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Deconstructed analogs of bupropion reveal structural requirements for transporter inhibition versus substrate-induced neurotransmitter release Abdelrahman R Shalabi, Donna Walther, Michael H Baumann, and Richard A. Glennon ACS Chem. Neurosci., Just Accepted Manuscript • DOI: 10.1021/acschemneuro.7b00055 • Publication Date (Web): 21 Feb 2017 Downloaded from http://pubs.acs.org on February 22, 2017
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Deconstructed analogs of bupropion reveal structural requirements for transporter inhibition versus substrate-induced neurotransmitter release
Abdelrahman R. Shalabi,1 Donna Walther,2 Michael H. Baumann,2 and Richard A. Glennon1
1
Department of Medicinal Chemistry, School of Pharmacy, Virginia Commonwealth
University, Richmond, Virginia 23298 USA
2
Designer Drug Research Unit, Intramural Research Program, National Institute on Drug
Abuse, National Institutes of Health, Baltimore, MD 21224, USA
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ABSTRACT Bupropion (1), an α-aminophenone uptake inhibitor at plasma membrane transporters for dopamine (DAT) and norepinephrine (NET), is a widely prescribed antidepressant and smoking cessation aid. Cathinone (2), a structurally simpler α-aminophenone, is a substrate-type releasing agent at the same transporters and a recognized drug of abuse. Our goal was to identify the structural features of α-aminophenones that govern the mechanistic transition from uptake inhibition to substrate-induced release. Deconstructed analogs of 1 were synthesized and compared for their ability to interact with DAT, NET, and the serotonin transporter (SERT) using in vitro assay methods. Bulky amine substituents resulted in compounds that function as DAT uptake inhibitors but not release agents, whereas smaller amine substituents result in relatively nonselective releasing agents at DAT and NET. Our findings add to empirical evidence supporting distinct molecular determinants for α-aminophenone- (i.e., cathinone-) related agents acting as transporter inhibitors versus those acting as releasers.
KEYWORDS: Dopamine transporter (DAT), Norepinephrine transporter (NET), Serotonin transporter (SERT), Cathinone analogs, Synthetic cathinones, Drug abuse
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Introduction Bupropion (1), a widely prescribed antidepressant and smoking cessation aid,1 is an αaminophenone very closely related in chemical structure to the illicit and controlled (U.S. Schedule I) substance cathinone (2) – a central nervous system stimulant. The structural difference between the two α-aminophenones is i) the presence of a tert-butyl amine group and ii) an aromatic meta-chloro substituent, present in the former but not the latter. Bupropion (1) is a non-transported inhibitor of uptake at dopamine (DAT) and norepinephrine (NET) transporters whereas cathinone (2) acts as a substrate at DAT and NET thereby inducing transporter-mediated neurotransmitter release.1,2
HN O
CH3 CH3 CH3 CH3
NH2 O
CH3
Cl 1
2
Based on its relatively selective actions at DAT and NET, bupropion (1) might be expected to possess at least some abuse liability. Indeed, although its abuse does not constitute a major problem, bupropion is the most commonly misused of all available antidepressants.3 The topic of bupropion abuse has been reviewed.3,4 Human subjects who abuse bupropion describe subjective effects as being much like those produced by cocaine.5,6 The euphoria associated with bupropion (1) is observed upon insufflation or intravenous injection of crushed material (i.e., crushed bupropion tablets), but not upon oral administration, suggesting slower kinetics or substantial metabolism following oral administration.4,7 Consequently, the pharmacokinetic character of oral bupropion might
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limit its abuse.7,8 On the other hand, the unavailability of more traditionally-abused stimulants to certain populations (e.g. those in correctional facilities) seems to make bupropion abuse attractive and more prevalent in these situations.4
Because bupropion (1) is a documented DAT/NET uptake inhibitor whereas cathinone (2) is a DAT/NET releasing agent, we “deconstructed” bupropion, piece-by-piece, to determine at what point, structurally, a non-transported uptake inhibitor (i.e., bupropion; 1) is converted to a transportable substrate (i.e., cathinone; 2) that induces transportermediated neurotransmitter release. That is, we addressed what structural features of bupropion (1) make it an uptake inhibitor at DAT/NET as opposed to the simpler αaminophenone cathinone (2), which is a releasing agent. To this end, we prepared and examined specific analogs, each of which bears a single structural modification relative to the parent agent bupropion (1): des-ketobupropion (3), des-chlorobupropion (4), and the N-isopropyl, N-methyl, and primary amine counterparts of bupropion (i.e. 5-7, respectively), that might answer this question.
