Synthesis, Molecular Pharmacology, and Structure-Activity Relationships of 3-(Indanoyl)indoles as Selective Cannabinoid Type 2 Receptor Antagonists

Harvey F. Fulo, Amal Shoeib, Christian V. Cabanlong, Alexander H. Williams, Chang-Guo Zhan,

Article

Synthesis, Molecular Pharmacology, and Structure−Activity

Relationships of 3‑(Indanoyl)indoles as Selective Cannabinoid Type

Receptor Antagonists

Paul L. Prather, and Gregory B. Dudley *

Cite This: J. Med. Chem. 2021, 64, 6381 −6396

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sı Supporting Information

ABSTRACT: Synthetic indole cannabinoids characterized by a 2′,2′-dimethylindan-5′-oyl

group at the indole C3 position constitute a new class of ligands possessing high aﬃnity for

human CB receptors at a nanomolar concentration and a good selectivity index. Starting from

the neutral antagonist 4, the eﬀects of indole core modiﬁcation on the pharmacodynamic proﬁle

of the ligands were investigated. Several N1 side chains aﬀorded potent and CB -selective neutral

antagonists, notably derivatives 26 (R = n-propyl, R = H) and 35 (R = 4-pentynyl, R = H).

Addition of a methyl group at C2 improved the selectivity for the CB receptor. Moreover, C2

indole substitution may control the CB activity as shown by the functionality switch in 35

(

antagonist) and 49 (R = 4-pentynyl, R = CH , partial agonist).

6,47

INTRODUCTION

nociception.

This seemingly contradictory potential for

■

agonists and inverse agonists in similar indications stems from

the ability of the CB receptors to mediate diﬀerent processes

depending on the conditions: recruitment of immune cells or

interference with the action of other chemoattractants.

Neutral antagonists would similarly be of great interest in

this regard. CB neutral antagonists are relatively rare and as

such have received less attention. However, there is some

evidence that CB neutral antagonists may be promising in the

treatment of obesity-associated inﬂammation, insulin resist-

The endocannabinoid system (ECS) is implicated in a large

spectrum of physiological and pathological processes,

especially ones related to the central and peripheral nervous

−10

systems.

Part of the complex intercellular communicating

system are two types of heptahelical G-protein coupled

receptors identiﬁed as cannabinoid receptor type 1 (CB )

and cannabinoid receptor type 2 (CB₂).

1−14

CB is expressed

but can also be

CB , which is

4,15

^p^rⁱ^m^a^rⁱ^l^yⁱⁿ^t^h^e^c^eⁿ^t^r^a^lⁿ^e^r^v^o^u^s^s^y^s^t^e^m1

6−19

found in locations outside the brain.

ance, and nonalcoholic fatty liver disease. Selective CB₂

sometimes called the peripheral cannabinoid receptor, is

receptor modulators are needed across the full spectrum of

functional activities for therapeutic targeting of the ECS.

Medicinal chemistry eﬀorts targeting the ECS have been

complicated by psychoactivity of CB

side eﬀects are associated with drugs of abuse. Many CB

expressed mainly in the cells associated with the immune

1,14

system.

CB receptors are also found in low concentrations

in activated microglia and inﬁltrated macrophages in the

0,21

-receptor agonists. These

brain,

gastrointestinal tract tissues and other cells such as

vascular endothelial cells, cardiomyocytes, and bone cells.

Small-molecule modulators of CB receptor activity are of

interest for therapeutic indications including neurological

disorders, pain, and inﬂammation. CB agonists have potential

agonists developed in academic and pharmaceutical research

labs have been hijacked for abuse in the recreational drug

23−26

market.

The illicit demand for cannabinoid agonists has

been met with regulatory controls on their supply, which

incidentally also restricts legitimate pharmaceutical research.

utility for treating neurological disorders such as reperfusion

35,36

brain injury; amyotrophic lateral and multiple sclerosis;

7,38

37,38

and Alzheimer’s,

diseases.

Parkinson’s,

and Huntington’s

7−39

Received: March 10, 2021

Published: April 23, 2021

CB₂agonists also exhibit anti-inﬂammatory

eﬀects and antinociceptive eﬀects in acute pain, persistent

inﬂammatory pain, postoperative pain, cancer pain, and

,40−42

neuropathic pain,

and treatment of gliomas.

agonists are also of interest for modulating inﬂammatory

and they may aid in cardioprotection

0,43−45

Interestingly, CB inverse

2021 American Chemical Society

381

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CB -selective ligands are attractive for chemotherapeutic

development due in part to the absence of known psychoactive

1,32

side eﬀects,

but many compounds designed to target CB₂

receptors have nonetheless ended up as Schedule I

controlled substances, presumably due to (oﬀ-target) CB₁

interactions.

While marijuana-like psychoactivity is a primary factor

behind the potential for abuse of many ECS modulators, ease

of synthesis is an auxiliary supply-side factor behind the illicit

production and distribution. For example, the simple two-step

synthesis of JWH-018 (Scheme 1) increases its potential for

Scheme 1. Synthesis of JWH-018

Figure 1. Rational design of a novel class of potent cannabinoids.

in Scheme 3. Starting from methyl isobutyrate (5), α-

substitution with 2-ﬂuorobenzyl bromide generates ester 6

(

81%), which on reduction with DIBAL-H is converted to

alcohol 7 in 88% yield. The alkyl iodide precursor (9) of

indane 10 is obtained through a modiﬁed Finkelstein reaction

in 81% overall yield. The two-step sequence involves

mesylation of the 1° alcohol (7), followed by nucleophilic

displacement with NaI-saturated N-methylpyrrolidone (NMP)

to provide iodide 9.

Reagents and conditions: (a) MeMgBr, Et O/THF, 0 °C, then 1-

naphthoyl Cl, reﬂux. (b) n-Pentyl Cl, KOH, DMSO, 80 °C.

9,52

abuse.

Many 1,3-disubstituted indoles like JWH-018 can

be made from indole and two simple electrophiles (e.g., acyl

and alkyl halides), and many similarly have emerged as drugs

of abuse. Conversely, less synthetically accessible cannabi-

noid ligands are less subject to abuse. Such compounds enable

one to explore the chemotherapeutic potential of ECS

modulators without the same risk of fueling the recreational

drug market.

There is a need for novel ECS modulators, particularly ones

having low potential for abuse. To this end, we designed

ligands that incorporate 2,2-dimethylindane into the known

indole cannabinoid framework. 2,2-Dimethylindanes blend the

benzenoid ring system of the traditional Huﬀman-(JWH)-type

compounds, with the bulky cycloalkane motifs of inﬂuential

CB -selective indole cannabinoids from Abbott. 2,2-Dime-

thylindane carboxylates can be crafted using recent synthetic

methodologies,

commercially available building blocks needed to prepare

many indole cannabinoid drugs of abuse. We expect our

designs to support development of novel ECS modulators

speciﬁcally for legitimate academic and pharmaceutical

research endeavors and without risk of “signiﬁcant diversion”

to illicit markets.

Indane-4-carboxylate 10 is prepared by treating 9 with tert-

butyllithium (t-BuLi), followed by ethyl chloroformate under

carefully controlled conditions. This transformation is

envisioned to proceed by a ﬁve-step, one-pot process, starting

with lithium−iodine exchange and lithiation ortho to the

ﬂuoro substituent to produce a dilithio intermediate. Release of

lithium ﬂuoride generates a benzyne-tethered alkyllithium that

undergoes 5-exo-dig cyclization to the 4-lithiated indane, which

is then trapped with ethyl chloroformate to produce 10.

Meanwhile, our lab was developing a synthesis of diyne

monoester 16 (Scheme 4). Dimedone (12) is condensed

with triﬂic anhydride and reacted with the lithium enolate of

ethyl acetate for a Claisen-type tandem addition/fragmentation

to generate β-keto ester 14, which is then dehydrated to give

16. Rhodium-catalyzed cyclotrimerization of 16 with TMS-

acetylene provides a mixture of regioisomers (17 and 18),

which can be hydrolyzed to 10 and then to 11. Acid 11 is

converted to the acid chloride and then coupled with N-

pentylindole to complete the synthesis of GBD-002 (3).

The synthesis of 3-(indan-5-oyl)indoles also begins with

dimedone (12, Scheme 5) and builds on prior synthetic

5,56

but they are less accessible than the

56,58

methodologies from the Dudley lab.

Vinylogous acyl

Indanoylindoles have not been examined previously as

cannabinoid ligands. The ﬁrst two indanoylindoles we

consideredGBD-002 and GBD-003, Figure 1indeed

exhibit high aﬃnities for both CB receptors, with GBD-003

triﬂate 13 is reduced with DIBAL-H to produce hydroxy-

cycloalkenyl triﬂate 19, which undergoes tandem fragmenta-

tion/oleﬁnation to furnish dienyne 20. Rhodium-catalyzed

cycloisomerization of 20, followed by oxidation with DDQ,

provides 21 in a one-pot process. Saponiﬁcation of 21 gives

acid 22, which is converted to the acid chloride for Friedel−

showing selectivity for CB (Figure 1). Functional assays

showed that GBD-002, like JWH-018, is a full agonist, whereas

positional isomer GBD-003 is a neutral antagonist (Table 2,

Crafts acylation of indole and 2-methylindole (Scheme 4, R =

vide infra). Considering that CB antagonists are promising but

H, Me). The resulting 3-acylindoles (23 and 24) were then N-

alkylated to produce a small collection of indanoylindoles,

designated as GBD-003 thru GBD-031.

We routinely produce gram quantities of indane-5-

carboxylate 21 as a synthetic building block for other projects

in the lab, and thus, it serves for us as a readily available and

convenient starting point for the synthesis of various

indanoylindoles. However, the four-step synthesis eﬀectively

ensures that indole cannabinoids derived from 21 will not be

rare, we focused on preparing a series of indanoylindoles based

on GBD-003 for preliminary molecular pharmacology and

structure−activity relationship (SAR) studies. Several CB -

selective neutral antagonists are described herein.

RESULTS AND DISCUSSION

Chemistry. We prepared indanoylindoles from carboxylates

0 and 21 (Scheme 2). The synthesis of GBD-002 is outlined

■

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Table 1. Screen of (Dimethylindanoyl)indole Derivatives for Binding to hCB and/or hCB Receptors

displacement of [ H]CP-55,940 (0.2 nM)

1 μM of each compound)

(

compound

R¹

R²

hCB₁

hCB₂

GBD-002 (3)

GBD-003 (4)

Pentyl

pentyl

methyl

propyl

butyl

hexyl

isobutyl

benzyl

4-ﬂuorobenzyl

E-2-pentenyl

Z-2-pentenyl

4-pentenyl

4-pentynyl

2-pentynyl

allyl

methyl

85.7 ± 5.2

93.4 ± 5.0

9.4 ± 3.1

94.1 ± 2.3

99.8 ± 0.9

22.2 ± 6.0

23.9 ± 13.0

90.5 ± 2.5

92.9 ± 1.3

83.6 ± 4.9

83.1 ± 2.4

84.6 ± 3.3

92.3 ± 2.5

86.9 ± 3.9

86.4 ± 5.0

95.2 ± 2.9

92.8 ± 1.6

65.9 ± 2.8

60.6 ± 5.4

56.0 ± 3.6

18.0 ± 10.9

73.9 ± 3.4

85.4 ± 3.9

83.6 ± 3.1

55.0 ± 5.9

51.2 ± 11.1

48.8 ± 0.0

73.7 ± 3.8

82.9 ± 3.4

86.4 ± 3.3

93.1 ± 3.6

30.2 ± 2.7

63.0 ± 6.8

GBD-001 (23)

GBD-004 (25)

GBD-005 (26)

GBD-006 (27)

GBD-007 (28)

GBD-008 (29)

GBD-009 (30)

GBD-010 (31)

GBD-011 (32)

GBD-012 (33)

GBD-013 (34)

GBD-014 (35)

GBD-015 (36)

GBD-016 (37)

GBD-017 (38)

GBD-018 (39)

GBD-020 (40)

GBD-021 (41)

GBD-022 (42)

GBD-023 (43)

GBD-024 (44)

GBD-025 (45)

GBD-026 (46)

GBD-027 (47)

GBD-028 (48)

GBD-029 (49)

GBD-030 (50)

GBD-031 (51)

10.8 ± 1.4

47.4 ± 0.3

82.7 ± 0.6

79.1 ± 3.0

42.2 ± 4.9

69.2 ± 2.3

88.0 ± 1.4

84.2 ± 1.2

60.7 ± 4.8

80.0 ± 3.8

71.0 ± 6.4

55.6 ± 5.7

16.0 ± 5.3

33.4 ± 3.3

19.4 ± 7.7

44.4 ± 1.9

43.5 ± 0.6

53.8 ± 0.8

26.2 ± 1.6

18.1 ± 13.3

38.4 ± 5.0

35.1 ± 3.3

43.0 ± 2.0

38.6 ± 1.8

41.9 ± 2.2

16.4 ± 3.1

51.9 ± 0.8

propargyl

CH CO CH CH

propyl

butyl

pentyl

hexyl

benzyl

4-ﬂuorobenzyl

E-2-pentenyl

Z-2-pentenyl

4-pentenyl

4-pentynyl

2-pentynyl

propargyl

This compound was selected for determination of K . This compound was selected for screening of functional activity.

Table 2. Calculated Binding Free Energies (kcal/mol) Compared to Experimental Binding Free Energy (kcal/mol)

rCB₁

rCB₂

ligand

K_i(nM)

ΔG_P_B

ΔG_e_x_p

ΔG_c_o_r_r

K_i(nM)

ΔG_P_B

ΔG_e_x_p

ΔG_c_o_r_r

ΔΔG_e_x_p

ΔΔG_c_o_r_r

GBD-002

GBD-003

GBD-005

GBD-013

GBD-014

GBD-029

1.46

38.80

2559.00

152.00

325.00

1904.00

−41.92

−34.00

−27.79

−32.25

−31.22

−27.89

−12.13

−10.17

−7.68

−9.36

−8.91

−7.85

−12.65

−10.07

−8.04

−9.50

−9.16

−8.07

0.23

1.36

4.90

130.00

15.60

24.00

62.50

−40.79

−35.81

−31.10

−35.26

−35.93

−34.65

−12.17

−11.41

−9.45

−12.28

−10.66

−9.12

−10.48

−10.70

−10.28

0.04

1.23

1.78

1.36

1.55

2.04

0.37

0.59

1.08

0.98

1.54

2.21

−10.72

−10.46

−9.89

rmsd

0.43

Calculated binding free energies obtained from the MM-PBSA calculations without any empirical correction. The experimental binding free

energy was converted from the experimental K using the thermodynamic equation ΔG = −RT ln (K ). Calculated binding free energies after

exp

empirical correction using eq 1. ΔΔG is the corrected binding free-energy diﬀerence [ΔG(CB R) − ΔG(CB R)]. ΔΔG = −RT ln (K

corr

exp

(

CB R)/K (CB R)) = ΔG (CB R) − ΔG (CB R). The root-mean-square deviation (RMSD) of the corrected ΔG binding energy values versus

exp

the experimental ΔG binding energy values.

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Scheme 2. Overview of the Synthetic Approach

replaced with diﬀerent lipophilic alkyl moieties including

28,30,50,53,59

chains based on other JWH cannabinoids

N-alkyl groups among reported indole cannabinoids such as Z-

-pentenyl, 2-pentynyl, 4-pentynyl, and propargyl. The

and novel

analogue lacking the N-substituent (23) showed little aﬃnity

for either receptor. A similar activity proﬁle was found when

the H was substituted with a methyl group (25). Replacement

with longer straight alkyl chains aﬀorded compounds with

good to excellent CBR aﬃnities, consistent with previously

reported links between chain length and receptor binding in

other synthetic indole cannabinoids. Compounds 26

propyl), 27 (butyl), and 4 (pentyl) exhibited increasing

(

CBR aﬃnity with increasing chain length until lower aﬃnity

was observed for the 6-carbon (28, hexyl) variant.

Introduction of an isobutyl substituent (29) led to poor

aﬃnity for CB (42% displacement) and good aﬃnity and

selectivity for CB₂(83% displacement). Comparison of

isobutyl 29 with its unbranched isomer (27, n-butyl) and

desmethyl analogue (26, n-propyl) suggests that this branching

cost-competitive with ones that have been co-opted by the

recreational drug market (e.g., JWH-018) and thus are unlikely

to be subject to widespread abuse as recreational drugs.

Receptor Binding Aﬃnity and SAR. The indanoylin-

doles thus prepared are high-aﬃnity cannabinoid ligands.

Initially, a screen to detect compounds with aﬃnity for human

CB (hCB ) and human CB (hCB ) receptors was conducted

disfavors CB receptor binding and enhances CB selectivity.

