Synthesis and Pharmacological Profiling of the ... - ACS Publications

Subscriber access provided by Binghamton University | Libraries

Letter

Synthesis and Pharmacological profiling of the metabolites of synthetic cannabinoid drugs APICA, STS-135, ADB-PINACA, and 5F-ADB-PINACA. Mitchell Longworth, Mark Connor, Samuel D. Banister, and Michael Kassiou ACS Chem. Neurosci., Just Accepted Manuscript • Publication Date (Web): 02 Jun 2017 Downloaded from http://pubs.acs.org on June 3, 2017

Just Accepted “Just Accepted” manuscripts have been peer-reviewed and accepted for publication. They are posted online prior to technical editing, formatting for publication and author proofing. The American Chemical Society provides “Just Accepted” as a free service to the research community to expedite the dissemination of scientific material as soon as possible after acceptance. “Just Accepted” manuscripts appear in full in PDF format accompanied by an HTML abstract. “Just Accepted” manuscripts have been fully peer reviewed, but should not be considered the official version of record. They are accessible to all readers and citable by the Digital Object Identifier (DOI®). “Just Accepted” is an optional service offered to authors. Therefore, the “Just Accepted” Web site may not include all articles that will be published in the journal. After a manuscript is technically edited and formatted, it will be removed from the “Just Accepted” Web site and published as an ASAP article. Note that technical editing may introduce minor changes to the manuscript text and/or graphics which could affect content, and all legal disclaimers and ethical guidelines that apply to the journal pertain. ACS cannot be held responsible for errors or consequences arising from the use of information contained in these “Just Accepted” manuscripts.

ACS Chemical Neuroscience is published by the American Chemical Society. 1155 Sixteenth Street N.W., Washington, DC 20036 Published by American Chemical Society. Copyright © American Chemical Society. However, no copyright claim is made to original U.S. Government works, or works produced by employees of any Commonwealth realm Crown government in the course of their duties.

Page 1 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

ACS Chemical Neuroscience

Synthesis and Pharmacological profiling of the metabolites of synthetic cannabinoid drugs APICA, STS-135, ADB-PINACA, and 5F-ADB-PINACA

Mitchell Longworth,a# Mark Connor,b# Samuel D. Banister,c Michael Kassioua*

a

School of Chemistry, The University of Sydney, NSW 2006, Australia; bDepartment of Biomedical Sciences, Macquarie University, NSW 2109, Australia; cDepartment of Pathology, Stanford University School of Medicine, CA 94305, USA;

*Corresponding author # Equal contribution

1 Environment ACS Paragon Plus


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Keywords: cannabinoid, THC, APICA, ADB-PINACA, metabolism Abstract: Synthetic cannabinoids (SCs) containing a 1-pentyl-1-H substituted indole or indazole have found abuse around the world and are associated with an array of serious side effects. These compounds are known to undergo extensive Phase 1 metabolism after ingestion with little understanding whether these metabolites are contributing to the cannabimimetic activity of the drugs. This work presents the synthesis and pharmacological characterization of the major metabolites of two high concern SCs; APICA and ADBPINACA. In a fluorometric assay of membrane potential, all metabolites that did not contain a carboxylic acid functionality retained potent activity at both the CB1 (EC50 = 14 – 783 nM) and CB2 (EC50 = 5.5 – 291 nM) receptors regardless of heterocyclic core or 3-carboxamide substituent. Of note were the 5-hydroxypentyl and 4-pentanone metabolites which showed significant increases in CB2 functional selectivity. These results suggest that the metabolites of SCs potentially contribute to the overall pharmacological profile of these drugs.


Page 2 of 26

Page 3 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Introduction: As of July 2015, 96 countries and territories have reported more than 540 novel psychoactive substances (NPSs) to the United Nations Office on Drugs and Crime (UNODC).1 Like the psychoactive phytocannabinoid ∆9-tetrahydrocannabinol (∆9-THC, 1, Fig. 1), many SCs possess activity at cannabinoid type 1 and type 2 (CB1 and CB2) receptors, and the in vitro and in vivo pharmacology of many of these SCs has been explored.2-4 While the metabolism of ∆9-THC is well understood, the rate of emergence of novel SCs impedes identification and pharmacological profiling of SCs and their metabolites. Forensic chemists and toxicologists have incrementally increased our knowledge of SC metabolism and developed methods for identification and quantitation of various SC metabolites,5-8 but attempts to understand the pharmacology of SC metabolites has been less systematic.

Unlike THC, which only contains one psychoactive metabolite (11-OH-THC, 2),9 it is hypothesized that the major oxidative metabolites of SCs retain cannabimimetic activity, contributing to the overall physiological profile of the drugs. JWH-018 (3) was one of the earliest recreational SCs detected,10 and the human urinary metabolites of JWH-018 were reported in 2010 at concentrations up to 83 ng/mL.11,12 Metabolic oxidation of JWH-018 was found to occur on the naphthalene and indole rings, as well as the pentyl chain. The terminally fluorinated analogue, AM-2201 (4), was found to generate common metabolites in some but not all cases. In humans, ω-hydroxy- (5-OH-JWH-018, 5), ω-1-hydroxy- (4-OHJWH-018, 6), and ω-carboxy-JWH-018 (5-COOH-JWH-018, 7) (Fig. 1) were found to be excreted in high concentrations (5 and 6 exclusively as glucuronic acid conjugates) following JWH-018 consumption.13 AM-2201 undergoes oxidative defluorination to give common metabolites 5 and 7, but not 6,14 as well as N-dealkylation to give naphthoylindole 8.15



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 4 of 26

Figure 1. Selected cannabinoids and metabolites.

