Deciphering Structure–Activity Relationships in a Series of 2,2-Dimethylchromans Acting as Inhibitors of Insulin Release and Smooth Muscle Relaxants

Article abstract of DOI:10.1002/cmdc.201700409

4,6-Disubstituted 2,2-dimethylchromans are reported as pharmacologically active compounds that mainly target the ATP-sensitive potassium channels. The present study is an attempt to characterize the impact of the nature of substituents introduced at the 4- and 6-positions of 2,2-dimethylchromans on their capacities to inhibit insulin release from pancreatic β-cells or to relax vascular smooth muscle cells, both biological responses that are supposed to reflect interaction with specific ion channels. From the core structure 4-amino-2,2-dimethylchroman, a progressive increase in the steric hindrance of the chemical functionalities introduced at the 4-position (amino, formamido, acetamido, arylureido/thioureido) and at the 6-position (amino, formamido, acetamido, alkoxycarbonylamino) led to a progressive magnification of the inhibitory effect on the insulin release process and, to a lesser extent, of the vasorelaxant activity. Moreover, the dextrorotatory enantiomer of 2,2-dimethylchroman compound 29 was more potent than its levorotatory counterpart for inhibiting the insulin secretory process. Additional pharmacological investigations suggested, however, that the myorelaxant activity of 11 and 15 resulted from a direct Ca²⁺ entry blockade.

Full text of DOI:10.1002/cmdc.201700409

Accepted Article
Title: Deciphering structure-activity relationships in a series of 2,2-
dimethylchromans acting as inhibitors of insulin release and
smooth muscle relaxants
Authors: Bernard Pirotte, Xavier Florence, Eric Goffin, and Philippe
Lebrun
This manuscript has been accepted after peer review and appears as an
Accepted Article online prior to editing, proofing, and formal publication
of the final Version of Record (VoR). This work is currently citable by
using the Digital Object Identifier (DOI) given below. The VoR will be
published online in Early View as soon as possible and may be different
to this Accepted Article as a result of editing. Readers should obtain
the VoR from the journal website shown below when it is published
to ensure accuracy of information. The authors are responsible for the
content of this Accepted Article.
To be cited as: ChemMedChem 10.1002/cmdc.201700409
Link to VoR: http://dx.doi.org/10.1002/cmdc.201700409
A Journal of
www.chemmedchem.org

10.1002/cmdc.201700409
ChemMedChem
Deciphering structure-activity relationships in a series of 2,2-dimethylchromans
acting as inhibitors of insulin release and smooth muscle relaxants
Prof. Bernard Pirotte1*, Dr. Xavier Florence1,2, Eric Goffin1, Prof. Philippe Lebrun2
1 Laboratoire de Chimie Pharmaceutique, Center for Interdisciplinary Research on Medicines
(CIRM), Université de Liège, Quartier Hôpital, Avenue Hippocrate 15, B-4000 Liège,
Belgium
2
Laboratoire de Physiologie et Pharmacologie, Université Libre de Bruxelles, Faculté de
Médecine, 808 Route de Lennik, B-1070 Bruxelles, Belgium
* Corresponding author
Keywords
2,2-Dimethylchromans; ATP-sensitive potassium channels; potassium channel openers;
insulin secretion; smooth muscle contractile activity
Abstract
4,6-Disubstituted 2,2-dimethylchromans are reported as pharmacologically active compounds
mainly targeting the ATP-sensitive potassium channels (KATP channels). The present study is
an attempt to characterize the impact of the nature of the substituent introduced at the 4- and
6-position of 2,2-dimethylchromans on their capacities to inhibit insulin release from
pancreatic β-cells or to relax vascular smooth muscle cells, both biological responses
supposed to reflect interaction with specific ion channels.
Starting from the core structure 4-amino-2,2-dimethylchroman, the progressive increase of
the steric hindrance of the chemical functions introduced at the 4-position (amino,
formamido, acetamido, arylureido/thioureido) and at the 6-position (amino, formamido,
1
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
acetamido, alkoxycarbonylamino) led to a progressive magnification of the inhibitory effect
on the insulin releasing process and, to a lesser extent, of the vasorelaxant activity.
Moreover, the dextrorotatory enantiomer of one selected 2,2-dimethylchroman racemic
compound (29) was more potent than its levorotatory counterpart at inhibiting the insulin
secretory process.
Additional pharmacological investigations suggested, however, that the myorelaxant activity
of (11) and (15) resulted from a direct Ca++ entry blockade.
Manuscript text
1. Introduction
Molecules containing the 2,2-dimethylchroman core structure are the object of a continuous
and intensive research in medicinal chemistry. Such heterocyclic drugs are known to be
found in several biologically active natural compounds [1-3], but also among synthetic
[4-7]
compounds expressing multiple pharmacological activities.
Regarding the synthetic
compounds, the well-known potassium channel opener cromakalim (1), an example of 4,6-
disubstituted 2,2-dimethylchroman, acted as a model for the design of an impressive number
of analogues; most of which developed as smooth muscle relaxants (vasodilators,
bronchodilators, oxytocic agents, …). [8-10]
The myorelaxant activity of cromakalim and of many analogues appears to be related to the
activation of potassium channels described as ATP-sensitive potassium channels (KATP
[8]
channels).
These channels are involved in major physiological processes, such as the
control of insulin release from pancreatic β-cells and the modulation of vascular smooth
[11-13]
muscle tone.
It is now well established that KATP channels are octameric complexes of
four pore-forming inwardly rectifying K+ channels (Kir6.1 or Kir6.2) and four regulatory
sulfonylurea receptor subunits (SUR1, SUR2A or SUR2B). [14] Different combinations of Kir
and SUR subunits lead to tissue-specific KATP channels; the pancreatic β-cell is known to
express a SUR1/Kir6.2 KATP channel subtype while the SUR2B/Kir6.1 combination is found
[15,16]
in vascular smooth muscle cells.
Modifications of the smooth muscle tone and insulin
secretory process may also be observed with drugs targeting the L-type voltage-operated
calcium channels. [17]
2
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
In such a context, we previously described novel series of cromakalim analogues.[18-26] Figure
1 reports typical examples of 2,2-dimethylchromans depicted by our research group. Most
drugs were characterized by the presence of an electronegative substituent at the 6-position of
the chroman nucleus (halogen atom, sulfonamide group, carbamate group) and by the
presence, at the 4-position, of an amide function or an urea/thiourea function as possible
isosteric substitute. Some compounds were described as KATP channel openers (PCOs) (i.e.
compound 4 [22]), while others were found to behave as PCOs as well as voltage-dependent
calcium channel blockers (i.e. compound 8 [26]) or only as calcium entry blockers (i.e.
compound 6 [25]).
