Configurational stability of bisindolylmaleimide cyclophanes: From conformers to the first configurationally stable, atropisomeric bisindolylmaleimides

Article abstract of DOI:10.1002/chem.200500520

The bisindolylmaleimides are selective protein kinase inhibitors that can adopt two limiting diastereomeric (syn and anti) conformations. The configurational stability of a range of substituted and macrocyclic bisindolylmaleimides was investigated by using appropriate techniques. With unconstrained bisindolylmaleimides, the size of the 2-indolyl substituents was found to affect configurational stability, though not sufficiently to allow atropisomeric bisindolylmaleimides to be obtained. However, with a tether between the two indole nitrogen atoms in place, the steric effect of 2-indolyl substituents was greatly exaggerated, leading to large differences in configurational sta bility. The rate of interconversion of the syn and anti conformers varied by over twenty orders of magnitude through substitution of a bisindolylmaleimide ring system, which was constrained within a macrocyclic ring. Indeed, the first examples of configurationally stable atropisomeric bisindolylmaleimides are reported; the half-life for epimerisation of these compounds at room temperature was estimated to be > 10⁷ years.

Full text of DOI:10.1002/chem.200500520

FULL PAPER
DOI: 10.1002/chem.200500520
Configurational Stability of Bisindolylmaleimide Cyclophanes: From
Conformers to the First Configurationally Stable, Atropisomeric
Bisindolylmaleimides
Simon Barrett,[a] Stephen Bartlett,[a, b] Amanda Bolt,[a, b] Alan Ironmonger,[a, b]
Catherine Joce,[a, b] Adam Nelson,*[a, b] and Thomas Woodhall[a, b]
Abstract: The bisindolylmaleimides are
selective protein kinase inhibitors that
can adopt two limiting diastereomeric
(syn and anti) conformations.The con-
figurational stability of a range of sub-
stituted and macrocyclic bisindolylma-
leimides was investigated by using ap-
propriate techniques.With unconstrain-
ed bisindolylmaleimides, the size of the
2-indolyl substituents was found to
affect configurational stability, though
not sufficiently to allow atropisomeric
bisindolylmaleimides to be obtained.
However, with a tether between the
two indole nitrogen atoms in place, the
steric effect of 2-indolyl substituents
was greatly exaggerated, leading to
large differences in configurational sta-
bility.The rate of interconversion of
the syn and anti conformers varied by
over twenty orders of magnitude
through substitution of a bisindolylma-
leimide ring system, which was con-
strained within
a macrocyclic ring.
Indeed, the first examples of configura-
tionally stable atropisomeric bisindolyl-
maleimides are reported; the half-life
for epimerisation of these compounds
at room temperature was estimated to
be >107 years.
Keywords: atropisomerism · config-
urational stability · conformation
analysis
kinases
· macrocycles · protein
Introduction
of some inhibitors of therapeutically important kinases has
been refined by the formation of macrocyclic analogues of
4; for example, the bisindolylmaleimide LY333531 (6) selec-
tively inhibits[5] the b isoforms of protein kinase C (PKCb)
(IC50 =4.7 nm for PKCbI and 5.9 nm for PKCbII), and the
macrocyclic analogues 7 target glycogen synthase kinase-3b
(GSK-3b) (with up to IC50 =22 nm).[6] PKCb is selectively
activated by elevated glucose in many vascular tissues, and
the bisindolylmaleimide 6 can produce significant improve-
ments in diabetic retinopathy, nephropathy, neuropathy and
cardiac dysfunction.[7] Indeed, the bisindolylmaleimide 6 is
undergoing phase III clinical trials as a therapeutic agent for
preventing diabetes complications (such as diabetic retinop-
athy) and left ventricular hypertrophy in heart failure.[8] The
mechanism of action of bisindolylmaleimides has been re-
vealed recently in molecular detail: structures of the com-
plexes of 10 with protein kinase A (PKA)[9] and those of 6,
8, 9a, 9b and 10 with 3-phosphoinositide-dependent protein
kinase-1 (PDK-1)[10] have been determined.
The indolocarbazole alkaloids, such as staurosporine (1),
K252a (2) and rebeccamycin (3), are potent (typically sub-
10 nm) inhibitors of a broad range of protein kinases.[1] X-
ray crystal structures of protein kinase–staurosporine com-
plexes reveal that, in each case, staurosporine is bound in an
ATP-binding pocket of the kinase, and that its lactam
mimics the adenine ring of ATP.[2] Despite their widespread
activity, the indolocarbazoles have been useful tools in the
discovery of selective kinase inhibitors.A fruitful strategy
has been to disrupt the planarity of the indolocarbazole ring
system to yield either bisindolylmaleimides[3] (e.g., 4) or di-
anilinophthalimides[4] (e.g., 5).The selectivity and potency
[a] S.Barrett, S.Bartlett, A.Bolt, A.Ironmonger, C.Joce,
Prof.A.Nelson, T.Woodhall
School of Chemistry, University of Leeds
Leeds LS2 9JT (UK)
E-mail: a.s.nelson@leeds.ac.uk
Bisindolylmaleimides 11 are not planar molecules, and a
30–408 angle between the planes of the maleimide and each
indole ring is typical.[11] Consequently, for simple bisindolyl-
maleimides 11 bearing achiral substituents R, R’ and R’’,
two diastereomeric (syn and anti) conformers are possible.