11

R

O

O

O

9 10

OH N

N

N 1

O

1

4

4

1

OH

5

R

5

O

5

JWH-018 (3; R = H) AM-2201 (4; R = F) 5-OH-JWH-018 (5; R = OH)

4 3

3

3

∆9-THC (1; R = H) 11-OH-THC (2; R = OH)

2

2

2

4-OH-JWH-018 (6)

OH

5-COOH-JWH-018 (7)

O O

O

NH 2

NH

NH

O N N

N

2

2

N H

1

1

3 5

8

4

4 3

R

APICA (9; R = H) STS-135 (10; R = F)

5

R

ADB-PINACA (11; R = H) 5F-ADB-PINACA (12; R = F)

Several phase I metabolites of JWH-018 and AM-2201, including common metabolite 5, possessed high affinity for CB1 receptors and exhibited efficacious agonist activity in vitro.16 Monohydroxylated JWH-018 metabolites also showed high affinity for human CB2 receptors.17 In contrast, the glucuronidated conjugate of 5-OH-JWH-018, a major human JWH-018 metabolite, was found to possess neutral antagonist activity at CB1 receptors.18 Additionally, when compared to ∆9-THC or CP 55,940, JWH-018 and 5-OH-JWH-018 are higher efficacy agonists in an assay of CB2 receptor inhibition of adenylyl cyclase (AC).17

In 2012, following the identification of numerous 3-acylindoles related to JWH-018, APICA (2NE1, SDB-001, 9) was reported as one of the first indole-3-carboxamide SCs, followed by its ω-fluorinated analogue, STS-135 (10) (Fig.1).19 APICA was found to exert potent agonist


Page 5 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


activity at CB1 and CB2 receptors in vitro,20 and to possess cannabimimetic activity in vivo.21 Despite the emergence of newer SCs, APICA and related adamantane-derived SCs persist, and continue to be associated with serious adverse effects.22

The phase I metabolites of APICA and STS-135 have been described.23-24 As expected, oxidation of the N-alkyl group was prominent, and several monohydroxylated metabolites were reported. Hydroxylation of the adamantane group was also extensive. Hydrolysis of the carboxamide occurs for APICA, a transformation that appears general for indole-3carboxamides but not 3-acylindoles,25 and 1-adamantylamine has been proposed as a simple APICA biomarker.26 Interestingly, 1-adamantylamine is itself a bioactive substance; the approved antiviral and anti-Parkinsonian drug amantadine.

ADB-PINACA (11) is a member of the recent, prevalent indazole-3-carboxamide SC class, and new variants continue to appear, including the anticipated 5-fluoro derivative 5F-ADBPINACA (12) (Fig. 1). ADB-PINACA exposure has been associated with severe adverse reactions, including neurotoxicity, cardiotoxicity, and death.27-28 Like APICA, ADBPINACA acts as a potent CB1 and CB2 receptor agonist in vitro and in vivo,29-30 but nothing is known of the pharmacology of its metabolites. The metabolism of closely related ABPINACA (differing by an isopropyl rather than tert-butyl side-chain) and its 5-fluorinated analogue (5F-AB-PINACA) was recently reported,31 enabling prediction of some likely ADB-PINACA metabolites. Another study in human liver microsomes confirmed that ABPINACA metabolism occurs mainly via hydroxylation of the pentyl chain,32 although hydrolysis of the terminal primary amide has also been reported.33



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Several reviews of the metabolism of SCs have appeared, focusing predominantly on methods for metabolite identification and quantitation in various matrices,34-36 however, there is growing interest in understanding the pharmacology of SC metabolites more systematically.37 Methods for the prediction of SC metabolites have shown good agreement with in vitro results, indicating that proactive pharmacological profiling of predicted SC metabolites is feasible for the latest SCs.38-39

The aim of the current work was to develop synthetic routes to several phase I metabolites of APICA, STS-135, ADB-PINACA, and 5F-ADB-PINACA and to characterize the pharmacology of these metabolites at human CB1 and CB2 receptors. Ultimately, we sought to elucidate whether apparent trends in the structure-activity relationships of acylindole SC metabolites translate to newer, prevalent indole- and indazole-3-carboxamide SC derivatives featuring a common 1-pentyl subunit.

Results and discussion: The synthesis of several phase I metabolites of APICA/STS-135 (13−18, Fig. 2) and ADB-PINACA/5F-ADB-PINACA (19−24), including diastereomers 19 and 20, was inspired by our previous synthetic routes to parent structures APICA/STS-135 and ADB-PINACA/5F-ADB-PINACA.29, 40

Figure 2. Representative phase I metabolites of APICA, STS-135, ADB-PINACA, and 5FADB-PINACA.


Page 6 of 26

Page 7 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


O

O

NH

O NH

O NH

OH O

NH

NH O

N

N

N

N OH

N H

O OH

4-OH-APICA (13)

4-CO-APICA (14)

NH

O

5-OH-APICA (15)

O O

O

N

NH

NH 2

OH

NH

NH 2

NH

O

NH 2

O

NH

N

OH

N

N

N

O OH

(4R)-4-OH-ADB-PINACA (19) (4S)-4-OH-ADB-PINACA (20)

18

O O

N

N

3-Ad-OH-APICA (17)

O O

N

N

N

OH

5-COOH-APICA (16)

O

NH 2

NH

N

4-CO-ADB-PINACA (21)

5-OH-ADB-PINACA (22)

O

OH

5-COOH-ADB-PINACA (23)

24

The synthesis of 4-OH-APICA enantiomers 4R-13 and 4S-13, 4-CO-APICA (14), 5-OHAPICA (15), Ad-OH-APICA (17) are shown in Scheme 1. In short, it was envisaged that 1alkylation of the esters of 1H-indole- and 1H-indazole-3-carboxylic acids with suitably protected alkyl bromides 25−27 would be followed by deprotection and chemoselective coupling of the carboxylic acid with the appropriate amines.