The present study is an attempt to decipher the impact of the structural elements introduced at
the 4- and the 6-positions of the 2,2-dimethylchroman core structure on the biological activity
(inhibition of insulin release and vascular smooth muscle relaxation) of selected compounds.
Both biological responses could reflect a possible interaction of the 2,2-dimethylchromans
with potassium and/or calcium channels.
Figure 2A illustrated the compounds synthesized in the present study in order to characterize
the critical importance of the nature of the substituent at the 4-position when the moiety
introduced at the 6-position remained constant (except for 9 devoid of substituent at the 6-
position). The impact of the nature of the substituent at the 6-position was also investigated
using compounds bearing a constant substituent at the 4-position (Fig. 2B).
Finally, these newly synthesized drugs bearing a chiral carbon atom at the 4-position, an
attempt was made to separate the two enantiomers of one selected 2,2-dimethylchroman
racemic compound (i.e. compound 29) and to examine potential differences in biological
activities (inhibition of insulin release and vascular smooth muscle relaxation).
2. Results and discussion
2.1. Chemistry
The synthesis of the new compounds (9-15) is described on schemes 1-3. The starting
compound 3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-one 21 was obtained from the
reaction of 2-hydroxyacetophenone 20 with acetone in the presence of pyrrolidine in
methanol (scheme 1). Compound 21 was converted into the corresponding oxime 22 by
means of hydroxylamine and the latter compound was reduced by hydrogenation in ethanolic
3
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
solution in the presence of Raney Nickel° to provide the target compound R/S-amino-3,4-
dihydro-2,2-dimethyl-2H-1-benzopyran 9 (scheme 1).
The key intermediate 6-bromo-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-ol 27, obtained
as previously described [20], reacted with cuprous cyanide in 1-methyl-2-pyrrolidinone to
provide the 6-cyano-substituted intermediate 28 (scheme 2). The latter compound was
converted into the corresponding 4-acetamido-substituted target compound 13 in the
conditions of the Ritter reaction [27] (scheme 2).
Compound 11, obtained as previously described [24], was acylated in the presence of a formic
acid-acetic anhydride mixture to provide compound 14, or with acetic anhydride to provide
compound 15 (scheme 3).
Lastly, the two enantiomers of the racemic intermediate R/S-4-amino-3,4-dihydro-2,2-
[26]
dimethyl-6-ethoxycarbonylamino-2H-1-benzopyran
were obtained after chromatographic
separation by preparative chiral HPLC using a Chiralcel OJ column. Each enantiomer then
reacted with 3-cyanophenyl isocyanate to provide the (+)-29 enantiomer [from (-)-4-amino-
3,4-dihydro-2,2-dimethyl-6-ethoxycarbonylamino-2H-1-benzopyran]
and
the
(-)-29
enantiomer [from (+)-4-amino-3,4-dihydro-2,2-dimethyl-6-ethoxycarbonylamino-2H-1-
benzopyran].
2.2. Biological results
The biological effects of the selected dimethylchromans (8-19), as well as of reference
compounds (±)-cromakalim and diazoxide, on the glucose-induced insulin release from rat
pancreatic islets and on the contractile activity of K+-depolarized rat aorta rings are reported
in Table 1.
The core structure by itself, i.e. the 2,4-dimethylchroman nucleus bearing exclusively an
amino group at the 4-position (compound 9), was devoid of any activity on the pancreatic β-
cells (inhibition of the insulin releasing process) and on the vascular smooth muscle cells
(myorelaxant activity).
By introducing a bulky ‘carbamate’ group at the 6-position, providing compounds 10 and 11,
a significant inhibitory activity on insulin release was observed at 50 µM for the isopropyl-
substituted compound 10 (25% inhibition of insulin release), while the tert-butyl-substituted
homologue 11 expressed an identical inhibitory effect at 10 µM (Table 1). Such a carbamate
4
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
group introduced at the 6-position was also responsible for a major increase in the
myorelaxant activity (Table 1).
By comparing 11 with 14 and 15, namely compounds with similar tert-butoxycarbonylamino
moiety at the 6-position, it was observed that conversion of the primary amine function
linked at the 4-position to an amide function (formamide 14 or acetamide 15) did not improve
the inhibitory activity on the insulin releasing process and the vasorelaxant effect (Table 1).
Looking at the substituent located at the 6-position for compounds bearing identical
acetamido group at the 4-position (see compounds 12, 13 and 15), it can be concluded that
the nature of the 6-substituent exerted a rather small impact on the myorelaxant activity,
although the 6-cyano-substituted compound 13 was recognized as the most potent vasodilator
(EC50 = 2.1 µM; Table 1). Such compounds (12, 13 and 15) also failed to exert a pronounced
effect on the glucose-induced insulin releasing process (Table 1).
When the substituent at the 4-position was a 3-cyanophenylthiourea moiety and the
substituent at the 6-position transformed from a primary amine function to the bulky tert-
butoxycarbonylamino group (see 16 to 19), a progressive improvement of the activity on
[24]
pancreatic β-cells was clearly noticed (16 < 17 < 18 < 19) (Table 1).
On the vascular
smooth muscle cells, such an improvement of activity was also observed (16,17 < 18 < 19)
(Table 1). [24]
Moreover, by comparing the myorelaxant properties of the 2,2-dimethylchromans 11 and 19,
it can be noticed that compound 11 devoid of the 3-cyanophenylthiourea group at the 4-
position was equipotent to the corresponding 3-cyanophenylthiourea-substituted compound
19 (Table 1).
Lastly, the conversion of the thiourea function of compound 19 into an urea function
(providing compound 8) was found to have no impact on the myorelaxant activity while the
inhibitory effect on the insulin releasing process was markedly improved. [26]
Compound 29, the ethoxy analogue of compound 8, was selected for further characterizing
the impact of the geometry of the carbon atom at the 4-position on the biological activity in
pancreatic β-cells and vascular smooth muscle cells (Table 1). As previously reported, the
ethoxy compound 29 was found to be less potent than its tert-butoxy analogue 8 on the
insulin releasing process, while the myorelaxant activity of the two compounds was roughly
similar. [26] The two enantiomers of the racemic compound 29 were separately prepared and
tested as inhibitors of the glucose-induced insulin releasing process and as vascular smooth
muscle relaxants (Table 1). The dextrorotatory enantiomer (+)-29 was found to be clearly
5
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
more potent than its levorotatory counterpart (-)-29 on pancreatic β-cells [IC50 near 1 µM for
(+)-29 and > 10 µM for (-)-29]. However, the difference in potency was less pronounced
regarding their vascular myorelaxant activity. At this stage, the absolute configuration of the
4-carbon atom of each separated enantiomer remained to be determined.
No critical information can be deduced from the virtual logP values (‘average’ logP values
[29]) calculated for these compounds (Table 1), except that the introduction of a bulky tert-
butyloxycarbonylamino group at the 6-position exhibited small impact on the calculated logP
values (see 9 versus 11; Table 1) while the introduction of a 3-cyanophenylurea/thiourea
moiety instead of an amino group at the 4-position considerably increased the lipophilicity of
the target compounds (see 11 versus 8 and 19; Table 1).