Compounds 30 (benzyl) and 31 (4-ﬂuorobenzyl) bearing

aromatic rings in the N-alkyl substituents displayed CBR

aﬃnities comparable to those of the aliphatic counterparts 27

(

butyl) and 28 (hexyl) but less than that of 4 (pentyl),

suggesting some steric tolerance of the receptors. Selectivity for

binding to either CB or CB receptors was modest at best in

these benzyl variants.

by determining the ability of a relatively high concentration (1

μM) of each compound to displace a 0.2 nM concentration of

the well-characterized nonselective CB /CB cannabinoid

radioligand [ H]CP-55,940 from its binding site (Table 1).

Under these conditions, the concentration of a compound

producing 50% displacement would approximate the aﬃnity

Replacement of the alkyl chain with a more hydrophilic

group has a profound eﬀect on the pharmacology of the

ligands. Compound 39, punctuated with an ester group of a

(

e.g., K ) for the receptor being examined, with higher

similar length [-CH

pentyl, showed a complete loss of aﬃnity and selectivity (only

19 and 18% displacement at CB and CB , respectively). This

observation is consistent with the results reported by Hynes et

2 2 3

C(O)OCH CH ] as compared with n-

displacement associated with greater aﬃnity for the CB

receptors. Compounds producing less than 50% displacement

are predicted to exhibit low aﬃnity (less than 1 μM). Selected

compounds showing promising aﬃnity and/or selectivity were

al., Frost et al., and Longworth et al. for fenchyl amide

indoles, cyclopropanecarbonyl indoles, and adamantyl amide

indoles, respectively, in which polar N1 side chains generally

selected for determination of K and selectivity index (SI)

(Figure 2) and/or were screened for functional activity (vide

infra; Table 3).

cause a decrease in CB

pentyl analogue.

receptor binding relative to the n-

GBD-002 and GBD-003 produced almost 100% displace-

ment of the radioligand from both hCB and hCB receptors in

Another set of compounds was synthesized to evaluate the

eﬀect of unsaturation in the alkyl chain, starting with

unsaturated versions of the three- and ﬁve-carbon straight

initial binding screens (Table 1) and (as predicted from these

initial screens) were subsequently determined to exhibit high

aﬃnity for cannabinoid receptors (CBRs) (Figure 2).

Speciﬁcally, GBD-002 exhibited a K (hCB ) of 1.46 ± 0.49

alkyl chains that appear to provide good CB aﬃnity and

selectivity. Addition of double bonds to the pentyl chain

yielded compounds 32−34, bearing the E-2-pentenyl, Z-2-

pentenyl, and 4-pentenyl moieties, respectively. Although less

potent than the n-pentyl analogue 4, the pentenyl compounds

nM and a K (hCB ) of 1.36 ± 0.52 nM; however, with an SI of

only 1.1 (nonselective). The positional isomer GBD-003 is also

a high-aﬃnity ligand for both receptors (hCB K = 38.8 ± 19.8

nM and hCB K = 4.9 ± 0.2 nM), with improved selectivity for

still retained good receptor aﬃnities, particularly at CB (Table

CB (SI = 7.9). Having determined that GBD-003 binds to

1). The Z-isomer (33) was more selective than the E-isomer

CB receptors in a relatively selective manner, we investigated

(32), and terminal alkene isomer 34 was more potent and

the broader series of indanoylindoles for binding to CB and

CB -selective. The alkynyl analogues 35 (4-pentynyl) and 36

CB₂receptors (Table 1). The N1-pentyl chain was ﬁrst

(2-pentynyl) showed slightly lower aﬃnities than the alkenyl

Scheme 3. Synthesis of Benzoate 10, the Precursor to 3-(Indan-4-oyl)indole GBD-002

Reagents and conditions: (a) 2-ﬂuorobenzyl bromide, LDA, THF, −78 °C to rt, 5 h, 81%. (b) DIBAL-H, THF, −78 °C to rt, 1 h, 88%. (c) MsCl,

Et N, CH Cl , 0 °C, 1 h, 91%. (d) NaI, NMP, 140 °C, 8 h, 89%. (e) (i) t-BuLi, n-pentane-Et O (4:1 by vol), −78 °C, 15 min; (ii) THF, −78 °C;

(iii) −78 °C to rt to −78 °C; (iv) ClCO Et, −78 °C (15 min) to rt (15 min).

384

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Scheme 4. Synthesis of the 3-(Indan-4-oyl)indole GBD-002

Reagents and conditions: (a) Tf O, pyridine, CH Cl , −78 °C to rt, 20 min, 97%. (b) (i) lithio ethyl acetate, THF, −78 °C, 20 min; (ii) LHMDS,

°C (10 min) to rt (1 h), 63%. (c) Tf O, aq. LiOH, hexanes, 0 °C to rt, 30 min, 88%. (d) LHMDS, THF, 0 °C to rt, 1 h, 63%. (e) (i)

[

Rh(cod) ]BF , (±)-BINAP, H , CH Cl , rt, 1 h; (ii) TMS-acetylene, CH Cl , rt, 1 h, 90%. (f) KI, TMSCl, H O, CH CN, 60 °C, 12 h, 68%. (g)

2 4 2 2 2 2 2 2 3

KOH, H O, EtOH, rt, 8 h, 95%. (h) (i) SOCl , DMF, CH Cl , rt, 8 h; (ii) N-pentylindole, HFIP, rt, 24 h, 68%.

Scheme 5. Synthesis of 3-(Indan-5-oyl)indoles

Reagents and conditions: (a) Tf O, pyridine, CH Cl , −78 °C to rt, 20 min, 97%. (b) DIBAL-H, THF, −78 °C, 1−2 h, 90%. (c) Ethyl-E-4-

(

diethoxyphosphoryl)but-2-enoate, LHMDS, THF, −78 °C (15 min) to rt to 60 °C (1 h), 82%. (d) (i) [Rh(nbd)Cl] , AgSbF , CH Cl , rt, 1 h;

ii) DDQ, rt, 2 h, 85%. (e) KOH, H O, EtOH, rt, 8 h, 96%. (f) (i) SOCl , DMF, CH Cl , rt, 8 h; (ii) indole or 2-methylindole, HFIP, rt, 24 h, 53−

8%. (g) Alkyl halide, NaH, DMSO, rt, 8 h.

analogues, although terminal alkyne 35 showed promising

aﬃnities (K ) for both CB receptors to determine an SI, as

selectivity for CB . In the three-carbon unsaturated series, the

outlined in Figure 2. Data for JWH-018 were reported

allyl (37) and propargyl (38) analogues exhibited modest CB

aﬃnities displacing only 61 and 56% of [ H]-CP-55,940,

previously and are provided for reference in Figure 1. The

N-pentyl indanoylindoles (3 and 4) were the most potent, and

respectively, compared to the corresponding saturated counter-

part 26 (propyl, 90% displacement).

We next examined N-substitution in the 2-methylindole

series, inspired by the potent CBR ligand WIN-55,212-2. The

N-(4-pentynyl)-2-methylindole 49 was the most CB -selective,

having an hCB K of 62.5 ± 10.0 nM and SI = 30.5.

Molecular Modeling. The six compounds highlighted in

Figure 2 were also examined computationally in combined

molecular docking/molecular dynamics (MD) simulations

(Figure 3), which correlated with experimentally measured

-methylindoles generally showed lower aﬃnity for both

receptors but better CB selectivity compared to the analogues

devoid of the additional substituent. Like the trend in the

binding aﬃnities (K , cf. Table 2 and Figure S1). The good

indole series, CB binding increased with the length of the N1-

correlation between experimental and computational data

suggests that this computational protocol provides the

opportunity to test future designs in silico.

alkyl chain up to 5 carbons (40−43), whereas CB aﬃnity was

largely unaﬀected by these modiﬁcations in chain length. Aung

et al. reported that the presence of a methyl group at C2 of

Binding of GBD-002 with CB R showed a strong hydrogen

the N1-propyl analogue enhanced CB selectivity (JWH-072,

SI = 6 vs JWH-015, SI = 24). However, in this indanoylindole

bond with the nearby S505 via the carbonyl oxygen and π−π

stacking interactions between the indole ring with F200 and

the indane ring with F268 (Figure 3A1). In the CB R model,

the indane ring interacted similar to the CB receptor in the

any apparent advantages in these initial assays, nor did the 2-

pentynyl (50) or propargyl (51) variants. On the other hand,

lipophilic pocket formed by F183, S285, and F87 resulting in

the retention of hydrogen bond (Figure 3A2). Meanwhile, the

nonconserved residues V261 and I110 (substituted with L482

the pentenyl derivatives (46−48) showed an increase in CB

aﬃnity without a substantive eﬀect on CB aﬃnity, leading to

and L193 in the CB receptor, respectively) allowed more

enhanced selectivity, and the 4-pentynyl side chain of 49

showed good aﬃnity and selectivity for CB₂.

Guided by preliminary screening data from Table 1, we

chose a small set of ligands and determined their binding

room for the indole core to relax and rotate, changing the

binding angle in the central transmembrane binding site. The

indane ring of the positional isomer GBD-003 induced a steric

strain in CB R, shifting the moiety closer to the ECL2 domain

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Article

pocket delimited by I271, L276, and Y275 and create eﬀective

van der Waals interactions (Figure 3C1). The truncation of the

lipophilic tail is not as deleterious within CB R as the scaﬀold

is still able to retain the hydrogen bond (Figure 3C2). The

greater loss of CB binding aﬃnity translates into greater CB

selectivity. GBD-013 and GBD-014 have similar binding

modes to GBD-003. Although the three compounds have N-

substituents with similar lengths, the double bond in GBD-013

weakens the hydrophobic interactions in CB R (Figure 3D2)

and more so in CB R (Figure 3D1), resulting in a 10-fold

selectivity. Hydrophobic interactions of the 5 C chain with the

hydrophobic channel in CB R further reduced with the

substitution of a triple bond (GBD-014, Figure 3E1,E2).

2-Methylation of the indole core (GBD-029) positioned the

ligand such that the additional methyl group clashes with L193

in CB R (Figure 3F1). This crowding disrupts the preferred

binding mode and substantially weakens the binding aﬃnity. In

CB R (Figure 3F2), on the other hand, the steric constraint is

less prominent as the rotation of the indole ring places the

methyl group in an open space between transmembrane

helices 5 and 6, thereby limiting the steric strain between the

ligand and protein, aﬀording a higher potency and selectivity

against CB2R.

The experimental and computational binding data are

compiled in Table 2 and depicted graphically in Figure S1.

Experimental binding aﬃnities (K ) were converted into free

energies of binding (ΔG ), and absolute magnitudes of raw

exp

calculated protein binding energies (ΔG ) were adjusted

Figure 2. Aﬃnities (K ) of selected derivatives for hCB and hCB

receptors.

using a standard correction (ΔG ) to align with the

corr

experimental data using eq 1

Table 3. [ S]-GTPγS Binding Assays of Selected and

Reference Compounds for Eﬃcacy at the hCB₂

Cannabinoid Receptors

ΔG_c_o_r_r= 0.3248 × ΔG_P_B+ 1.0477

(1)

The diﬀerence between the CB R and CB R binding

energies deﬁnes selectivity as measured experimentally

compound

[ S]-GTPγS speciﬁc binding (control = 100%)

(

ΔΔG ) and estimated computationally (ΔΔG_c_o_r_r). We see

exp

CP-55,950

AM-630

131 ± 6**

67 ± 4**

134 ± 5*

108 ± 9

99 ± 3

102 ± 4

102 ± 9

87 ± 8

good correlation (R = 0.94) between the calculated (ΔG )

and measured (ΔG ) binding energies (Figure S1, top),

exp

GBD-002 (3)

GBD-003 (4)

GBD-005 (26)

GBD-006 (27)

GBD-007 (28)

GBD-009 (30)

GBD-010 (31)

GBD-011 (32)

GBD-012 (33)

GBD-013 (34)

GBD-014 (35)

GBD-029 (49)

suggesting that SARs can be used to estimate binding aﬃnities

(

and selectivities, Figure S1, bottom) with a reasonable degree

of conﬁdence.

CB R Functional Activity. Several compounds in the

indole series and 49, the most selective in the 2-methylindole

series, were subjected to in vitro pharmacological evaluation in

90 ± 3

a [ S]GTPγS binding assay to evaluate the intrinsic functional

81 ± 2**

106 ± 7

92 ± 2*

96 ± 5

activity. The CBR-mediated G-protein activities of the selected

compounds at CB R as well as reference ligands CP-55,940

(

CB full agonist) and AM-620 (CB inverse agonist) were

measured by examining the ability of a single, high (near

121 ± 8

receptor-saturating) 10 μM concentration to modulate

The results are expressed as the percentages of stimulation of [ S]-

[ S]GTPγS binding in hCB2-transfected CHO-cells as

GTPγS binding (the basal value set at 100%) obtained for a

concentration of ligands of 10 μM. Data are the mean ± standard

error of the mean of three experiments. Statistical signiﬁcance from

previously described.

The molecular geometry of the indanoyl moiety has a

striking eﬀect on hCB R-mediated G-protein activation (Table

00% (basal) as assessed by one-sample t-test (*P < 0.05 and **P <

.01).

). Consistent with other indole cannabinoids, GBD-002

increased [ S]GTPγS binding, indicative of the full agonist

property (134 ± 5% basal). When the indane point of

attachment was moved from C4 (GBD-002) to C5 (GBD-

003), the indanoylindane evoked little if any agonist response

(108 ± 9% basal) compared to the reference full agonist CP-

55,940 (131 ± 6% basal). This lack of signiﬁcant agonist

activity suggests that GBD-003 may act as a neutral hCB₂R

antagonist. Co-incubation of GBD-003 in CHO-hCB2

membranes produced a concentration-dependent reduction

(

Figure 3B1). The shift rotated the carbonyl group away from

S505 which makes hydrogen bonding unfavorable. In CB₂R

however, the loss in aﬃnity was not observed as I110 and V261

still allowed strong π−π stacking interactions with F183 and

hydrogen bonds with S285 (Figure 3B2).

A >60-fold reduction in CB R aﬃnity was observed in GBD-

05, with the propyl tail being too short to occupy the binding

386

Journal of Medicinal Chemistry

J. Med. Chem. 2021, 64, 6381−6396

Article

Figure 3. Binding modes of selected derivatives in CB (A1−F1) and CB (A2−F2) receptors. (A1) GBD-002 (3), CB ; (A2) GBD-002 (3), CB ;

(

B1) GBD-003 (4), CB ; (B2) GBD-003 (4), CB ; (C1) GBD-005 (26), CB ; (C2) GBD-005 (26), CB ; (D1) GBD-013 (34), CB ; (D2) GBD-

1 2 1 2 1

13 (34), CB ; (E1) GBD-014 (35), CB ; (E2) GBD-014 (35), CB ; (F1) GBD-029 (49), CB ; (F2) GBD-029 (49), CB . Original crystal

2 1 2 1 2

structures of CB R (5XR8) and CB R (5ZTY) were obtained from the Protein Data Bank for modeling. Pivotal to the diﬀerence in binding modes

of these compounds between CB R and CB R are the residue changes L193/I110 and L481/V261 (in CB R and CB R, respectively). These

substitutions allow for the central indole scaﬀold of the GBD compounds to rotate, allowing for the dimethylindane moiety and its connecting

ketone to engage in hydrogen bonding with S285. Modiﬁcations to the aliphatic chain such as truncation (C1-2) and dehydrogenation (D1-2, E1-

) reduce hydrophobic interactions with the nearby hydrophobic pockets in CB R and CB R. The resulting indole moiety rotation due to the

1 2

residue changes between CB R and CB R also allows for substitutions (F1−F2) on the indole moiety with CB2R that are otherwise unavailable in

CB R.

in [ S]GTPγS binding elicited by CP-55,940 (Figure 4). The

ability to reverse the response of a full agonist (CP-55,940,

IC₅₀= 1.8 μM, K = 9.7 nM) indicates that GBD-003 is best

characterized as a CB R antagonist.

Other indanoylindoles examined are best broadly charac-

terized functionally as neutral antagonists or perhaps weak

inverse agonists. The N-alkyl derivatives (26−28), similar to

the results from receptor binding studies, suggest that the

length of the alkyl chain does not inﬂuence functional activity.

Arylmethyl derivatives (30, benzyl and 31, p-ﬂuorobenzyl)

reduced [ S]GTPγS binding to 87 ± 8% and 90 ± 3% of

basal, respectively, consistent with weak inverse agonism. Two

of the N-pentenyl isomers (32 and 34) behaved as weak

inverse agonists (81 ± 2% basal and 92 ± 2% basal,

respectively), whereas Z-isomer 33 registered as a weak

agonist/antagonist (106 ± 7% basal) in our screening, and

N-pentynyl 35 insigniﬁcantly altered the basal binding (96 ±

Figure 4. GBD-003 acts as an antagonist at CHO-hCB2 receptors,

producing concentration-dependent reduction of G-protein activation

produced by the hCB2 receptor agonist CP-55,940.