A one-pot N-alkylation using the appropriate synthon, followed by addition of trifluoroacetic anhydride to indole (29), provided 30–33, which were subsequently treated with hydroxide to furnish carboxylic acids 34–37. The hydrolysis of the acetal protecting group to yield 30 can likely be attributed to the generation of trifluoroacetic acid in situ. Coupling of 37 to 3hydroxy-1-aminoadamantane with HOBt and EDC provided 17, while 34–36 were reacted with 1-aminoadamantane in the presence of PyBOP to give 14, 38 and 39 respectively. Hydrogenolysis of 38 and 39 afforded (4R)-13 and 15 respectively. Finally, a Mitsunobu esterification of (4R)-13 gave the inverted acetate ester 40, saponification of which afforded the (4S)-13 enantiomer.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 8 of 26

N-Dealkylated APICA metabolite, 18, and 5-COOH-APICA (16) were synthesized from indole-3-carboxylic acid (41), converting it to the corresponding acid chloride by treatment with oxalyl chloride, and reacting with 1-aminoadamantane to give 18. Alkylation of 18 with the methyl ester of 1-bromo-5-pentanoic acid (28), and subsequent hydrolysis upon work-up, provided 16.

Scheme 1. Synthesis of APICA metabolites 13, 14, 15, 16, 17 and 18.a

O

O

CF3

a

b

N H

O

OH

O

NH

d or e

N R

NH

g

N R

N R

N

OH 30: 31: 32: 33:

29

R = (CH 2)3C(O)CH3 R = (CH 2)3CH(OBn)CH3 R = (CH 2)5OBn R = (CH 2) 4CH3

34: 35: 36: 37: 38:

R = (CH 2)3C(O)CH3 R = (CH 2)3CH(OBn)CH3 R = (CH 2)5OBn R = (CH 2) 4CH3 R=H c

1

3 2

O

1

5

4

Br

O

3

= (CH 2) 3C(O)CH 3 = (CH 2) 3CH(OBn)CH3 = (CH 2) 5OBn =H = (CH 2) 4COOH

15

f

g

5

4

OH

Br 2

14: R 39: R 40: R 18: R 16: R

O

OBn

NH

O

O

NH

NH

h 26

25

N 1

5

3

Br

1

OBn 2

4

27

Br

5 2

N

N

O

3

OH

OR

O

4

28

17

(4R)-13 i

a

41: R = OAc (4S)-13: R = H

Reagents and conditions: (a)(i) 25, 26 or 37, NaH, DMF, 0 ºC–rt, 1 h; (ii) (CF3CO)2O, 0 ºC–

rt, 1 h, 60–94%; (b) 3 M aq. NaOH, MeOH, ∆, 14 h, 75–95%; (c) 1-amino-3hydroxyadamantane hydrochloride, EDC, HOBt, DIPEA, DMSO, rt, 14 h, 41%; (d) 1aminoadamantane hydrochloride, PyBOP, DIPEA, DMSO, rt, 2 h, 46–97%; (e) for 48 only; (i) (COCl)2, DMF, 0 °C–rt, 1 h; (ii) 1-aminoadamantane, DIPEA, CH2Cl2, rt, 58%; (f)(i) 28, NaH, DMF, 0 °C–rt; (ii) 2 M aq. NaOH, 1 h, 41%; (g) 10% Pd/C, H2(g), MeOH, rt, 2 h, 94–


Page 9 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


98%; (h) DIAD, PPh3, AcOH, THF, 0 ºC–rt, 16 h, 95%; (i) 1 M aq. NaOH, MeOH, rt, 30 min, 77%.

The

synthesis

of

ADB-PINACA/5F-ADB-PINACA

metabolites

utilized

similar

methodologies described for APICA/STS-135 metabolites. The synthesis of diastereomers (4R)-4-OH-ADB-PINACA (19) and (4S)-4-OH-ADB-PINACA (20), as well as 4-CO-ADBPINACA (21), and 5-OH-ADB-PINACA (22), are depicted in Scheme 2. Regioselective 1alkylation of methyl 1H-indazole-3-methyl carboxylate (42) with the appropriate synthons 25, 26 or 27, afforded 43, (4R)-44, and 45 respectively, followed by ester hydrolysis to furnish the corresponding acids 46, (4R)-47, and 48. In the case of 43, additional reaction time in the presence of aqueous acid was required for acetal hydrolysis. Carboxylic acids 46, (4R)-47, and 48 were coupled with L-tert-leucinamide to give 21, 49, and 50 respectively. Hydrogenolysis of O-benzyl protected alcohols 49 and 50 furnished 20 and 22 respectively. As with the corresponding APICA metabolites, 20 was subjected to a Mitsunobu esterification to invert the alcohol stereocenter yielding acetate 51, the hydrolysis of which provided diastereomer 19.

Scheme 2. Synthesis of ADB-PINACA metabolites 19–22.a



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 10 of 26

O O

O

O N

N H

a

O

OR' c

N N R

O O

NH 2

NH

NH

d

N N R

NH 2

N N

OH 42 b

43: R = (CH 2)3C(O)CH3; R' = CH3 (4R)-44: R = (CH 2) 3CH(OBn)CH3; R' = CH3 45: R = (CH 2)5OBn; R' = CH 3

21: R = (CH 2) 3C(O)CH3 49: R = (CH 2) 3CH(OBn)CH3 50: R = (CH 2) 5OBn

46: R = (CH 2)3CH(O)CH3; R' = H (4R)-47: R = (CH 2) 3CH(OBn)CH3; R' = H 48: R = (CH 2)5OBn; R' = H

22

d

O O

NH

O O

NH 2

NH

NH 2

e N

N

N

N OH

20

a

OR

f

51: R = OAc 19: R = H

Reagents and conditions: (a)(i) 25, 26, or 27, t-BuOK, THF, 0 ºC–rt, 48 h; (ii) ∆, 24 h, 54–

84%; (b)(i) 3 M aq. NaOH, MeOH, ∆, 14 h, 71–90%; (ii) 1 M aq. HCl, MeOH, rt, 54% (43 only); (c) L-tert-leucinamide, EDC, HOBt, DMSO, rt, 14 h, 64–84%; (d) 10% Pd/C, H2(g), MeOH, rt, 2 h, 88–99%; (e) DIAD, PPh3, AcOH, THF, 0 ºC–rt, 16 h, 94%; (f) 1 M aq. NaOH, MeOH, rt, 30 min, 99%.