In order to decipher the mechanism of action of compounds 11 and 15 on the vascular
smooth muscle cells, the myorelaxant activity was further characterized on rat aortic rings
precontracted by 30 mM KCl in the presence of glibenclamide (1 µM or 10 µM) or
precontracted by 80 mM extracellular KCl.
The concomitant presence of the KATP channel blocker glibenclamide (1 µM or 10 µM) in the
bathing solution failed to affect the myorelaxant properties of 11 and 15, as it was observed
[25,28]
for the calcium entry blocker verapamil (Table 2).
By contrast, the KATP channel
blocker glibenclamide induced a dose-dependent reduction in the vasorelaxant response to
the potassium channel openers (±)-cromakalim and diazoxide (Table 2).
Table 2 also indicates that, when the aorta rings were precontracted by 80 mM KCl, the
vasorelaxant potency of compound 11 or 15 was unaffected. In the presence of 80 mM KCl
in the bathing medium, however, the myorelaxant effects of the potassium channel openers
(±)-cromakalim and diazoxide were drastically reduced whilst those of the calcium entry
blocker verapamil were unaffected (Table 2). [28]
These findings indicate that, on vascular smooth muscle cells, 11 and 15 mainly behaved as
calcium entry blockers.
Further electrophysiological investigations are required to confirm the target site of such
compounds at the plasma membrane level.
3. Conclusions
The aim of this study was to explore the impact of the nature of the substituent introduced at
the 4- and 6-position of 2,2-dimethylchromans on their abilities to exert either an inhibitory
6
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
effect on the insulin releasing process or to relax vascular smooth muscle; both biological
responses reflecting a possible interaction with specific potassium and/or calcium channels.
The simple core structure, 4-amino-2,2-dimethylchroman 9, was devoid of activity on the
endocrine and smooth muscle tissues. The introduction of a bulky alkoxycarbonylamino
group at the 6-position, providing ‘carbamate’ derivatives 10 and 11, was decisive to generate
active compounds.
Keeping the bulky tert-butyloxycarbonylamino group at the 6-position, conversion of the 4-
amino group by acylation failed to improve the biological activities (see compounds 14 and
15) while the presence of an arylureido group at the 4-position led to a remarkable increase of
activity on the endocrine tissue with no change in the myorelaxant properties (see compound
8). Such findings confirm the critical importance for the presence of a bulky carbamate group
at the 6-position associated with an arylureido group at the 4-position to generate compounds
exhibiting a marked inhibitory activity on the insulin releasing process.
Moreover, the geometry of the carbon atom at the 4-position of a 6-alkoxycarbonylamino-
2,2-dimethylchroman bearing a 4-arylurea moiety was found to be crucial for eliciting
inhibitory activity on the endocrine tissue (the dextrorotatory enantiomer being more potent
than the levorotatory enantiomer).
Interestingly, the carbamate derivatives 11 and 15 exerted a myorelaxant activity related to a
mechanism similar to calcium channel blockers.
4. Experimental section
4.1. Chemistry
Reagents and solvents were purchased from usual commercial suppliers and were used
without further purification. Yields reported refer to purified products. All reactions were
routinely checked by thin-layer chromatography (TLC) on silica gel 60 F254 (Merck) and
visualization was performed by UV light (254 nm). Melting points were determined on a
Stuart SMP3 apparatus in open capillary tubes and were uncorrected. IR spectra were
1
recorded as KBr pellets on a Perkin–Elmer 1000 FTIR spectrophotometer. The H NMR
spectra were recorded on a Bruker Avance (500 MHz) instrument using DMSO-d6 as solvent
and tetramethylsilane (TMS) as internal standard; chemical shifts were reported in δ values
(ppm) relative to internal TMS. The abbreviation s = singlet, d = doublet, t = triplet, q =
7
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
quadruplet, m = multiplet and b = broad signal were used throughout. Elemental analyses (C,
H, N, S) were determined on a Thermo Flash EA 1112 series elemental analyzer and were
within ±0.4% of the theoretical values, except otherwise specified (in italic).
R/S-4-amino-6-tert-butylcarbonylamino-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran
(11),
bromophenyl acetate (24), 5-bromo-2-hydroxyacetophenone, (25), 6-bromo-3,4-dihydro-2,2-
dimethyl-2H-1-benzopyran-4-one (26) and R/S-6-bromo-3,4-dihydro-2,2-dimethyl-2H-1-
benzopyran-4-ol (27) were obtained according to previously described procedures. [20,24]
4.1.1. 3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-one (21). 2-Hydroxyacetophenone (20)
(2.00 g, 1.76 mL, 14.7 mmol) was added to a solution of pyrrolidine (1.72 mL, 21 mmol),
acetone (3.62 mL, 49 mmol) and methanol (20 mL). The mixture was stirred overnight at
room temperature. On the next day, the solvents were removed under reduced pressure.
Water was added to the residue and the mixture was adjusted to pH 1 with concentrated
hydrochloric acid. The resulting precipitate was collected by filtration. The crude product
was then dissolved in methanol. The solution was treated with charcoal, filtered and water
was added to the filtrate. The title compound was collected by filtration, washed with water
and dried. The crude product was purified by chromatography on a silica gel 60 column
1
eluted by chloroform (1.4 g, 54 %): mp: 88-90 °C; IR (KBr): υ 1687 (C=O) cm-1; H NMR
(DMSO-d6, 500 MHz): δ 1.39 (s, 6H, CH3), 2.79 (s, 2H, CH2), 6.98 (d, J = 8.3 Hz, 1H, 8-H),
7.02 (t, J = 7.6 Hz, 1H, 6-H), 7.55 (t, J = 7.8 Hz, 1H, 7-H), 7.72 (d, J = 7.8 Hz, 1H, 5-H); 13C
NMR (DMSO-d6): δ 26.0 (CH3), 47.9 (C-3), 79.3 (C-2), 118.2 (C-8), 119.8 (C-4a), 120.6 (C-
6), 125.8 (C-5), 136.3 (C-7), 159.4 (C-8a), 192.0 (C-4). Anal. Calcd for C11H12O2: C 74.98,
H, 6.86, found: C 74.64, H 6.96.