[b] S.Bartlett, A.Bolt, A.Ironmonger, C.Joce, Prof.A.Nelson,
T.Woodhall
Astbury Centre for Structural Molecular Biology
University of Leeds, Leeds LS2 9JT (UK)
Fax : (+44)113-343-6565
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6277

and (b) as the size of the macrocycle decreases.[14] In this
paper, we present our investigation of the configurational
stability of simple (11) and macrocyclic (12) bisindolylmale-
imides, and we conclude that atropisomeric bisindolylmale-
imides 12 may be prepared, provided that the 2-indolyl sub-
stituents, R, are sufficiently large.
Results
Synthesis of simple bisindolylmaleimides: The preparation
of simple bisindolylmaleimides with R=H and Me has been
described previously.[15] The bisindolylmaleimides 16Ph and
16Bn (R=Ph and Bn) were prepared in an analogous
manner (Scheme 1).Hence, base-catalysed cyclisation of the
2-alkynyl anilines 14, prepared by substitution of 2-iodo ani-
line, gave the indoles 15.[16] Treatment of the indoles 15 with
ethyl magnesium bromide, and reaction with N-benzyl 3,4-
dichloromaleimide, gave the bisindolylmaleimides 16.[15]
Rotation about either of the
indolylmaleimide bonds results
in interconversion between the
conformers.We have shown
that simple bisindolylmale-
imides 11 with R=H and Me
are not, in general, atropiso-
meric at ambient temperatures
(the half-lives for epimerisa-
Scheme 1.Preparation of simple bisindolylmaleimides.
tion are much less than an ar-
bitrary 1000 s[12]).[13]
Macrocyclic bisindolylmaleimides 12 are structurally relat-
ed to [2.n]metacyclophanes (such as 13), which may also
populate two diastereomeric conformations.The configura-
tional stability of cyclophanes is rather sensitive to substitu-
tion and the size of the macrocyclic ring: the transition state
for isomerisation of a cyclophane is destabilised by steric ef-
fects, and configurational stability increases (a) as the size of
the substituent(s) passed through the larger ring increases
Variable temperature (VT) NMR experiments on simple
bisindolylmaleimides: We investigated the interconversion
of the simple bisindolylmaleimides 16Ph, 16Bn and 17Me
1
by using variable temperature 500 MHz H NMR spectros-
copy.In each case, the choice of solvent was dictated by the
temperature range for which intermediate exchange was ob-
served, and also by the solubility of each compound.The
syn and anti conformers of these bisindolylmaleimides inter-
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Stability of Bisindolylmaleimide Cyclophanes
FULL PAPER
Table 1.Populations of the syn and anti conformers of the bisindolylmaleimides.
convert slowly enough at tem-
peratures below approximately
273 K for them to give rise to
two sets of discrete signals in
their 500 MHz 1H NMR spec-
tra.The signals corresponding
to the syn and anti conformers
were assigned by careful in-
spection of the region of the
spectrum corresponding to the
benzyl substituent on the
imide: the benzylic protons are
enantiotopic in the syn con-
former (HA and HA’) and give
rise to a singlet, and are dia-
stereotopic in the anti con-
Compound
R
Solvent,
T [K]
syn
anti
DH8
DS8
DG8
[b]
[b]
[b]
[%][a]
[%][a]
[kJmolꢀ1
]
[Jmolꢀ1 Kꢀ1
]
[kJmolꢀ1
]
16Ph
Ph
CD2Cl2,
243
CD2Cl2,
253
[D8]toluene,
298
[D6]DMSO,
300
41
39
65
57
35
59
61
35
43
65
ꢀ7.9 ꢁ1.0
2.7ꢁ0.5
ꢀ29ꢁ4
14ꢁ2
0. ꢁ0.2
1. ꢁ0.3
8
6
16Bn
Bn
Me
Me
Me
[c]
[c]
17Me
–
–
1.4ꢁ0.2[d]
0.8ꢁ0.1
19Me (n=9)
19Me (n=10)
5.0ꢁ1.5
4.4ꢁ0.5
1
14.2ꢁ3.0
20ꢁ2
[D6]DMSO,
300
ꢀ1.5ꢁ0.1
[a] At temperature T, determined by integration of the 500 MHz H NMR spectrum.[b] At 298 K, determined
by extrapolation of the relative populations of the slowly exchanging conformers.[c] Not determined.[d] De-
1
termined by integration of the 500 MHz H NMR spectrum recorded at 298 K.
former (HB and HC) and give rise to a pair of doublets (see
Figure 1).The relative populations of the syn and anti con-
formers were determined as a func-
tion of temperature by integration of
1
the 500 MHz H NMR spectrum over
a range of 40 K in the slow exchange
regime (see Table 1).
