramine] is the initial concentration of [3H]imipramine, and Kdimipramine is the [3H]imipramine dissociation constant for the h34 AChR (0.41 µM;[6]). The apparent Ki and nH values were summarized in table 1.


Homology modeling and molecular docking

Sequence alignment of the Torpedo AChR subunits and the h3 and h4 subunits was first performed using the ClustalW2 server (www.ebi.ac.uk/Tools/msa/clustalw2). The residue numbering of this alignment corresponds to the h3 and h4 sequence from the Protein knowledgebase (i.e., UniProtKB) implemented in the ExPASy Molecular Biology Server[17]. The cryo-electron microscopy structure of the Torpedo AChR determined at ~4 Å resolution (PDB ID: 2BG9)[18] was used as target for molecular modeling. Models of the h34 AChR were subsequently constructed by homology modeling methods using the Torpedo AChR as template, as previously described[6,10,13,19-21]. Modeller 9.9 was used to obtain 100 homology models, and subsequently, their Discrete Optimized Protein Energy profiles[22] were assessed. The best model was subjected to model quality assessment, using the web-based tools of Verify3D[23] and ProCheck[24].

    For the molecular docking procedure, both (S,S)- and (R,R)-enantiomers, in the protonated and neutral forms, were first modeled using HyperChem 6.03 (HyperCube Inc., Gainesville, FL, USA), optimized using the semi-empirical method AM1, and then transferred for the docking simulation as previously described[6,10,13,19-21]. Molegro Virtual Docker (MVD, v 5.0.0, Molegro ApS Aarhus, Denmark) was used for docking simulations of flexible ligands into the rigid target of the AChR model. The docking space was defined to cover all transmembrane domains (TMDs) (i.e., M1-M4), the ion channel lumen, and the extracellular (EXD)-TMD junction. The subsequent docking simulations were performed using the settings previously described[6,10,13,19-21]. The lower energy conformations [i.e., representing the lowest value of the scoring function (MolDockScore)] were selected from each cluster of superposed poses.

Molecular dynamics

To test the stability of each enantiomer in its predicted docking sites, 15-ns MD simulations were performed using the Yasara package (v 11.11.2, Yasara Biosciences, Graz, Austria) and the settings described previously[1,10,13,21].

    The root mean square deviation (RMSD) (see legend from figure 4) values were estimated using the Yasara software, as previously described[10,13]. These values represent the inter-molecular conformational changes and the translation of the whole molecule in the binding site. Poses obtained every 25-ps simulation snapshot were extracted from the simulation trajectory of 15-ns total time and compared to its reference position obtained by docking.




[3H]Imipramine competition binding experiments using h34 AChRs in different conformational states

The binding affinity of each (S,S)- and (R,R)-enantiomer for the imipramine binding sites at the h34 AChR was determined by [3H]imipramine competition binding experiments, when the receptor is in the resting (-BTx-bound) or desensitized (epibatidine-bound) state (Figure 2). The enantiomers displace only ~50% [3H]imipramine binding at the highest concentrations used. The apparent Ki values suggest that both enantiomers bind with very low affinity to the [3H]imipramine binding sites at h34 AChRs in either the resting or desensitized state (Table 1). The observed nH values are lower than unity (close to 0.5) (Table 1), suggesting a negative cooperative interaction between imipramine and each compound.




Molecular docking results

The (S,S)- and (R,R)-enantiomer in the protonated state interacts with mixed luminal/nonluminal and non-luminal binding domains (i.e., intersubunit site) (see 2D and 3D models in figure 3). Since the docking results in the neutral state are the same as that for the protonated state, the results in the neutral state are not shown. The docking results indicate that the mixed luminal/non-luminal (Figure 3C) and intersubunit sites (Figure 3D) for each enantiomer do not overlap the imipramine binding sites. In the mixed luminal/non-luminal site, the enantiomers mainly interact by van der Waals contacts with luminal residues, including 3-Val258 and 4-Leu259 (position 17; non-polar ring), 4-Phe255 and 3-Val254 (position 13; valine ring), as well as with non-luminal residues, comprising 4-Ile263 (position 21), 3-Thr260 (position 19), 4-Ile260 (position 18), 3-Leu257 (position 16), and 4-Phe256 (position 14) (Table 2). In addition, the molecule is stabilized by electrostatic interactions with 3-Glu261 (position 20) at the outer ring.