The synthesis of 5-COOH-ADB-PINACA (23) required an orthogonal protecting group strategy and is depicted in Scheme 3. Indazole-3-carboxylic acid (52) was esterified with benzyl alcohol using DCC to give 53. Regioselective alkylation of 53 with 28 afforded 54, and was followed by hydrogenolysis to give carboxylic acid 55. Coupling acid 55 with L-tertleucinamide afforded 56, the hydrolysis of which furnished metabolite 23.


Page 11 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Analogous to the preparation of 46, (4R)-47, and 48, alkylation of 42 with 1-bromopentane furnished 57, and subsequent hydrolysis gave 58. Coupling of 58 to methyl L-tert-leucinate using PyBOP afforded 59, which was subsequently hydrolyzed to give 24.

Scheme 3. Synthesis of ADB-PINACA/5F-ADB-PINACA metabolites 23 and 24.a O O

O

OR b

N

N

N

N

O 52: R = H 53: R = Bn

c

NH 2

NH

d

N

N H

a

O

OR

O

OR

O

54: R = Bn 55: R = H

56: R = Me 23: R = H

e

O O

O

O N

f

N

N H

g

e

57: R = Me 58: R = H

NH

OR

N

N

42

a

O

OR

N

e

59: R = Me 24: R = H

Reagents and conditions: (a) BnOH, DCC, DMAP, CH2Cl2, r.t., 3 h, 74%; (b)

Br(CH2)4COOCH3, K2CO3, KI, CH3CN, ∆, 24 h, 50%; (c) 10% Pd/C, MeOH, H2(g), r.t., 2 h, 68%; (d) L-tert-leucinamide·HCl, HOBt, EDC, DMF, r.t., 4 h, 73%; (e) 2 M aq. NaOH, MeOH, ∆, 2 h, 67–85%; (f) Br(CH2)4CH3, t-BuOK, THF, 0°C–rt, 48 h, 84%; (g) methyl Ltert-leucinate hydrochloride, PyBOP, DIPEA, DMSO, 0 °C–rt, 2 h, 76%.

The cannabimimetic activity of APICA and ADB-PINACA metabolites at CB1 and CB2 were compared to CP 55,940 and the parent compounds, with the results shown in Tables 1 and 2. AtT-20 neuroblastoma cells were transfected with CB1 or CB2, and the activities of CP



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 12 of 26

55,940, APICA, ADB-PINACA and their respective metabolites were assessed using a fluorometric imaging reading plate reader (FLIPR) membrane potential assay. G proteingated inwardly rectifying K+ channels (GIRK) are expressed endogenously on AtT-20 cells which are activated by co-expressed CB1 and CB2 receptors.4 The effects of 9–24 were compared to CP-55,940, an efficacious CB1 and CB2 agonist, which produced a maximal decrease in fluorescence, corresponding to cellular hyperpolarization, of 26% ± 1% in AtT20-CB1 cells and 32% ± 2% in AtT-20-CB2 cells.

Table 1. Functional activity of CP 55,940, 14, and 16–21 at CB1 and CB2 receptors. Compound

hCB2

hCB1

CB2 sel.*

pEC50 ± SEM

Max ± SEM

pEC50 ± SEM

(EC50, nM)

(%CP 55,940)

(EC50, nM)

7.76 ± 0.05 (18)

-

7.66 ± 0.05 (22)

-

0.8

6.93 ± 0.09 (118)

109 ± 5

7.44 ± 0.09 (37)

92 ± 4

3.2

6.29 ± 0.05 (512)

109 ± 3

6.58 ± 0.20 (264)

113 ± 9

1.9

Max

±

SEM

(%CP 55,940)

CP 55,940

APICA (9)

(4R)-4-OH-APICA (4R-13)


Page 13 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


6.45 ± 0.05 (353)

106 ± 3

6.54 ± 0.07 (291)

109 ± 3

1.2

6.40 ± 0.07 (400)

100 ± 4

8.05 ± 0.11 (8.9)

99 ± 5

45

6.10 ± 0.13 (787)

99 ± 8

8.03 ± 0.11 (9.2)

94 ± 5

86

n.d.

82 (at 30 µM)

6.54 ± 0.16 (291)

89 ± 9

>34

7.63 ± 0.10 (24)

119 ± 5

8.14 ± 0.13 (7.3)

94 ± 4

3.3

(4S)-4-OH-APICA (4S-13)

4-CO-APICA (14)

5-OH-APICA (15)

5-COOH-APICA (16)

3-Ad-OH-APICA (17)



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

5.44 ± 0.23 (3670)

95 ± 16

7.30 ± 0.08 (50)

Page 14 of 26

96 ± 4

73

18 *CB2 selectivity expressed as the ratio of CB1 EC50 to CB2 EC50. n.d. = not determined

In comparison to APICA itself (CB1 EC50 = 118 nM), all APICA metabolites, except for 17 (EC50 = 24 nM), showed reduced activity at CB1. The N-dealkylated metabolite (18, CB1 EC50 = 3673 nM) and terminally carboxylated metabolite (16, CB1 EC50 > 10 µM) showed the most substantial decrease in potency. Despite varying potencies, all APICA metabolites were shown to be highly efficacious CB1 agonists with 17 eliciting a greater response (119%) than CP 55,940.