4.1.2. 3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-hydroxyimine (22). Potassium carbonate
(23.80 g, 0.172 mol) and hydroxylamine hydrochloride (11.90 g, 0.172 mol) were added to a
stirred solution of 3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-one (21) (15.00 g, 0.085
mol) in ethanol (250 ml). The suspension was refluxed for 3 h. The reaction was then poured
onto ice and, after complete melting of ice, water was added to obtain a final volume of 195
ml. The title compound was collected by filtration, washed with water and dried (14,97 g, 92
%): mp: 123-125 °C; IR (KBr): υ 931 (N-O), 1260 (C-O), 1650 (C=N), 2935 (aliphatic C-H),
2977 (aromatic C-H), 3264 (O-H) cm-1; 1H NMR (DMSO-d6, 500 MHz): δ 1.30 (s, 6H, CH3),
8
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
2.77 (s, 2H, CH2), 6.82 (d, J = 8.2 Hz, 1H, 8-H), 6.89 (t, J = 7.7 Hz, 1H, 6-H), 7.24 (t, J = 7.8
Hz, 1H, 7-H), 7.74 (d, J = 7.8 Hz, 1H, 5-H), 11.19 (s, 1H, =N-OH). Anal. calcd for
C11H13NO2: C 69.09, H 6.85, N 7.32, found: C 69.40, H 6.77, N 7.34.
4.1.3. R/S-4-amino-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran (9). Raney-Nickel (3 g) was
added to a stirred solution of 3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-hydroxyimine
(22) (10.00 g, 0.052 mol) in methanol. The solution was stirred in a sealed hydrogenator
under a hydrogen pressure of 5 bars. When the reaction was ended, the catalyst was filtered
off and the filtrate was evaporated under reduced pressure. The crude product was purified by
chromatography on a silica gel 60 column eluted by a chloroform / ethyl acetate mixture
(15:5). Fractions containing the title compound were pooled and the solvent was evaporated
under reduced pressure. The resulting oil was dissolved in dry ether and treated by gazeous
HCl. The resulting chlorhydrate was collected by filtration, washed with diethyl ether and
dried (8.35 g, 90 %): mp: 268 °C; IR (KBr): υ 1260 (C-O), 2935 (aliphatic C-H), 2977
(aromatic C-H), 3436 (N-H2) cm-1; 1H NMR (DMSO-d6, 500 MHz): δ 1.22 (s, 3H, CH3), 1.42
(s, 3H, CH3), 1.82 (t, J = 12.3 Hz, 1H, CH2), 2.32 (dd, J = 13.1/6.4 Hz, 1H, CH2), 4.49 (dd, J
= 11.5/6.4 Hz, 1H, CH-NH2), 6.80 (d, J = 7.7 Hz, 1H, 8-H), 6.96 (t, J = 7.1 Hz, 1H, 6-H),
13
7.24 (t, J = 7.2 Hz, 1H, 7-H), 7.69 (d, J = 7.7 Hz, 1H, 5-H), 8.80 (s, 3H, CH-NH2.HCl); C
NMR (DMSO-d6): δ 23.6 (CH3), 29.3 (CH3), 37.4 (C-3), 43.2 (C-4), 74.6 (C-2), 117.3 (C-8),
118.7 (C-4a), 120.2 (C-6), 127.1 (C-5), 129.8 (C-7), 153.1 (C-8a). Anal. calcd for
C11H15NO.HCl: C 61.82, H 7.55, N 6.55, found: C 61.82, H 7.59, N 6.77.
4.1.4. R/S-6-cyano-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-ol (28). A mixture of R/S-
[20]
6-bromo-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-ol (27)
(9.00 g, 0.035 mol) and
Copper(I) cyanide (6.30 g, 0.070 mol) in 1-Methyl-2-pyrrolidinone (180 mL) was refluxed
for 6 hours. After completion of the reaction, the solvents were removed under reduced
pressure. Water was added to the residue and the solution was extracted by ethyl acetate. The
organic layer was treated with charcoal, dried over MgSO4 and evaporated under reduced
pressure. The resulting oil was used in the next step without further purification (7.11 g, 100
%), IR (KBr): υ 2224 (C≡N), 2978 (aliphatic C-H), 3436 (O-H) cm-1.
9
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
4.1.5. R/S-4-(N-methylcarbonylamino)-6-cyano-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran
(13). A suspension of R/S-6-cyano-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran-4-ol (28)
(7.00 g, 34.44 mmol) in acetonitrile (60 mL) was added dropwise to a stirred solution of
acetonitrile (12 mL) in 98% sulfuric acid (3 mL) kept between -10 and 0 °C. Stirring was
continued for 1 h at room temperature. The solution was poured into cold water and the
solution was concentrated under reduced pressure. The resulting precipitate was collected by
filtration, washed with water and dried. The crude product was solubilized in a minimal
volume of ethyl acetate and the title compound was precipitated by the addition of 3 volumes
of petroleum ether (bp : 40-46 °C). Final compound was collected by filtration, washed by
petroleum ether (bp : 40-46 °C) and dried (1.85 g, 22 %): mp: 163-165 °C; IR (KBr): υ 1656
(C=O), 2226 (C≡N), 2933, 2950, 2978 (aliphatic C-H), 3076 (aromatic C-H), 3256 (N-H)
1
cm-1; H NMR (DMSO-d6, 500 MHz): δ 1.28 (s, 3H, CH3), 1.41 (s, 3H, CH3), 1.71 (m, 1H,
CH2), 1.94 (s, 3H, -NHCOCH3), 2.08 (m, 1H, CH2), 5.07 (m, 1H, CH), 6.89 (d, J = 8.5 Hz,
1H, 8-H), 7.54 (s, 1H, 5-H), 7.58 (d, J = 8.5 Hz, 1H, 7-H), 8.26 (d, J = 8.4 Hz, 1H,
13
NHCOCH3); C NMR (DMSO-d6): δ 22.7 (COCH3), 24.3 (CH3), 29.0 (CH3), 38.1 (C-3),
40.9 (C-4), 77.1 (C-2), 102.1 (C-6), 118.1 (C-8), 119.1 (CN), 124.6 (C-4a), 132.2-132.5 (C-
5/C-7), 157.2 (C-8a), 169.6 (CO). Anal. calcd for C14H16N2O2: C 68.83, H 6.60, N 11.47,
found: C 68.63, H 7.15, N 11.45.
4.1.6.