The barrier to interconversion be-
tween the syn and anti conformers
was determined by analysis of
500 MHz 1H NMR spectra recorded
over a temperature range (of between 20 and 40 K) in the
intermediate exchange regime (see Table 2).[17] The rate of
equilibration, and hence krot,
Figure 1.Enantiotopic and diastereotopic benzylic protons in the bisindo-
lylmaleimides 11.
was estimated by comparison
Table 2.Results of VT-NMR studies on the bisindolylmaleimides.
of the experimental spectra
with the simulated spectra gen-
erated by gNMR analysis[18]
using populations extrapolated
from the slow exchange
regime.Kinetic data for the
isomerisation of the anti con-
formers of 16Ph, 16Bn and
17Me is summarised in
Table 3.
Compound
R
Solvent
T range[a]
[K]
K[b]
krot
]
DG°
[b]
[c]
[sꢀ1
[kJmolꢀ1
]
16Ph
16Bn
17Me
19Me (n=9)
19Me (n=10)
Ph
CD2Cl2
CD2Cl2
[D8]toluene
[D6]DMSO
[D6]DMSO
253–283
253–291
263–343
323–373
300–373
0.71ꢁ0.10
1.90ꢁ0.05
0.57ꢁ0.05
0.72ꢁ0.05
1.8ꢁ0.2
170ꢁ30
120ꢁ30
60.3ꢁ0.5
61.1ꢁ0.3
54.8ꢁ0.3
74.3ꢁ0.3
70.3ꢁ0.3
Bn
Me
Me
Me
1600ꢁ200
0.58ꢁ0.1
2.0ꢁ0.3
[a] Temperature range of VT-NMR experiments.[b] At 298 K.[c] For conversion of the anti conformer into
the syn conformer, extrapolated to 298 K.
Table 3.Kinetic data for the epimerisation of the bisindolylmaleimides.
Synthesis of macrocyclic bisin-
dolylmaleimides: The synthe-
ses of the macrocyclic bisindo-
lylmaleimides 18H, 18Me,
19H and 19Me (n=6–10) have
been described previously.[15]
[a]
[b]
[c]
1
Compound
R
Solvent
Method
krot [sꢀ1
]
DG° [kJmolꢀ1
]
t =
2
16Ph
16Bn
17Me
18H (n=6)
18Me (n=6)
18Me (n=8)
19Me (n=9)
19Me (n=10)
18Ph (n=10)
18Bn (n=10)
Ph
Bn
Me
H
H
H
Me
Me
Ph
Bn
CD2Cl2
CD2Cl2
[D8]toluene
CD2Cl2
hexane-iPrOH
hexane-iPrOH
[D6]DMSO
[D6]DMSO
[D6]DMSO
[D6]DMSO
VT-NMR
VT-NMR
VT-NMR
VT-NMR
chiral HPLC
chiral HPLC
VT-NMR
VT-NMR
NMR[g]
170ꢁ30
60.3ꢁ0.5
61.1ꢁ0.3
54.8ꢁ0.3
36.6ꢁ0.3[d]
>90
4.1ꢁ0.3 ms
5.8ꢁ0.5 ms
0.43ꢁ0.05 ms
0.3 ms[e]
120ꢁ30
1600ꢁ200
1.1”106[e]
<1”10ꢀ3
<1”10ꢀ3
0.58ꢁ0.1
2.9ꢁ0.3
>10 min[f]
>10 min[f]
1.2ꢁ0.1 s
0.23ꢁ0.03 s
>107 yr[e]
The
bisindolylmaleimides
>90
16Ph and 16Bn were treated
with sodium hydride in DMF,
and the resulting anions were
reacted with 1,10-dibromode-
cane to give the corresponding
74.3ꢁ0.3
70.3ꢁ0.3
>160
<1”10ꢀ17[e]
<1”10ꢀ17[e]
NMR[g]
>160
>107 yr[e]
[a] At 298 K.[b] For the conversion of the anti conformer into the syn conformer at 298 K.[c] Estimated half-
life for the epimerisation of the anti conformer at 298 K.[d] DG° is given at the coalescence temperature
(203 K).[e] Estimated value, on the assumption that DS° is small.[f] The enantiomeric anti conformers could
be separated by chiral analytical HPLC.[g] Epimerisation did not occur when the syn and anti atropisomers
were heated at 433 K for five days.
macrocyclic
bisindolylmale-
imides 18Ph (n=10) and 18Bn
(n=10) (Scheme 2).The mac-
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6279

A.Nelson et al.
ies had revealed that the anti conformer of 18H (n=6) was
>20 kJmolꢀ1 more stable that the syn conformer, the dia-
stereotopicity must stem from slow interconversion of the
enantiomeric anti conformers.This hypothesis is summarised
in Scheme 3.At the coalescence temperature, the barrier to
racemisation, DG°, of the anti conformer of 18H (n=6) was
found to be 36.6ꢁ0.3 kJmolꢀ1 (see Table 3).The NMR
spectra of the larger macrocycles 18H (n=7–10) were also
recorded at 200 K; however, for these compounds, the dia-
stereotopicity of the protons in the tether was not revealed,
suggesting that conformational
interconversion in these cases
was faster.