HN
CH3 CH3 CH3 CH3
HN O
CH3 CH3 CH3 CH3
CH3 HN O
CH3
Cl 3
CH3
HN O
CH3
Cl 4
5
CH3
NH2 O
CH3
Cl
Cl 6
7
This study examined the ability of bupropion (1) and its deconstructed analogs 3-7 to inhibit neurotransmitter uptake and stimulate release at DAT and NET. The action of the compounds at the serotonin transporter (SERT) was also examined for comparison.
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The more behaviorally potent isomer of cathinone (2), S(-)cathinone, was included as a comparator of known pharmacology.
Synthesis Compound 3 was prepared in a one-pot reaction from 1-(3-chlorophenyl)-2-propanone and tert-butylamine using a reductive amination procedure (TiCl4/NaCNBH3) developed by Barney et al.9 for the synthesis of hindered amines. The des-chloro analog 4 was obtained by the simple reaction of α-chloropropiophenone with tert-butylamine using conditions comparable to what we have previously reported for the synthesis of related compounds, and 5 was prepared by a literature method,10 but isolated as its (previously unknown, and more water soluble) HCl salt. The α-methylaminopropiophenone 6 was not known at the time we prepared it (Scheme 1), but was subsequently reported by Blough et al.11 using a somewhat different method of preparation involving an Nprotected amine. Primary amine 7 was obtained by reduction of azide 9 (Scheme 1) which was prepared following the general method of Pulici et al.12 for a related compound. Although 7 has been identified as a possible bupropion metabolite in rabbit liver homogenates by spectroscopic methods,13 this finding has not been confirmed in other animal species,8 and 7 has not been hitherto synthesized or characterized.
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Scheme 1. Synthesis of compounds 6 and 7.a
a
Reagents and conditions: (i) CH3NH2, room temperature, 2 h; (ii) NaN3, room temperature, 20 h; (iii) SnCl2, room temperature, 2 h.
Results Uptake inhibition findings Figure 1 depicts the effects of test compounds on inhibition of uptake at DAT, NET and SERT in rat brain synaptosomes. Table 1 shows potency estimates expressed as IC50 values; IC50 represents the drug concentration required to inhibit uptake by 50%. Consistent with previous reports, bupropion (1) was found to act as a fully efficacious uptake inhibitor at DAT (IC50 = 305 nM). Given differences in species and techniques across previous studies, the DAT potency for bupropion (1) obtained here is comparable to what others have reported: IC50 = 550 nM,14 630 nM,15 and 945 nM.10 It would appear that the various bupropion substituents make only a minor contribution to DAT uptake potency, with compounds 5-7 being equipotent with bupropion (1), and
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compounds 3 and 4 being about three to four times less potent. Carroll et al.10 have previously found that 4 and 5 are half as potent as 1 at inhibiting DAT uptake.
At NET, bupropion (1) was about 10-fold less potent as an uptake inhibitor (IC50 = 3,715 nM) compared to its action at DAT (IC50 = 305 nM; Table 1). Here too, given differences in species and techniques, the NET potency of bupropion we found is reasonably analogous to what has been previously reported: IC50 = 443 nM,10 1,900 nM,14 2,300 nM.15 In general, compounds with bulky terminal amine substituents were more potent as uptake inhibitors at DAT than at NET (i,e., 1, 3-5), whereas compounds with reduced bulk at this position displayed nearly equal potency at DAT and NET (e.g., 6 and 7).
Consistent with prior literature,15,16 bupropion (1) was essentially inactive as a 5-HT uptake inhibitor at SERT (IC50 >10,000 nM; Table 1). Compound 3 was weakly active at SERT while 4 was inactive. Compounds 5-7 displayed efficacious inhibition at SERT but at lower potency when compared to effects at DAT. Finally, the behaviorally more potent isomer of cathinone, S(-)2, was weak at SERT producing only partial inhibition of uptake at 10 µM (Table 1).
Release findings Figure 2 illustrates the effects of test compounds on transporter-mediated release of tritiated substrates at DAT, NET and SERT. Table 1 shows potency estimates as EC50 values; EC50 is the drug concentration required to evoke a release response that is 50% of maximum. Interestingly, bupropion (1) and compounds 3-5 lacked activity as DAT
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releasing agents as demonstrated by the flattened appearance of their release curves. For these experiments, release efficacy 10,000
SERT >10,000 5,113 (467) >10,000
>10,000
DAT
NET
SERT
IAb
IA
IA
IA
IA
IA
IA
IA
IA
IA
IA
a
All compounds were examined as their salts; see experimental section for specific salts used.
b
IA = Inactive as defined by
