Identification of Atuveciclib (BAY 1143572), the First Highly Selective, Clinical PTEFb/CDK9 Inhibitor for the Treatment of Cancer

Article abstract of DOI:10.1002/cmdc.201700447

Selective inhibition of exclusively transcription-regulating PTEFb/CDK9 is a promising new approach in cancer therapy. Starting from lead compound BAY-958, lead optimization efforts strictly focusing on kinase selectivity, physicochemical and DMPK properties finally led to the identification of the orally available clinical candidate atuveciclib (BAY 1143572). Structurally characterized by an unusual benzyl sulfoximine group, BAY 1143572 exhibited the best overall profile in vitro and in vivo, including high efficacy and good tolerability in xenograft models in mice and rats. BAY 1143572 is the first potent and highly selective PTEFb/CDK9 inhibitor to enter clinical trials for the treatment of cancer.

Full text of DOI:10.1002/cmdc.201700447

Accepted Article
Title: Identification of Atuveciclib (BAY 1143572), the First Highly
Selective, Clinical PTEFb/CDK9 Inhibitor for the Treatment of
Cancer
Authors: Ulrich Lücking, Arne Scholz, Philip Lienau, Gerhard
Siemeister, Dirk Kosemund, Rolf Bohlmann, Hans Briem,
Ildiko Terebesi, Kirstin Meyer, Katja Prelle, Karsten Denner,
Ulf Boemer, Martina Schäfer, Knut Eis, Ray Valencia,
Stuart Ince, Franz von Nussbaum, Dominik Mumberg, Karl
Ziegelbauer, Bert Klebl, A. Choidas, Peter Nussbaumer, M.
Baumann, Carsten Schultz-Fademrecht, Gerd Rühter, Jan
Eickhoff, and Michael Brands
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.201700447
Link to VoR: http://dx.doi.org/10.1002/cmdc.201700447
A Journal of
www.chemmedchem.org

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
Identification of Atuveciclib (BAY 1143572), the First Highly
Selective, Clinical PTEFb/CDK9 Inhibitor for the Treatment of
Cancer
Ulrich Lücking,*[a] Arne Scholz,[a] Philip Lienau,[a] Gerhard Siemeister,[a] Dirk Kosemund,[a] Rolf
Bohlmann,[a] Hans Briem,[a] Ildiko Terebesi,[a] Kirstin Meyer,[a] Katja Prelle,[a] Karsten Denner,[a] Ulf
Bömer,[a] Martina Schäfer,[a] Knut Eis,[a] Ray Valencia,[a] Stuart Ince,[a] Franz von Nussbaum,[a] Dominik
Mumberg,[a] Karl Ziegelbauer,[a] Bert Klebl,[b] Axel Choidas,[b] Peter Nussbaumer,[b] Matthias
Baumann,[b] Carsten Schultz-Fademrecht,[b] Gerd Rühter,[b] Jan Eickhoff,[b] and Michael Brands[a]
[a]
[b]
Dr. U. Lücking (0000-0003-2466-5800), Dr. A. Scholz, Dr. P. Lienau, Dr. G. Siemeister, Dr. D. Kosemund, Dr. R. Bohlmann, Dr. H. Briem, Dr. I. Terebesi,
Dr. K. Meyer, Dr. K. Prelle, Dr. K. Denner, Dr. U. Bömer, Dr. M. Schäfer, Dr. K. Eis, Dr. R. Valencia, Dr. S. Ince, Dr. F. von Nussbaum, Dr. D. Mumberg, Dr.
K. Ziegelbauer, Dr. M. Brands
Bayer AG
Pharmaceuticals Division, Drug Discovery
Müllerstr. 178, 13353 Berlin (Germany)
E-mail: ulrich.luecking@bayer.com
Dr. B. Klebl, Dr. A. Choidas, Dr. P. Nussbaumer, Dr. M. Baumann, Dr. C. Schultz-Fademrecht, Dr. G. Rühter, Dr. J. Eickhoff
Lead Discovery Center GmbH (LDC)
Otto-Hahn-Str. 15, 44227 Dortmund (Germany)
Abstract: Selective inhibition of exclusively transcription-regulating
PTEFb/CDK9 is a promising new approach in cancer therapy.
Starting from lead compound BAY-958, lead optimization efforts
strictly focusing on kinase selectivity, physicochemical and DMPK
properties finally led to the identification of the orally available clinical
candidate atuveciclib (BAY 1143572). Structurally characterized by
an unusual benzyl sulfoximine group, BAY 1143572 exhibited the
best overall profile in vitro and in vivo, including high efficacy and
good tolerability in xenograft models in mice and rats. BAY 1143572
is the first potent and highly selective PTEFb/CDK9 inhibitor that
entered clinical trials for the treatment of cancer.
assembly of the preinitiation complex at the promoter region and
phosphorylation of Ser5 and Ser7 of the CTD by CDK7/cyclin H.
For most genes, RNA polymerase II stops mRNA transcription
after it has moved 20–40 nucleotides along the DNA template.
This promoter-proximal pausing of RNA polymerase II is
mediated by negative elongation factors and is recognized as a
major control mechanism to regulate expression of rapidly
induced genes in response to a variety of stimuli.[3] PTEFb is
crucially involved in overcoming promoter-proximal pausing of
RNA polymerase II and transition into a productive elongation
state by phosphorylation of Ser2 of the CTD, as well as by
phosphorylation and inactivation of negative elongation factors.
Deregulated kinase activity of CDK9 of the PTEFb heterodimer
is associated with a variety of human pathological conditions
such as hyperproliferative diseases (e.g., cancer), virally
induced infectious diseases and cardiovascular diseases.[4]
PTEFb-mediated transcription of short-lived, antiapoptotic
survival proteins, such as Mcl-1, and oncogenes, such as c-
MYC, plays a critical role in cancer cell growth and survival. In
addition, these proteins exhibit important functions in the
development of resistance to chemotherapy. Inhibition of
PTEFb/CDK9 results in the rapid depletion of short-lived mRNA
transcripts of these important survival proteins and oncogenes.
Thus, selective, transient inhibition of exclusively transcription-
regulating PTEFb/CDK9 is a promising new approach in cancer
therapy.[5]
Introduction
The cyclin-dependent kinase (CDK) family consists of members
that are key regulators of the cell-division cycle (cell-cycle
CDKs), involved in regulation of gene transcription
(transcriptional CDKs), and of members with other functions
(e.g., CDK5). Whereas cell-cycle CDKs 1, 2, 3, 4 and 6 are
required for the correct timing and order of events of the cell-
division cycle, transcriptional CDK7 and CDK9 regulate the
activity of RNA polymerase II via phosphorylation of the carboxy-
terminal domain (CTD). Since their discovery, CDKs have been
considered strong prospective targets for a new generation of
anticancer drugs.[1] Although numerous pharmaceutical
companies have initiated drug discovery efforts to identify low-
molecular-weight CDK inhibitors for cancer therapy, most pan-
CDK inhibitors have failed rigorous clinical testing. Nevertheless,
the recent approvals of the selective CDK4/6 inhibitors
palbociclib and ribociclib demonstrate that CDK inhibitors with
narrow selectivity profiles can have therapeutic clinical utility.[1c,2]
We now report the identification of atuveciclib (BAY 1143572,
Figure 1), the first potent and highly selective PTEFb/CDK9
inhibitor that entered clinical trials. Starting from lead compound
BAY-958, which displayed potent PTEFb-inhibitory activity and
high kinase selectivity in vitro, lead optimization by
a
collaborative effort involving medicinal chemistry, pharmacology,
DMPK, structural biology and computational chemistry led to the
identification of the orally available clinical candidate
BAY 1143572. Structurally characterized by an unusual benzyl
sulfoximine group, BAY 1143572 exhibited the most promising
overall profile with respect to potency, kinase selectivity,
Positive transcription elongation factor
b
(PTEFb) is
a
heterodimer of CDK9 and one of four cyclin partners, cyclin T1,
cyclin K, cyclin T2a or cyclin T2b. Whereas CDK9 is exclusively
involved in transcriptional regulation, CDK7 in addition
participates in cell-cycle regulation as a CDK-activating kinase.