% basal), consistent with an antagonist proﬁle. Interestingly,

the addition of a methyl group to C2 of 35 to produce

387

J. Med. Chem. 2021, 64, 6381−6396

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Article

compound 49 shifted the CB R activity from the antagonist to

the agonist (121 ± 8% basal).

C{ H}NMR (100 MHz, CDCl

): δ 24.71, 38.64, 38.65, 43.64,

51.80, 115.21 (d, JC−F = 20 Hz), 123.64 (d, JC−F = 4 Hz), 124.88 (d,

J_C₋_F= 16 Hz), 128.30 (d, J_C₋_F= 8 Hz), 132.21 (d, J_C₋_F= 5 Hz),

61.46 (d, J

= 244 Hz), 177.79. HRMS (ESI, m/z): for C H FO

C−F 12 16 2

CONCLUSIONS

■

[M + H] calcd, 211.1129; found, 211.1124.

3-(2-Fluorophenyl)-2,2-dimethylpropan-1-ol (7). A mixture of 6

(2.0 g, 9.5 mmol) and THF (43 mL) was ﬁrst cooled and stirred at

We identify (2,2-dimethylindanoyl)indoles as a promising new

class of indole cannabinoids comprising potent and CB₂-

selective ligands and presumably having lower potential for

abuse as recreational drugs compared to other indole

cannabinoids. Similar to the reference compound JWH-018,

GBD-002 (3) and GBD-003 (4) displayed excellent aﬃnities

for both CB receptors. The latter (GBD-003) is moderately

−

78 °C for 15 min. At the same temperature, DIBAL-H (1.0 M in

toluene, 20.9 mL, 20.9 mmol) was then added dropwise. After stirring

for 1 h, the mixture was warmed to ambient temperature and ether

(

21 mL) was added. The solution was quenched in an ice bath with

water (0.8 mL), then 15% NaOH (0.8 mL), and ﬁnally water (2.1

mL). The resulting heterogeneous mixture was removed from the

bath, dried over MgSO , ﬁltered, and concentrated under reduced

selective for CB and, in contrast to most indole cannabinoid

ligands, can be characterized as a neutral antagonist. We

developed a preliminary SAR proﬁle for these new (2,2-

dimethylindanoyl)indoles. The observed patterns on aﬃnities

and selectivity in both the indole and methylindole series are

generally in accordance with a number of previously reported

SAR investigations of indole-derived cannabinoids, with the

major exception of their unusual antagonist functional activity.

The N-alkyl substituent provides opportunities to alter aﬃnity

and selectivity while broadly retaining antagonist activity.

Other changes to the indanoylindole core structure had larger

eﬀects on functional activity, with GBD-029 (2-methylindole)

and GBD-002 (indanoyl positional isomer) exhibiting agonist

activity. The indanoylindole derivatives reported here provide

pressure. Puriﬁcation by automatic ﬂash column chromatography on

silica gel (gradient elution from 5 to 40% EtOAc-hexanes) aﬀorded a

white, waxy solid (71% yield). H NMR (400 MHz, CDCl

): δ 0.90

(

−

s, 6H), 1.69 (br s, 1H), 2.63 (s, 2H), 3.31 (d, J = 8 Hz, 2H), 6.99−

.08 (m, 2H), 7.15−7.22 (m, 2H). FNMR (376 MHz, CDCl ): δ

116.17. C{ H}NMR (100 MHz, CDCl ): δ 24.03, 36.92, 36.93,

7.03, 71.02, 115.25 (d, J_C_‑_F= 23 Hz), 123.67 (d, J_C₋_F= 3 Hz),

25.77 (d, J_C₋_F= 16 Hz), 127.96 (d, J_C₋_F= 8 Hz), 133.13 (d, J_C₋_F= 5

Hz), 161.68 (d, J

= 242 Hz). HRMS (ESI, m/z): for C H FO

C−F

11 14

−

[

M − H] calcd, 181.1034; found, 181.1030.

3-(2-Fluorophenyl)-2,2-dimethylpropyl Methanesulfonate (8).

Triethylamine (4.6 mL, 32.9 mmol) was added to a solution of 7 (4.0

g, 22.0 mmol) and dichloromethane (73 mL). The mixture was

cooled to 0 °C, and then, methanesulfonyl chloride (2.1 mL, 26.3

mmol) was added. After stirring for 1 h, the mixture was warmed to

ambient temperature and diluted with dichloromethane. The mixture

was successively washed with cold water, chilled 10% HCl, saturated

NaHCO , and brine. The organic layer was dried over Na SO and

a new template for future development of CB -selective neutral

antagonists.

EXPERIMENTAL SECTION

Chemistry. All the chemicals were used as received unless

otherwise stated. Diethyl ether (Et O), tetrahydrofuran (THF),

concentrated under reduced pressure, producing a colorless, viscous

■

liquid (91% yield). The product was used immediately in the next

step without puriﬁcation. H NMR (400 MHz, CDCl ): δ 1.00 (s,

methylene chloride (DCM), and toluene were dried over a column of

molecular sieves under argon. All reactions were carried out under an

inert nitrogen atmosphere unless otherwise stated. Crude products

were puriﬁed in a Biotage Isolera One Flash Puriﬁcation System using

Biotage prepacked cartridges (50 μm irregular silica). Yields refer to

isolated material following silica gel chromatography. The purity of

6H), 2.68 (s, 2H), 3.03 (s, 3H), 3.92 (s, 2H), 7.01−7.11 (m, 2H),

7.13−7.25 (m, 2H). FNMR (376 MHz, CDCl ): δ −115.22.

C{ H}NMR (100 MHz, CDCl

76.86, 115.39 (d, JC−F = 23 Hz), 123.75 (d, JC−F = 4 Hz), 124.39 (d,

C−F = 16 Hz), 128.38 (d, JC−F = 8 Hz), 132.80 (d, JC−F = 4 Hz),

161.43 (d, JC−F = 244 Hz).

1-Fluoro-2-(3-iodo-2,2-dimethylpropyl)benzene (9). NaI (16.9

g, 113.0 mmol) was added to a solution of 8 (5.9 g, 22.6 mmol) and

NMP (45 mL). The mixture was heated to 140 °C overnight. After

cooling to ambient temperature, the orange mixture was diluted with

water, followed by extraction with pentane. The organic layer was

): δ 23.84, 35.82, 37.00, 37.11,

the compounds tested was determined by quantitative H NMR using

the absolute internal calibrant method (see the Supporting

Information ). H NMR and C NMR spectra were recorded on a

JEOL 400 MHz spectrometer using CDCl and dimethyl sulfoxide-d

(

DMSO-d ) as the deuterated solvent [≥99.8 atom % D, contains

.03% (v/v) TMS]. The chemical shifts (δ) were reported in parts per

then successively washed with saturated Na 2O , saturated CuSO ,

S 3 4

million (ppm) relative to the internal standard TMS. High-resolution

mass spectroscopy (HRMS) data were obtained on a UHR-TOF

maXis 4G instrument (Bruker Daltonics, Bremen, Germany) using

electrospray ionization (ESI). Melting points (mp) were taken in

open capillaries on a Stuart melting point apparatus SMP11 and are

uncorrected.

water, and brine. Puriﬁcation of the crude product by automatic ﬂash

column chromatography on silica gel (gradient elution from 2 to 20%

EtOAc-hexanes) aﬀorded a colorless, viscous liquid (89% yield). H

NMR (400 MHz, CDCl

6.99−7.10 (m, 2H), 7.19−7.28 (m, 2H). FNMR (376 MHz,

): δ 1.05 (s, 6H), 2.72 (s, 2H), 3.20 (s, 2H),

CDCl ): δ −114.86. C{ H}NMR (100 MHz, CDCl ): δ 24.14,

Methyl 3-(2-Fluorophenyl)-2,2-dimethylpropanoate (6).

26.79, 26.80, 35.28, 38.66, 115.33 (d, J = 23 Hz), 123.64 (d, J_C₋_F=

C‑F

mixture of diisopropylamine (3.0 mL, 21.5 mmol) and THF (98

4 Hz), 125.25 (d, J_C₋_F= 15 Hz), 128.22 (d, J_C₋_F= 8 Hz), 132.44 (d,

J_C₋_F= 5 Hz), 161.43 (d, J_C₋_F= 244 Hz).

Ethyl 2,2-Dimethyl-2,3-dihydro-1H-indene-4-carboxylate (10).

mL) was ﬁrst cooled and stirred at −78 °C for 15 min. Next, n-BuLi

(1.6 M in hexanes, 13.5 mL, 21.5 mmol) was added dropwise via a

syringe. The mixture was stirred for another 15 min at −78 °C, 25

min at 0 °C, and 15 min at −78 °C. Methyl isobutyrate (2.2 mL, 19.6

mmol) was then added at the same temperature. After 30 min, 2-

ﬂuorobenzyl bromide (2.7 mL, 21.5 mmol) was ﬁnally added. The

mixture was stirred at −78 °C for 1 h and then at ambient

temperature for 5 h. After the reaction, the yellow solution was

extracted with ether (3 × 50 mL). The organic layer was washed with

brine, dried over Na SO , and concentrated under reduced pressure.

Method 1: Compound 9 (4.5 g, 15.5 mmol) was ﬁrst dissolved in 155

mL of anhydrous n-pentane-diethyl ether solution (4:1 by volume).

The solution was cooled to −78 °C, after which t-BuLi (1.9 M

solution in pentane, 26.1 mL, 49.5 mmol) was added dropwise. The

solution was stirred at the same temperature for 15 min, and then, dry

THF (7.8 mL) was added, turning the blue solution to a yellow,

heterogeneous mixture. Next, the reaction mixture was warmed to

room temperature (rt), stirred for 30 min, and cooled back to −78

°C. Ethyl chloroformate (15.3 mL, 154.7 mmol) was added dropwise,

and the mixture was stirred for 15 min before warming up to rt and

stirring for another 15 min. The mixture was quenched with water and

extracted with diethyl ether (3 × 100 mL). The organic layer pool was

dried over Na SO and concentrated under reduced pressure,

Puriﬁcation by automatic ﬂash column chromatography on silica gel

gradient elution from 2 to 20% EtOAc-hexanes) aﬀorded a colorless,

(

viscous liquid (84% yield). H NMR (400 MHz, CDCl ): δ 1.19 (s,

H), 2.92 (s, 2H), 3.67 (s, 3H), 6.98−7.06 (m, 2H), 7.07−7.13 (m,

H), 7.16−7.23 (m, 1H). FNMR (376 MHz, CDCl ): δ −115.90.

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J. Med. Chem. 2021, 64, 6381−6396

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Article

producing a brown, viscous liquid which was used in the next step

without puriﬁcation and characterization.

Method 2: A mixture of TMS-esters 17 and 18 (20.3 mg, 0.07

C{ H}NMR (100 MHz, CDCl ): δ 28.03, 33.42, 42.37, 50.58,

116.84, 118.38, 166.09, 197.55.

Ethyl 5,5-Dimethyl-3-oxooct-7-ynoate (14). LHMDS (1.0 M

solution in THF, 9.1 mL, 9.1 mmol) was ﬁrst cooled to −78 °C for 10

min. Ethyl acetate (0.9 mL, 9.1 mmol) was then added, and the

mixture was stirred for 15 min. In a separate ﬂask, a mixture of 13 (2.1

g, 7.6 mmol) in THF (30 mL) was also cooled to −78 °C. To this

mixture, the ethyl lithioacetate solution was added slowly. After 20

min, the mixture was warmed to 0 °C, and then, LHMDS (1.0 M

solution in THF, 22.7 mL, 22.7 mmol) was added dropwise over a

period of 20 min. The resulting red solution was warmed to ambient

mmol), KI (58.0 mg, 0.35 mmol), H O (6 μL, 0.35 mmol), and

acetonitrile (0.7 mL) was mixed in a vial. Trimethylsilyl chloride (44

μL, 0.35 mmol) was then added dropwise, and the mixture was stirred

at 60 °C. After 12 h, the mixture was quenched with saturated

NaHCO and extracted with ethyl acetate. The organic layer pool was

washed brine, dried over Na SO , and concentrated under reduced

pressure. The crude product was puriﬁed by automatic ﬂash column

chromatography on silica gel (isocratic elution: 5% EtOAc-hexanes)

to aﬀord 10 as a colorless, viscous liquid (10.4 mg, 68% yield). H

temperature and stirred for 1 h. Saturated NH Cl was added, and the

mixture was extracted with ether (3 × 20 mL). The organic layer was

washed with brine, dried over Na SO , and concentrated under

NMR (400 MHz, CDCl ): δ 1.15 (s, 6H), 1.39 (t, 3H), 2.74 (s, 2H),

.10 (s, 2H), 4.35 (q, 2H), 7.19 (t, 1H), 7.33 (d, 1H), 7.81 (d, 1H).

C{ H}NMR (100 MHz, CDCl ): δ 14.38, 28.85, 39.52, 47.27,

reduced pressure. The crude product was puriﬁed by automatic ﬂash

8.79, 60.56, 126.08, 127.04, 127.86, 128.86, 145.10, 145.95, 167.25.

column chromatography on silica gel (isocratic elution: 5% EtOAc-

HRMS (ESI, m/z): for C H O [M + H] calcd, 219.1375; found,

hexanes) to aﬀord the β-ketoester as a yellow, viscous liquid (1.0 g,

219.1380.

63% yield). H NMR (400 MHz, CDCl ): δ 1.08 (s, 6H), 1.26 (t,

,2-Dimethyl-2,3-dihydro-1H-indene-4-carboxylic acid (11).

3H), 1.99 (t, 1H), 2.26 (d, 2H), 2.57 (s, 2H), 3.41 (s, 2H), 4.17 (q,

Ester 10 (crude from method 1 or puriﬁed from method 2) was

2H). C{ H}NMR (100 MHz, CDCl ): δ 14.09, 26.93, 31.14, 33.42,

ﬁrst dissolved in ethanol (0.2 M). KOH (2.0 M solution in H O, 3.0

50.95, 51.55, 61.32, 70.54, 81.90, 167.12, 201.92.

eq) was then added, and the mixture was stirred overnight at ambient

temperature. The mixture was then cooled in an ice bath and acidiﬁed

with concentrated HCl until pH 1. The solids were ﬁltered and

washed with cold water. The resulting white powder (Method 1: 17%

Ethyl (Z)-5,5-Dimethyl-3-(((triﬂuoromethyl)sulfonyl)oxy)oct-2-

en-7-ynoate (15). A mixture of 14 (0.97 g, 4.6 mmol) and hexanes

(23 mL) was cooled in an ice bath. Saturated aqueous LiOH (1.5

mL/mmol β-ketoester) was added, and the resulting suspension was

stirred for 5 min. Triﬂic anhydride (1.9 mL, 11.5 mmol) was then

added slowly. The mixture was removed from the bath and stirred for

30 min. After the reaction, the mixture was diluted with water and

extracted with ether (3 × 10 mL). The organic layer was washed with

over two steps, Method 2: 95%) was used in the next step without

further puriﬁcation. H NMR (400 MHz, CDCl ): δ 1.16 (s, 6H),

.76 (s, 2H), 3.16 (s, 2H), 7.23 (t, 1H), 7.39 (d, 1H), 7.90 (d, 1H).

C{ H}NMR (100 MHz, CDCl ): δ 28.95, 39.69, 47.34, 49.04,

brine, dried over Na SO , and concentrated under reduced pressure.

25.90, 126.36, 128.81, 129.99, 145.42, 147.17, 172.84. HRMS (ESI,

−

The crude product was puriﬁed by automatic ﬂash column

chromatography on silica gel (isocratic elution: 5% EtOAc-hexanes)

m/z): for C H O [M − H] calcd, 189.0921; found, 189.0914. mp

decomposes at 277 °C.

2,2-dimethyl-2,3-dihydro-1H-inden-4-yl) (1-pentyl-1H-indol-3-

to aﬀord the triﬂate as a light-yellow, viscous liquid (1.39 g, 88%

(

yl)methanone (GBD-002, 3). A mixture containing 11 (164.8

mg, 0.87 mmol) and dichloromethane (5.1 mL) was cooled to 0 °C.