The barrier to conformation-
al interconversion increased as
the size of the 2-indolyl sub-
stituent increased.The syn and
anti conformers of the macro-
cycles 19Me (n=9 and 10) in-
terconvert slowly enough in
[D6]DMSO at around, and just
above, room temperature to
give rise to two sets of discrete
Scheme 2.Synthesis of atropisomeric, macrocyclic bisindolylmaleimides.
signals in their 500 MHz
rocyclic bisindolylmaleimides were separable atropisomers
whose relative configuration could be assigned by careful
analysis of their 500 MHz 1H NMR spectra: the benzylic
protons give rise to a singlet in the syn atropisomer, and to
a pair of doublets in the anti atropisomer (see Figures 1 and
2).In addition, the relative configuration of the bisindolyl-
maleimide syn-18Ph was determined by X-ray crystallogra-
phy (Figure 3).
Figure 2.Partial 500 MHz 1H NMR spectra of the bisindolylmaleimides
syn- and anti-18Bn.The signal(s) corresponding to the benzylic protons
on the imide nitrogen are at approximately 4.8 ppm in each case.
Figure 3.X-ray crystal structure of the bisindolylmaleimide syn-18Ph.
Investigation into the configurational stability of macrocyclic
bisindolylmaleimides: As a starting point, we investigated
the configurational stability of macrocyclic bisindolylmale-
imides 18H.The syn and anti conformations of the macrocy-
cles 18H were in fast exchange on the NMR timescale at
298 K.However, at low temperature, the region correspond-
ing to the tether of 18H (n=6) in its 500 MHz 1H NMR
spectrum broadened dramatically: below the coalescence
temperature, 203 K, the methylene protons adjacent to
indole nitrogens (NCHAHB) were rendered diastereotopic
on the NMR timescale.Because molecular modelling stud-
Scheme 3.Exchange of H A and HB by racemisation of the anti conformer
of 18H.
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Stability of Bisindolylmaleimide Cyclophanes
FULL PAPER
1H NMR spectra (Figure 4).The syn and anti conformers
were assigned, and the relative populations of the conform-
ers were determined as a function of temperature in the
slow exchange regime (see Table 1).Analysis of the NMR
spectra that had been recorded over a range of higher tem-
peratures, and were, therefore, dramatically broadened, al-
lowed krot and DG° to be extracted (Tables 2 and 3).The
barriers, DG°, to isomerisation of the anti conformers of
19Me (n=10) and 19Me (n=9) were 70.3ꢁ0.3 and 74.3ꢁ
0.3 kJmolꢀ1, respectively (see Table 3).
py.Indeed, analysis of the macrocycles 18Me (n=6) and
18Me (n=8) by performing chiral analytical HPLC revealed
two peaks in each case, demonstrating that the half-lives of
the enantiomeric anti conformers at 298 K were greater
than the separation of the peaks (>10 min).[19] The barrier
to racemisation of the anti conformer of 18Me (n=6 and 8)
was concluded to be at least 90 kJmolꢀ1.
The arbitrary boundary[12] between conformational iso-
mers and atropisomers was crossed by further increasing the
size of the 2-indolyl substituents.Although we had been
able to separate the syn and anti atropisomers of 18Ph (n=
10) and 18Bn (n=10) by preparative HPLC, the effect of
the larger 2-indoyl groups was remarkable.The anti and syn
atropisomers of 18Ph (n=10) and 18Bn (n=10) were
heated at 433 K for five days in [D6]DMSO, and the samples
1
were reanalysed by performing 500 MHz H NMR spectros-
[20]
copy.There was no evidence for epimerisation
of either
of the atropisomers, indicating half-lives of at least one
month at 433 K, and, hence, estimated half-lives of at least
107 years at 298 K.A lower limit for the barrier, DG°, to
conformational isomerisation of 18Ph (n=10) and 18Bn
(n=10) is 160 kJmolꢀ1.
Evidence for “rocking” between unsymmetrical syn and
anti conformers: Perfectly symmetrical conformers of the
bisindoylmaleimides described in this paper would have
identical indole rings: a C2-symmetrical anti conformer has
homotopic indoles, and a Cs-symmetrical syn conformer has
enantiotopic indoles.The 500 MHz 1H NMR spectra ob-
tained for the bisindolylmaleimides 16Bn and 19Me (n=
10) that had been cooled to around 250 K had two sets of
discrete signals that corresponded to the slowly intercon-
verting syn and anti conformers.For each conformer, there
was only one set of resonances for each pair of indole rings.
The conformers were either symmetrical, or were still inter-
converting rapidly on the NMR timescale.
Cooling the samples further resulted in the dramatic
broadening of some signals: all of the indolyl signals of
16Bn, and the indolyl signals corresponding to the syn con-
former of 19Me (n=10), were broadened.We conclude that
this line broadening demonstrates that some of the conform-
ers are, in fact, unsymmetrical; interconversion between
these unsymmetrical conformers had simply been fast
enough at higher temperatures (>ca.250 K) for the indole
rings to appear to be identical.