Transcription of genes by RNA polymerase II is initiated by
1
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
physicochemical and DMPK properties, and antitumour efficacy
in animal models.
Additionally, there was excellent tolerability upon treatment with
BAY-958 hydrochloride, with a body weight change of less than
10% and no fatal toxicities. Nevertheless, due to the unfavorable
physicochemical and DMPK properties of BAY-958, we elected
to search for a compound with an improved overall profile.
BAY1143572
BAY-958
Figure 1. Structures of lead compound BAY-958 and clinical candidate
atuveciclib (BAY 1143572).
Results and Discussion
Search for low-molecular-weight PTEFb/CDK9 inhibitors
Figure 2. Antitumor efficacy of BAY-958 hydrochloride in an MOLM-13 human
AML model in mice. Treatments were started
3 days after tumor cell
Attracted by the encouraging biological rationale, we set course
to identify a potent, highly selective and orally applicable
PTEFb/CDK9 inhibitor exclusively targeting oncogenic
transcription for the treatment of cancer. Special attention was
paid to the aspect of high selectivity with respect to kinases in
general as well as within the CDK family, in particular high CDK9
selectivity. Such CDK9 selectivity was anticipated as being
crucial for the differentiation of a potential candidate from cell-
cycle-addressing selective CDK inhibitors (like the CDK4/6
inhibitor palbociclib[6]) and from pan-CDK inhibitors (like
dinaciclib[7] or roniciclib[8]), since a differentiated activity and
tolerability profile can be expected. Furthermore, favorable
physicochemical and DMPK properties, which have been key
hurdles in many kinase inhibitor projects,[9] were of high
importance.
inoculation. (A) Tumor growth. Asterisks indicate statistical significance in
comparison to the vehicle control, calculated using the mean tumor areas at
the time point when the vehicle group was sacrificed (*p <0.047, ***p <0.001).
(B) Body weight change expressed as a percentage of the starting weight.
Structural variation of lead compound BAY-958
Lead compound BAY-958 and structurally related derivatives
were easily accessible by straightforward chemistry (Scheme 1).
In the first step, commercial 2,4-dichloro-1,3,5-triazine (1) was
reacted with a suitable aniline under basic conditions to give the
corresponding N-aryl-4-chloro-1,3,5-triazin-2-amine 2. In the
second step, the crude coupling product 2 was reacted with a
boronic acid derivative to give the desired product 3.
In the search for a suitable lead structure, triazine BAY-958
(LDC 526, Figure 1) attracted our attention, since this compound
proved to be a potent PTEFb/CDK9 inhibitor that also displayed
very high kinase selectivity, even within the CDK family (in-
house kinase panel: IC50 CDK9/CycT1: 11 nM, selectivity vs
CDK2: 98, see Table 3; Millipore panel: IC50 CDK9/CycT1: 5 nM,
selectivity vs other CDKs: >90, see Table 4). BAY-958 also
exhibited good antiproliferative activity in vitro, for example
against HeLa cells (IC50: 1000 nM) and MOLM-13 cells (IC50: 280
nM). In vitro pharmacokinetic studies with BAY-958
demonstrated high metabolic stability in rat hepatocytes (ratHep)
and liver microsomes (ratLM), resulting in a low predicted blood
clearance (CLb) of 0.33 L/h/kg and 0.48 L/h/kg, respectively (see
Table 3).
On the other hand, BAY-958 has a rather low aqueous solubility
of 11 mg/L at pH 6.5. Moreover, we recorded a very moderate
permeability coefficient (Papp A–B) of 22 nm/s and a high efflux
ratio of 15 in Caco-2 cells, as well as a blood/plasma partitioning
in rats of about 3:1 (see Table 3). Pharmacokinetic studies in
rats in vivo revealed a low blood clearance (CLb: 0.5 L/h/kg), a
high volume of distribution (Vss: 1.4 L/kg) and a short half-life
(t1/2: 0.7 h) of BAY-958. After oral administration, a low
bioavailability of only 10% was observed.
a)
b)
1
2
3
Scheme 1. General synthesis of triazine-based PTEFb inhibitors. Reagents
and conditions: a) R1NH2, DIPEA, THF/iPrOH (1:1), –40 °C to 0 °C; b)
R2B(OR)2, Pd(dppf)Cl2·DCM, K3PO4, dioxane/water (10:1), 140–145 °C,
microwave oven or R2B(OR)2, Pd(PPh3)4, K2CO3 (aq), 1,2-dimethoxyethane,
100 °C.
The binding mode of lead compound BAY-958 with CDK9 was
investigated by docking experiments (Figure 3). According to
these modeling studies, binding to the hinge region of CDK9 is
mediated by the triazine core and the aniline NH. The benzyl
sulfonamide moiety is directed towards the exit of the ATP
binding pocket, whereas the methoxyphenyl substituent points
towards the ribose pocket. Favorable interactions include two
hydrogen bonds to the hinge region and two hydrogen bonds
formed by the amino part of the sulfonamide group, as well as a
-stacking interaction of the methoxyphenyl moiety with the
Phe103 gatekeeper residue. In addition, a weak hydrogen bond
between the para-fluoro substituent and the catalytic Lys48 may
be postulated.[10] However, the high CDK9 selectivity of BAY-958,
versus other CDKs, could not be rationalized.
Due to the slow absorption and low bioavailability of BAY-958
observed in the rat pharmacokinetic studies, the hydrochloride of
BAY-958 was used in a subsequent mouse xenograft study to
analyze the in vivo efficacy. Daily oral administration of BAY-958
hydrochloride at 30 or 40 mg/kg resulted in a marked inhibition
of tumor growth with treatment-to-control (T/C) ratios of 0.16 and
0.12, respectively, at the end of the experiment (Figure 2).
2
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
32
19
1340
5
49
12
2200
6
Figure 3. Docking mode of BAY-958 in complex with CDK9. The compound
was docked into a published X-ray complex of CDK9/CycT1 (PDB code 3MY1)
using the Glide docking program. The sulfonamide moiety forms two hydrogen
bonds, one to Glu107 (main chain) and the other to Asp109 (side chain). For
the substituted phenyl ring attached to the triazine, a -stacking interaction
with Phe103 and a weak hydrogen bond to Lys48 may be postulated.
7
22
<1
550
11000
2900
7
8
9
51
Analyzing the chemical structure of lead compound BAY-958,
the chemistry team was well-disposed towards inclusion of the
central triazine core. The benzyl sulfonamide group, however,
raised concerns: due to its polarity, high number of heteroatoms
and multiple hydrogen-bonding properties, it was suspected of
contributing significantly to the low aqueous solubility of BAY-
958, the rather moderate Caco-2 permeability and the high efflux
ratio. Furthermore, the acidity properties and potential instability
of the benzyl sulfonamide group were considered and ultimately
triggered the evaluation of potential structural alternatives at this
position (Table 1).
47
43
85
1000
20
21
>20
13
3200
10
11
12
Table 1. Selected examples of lead optimization efforts on the western side
and corresponding key in vitro properties.
ND
Compound
R
CDK9/CycT1
IC50 [nM]
Selectivity
vs CDK2,
ratio of
IC50
HeLa[a]
IC50 [nM]
890
values
11
24
98
67
1000
1500
BAY-958
67
19
4200
13
4
[a] Cells were treated with test compounds for 96 h; ND: not determined.