R/S-6-tert-butyloxycarbonylamino-3,4-dihydro-2,2-dimethyl-4-formamido-2H-1-
benzopyran (14). A mixture of formic acid (3.0 mL, 79.5 mmol) and acetic anhydride (6.0
mL, 63.5 mmol) was heated at 60 °C for 30 minutes. R/S-4-amino-6-tert-
butylcarbonylamino-3,4-dihydro-2,2-dimethyl-2H-1-benzopyran (11) [24] (1.00 g, 3.59 mmol)
was added and the mixture was stirred 60 minutes at room temperature. After completion of
the reaction, water was added and the resulting precipitate was collected by filtration, washed
1
with water and dried (5.25 g, 88 %): mp: 165-166.5 °C; H NMR (DMSO-d6, 500 MHz): δ
1.22 (s, 3H, CH3), 1.34 (s, 3H, CH3), 1.44 (s, 9H, NHCOOC(CH3)3), 1.67 (m, 1H, CH2), 2.00
(m, 1H, CH2), 5.09 (m, 1H, CH), 6.63 (d, J = 8.8 Hz, 1H, 8-H), 7.13 (d, J = 8.1 Hz, 1H, 7-H),
7.34 (s, 1H, 5-H), 8.20 (s, 1H, CHO), 8.42 (d, J = 8.8 Hz, 1H, NH), 9.08 (bs, 1H,
NHCOOC(CH3)3); 13C NMR (DMSO-d6): δ 24.1 (CH3), 28.1 (C(CH3)3), 29.1 (CH3), 39.3 (C-
3), 40.3 (C-4), 74.7 (C-2), 78.6 (C(CH3)3), 116.6 (C-8), 117.2 (C-5), 119.6 (C-7), 121.8 (C-
4a), 132.1 (C-6), 148.5 (C-8a), 152.9 (COO), 161.0 (COH). Anal. (C17H24N2O4) theoretical:
C, 63.73; H, 7.55; N, 8.74. Found: C, 63.87; H, 7.71; N, 8.90.
10
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
4.1.7.
R/S-6-tert-butyloxycarbonylamino-3,4-dihydro-2,2-dimethyl-4-acetamido-2H-1-
benzopyran (15). R/S-4-amino-6-tert-butyloxycarbonylamino-3,4-dihydro-2,2-dimethyl-2H-
1-benzopyran (11) (0.20 g, 0.72 mmol) was dissolved in a ice cooled solution of sodium
acetate (0.60 g, 7.31 mmol) in glacial acetic acid (5.0 mL, 87.3 mmol), and acetic anhydride
(0.20 mL, 2.12 mmol) was added. The mixture was stirred 60 minutes at room temperature.
After completion of the reaction, water was added and the resulting precipitate was collected
1
by filtration, washed with water and dried (5.25 g, 88 %): mp: 146-149 °C; H NMR
(DMSO-d6, 500 MHz): δ 1.21 (s, 3H, CH3), 1.34 (s, 3H, CH3), 1.44 (s, 9H,
NHCOOC(CH3)3), 1.64 (m, 1H, CH2), 1.90 (s, 3H, COCH3), 1.97 (m, 1H, CH2), 5.01 (m, 1H,
CH), 6.61 (d, J = 8.7 Hz, 1H, 8-H), 7.19 (d, J = 8 Hz, 1H, 7-H), 7.23 (s, 1H, 5-H), 8.20 (d, J =
13
8.6 Hz, 1H, NH), 9.05 (bs, 1H, NHCOOC(CH3)3); C NMR (DMSO-d6): δ 22.7 (COCH3),
24.1 (CH3), 28.1 (C(CH3)3), 29.2 (CH3), 39.4 (C-3), 41.4 (C-4), 74.7 (C-2), 78.6 (C(CH3)3),
116.5 (C-8), 117.5 (C-5), 119.5 (C-7), 122.5 (C-4a), 132.0 (C-6), 148.5 (C-8a), 152.9 (COO),
169.1 (COCH3). Anal. calcd for C18H26N2O4: C 64.65, H 7.84, N 8.38, found: C 64.31, H
7.89, N, 8.63.
4.1.8.
(-)-N-3-Cyanophenyl-N’-(6-ethoxycarbonylamino-3,4-dihydro-2,2-dimethyl-2H-1-
benzopyran-4-yl)urea [(-)-29] and (+)-N-3-Cyanophenyl-N’-(6-ethoxycarbonylamino-3,4-
dihydro-2,2-dimethyl-2H-1-benzopyran-4-yl)urea [(+)-29]. R/S-4-amino-3,4-dihydro-2,2-
[26]
dimethyl-6-ethoxycarbonylamino-2H-1-benzopyran
was dissolved in a mixture of
isopropanol/heptane (1 :1), injected (24 mL/16 minutes) on a Chiralcel° OJ column (50*500
cm) and eluted with a mobile phase of isopropanol/heptane/diethylamine (50 :50 :1). The two
enantiomers (compound A: Rt = 4.2 min; compound B: Rt = 7.6 min) were obtained with an
optical purity of 99.7% (compound A : [a]D20 = +65° ; compound B : [a]D20 = -65°).
(+)-4-Amino-3,4-dihydro-2,2-dimethyl-6-ethoxycarbonylamino-2H-1-benzopyran
(compound A) (0.50 g, 1.89 mmol) was dissolved in methylene chloride (8 mL) and then
supplemented with 3-cyanophenyl isocyanate (0.30 g, 2.08 mmol, 1.1 equiv.). The reaction
mixture was stirred during 30 min, after which the resulting precipitate of the title compound
(-)-29 was collected by filtration, washed with methylene chloride and dried (yields: 85%):
20
1
mp: 167-169 °C; [a]D = -36° ; IR (KBr): υ 3343 (N-H), 2231 (CN), 1673 (C=O) cm-1; H
NMR (DMSO-d6, 500 MHz): δ 1.19 (t, J = 7 Hz, 3H, CH3-CH2), 1.25 (s, 3H, CH3), 1.36 (s,
3H, CH3), 1.74 (t, J = 12 Hz, 1H, CH2), 2.08 (dd, J = 13 Hz/6 Hz, 1H, CH2), 4.06 (q, J = 7
11
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Hz, 2H, CH3-CH2), 4.94 (dd, J = 16 Hz/9.5 Hz, 1H, CH), 6.66 (d, J = 8.8 Hz, 1H, 8-H), 6.69
(d, J = 8.7 Hz, 1H, CH-NH-CO-NH-), 7.22 (d, J = 7.8 Hz, 1H, 7-H), 7.36-7.39 (m, 2H, 5-H
and 4’-H), 7.46 (t, J =8 Hz, 1H, 5’-H), 7.64 (d, J = 8.3 Hz, 1H, 6’-H), 8.01 (s, 1H, 2’-H), 8.86
(s, 1H, CH-NH-CO-NH), 9.34 (s, 1H, NHCOOCH2CH3). Anal. calcd for C22H24N4O4: C
64.69, H 5.92, N 13.72, found: C 64.59, H 5.99, N 13.76.