3-Ad-OH-APICA also proved to be the most potent metabolite at CB2 (EC50 = 7.3 nM). Interestingly, pentyl oxidized metabolites 14 and 15 exhibited potent activity at the CB2 receptor (EC50 = 8.9 and 9.2 nM respectively), exceeding that of APICA (CB2 EC50 = 37 nM) and displaying a substantial increase in selectivity for CB2 (45 and 85 times more selective for CB2 than CB1 respectively). In contrast, 4R-13 and 4S-13 showed lower efficacy (264 and 291 nM respectively) and selectivity (1.9 and 1.2 respectively) for CB2. Despite being less efficacious, 5-COOH-APICA (CB2 EC50 = 291 nM) and dealkylated APICA (CB2 EC50 = 50 nM) also displayed significant selectivity for the CB2 receptor (>34 and 73 respectively). The hyperpolarization produced by each of the APICA metabolites was comparable to CP-55,940, indicating they act as high efficacy agonists at CB2.

ADB-PINACA metabolites showed similar results to their APICA counterparts. Despite showing decreased potency compared to the parent compound (CB1 EC50 = 1.3 nM),


Page 15 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


metabolites 19–22, arising from oxidation of the pentyl chain, retained potent activity at CB1 (EC50 = 14–58 nM) while the carboxylic acid metabolites 23 and 24 showed greatly diminished activity (CB1 EC50 = 333 nM and >10 µM respectively). Excluding 5-COOHADB-PINACA (84% maximum response of CP-55,940), all metabolites showed a response comparable to ADB-PINACA at CB1. All metabolites except 5-COOH-ADB-PINACA exhibited similar functional activity (5.5 – 65 nM) and generally an increase in selectivity for CB2 compared to ADB-PINACA. Most notably were 21 and 22 (EC50 = 3.2 and 5.5 nM respectively), which showed a reversal in receptor selectivity compared to the parent compound (5.9 and 11 respectively). Hydroxylated metabolites 19, 20 and 22 proved to be highly efficacious CB2 agonists whereas all other ADB-PINACA metabolites showed submaximal responses in compared to CP-55,940.

The concentration of APICA and ADB-PINACA metabolites generated following ingestion is unknown except for 2328, however, based on the findings that SC metabolites can often be found in amounts in the range of 20-100 ng/ml (corresponding to a concentration of approximately 5-30 nM for the compounds reported here),41 it is likely that some of the metabolites will be exerting significant agonist actions on their own, and these may contribute to persistent intoxication.

Table 2. Functional activity of CP 55,940 and 11, and 19–24 at CB1 and CB2 receptors. Compound

hCB1

hCB2

pEC50 ± SEM

Max ± SEM

pEC50 ± SEM

(EC50, nM)

(%CP 55,940)

(EC50, nM)


Max

CB2 sel.* ±

SEM

(%CP 55,940)


1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Page 16 of 26

7.76 ± 0.05 (18)

-

7.66 ± 0.05 (22)

-

0.8

8.91 ± 0.07 (1.3)

121 ± 5

8.59 ± 0.12 (2.6)

103 ± 6

0.5

7.85 ± 0.05 (14)

113 ± 4

7.73 ± 0.08 (19)

111 ± 4

0.8

7.77 ± 0.04 (17)

116 ± 4

7.89 ± 0.08 (13)

109 ± 4

0.8

7.19 ± 0.11 (65)

123 ± 6

7.98 ± 0.08 (11)

78 ± 3

5.9

CP 55,940

ADB-PINACA (11)

(4R)-4-OH-ADBPINACA (19)

(4S)-4-OH-ADBPINACA (20)

4-CO-ADBPINACA (21)


Page 17 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


7.24 ± 0.08 (58)

116 ± 5

8.26 ± 0.07 (5.5)

96 ± 4

11

>10 µM

84 (at 30 µM)

n.d.

81 (at 30 µM)

n.d.

6.48 ± 0.66 (333)

140 ± 40

7.19 ± 0.11 (65)

91 ± 4

5.1

5-OH-ADBPINACA (22)

5-COOH-ADBPINACA (23)

L-COOH-ADBPINACA (24) *CB2 selectivity expressed as the ratio of CB1 EC50 to CB2 EC50. n.d. = not determined This work presents an effective strategy to the synthesis of a range of N-pentyl oxidized metabolites of newer generation SC designer drugs. These synthetic approaches could be of use to forensic chemists in the synthesis of chemical reference standards of major SCs. All metabolites were also shown to retain some cannabimimetic activity in vitro as shown in a FLIPR membrane potential assay, suggesting they could be contributing to the physiological effects of the drugs.

An intriguing approach to addressing the activity of synthetic

cannabinoids and metabolites is the recently reported luminescent assay of arrestin recruitment by CB receptors expressed in HEK293 cells.42 Different cannabinoids are likely



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

to recruit arrestin more or less well than they couple to inwardly rectifying K channels (the basis of our assay),43 and it is possible that there will be absolute bias for one pathway over the other, but the 2 approaches are complementary, and it would be intriguing to compare the same compounds in the 2 systems.

The preliminary findings from this work suggest that oxidation on the N-pentyl chain regardless of heterocyclic core generally results in a preference for CB2 functionality. Most notably were the 5-hydroxylated and 4-carbonylated metabolites, which showed significant increases in CB2 selectivity for both series of compounds. It is also apparent the presence of a carboxylic acid at the terminus of the N-pentyl chain, causes a drastic decrease in cannabimimetic activity. These results suggest the N-pentyl oxidized metabolites of high concern SCs potentially contribute to the overall pharmacological and toxicological profiles. Further investigation into the biodistribution of these metabolites should provide further insights as to whether they act in a synergistic manner with their parent compounds.