3
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
Sulfones aroused our attention, since analogue 4 exhibited a
similar CDK9-inhibitory activity as BAY-958 in our biochemical
assay in vitro [IC50 CDK9/CycT1 (in-house), 4: 24 nM vs BAY-
958: 11 nM, see Table 3], even though the sulfone group cannot
form similar hydrogen bonds as the amino part of the
sulfonamide group (Figure 3). Sulfone 4 also exhibited similar
selectivity against cell-cycle kinase CDK2 (ratio of IC50 values
CDK2/CDK9, 4: 67 vs BAY-958: 98) and antiproliferative activity
against HeLa cells (IC50 4: 1.5 M vs BAY-958: 1.0 M). To
address the initial concerns regarding the stability and acidity of
a polar functional group at the benzylic position, analogues 5
and 6 were synthesized; however, in this sulfone series, the
removal of the methylene group from 4 (compound 5) or the
introduction of an additional methylene group to 4 (compound 6)
resulted in both reduced PTEFb/CDK9 activity and significantly
reduced selectivity against cell-cycle kinase CDK2 (Table 1).
This finding was quite surprising, as the sulfone group is
directed towards the exit of the ATP binding pocket. Furthermore,
the introduction of methyl or even fluoro substituent(s) at the
benzylic position was not well-tolerated with regard to activity
against CDK9 (compounds 10, 11, 13), as well as selectivity
against CDK2 (compounds 10–13). Inclusion of an annulated
ring system (compounds 7, 8) also resulted in poor CDK9
selectivity.
1800
14
ND
16
17
18
130
6
2900
270
16
>3000
Extensive structural variation on the eastern side of lead
compound BAY-958 also revealed a rather steep SAR with
respect to selectivity against CDK2 and potency (Table 2).
620
20
>3000
19
Table 2. Selected examples of lead optimization efforts on the eastern side
and corresponding key in vitro properties.
1700
>11
ND
20
Compound
R
CDK9/CycT1
IC50 [nM]
Selectivity
vs CDK2,
ratio of IC50
values
HeLa[a]
IC50 [nM]
3600
<1
3000
21
11
98
1000
BAY-958
15000
1
ND
22
23
27
62
1600
3800
14
15
4
34
100
23
[a] Cells were treated with test compounds for 96 h; ND: not determined.
108
The para-fluoro substituent on the phenyl ring of BAY-958
proved to be beneficial for antiproliferative activity in vitro:
analogues 14 (p-H) and 15 (p-Cl) both displayed reduced IC50
values against HeLa cells (IC50 14: 1.6 M, 15: 3.8 M vs BAY-
4
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
958: 1.0 M). Pyridyl analogue 16 exhibited significantly reduced
activity against CDK9 in the biochemical assay (IC50 16: 1800
nM). Moreover, an ortho-alkoxy substituent was crucial for the
desired CDK9 selectivity, as well as in vitro potency; various
functional groups were evaluated as potential structural
alternatives, but all such analogues displayed significantly
reduced selectivity against CDK2 and inhibitory activity against
CDK9 (see Table 2 for selected examples; e.g., 17–22).
Introduction of a sterically larger alkoxy substituent, especially a
benzyloxy group, resulted in significantly improved potency;
however, most of these compounds suffered from very low
metabolic stability in vitro. For instance, whilst benzyl analogue
23 proved to be a highly potent CDK9 inhibitor, it exhibited low
metabolic stability in liver microsome preparations of human,
mouse and rat origin, with recovery of the parent compound
group has rarely been used in life science approaches, even
though it offers a unique combination of interesting properties,
namely high stability, favorable physicochemical properties,
hydrogen-bond acceptor/donor functionalities and structural
diversity.[15] Lately, however, there has been a rapidly increasing
interest in sulfoximines as pharmacophores in the life
sciences.[15,16] Our prior, long-standing interest in sulfoximines in
medicinal chemistry had been exclusively focused on aryl
sulfoximines;[17] nevertheless, we then decided to evaluate a
benzyl sulfoximine analogue of BAY-958.
In the corresponding synthesis (Scheme 2), thioether 25,
prepared from the commercial benzyl chloride 24, was oxidized
and the resulting sulfoxide 26 was converted into sulfoximine 27
using the rhodium-catalyzed method of Okamura and Bolm.[18]
Next, a protecting group was introduced at the sulfoximine
after incubation for 60 minutes of 0%, 0.2% and 0%, respectively. nitrogen to provide 28. The nitro group of 28 was reduced and
the resulting aniline 29 reacted with 2,4-dichloro-1,3,5-triazine
Identification of BAY 1143572
(1) under basic conditions. The crude coupling product 30 was
used in a subsequent Suzuki reaction to yield compound 31.
Finally, deprotection and chiral HPLC separation provided the
(R)-sulfoximine BAY 1143572 and its enantiomer 32.[19] The
stereochemistry at the sulfur of the sulfoximine group was
determined by X-ray crystallography of the corresponding N-
acetyl derivative 33 (Figure 4).
Since the late discovery of the sulfoximine group in 1949,[11]
sulfoximine chemistry[12] has been rather a niche discipline.
Applications have mainly centered around the use of
sulfoximines as either chiral auxiliaries[13] or ligands in
asymmetric catalysis.[14] Until very recently, the sulfoximine
a)
b)
c-d)
27
25
26
24
e)
f)
g)
28
29
30
h)
i-j)
31
32 (S) and BAY 1143572 (R): R1 = H
k)
33: R1 = Ac
Scheme 2. Synthesis of BAY 1143572. Reagents and conditions: a) NaSMe, EtOH, –15 °C, then RT, 3 h; b) H5IO6, FeCl3, MeCN, RT, 90 min, 70% (2 steps); c)
CF3C(O)NH2, PhI(OAc)2, MgO, Rh2(OAc)4, DCM, RT, 16 h; d) K2CO3, MeOH, RT, 1 h, 79% (2 steps); e) ClC(O)OEt, pyridine, 0 °C to RT, 24 h; f) TiCl3, THF, RT,
18 h, 94% (2 steps); g) 2,4-dichloro-1,3,5-triazine (1), DIPEA, THF/iPrOH (1:1), –40 °C to 0 °C, 3 h; h) 4-fluoro-2-methoxyphenylboronic acid, Pd(PPh3)4, K2CO3
(aq), 1,2-dimethoxyethane, 100 °C, 80 min, 36%; i) NaOEt, EtOH, 60 °C, 7 h, 90%; j) preparative chiral HPLC; k) AcCl, TEA, DCM, 0 °C to RT, 3 h, 57%.
5
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
potential CYP1A2 in vitro induction liability. Induction of CYP3A4
in vitro was not observed.
Table 3. Properties of PTEFb inhibitors BAY-958, benzyl sulfone 4 and
BAY 1143572.
BAY-958
11
4
BAY 1143572
13
24
CDK9/CycT1, IC50 [nM]
Selectivity vs CDK2,
ratio of IC50 values
98
67
100
HeLa, IC50 [nM] [a]
MOLM-13, IC50 [nM] [a]
TPSA[b]
1000
280
120.0
11
1500
130
94.1
4
920
310
100.9
479
35
Figure 4. X-ray structure of N-acetyl derivative 33.
Sw, pH 6.5 [mg/L][c]
Papp A–B [nm/s]
In comparison to BAY-958 and benzyl sulfone 4, benzyl
sulfoximine BAY 1143572 clearly exhibited the best overall
profile in vitro and in vivo (Table 3). BAY 1143572 is a potent
and highly selective CDK9 inhibitor (IC50 CDK9/CycT1: 13 nM,
ratio of IC50 values CDK2/CDK9: 100). The selectivity of
BAY 1143572 within the CDK family is even higher than that of
lead compound BAY-958 (Table 4). Outside the CDK family,
submicromolar inhibitory activity was only recorded against
GSK3 kinase(IC50 GSK345 nM, GSK387 nM) (see the
Supporting Information). BAY 1143572 also demonstrated
comparable antiproliferative activity as lead compound BAY-958,
for example against HeLa cells (IC50 BAY 1143572: 920 nM,
BAY-958: 1000 nM) and MOLM-13 cells (IC50 BAY 1143572: 310
nM, BAY-958: 280 nM). In comparison to BAY 1143572, the (S)-
enantiomer 32 revealed very similar in vitro properties, well
within the limits of measurement accuracy; however, with
multiple batches of enantiomer 32 there was a trend toward a
slightly reduced activity against CDK9 in the biochemical assay
(IC50 CDK9/CycT1: 16 nM) and antiproliferative activity against
HeLa cells (IC50: 1100 nM). Surprisingly, BAY 1143572 has a
high aqueous solubility of 479 mg/L, whereas both BAY-958 and
benzyl sulfone 4 have a low aqueous solubility of 11 mg/L and 4
mg/L, respectively. Nevertheless, BAY 1143572 demonstrated
improved Caco-2 permeability and a reduced efflux ratio (Papp
A–B: 35 nm/s, ER: 6) relative to lead compound BAY-958 (Papp
A–B: 22 nm/s, ER: 15).