We have previously shown that the syn conformers of bis-
indolylmaleimides are generally unsymmetrical, a phenom-
enon that is also apparent in the X-ray crystal structure of
syn-18Ph (n=10) (Figure 3).The “rocking” between unsym-
metrical syn conformers is illustrated in Figure 5; this pro-
cess does not require the 2-indolyl substituent (R=Ph) to
eclipse the maleimide C=C bond, and so the associated acti-
vation barrier is rather low.The activation barrier could not
be estimated because the slow exchange regime was inacces-
sible, and the separation of the peaks, dn, could not, there-
fore, be determined.[17,18] The effect of the “rocking” of the
syn conformer of 18Me (n=10) impacts dramatically on the
Figure 4.Variable temperature 500 MHz 1H NMR spectra of the bisindo-
lylmaleimide 19Me (n=10) recorded in [D6]DMSO.The rate of equili-
bration at each temperature, estimated by comparison of experimental
and simulated spectra, is indicated.
The further reduction in the size of the macrocyclic ring
had a remarkable effect on the barrier to conformational in-
terconversion.The 500 MHz 1H NMR spectra of 18Me (n=
6–8) in [D8]toluene revealed that only the chiral, anti con-
former was populated at 298 K.In this case, the barrier to
anti!syn isomerisation, DG°, may be assessed indirectly by
tracking the racemisation of the anti conformer (for which
the syn conformer is a presumed intermediate).The benzylic
protons were diastereotopic on the NMR timescale, and
were used as a handle to assess the rate of racemisation.
The signals corresponding to the benzylic protons did not
coalesce as the sample was heated to 373 K.Interconversion
between the enantiomeric anti conformers of 18Me (n=6–
8) was, therefore, sufficiently slow that its rate could not be
determined by using variable temperature NMR spectrosco-
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A.Nelson et al.
two inter-ring bonds (i.e., direct isomerisation between the
enantiomeric anti conformers).The conformation of bisin-
dolylmaleimides may be described in terms of the dihedral
angles, q and f, which are defined in Figure 6.A description
of the racemisation process in terms of the “traverse” of a
Ramachandran-like plot is illustrated in Figure 7.
Figure 5.“Rocking” process between unsymmetrical syn conformers.
Figure 6.Definitions of the dihedral angles q and f.
1
indole region of its 500 MHz H NMR spectrum.On cooling
from 243 K to 203 K, the indole rings in the syn conformer
become inequivalent as the “rocking” process becomes com-
parable with the NMR timescale, and its NMR resonances
broaden accordingly.
Discussion
The proposed mechanism for the racemisation of a symmet-
rical anti bisindolylmaleimide involves the stepwise rotation
about the two indole–maleimide bonds (Scheme 4).Rota-
tion about one of the inter-ring bonds of an anti bisindolyl-
maleimide gives an unsymmetrical syn bisindolylmaleimide
that is able to equilibrate easily with the enantiomeric con-
former (ent-syn) (see the “rocking” process described in
Figure 5).The racemisation process is completed by rotation
about the other inter-ring bond of the ent-syn conformation
to give the enantiomeric anti bisindolylmaleimide conformer
(ent-anti).The variable temperature NMR studies did not
produce any evidence for correlated[21] rotation about the
Figure 7.Traversing the Ramachandran plot during the racemisation of a
symmetrical, anti bisindolylmaleimide.
The anti!syn isomerisation of simple bisindolylmale-
imides is significantly easier than rotation around the inter-
aryl bond of a similarly substituted biphenyl.
The difficulty in racemisation of tetrasubstitut-
ed biphenyls 20 (with A¼6 B¼6 H and C¼6 D¼6
H) is generally sufficient to prevent resolu-
tion.[22] There is a substituent in each of the
ortho positions flanking the [3,3’]bipyrrolyl
bonds of 16Ph, 16Bn and 17Me (see Figure 8,
which highlights the bipyrrolyl unit), and yet,
their syn and anti conformers are far from
being atropisomeric.
The relatively low activation barrier may stem partly from
the conjugation between a maleimide ring and an indole
ring in the transition state, which may be more stabilising
than the conjugation between two phenyl rings.However,
the sizes of the substituents and the geometry of the [3,3’]bi-
pyrrolyl ring system may also be important.The small size
of the maleimide oxo group is rivalled only by a hydrogen
atom (compare the length of the carbonyl moiety, 1.21 Š,
with the lengths of other bonds to “small”[22] substituents:
ꢀ
ꢀ
Scheme 4.A mechanism for racemisation of anti bisindolylmaleimides.
1.39 Š for Ph F, 1.45 Š for Ph OH).Nevertheless, increas-
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Chem. Eur. J. 2005, 11, 6277 – 6285

Stability of Bisindolylmaleimide Cyclophanes
FULL PAPER
configurationally stable atropisomers with half-lives of
greater than 107 years!
Conclusion
Figure 8.The [3,3 ’]-bipyrrolyl unit.
With unconstrained bisindolylmaleimides, the size of the 2-
indolyl substituents does affect configurational stability, but
not sufficiently to allow atropisomeric bisindolylmaleimides
to be obtained.However, with a tether in place, as in 18 and
19, the steric effect of 2-indolyl substituents is greatly exag-
gerated, leading to large differences in configurational sta-
bility (Figure 9).The rate of interconversion of the syn and
anti conformers may be varied by over twenty orders of
magnitude through substitution of a bisindolylmaleimide
ring system, which is constrained within a macrocyclic ring.