Supplementary Information: Full experimental detail for all synthesized compounds including 1H and 13C NMR spectra of final compounds are included.

Corresponding Author Michael Kassiou. E-mail: [email protected]

Author Contributions M.K. and M.L. conceived and designed the project. M.L. and S.D.B carried out the organic synthesis and analyzed all the compounds. M.C. carried out and analyzed the in vitro experiments. M.L. and S.D.B. wrote the manuscript with the help of M.C. and M.K. 18 Environment ACS Paragon Plus

Page 18 of 26

Page 19 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


Notes The authors declare no competing financial interest.

Acknowledgments Work was supported by NHMRC Project Grant 1107088 awarded to M.K. and M.C. The authors thank Dr. Nick Proschogo for HRMS measurements.

Abbreviations SC, synthetic cannabinoid; CB1, cannabinoid receptor 1; CB2, cannabinoid receptor 2; NPS, novel psychoactive substance; UGT, uridine diphosphate-glucuronosyltransferase; AC, adenylate

cyclase;

HOBt,

dimethylaminopropyl)carbodiimide;

hydroxybenzotriazole; PyBOP,

EDC,

1-ethyl-3-(3benzotriazol-1-yl-

oxytripyrrolidinophosphonium hexafluorophosphate; DMF, dimethylformamide; DIPEA, diisopropylethylamine; DMSO, dimethylsulfoxide; THF, tetrahydrofuran; DIAD, diisopropyl azodicarboxylate; DCC, N,N’-dicyclohexylcarbodiimide; FLIPR, fluorometric imaging reading plate reader; GIRK, G protein-gated inwardly rectifying K+ channels.

References 1.

Qian, Z.; Hua, Z.; Liu, C.; Jia, W., Four types of cannabimimetic indazole and indole

derivatives, ADB-BINACA, AB-FUBICA, ADB-FUBICA, and AB-BICA, identified as new psychoactive substances. Forensic Toxicol 2016, 34, 133-143.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

2.

Wiley, J. L.; Marusich, J. A.; Martin, B. R.; Huffman, J. W., 1-Pentyl-3-

phenylacetylindoles and JWH-018 share in vivo cannabinoid profiles in mice. Drug Alcohol Depen. 2012, 123 (1-3), 148-53. 3.

Gatch, M. B.; Forster, M. J., Delta9-Tetrahydrocannabinol-like effects of novel

synthetic cannabinoids found on the gray market. Behav Pharmacol 2015, 26 (5), 460-8. 4.

Banister, S. D.; Longworth, M.; Kevin, R.; Sachdev, S.; Santiago, M.; Stuart, J.;

Mack, J. B.; Glass, M.; McGregor, I. S.; Connor, M.; Kassiou, M., Pharmacology of Valinate and tert-Leucinate Synthetic Cannabinoids 5F-AMBICA, 5F-AMB, 5F-ADB, AMBFUBINACA, MDMB-FUBINACA, MDMB-CHMICA, and Their Analogues. ACS Chem Neurosci 2016, 7 (9), 1241-54. 5.

Wohlfarth, A.; Scheidweiler, K. B.; Chen, X.; Liu, H.-f.; Huestis, M. A., Qualitative

Confirmation of 9 Synthetic Cannabinoids and 20 Metabolites in Human Urine Using LCMS/MS and Library Search. Anal. Chem. 2013, 85 (7), 3730-3738. 6.

Jang, M.; Shin, I.; Kim, J.; Yang, W., Simultaneous quantification of 37 synthetic

cannabinoid metabolites in human urine by liquid chromatography-tandem mass spectrometry. Forensic Toxicology 2015, 33 (2), 221-234. 7.

Watanabe, S.; Kuzhiumparambil, U.; Winiarski, Z.; Fu, S., Biotransformation of

synthetic cannabinoids JWH-018, JWH-073 and AM2201 by Cunninghamella elegans. Forensic Sci Int 2016, 261, 33-42. 8.

Diao, X.; Huestis, M. A., Approaches, Challenges, and Advances in Metabolism of

New Synthetic Cannabinoids and Identification of Optimal Urinary Marker Metabolites. Clin Pharmacol Ther 2016. 9.

Maurer, H. H.; Sauer, C.; Theobald, D. S., Toxicokinetics of drugs of abuse: current

knowledge of the isoenzymes involved in the human metabolism of tetrahydrocannabinol, cocaine, heroin, morphine, and codeine. Ther Drug Monit 2006, 28 (3), 447-53.


Page 20 of 26

Page 21 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


10.

Auwarter, V.; Dresen, S.; Weinmann, W.; Muller, M.; Putz, M.; Ferreiros, N., 'Spice'

and other herbal blends: harmless incense or cannabinoid designer drugs? J. Mass Spectrom. 2009, 44 (5), 832-837. 11.

Sobolevsky, T.; Prasolov, I.; Rodchenkov, G., Detection of JWH-018 metabolites in

smoking mixture post-administration urine. Forensic Science International 2010, 200 (1–3), 141-147. 12.

Moran, C. L.; Le, V.-H.; Chimalakonda, K. C.; Smedley, A. L.; Lackey, F. D.; Owen,

S. N.; Kennedy, P. D.; Endres, G. W.; Ciske, F. L.; Kramer, J. B.; Kornilov, A. M.; Bratton, L. D.; Dobrowolski, P. J.; Wessinger, W. D.; Fantegrossi, W. E.; Prather, P. P.; James, L. P.; Radominska-Pandya, A.; Moran, J. H., Quantitative Measurement of JWH-018 and JWH-073 Metabolites Excreted in Human Urine. Analytical chemistry 2011, 83 (11), 4228-4236. 13.