22
143
1.2
15
6
Efflux ratio
0.33
0.48
0.50
1.4
0.79
0.58
2.7
0.17
0.15
1.1
CLb, ratHep [L/h/kg]
CLb, ratLM [L/h/kg]
CLb, rat in vivo, iv [L/h/kg]
Vss, rat in vivo, iv [L/kg]
t1/2, rat in vivo, iv [h]
1.5
1.0
0.7
0.3
0.6
AUC,[d] rat in vivo, p.o.
[mg·h/L]
0.11
0.16
0.28
Cmax,
[d] rat in vivo, p.o.
[mg/L]
0.029
0.059
0.058
10
3.0
>20
53
1.2
>20
54
1.1
>20
F, rat in vivo, p.o. [%]
Blood/plasma ratio (rat)
CYP inhibition [M]
CYP1A2 induction
NOEL[e]
5 g/L
no (up to
370 g/L)
no (up to
370 g/L)
[a] Cells were treated with test compounds for 96 h. [b] TPSA: topological
polar surface area.[20] [c] The solid state of the test compounds was not
characterized. [d] Normalized to 1 mg/kg. [e] NOEL: no-observed-effect-level.
In an in vivo pharmacokinetic study in rats, BAY 1143572
showed low blood clearance (CLb 1.1 L/h/kg), whereas benzyl
sulfone 4 exhibited a significantly higher blood clearance (2.7
L/h/kg) (Table 3). The volumes of distribution (Vss) of BAY-958
(1.4 L/kg), benzyl sulfone 4 (1.5 L/kg) and BAY 1143572 (1.0
L/kg) proved to be rather comparable but, in contrast to lead
compound BAY-958, benzyl sulfone 4 and BAY 1143572
showed significantly improved oral bioavailability (4: 53%,
Table 4. CDK-inhibitory activity of lead compound BAY-958 and clinical
candidate BAY 1143572 in the Merck Millipore KinaseProfiler panel.
IC50 [nM]
BAY-958
5
BAY 1143572
6
CDK9/CycT1(h)
CDK1/CycB(h)
CDK2/CycE(h)
CDK3/CycE(h)
CDK5/p35(h)
BAY 1143572: 54%). Furthermore, benzyl sulfone
4 and
690
1100
BAY 1143572 exhibited blood/plasma ratios of about 1.
Compared to BAY 1143572, its enantiomer 32 revealed very
similar rat PK properties in vivo (CLb: 1.2 L/h/kg, Vss: 1.2 L/kg,
t1/2: 0.6 h, F: 53%). All three key CDK9 inhibitors, BAY-958,
benzyl sulfone 4 and BAY 1143572, did not show significant
inhibition of cytochrome P450 activity, with IC50 values >20 M;
however, the switch from benzyl sulfonamide BAY-958 to benzyl
sulfone 4 and benzyl sulfoximine BAY 1143572 removed a
470
1000
570
890
800
1600
4400
>10000
>10000
>10000
CDK6/CycD3(h)
CDK7/CycH/MAT1(h)
6
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
For the subsequent in vivo studies, two xenograft models of
human acute myeloid leukemia (AML) were selected since it is
well known from the preclinical literature[21] that this indication is
dependent on oncogenic transcription and also the only
indication where nonselective CDK inhibitors with CDK9
inhibitory activity have reached randomized phase 2 clinical
trials (NCT01349972, NCT00634244, NCT00795002). Of note,
BAY 1143572 did not show relevant activity against Flt3 in the
Merck Millipore KinaseProfiler panel (see the Supporting
Information); therefore, the possibility that in vivo activity of the
compound might be partially mediated by Flt3 inhibition in Flt3-
alterated MOLM-13 and MV4-11 cells can be excluded.
In in vivo efficacy studies in the MOLM-13 xenograft model in
mice, BAY 1143572 demonstrated greater potency and higher
antitumor efficacy than benzyl sulfone 4 (Figure 5A and Table 5).
Daily treatment with benzyl sulfone 4 at the maximum tolerated
dose of 20 mg/kg revealed no efficacy (T/C ratio of 0.73, p =
0.073). In contrast, daily administration of BAY 1143572 at 6.25
or 12.5 mg/kg resulted in a dose-dependent antitumor efficacy
with a T/C ratio of 0.64 and 0.49, respectively (p <0.001). In a
separate experiment with a higher daily dose of 20 or 25 mg/kg
BAY 1143572, antitumor efficacy with a T/C ratio of 0.41 and
0.31, respectively, was observed (p <0.001). The 25 mg/kg once
daily dose is the maximum tolerated dose in nude mice.
Furthermore, BAY 1143572 administered at 25 or 35 mg/kg, 3
days on/2 days off, resulted in a T/C ratio of 0.33 and 0.20,
respectively (p <0.001), suggesting a possible flexibility for the
selection of a clinical dosing regimen (Figure 5B and Table 5).
Treatment with BAY 1143572 was well-tolerated, as
demonstrated by less than 10% mean body weight reduction
throughout the study.
(maximum tolerated dose) resulted in almost complete tumor
remission, followed by delayed regrowth after cessation of the
treatment (Figure 5C). At the conclusion of the experiment (57
days after initial inoculation), there was no tumor regrowth in
nine of the 12 test animals.
Table 5. Antitumor efficacy of compound 4 and BAY 1143572 in human AML
models in female NMRI nu/nu mice (MOLM-13) and athymic rats (MV4-11).
[a]
Animal
model
Treatment
Dose
Schedule
T/Carea
Fatal
toxicity
Max
BWC
(%)[b]
Vehicle
0
QD
QD
1.00
0.91
0/10
0/10
0/10
0/10
1/10
1/13
0/13
0/13
0/13
0/13
1/12
0/12
+13
+11
+11
+10
+10
0
MOLM-
13
model
in mice
mg/kg
10
mg/kg
20
mg/kg
6.25
mg/kg
12.5
mg/kg
0
mg/kg
20
mg/kg
25
mg/kg
25
mg/kg
35
Compound 4
QD
0.73
BAY 1143572
QD
0.64*
0.49*
1.00
QD
Vehicle
QD
MOLM-
13
model
in mice
BAY 1143572
QD
0.41*
0.31*
0.33*
0.20*
1.00
–1
QD
–3
3 on/2 off
3 on/2 off
QD
–2
–4
mg/kg
0
mg/kg
12
Vehicle
–1
MV4-
11
BAY 1143572
QD×14
0.05*
–1
model
in rats
mg/kg
[a] Treatment-to-control (T/C) ratios and statistical significances were
calculated using the mean tumor areas at the time point when the vehicle
group was sacrificed. An asterisk indicates statistical significance (p <0.001) in
comparison to the vehicle control. [b] The maximum body weight change
(BWC) expressed as a percentage of the starting weight, for duration of the
treatment.