Indeed, the macrocycles 18Ph (n=10) and 18Bn (n=10)
are the first examples of configurationally stable atropiso-
meric bisindolylmaleimides.
ing the size of the 2-indolyl R group has a remarkably small
effect on the activation barriers to bond rotation: with
16Bn, 16Ph and 17Me, R varies from Bn to Ph to Me, and
yet, the activation barriers (DG°) vary from only 61.1 to
60.3 to 54.8 kJmolꢀ1.The variable temperature NMR ex-
periments were conducted in CD2Cl2 (for 16Bn and 16Ph)
and in [D8]toluene (for 17Me).However, the rates of rota-
tion about a single bond between sp2 centres, for which no
change in hybridisation in the transition state occurs, gener-
ally vary by less than ten-fold;[23,24] the lower barrier to epi-
merisation of 17Me, though not great, is, therefore, likely to
depend largely on the smaller size of the methyl substitu-
ents.Nevertheless, the impact of the size of these groups on
the activation barrier is relatively small, and may stem from
the internal bond angles of a bisindolylmaleimide[11] (107
and 1088 in the crystal structure of 16H), which are marked-
ly smaller than those of a biphenyl[25] (1198).The carbon
atoms that are ortho to the bipyrrolyl bond are, therefore,
significantly further apart (2.96 and 3.30 Š[11]) than for bi-
phenyl (2.92 Š), which may reduce steric effects in the tran-
sition state.
Tethering the bisindolylmaleimide in a macrocyclic ring—
as in 18 and 19—had a large impact on configurational sta-
bility.The presence of the tether greatly exaggerates steric
effects in the transition state for conformational interconver-
sion.Decreasing the length of the tether from ten to eight
methylene groups had a large effect on configurational sta-
bility: with a indolyl 2-methyl substituent, as in 18Me or
19Me, the conformational half-life varied from 200 ms (with
n=10) to >10 min (with n=8).More dramatically, howev-
er, the presence of a tether greatly exaggerated the effect of
2-indolyl substitution.Changing the 2-indolyl substituents
from hydrogen to methyl to phenyl increased configuration-
al stability by over twenty orders of magnitude: from con-
formers that interconverted on the microsecond timescale to
Experimental Section
Preparative LC-MS was conducted by using a Waters 2525 binary gradi-
ent pump and detection with a Micromass ZQ mass spectrometer; an
XTerraꢁ preparative HPLC column (19”50 mm) was used.Preparative
HPLC was generally conducted by using a Gilson HPLC machine and a
gradient of 90!95% MeCN in H2O over 30 min, detecting at 200 nm on
a Thermohypersil 250”21.2 mm, 8 m, Hyperprepꢁ HS C18 column.The
configurational stability of the bisindolylmaleimides was assessed by per-
forming analytical HPLC by using an Ultron chiral column (150”4.6 mm
ES-OVM).The bisindolylmaleimides 16[15a] were prepared from the cor-
responding commercially available or known[16,26] indoles by the following
general procedure: Ethyl magnesium bromide (300 mL of a 3m solution
in ether, 1.0 mmol) was added dropwise to a solution of the indole 15
(200 mg, 1.0 mmol) in THF (0.9 mL) and toluene (2.5 mL). The solution
was heated to 508C for 1 h and 2,3-dichloro-N-benzylmaleimide (89 mg,
0.4 mmol) in toluene (1.9 mL) was added dropwise. The reaction mixture
was heated to reflux for 54 h, cooled and quenched by addition of satu-
rated aqueous ammonium chloride solution (8 mL) and stirred for
30 min.Water (30 mL) was added and the resulting precipitate was fil-
tered, washed with water (3”10 mL) and dried under vacuum to afford
the required bisindolylmaleimide.
1-Benzyl-3,4-bis-(2-phenyl-1H-indol-3-yl)-pyrrole-2,5-dione (16Ph): The
crude precipitate was washed with water (3”10 mL) to give the bisindo-
lylmaleimide 16Ph (190 mg, 95%) as deep red plates.Mp..137–139 8C;
Rf =0.30 (1:1 diethyl ether/petroleum ether); 1H NMR (500 MHz,
CDCl3): d=8.10 (brs, 2H; NH), 7.40–7.29 (m, 8H; indolyl 4-H, p-Ph, m-
Figure 9.Typical half-lives of the anti conformers of bisindolylmaleimides at 298 K.
Chem. Eur. J. 2005, 11, 6277 – 6285
ꢀ 2005 Wiley-VCH Verlag GmbH & Co.KGaA, Weinheim
www.chemeurj.org
6283

A.Nelson et al.
Bn, o-Bn), 7.18–7.08 (m, 9H; indolyl 5-H, indolyl 7-H, p-Bn, m-Ph),
6.98–6.85, (m, 6H; indolyl 6-H, o-Ph), 4.76 ppm (s, 2H; CH2); 13C NMR
(125 MHz, CDCl3): d=137.8, 132.9, 128.9, 128.4, 128.3, 127.8, 123.1,
120.8, 111.3, 104.3, 42.0 ppm (eight carbon signals missing or overlap-
ped); IR (film): n˜ =3343, 2962, 2924, 2853, 1697 cmꢀ1; MS (ES): m/z (%):
570 (100) [M++Na]; HRMS: m/z calcd for C39H28N3O2 [M++Na]:
570.2182; found: 570.2178.