Thionyl chloride (0.16 mL, 2.2 mmol) was added dropwise, followed

by a catalytic amount of DMF (7 μL, 0.09 mmol). After stirring at

ambient temperature for 8 h, the mixture was concentrated under

reduced pressure and redissolved in 0.6 mL of 1,1,1,3,3,3-

hexaﬂuoroisopropanol (HFIP). The solution was added dropwise to

a mixture of N-pentylindole (486.6 mg, 2.6 mmol) and 0.6 mL of

HFIP. The mixture was stirred at ambient temperature for 24 h and

concentrated under reduced pressure. Puriﬁcation by automatic ﬂash

column chromatography on silica gel (gradient elution from 5 to 40%

yield). H NMR (400 MHz, CDCl

): δ 1.09 (s, 6H), 1.32 (t, 3H),

2.09 (t, 1H), 2.16 (d, 2H), 2.40 (s, 2H), 4.26 (q, 2H), 5.84 (s, 1H).

C{ H}NMR (100 MHz, CDCl ): δ 14.00, 26.72, 31.61, 34.82,

45.00, 61.33, 71.51, 80.85, 115.01, 116.70, 119.88, 156.43, 162.40.

Ethyl 5,5-Dimethylocta-2,7-diynoate (16). To a mixture of 15

(1.2 g, 3.5 mmol) in THF (17 mL) cooled at 0 °C, LHMDS (1.0 M

solution in THF, 3.8 mL, 3.8 mmol) was added. The solution was

warmed to ambient temperature and stirred for 1 h. After the reaction,

saturated NH Cl was added, and the mixture was extracted with ether

(3 × 10 mL). The organic layer was washed with brine, dried over

Na SO , and concentrated under reduced pressure. The crude

EtOAc-hexanes) aﬀorded a colorless oil (68% yield). H NMR (400

product was puriﬁed by automatic ﬂash column chromatography on

silica gel (gradient elution from 2 to 20% EtOAc-hexanes) to aﬀord

the diyne as a light-yellow oil (0.42 g, 63% yield). H NMR (400

MHz, CDCl ): δ 0.88 (t, 3H), 1.13 (s, 6H), 1.26−1.40 (m, 4H), 1.86

(

quint, 2H), 2.78 (s, 2H), 2.91 (s, 2H), 4.12 (t, 2H), 7.19−7.24 (m,

H), 7.29−7.35 (m, 3H), 7.36−7.42 (m, 2H), 7.46 (s, 1H), 8.39−

MHz, CDCl ): δ 1.11 (s, 6H), 1.31 (t, 3H), 2.03 (t, 1H), 2.22 (s,

.44 ppm (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 13.91,

2H), 2.37 (s, 2H), 4.22 (q, 2H). C{ H}NMR (100 MHz, CDCl ): δ

2.21, 28.69, 28.97, 29.56, 40.03, 47.05, 47.14, 47.53, 109.83, 116.34,

22.60, 122.79, 123.41, 125.68, 126.09, 126.56, 127.05, 136.86,

14.04, 26.39, 30.87, 31.13, 34.09, 61.83, 70.76, 75.14, 81.29, 86.70,

153.72.

Ethyl 2,2-Dimethyl-5-(trimethylsilyl)-2,3-dihydro-1H-indene-4-

37.32, 137.61, 142.78, 144.92, 191.88. HRMS (ESI, m/z): for

C H NO [M + H] calcd, 360.2322; found, 360.2317. Purity

carboxylate (17/18). In a vial, [Rh(COD)

mmol), (±)-BINAP (24.6 mg, 0.04 mmol), and CH

]BF

(16.2 mg, 0.04

Cl (7.9 mL)

were stirred for 1 h at ambient temperature under a H atmosphere.

The mixture was concentrated and redissolved in CH Cl (2.0 mL).

(

qNMR), 100.4%.

,5-Dimethyl-3-oxocyclohex-1-en-1-yl triﬂuoromethanesulfo-

nate, VAT (13). Pyridine (11.9 mL, 146.8 mmol) was added to a

suspension of dimedone (10.3 g, 73.4 mmol) in dry dichloromethane

TMS-acetylene (0.25 mL, 1.7 mmol) was added, followed by a

(

367 mL) at ambient temperature. The mixture was cooled to −78 °C

solution of diyne 16 (152.0 mg, 0.79 mmol) in CH Cl (7.9 mL).

and stirred for 10 min. Triﬂic anhydride (13.6 mL, 80.7 mmol) was

then added slowly. After 20 min, the mixture was removed from the

bath and warmed to ambient temperature. The mixture was quenched

with 1 M HCl after complete consumption of the starting material

and extracted with diethyl ether (3 × 50 mL). The organic layer pool

was washed with saturated Na CO and brine, dried over Na SO ,

After stirring for 1 h at ambient temperature, the mixture was

concentrated and puriﬁed by automatic ﬂash column chromatography

on silica gel (isocratic elution: 2% EtOAc-hexanes) to aﬀord a mixture

of TMS-esters as a colorless, viscous liquid (206 mg, 90% yield). H

NMR (400 MHz, CD CO): ortho isomer: δ 0.29 (s, 9H), 1.14 (s,

6H), 1.40 (t, 3H), 2.73 (s, 2H), 2.91 (s, 2H), 4.36 (q, 2H), 7.25 (d,

1H), 7.42 (d, 1H). Meta isomer: δ 0.28 (s, 9H), 1.15 (s, 6H), 1.40 (t,

3H), 2.74 (s, 2H), 3.09 (s, 2H), 4.36 (q, 2H), 7.47 (d, 1H), 7.95 (d,

and concentrated under reduced pressure. The crude product was

puriﬁed by automatic ﬂash column chromatography on silica gel

(gradient elution from 2% to 20% EtOAc-hexanes) to aﬀord VAT as a

1H). C{ H}NMR (100 MHz, CDCl ): δ 1.38, 15.37, 15.45, 29.90,

colorless, viscous liquid (19.3 g, 97% yield). H NMR (400 MHz,

CDCl ): δ 1.14 (s, 6H), 2.32 (s, 2H), 2.55 (s, 2H), 6.08 (s, 1H).

29.96, 40.42, 40.43, 48.28, 48.49, 49.30, 49.99, 61.58, 61.90, 127.60,

127.67, 133.91, 134.32, 134.65, 135.22, 138.57, 139.19, 144.47,

389

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

Article

45.51, 146.68, 147.62, 168.62, 170.96. HRMS (ESI, m/z): for

NMR (400 MHz, CDCl ): δ 1.16 (s, 6H), 2.77 (s, 4H), 7.25 (d, 1H),

C H O Si [M + H] calcd, 291.1768; found, 291.1775.

7.89−7.93 (m, 2H). C{ H}NMR (100 MHz, CDCl ): δ 28.69,

-Hydroxy-5,5-dimethyl-cyclohex-1-en-1-yl Triﬂuoromethane

40.65, 47.36, 47.93, 124.80, 126.56, 127.40, 128.73, 144.16, 150.62,

−

Sulfonate, VHAT (19). A mixture of 13 (19.3 g, 70.9 mmol) and

dry THF (322 mL) was cooled to −78 °C. DIBAL-H (1.0 M solution

in toluene, 92.2 mL, 92.2 mmol) was added slowly, and the resulting

mixture was stirred for 10 min. The mixture was warmed to ambient

temperature until complete consumption of the starting material and

then diluted with ether (∼160 mL) before cooling in an ice bath.

Water (3.7 mL) was added slowly, followed by 15% aqueous NaOH

172.73. HRMS (ESI, m/z): for C H O [M − H] calcd, 189.0921;

found, 189.0915. mp 158−160 °C.

General Procedure for the Synthesis of Indoles 23 and 24.

A mixture containing 22 (500.0 mg, 2.6 mmol) and dichloromethane

(15.5 mL) was cooled to 0 °C. Thionyl chloride (0.4 mL, 5.3 mmol)

was added dropwise, followed by a catalytic amount of DMF (0.02

mL, 0.3 mmol). After stirring at ambient temperature for 24 h, the

mixture was concentrated under reduced pressure and redissolved in

1.5 mL of 1,1,1,3,3,3-HFIP. The acid chloride solution was added

dropwise to a mixture of indole (2.8 mmol) and 2.0 mL of HFIP. The

mixture was stirred at ambient temperature for 24 h, diluted with

DCM, concentrated under reduced pressure, and puriﬁed by

automatic ﬂash column chromatography on silica gel (gradient

elution from 1 to 10% MeOH−CH Cl ).

(

3.7 mL) and water (9.2 mL). The mixture was removed from the

bath and dried over MgSO . After 15 min of stirring, the mixture was

ﬁltered. The ﬁltrate was concentrated under reduced pressure and

ﬁnally puriﬁed by automatic ﬂash column chromatography on silica

gel (gradient elution from 5% to 40% EtOAc-hexanes) to aﬀord

VHAT as a colorless, viscous liquid (14.5 g, 75% yield). H NMR

(

400 MHz, CDCl ): δ 0.99 (s, 3H), 1.09 (s, 3H), 1.33 (dd, 1H), 1.83

dd, 1H), 1.96 (broad, s, 1H), 2.01 (d, 1H), 2.27 (d, 1H), 4.43−4.46

m, 1H), 5.80 (s, 1H). C{ H}NMR (100 MHz, CDCl ): δ 25.95,

(

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1H-indol-3-yl)-

methanone (GBD-001, 23). Obtained as white, powdery crystals

0.55, 32.30, 41.08, 44.17, 65.57, 116.91, 120.35, 149.82.

(58% yield). H NMR (400 MHz, DMSO-d ): δ 1.15 (s, 6H), 2.77

Ethyl-E-4-(diethoxyphosphoryl)but-2-enoate. Triethyl phos-

(overlapping s, 4H), 7.19−7.27 (m, 2H), 7.31 (d, 1H), 7.50−7.53 (m,

phite (10.0 g, 38.8 mmol) and ethyl 4-bromocrotonate (7.9 g, 46.6

mmol) were combined in a pressure ﬂask and heated to 140 °C

overnight. The mixture was cooled to ambient temperature,

transferred to a ﬂask, and distilled until complete removal of ethyl

bromide. The crude product was a brown, viscous liquid and used in

the next step without puriﬁcation. The H NMR is consistent with the

literature report.

1H), 7.55−7.58 (m, 1H), 7.59 (d, 1H), 7.93 (s, 1H), 8.22−8.25 (m,

1H), 12.00 ppm (s, 1H). C{ H}NMR (100 MHz, DMSO-d ): δ

28.45, 40.12, 46.94, 47.10, 112.20, 115.13, 121.50, 121.75, 123.01,

124.40, 124.72, 126.36, 126.99, 135.40, 136.66, 138.84, 143.34,

−

146.69, 190.08. HRMS (ESI, m/z): for C H NO [M − H] calcd,

288.1394; found, 288.1394. mp 224−225 °C. Purity (qNMR),

100.1%.

Ethyl (2E, 4E)-7,7-Dimethyldeca-2,4-dien-9-ynoate (20).

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1H-indol-3-

mixture of 19 (5.3 g, 19.5 mmol), ethyl-E-4-(diethoxyphosphoryl)-

but-2-enoate (7.3 g, 29.2 mmol), and dry THF (97 mL) was cooled

to −78 °C. LHMDS (1.0 M solution in THF, 42.8 mL, 42.8 mmol)

was added slowly, and the resulting mixture was stirred for 15 min.

The mixture was warmed to ambient temperature and then at 60 °C

for 1 h or until complete consumption of the starting material. After

cooling to ambient temperature, 1 M HCl (∼97 mL) was added, and

the mixture was extracted with ethyl acetate (3 × 25 mL). The

organic layer was washed with water and brine, dried over Na SO ,

yl)methanone (GBD-019, 24). Obtained as white, powdery crystals

(53% yield). H NMR (400 MHz, DMSO-d ): δ 1.13 (s, 6H), 2.72 (s,

2H), 2.76 (s, 2H), 7.00 (t, 1H), 7.10 (t, 1H), 7.27 (d, 1H), 7.24 (d,

1H), 7.36 (d, 1H), 7.38 (d, 1H), 7.43 (s, 1H), 11.87 (s, 1H).

C{ H}NMR (100 MHz, DMSO-d ): δ 14.13, 28.30, 40.21, 46.83,

47.07, 111.18, 112.75, 120.01, 120.74, 121.67, 124.35, 124.51, 126.77,

127.35, 134.92, 139.72, 143.27, 143.80, 146.77, 191.84. HRMS (ESI,

m/z): for C H NO [M + H] calcd, 304.1696; found, 304.1636. mp

239−241 °C.

and concentrated under reduced pressure. The crude product was

puriﬁed by automatic ﬂash column chromatography on silica gel

General Procedure for the Synthesis of 4, 26−34, 36, 37,

and 39. NaH (27.6 mg, 0.69 mmol) and DMSO (1.0 mL) were

mixed thoroughly in a round-bottom ﬂask. In a separate pear-shaped

ﬂask, 23 (100 mg, 0.35 mmol) and DMSO (1.5 mL) were heated to

∼40 °C to form a solution and then added slowly to the NaH

suspension. After stirring at ambient temperature for 30 min, the alkyl

halide (0.42 mmol) was added. The resulting mixture was stirred

overnight at ambient temperature. The reaction mixture was

quenched with water and extracted with ethyl acetate (2 × 10 mL).

The organic layer was washed with brine, dried over Na SO ,

(gradient elution from 2 to 20% EtOAc-hexanes) to aﬀord dienyne as

a light-yellow, viscous liquid (3.2 g, 75% yield). H NMR (400 MHz,

CDCl ): δ 0.10 (s, 6H), 1.30 (t, 3H), 2.03 (t, 1H), 2.08 (d, 2H), 2.19

(

d, 2H), 4.21 (q, 2H), 5.82 (d, 1H) 6.10−6.16 (m, 1H), 6.22 (dd,

H), 7.28 (dd, 1H). C{ H}NMR (100 MHz, CDCl ): δ 14.30,

6.66, 31.51, 34.31, 44.43, 60.19, 70.32, 81.99, 119.83, 131.13, 140.44,

44.57, 167.19.

Ethyl 2,2-Dimethyl-2,3-dihydro-1H-indene-5-carboxylate (21).

To a mixture of 20 (3.2 g, 14.6 mmol) in dry dichloromethane (291

mL), di-μ-chlorobis(norbornadiene)dirhodium [Rh(nbd)Cl]₂(0.14

g, 0.29 mmol) was added, followed by silver hexaﬂuoroantimonate

concentrated under reduced pressure, and puriﬁed by automatic ﬂash

column chromatography on silica gel (gradient elution from 5 to 40%

EtOAc-hexanes).

(

AgSbF , 0.43 g, 1.24 mmol). The resulting mixture was stirred at

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-pentyl-1H-indol-3-

ambient temperature for 1 h. DDQ (4.1 g, 17.5 mmol) was added,

and stirring was continued until complete oxidation (1−2 h). After

the reaction, the brown mixture was ﬁltered in basic alumina. The

ﬁltrate was concentrated under reduced pressure and puriﬁed by

automatic ﬂash column chromatography on silica gel (gradient elution

yl)methanone (GBD-003, 4). Obtained as white crystals (88%

yield). H NMR (400 MHz, CDCl ) δ 0.89 (t, 3H), 1.19 (s, 6H),

1.26−140 (m, 4H), 1.87 (quintet, 2H), 2.79 (s, 4H), 4.15 (t, 2H),

7.22−7.27 (m, 1H), 7.28−7.35 (m, 2H), 7.36−7.41 (m, 1H), 7.57−

7.62 (m, 2H), 7.64 (s, 1H), 8.39−8.45 (m, 1H). C{ H}NMR (100

from 1 to 10% EtOAc-hexanes) to aﬀord the ester as a colorless,

MHz, CDCl ) δ 13.93, 22.22, 28.72, 28.98, 29.57, 40.48, 47.10, 47.52,

viscous liquid (2.7 g, 85% yield). H NMR (400 MHz, CDCl ): δ

47.73, 109.77, 115.67, 122.40, 122.80, 123.31, 124.27, 125.15, 127.30,

.15 (s, 6H), 1.38 (t, 3H), 2.75 (s, 4H), 4.35 (q, 2H), 7.21 (d, 1H),

127.45, 136.68, 136.75, 139.21, 143.88, 147.37, 191.20. HRMS (ESI,

.81−7.86 (m, 2H). C{ H}NMR (100 MHz, CDCl ): δ 14.38,

m/z) for C H NO [M + H] calcd, 360.2322, found 360.2318. mp

25 30

8.60, 40.51, 47.31, 47.74, 60.70, 124.53, 125.79, 127.88, 128.52,

43.84, 149.27, 167.09.

62−64 °C. Purity (qNMR), 99.9%.

(2,2-dimethyl-2,3-dihydro-1H-inden-5-yl) (1-propyl-1H-indol-3-

,2-Dimethyl-2,3-dihydro-1H-indene-5-carboxylic acid (22).