    In the intersubunit site, each enantiomer interacts predominantly by van der Waals contacts with nonpolar residues at M1 (i.e., 3-Ile219, 3-Pro220, 3-Leu223, 3-Ile224, and 3-Leu227), M2 [i.e., 4-Leu250 (position 8), 4-Thr254 (position 12), 4-Leu258 (position 16), 3-Leu252 (position 10), 3-Phe255 (position 14), 3-Leu259 (position 18)], and M3 (i.e., 4-Ile289) (Table 2).



Molecular dynamics results

The MD results for each protonated enantiomer indicate that the interactions with either the mixed luminal/non-luminal or intersubunit binding site at the h34 AChR are stable during the 15-ns MD simulations (Figure 4). The RMSD values that represent the inter-molecular conformational changes and the translation of the ligand within the binding site were calculated for each studied enantiomer. The higher RMSD value for the (S,S)-enantiomer (Figure 4A) at the mixed luminal/non-luminal binding pocket than that for the (R,R)-enantiomer (Figure 4B) suggests that the (S,S)-enantiomer oscillates within the binding pocket more than the (R,R)-enantiomer. Despite the observed molecular oscillation, each molecule remains stable within the binding site during the 15-ns MD simulation.



In this work, we determined the binding affinity of novel N,6-dimethyltricyclo[,6]decan-2-amine enantiomers for the imipramine sites and the most important structural components of  their binding sites at the h34 AChR.

    The radioligand binding competition results indicate that both enantiomers bind with very low affinity (apparent Ki ~0.5-0.8 mM) to the [3H]imipramine sites, but each enantiomer has opposite preference for the resting or desensitized AChR (Table 1). The observed low affinity is similar to that obtained for the h42 AChR[10] and for mecamylamine-induced inhibition of [3H]imipramine binding to resting and desensitized h34 AChRs, where IC50 ~0.2 mM and nH ~0.5 values were also observed[6]. These results support the notion that the enantiomers interact with the imipramine sites mainly by allosteric mechanisms. Our docking results (Figure 3) are in agreement with the [3H]imipramine binding results showing that the sites for the enantiomers are different to that for imipramine.

    We previously characterized the imipramine and mecamylamine binding sites in the h34 AChR by pharmacologic and molecular docking methods[6]. In that work, we used a docking space centered on the ion channel that ensured covering only the M2 segments from each subunit, including the ion channel. However, increasing evidence indicates that the AChR TMD might be involved in the interaction and function of several allosteric modulators[4,10,13,25-30]. Based on these reports, we used the whole h34-TMD for the searching of potential binding sites for our novel enantiomers. The molecular docking and MD results suggest that the (S,S)- (Figure 3) and (R,R)-enantiomer (data not shown) interacts with two non-overlapping binding sites, the mixed luminal/non-luminal and intersubunit sites. These sites coincide with that previously described at other ion channels. For example, the intersubunit site for the enantiomers overlaps the intrasubunit pocket for desflurane and propofol at the proton-activated ion channel from the bacteria Gleobacter violaceus (GLIC) (i.e., GLIC-Ile202 corresponding to 3-Ile218)[31], two intrasubunit sites for halothane at the 42 AChR (i.e., 2-Leu226 and 2-Ile261 corresponding to 3-Leu227 and 4-Ile263, respectively)[26], the non-luminal site 2 described for (S)-(+)-mecamylamine at the muscle AChR (i.e., 1-Ile219 corresponding to 3-Ile219)[13], and the ketamine and halothane binding sites determined at h7 AChRs (i.e., 7-Ile217 corresponding to 3-Ile219)[27]. On the other hand, the mixed luminal/non-luminal site for the enantiomers overlaps the luminal site 2 determined for (S)-(+)-mecamylamine at the h(4)2(2)3-TMDs (i.e., 2-Lys260 corresponding to 3-Glu261 at position 20)[28].

    Recent studies demonstrated that the N,6-dimethyltricyclo [,6]decan-2-amine enantiomers produce antidepressant-like activity in mice in a gender-dependent manner[10], and interact with luminal and non-luminal binding sites at the h42 AChR[10]. The present work supports two hypotheses: (1) that 4-containing AChRs are involved in the antidepressant-like activity mediated by the (S,S)- and (R,R)-enantiomers[9], and (2) that, as previously suggested for the h42 AChR[10], the interaction of these enantiomers with the intersubunit locus may impede the rotation of the M2 segment, blocking or disrupting the switching of the hydrophobic residues located along the closed ion channel, finally maintaining the receptor in a nonconducting conformation state. In this regard, the antidepressant-like activity elicited by both enantiomers might be associated with novel negative allosteric modulatory mechanisms, where mixed luminal/non-luminal and intersubunit domains play an important role.