(-)-4-Amino-3,4-dihydro-2,2-dimethyl-6-ethoxycarbonylamino-2H-1-benzopyran (compound
B) (0.50 g, 1.89 mmol) was dissolved in methylene chloride (8 mL) and then supplemented
with 3-cyanophenyl isocyanate (0.30 g, 2.08 mmol, 1.1 equiv.). The reaction mixture was
stirred during 30 min, after which the resulting precipitate of the title compound (+)-29 was
collected by filtration, washed with methylene chloride and dried (yields: 82%): mp: 167-169
°C; [a]D20 = +36° ; IR (KBr): υ 3341 (N-H), 2231 (CN), 1672 (C=O) cm-1; 1H NMR (DMSO-
d6, 500 MHz): δ 1.19 (t, J = 7 Hz, 3H, CH3-CH2), 1.25 (s, 3H, CH3), 1.36 (s, 3H, CH3), 1.74
(t, J = 12 Hz, 1H, CH2), 2.08 (dd, J = 13 Hz/6 Hz, 1H, CH2), 4.06 (q, J = 7 Hz, 2H, CH3-
CH2), 4.94 (dd, J = 16 Hz/9.5 Hz, 1H, CH), 6.66 (d, J = 8.8 Hz, 1H, 8-H), 6.69 (d, J = 8.7 Hz,
1H, CH-NH-CO-NH-), 7.22 (d, J = 7.8 Hz, 1H, 7-H), 7.36-7.39 (m, 2H, 5-H and 4’-H), 7.46
(t, J =8 Hz, 1H, 5’-H), 7.64 (d, J = 8.3 Hz, 1H, 6’-H), 8.01 (s, 1H, 2’-H), 8.86 (s, 1H, CH-
NH-CO-NH), 9.34 (s, 1H, NHCOOCH2CH3). Anal. calcd for C22H24N4O4: C 64.69, H 5.92,
N 13.72, found: C 64.58, H 6.02, N 13.73.
4.2. Biological assays
(±)-Cromakalim (Tocris, UK), diazoxide (Sigma Chemical Co, USA) and verapamil (Sigma
Chemical Co, USA) were used as reference compounds. All experiments were performed
with aortae or pancreatic islets isolated from adult fed Wistar Rats (Charles River
Laboratories, Belgium). The laboratory animal care was approved by the local ethic's
committee of the Université Libre de Bruxelles.
4.2.1. Measurement of insulin secretion from incubated rat pancreatic islets. Pancreatic islets
were isolated by the collagenase method and freshly isolated islets were used for
measurements of insulin secretion. [28] Groups of 10 islets, each derived from the same batch
of islets, were pre-incubated for 30 min at 37 °C in 1 ml of a physiological salt medium (in
mM: NaCl 115, KCl 5, CaCl2 2.5, MgCl2 1, NaHCO3 24) supplemented with 2.8 mM
12
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
glucose, 0.5% (w/v) albumin (RIA grade) and equilibrated against a mixture of O2 (95 %)
and CO2 (5%). The islets were then incubated at 37 °C for a further 90 min in 1 ml of the
same medium containing 16.7 mM glucose and, in addition, either the reference compound or
the required chroman derivative. The release of insulin was measured radioimmunologically
using rat insulin as a standard. Residual insulin secretion was expressed as a percentage of
the value recorded in control experiments (100%); that is in the absence of drug and presence
of 16.7 mM glucose. [28]
4.2.2. Measurement of myorelaxant activity on rat aorta rings. The rat thoracic aorta was
removed, cut into transverse rings (3-4 mm), and adhering fat and connective tissue was
[28]
detached.
After removal of the endothelium, the segments were suspended under 1.5 g
tension in an organ bath containing 20 ml of a buffered physiological solution (in mM: NaCl
118, KCl 4.7, CaCl2 2.5, NaHCO3 25, KH2PO4 1.2, MgSO4 1.2, glucose 5). The solution was
maintained at 37 °C and continuously oxygenated with a mixture of 95% O2 and 5% CO2.
After equilibration for 60 min, isometric contractions were measured with a force-
displacement transducer. Contractile activity was induced by increasing the extracellular
concentration of K+ (30 or 80 mM KCl). When the tension was stabilized, drugs were added
cumulatively until maximal relaxation or until a maximum concentration of 200 µM. Some
experiments were repeated in the continuous presence of 1 or 10 µM glibenclamide (Sigma
Chemical Co) in the physiological medium. The contractile responses were expressed as the
percentage of the contractile response to KCl (100 %). The EC50 values (concentration
evoking 50% inhibition of the plateau phase induced by KCl) were assessed from
concentration–response curves using Datanalyst software (EMKA Technologies, France). [28]
4.2.3. Statistical evaluation. The statistical significance of differences between mean data
was assessed by using the Student’s t-test or by analysis of variance followed for multiple
comparisons by a Bonferroni test procedure. The biological results were considered as
statistically different when p value was <0.05.
4.2.4. Predicted partition coefficients. The program ALOGPS 2.1 (VCCLAB, Virtual
Computational Chemistry Laboratory, http://www.vcclab.org, 2005) was used for the
13
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
calculation of AC log P values. [29] Such a program is available at the Virtual Computational
Chemistry Laboratory and used algorithms have been previously described in the literature.
[19]
Author information : Corresponding Author
* Bernard Pirotte
Phone: + 32 4 366 43 65. Fax : + 32 4 366 43 62. E-mail : b.pirotte@ulg.ac.be
Acknowledgements
This study was supported in part by grants from the “National Fund for Scientific Research”
(F.N.R.S., Belgium) from which P. Lebrun is a Research Director. The authors gratefully
acknowledge the technical assistance of S. Counerotte, F. Leleux, and A.-M. Vanbellinghen.
The preparative chiral chromatography was performed at UCB Pharma (Braine l’Alleud,
Belgium) by V. Pinilla and D. Sénéchal, and the circular dichroism experiment was realized
at the Center for Protein Engineering (CIP, University of Liege, Belgium) by A. Matagne;
these collaborators are gratefully acknowledged.
Abbreviations used
ATP: adenosine triphosphate; KATP channel: ATP-sensitive potassium channel; PCO:
potassium channel opener; TLC: thin layer chromatography; TMS: tetramethylsilane.
References
[1] Y. Zheng, B. Zhao, C. Lu, X. Lin, Z. Zheng, W. Su, Nat. Prod. Commun. 2009, 4, 501-
506.
[2] S. Li, H.-Y. Tian, W.-C. Ye, R.-W. Jiang, Biochem. Syst. Ecol. 2011, 39, 64-67.
14
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
[3] S.A.T. Fobofou, K. Franke, A. Porzel, W. Brandt, L.A. Wessjohann, J. Nat. Prod. 2016,
79, 743-753.
[4] R.M. Reilly, H.A. McDonald, P.S. Puttfarcken, S.K. Joshi, L. Lewis, M. Pai, P.H.
Franklin, J.A. Segreti, T.R. Neelands, P. Han, J. Chen, P.W. Mantyh, J.R. Ghilardi, T.M.
Turner, E.A. Voight, J.F. Daanen, R.G. Schmidt, A. Gomtsyan, M.E. Kort, C.R. Faltynek,
P.R. Kym, J. Pharmacol. Exp. Ther. 2012, 342, 416-428.
[5] E.A. Voight, A.R. Gomtsyan, J.F. Daanen, R.J. Perner, R.G. Schmidt, E.K. Bayburt, S.