Compound 21 (3.7 g, 17.1 mmol) was ﬁrst dissolved in ethanol

82 mL). Aqueous KOH (9.1 mL) was then added, and the mixture

yl)methanone (GBD-005, 26). Obtained as white, powdery crystals

(83% yield). H NMR (400 MHz, CDCl ): δ 0.95 (t, 3H), 1.19 (s,

(

6H), 1.91 (sextet, 2H), 2.79 (s, 4H), 4.13 (t, 2H), 7.23−7.27 (m,

was stirred overnight at ambient temperature. After the reaction, the

mixture was neutralized with 3 M HCl. The solids were ﬁltered and

1H), 7.29−7.35 (m, 2H), 7.36−7.42 (m, 1H), 7.58−7.62 (m, 2H),

7.64 (s, 1H), 8.39−8.45 (m, 1H). C{ H}NMR (100 MHz, CDCl ):

washed with cold water. The crude product was white, ﬂuﬀy crystals

δ 11.42, 23.21, 28.73, 40.48, 47.52, 47.73, 48.74, 109.79, 115.67,

(

2.8 g, 85%) and used in the next step without further puriﬁcation. H

122.42, 122.82, 123.32, 124.26, 125.13, 127.28, 127.46, 136.76,

390

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

Article

36.78, 139.21, 143.90, 147.36, 191.22. HRMS (ESI, m/z): for

(Z)-(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(pent-2-en-1-

C H NO [M + H] calcd, 332.2009; found, 332.2006. mp 91−93

yl)-1H-indol-3-yl)methanone (GBD-012, 33). Obtained as white,

C. Purity (qNMR), 99.2%.

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-butyl-1H-indol-3-

yl)methanone (GBD-006, 27). Obtained as white, powdery crystals

powdery crystals (68% yield). H NMR (400 MHz, CDCl

): δ 1.07

(

(t, 3H), 1.17 (s, 6H), 2.24 (quint, 2H), 2.78 (s, 4H), 4.78 (app. d,

2H), 5.51−5.59 (m, 1H, J = 10.8 Hz, 6.8 Hz, 0.8 Hz), 5.68−5.76 (m,

1H, J = 10.7 Hz, 7.4 Hz, 0.8 Hz), 7.22−7.26 (m, 1H), 7.29−7.34 (m,

2H), 7.35−7.40 (m, 1H), 7.56−7.62 (m, 2H), 7.63 (s, 1H), 8.37−

(85% yield). H NMR (400 MHz, CDCl ): δ 0.93 (t, 3H), 1.19 (s,

H), 1.36 (sextet, 2H), 1.86 (quintet, 2H), 2.80 (s, 4H), 4.17 (t, 2H),

.23−7.27 (m, 1H), 7.29−7.35 (m, 2H), 7.36−7.41 (m, 1H), 7.58−

8.43 (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 14.00, 20.96,

.62 (m, 2H), 7.64 (s, 1H), 8.39−8.44 ppm (m, 1H). C{ H}NMR

28.72, 40.48, 43.79, 47.52, 47.73, 109.83, 115.91, 122.54, 122.58,

122.82, 123.36, 124.28, 125.14, 127.29, 127.60, 136.18, 136.83,

136.85, 139.17, 143.87, 147.38, 191.18. HRMS (ESI, m/z): for

(100 MHz, CDCl ) δ 13.63, 20.09, 28.73, 31.92, 40.48, 46.86, 47.52,

7.73, 109.77, 15.69, 122.41, 122.82, 123.31, 124.27, 125.14, 127.28,

27.45, 136.68, 136.76, 139.21, 143.90, 147.37, 191.20. HRMS (ESI,

C H NO [M + H] calcd, 358.2165; found, 358.2162. mp 70−71

m/z): for C H NO [M + H] calcd, 346.2165; found, 346.2163. mp

°C. Purity (qNMR), 99.5%.

0−72 °C. Purity (qNMR), 100.1%.

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-hexyl-1H-indol-3-

yl)methanone (GBD-007, 28). Obtained as white, powdery crystals

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(pent-4-en-1-yl)-

(

1H-indol-3-yl)methanone (GBD-013, 34). Obtained as light-yellow,

powdery crystals (74% yield). H NMR (400 MHz, CDCl ): δ 1.18

(82% yield). H NMR (400 MHz, CDCl ): δ 0.87 (t, 3H), 1.19 (s,

(s, 6H), 1.97 (quint, 2H), 2.09 (q, 2H), 2.79 (s, 4H), 4.16 (t, 2H),

5.01−5.04 (m 1H), 5.05−5.09 (m, 1H), 5.72−5.84 (ddt, 1H), 7.22−

7.26 (m, 1H), 7.29−7.34 (m, 2H), 7.35−7.40 (m, 1H), 7.56−7.61

H), 1.31 (m, 6H), 1.87 (quintet, 2H), 2.80 (s, 2H), 4.15 (t, 2H),

.23−7.27 (m, 1H), 7.29−7.36 (m, 2H), 7.37−7.41 (m, 1H), 7.57−

.62 (m, 2H), 7.64 (s, 1H), 8.39−8.44 ppm (m, 1H). C{ H}NMR

(m, 2H), 7.63 (s, 1H), 8.39−8.44 ppm (m, 1H). C{ H}NMR (100

(100 MHz, CDCl ): δ 13.97, 22.50, 26.54, 28.73, 29.83, 31.28, 40.48,

MHz, CDCl ): δ 28.72, 28.77, 30.67, 40.48, 46.24, 47.53, 47.73,

7.12, 47.53, 47.74, 109.77, 115.68, 122.41, 122.82, 123.31, 124.27,

25.15, 127.30, 127.47, 136.67, 136.75, 139.23, 143.88, 147.36,

109.75, 115.80, 116.17, 122.46, 122.86, 123.37, 124.28, 125.12,

127.26, 127.48, 136.69, 136.71, 136.79, 139.18, 143.88, 147.39,

91.20. HRMS (ESI, m/z): for C H NO [M + H] calcd, 374.2478;

191.18. HRMS (ESI, m/z): for C H NO [M + H] calcd, 358.2165;

25 28

found, 374.2477. mp 61−63 °C. Purity (qNMR), 99.7%.

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-isobutyl-1H-indol-

-yl)methanone (GBD-008, 29). Obtained as yellowish oil (53%

found, 358.2162. mp 78−79 °C. Purity (qNMR), 99.3%.

(

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(pent-2-yn-1-yl)-

1H-indol-3-yl)methanone (GBD-015, 36). Obtained as light-yellow,

yield). H NMR (400 MHz, CDCl ): δ 0.94 (d, 6H), 1.18 (s, 6H),

powdery crystals (55% yield). H NMR (400 MHz, CDCl ): δ 1.13

.23 (m, 1H), 2.79 (s, 4H), 3.94 (d, 2H), 7.22−7.26 (m, 1H), 7.28−

(t, 3H), 1.19 (s, 6H), 2.18−2.25 (qt, 2H), 2.79 (s, 4H), 4.88 (t, 2H),

.33 (m, 2H), 7.34−7.39 (m, 1H), 7.54−7.61 (m, 2H), 7.64 (s, 1H),

7.23−7.27 (m, 1H), 7.31−7.38 (m, 2H), 7.42−7.48 (m, 1H), 7.60−

.39−8.44 ppm (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 20.19,

7.68 (m, 2H), 7.77 (s, 1H), 8.39−8.45 (m, 1H). C{ H}NMR (100

8.72, 29.15, 40.46, 47.51, 47.71, 54.68, 109.97, 115.54, 122.37,

22.76, 123.29, 124.25, 125.16, 127.29, 127.40, 137.00, 137.16,

39.18, 143.89, 147.38, 191.23. HRMS (ESI, m/z): for C H NO [M

H] calcd, 346.2165; found, 346.2161. Purity (qNMR), 98.5%.

1-Benzyl-1H-indol-3-yl) (2,2-dimethyl-2,3-dihydro-1H-inden-5-

yl)methanone (GBD-009, 30). Obtained as white, powdery crystals

MHz, CDCl ): δ 12.37, 13.58, 28.71, 37.02, 40.49, 47.53, 47.73,

71.93, 88.93, 109.79, 116.00, 122.70, 122.88, 123.53, 124.31, 125.15,

127.33, 127.63, 136.07, 136.48, 139.05, 143.85, 147.47, 191.18.

HRMS (ESI, m/z): for C H NO [M + H] calcd, 356.2009; found,

(

332.2007. mp 98−99 °C. Purity (qNMR), 96.8%.

(1-Allyl-1H-indol-3-yl) (2,2-dimethyl-2,3-dihydro-1H-inden-5-yl)-

(

86% yield). H NMR (400 MHz, CDCl ): δ 1.17 (s, 6H), 2.78 (s,

H), 5.37 (s, 2H), 7.11−7.15 (m, 2H), 7.21−7.35 (m, 7H), 7.58−

.62 (app. d, 1H), 7.64 (s, 2H), 8.42−8.46 ppm (m, 1H). C{ H}-

methanone (GBD-016, 37). Obtained as yellowish oil (81% yield).

H NMR (400 MHz, CDCl ): δ 1.18 (s, 6H), 2.79 (s, 4H), 4.76 (m,

2H), 5.10−5.17 (m, 1H), 5.23−5.28 (m, 1H), 5.94−6.04 (m, 1H),

NMR (100 MHz, CDCl ): δ 28.71, 40.47, 47.50, 47.71, 50.74, 110.14,

7.22−7.26 (m, 1H), 7.29−7.38 (m, 3H), 7.57−7.61 (m, 2H), 7.64 (s,

16.25, 122.64, 122.82, 123.63, 124.31, 125.16, 126.78, 127.31,

27.52, 128.10, 128.98, 135.92, 136.96, 137.02, 139.01, 143.91,

47.52, 191.24. HRMS (ESI, m/z): for C H NO [M + H] calcd,

80.2009; found, 380.2005. mp 111−114 °C. Purity (qNMR), 99.6%.

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(4-ﬂuorobenzyl)-

H-indol-3-yl)methanone (GBD-010, 31). Obtained as white,

1H), 8.41−8.45 ppm (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ

28.67, 40.43, 47.46, 47.66, 49.33, 109.97, 116.01, 118.39, 122.50,

122.74, 123.42, 124.25, 125.04, 127.23, 127.39, 132.07, 136.60,

136.80, 139.04, 143.87, 147.40, 191.17. HRMS (ESI, m/z): for

(

C H NO [M + H] calcd, 330.1852; found, 330.1848. Purity

(

qNMR), 98.3%.

powdery crystals (90% yield). H NMR (400 MHz, CDCl ): δ 1.18

Ethyl 2-(3-(2,2-Dimethyl-2,3-dihydro-1H-indene-5-carbonyl)-1H-

(

s, 6H), 2.78 (s, 4H), 5.34 (s, 2H), 6.98−7.04 (m, 2H), 7.09−7.14

indol-1-yl)acetate (GBD-018, 39). Obtained as yellowish oil (28%

yield). H NMR (400 MHz, CDCl ): δ 1.18 (s, 6H), 1.26 (t, 3H),

m, 2H), 7.24 (m, 1H), 7.28−7.35 (m, 3H), 7.58−7.65 (m, 3H),

.40−8.43 ppm (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 28.70,

2.78 (s, 4H), 4.22 (q, 2H), 4.86 (s, 2H), 7.22−7.36 (m, 4H), 7.59−

0.49, 47.50, 47.71, 50.09, 110.02, 115.86, 116.06, 116.40, 122.72,

22.90, 123.72, 124.33, 125.15, 127.31, 127.55, 131.64, 128.50,

28.58, 136.68, 136.87, 138.95, 143.96, 147.62, 163.65, 191.20.

7.63 (m, 2H), 7.65 (s, 1H), 8.40−8.45 ppm (m, 1H). C{ H}NMR

(100 MHz, CDCl ): δ 14.10, 28.69, 40.47, 47.48, 47.70, 48.15, 62.11,

109.20, 116.80, 122.76, 122.91, 123.85, 124.32, 125.11, 127.21,

127.34, 137.10, 137.32, 138.89, 143.90, 147.60, 167.49, 191.28.

FNMR (376 MHz, CDCl ): δ −113.65. HRMS (ESI, m/z): for

C H FNO [M + H] calcd, 398.1915; found, 398.1911. mp 124−

HRMS (ESI, m/z): for C H NO [M + H] calcd, 376.1907; found,

24 26

26 °C. Purity (qNMR), 100.5%.

E)-(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(pent-2-en-1-yl)-

H-indol-3-yl)methanone (GBD-011, 32). Obtained as white,

376.1904. Purity (qNMR), 96.5%.

(

(2,2-dimethyl-2,3-dihydro-1H-inden-5-yl) (1-methyl-1H-indol-3-

yl)methanone (GBD-004, 25). NaH (14.2 mg, 0.36 mmol) and

DMSO (1.0 mL) were mixed thoroughly in a round-bottom ﬂask. In a

separate pear-shaped ﬂask, 23 (51.4 mg, 0.18 mmol) and DMSO (1.5

mL) were heated to ∼40 °C to form a solution and then added slowly

to the NaH suspension. After stirring at ambient temperature for 30

min, (chloromethyl)trimethylsilane (0.03 mL, 0.21 mmol) was added.

The resulting mixture was stirred overnight at ambient temperature.

The reaction was quenched with water and extracted with ethyl

acetate (2 × 5 mL). The organic layer was washed with brine, dried

over Na SO , and concentrated under reduced pressure. Puriﬁcation

powdery crystals (88% yield). H NMR (400 MHz, CDCl ): δ 0.98

(

t, 3H), 1.18 (s, 6H), 2.06 (quint, 2H), 2.79 (s, 4H), 4.69−4.74 (app.

d, 2H), 5.56−5.65 (m, 1H, J = 15.3 Hz, 6.0 Hz, 1.4 Hz), 5.71−5.80

(

m, 1H, J = 15.4 Hz, 6.2 Hz, 1.4 Hz), 7.22−7.27 (m, 1H), 7.29−7.35

(m, 2H), 7.36−7.42 (m, 1H), 7.58−7.62 (m, 2H), 7.65 (s, 1H),

.39−8.44 (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 13.21,

5.18, 28.72, 40.48, 47.52, 47.72, 48.94, 110.05, 115.86, 122.46,

22.70, 122.77, 123.33, 124.28, 125.14, 127.30, 127.52, 136.47,

36.85, 137.27, 139.17, 143.87, 147.38, 191.20. HRMS (ESI, m/z):

for C H NO [M + H] calcd, 358.2165; found, 358.2163. mp 80−

by automatic ﬂash column chromatography on silica gel (gradient

elution from 5 to 40% EtOAc-hexanes) aﬀorded white, powdery

82 °C. Purity (qNMR), 99.1%.

391

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

Article

crystals (64% yield). H NMR (400 MHz, CDCl ): δ 1.18 (s, 6H),

47.76, 109.33, 113.94, 121.02, 121.09, 121.70, 124.25, 125.42, 127.31,

127.88, 135.79, 139.58, 143.72, 143.77, 147.84, 193.28. HRMS (ESI,

.79 (s, 4H), 3.85 (s, 3H), 7.22−7.26 (m, 1H), 7.31−7.39 (m, 3H),

(

.56 (s, 1H), 7.57−7.62 (app. d, 1H), 7.63 (s, 1H), 8.40−8.44 ppm

m/z): for C H NO [M + H] calcd, 360.2322; found, 360.2318.

m, 1H). ¹³C{ H}NMR (100 MHz, CDCl ) δ 28.71, 33.53, 40.48,

Purity (qNMR), 97.3%.

7.51, 47.71, 109.53, 115.75, 122.51, 122.74, 123.47, 124.28, 125.06,

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-pentyl-

27.03, 127.28, 137.46, 137.61, 139.15, 143.86, 147.36, 191.18.

1H-indol-3-yl)methanone (GBD-022, 42). Obtained as yellowish

HRMS (ESI, m/z): for C H NO [M + H] calcd, 304.1696; found,

oil (74% yield). H NMR (400 MHz, CDCl

): δ 0.92 (t, 3H), 1.17 (s,

304.1693. mp 92−95 °C. Purity (qNMR), 99.9%.

6H), 1.34−1.43 (m, 4H), 1.80 (quintet, 2H), 2.59 (s, 3H), 2.75 (s,

General Procedure for the Synthesis of 35 and 38. To a

2H), 2.78 (s, 2H), 4.12 (t, 2H), 7.05 (m, 1H), 7.15−7.21 (m, 2H),

mixture of 23 (100 mg, 0.35 mmol) and DMSO (2.0 mL) was added

KOH (38.8 mg, 0.69 mmol). After stirring at ambient temperature for

was stirred for 2 h at ambient temperature. The reaction was

quenched with water and extracted with ethyl acetate (2 × 10 mL).