Chimalakonda, K. C.; Moran, C. L.; Kennedy, P. D.; Endres, G. W.; Uzieblo, A.;

Dobrowolski, P. J.; Fifer, E. K.; Lapoint, J.; Nelson, L. S.; Hoffman, R. S.; James, L. P.; Radominska-Pandya, A.; Moran, J. H., Solid-phase extraction and quantitative measurement of omega and omega-1 metabolites of JWH-018 and JWH-073 in human urine. Anal Chem 2011, 83 (16), 6381-8. 14.

Chimalakonda, K. C.; Seely, K. A.; Bratton, S. M.; Brents, L. K.; Moran, C. L.;

Endres, G. W.; James, L. P.; Hollenberg, P. F.; Prather, P. L.; Radominska-Pandya, A.; Moran, J. H., Cytochrome P450-mediated oxidative metabolism of abused synthetic cannabinoids found in K2/Spice: identification of novel cannabinoid receptor ligands. Drug Metab Dispos 2012, 40 (11), 2174-84. 15.

Sobolevsky, T.; Prasolov, I.; Rodchenkov, G., Detection of urinary metabolites of

AM-2201 and UR-144, two novel synthetic cannabinoids. Drug Test Anal 2012, 4 (10), 74553.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

16.

Page 22 of 26

Brents, L. K.; Reichard, E. E.; Zimmerman, S. M.; Moran, J. H.; Fantegrossi, W. E.;

Prather, P. L., Phase I hydroxylated metabolites of the K2 synthetic cannabinoid JWH-018 retain in vitro and in vivo cannabinoid 1 receptor affinity and activity. PLoS One 2011, 6 (7), e21917. 17.

Rajasekaran, M.; Brents, L. K.; Franks, L. N.; Moran, J. H.; Prather, P. L., Human

metabolites of synthetic cannabinoids JWH-018 and JWH-073 bind with high affinity and act as potent agonists at cannabinoid type-2 receptors. Toxicol Appl Pharmacol 2013, 269 (2), 100-8. 18.

Seely, K. A.; Brents, L. K.; Radominska-Pandya, A.; Endres, G. W.; Keyes, G. S.;

Moran, J. H.; Prather, P. L., A major glucuronidated metabolite of JWH-018 is a neutral antagonist at CB1 receptors. Chem. Res. Toxicol. 2012, 25 (4), 825-827. 19.

Langer, N.; Lindigkeit, R.; Schiebel, H. M.; Ernst, L.; Beuerle, T., Identification and

quantification of synthetic cannabinoids in 'spice-like' herbal mixtures: a snapshot of the German situation in the autumn of 2012. Drug Test Anal 2014, 6 (1-2), 59-71. 20.

Banister, S. D.; Wilkinson, S. M.; Longworth, M.; Stuart, J.; Apetz, N.; English, K.;

Brooker, L.; Goebel, C.; Hibbs, D. E.; Glass, M.; Connor, M.; McGregor, I. S.; Kassiou, M., The

synthesis

and

pharmacological

evaluation

of

adamantane-derived

indoles:

cannabimimetic drugs of abuse. ACS Chem. Neurosci. 2013, 4 (7), 1081-1092. 21.

Cannizzaro, C.; Malta, G.; Argo, A.; Brancato, A.; Roda, G.; Casagni, E.; Fumagalli,

L.; Valoti, E.; Froldi, R.; Procaccianti, P.; Gambaro, V., Behavioural and pharmacological characterization of a novel cannabinomimetic adamantane-derived indole, APICA, and considerations on the possible misuse as a psychotropic spice abuse, in C57bl/6J mice. Forensic Sci Int 2016, 265, 6-12.


Page 23 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


22.

McIlroy, G.; Ford, L.; Khan, J. M., Acute myocardial infarction, associated with the

use of a synthetic adamantyl-cannabinoid: a case report. BMC Pharmacol Toxicol 2016, 17 (1), 2. 23.

Sobolevsky, T.; Prasolov, I.; Rodchenkov, G., Study on the phase I metabolism of

novel synthetic cannabinoids, APICA and its fluorinated analogue. Drug Test Anal 2015, 7 (2), 131-42. 24.

Gandhi, A. S.; Wohlfarth, A.; Zhu, M.; Pang, S.; Castaneto, M.; Scheidweiler, K. B.;

Huestis, M. A., High-resolution mass spectrometric metabolite profiling of a novel synthetic designer drug, N-(adamantan-1-yl)-1-(5-fluoropentyl)-1H-indole-3-carboxamide (STS-135), using cryopreserved human hepatocytes and assessment of metabolic stability with human liver microsomes. Drug Test Anal 2015, 7 (3), 187-98. 25.

Davies, B. B.; Bayard, C.; Larson, S. J.; Zarwell, L. W.; Mitchell, R. A.,

Retrospective Analysis of Synthetic Cannabinoid Metabolites in Urine of Individuals Suspected of Driving Impaired. J Anal Toxicol 2016, 40 (2), 89-96. 26.

Ford, L. T.; Berg, J. D., 1-Adamantylamine a simple urine marker for screening for

third generation adamantyl-type synthetic cannabinoids by ultra-performance liquid chromatography tandem mass spectrometry. Ann Clin Biochem 2016. 27.

Monte, A. A.; Bronstein, A. C.; Cao, D. J.; Heard, K. J.; Hoppe, J. A.; Hoyte, C. O.;

Iwanicki, J. L.; Lavonas, E. J., An outbreak of exposure to a novel synthetic cannabinoid. N Engl J Med 2014, 370 (4), 389-90. 28.

Schwartz, M. D.; Trecki, J.; Edison, L. A.; Steck, A. R.; Arnold, J. K.; Gerona, R. R.,

A Common Source Outbreak of Severe Delirium Associated with Exposure to the Novel Synthetic Cannabinoid ADB-PINACA. J Emerg Med 2015, 48 (5), 573-80. 29.