DiDomenico, H.A. McDonald, P.S. Puttfarcken, J. Chen, T.R. Neelands, B.R. Bianchi, P.
Han, R.M. Reilly, P.H. Franklin, J.A. Segreti, R.A. Nelson, Z. Su, A.J. King, J .S.
Polakowski, S.J. Baker, D.M. Gauvin, L.R. Lewis, J.P. Mikusa, S.K. Joshi, C.R. Faltynek,
P.R. Kym, M.E. Kort, J. Med. Chem. 2014, 57, 7412-7424.
[6] K.W. Hunt, A.W. Cook, R.J. Watts, C.T. Clark, G. Vigers, D. Smith, A.T. Metcalf, I.W.
Gunawardana, M. Burkard, A.A. Cox, M.K. Geck Do, D. Dutcher, A.A. Thomas, S. Rana,
N.C. Kallan, R.K. DeLisle, J.P. Rizzi, K. Regal, D. Sammond, R. Groneberg, M. Siu, H.
Purkey, J.P. Lyssikatos, A. Marlow, X. Liu, T.P. Tang, J. Med. Chem. 2013, 56, 3379-3403.
[7] A. Dhawan, S. Balwani, A.K. Prasad , B. Ghosh, V.S. Parmar, J. Heterocycl. Chem. 2014,
51, 1712-1719.
[8] S. Sebille, P. De Tullio, S. Boverie, M.-H. Antoine, P. Lebrun, B. Pirotte, Curr. Med.
Chem. 2004, 11, 1213-1222.
[9] V. Cecchetti, O. Tabarrini, S. Sabatini, Curr. Top. Med. Chem. 2006, 6, 1049-1068.
[10] X. Zhang, J. Zhao, S. Kang, M. Yi, S. You, D.S. Shin, D.K. Kim, Int. J. Oncol. 2011,
39, 1609-1617.
[11] D.L. Cook, C.N. Hales, Nature 1984, 311, 271-273.
[12] N. Inagaki, S. Seino, Jpn. J. Physiol. 1998, 48, 397-412.
15
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
[13] A. Tinker, Q. Aziz, A. Thomas, Br. J. Pharmacol. 2014, 171, 12-23.
[14] S. Seino, Annu. Rev. Physiol. 1999, 61, 337-362.
[15] D. Tricarico, A. Mele, A.L. Lundquist, R.R. Desai, A.L. George, D.C. Camerino, Proc.
Natl. Acad. Sci. 2006, 103, 1118-1123.
[16] N.Q. Shi, B. Ye, J.C. Makielski, J. Mol. Cell. Cardiol. 2005, 39, 51-60.
[17] T. Godfraind, Frontiers in Pharmacology 2017, 8, 1-25.
[18] X. Florence, S. Sebille, P. de Tullio, P. Lebrun, B. Pirotte, Bioorg. Med. Chem. 2009, 17,
7723-7731.
[19] S. Khelili, N. Kihal, M. Yekhlef, P. de Tullio, P. Lebrun, B. Pirotte, Eur. J. Med. Chem.
2012, 54, 873-878.
[20] S. Sebille, P. de Tullio, B. Becker, M.-H. Antoine, S. Boverie, B. Pirotte, P. Lebrun, J.
Med. Chem. 2005, 48, 614-621.
[21] S. Khelili, P. Lebrun, P. de Tullio, B. Pirotte, Bioorg. Med. Chem. 2006, 14, 3530-3534.
[22] S ; Sebille, D. Gall, P. de Tullio, X. Florence, P. Lebrun, B. Pirotte, J. Med. Chem. 2006,
49, 4690-4697.
[23] S. Sebille, P. de Tullio, X. Florence, B. Becker, M.-H. Antoine, C. Michaux, J. Wouters,
B. Pirotte, P. Lebrun, Bioorg. Med. Chem. 2008, 16, 5704-5719.
[24] X. Florence, S. Dilly, P. de Tullio, B. Pirotte, P. Lebrun, Bioorg. Med. Chem. 2011, 19,
3919-3928.
[25] X. Florence, V. Desvaux, E. Goffin, P. de Tullio, B. Pirotte, P. Lebrun, Eur. J. Med.
Chem. 2014, 80, 36-46.
16
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
[26] B. Pirotte, X. Florence, E. Goffin, M.B. Medeiros, P. de Tullio, P ; Lebrun, Eur. J. Med.
Chem. 2016, 121, 338-351.
[27] M.Y. Lebedev, M.B. Erman, Tetrahedron Lett. 2002, 43, 1397-1399.
[28] P. Lebrun, B. Becker, N. Morel, P. Ghisdal, M.-H. Antoine, P. de Tullio, B. Pirotte,
Biochem. Pharmacol. 2008, 75, 468-475.
[29] I. Tetko, J. Gasteiger, R. Todeschini, A. Mauri, D. Livingstone, P. Ertl, V. Palyulin, E.
Radchenko, N. Zefirov, A. Makarenko, V. Tanchuk, V. Prokopenko, J. Comput.-Aided Mol.
Des. 2005, 19, 453-463.
17
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Legends for figures
Figure 1. Chemical structures of cromakalim (as the most active enantiomer levcromakalim)
(1) and previously described 2,2-dimethylchromans (2-8) acting as inhibitors of insulin
release and/or smooth muscle relaxants.
Figure 2. Chemical structures of the 4-amino-2,2-dimethylchroman derivatives (9-19) tested
as putative inhibitors of insulin release and/or smooth muscle relaxants. Figure 2A: structural
evolution from 4-amino-2,2-dimethylchroman 9 to compound 8 and 29. Figure 2B: structural
evolution from the 6-amino-substituted arylthiourea 16 to the 6-tertbutoxycarbonylamino-
substituted arylurea 8.
18
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Figures
H
N
CH3
O
N
S
H
H
O
N
N
O
6
NC
OH
Cl
Cl
4
O
O
1
2
3
Cl
H
H
N
H
N
N
Cl
N
Cl
N
H
H
H
H
N
N
S
O
S
H3C
Cl
F
S
O
O
O
O
O
6
5
4
CN
H
N
H
N
N
N
CN
H
H
H
N
S
H
O
O
O
N
O
O
O
O
8
7
Figure 1
19
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
H
N
CN
R'
H
H
H
N
N
O
O
H
N
NH2
O
H
N
N
O
O
NH2
O
O
O
R
R
R
O
O
O
O
9
10: -R = -CH(CH3)2
11: -R = -C(CH3)3
14: -R = C(CH3)3; -R' = -H
15: -R = C(CH3)3; -R' = -CH3
8: -R = -C(CH3)3
29: -R = -CH2CH3
CH3
H
N
O
R
O
12: -R = -Cl
13: -R = -CN
Figure 2A
H
N
H
N
H
N
CN
N
CN
N
CN
H
H
H
S
H
N
S
H
N
N
S
H2N
R
O
8
O
O
O
O
O
17: -R = -H
18: -R = -CH3
16
19
Figure 2B
20
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Schemes
OH
O
O
O
N
O
NH2
i
iii
ii
CH3
OH
O
9
20
21
22
Scheme 1
Synthetic pathway to compound 9
Reagents: (i): acetone, pyrrolidine, methanol; (ii): NH2OH.HCl, K2CO3, ethanol;
(iii): H2, Raney Nickel®, ethanol.