To further substantiate the antitumor efficacy, BAY 1143572 was
evaluated in an MV4-11 human acute myeloid leukemia model
in nude rats. Of note, the antiproliferative activity of
BAY 1143572 against MV4-11 cells (IC50: 890 nM) and HeLa
cells (IC50: 920 nM) in vitro is comparable, and is slightly
improved against MOLM-13 cells (IC50: 310 nM). Daily oral
administration of BAY 1143572 at 12 mg/kg for 14 days
Figure 5. Antitumor efficacy in two AML models in mice and rats. A, B: Antitumor efficacy of BAY 1143572 in an MOLM-13 human AML model in mice. (A) Tumor
growth in mice treated with compound 4 or BAY 1143572. Treatments were started 3 days after tumor cell inoculation. (B) Tumor growth in mice treated with
BAY 1143572 once daily (QD) or with an intermittent 3 days on/2 days off dosing schedule. Treatments were started 4 days after tumor cell inoculation. C:
Antitumor efficacy of BAY 1143572 in an MV4-11 human AML model in rats. Tumor growth in rats treated with vehicle or BAY 1143572. Treatments were started
13 days after tumor cell inoculation and were continued for 14 days, until the vehicle group was sacrificed. Asterisks indicate statistical significance in comparison
to the vehicle control, calculated using the mean tumor areas at the time point when the vehicle group was sacrificed (***p <0.001, ns = not significant).
Due to the promising overall profile in vitro and in vivo, benzyl
sulfoximine BAY 1143572 was selected as the development
candidate and entered phase 1 clinical trials in patients with
advanced
cancer
and
leukemia
(NCT01938638,
NCT02345382).[22]
7
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
substrate was evaluated by measurement of the resonance energy
transfer from the europium chelate to the streptavidin-XL. Therefore, the
fluorescence emissions at 620 nm and 665 nm, after excitation at 350 nm,
Conclusions
The benzyl sulfoximine atuveciclib (BAY 1143572) is a potent
and highly selective, oral PTEFb/CDK9 inhibitor. During lead
optimization, a surprisingly steep SAR with regard to the key
optimization parameters kinase selectivity and potency was
recorded. BAY 1143572 clearly exhibited the most promising
overall profile with respect to potency, selectivity,
physicochemical properties, and in vivo PK as well as in vivo
potency. Notably, in contrast to lead compound BAY-958 and
benzyl sulfone 4, BAY 1143572 combines high aqueous
solubility with moderate Caco-2 permeability and moderate
efflux, as well as low blood clearance and moderate
bioavailability in rats in vivo. Surprisingly, the switch from the
benzyl sulfonamide BAY-958 to the benzyl sulfoximine
BAY 1143572 removed an in vitro CYP1A2 induction liability.
BAY 1143572 is efficacious in human xenograft tumor models of
acute myeloid leukemia in both mice and rats. The compound is
active upon once-daily dosing, as well as upon intermittent
dosing schedules, providing valuable options to optimize human
dosing schedules with respect to efficacy and tolerability.
BAY 1143572 is the first selective PTEFb/CDK9 inhibitor that
entered clinical evaluation. The introduction of the uncommon
benzyl sulfoximine group was crucial for overcoming hurdles in
this project, which underlines the constant need for novel
chemical functionalities and methodologies as a means to solve
biological problems.
were measured with
a TR-FRET reader [e.g., PHERAstar (BMG
Labtechnologies, Offenburg, Germany) or ViewLux (Perkin-Elmer)]. The
ratio of the emissions at 665 nm and 620 nm was taken as the measure
of the amount of phosphorylated substrate. The data were normalized
(enzyme reaction without inhibitor = 0% inhibition, all other assay
components but no enzyme
= 100% inhibition). Usually the test
compound was tested on the same microtiter plate at 11 different
concentrations in the range of 20 M to 0.1 nM (20 M, 5.9 M, 1.7 M,
0.51 M, 0.15 M, 44 nM, 13 nM, 3.8 nM, 1.1 nM, 0.33 nM and 0.1 nM; the
dilution series was prepared separately before the assay on the 100-fold
concentrated solution in DMSO by serial 1:3.4 dilutions) in duplicate for
each concentration. IC50 values were calculated using a 4-parameter fit.
CDK2/CycE
Recombinant fusion proteins of GST and human CDK2 and of GST and
human CycE, expressed in insect cells (Sf9) and purified by Glutathione-
Sepharose affinity chromatography, were purchased from ProQinase
GmbH (Freiburg, Germany). As substrate for the kinase reaction, the
biotinylated peptide biotin–Ttds–YISPLKSPYKISEG (C-terminus in amide
form) was used, which is commercially available (e.g., from JERINI
Peptide Technologies, Berlin, Germany). For assays, 50 nL of a 100-fold
concentrated solution of the test compound in DMSO was pipetted into a
black, low volume, 384-well microtiter plate (Greiner Bio-One,
Frickenhausen, Germany); 2 L of a solution of CDK2/CycE in aqueous
assay buffer [50 mM Tris/HCl pH 8.0, 10 mM MgCl2, 1.0 mM dithiothreitol,
0.1 mM sodium ortho-vanadate, 0.01% (v/v) Nonidet P-40 (Sigma)] was
added, and the mixture was incubated for 15 min at 22 °C to allow pre-
binding of the test compound to the enzyme before the start of the kinase
reaction. Then, the kinase reaction was started by the addition of 3 L of
a solution of ATP (16.7 M; final concn in the 5 L assay volume: 10 M)
and substrate (1.25 M; final concn in the 5 L assay volume: 0.75 M) in
assay buffer, and the resulting mixture was incubated for 25 min at 22 °C.
The concentration of CDK2/CycE was adjusted depending on the activity
of the enzyme lot and was chosen appropriate to have the assay in the
linear range; typical concentrations were in the order of 130 ng/mL. The
reaction was stopped by the addition of 5 L of a solution of TR-FRET
detection reagents [0.2 M streptavidin-XL665 (Cisbio Bioassays,
Codolet, France), 1 nM anti-RB(pSer807/pSer811) antibody (BD
Pharmingen, Cat. no. 558389) and 1.2 nM LANCE EU-W1024 labeled
anti-mouse IgG antibody (Perkin-Elmer, product no. AD0077)] in an
aqueous EDTA solution [100 mM EDTA, 0.2% (w/v) BSA in 100 mM
HEPES/NaOH pH 7.0]. The resulting mixture was incubated for 1 h at
22 °C to allow the formation of a complex between the phosphorylated
biotinylated peptide and the detection reagents. Subsequently, the
amount of phosphorylated substrate was evaluated by measurement of
the resonance energy transfer from the europium chelate to the
streptavidin-XL. Therefore, the fluorescence emissions at 620 nm and
665 nm, after excitation at 350 nm, were measured with a TR-FRET
reader [e.g., PHERAstar (BMG Labtechnologies, Offenburg, Germany) or
ViewLux (Perkin-Elmer)]. The ratio of the emissions at 665 nm and 620
nm was taken as the measure of the amount of phosphorylated substrate.
The data were normalized (enzyme reaction without inhibitor = 0%
inhibition, all other assay components but no enzyme = 100% inhibition).
Usually the test compound was tested on the same microtiter plate at 11
different concentrations in the range of 20 M to 0.1 nM (20 M, 5.9 M,
1.7 M, 0.51 M, 0.15 M, 44 nM, 13 nM, 3.8 nM, 1.1 nM, 0.33 nM and 0.1
nM; the dilution series was prepared separately before the assay on the
100-fold concentrated solution in DMSO by serial 1:3.4 dilutions) in
duplicate for each concentration. IC50 values were calculated using a 4-
parameter fit.