The organic layer was washed with brine, dried over Na SO ,

concentrated under reduced pressure, and puriﬁed by automatic ﬂash

column chromatography on silica gel (gradient elution from 5 to 40%

EtOAc-hexanes).

7.33 (app. t, 2H), 7.53 (app. d, 1H), 7.61 (s, 1H). C{ H}NMR (100

MHz, CDCl ): δ 12.42, 13.95, 22.41, 28.65, 29.13, 29.43, 40.51,

h, the alkyl halide (0.52 mmol) was added. The resulting mixture

43.34, 47.41, 47.75, 109.33, 113.92, 121.02, 121.08, 121.71, 124.24,

125.41, 127.31, 127.86, 135.78, 139.59, 143.70, 143.75, 147.82,

193.26. HRMS (ESI, m/z): for C26

32NO [M + H] calcd, 374.2478;

found, 374.2477. Purity (qNMR), 96.2%.

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-hexyl-2-methyl-1H-

indol-3-yl)methanone (GBD-023, 43). Obtained as yellowish oil

(

62% yield). H NMR (400 MHz, CDCl ): δ 0.89 (t, 3H), 1.17 (s,

(

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(pent-4-yn-1-yl)-

6H), 1.28−1.44 (m, 6H), 1.79 (quintet, 2H), 2.59 (s, 3H), 2.74 (s,

H-indol-3-yl)methanone (GBD-014, 35). Obtained as light-yellow,

2H), 2.78 (s, 2H), 4.12 (t, 2H), 7.04 (m, 1H), 7.15−7.21 (m, 2H),

powdery crystals (79% yield). H NMR (400 MHz, CDCl ): δ 1.18

7.32 (app. t, 2H), 7.52 (app. d, 1H), 7.59 (s, 1H). C{ H}NMR (100

(

s, 6H), 2.03−2.12 (m, 3H), 2.17−2.22 (td, 2H, J = 6.4 Hz, 2.5 Hz),

MHz, CDCl ): δ 12.44, 14.00, 22.55, 26.72, 28.65, 29.70, 31.47,

40.52, 43.39, 47.42, 47.76, 109.32, 113.93, 121.02, 121.09, 121.71,

.79 (s, 4H), 4.34 (t, 2H), 7.22−7.28 (m, 1H), 7.29−7.36 (m, 2H),

.39−7.45 (m, 1H), 7.58−7.63 (d, 1H), 7.65 (m, 2H), 8.39−8.46

124.25, 125.43, 127.32, 127.87, 135.78, 139.60, 143.71, 143.75,

ppm (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 15.64, 28.06,

147.83, 193.28. HRMS (ESI, m/z): for C H NO [M + H] calcd,

27 34

8.71, 40.49, 45.29, 47.53, 47.73, 70.21, 82.30, 109.68, 115.84, 122.55,

22.91, 123.48, 124.32, 125.20, 127.33, 127.53, 136.59, 136.97,

39.06, 143.88, 147.47, 191.15. HRMS (ESI, m/z): for C H NO [M

388.2635; found, 388.2632. Purity (qNMR), 98.1%.

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-benzyl-2-methyl-

1H-indol-3-yl)methanone (GBD-024, 44). Obtained as yellowish

H] calcd, 356.2009; found, 356.2006. mp 84−85 °C. Purity

oil (56% yield). H NMR (400 MHz, CDCl

): δ 1.18 (s, 6H), 2.54 (s,

(

qNMR), 100.4%.

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(prop-2-yn-1-yl)-

H-indol-3-yl)methanone (GBD-017, 38). Obtained as light-yellow

3H), 2.76 (s, 2H), 2.79 (s, 2H), 5.40 (s, 2H), 7.01−7.11 (m, 3H),

(

7.13−7.23 (m, 2H), 7.24−7.33 (m, 4H), 7.39−7.43 (app. d, 1H),

7.54−7.58 (app. d, 1H), 7.63 (br s, 1H). C{ H}NMR (100 MHz,

oil (66% yield). H NMR (400 MHz, CDCl ): δ 1.19 (s, 6H), 2.49 (t,

CDCl ): δ 12.48, 28.65, 40.53, 46.62, 47.42, 47.77, 109.54, 114.45,

H), 2.80 (s, 4H), 4.93 (d, 2H), 7.23−7.28 (m, 1H), 7.32−7.41 (m,

121.16, 121.36, 122.14, 124.30, 125.47, 125.99, 127.31, 127.69,

H), 7.43−7.49 (m, 1H), 7.59−7.67 (m, 2H), 7.74 (s, 1H), 8.39−

127.96, 128.97, 136.28, 136.31, 139.39, 143.79, 143.95, 148.06,

.45 ppm (m, 1H). C{ H}NMR (100 MHz, CDCl ): δ 28.71,

193.34. HRMS (ESI, m/z): for C H NO [M + H] calcd, 394.2165;

28 28

6.53, 40.50, 47.50, 47.73, 75.02, 76.29, 109.64, 116.47, 122.87,

22.97, 123.76, 124.34, 125.14, 127.33, 127.56, 135.81, 136.41,

38.89, 143.96, 147.63, 191.21. HRMS (ESI, m/z): for C H NO [M

found, 394.2162. Purity (qNMR), 97.4%.

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-(4-ﬂuorobenzyl)-2-

methyl-1H-indol-3-yl)methanone (GBD-025, 45). Obtained as

H] calcd, 328.1696; found, 328.1695. Purity (qNMR), 100.0%.

yellowish oil (52% yield). H NMR (400 MHz, CDCl

): δ 1.18 (s,

General Procedure for the Synthesis of 40−48 and 50. NaH

6H), 2.53 (s, 3H), 2.76 (s, 2H), 2.79 (s, 2H), 5.36 (s, 2H), 6.98−7.02

(

26.4 mg, 0.66 mmol) and DMSO (1.0 mL) were mixed thoroughly

(m, 4H), 7.06−7.11 (m, 1H), 7.14−7.27 (m, 3H), 7.39−7.43 (app. d,

in a round-bottom ﬂask. In a separate pear-shaped ﬂask, 24 (100 mg,

1H), 7.54−7.57 (app. d, 1H), 7.63 (br s, 1H). C{ H}NMR (100

0.33 mmol) and DMSO (1.5 mL) were heated to ∼40 °C to form a

MHz, CDCl ): δ 12.44, 28.64, 40.54, 45.98, 47.41, 47.77, 109.40,

solution and then added slowly to the NaH suspension. After stirring

at ambient temperature for 30 min, the alkyl halide (0.40 mmol) was

added. The resulting mixture was stirred overnight at ambient

temperature. The reaction mixture was quenched with water and

extracted with ethyl acetate (2 × 10 mL). The organic layer was

washed with brine, dried over Na SO , concentrated under reduced

114.59, 115.83, 116.04, 121.23, 121.45, 122.24, 124.32, 125.47,

127.32, 127.96, 127.72, 127.64, 132.01, 136.14, 139.29, 143.86,

143.83, 148.16, 163.41, 193.29. FNMR (376 MHz, CDCl

−114.40. HRMS (ESI, m/z): for C28

27FNO [M + H]⁺calcd,

412.2071; found, 412.2068. Purity (qNMR), 96.8%.

): δ

(E)-(2,2-dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-(pent-

-en-1-yl)-1H-indol-3-yl)methanone (GBD-026, 46). Obtained as

pressure, and puriﬁed by automatic ﬂash column chromatography on

silica gel (gradient elution from 5 to 40% EtOAc-hexanes).

yellowish oil (78% yield). H NMR (400 MHz, CDCl ): δ 0.94 (t,

H), 1.17 (s, 6H), 2.02 (quint, 2H), 2.56 (s, 3H), 2.75 (s, 2H), 2.78

(

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-propyl-

H-indol-3-yl)methanone (GBD-020, 40). Obtained as yellowish

(s, 2H), 4.69−4.73 (m, 2H), 5.48−5.58 (m, 2H), 7.03−7.09 (m, 1H),

oil (72% yield). H NMR (400 MHz, CDCl ): δ 1.01 (t, 3H), 1.17 (s,

7.15−7.22 (m, 2H), 7.31 (dd, 1H), 7.37 (dd, 1H), 7.54 (d, 1H), 7.61

6H), 1.85 (sextet, 2H), 2.59 (s, 3H), 2.75 (s, 2H), 2.78 (s, 2H), 4.11

(s, 1H). C{ H}NMR (100 MHz, CDCl ): δ 12.32, 13.19, 25.07,

(

t, 2H), 7.05 (app. t, 1H), 7.15−7.21 (m, 2H), 7.33 (app. t, 2H), 7.53

28.62, 40.49, 44.92, 47.38, 47.73, 109.41, 114.02, 121.02, 121.09,

121.79, 122.51, 124.23, 125.41, 127.24, 127.87, 135.38, 135.84,

139.51, 143.69, 143.88, 147.85, 193.27. HRMS (ESI, m/z): for

(app. d, 1H), 7.60 (s, 1H). C{ H}NMR (100 MHz, CDCl ): δ

1.48, 12.43, 22.99, 28.64, 40.50, 44.84, 47.40, 47.74, 109.36, 113.92,

21.02, 121.06, 121.70, 124.24, 125.41, 127.28, 127.86, 135.83,

C H NO [M + H] calcd, 372.2322; found, 372.2318. Purity

39.56, 143.70, 143.79, 147.83, 193.28. HRMS (ESI, m/z): for

(qNMR), 95.9%.

C H NO [M + H] calcd, 346.2165; found, 346.2162. Purity

(Z)-(2,2-dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-(pent-

(

qNMR), 98.8%.

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (1-butyl-2-methyl-1H-

indol-3-yl)methanone (GBD-021, 41). Obtained as yellowish oil

2-en-1-yl)-1H-indol-3-yl)methanone (GBD-027, 47). Obtained as

(

white, powdery crystals (73% yield). H NMR (400 MHz, CDCl

): δ

1.11 (t, 3H), 1.17 (s, 6H), 2.29 (quint, 2H), 2.58 (s, 3H), 2.75 (s,

2H), 2.78 (s, 2H), 4.78−4.83 (m, 2H), 5.32−5.40 (m, 1H), 5.57−

5.65 (m, 1H), 7.04−7.09 (m, 1H), 7.16−7.22 (m, 2H), 7.30 (dd,

(

71% yield). H NMR (400 MHz, CDCl ): δ 0.98 (t, 3H), 1.16 (s,

H), 1.43 (sextet, 2H), 1.78 (quintet, 2H), 2.59 (s, 3H), 2.74 (s, 2H),

.78 (s, 2H), 4.13 (t, 2H), 7.04 (app. t, 1H), 7.15−7.21 (m, 2H), 7.32

1H), 7.36 (dd, 1H), 7.53 (d, 1H), 7.61 (s, 1H). C{ H}NMR (100

dd, 2H), 7.52 (app. d, 1H), 7.59 (s, 1H). ¹³C{ H}NMR (100 MHz,

MHz, CDCl ): δ 12.39, 14.03, 21.07, 28.65, 40.52, 40.63, 47.41,

(

CDCl ): δ 12.42, 13.83, 20.31, 28.65, 31.81, 40.52, 43.15, 47.41,

47.76, 109.24, 114.15, 121.12, 121.16, 121.81, 123.38, 124.26, 125.44,

392

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

The Supporting Information is available free of charge at

Article

27.37, 127.91, 134.90, 135.76, 139.50, 143.67, 143.73, 147.91,

contained 100 or 50 μg of membrane protein prepared from CHO-

93.26. HRMS (ESI, m/z) for C H NO [M + H] : calcd, 372.2322;

hCB1 or CHO-hCB cells, respectively. Reactions were incubated for

found, 372.2322. mp 65−66 °C. Purity (qNMR), 100.5%.

90 min at rt. Nonspeciﬁc binding was deﬁned as binding observed in

the presence of 10 μM of the nonselective CB /CB ligand WIN-

(

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-(pent-4-

en-1-yl)-1H-indol-3-yl)methanone (GBD-028, 48). Obtained as

yellowish oil (48% yield). H NMR (400 MHz, CDCl ): δ 1.17 (s,

55,212-2. Reactions were terminated by rapid vacuum ﬁltration

through Whatman GF/B glass ﬁber ﬁlters, followed by three washes

with the ice-cold binding buﬀer. Filters were then immediately placed

H), 1.90 (quintet, 2H), 2.18 (q, 2H), 2.58 (s, 3H), 2.74 (s, 2H),

.78 (s, 2H), 4.14 (t, 2H), 5.03−5.14 (m, 2H), 5.78−5.90 (ddt, 1H),

.02−7.08 (m, 1H), 7.15−7.21 (m, 2H), 7.29−7.36 (app. t, 2H), 7.53

into scintillation vials with 4 mL of the scintiverse BD cocktail

scintillation ﬂuid (Fisher Scientiﬁc, Fair Lawn, NJ). Samples were

incubated overnight in the scintillation ﬂuid, and vortexed and bound

reactivity was determined by employing a liquid scintillation

spectrophotometer (Tri Carb 2100 TR Liquid Scintillation Analyzer,

Packard Instrument Company, Meriden, CT).

(app. d, 1H), 7.60 (s, 1H). C{ H}NMR (100 MHz, CDCl ): δ

2.41, 28.58, 28.64, 30.90, 40.50, 42.68, 47.41, 47.75, 109.27, 114.04,

15.95, 121.08, 121.11, 121.76, 124.25, 125.41, 127.31, 127.87,

35.75, 136.94, 139.52, 143.64, 143.72, 147.87, 193.25. HRMS (ESI,

m/z): for C H NO [M + H] calcd, 372.2322; found, 372.2319.

[ S]GTPγS Binding. All [ S]GTPγS assays were conducted as

Purity (qNMR), 99.7%.

described previously in a buﬀer containing 20 mM N-(2-

hydroxyethyl)piperazine-N′-ethanesulfonic acid (HEPES), 100 mM

(

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-(pent-2-

yn-1-yl)-1H-indol-3-yl)methanone (GBD-030, 50). Obtained as

yellowish oil (22% yield). H NMR (400 MHz, CDCl ): δ 1.09 (t,

NaCl, 10 mM MgCl , 0.05% BSA, and 20 units/L of adenosine

deaminase. Assays were performed in triplicate in a ﬁnal volume of 1

H), 1.17 (s, 6H), 2.15 (qt, 2H), 2.62 (s, 3H), 2.75 (s, 2H), 2.78 (s,

mL with all reactions containing 0.1 nM [ S]GTPγS and an

experimental drug or drug vehicle and 50 or 25 μg of membrane

protein prepared from CHO-hCB₂cells, respectively. Nonspeciﬁc

binding was deﬁned by inclusion of 10 μM of nonradiolabeled

GTPγS. Reactions were incubated at 30 °C for 30 min and terminated

by rapid vacuum ﬁltration through Whatman GF/B glass ﬁber ﬁlters

and followed by four 1 mL washes with the ice-cold ﬁltration buﬀer

H), 4.84 (t, 2H), 7.05−7.11 (m, 1H), 7.17−7.25 (m, 2H), 7.36−

.42 (m, 2H), 7.54 (app. d, 1H), 7.62 (s, 1H). C{ H}NMR (100

MHz, CDCl ): δ 12.29, 12.33, 13.59, 28.64, 33.04, 40.51, 47.39,

7.75, 72.65, 86.73, 109.33, 114.42, 121.08, 121.31, 122.02, 124.27,

25.46, 127.26, 127.94, 135.57, 139.36, 143.38, 143.74, 148.01,

93.23. HRMS (ESI, m/z): for C H NO [M + H] calcd, 370.2165;

found, 370.2163. Purity (qNMR), 99.4%.

(20 mM HEPES, pH 7.4, 0.05% BSA). Filters were then immediately

General Procedure for the Synthesis of 49 and 51. To a

mixture of 24 (100 mg, 0.33 mmol) and DMSO (2.0 mL) was added

KOH (37.0 mg, 0.66 mmol). After stirring at ambient temperature for

placed into scintillation vials with 4 mL of the scintiverse

cocktail scintillation ﬂuid (Fisher Scientiﬁc, Fair Lawn, NJ). Samples

were incubated overnight in the scintillation ﬂuid, and vortexed and

bound reactivity was determined by employing a liquid scintillation

spectrophotometer (Tri Carb 2100 TR Liquid Scintillation Analyzer,

Packard Instrument Company, Meriden, CT).

Data Analysis. Curve ﬁtting was conducted utilizing GraphPad

Prism v6.0b (GraphPad Software, Inc.; San Diego, CA). Nonlinear

regression for one-site competition was used to determine the IC50 for

competition receptor binding. The Cheng-Prusoﬀ equation was

^e^m^p^l^o^y^e^d^t^o^c^oⁿ^v^e^r^t^t^h^e^e^x^p^e^rⁱ^m^eⁿ^t^a^l^I^C50 values obtained from

h, the alkyl halide (0.49 mmol) was added. The resulting mixture

was stirred for 2 h at ambient temperature. The reaction mixture was

quenched with water and extracted with ethyl acetate (2 × 10 mL).

