2
P. S. Humphries et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Figure 1. KL001 and chosen lead scaffold 1.
Scheme 2. Reagents and conditions: (a) MeSO2Cl, pyridine or Si-imidazole, CH2Cl2,
rt, 16–48 h; (b) 3, Cs2CO3, DMA, 100 °C, 16 h.
binding site of CRY2 and interferes with the binding of FBXL3 to
CRY2.26 In order to determine the structural requirements for
KL001 activity and the molecular basis for highly potent deriva-
tives, we wished to synthesize compounds represented by the gen-
eral structure 1 (Fig. 1). Herein, we report the synthesis, structure–
activity relationships (SAR), and in vivo activity of this new class of
compounds.27
The synthetic route for the preparation of compounds 4–14 is
shown in Scheme 1. Coupling of commercially available epibromo-
hydrin and carbazole 2 afforded 3. Condensing furfurylamine with
epoxide 3 followed by addition of a variety of sulfonyl chlorides
allowed for an efficient variation of sulfonamide R2 substituents
giving final products 4–14.
Scheme 3. Reagents and conditions: (a) (NH2)2SO2, pyridine, reflux, 16 h; (b) 3,
NaH, DMF, 70 °C, 16 h.
In an effort to look at the effects of variation of the amine R1
moiety, a new synthetic route was required (Scheme 2). Methane-
sulfonyl chloride was added to a variety of amines 15 to afford 16.
The resulting sulfonamides 16 were then condensed with epoxide
3 under basic conditions giving final products 17–37.
was calculated in an effort to move toward a quality drug candi-
date with high potency, low dose and an adequate safety profile.
LipE is a parameter used in drug design to evaluate the quality of
compounds, linking potency and lipophilicity in an attempt to esti-
mate druglikeness.28–30
Initially, we decided to investigate the effects of varying the sul-
fonamide R2 substituents of 1, Table 1. Compounds 4–14 demon-
strate that Per2 potency is sensitive to the size of the
substituent. Small substituents yielded analogs with submicromo-
lar potency (e.g. KL001 and compound 4). Larger cyclic moieties
(compounds 9 and 13) resulted in significant reductions in Per2
affinity. Methyl, ethyl and 2-methoxyethyl were preferable (com-
pounds KL001, 4 and 12) from a LipE standpoint, due to their bal-
ance of Per2 potency and lipophilicity.
The optimal lipophilic efficiency, molecular weight and in vitro
metabolism (data not shown) associated with the methyl sub-
stituent (e.g. KL001) prompted us to hold this moiety constant
and make changes to the R1 substituents (Table 2). Per2 potency
for compounds 17–37 demonstrate that large changes in this
region can be tolerated. Removing the R1 substituent of KL001
(compound 17) afforded an increase in potency and LipE. Per2
potency of ꢀ100 nM could be achieved with two analogs (com-
pounds 22 and 24) although only minor increases in LipE resulted
from these changes. A lipophilicity lowering strategy in this region
was also successful as compounds 27, 28 and 31 all resulted in
acceptable LipE values although at the expense of Per2 potency.
Interestingly, replacing a phenyl moiety (compound 32) with a 2-
pyridyl substituent (compound 33) resulted in a significant reduc-
tion in Per2 potency possibly due to an unfavorable conformational
change caused by an interaction with this new moiety and the sul-
fonamide functionality.
Attempts to further increase potency via conformational
restriction required molecules where R1 and R2 are joined as part
of
a ring (Scheme 3). Commercially available sultams (39,
X = CH2) and synthesized cyclic sulfamides (39, X = NR3) were
introduced to this effect. Diamines 38 were condensed with sul-
famide to afford intermediates 39. Cyclic sulfonamides and sul-
famides 39 were then condensed with epoxide 3 under basic
conditions giving final products 40–47.
The newly synthesized compounds were evaluated for their
effects on circadian rhythms in a human osteosarcoma U2OS cell
line harboring a Per2-dLuc luciferase reporter.23 Continuous treat-
ment with these compounds causes period lengthening and ampli-
tude reduction in a dose-dependent manner. The amplitude effect
of these derivatives was analyzed by testing 8 points of threefold
dilution series to obtain EC50 values. Lipophilic efficiency (LipE)
Given the increased potency and LipE associated with small R1
and R2 substituents we decided to attempt a conformational
restriction strategy (Table 3). The rationale behind conformational
restriction is to keep a molecule in its bioactive conformation while
eliminating alternative conformations. Cyclic sulfonamides (com-
pounds 40–43) all showed excellent Per2 potency and LipE values.
Variations in ring size (compounds 40, 41 and 42) resulted in no
change in Per2 potency. One example of lipophilicity lowering
(compound 43) afforded very high Per2 potency and LipE. On mov-
ing from cyclic sulfonamides to cyclic sulfamides (compounds 44–
47) we also observed acceptable Per2 affinity. Within the sul-
famides, we did observe a trend toward improved Per2 affinity
Scheme 1. Reagents and conditions: (a) epibromohydrin, KOH, DMF, 0 °C to rt,
16 h, 71%; (b) furfurylamine, EtOH, 40 °C, 16 h, 91%; (c) R2SO2Cl, pyridine, CH2Cl2,
0 °C to rt, 16 h.