Experimental Section
Kinase assays
CDK9/CycT1
Recombinant full-length His-tagged human CDK9 and CycT1, expressed
in insect cells and purified by Ni-NTA affinity chromatography, were
purchased from Invitrogen (Cat. no. PV4131). As substrate for the kinase
reaction, the biotinylated peptide biotin–Ttds–YISPLKSPYKISEG (C-
terminus in amide form) was used, which is commercially available (e.g.,
from JERINI Peptide Technologies, Berlin, Germany). For assays, 50 nL
of a 100-fold concentrated solution of the test compound in DMSO was
pipetted into a black, low volume, 384-well microtiter plate (Greiner Bio-
One, Frickenhausen, Germany); 2 L of a solution of CDK9/CycT1 in
aqueous assay buffer [50 mM Tris/HCl pH 8.0, 10 mM MgCl2, 1.0 mM
dithiothreitol, 0.1 mM sodium ortho-vanadate, 0.01% (v/v) Nonidet P-40
(Sigma)] was added, and the mixture was incubated for 15 min at 22 °C
to allow pre-binding of the test compound to the enzyme before the start
of the kinase reaction. Then, the kinase reaction was started by the
addition of 3 L of a solution of adenosine triphosphate (ATP, 16.7 M;
final concn in the 5 L assay volume: 10 M) and substrate (1.67 M; final
concn in the 5 L assay volume: 1 M) in assay buffer, and the resulting
mixture was incubated for 25 min at 22 °C. The concentration of
CDK9/CycT1 was adjusted depending on the activity of the enzyme lot
and was chosen appropriate to have the assay in the linear range: typical
concentrations were in the order of 1 g/mL. The reaction was stopped
by the addition of 5 L of a solution of TR-FRET detection reagents [0.2
M streptavidin-XL665 (Cisbio Bioassays, Codolet, France), 1 nM anti-
RB(pSer807/pSer811) antibody (BD Pharmingen, Cat. no. 558389) and
1.2 nM LANCE EU-W1024 labeled anti-mouse IgG antibody (Perkin-
Elmer, product no. AD0077)] in an aqueous EDTA solution [100 mM
EDTA, 0.2% (w/v) bovine serum albumin (BSA) in 100 mM HEPES/NaOH
pH 7.0]. The resulting mixture was incubated for 1 h at 22 °C to allow the
formation of a complex between the phosphorylated biotinylated peptide
and the detection reagents. Subsequently, the amount of phosphorylated
Merck Millipore CDK assays
8
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
Assays were performed according to the Merck Millipore KinaseProfiler
standard protocols, with an ATP concentration of 10 M.
blood clearance (CLb) and the maximal oral bioavailability (Fmax) were
calculated.
Proliferation assay
The inhibitory potency of the test compounds towards cytochrome P450
dependent metabolic pathways was determined in human liver
microsomes by applying individual CYP isoform-selective standard
probes (CYP1A2, phenacetin; CYP2C8, amodiaquine; CYP2C9,
diclofenac; CYP2D6, dextromethorphan; CYP3A4, midazolam).
Reference inhibitors were included as positive controls. Incubation
conditions (protein and substrate concentration, incubation time) were
optimized with regard to linearity of metabolite formation. Assays were
processed in 96-well plates at 37 °C by using a Genesis Workstation
(Tecan, Crailsheim, Germany). After protein precipitation, the metabolite
formation was quantified by LC-MS/MS analysis followed by inhibition
evaluation and IC50 calculation.
HeLa human cervical tumor cells (CCL-2) were obtained from the
American Type Culture Collection (Manassas, USA) and MOLM-13
human acute myeloid leukemia cells (ACC 554) were obtained from the
German Collection of Microorganisms and Cell Cultures (Braunschweig,
Germany). Authentication of cell lines was conducted at the German
Collection of Microorganisms and Cell Cultures via PCR-based DNA
profiling of polymorphic short tandem repeats. Cells were propagated
under the suggested growth conditions in a humidified 37 °C incubator.
Proliferation assays were conducted in 96-well plates at densities of 3000
(HeLa) and 5000 (MOLM-13) cells per well in the growth medium
containing 10% fetal calf serum (FCS). Cells were treated in
quadruplicate with serial dilutions of test compounds for 96 h. Relative
cell numbers were quantified by crystal violet staining (HeLa)[23] or
CellTitre-Glo Luminescent Cell Viability Assay (Promega) (MOLM-13).
IC50 values (inhibitory concentration at 50% of maximal effect) were
determined by means of a 4-parameter fit on measurement data which
were normalized to vehicle (DMSO) treated cells (= 100%) and
measurement readings taken immediately before compound exposure (=
0%).
To evaluate the CYP induction potential in vitro, cultured human
hepatocytes from three separate livers were treated once daily for three
consecutive days with vehicle control, one of eight concentrations of test
compound and known human CYP inducers (e.g., omeprazole,
phenobarbital, rifampin). After treatment, the cells were incubated in situ
with the appropriate marker substrates for the analysis of CYP3A4 and
CYP1A2 activity by LC-MS/MS.
Caco-2 cells (purchased from the German Collection of Microorganisms
and Cell Cultures, Braunschweig, Germany) were seeded at a density of
4.5 × 104 cells per well on 24-well insert plates, 0.4 m pore size, and
grown for 15 days in DMEM supplemented with 10% FCS, 1% GlutaMAX
(100x, GIBCO), 100 U/mL penicillin, 100 g/mL streptomycin (GIBCO)
and 1% nonessential amino acids (100x). Cells were maintained at 37 °C
in a humidified 5% CO2 atmosphere. The medium was changed every 2–
3 d. Before running the permeability assay, the culture medium was
replaced with an FCS-free HEPES/carbonate transport buffer pH 7.2. For
assessment of monolayer integrity, the transepithelial electrical
resistance was measured. Test compounds were predissolved in DMSO
and added either to the apical or basolateral compartment at a final
concentration of 2 M. Before and after incubation for 2 h at 37 °C,
samples were taken from both compartments. LC-MS/MS analysis of
compound content was undertaken after precipitation with MeOH.
Permeability (Papp) was calculated in the apical to basolateral (A–B) and
basolateral to apical (B–A) directions. The efflux ratio basolateral (B) to
apical (A) was calculated by dividing Papp B–A by Papp A–B.
Equilibrium shake flask solubility assay
The thermodynamic solubility of compounds in water was determined by
an equilibrium shake flask method.[24] A saturated solution of the test
compound was prepared and the solution was mixed for 24 h to ensure
that equilibrium was reached. The solution was centrifuged to remove the
insoluble fraction and the concentration of the compound in solution was
determined using a standard calibration curve. To prepare the test
sample, solid compound (2 mg) was weighed in a 4-mL glass vial.
Phosphate buffer pH 6.5 (1 mL) was added and the suspension was
stirred for 24 h at RT. Then, the solution was centrifuged. To prepare the
sample for the standard calibration, solid sample (2 mg) was dissolved in
MeCN (30 mL). After sonication, the solution was diluted with water to 50
mL. Sample and standard were quantified by HPLC with UV detection.
For each test sample, two injection volumes (5 and 50 L) in triplicate
were made. Three injection volumes (5 L, 10 L and 20 L) were made
for the standard. HPLC conditions: column: Xterra MS C18 2.5 m, 4.6 ×
30 mm; injection volume: sample: 3 × 5 L and 3 × 50 L, standard: 5 L,
10 L and 20 L; flow: 1.5 mL/min; mobile phase: acidic gradient: eluent
A: water/0.01% TFA, eluent B: MeCN/0.01% TFA, 0 min 95% A 5% B; 0–
3 min 35% A 65% B linear gradient, 3–5 min 35% A 65% B isocratic, 5–6
min 95% A 5% B isocratic; UV detection: wavelength near the absorption
maximum (between 200 and 400 nm). The areas of sample and standard
injections, as well as the calculation of the solubility values (in mg/L),
were determined using Waters Empower 2 FR software.
For rat PK studies, test compounds were administered to male Wistar
rats intravenously at low doses of 0.3 to 0.5 mg/kg and intragastrically at
doses of 0.6 to 1 mg/kg formulated as solutions using solubilizers such
as PEG 400 in well-tolerated amounts. Animals were catheterized at the
jugular vein and samples were taken via the catheter. Plasma samples
were collected after 2 min (only intravenously), 8 min, 15 min, 30 min, 45
min, 1 h, 2 h, 4 h, 6 h, 8 h and 24 h post-application, and precipitated
with ice-cold MeCN (1:5). Supernatants were analyzed by LC-MS/MS.