(CH2)4CH2); 13C NMR (75 MHz, CDCl3): d=142.5, 137.0, 132.1, 130.4,
128.8, 128.5, 122.6, 121.2, 120.5, 111.1, 43.5, 29.4, 27.0, 26.8, 24.7 ppm
(four carbon signals missing or overlapped); IR (film): n˜ =2926, 2854,
1760, 1626 cmꢀ1; MS (ES) m/z (%): 641 (100) [M++Na]; HRMS: m/z
calcd for C42H38N2O3 [M++Na]: 641.2780; found: 641.2808.
23-Benzyl-26,27-dibenzyl-6,7,8,9,10,11,12,13,14,15-decahydro-22H-
5,25:16,21-dimethenodibenzo[b,h]pyrrolo[3,4-e,1,10]diazacycloicosine-
22,24-dione anti- and syn-18Bn: By using the same general method, the
bisindolylmaleimide 16Bn (50 mg, 0.084 mmol) and 1,10-dibromodecane
(19 ml, 0.084 mmol) gave a crude product that was purified by performing
flash chromatography, eluting with 15:85 diethyl ether/petroleum ether,
and preparative LC-MS, to give the macrocycle syn-18Bn (24 mg, 38%)
as a red film. Rf =0.36 (3:7 diethyl ether/petroleum ether); 1H NMR
(300 MHz, CDCl3): d=7.59 (d, J=7.8 Hz, 2H; indolyl 4-H), 7.34–7.23
(m, 11H; aromatic), 7.16 (td, J=7.4, 0.9 Hz, 2H; p-Ph), 7.09–7.05 (m,
4H; aromatic), 6.69–6.65 (m, 4H; aromatic), 4.82 (d, J=14.9 Hz, 1H;
NCHAPh), 4.76 (d, J=14.9 Hz, 1H; NCHBPh), 4.02–3.97 (m, 2H;
NCHA), 3.79–3.73 (m, 2H; NCHB), 3.76 (d, J=16.9 Hz, 2H; indolyl 2-
CHAHB), 3.30 (d, J=16.9 Hz, 2H; indolyl 2-CHAHB), 1.74–0.82 ppm (m,
16H); 13C NMR (75 MHz, CDCl3): d=171.6, 139.7, 138.6, 137.5, 133.1,
129.0, 128.8, 128.6, 127.8, 126.8, 126.2, 122.1, 121.0, 120.8, 110.4, 106.2,
77.6, 42.9, 42.1, 32.6, 28.1, 26.7, 26.6, 24.5 ppm (two carbon signals missing
or overlapped); IR (film): n˜ =2954, 1704, 1396 cmꢀ1; MS (ES) m/z (%):
736.3 (100) [M++H]; HRMS: m/z calcd for C51H49N3O2 [M++H]:
736.3903; found: 736.3911.
1-Benzyl-3,4-bis-(2-benzyl-1H-indol-3-yl)-pyrrole-2,5-dione (16Bn): The
crude product was purified by conducting flash chromatography, eluting
with 1:1 diethyl ether/petroleum ether and then EtOAc, to yield the bis-
indolylmaleimide 16Bn (1.37 g, 32%) as red/orange prisms. M.p. 270–
2728C (from EtOAc); Rf =0.32 (1:1 diethyl ether/petroleum ether);
1H NMR (300 MHz, CDCl3): d=7.80 (s, 2H; NH), 7.55 (d, J=7.2 Hz,
2H; indolyl 4-H), 7.42–6.70 (m, 21H; aromatic), 4.91 (s, 2H; CH2),
3.83 ppm (brs, 4H; 2PhCH2); 13C NMR (75 MHz, CDCl3): d=171.2,
139.2, 137.4, 137.1, 135.5, 131.7, 129.2, 128.9, 128.8, 128.7, 127.7, 126.9,
126.7, 122.2, 120.5, 110.7, 104.5, 42.0, 34.1 ppm (one carbon signal missing
or overlapped); IR (film): n˜ =3375, 3060, 1692, 1455, 1431, 1398,
741 cmꢀ1; MS (ES): m/z (%): 598.0 (100) [M++H]; HRMS: m/z calcd for
C41H31N3O2 [M++H]: 598.2495; found: 598.2509.