Abbreviations: AChR, nicotinic acetylcholine receptor; EXD, extracellular domain; NCA, noncompetitive antagonist; RMSD, root mean square deviation; MD, molecular dynamics; TCAs, tricyclic antidepressants; TMD, transmembrane domain; SSRIs, serotonin selective reuptake inhibitors; -BTx, -bungarotoxin; RT, room temperature; BS, binding saline; Ki, inhibition constant; Kd, dissociation constant; IC50, ligand concentration that produces 50% inhibition of binding; nH, Hill coefficient; DMEM, Dulbecco's Modified Eagle Medium; FBS, fetal bovine serum.



This work was supported the TEAM research subsidy from the Foundation for Polish Science (to K.J.), and from the National Science Center, Poland (SONATA funding, UMO-2013/09/D/NZ7/04549) [to K.T-D. (PI) and H.R.A. (Co-PI)]. Dr. Targowska-Duda was supported by the TEAM program to work at Dr. Arias laboratory.



The Author has no conflicts of interest to declare.



1    Arias HR. Is the inhibition of nicotinic acetylcholine receptors by bupropion involved in its clinical actions? Int J Biochem Cell Biol 2009; 41: 2098C108.

2    Shytle RD, Sheehan D, Sanberg P, Arias HR. Neuronal nicotinic receptors as therapeutic targets for mood disorders. In: Arias HR (Ed), Pharmacology of nicotinic acetylcholine receptors from the basic and therapeutic perspectives. Kerala, India: Research Signpost. 2011: 187-198.

3    Targowska-Duda KM, Arias HR, Jozwiak K, Application of in silico methods to support experimental data: Interactions of antidepressants with nicotinic acetylcholine receptors. Open Conf Proc J 2013; 4: 11-22.

4    Arias HR, Fedorov NB, Benson LC, Lippiello PM, Gatto GJ, Feuerbach D, Ortells MO. Functional and structural interaction of (-)-reboxetine with the human 42 nicotinic acetylcholine receptor. J Pharmacol Exp Ther 2013; 344: 113-23.

5    Arias HR, Biala G, Kruk-Slomka M, Targowska-Duda KM. Interaction of nicotinic receptors with bupropion: Structural, functional, and pre-clinical perspectives. Recept Clin Investig 2014; 1: 38-53.

6    Arias HR, Targowska-Duda KM, Feuerbach D, Sullivan CJ, Maciejewski R, Jozwiak K. Different interaction between tricyclic antidepressants and mecamylamine with the human 34 nicotinic acetylcholine receptor ion channel. Neurochem Int 2010; 56: 642-9.

7    Arias HR, Feuerbach D, Targowska-Duda KM, Russell M, Jozwiak K. Interaction of selective serotonin reuptake inhibitors with neuronal nicotinic acetylcholine receptors. Biochemistry (Mosc) 2010; 49: 5734-42.

8    Fryer JD, Lukas RJ. Noncompetitive functional inhibition at diverse, human nicotinic acetylcholine receptor subtypes by bupropion, phencyclidine, and ibogaine. J Pharmacol Exp Ther 1999; 288: 88-92.

9    Targowska-Duda KM, Jozwiak K, Arias HR. Role of the nicotinic receptor 4 subunit in the antidepressant activity of novel N,6-dimethyltricyclo[,6]decan-2-amine enantiomers. Neurosci Lett 2013; 553: 186-90.

10   Arias HR, Targowska-Duda KM, Jozwiak K. N,6-dimethyltricyclo[,6]decan-2-amine enantiomers interacts with the human 4b2 nicotinic receptor to luminal and non-luminal binding sites. OA Biochemistry 2013; 1: 11

11   Grady SR, Moretti M, Zoli M, Marks MJ, Zanardi A, Pucci L, Clementi F, Gotti C. Rodent habenulo-interpeduncular pathway expresses a large variety of uncommon nAChR subtypes, but only the 34* and 3 34* subtypes mediate acetylcholine release. J Neurosci Off J Soc Neurosci 2009; 29: 2272-82.

12   Radhakrishnan R, Santamara A, Escobar L, Arias HR. The 4 nicotinic receptor subunit modulates the chronic antidepressant effect mediated by bupropion. Neurosci Lett 2013; 555: 68-72.

13   Arias HR, Targowska-Duda KM, Feuerbach D, Jozwiak K. Mecamylamine inhibits muscle nicotinic acetylcholine receptors by competitive and noncompetitive mechanisms. OA Biochemistry 2013; 1: 7.

14   Krashia P, Moroni M, Broadbent S, Hofmann G, Kracun S, Beato M, Groot-Kormelink PJ, Sivilotti LG. Human 34 neuronal nicotinic receptors show different stoichiometry if they are expressed in Xenopus oocytes or mammalian HEK293 cells. PloS One 2010; 5: e13611.