Pharmacokinetic parameters were based on the plasma concentration–
time data and calculated (e.g., using the linear-log trapezoidal rule for
AUC estimation) with KinEx, an Excel-based program.
Pharmacokinetic studies
For the metabolic stability assay in rat hepatocytes, liver cells were
distributed in Williams’ Medium E containing 5% FCS to glass vials at a
density of 1.0 × 106 vital cells/mL. The test compound was added at a
final concentration of 1 M. During incubation, the hepatocyte
suspensions were continuously shaken at 580 rpm and aliquots were
removed at 2, 8, 16, 30, 45 and 90 min, to which an equal volume of cold
MeCN was immediately added. Samples were frozen at –20 °C overnight,
then centrifuged for 15 min at 3000 rpm. The supernatants were
analyzed by liquid chromatography–tandem mass spectrometry (LC-
MS/MS) using an Agilent 1200 HPLC system with an Ascentis Express
column and water/MeCN (isocratic) as eluent. The half-life of a test
compound was determined from the concentration–time plot and the
intrinsic clearances were calculated. Together with the additional
parameters liver blood flow and amount of liver cells in vivo and in vitro,
and application of the ‘well-stirred’ liver model,[25] the hepatic in vivo
MOLM-13 and MV4-11 human AML tumor models in mice and rats
All animal experiments were conducted in accordance with the German
Animal Welfare Law and approved by local authorities. For the acute
myeloid leukemia (AML) mouse model, 2 × 106 MOLM-13 human AML
cells were suspended in 100% MatrigelTM Basement Membrane Matrix
(BD Biosciences) and inoculated subcutaneously to the left flank of
female NMRI nu/nu mice (18–21 g, 5–6 weeks, Taconic M&B). For the
AML model in rats, 2 × 106 MV4-11 human AML cells were suspended in
100% MatrigelTM and inoculated subcutaneously to the left flank of
female athymic nude rats (160–200 g, 5–6 weeks, Harlan). Animals were
stratified into treatment and control groups (n = 8–13/group for mice,
n = 12/group for rats) based on primary tumor size. Treatments were
9
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
started 3–13 days after tumor cell inoculation when the average tumor
sizes were 23–38 mm2 and 43 mm2 for mice and rats, respectively.
ppm, multiplicity (s = singlet, d = doublet, t = triplet, q = quartet, br =
broad, m = multiplet) and integration. High-resolution mass spectra were
recorded on a Xevo® G2-XS Tof (Waters) instrument. Low-resolution
mass spectra (electrospray ionization) were obtained via HPLC-MS (ESI)
using a Waters Acquity UPLC system equipped with an SQ 3100 Mass
Detector; column: Acquity UPLC BEH C18 1.7 m, 50 × 2.1 mm; eluent
A: H2O + 0.05% formic acid (99%), eluent B: MeCN + 0.05% formic acid
(99%); gradient: 0–0.5 min 5% B, 0.5–2.5 min 5–100% B, 2.5–4.5 min
100% B; total run time: 5 min; flow: 0.5 mL/min. Melting points were
determined with a Büchi B-540 melting point apparatus. Optical rotations
were recorded on a JASCO P-2000 polarimeter. The purity of all target
compounds was >97%, as determined by 1H NMR spectroscopy.
BAY 1143572 was administered either using a once daily (QD) or an
intermittent
3 days on/2 days off treatment schedule. BAY-958
hydrochloride and compound 4 were administered QD. PEG 400/water
80:10 was used as the vehicle control. Unless otherwise indicated, all
treatments were administered orally (p.o.) and were continued until the
end of the experiment. Body weight and tumor areas (longest diameter
multiplied by its perpendicular) measured by caliper were determined at
least twice weekly. Treatment-to-control (T/C) ratios were calculated by
dividing the mean tumor area of the treatment group by the mean tumor
area of the vehicle group at the time point when the vehicle group was
sacrificed.
3-[(4-Chloro-1,3,5-triazin-2-yl)amino]benzenemethanesulfonamide
All statistical analyses were performed using statistical software R
(version 3.3.2) by comparing the treatment groups to the vehicle group at
the time point when the vehicle group was sacrificed. Statistical analysis
of the log-transformed tumor area data was performed using one-way
ANOVA followed by Dunnett’s contrasts or Kruskal–Wallis test followed
by Dunn’s test with Holm–Bonferroni correction. In the MV4-11 model
with only two groups, the analysis was performed using Student’s t test.
In all cases, p <0.05 was considered as being statistically significant.
To a solution of 2,4-dichloro-1,3,5-triazine (1; 900 mg, 6.0 mmol) in
anhyd THF/iPrOH (1:1, 8 mL) at –20 °C under N2 atmosphere was added
a cooled solution (–20 °C) of (3-aminophenyl)methanesulfonamide (1117
mg, 6.0 mmol) and DIPEA (2.07 mL, 12 mmol) in anhyd THF/iPrOH (1:1,
8 mL). The reaction mixture was stirred for 2 h at –10 °C. The mixture
was concentrated under reduced pressure and the crude product was
dried in vacuo for 15 h. The white solid was used in the next step without
further purification: 1H NMR (400 MHz, [D6]DMSO):  = 4.26 (s, 2H), 6.89
(s, 2H), 7.15 (d, 1H), 7.36 (d, 1H), 7.66 (d, 1H), 8.64 (s, 1H), 9.75 (br s,
1H), 10.83 ppm (s, 1H); MS (ESI) m/z: 300 [M + H]+.
Docking
Docking calculations were performed using Glide[26] in standard precision
mode and default settings. The X-ray complex of CDK9/CycT1 with a
benzimidazole inhibitor (PDB code 3MY1) was used.[27] Prior to the
docking procedure, all water molecules and ligands in the protein–ligand
complex were removed. Ligands were pre-processed using LigPrep.[28]
Docking poses were selected based on Glide scoring.
1-(3-{[4-(4-Fluoro-2-methoxyphenyl)-1,3,5-triazin-2-yl]amino}phenyl)-
methanesulfonamide (BAY-958)
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (1767 mg), 4-fluoro-2-methoxyphenylboronic acid
(1504 mg, 8.85 mmol) and K3PO4 (2505 mg, 11.82 mmol) in
dioxane/water (10:1, 66 mL) was degassed with a stream of nitrogen for
15 min. Pd(dppf)Cl2·DCM (489 mg, 0.6 mmol) was added and the
reaction mixture was heated for 90 min at 145 °C in a microwave oven.
The mixture was diluted with EtOAc and washed with sat. aq NaHCO3.
The organic phase was dried (Na2SO4), filtered and concentrated. The
crude was purified by flash chromatography (EtOAc/MeOH, 100:0 to 5:1).
Finally, precipitation from EtOAc yielded BAY-958 as a white solid (129
mg, 0.33 mmol): mp: 234 °C; 1H NMR (400 MHz, [D6]DMSO):  = 3.87 (s,
3H), 4.22 (s, 2H), 6.82–6.91 (m, 3H), 7.07 (m, 2H), 7.34 (t, 1H), 7.72 (s,
1H), 7.89 (br s, 2H), 8.78 (s, 1H), 10.32 ppm (s, 1H); ESI-HRMS: m/z
[M + H]+ calcd for C17H17FN5O3S: 390.1036, found: 390.1039.