The organic layer was washed with brine, dried over Na SO ,

concentrated under reduced pressure, and puriﬁed by automatic ﬂash

column chromatography on silica gel (gradient elution from 5 to 40%

EtOAc-hexanes).

(

2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-(pent-4-

yn-1-yl)-1H-indol-3-yl)methanone (GBD-029, 49). Obtained as

yellowish oil (63% yield). H NMR (400 MHz, CDCl ): δ 1.17 (s,

competition receptor binding experiments to K

values (a quantitative

measure of receptor aﬃnity). Statistical signiﬁcance of values from

H), 2.02 (quintet, 2H), 2.10 (t, 1H), 2.26−2.32 (td, 2H), 2.61 (s,

H), 2.75 (s, 2H), 2.78 (s, 2H), 4.29 (t, 2H), 7.03−7.09 (m, 1H),

.16−7.22 (m, 2H), 7.33−7.40 (m, 2H), 7.53 (app. d, 1H), 7.60 (s,

100% (basal) in the [ S]GTPγS assay was assessed by the one-

sample t-test (*P < 0.05 and **P < 0.01).

Molecular Modeling. To elucidate the binding modes of the

GBD series of compounds, a hybrid docking/MD simulation

approach was utilized. This methodology has been previously used

accurately and precisely to estimate the binding aﬃnity and selectivity

H). ¹³C{ H}NMR (100 MHz, CDCl ): δ 12.37, 15.95, 28.17, 28.62,

0.49, 41.83, 47.38, 47.73, 69.89, 82.75, 109.23, 114.16, 121.10,

21.17, 121.88, 124.25, 125.41, 127.29, 127.87, 135.79, 139.42,

43.65, 143.73, 147.94, 193.23. HRMS (ESI, m/z): for C H NO [M

of compounds toward the cannabinoid receptors. In summary, the

selected ligands (Figure 2) GBD-002 and GBD-003 had their lowest

H] calcd, 370.2165; found, 370.2163. Purity (qNMR), 100.5%.

energy conformation estimated using OpenEye OMEGA. These

(2,2-Dimethyl-2,3-dihydro-1H-inden-5-yl) (2-methyl-1-(prop-2-

yn-1-yl)-1H-indol-3-yl)methanone (GBD-031, 51). Obtained as

colorless oil (32% yield). H NMR (400 MHz, CDCl ): δ 1.17 (s,

ligands were then prepared with Autodock Tools to the PDBQT

69,70

format for use with Autodock Vina.

structures (PDB ID: 5XR8 for CB

for the initial docking, and the grid center was placed upon the

and CB

R and 5ZTY for CB R) were used

1 2

receptor

H), 2.33 (t, 1H), 2.63 (s, 3H), 2.75 (s, 2H), 2.79 (s, 2H), 4.89 (d,

H), 7.07−7.11 (m, 1H), 7.18−7.25 (m, 2H), 7.36−7.43 (m, 2H),

71,72

.54 (app. d, 1H), 7.62 (s, 1H). C{ H}NMR (100 MHz, CDCl ): δ

original crystal structure ligand.

The resulting binding poses

2.25, 28.65, 32.61, 40.53, 47.40, 47.77, 73.17, 77.22, 109.12, 114.80,

21.23, 121.55, 122.27, 124.32, 125.50, 127.30, 127.99, 135.50,

(Figure 3A1,A2,B1,B2) then had their atomic charges calculated using

73,74

the AM1-BCC method in the Antechamber module of AMBER16

and were subsequently energy-minimized using the SANDER module

39.17, 143.01, 143.81, 148.21, 193.22. HRMS (ESI, m/z): for

of AMBER16. The energy-minimized binding structures then had

C H NO [M + H] calcd, 342.1852; found, 342.1849. Purity

their binding free energy estimated using the MMPBSA.py module

(qNMR), 97.2%.

of AMBER16. The rest of the GBD series of compounds were then

Pharmacology. CP-55,940, rimonabant, and AM-630 were

modeled using PyMol to revise the structure of GBD-003 within

either CB R or CB R as the initial binding structure. The initial

obtained from Tocris Bioscience (Bristol, UK). All drugs were

prepared as a stock solution in 100% DMSO at a concentration of 100

mM, divided into aliquots, and maintained at −4 °C until use.

binding structures of these compounds were similarly energy-

minimized and had their binding free energy estimated by using the

same MMPBSA.py module.

[

H]CP-55,950 (168 Ci/mmol) and [ S]GTPγS (1250 Ci/mmol)

were purchased from Perkin Elmer (Boston, MA). All other reagents

were obtained from Fisher Scientiﬁc Inc. (Pittsburgh, PA).

Competition Receptor Binding. Increasing concentrations of

the GBD compounds were incubated with 0.2 nM of the nonselective

ASSOCIATED CONTENT

sı Supporting Information

■

CB /CB agonist [ H]CP-55,940 in a ﬁnal volume of 1 mL of the

binding buﬀer (50 mM Tris, 0.05% bovine serum albumin, 5 mM of

https://pubs.acs.org/doi/10.1021/acs.jmedchem.1c00442.

MgCl , pH 7.4) as described previously. Each binding assay

393

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

(2) Finn, D. P. Endocannabinoid-mediated Modulation of Stress

Article

Molecular formula strings (CSV )

Responses: Physiological and Pathophysiological Significance. Im-

munobiology 2010, 215, 629 −646.

Graphs correlating experimental and calculated binding

energies, copies of H and C NMR spectra and qNMR

(3) Bab, I.; Zimmer, A. Cannabinoid Receptors and the Regulation

analysis, and PDB ﬁles for docking models (PDF)

of Bone Mass. Br. J. Pharmacol. 2008 , 153 , 182 −188.

(4) Oesch, S.; Gertsch, J. Cannabinoid Receptor Ligands as Potential

AUTHOR INFORMATION

Corresponding Author

Gregory B. Dudley − C. Eugene Bennett Department of

Chemistry, West Virginia University, Morgantown, West

Virginia 26506, United States; orcid.org/0000-0001-

Anticancer Agents High Hopes for New Therapies? J. Pharm.

■

Pharmacol. 2009, 61, 839−853.

(5) Karsak, M.; Gaffal, E.; Date, R.; Wang-Eckhardt, L.; Rehnelt, J.;

Petrosino, S.; Starowicz, K.; Steuder, R.; Schlicker, E.; Cravatt, B.;

Mechoulam, R.; Buettner, R.; Werner, S.; Di Marzo, V.; Tuting, T.;

Zimmer, A. Attenuation of Allergic Contact Dermatitis Through the

Endocannabinoid System. Science 2007 , 316 , 1494 −1497.

133-3747 ; Email: gregory.dudley@mail.wvu.edu

(6) Guindon, J.; Hohmann, A. G. Cannabinoid CB2 receptors: a

Authors

therapeutic target for the treatment of inflammatory and neuropathic

Harvey F. Fulo − C. Eugene Bennett Department of

Chemistry, West Virginia University, Morgantown, West

Virginia 26506, United States

Amal Shoeib − Department of Pharmacology and Toxicology,

University of Arkansas for Medical Sciences, Little Rock,

Arkansas 72205, United States

Christian V. Cabanlong − Department of Pharmacology and

Toxicology, University of Arkansas for Medical Sciences, Little

Rock, Arkansas 72205, United States

Alexander H. Williams − Department of Pharmaceutical

Sciences, University of Kentucky, Lexington, Kentucky 40536,

United States

Chang-Guo Zhan − Department of Pharmaceutical Sciences,

University of Kentucky, Lexington, Kentucky 40536, United

States

Paul L. Prather − Department of Pharmacology and

Toxicology, University of Arkansas for Medical Sciences, Little

Rock, Arkansas 72205, United States

pain. Br. J. Pharmacol. 2008, 153, 319−334.

(7) Malfitano, A. M.; Ciaglia, E.; Gangemi, G.; Gazzerro, P.; Laezza,

C.; Bifulco, M. Update on the Endocannabinoid System as an

Anticancer Target. Expert Opin. Ther. Targets 2011, 15, 297−308.

(8) Cianchi, F.; Papucci, L.; Schiavone, N.; Lulli, M.; Magnelli, L.;

Vinci, M. C.; Messerini, L.; Manera, C.; Ronconi, E.; Romagnani, P.;

Donnini, M.; Perigli, G.; Trallori, G.; Tanganelli, E.; Capaccioli, S.;

Masini, E. Cannabinoid Receptor Activation Induces Apoptosis

Through Tumor Necrosis Factor Alpha-mediated Ceramide de

novo Synthesis in Colon Cancer Cells. Clin. Cancer Res. 2008, 14,

9) Velasco, G.; Sánchez, C.; Guzmán, M. Towards the Use of

691−7700.

Cannabinoids as Antitumour Agents. Nat. Rev. Cancer 2012, 12, 436−

44.

10) Pertwee, R. G. Targeting the Endocannabinoid System with

Cannabinoid Receptor Agonists: Pharmacological Strategies and

Therapeutic Possibilities. Philos. Trans. R. Soc. Lond. B Biol. Sci.

2012, 367, 3353−3363.

(11) Munro, S.; Thomas, K. L.; Abu-Shaar, M. Molecular

Characterization of a Peripheral Receptor for Cannabinoids. Nature

Complete contact information is available at:

https://pubs.acs.org/10.1021/acs.jmedchem.1c00442

12) Reggio, P. Endocannabinoid Binding to the Cannabinoid

993, 365, 61−65.

Receptors: What is Known and What Remains Unknown. Curr. Med.

Chem. 2010, 17, 1468−1486.

Author Contributions

All authors have given approval to the ﬁnal version of the

manuscript.

(

13) Matsuda, L. A.; Lolait, S. J.; Brownstein, M. J.; Young, A. C.;

Bonner, T. I. Structure of a Cannabinoid Receptor and Functional

Expression of the Cloned cDNA. Nature 1990 , 346 , 561 −564.

Funding

14) Svízenska, I.; Dubovy, P.; Sulcova, A. Cannabinoid Receptors 1

This work was supported in part by the funding from the

National Institutes of Health (R01 DA039143) for PLP, with

additional support from West Virginia University and the

Eberly Family Foundation. Molecular modeling studies were

supported by the National Institutes of Health (P20

GM130456 and T32 DA016176).

and 2 (CB and CB ), Their Distribution, Ligands and Functional

Involvement in Nervous System Structures  A Short Review.

Pharmacol. Biochem. Behav. 2008 , 90 , 501 −511.

(15) Ameri, A. The Effects of Cannabinoids on the Brain. Prog.

Neurobiol. 1999, 58, 315−348.

(16) Das, S. K.; Paria, B. C.; Chakraborty, I.; Dey, S. K. Cannabinoid

Notes

Ligand-Receptor Signaling in the Mouse Uterus. Proc. Natl. Acad. Sci.

U.S.A. 1995, 92, 4332−4336.

The authors declare no competing ﬁnancial interest.

(

17) Galiegue, S.; Mary, S.; Marchand, J.; Dussossoy, D.; Carriere,

D.; Carayon, P.; Bouaboula, M.; Shire, D.; Fur, G.; Casellas, P.

Expression of Central and Peripheral Cannabinoid Receptors in

Human Immune Tissues and Leukocyte Subpopulations. Eur. J.

Biochem. 1995, 232, 54−61.

ACKNOWLEDGMENTS

■

Prof Steven Kinsey (University of Connecticut) is gratefully

acknowledged for helpful discussions and assistance with early

development of this project.

(18) Gerard, C. M.; Mollereau, C.; Vassart, G.; Parmentier, M.

Molecular Cloning of a Human Cannabinoid Receptor Which is Also

ABBREVIATIONS

Expressed in Testis. Biochem. J. 1991, 279, 129−134.

■

(19) Wenger, T.; Ledent, C.; Csernus, V.; Gerendai, I. The Central

ECS, endocannabinoid system; HFIP, 1,1,1,3,3,3-hexaﬂuor-

oisopropanol; BINAP, (2,2′-bis(diphenylphosphino)-1,1′-bi-

naphthyl); SI, selectivity index; nbd, norbornadiene; n-BuLi,

n-butyllithium; t-BuLi, tert-butyllithium; MsCl, mesyl chloride;

Cannabinoid Receptor Inactivation Suppresses Endocrine Reproduc-

tive Functions. Biochem. Biophys. Res. Commun. 2001, 284, 363−368.

(

20) Benito, C.; Tolón, R. M.; Pazos, M. R.; Nunez, E.; Castillo, A.

I.; Romero, J. Cannabinoid CB ₂ Receptors in Human Brain

Tf O, triﬂic anhydride

Inflammation. Br. J. Pharmacol. 2008, 153, 277−285.

(21) Onaivi, E. S.; Ishiguro, H.; Gong, J.-P.; Patel, S.; Perchuk, A.;

REFERENCES

■

Meozzi, P. A.; Myers, L.; Mora, Z.; Tagliaferro, P.; Gardner, E.;

Brusco, A.; Akinshola, B. E.; Liu, Q.-R.; Hope, B.; Iwasaki, S.;

Arinami, T.; Teasenfitz, L.; Uhl, G. R. Discovery of the Presence and

1) Di Marzo, V. Targeting the Endocannabinoid System: To

Enhance or Reduce? Nat. Rev. Drug Discovery 2008, 7, 438−455.

394

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

(38) Pacher, P.; Bátkai, S.; Kunos, G. The Endocannabinoid System

Article

Functional Expression of Cannabinoid CB Receptors in Brain. Ann.

Associated Glia in Alzheimer ’s Disease Brains. J. Neurosci. 2003, 23,

11136−11141.

N.Y. Acad. Sci. 2006, 1074, 514−536.

(22) Maccarrone, M.; Bab, I.; Bíró, T.; Cabral, G. A.; Dey, S. K.; Di

as an Emerging Target of Pharmacotherapy. Pharmacol. Rev. 2006, 58,

389−462.

Marzo, V.; Konje, J. C.; Kunos, G.; Mechoulam, R.; Pacher, P.;

Sharkey, K. A.; Zimmer, A. Endocannabinoid Signaling at the

Periphery: 50 years After THC. Trends Pharmacol. Sci. 2015, 36, 277−

(39) Sagredo, O.; García-Arencibia, M.; de Lago, E.; Finetti, S.;

Decio, A.; Fernández-Ruiz, J. Cannabinoids and Neuroprotection in

Basal Ganglia Disorders. Mol. Neurobiol. 2007, 36 , 82 −91.

(40) Whiteside, G.; Lee, G.; Valenzano, K. The Role of the

Cannabinoid CB2 Receptor in Pain Transmission and Therapeutic

Potential of Small Molecule CB2 Receptor Agonists. Curr. Med. Chem.

96.

23) Maxwell, J. C. Psychoactive Substances  Some New, Some

Old: A Scan of the Situation in the U.S. Drug Alcohol Depend. 2014,

34, 71−77.

24) Ford, B. M.; Tai, S.; Fantegrossi, W. E.; Prather, P. L. Synthetic

Pot: Not Your Grandfather ’s Marijuana. Trends Pharmacol. Sci. 2017,

8, 257−276.

25) Banister, S. D.; Wilkinson, S. M.; Longworth, M.; Stuart, J.;

41) Cheng, Y.; Hitchcock, S. A. Targeting Cannabinoid Agonists

007, 14, 917−936.

for Inflammatory and Neuropathic Pain. Expert Opin. Invest. Drugs

007, 16, 951−965.

42) Yao, B. B.; Hsieh, G. C.; Frost, J. M.; Fan, Y.; Garrison, T. R.;

(

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: Canna-

bimimetic Drugs of Abuse. ACS Chem. Neurosci. 2013, 4, 1081−1092

and the references cited therein.

Daza, A. V.; Grayson, G. K.; Zhu, C. Z.; Pai, M.; Chandran, P.;

Salyers, A. K.; Wensink, E. J.; Honore, P.; Sullivan, J. P.; Dart, M. J.;

Meyer, M. D. In Vitro and In Vivo Characterization of A-796260: A

Selective Cannabinoid CB Receptor Agonist Exhibiting Analgesic

(

26) Wiley, J. L.; Marusich, J. A.; Lefever, T. W.; Grabenauer, M.;

Activity in Rodent Pain Models. Br. J. Pharmacol. 2008, 153, 390−

Moore, K. N.; Thomas, B. F. Cannabinoids in Disguise: Δ -

Tetrahydrocannabinol-like Effects of Tetramethylcyclopropyl Ketone

Indoles. Neuropharmacology 2013, 75, 145−154 and the references

cited therein.