15   Moore MA, McCarthy MP. Snake venom toxins, unlike smaller antagonists, appear to stabilize a resting state conformation of the nicotinic acetylcholine receptor. Biochim Biophys Acta 1995; 1235: 336-42.

16   Cheng Y, Prusoff WH. Relationship between the inhibition constant (K1) and the concentration of inhibitor which causes 50 per cent inhibition (I50) of an enzymatic reaction. Biochem Pharmacol 1973; 22: 3099-108.

17   Gasteiger E, Gattiker A, Hoogland C, Ivanyi I, Appel RD, Bairoch A. ExPASy: The proteomics server for in-depth protein knowledge and analysis. Nucleic Acids Res 2003; 31: 3784-8.

18   Unwin N. Refined structure of the nicotinic acetylcholine receptor at 4Å resolution. J Mol Biol 2005; 346: 967-89.

19   Arias HR, Feuerbach D, Targowska-Duda KM, Russell M, Jozwiak K. Interaction of selective serotonin reuptake inhibitors with neuronal nicotinic acetylcholine receptors. Biochemistry (Mosc) 2010; 49: 5734-42.

20   Arias HR, Rosenberg A, Targowska-Duda KM, Feuerbach D, Jozwiak K, Moaddel R, et al. Tricyclic antidepressants and mecamylamine bind to different sites in the human a4b2 nicotinic receptor ion channel. Int J Biochem Cell Biol 2010; 42: 1007-18.

21   Arias HR, Feuerbach D, Targowska-Duda KM, Aggarwal S, Lapinsky DJ, Jozwiak K. Structural and functional interaction of ()-2-(N-tert-butylamino)-3-iodo-4-azidopropiophenone, a photoreactive bupropion derivative, with nicotinic acetylcholine receptors. Neurochem Int 2012; 61: 1433-41.

22   Eswar N, Webb B, Marti-Renom MA, Madhusudhan MS, Eramian D, Shen MY, Pieper U, Sali A. Comparative protein structure modeling using Modeller. Curr Protoc Bioinformatics John Wiley & Sons, Inc., Supplement 15. 2006; 5.6.1-5.6.30.

23   Eisenberg D, Lthy R, Bowie JU. VERIFY3D: assessment of protein models with three-dimensional profiles. Methods Enzymol 1997; 277: 396-404.

24   Laskowski RA, MacArthur MW, Moss DS, Thornton JM. PROCHECK: a program to check the stereochemical quality of protein structures. J. Appl. Cryst. 1993; 26: 283-291.

25   Young GT, Zwart R, Walker AS, Sher E, Millar NS. Potentiation of 7 nicotinic acetylcholine receptors via an allosteric transmembrane site. Proc Natl Acad Sci U S A 2008; 105:14686-91.

26   Bondarenko V, Mowrey D, Liu LT, Xu Y, Tang P. NMR resolved multiple anesthetic binding sites in the TM domains of the 42 nAChR. Biochim Biophys Acta 2013; 1828: 398-404. doi:10.1016/j.bbamem.2012.09.014.

27   Bondarenko V, Mowrey DD, Tillman TS, Seyoum E, Xu Y, Tang P. NMR structures of the human 7 nAChR transmembrane domain and associated anesthetic binding sites. Biochim Biophys Acta 2014; 1838: 1389C95.

28   Bondarenko V, Targowska-Duda KM, Jozwiak K, Tang P, Arias HR. Molecular interactions between mecamylamine enantiomers and the transmembrane domain of the human 42 nicotinic receptor. Biochemistry (Mosc) 2014; 53: 908-18.

29   Arias HR. Positive and negative modulation of nicotinic receptors. Adv Protein Chem Struct Biol 2010; 80: 153-203.

30   Arias HR. Allosteric modulation of nicotinic acetylcholine receptors. In: Arias HR (Ed). Pharmacology of Nicotinic Acetylcholine Receptors from the Basic and Therapeutic Perspectives. Kerala, India: Research Signpost. 2011: 151-173.

31   Nury H, Van Renterghem C, Weng Y, Tran A, Baaden M, Dufresne V, et al. X-ray structures of general anaesthetics bound to a pentameric ligand-gated ion channel. Nature 2011; 469: 428-31.


Peer reviewer: Chau H. Nguyen, PhD, Assistant Professor, Department of Pharmaceutical Sciences, D'Youville College School of Pharmacy, DAC 440, 320 Porter Avenue, Buffalo, NY, 14201, USA.




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