23-Benzyl-26,27-diphenyl-6,7,8,9,10,11,12,13,14,15-decahydro-22H-
5,25:16,21-dimethenodibenzo[b,h]pyrrolo[3,4-e,1,10]diazacycloicosine-
22,24-dione anti- and syn-18Ph: Sodium hydride (60% dispersion in
mineral oil, 32 mg, 0.8 mmol) was added portionwise to a solution of the
bisindolylmaleimide (16Ph) (150 mg, 0.3 mmol) in DMF (12 mL) at 08C,
and the mixture was stirred at room temperature for 15 min.1,10-Dibro-
modecane (60 mL, 0.3 mmol) was added, the reaction mixture was stirred
for 6 h, then quenched by the addition of water (100 mL).Ethyl acetate
(150 mL) was added, the organic layers were washed with water (2”
100 mL), dried (MgSO4) and evaporated under reduced pressure.Purifi-
cation was achieved by conducting flash column chromatography (gradi-
ent elution 20:80!35:65 diethyl ether/petroleum ether) to yield the mac-
Also obtained, after further preparative HPLC, was the macrocycle anti-
18Bn (anti, 8 mg, 13% yield) as a red film. Rf =0.42 (3:7 diethyl ether/
petroleum ether); 1H NMR (300 MHz, CDCl3): d=7.40–6.87 (m, 23H;
aromatic), 4.81 (s, 2H; NCH2Ph), 4.14 (d, J=16.7 Hz, 2H; indolyl 2-
CHAHB), 3.57 (d, J=16.7 Hz, 2H; indolyl 2-CHAHB), 3.91–3.86 (m, 2H;
2NCHAHB), 3.69–3.63 (m, 2H; 2NCHAHB), 1.65–0.80 ppm (m, 16H;
alkyl chain); 13C NMR (75 MHz, CDCl3): d=170.9, 137.6, 137.0, 128.5,
128.4, 128.1, 127.8, 127.3, 126.7, 126.3, 122.1, 121.4, 120.5, 109.4, 77.2,
43.7, 30.9, 28.9, 26.8, 24.7, 24.3 ppm (four carbon signals missing or over-
lapped); IR (film): n˜ =2926, 2855, 1701, 1419, 1395, 1347 cmꢀ1; MS (ES)
m/z (%): 736.2 (100) [M++H]; HRMS: m/z calcd for C51H49N3O2 [M+
+H]: 736.3903; found: 736.3922.
rocycle syn-18Ph (45 mg, 24%) as red cubes.Mp..304–306
8C (from
DMSO); Rf =0.2 (1:3 diethyl ether/petroleum ether); 1HNMR
(500 MHz, CDCl3): d=7.37–7.30 (m, 4H; indolyl 4-H, p-Ph), 7.29–7.19
(m, 9H; m-Ph, o-Bn, m-Bn, p-Bn), 7.17–7.10 (m, 4H; indolyl 5-H, indolyl
7-H), 6.92–6.86 (m, 6H; indolyl 6-H, o-Ph), 4.59 (s, 2H; CH2Bn), 4.16
2
2
(dt, J=14.6, 6.9 Hz, 2H; NCHA), 4.08 (dt, J=14.6, 6.9 Hz, 2H; NCHB),
1.43–1.33 (m, 4H; NCH2CH2), 1.26–1.23 (m, 4H; N(CH2)2CH2), 1.06–
0.90 (m, 4H; N(CH2)3CH2), 0.86–0.68 ppm (m, 4H; N(CH2)4CH2);
13C NMR (75 MHz, CDCl3): d=170.7, 141.6, 137.6, 137.4, 137.1, 136.6,
132.7, 130.9, 129.9, 127.8, 127.1, 122.5, 121.9, 120.7, 111.2, 105.8, 43.8,
41.8, 28.9, 27.4, 27.1, 25.1 ppm (two carbon signals missing or overlap-
ped); IR (film): n˜ =2928, 2855, 1704, 1545 cmꢀ1; MS (ES): m/z (%): 708
(100) [M++H]; HRMS: m/z calcd for C49H45N3O2 [M++H]: 708.3590;
found: 708.3599.
Acknowledgements
We thank EPSRC and the University of Leeds for funding, the EPSRC
National Mass Spectrometry Centre, Swansea, for mass spectrometry
measurements, Stuart Warriner, Julie Fisher and Andrew Leach for help-
ful discussions, and Jacqueline Colley for HPLC analysis.
Also obtained was the macrocycle anti-18Ph (10 mg, 5% yield); Rf =0.3
1
(1:3 diethyl ether/petroleum ether); H NMR (500 MHz, CDCl3): d=7.44
(d, J=7.8 Hz, 2H; indolyl 4-H), 7.36 (t, J=7.5 Hz, 2H; p-Ph), 7.35–7.27
(m, 2H; Bn), 7.24–7.22 (m, 3H; Bn), 7.12 (appt, J=7.8 Hz, 2H; indolyl
7-H), 6.95–6.87 (m, 6H; H-5, m-Ph), 6.73–6.64 (m, 6H; indolyl 6-H, o-
Ph), 4.85 (d, 2J=14.9 Hz, 1H; CHABn), 4.81 (d, 2J=14.9Hz, 1H;
CHBBn), 3.89–3.84 (m, 2H; NCHAHB), 3.64–3.61 (m, 2H; NCHAHB),
2.19–2.14 (m, 2H; NCH2CH2), 2.12–1.86 (m, 2H; NCH2CH2), 1.64–1.04
(m, 8H; N(CH2)2CH2, N(CH2)3CH2), 0.88–0.65 ppm (m, 4H; N-
(CH2)4CH2); IR (film): n˜ =2928, 2855, 1704, 1545 cmꢀ1; MS (ES) m/z
(%): 708 (100) [M++H]; HRMS: m/z calcd for C49H45N3O2 [M++H]:
708.3590; found: 708.3598.
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Received: May 10, 2005
Published online: August 1, 2005
Chem. Eur. J. 2005, 11, 6277 – 6285
ꢀ 2005 Wiley-VCH Verlag GmbH & Co.KGaA, Weinheim
www.chemeurj.org
6285