Banister, S. D.; Moir, M.; Stuart, J.; Kevin, R. C.; Wood, K. E.; Longworth, M.;

Wilkinson, S. M.; Beinat, C.; Buchanan, A. S.; Glass, M.; Connor, M.; McGregor, I. S.;



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

Kassiou, M., Pharmacology of Indole and Indazole Synthetic Cannabinoid Designer Drugs AB-FUBINACA, ADB-FUBINACA, AB-PINACA, ADB-PINACA, 5F-AB-PINACA, 5FADB-PINACA, ADBICA, and 5F-ADBICA. ACS Chem Neurosci 2015, 6 (9), 1546-59. 30.

Gatch, M. B.; Forster, M. J., Delta(9)-Tetrahydrocannabinol-like effects of novel

synthetic cannabinoids in mice and rats. Psychopharmacology (Berl) 2016, 233 (10), 190110. 31.

Wohlfarth, A.; Castaneto, M. S.; Zhu, M.; Pang, S.; Scheidweiler, K. B.; Kronstrand,

R.; Huestis, M. A., Pentylindole/Pentylindazole Synthetic Cannabinoids and Their 5-Fluoro Analogs Produce Different Primary Metabolites: Metabolite Profiling for AB-PINACA and 5F-AB-PINACA. AAPS J 2015, 17 (3), 660-77. 32.

Takayama, T.; Suzuki, M.; Todoroki, K.; Inoue, K.; Min, J. Z.; Kikura-Hanajiri, R.;

Goda, Y.; Toyo'oka, T., UPLC/ESI-MS/MS-based determination of metabolism of several new illicit drugs, ADB-FUBINACA, AB-FUBINACA, AB-PINACA, QUPIC, 5F-QUPIC and alpha-PVT, by human liver microsome. Biomed Chromatogr 2014, 28 (6), 831-8. 33.

Thomsen, R.; Nielsen, L. M.; Holm, N. B.; Rasmussen, H. B.; Linnet, K.;

Consortium, I., Synthetic cannabimimetic agents metabolized by carboxylesterases. Drug Test Anal 2015, 7 (7), 565-76. 34.

Elsohly, M. A.; Gul, W.; Wanas, A. S.; Radwan, M. M., Synthetic cannabinoids:

analysis and metabolites. Life Sci 2014, 97 (1), 78-90. 35.

Castaneto, M. S.; Wohlfarth, A.; Desrosiers, N. A.; Hartman, R. L.; Gorelick, D. A.;

Huestis, M. A., Synthetic cannabinoids pharmacokinetics and detection methods in biological matrices. Drug Metab Rev 2015, 47 (2), 124-74. 36.

Namera, A.; Kawamura, M.; Nakamoto, A.; Saito, T.; Nagao, M., Comprehensive

review of the detection methods for synthetic cannabinoids and cathinones. Forensic Toxicol 2015, 33 (2), 175-194.


Page 24 of 26

Page 25 of 26

1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60


37.

Fantegrossi, W. E.; Moran, J. H.; Radominska-Pandya, A.; Prather, P. L., Distinct

pharmacology and metabolism of K2 synthetic cannabinoids compared to Delta(9)-THC: mechanism underlying greater toxicity? Life Sci 2014, 97 (1), 45-54. 38.

Strano-Rossi, S.; Anzillotti, L.; Dragoni, S.; Pellegrino, R. M.; Goracci, L.; Pascali,

V. L.; Cruciani, G., Metabolism of JWH-015, JWH-098, JWH-251, and JWH-307 in silico and in vitro: a pilot study for the detection of unknown synthetic cannabinoids metabolites. Anal Bioanal Chem 2014, 406 (15), 3621-36. 39.

Adams, A. J.; Banister, S. D.; Irizarry, L.; Trecki, J.; Schwartz, M.; Gerona, R.,

"Zombie" Outbreak Caused by the Synthetic Cannabinoid AMB-FUBINACA in New York. N Engl J Med 2017, 376 (3), 235-242. 40.

Banister, S. D.; Stuart, J.; Kevin, R. C.; Edington, A.; Longworth, M.; Wilkinson, S.

M.; Beinat, C.; Buchanan, A. S.; Hibbs, D. E.; Glass, M.; Connor, M.; McGregor, I. S.; Kassiou, M., Effects of Bioisosteric Fluorine in Synthetic Cannabinoid Designer Drugs JWH018, AM-2201, UR-144, XLR-11, PB-22, 5F-PB-22, APICA, and STS-135. ACS Chem Neurosci 2015, 6 (8), 1445-58. 41.

Presley, B. C.; Gurney, S. M.; Scott, K. S.; Kacinko, S. L.; Logan, B. K., Metabolism

and toxicological analysis of synthetic cannabinoids in biological fluids and tissues. Forensic Sci Rev 2016, 28 (2), 103-69. 42.

Cannaert, A.; Storme, J.; Franz, F.; Auwärter, V.; Stove, C. P., Detection and Activity

Profiling of Synthetic Cannabinoids and Their Metabolites with a Newly Developed Bioassay. Analytical Chemistry 2016, 88 (23), 11476-11485. 43.

Ibsen, M. S.; Connor, M.; Glass, M., Cannabinoid CB1 and CB2 Receptor Signaling

and Bias. Cannabis and Cannabinoid Research 2017, 2 (1), 48-60.



1 2 3 4 5 6 7 8 9 10 11 12 13 14 15 16 17 18 19 20 21 22 23 24 25 26 27 28 29 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54 55 56 57 58 59 60

105x50mm (300 x 300 DPI)

ACS Paragon Plus Environment

Page 26 of 26

Synthesis and Pharmacological Profiling of the ... - ACS Publications

Recommend Documents