(

01.

43) Ramirez, B. G.; Blazquez, C.; Gomez del Pulgar, T.; Guzman,

M.; de Ceballos, M. L. Prevention of Alzheimer ’s Disease Pathology

by Cannabinoids: Neuroprotection Mediated by Blockade of

Microglial Activation. J. Neurosci. 2005, 25, 1904−1913.

(27) Nutt, D. J.; King, L. A.; Nichols, D. E. Effects of Schedule I

Drug Laws on Neuroscience Research and Treatment Innovation.

(44) Body-Malapel, M.; El-Bakali, J.; Mérour, E.; Stern, E.; Millet,

Nat. Rev. Neurosci. 2013, 14, 577−585.

R.; Desreumaux, P. 741 4-Oxo-1,4-Dihydroquinoline-3-Carboxamides

Derivatives As New Potent and Selective Cb2 Agonists with Anti-

Inflammatory and Analgesic Properties in the Gut. Gastroenterology

(28) Huffman, J. W.; Zengin, G.; Wu, M.-J.; Lu, J.; Hynd, G.;

Bushell, K.; Thompson, A. L. S.; Bushell, S.; Tartal, C.; Hurst, D. P.;

Reggio, P. H.; Selley, D. E.; Cassidy, M. P.; Wiley, J. L.; Martin, B. R.

Structure-activity relationships for 1-alkyl-3-(1-naphthoyl)indoles at

the cannabinoid CB1 and CB2 receptors: steric and electronic effects

of naphthoyl substituents. New highly selective CB2 receptor agonists.

Bioorg. Med. Chem. 2005, 13, 89−112.

(

008, 134, A−107.

45) Benito, C.; Romero, J. P.; Tolón, R. M.; Clemente, D.;

Docagne, F.; Hillard, C. J.; Guaza, C.; Romero, J. Cannabinoid CB ₁

and CB Receptors and Fatty Acid Amide Hydrolase are Specific

Markers of Plaque Cell Subtypes in Human Multiple Sclerosis. J.

Neurosci. 2007, 27, 2396−2402.

(29) Drug Enforcement Administration. Schedules of Controlled

Substances: Temporary Placement of Five Synthetic Cannabinoids

(46) Pasquini, S.; Ligresti, A.; Mugnaini, C.; Semeraro, T.; Cicione,

into Schedule I. Fed. Regist. 2011, 76, 11075−11078.

L.; De Rosa, M.; Guida, F.; Luongo, L.; De Chiaro, M.; Cascio, M. G.;

Bolognini, D.; Marini, P.; Pertwee, R.; Maione, S.; Marzo, V. D.;

Corelli, F. Investigations on the 4-Quinolone-3-carboxylic acid Motif.

(30) Frost, J. M.; Dart, M. J.; Tietje, K. R.; Garrison, T. R.; Grayson,

G. K.; Daza, A. V.; El-Kouhen, O. F.; Yao, B. B.; Hsieh, G. C.; Pai, M.;

Zhu, C. Z.; Chandran, P.; Meyer, M. D. Indol-3-ylcycloalkyl Ketones:

Effects of N1 Substituted Indole Side Chain Variations on

CB2Cannabinoid Receptor Activity. J. Med. Chem. 2010, 53, 295−

3. Synthesis, Structure-affinity Relationships, and Pharmacological

Characterization of 6-Substituted 4-quinolone-3-carboxamides as

Highly Selective Cannabinoid-2 Receptor Ligands. J. Med. Chem.

(

15.

(

010, 53, 5915−5928 and the references cited therein.

31) Malan, T. P.; Ibrahim, M. M.; Lai, J.; Vanderah, T. W.;

47) Lunn, C. A.; Fine, J. S.; Rojas-Triana, A.; Jackson, J. V.; Fan, X.;

Makriyannis, A.; Porreca, F. CB2 cannabinoid receptor agonists: pain

relief without psychoactive effects? Curr. Opin. Pharmacol. 2003, 3,

Kung, T. T.; Gonsiorek, W.; Schwarz, M. A.; Lavey, B.; Kozlowski, J.

A.; Narula, S. K.; Lundell, D. J.; Hipkin, R. W.; Bober, L. A. A Novel

Cannabinoid Peripheral Cannabinoid Receptor-Selective Inverse

Agonist Blocks Leukocyte Recruitment in Vivo. J. Pharmacol. Exp.

Therapeut. 2006, 316, 780−788.

32) Manera, C.; Tuccinardi, T.; Martinelli, A. Indoles and Related

2−67.

Compounds as Cannabinoid Ligands. Mini Rev. Med. Chem. 2008, 8,

70−387.

33) Zhang, M.; Adler, M. W.; Abood, M. E.; Ganea, D.; Jallo, J.;

(

(48) Savinainen, J. R.; Kokkola, T.; Salo, O. M. H.; Poso, A.;

Järvinen, T.; Laitinen, J. T. Identification of WIN55212-3 as a

competitive neutral antagonist of the human cannabinoid CB2

receptor. Br. J. Pharmacol. 2005, 145, 636−645.

Tuma, R. F. CB2 receptor activation attenuates microcirculatory

dysfunction during cerebral ischemic/reperfusion injury. Microvasc.

Res. 2009, 78, 86−94.

(49) Deveaux, V.; Cadoudal, T.; Ichigotani, Y.; Teixeira-Clerc, F.;

(34) Shoemaker, J. L.; Seely, K. A.; Reed, R. L.; Crow, J. P.; Prather,

Louvet, A.; Manin, S.; Nhieu, J. T.-V.; Belot, M. P.; Zimmer, A.; Even,

P.; Cani, P. D.; Knauf, C.; Burcelin, R.; Bertola, A.; Le Marchand-

Brustel, Y.; Gual, P.; Mallat, A.; Lotersztajn, S. Cannabinoid CB2

Receptor Potentiates Obesity-Associated Inflammation, Insulin

Resistance and Hepatic Steatosis. PloS One 2009, 4, No. e5844.

(50) Frost, J. M.; Dart, M. J.; Tietje, K. R.; Garrison, T. R.; Grayson,

G. K.; Daza, A. V.; El-Kouhen, O. F.; Miller, L. N.; Li, L.; Yao, B. B.;

Hsieh, G. C.; Pai, M.; Zhu, C. Z.; Chandran, P.; Meyer, M. D. Indol-

3-yl-tetramethylcyclopropyl Ketones: Effects of Indole Ring Sub-

stitution on CB2Cannabinoid Receptor Activity. J. Med. Chem. 2008,

51, 1904−1912.

P. L. The CB2 cannabinoid agonist AM-1241 prolongs survival in a

transgenic mouse model of amyotrophic lateral sclerosis when

initiated at symptom onset. J. Neurochem. 2007 , 101 , 87 −98.

(35) Croxford, J. L.; Miller, S. D. Immunoregulation of a viral model

of multiple sclerosis using the synthetic cannabinoid R(+)-

WIN55,212. J. Clin. Invest. 2003, 111, 1231−1240.

(36) Fernández-Ruiz, J.; Romero, J.; Velasco, G.; Tolón, R. M.;

Ramos, J. A.; Guzmán, M. Cannabinoid CB2 receptor: a new target

for controlling neural cell survival? Trends Pharmacol. Sci. 2007, 28,

(

9−45.

37) Benito, C.; Nun

ez, E.; Tolón, R. M.; Carrier, E. J.; Rábano, A.;

(51) Drug Enforcement Administration. Schedules of Controlled

Substances: Placement of UR-144, XLR11, and AKB48 into Schedule

I. Fed. Regist. 2016, 81, 29142−29145.

Hillard, C. J.; Romero, J. Cannabinoid CB2Receptors and Fatty Acid

Amide Hydrolase Are Selectively Overexpressed in Neuritic Plaque-

395

J. Med. Chem. 2021, 64, 6381−6396

Journal of Medicinal Chemistry

Crooks, P. A.; Zhan, C.-G. Binding Modes and Selectivity of

Article

(

52) Regulatory agencies including the US Drug Enforcement

(67) Yang, J.-F.; Williams, A. H.; Penthala, N. R.; Prather, P. L.;

Cannabinoid 1 (CB1) and Cannabinoid 2 (CB2) Receptor Ligands.

ACS Chem. Neurosci. 2020, 11, 3455−3463.

(68) Hawkins, P. C. D.; Skillman, A. G.; Warren, G. L.; Ellingson, B.

A.; Stahl, M. T. Conformer generation with OMEGA: algorithm and

validation using high quality structures from the Protein Databank

and Cambridge Structural Database. J. Chem. Inf. Model. 2010, 50,

Agency consider “history and current pattern of abuse”. “evidence that

individuals are taking the drug or other substance in amounts

suﬃcient to create a hazard”; and “signiﬁcant diversion of the drug or

other substance from legitimate drug channels” as key “indicators that

a drug or other substance has a potential for abuse”; see: Drugs of

Abuse: A DEA Resource Guide, 2020 Edition, April 13, 2020. see

https://www.dea.gov/documents/2020/04/13/drugs-abuse (ac-

cessed Apr 4 2021).

(

72−584.

69) Morris, G. M.; Huey, R.; Lindstrom, W.; Sanner, M. F.; Belew,

53) Wiley, J. L.; Marusich, J. A.; Huffman, J. W. Moving Around the

R. K.; Goodsell, D. S.; Olson, A. J. AutoDock4 and AutoDockTools4:

Automated docking with selective receptor flexibility. J. Comput.

Chem. 2009, 30, 2785−2791.

Molecule: Relationship Between Chemical Structure and in vivo

Activity of Synthetic Cannabinoids. Life Sci. 2014, 97, 55−63.

(

Falciani, C.; Pedani, V.; Pesco, N.; Guida, F.; Luongo, L.; Varani, K.;

Borea, P. A.; Maione, S.; Di Marzo, V.; Corelli, F. Design, Synthesis,

and Pharmacological Characterizationof Indol-3- ylacetamides, Indol-

54) Pasquini, S.; Mugnaini, C.; Ligresti, A.; Tafi, A.; Brogi, S.;

(70) Trott, O.; Olson, A. J. AutoDock Vina: improving the speed

and accuracy of docking with a new scoring function, efficient

optimization, and multithreading. J. Comput. Chem. 2010, 31, 455−

(

61.

-yloxoacetamides, and Indol-3-ylcarboxamides: Potent and Selective

CB2 Cannabinoid Receptor Inverse Agonists. J. Med. Chem. 2012, 55,

391−5402.

55) Tavakoli, A.; Dudley, G. B. Synthesis of 4,4-Dimethyl-1,6-

heptadiyne and Alcyopterosin O. Org. Lett. 2020 , 22 , 8947 −8951.

56) Kramer, N. J.; Hoang, T. T.; Dudley, G. B. Reaction Discovery

71) Hua, T.; Vemuri, K.; Pu, M.; Qu, L.; Han, G. W.; Wu, Y.; Zhao,

S.; Shui, W.; Li, S.; Korde, A.; Laprairie, R. B.; Stahl, E. L.; Ho, J.-H.;

Zvonok, N.; Zhou, H.; Kufareva, I.; Wu, B.; Zhao, Q.; Hanson, M. A.;

Bohn, L. M.; Makriyannis, A.; Stevens, R. C.; Liu, Z.-J. Crystal

structure of the human cannabinoid receptor CB1. Cell 2016, 167,

Using Neopentylene-Tethered Coupling Partners: Cycloisomeriza-

50−762.e714.

tion/Oxidation of Electron-Deficient Dienynes. Org. Lett. 2017, 19,

72) Li, X.; Hua, T.; Vemuri, K.; Ho, J.-H.; Wu, Y.; Wu, L.; Popov,

P.; Benchama, O.; Zvonok, N.; Locke, K. a.; Qu, L.; Han, G. W.; Iyer,

M. R.; Cinar, R.; Coffey, N. J.; Wang, J.; Wu, M.; Katritch, V.; Zhao,

S.; Kunos, G.; Bohn, L. M.; Makriyannis, A.; Stevens, R. C.; Liu, Z.-J.

Crystal structure of the human cannabinoid receptor CB2. Cell 2019,

57) Bailey, W. F.; Longstaff, S. C. Generation and Cyclization of a

636−4639.

Benzyne-Tethered Alkyllithium: Preparation of 4-Substituted Indans.

J. Org. Chem. 1998, 63, 432−433.

58) Hoang, T. T.; Dudley, G. B. Synthesis of High-Value 1,6-

76, 459−467 e13.

73) Wang, J.; Wang, W.; Kollman, P. A.; Case, D. A. Antechamber:

Enynes by Tandem Fragmentation/Olefination. Org. Lett. 2013, 15,

026−4029.

59) Aung, M. M.; Griffin, G.; Huffman, J. W.; Wu, M.-J.; Keel, C.;

an accessory software package for molecular mechanical calculations.

J. Am. Chem. Soc 2001, 222, U403.

(

(74) Pearlman, D. A.; Case, D. A.; Caldwell, J. W.; Ross, W. S.;

Yang, B.; Showalter, V. M.; Abood, M. E.; Martin, B. R. Influence of

^t ^h ^e ^N ^-¹ ^A ^l ^k ^y ^l ^C ^h ^a ⁱ ⁿ ^L ^e ⁿ ^g ^t ^h ^o ^f ^C ^a ⁿ ⁿ ^a ^b ⁱ ^m ⁱ ^m ^e ^t ⁱ ^c ^I ⁿ ^d ^o ^l ^e ^s ^u ^p ^o ⁿ ^C ^B 1

and CB Receptor Binding. Drug Alcohol Depend. 2000, 60, 133−140.

(

J.; Chen, B.-C.; Zhao, R.; Kiener, P. A.; Chen, X.; Turk, L. A.; Patil-

Koota, V.; Gillooly, K. M.; Shuster, D. J.; McIntyre, K. W. C-3 Amido-

Indole Cannabinoid Receptor Modulators. Bioorg. Med. Chem. Lett.

Cheatham, T. E., III; DeBolt, S.; Ferguson, D.; Seibel, G.; Kollman, P.

AMBER, a package of computer programs for applying molecular

mechanics, normal mode analysis, molecular dynamics and free

energy calculations to simulate the structural and energetic properties

of molecules. Comput. Phys. Commun. 1995, 91, 1−41.

(75) Miller, B. R., III; McGee, T. D., Jr.; Swails, J. M.; Homeyer, N.;

Gohlke, H.; Roitberg, A. E. MMPBSA py: an efficient program for

end-state free energy calculations. J. Chem. Theory Comput. 2012, 8,

3314−3321.

60) Hynes, J.; Leftheris, K.; Wu, H.; Pandit, C.; Chen, P.; Norris, D.

61) Longworth, M.; Connor, M.; Banister, S. D.; Kassiou, M.

002, 12, 2399−2402.

Synthesis and Pharmacological Profiling of the Metabolites of

Synthetic Cannabinoid Drugs APICA, STS-135, ADB-PINACA, and

(76) DeLano, W. L. Pymol: An open-source molecular graphics tool.

CCP4 Newsl. protein crystallogr. 2002, 40, 82−92.

(

FADB-PINACA. ACS Chem. Neurosci. 2017, 8, 1673−1680.

62) Brents, L. K.; Gallus-Zawada, A.; Radominska-Pandya, A.;

Vasiljevik, T.; Prisinzano, T. E.; Fantegrossi, W. E.; Moran, J. H.;

Prather, P. L. Mono-hydroxylated Metabolites of the K2 Synthetic

Cannabinoid JWH-073 Retain High Cannabinoid 1 Receptor (CB R)

Affinity and Exhibit Neutral Antagonist to Partial Agonist Activity.

Biochem. Pharmacol. 2012, 83, 952−961.

(63) Klamo, S. B.; Wendt, O. F.; Henling, L. M.; Day, M. W.;

Bercaw, J. E. Dialkyl and Methyl-Alkyl Zirconocenes: Synthesis and

Characterization of Zirconocene-Alkyls That Model the Polymeryl

Chain in Alkene Polymerizations. Organometallics 2007, 26, 3018−

64) Vekariya, R. H.; Aubé, J. Hexafluoro-2-propanol-Promoted

030.

Intermolecular Friedel −Crafts Acylation Reaction. J. Org. Lett. 2016,

8, 3534−3537.

65) Hudlicky, T.; Frazier, J. O.; Seoane, G.; Tiedje, M.; Seoane, A.;

Kwart, L. D.; Beal, C. Topological selectivity in the intramolecular [4

(

1] pyrroline annulation. Formal total stereospecific synthesis of (.

-.)-supinidine, (.+-.)-isoretronecanol, and (.+-.)-trachelanthamidine.

J. Am. Chem. Soc. 1986, 108, 3755−3762.

(66) 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,

No. e21917.

396