O
O
O
i
iii
ii
Br
Br
Br
Br
O
CH3
OH
OH
O
CH3
23
24
26
25
iv
CH3
O
H
N
O
OH
O
OH
O
v
vi
NC
NC
Br
13
28
27
Scheme 2
Synthetic pathway to compound 13
Reagents: (i): (CH3CO)2O, H2SO4; (ii): AlCl3; (iii): acetone, pyrrolidine, methanol; (iv):
NaBH4, CH3OH; (v): CuCN, 1-methyl-2-pyrrolidinone; (vi): CH3CN, H2SO4.
21
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
R
H
i or ii
H
N
NH2
O
H
N
N
O
O
O
O
O
O
14 : R = H
15 : R = CH3
11
Scheme 3
Synthetic pathway to compounds 14 and 15
Reagents: (i): HCOOH, Ac2O; (ii): Ac2O, HOAc.
22
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Tables
Table 1. Calculated logP values and effects of selected dimethylchromans, (±)-cromakalim
and diazoxide on the glucose-induced insulin release from rat pancreatic islets and on the
contractile activity of K+ depolarized rat aorta rings.
H
CH3
O
R
N
CN
H
H
H
NH2
O
H
N
NH2
O
N
O
H
N
N
O
O
N
O
X
O
R
O
R
R
O
O
9
10,11
12,13
14,15
16-19, 8, 29
Residual insulin secretion (%) b
Myorelaxant
activity
Average
logPa
Cpd.
9
-R
X
EC50 (µM) c
50 µM
10 µM
1 µM
-
-
1.61
99.7 ± 4.9 (23)
97.1 ± 5.2 (30)
180.0 ± 14.0 (5)
10
11
-CH(CH3)2
-C(CH3)3
-
-
1.87
2.16
75. 4 ± 4.5 (21)
57.7 ± 2.5 (24)
93.6 ± 4.1 (32)
73.3 ± 3.0 (30)
12.1 ± 2.7 (7)
5.8 ± 1.1 (5)
12
13
-Cl
-
-
2.53
1.63
100.8 ± 5.6 (15)d
85.6 ± 5.2 (22)
6.6 ± 0.5 (10)d
2.1 ± 0.1 (5)
-CN
14
15
-H
-
-
2.35
2.52
68.4 ± 3.1 (22)
65.7 ± 3.1 (24)
86.8 ± 3.5 (32)
90.1 ± 3.6 (32)
11.5 ± 1.7 (6)
7.8 ± 0.8 (5)
-CH3
16d
17d
18d
19d
8d
-NH2
S
S
S
S
O
2.69
2.61
2.76
3.99
3.61
96.9 ± 3.4 (23)d
71.3 ± 3.3 (31)d
60.9 ± 3.3 (21)d
13.3 ± 1.8 (19)d
7.4 ± 1.0 (25)d
>30.0 (4)d
>30.0 (4)d
11.9 ± 1.4 (4)d
8.5 ± 1.0 (7)d
10.5 ± 1.8 (4)d
-NHCOH
-NHCOCH3
-NHCOOC(CH3)3
-NHCOOC(CH3)3
72.2 ± 4.3 (24)d
15.6 ± 1.2 (20)d
29d
-NHCOOCH2CH3
-NHCOOCH2CH3
-NHCOOCH2CH3
O
O
O
2.99
2.99
2.99
5.9 ± 0.8 (23)d
66.5 ± 2.7 (24)
10.9 ± 1.0 (24)
81.0 ± 4.0 (23)d
86.1 ± 3.3 (15)
53.0 ± 3.2 (16)
14.2 ± 1.8 (5)d
17.7 ± 1.0 (6)
8.0 ± 1.9 (4)
(-)-29
(+)-29
1.82
n.d.
87.1 ± 3.6 (22)
30.4 ± 2.2 (24)
96.4 ± 6.5 (16)
80.8 ± 3.7 (32)d
0.13 ± 0.01 (7)d
26.1 ± 2.9 (4)d
(±)-cromakalim
diazoxide
a Calculated logP value according to ref. 22. b Percentage of residual insulin release from rat pancreatic islets
c
incubated in the presence of 16.7 mM glucose (mean ± SEM (n)). EC50: drug concentration giving 50%
d
relaxation of the 30 mM KCl-induced contraction of rat aorta rings (mean ± SEM (n)). Published compounds
and results (ref 17-19).
23
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Table 2. Myorelaxant effects of 11, 15, (±)-cromakalim, diazoxide and verapamil on 30 mM
or 80 mM KCl-precontracted rat aorta rings incubated in the absence or presence of
glibenclamide.
Myorelaxant activity
EC50 (µM)a
Compounds
KCl 30 mM
KCl 30 mM
KCl 30 mM
+
KCl 80 mM
+
1 µM Gliben.b
10 µM Gliben.b
6.8 ± 1.4 (4)
8.2 ± 1.9 (9)
6.9 ± 1.5 (6)
8.8 ± 1.1 (5)
5.7 ± 0.5 (6)
7.9 ± 0.7 (6)
11
15
6.6 ± 1.2 (11)
13.3 ± 0.9 (10)
9.6 ± 1.7 (4)
11.3 ± 2.1 (10)
0.22 ± 0.07 (6)c
0.17 ± 0.01 (4)c
2.5 ± 0.7 (9)c
153.7 ± 8.0 (7)c
0.038 ± 0.007 (6)c
28.8 ± 6.0 (5)c
(±)-Cromakalim
Diazoxide
137.7 ± 10.4 (10)c
34.8 ± 2.7 (6)c
23.8 ± 5.0 (6)c
279.8 ± 11.8 (4)c
> 300 (8)c
0.031 ± 0.004 (4)c
0.035 ± 0.003 (5)c
a
b
c
EC50: drug concentration giving 50% relaxation (mean ± SEM (n)). Gliben.: glibenclamide. Published
results (Ref 8).
24
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700409
ChemMedChem
Graphical abstract
H
N
CN
CH3
O
H
H
NH2
O
H
N
NH2
O
H
N
N
O
O
H
N
N
O
O
O
O
O
O
O
9
11
8
15
Inhibitory activity on insulin release: 9 < 11 ≈ 15 <<< 8
Myorelaxant activity on vascular smooth muscle: 9 << 11 ≈ 15 ≈ 8
25
This article is protected by copyright. All rights reserved.