Synthetic procedures
General methods and materials
Commercially available reagents and anhydrous solvents were used as
supplied, without further purification. All air- and moisture-sensitive
reactions were carried out in oven-dried (at 120 °C) glassware under an
inert atmosphere of argon. A Biotage® Initiator Classic microwave reactor
was used for reactions conducted in a microwave oven. Reactions were
monitored by TLC and UPLC analysis with a Waters Acquity UPLC MS
Single Quad system; column: Acquity UPLC BEH C18 1.7 m,
50 × 2.1 mm; basic conditions: eluent A: H2O + 0.2 vol% aq NH3 (32%),
eluent B: MeCN; gradient: 0–1.6 min 1–99% B, 1.6–2.0 min 99% B; flow:
0.8 mL/min; acidic conditions: eluent A: H2O + 0.1 vol% formic acid
(99%), eluent B: MeCN; gradient: 0–1.6 min 1–99% B, 1.6–2.0 min 99%
B; flow: 0.8 mL/min; temperature: 60 °C; DAD scan: 210–400 nm.
Analytical TLC was carried out on aluminum-backed plates coated with
Merck Kieselgel 60 F254, with visualization under UV light at 254 nm.
Flash chromatography was carried out using a Biotage® Isolera™ One
system with 200–400 nm variable detector. Preparative HPLC was
carried out with a Waters AutoPurification MS Single Quad system;
column: Waters XBridge C18 5 m, 100 × 30 mm; basic conditions:
eluent A: H2O + 0.2 vol% aq NH3 (32%), eluent B: MeCN; gradient: 0–0.5
min 5% B, flow: 25 mL/min; 0.51–5.50 min 10–100% B, flow: 70 mL/min;
5.51–6.5 min 100% B, flow: 70 mL/min; acidic conditions: eluent A:
H2O + 0.1 vol% formic acid (99%), eluent B: MeCN; gradient: 0–0.5 min
5% B, flow: 25 mL/min; 0.51–5.50 min 10–100% B, flow: 70 mL/min;
5.51–6.5 min 100% B, flow: 70 mL/min; temperature: 25 °C; DAD scan:
210–400 nm. NMR spectra were recorded at ambient temperature
(22 ± 1 °C), unless otherwise noted, on Bruker Avance III HD
spectrometers. 1H NMR spectra were obtained at 300, 400, 500 or 600
MHz, and referenced to the residual solvent signal (7.26 ppm for CDCl3,
2.50 ppm for [D6]DMSO). 13C NMR spectra were obtained at 125 MHz
and also referenced to the residual solvent signal (39.52 ppm for
[D6]DMSO). 1H NMR data are reported as follows: chemical shift () in
1-(3-{[4-(4-Fluoro-2-methoxyphenyl)-1,3,5-triazin-2-yl]amino}phenyl)-
methanesulfonamide hydrochloride (BAY-958·HCl)
BAY-958 (914 mg, 2.35 mmol) was suspended in aq 1 N HCl (2.35 mL)
and the mixture was stirred for 4 h at RT, then concentrated under
reduced pressure to give the desired hydrochloride as a white solid (980
mg, 2.30 mmol, 98%): mp: 180 °C; 1H NMR (400 MHz, [D6]DMSO):  =
3.90 (s, 3H), 4.23 (s, 2H), 6.91 (m, 3H), 7.13 (m, 2H), 7.37 (m, 1H), 7.81
(m, 3H), 8.82 (s, 1H), 10.83 ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd
for C17H17FN5O3S: 390.1036, found: 390.1038.
1-[(Methylsulfanyl)methyl]-3-nitrobenzene
Sodium methanethiolate (13.5 g, 192 mmol) was added in two portions to
a stirred solution of 1-(chloromethyl)-3-nitrobenzene (30.0 g, 175 mmol)
in EtOH (360 mL) at –15 °C. The cold bath was removed and the mixture
was stirred at RT for 3 h. Then, it was diluted with brine and extracted
with EtOAc (2 ×). The combined organic phases were washed with water,
dried (Na2SO4), filtered and concentrated to give the desired product
(32.2 g) that was used without further purification: 1H NMR (400 MHz,
CDCl3):  = 2.01 (s, 3H), 3.75 (s, 2H), 7.50 (m, 1H), 7.66 (m, 1H), 8.11 (m,
1H), 8.18 ppm (m, 1H).
10
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
1-[(Methylsulfonyl)methyl]-3-nitrobenzene
A mixture of crude 4-chloro-N-[3-(methylsulfonyl)phenyl]-1,3,5-triazin-2-
amine (450 mg), 4-fluoro-2-methoxyphenylboronic acid (403 mg, 2.37
mmol) and Pd(PPh3)4 (274 mg, 0.24 mmol) in 1,2-dimethoxyethane (7.3
mL) and aq 2 M K2CO3 (1.6 mL) was degassed using argon. The mixture
was stirred under argon for 90 min at 100 °C. After cooling, the mixture
was diluted with EtOAc and washed with brine. The organic phase was
filtered using a Whatman filter and concentrated. The residue was
purified by preparative HPLC (acidic conditions) to give 5 as a white solid
(152 mg, 0.41 mmol): 1H NMR (400 MHz, [D6]DMSO):  = 3.20 (s, 3H),
3.88 (br s, 3H), 6.91 (td, 1H), 7.12 (dd, 1H), 7.58–7.68 (m, 2H), 7.91 (br s,
1H), 8.24 (br s, 1H), 8.42 (s, 1H), 8.88 (s, 1H), 10.68 ppm (br s, 1H); ESI-
HRMS: m/z [M + H]+ calcd for C17H16FN4O3S: 375.0927, found: 375.0924.
3-Chloroperoxybenzoic acid (77%; 26.9 g, 120 mmol) was added to a
stirred solution of 1-[(methylsulfanyl)methyl]-3-nitrobenzene (10.0 g) in
DCM (1305 mL) at 0 °C. The mixture was stirred at 0 °C for 30 min and
then at RT for 2.5 h. Then, it was diluted with water (300 mL), and
NaHCO3 (11.0 g) was added. The mixture was extracted with DCM (2 ×).
The combined organic phases were filtered using a Whatman filter and
concentrated. The residue was purified by flash chromatography
(DCM/EtOH, 95:5) and finally recrystallized from EtOAc to give the
desired product (6.2 g, 28.9 mmol): 1H NMR (400 MHz, [D6]DMSO):  =
2.93 (s, 3H), 4.68 (s, 2H), 7.69 (m, 1H), 7.83 (m, 1H), 8.22 (m, 1H), 8.28
ppm (m, 1H).
4-Chloro-N-{3-[2-(methylsulfonyl)ethyl]phenyl}-1,3,5-triazin-2-amine
3-[(Methylsulfonyl)methyl]aniline
DIPEA (0.35 mL, 2.01 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 150 mg, 1.00 mmol) in THF/iPrOH (1:1, 2 mL)
at –40 °C. Then, a solution of 3-[2-(methylsulfonyl)ethyl]aniline (200 mg,
1.00 mmol) in THF/iPrOH (1:1, 1 mL) was added at this temperature.
Under stirring, the temperature of the reaction mixture was slowly raised
over 2 h to 0 °C. The mixture was concentrated in vacuo to give the
crude product (458 mg) that was used without further purification.
TiCl3 solution (ca. 15% in ca. 10% HCl, 162 mL) was added to a stirred
solution of 1-[(methylsulfonyl)methyl]-3-nitrobenzene (5.1 g, 23.8 mmol)
in THF (250 mL) at RT, and the mixture was stirred for 16 h. The pH was
increased to 10 by the addition of 1 N NaOH, then the reaction mixture
was extracted with EtOAc (2 ×). The combined organic phases were
washed with brine, filtered using a Whatman filter and concentrated to
give the desired product (4.5 g) that was used without further purification:
1H NMR (400 MHz, [D6]DMSO):  = 2.83 (s, 3H), 4.23 (s, 2H), 5.13 (br,
2H), 6.51 (m, 3H), 6.97 ppm (m, 1H).
4-(4-Fluoro-2-methoxyphenyl)-N-{3-[2-(methylsulfonyl)ethyl]phenyl}-
1,3,5-triazin-2-amine (6)
4-Chloro-N-{3-[(methylsulfonyl)methyl]phenyl}-1,3,5-triazin-2-amine
A mixture of crude 4-chloro-N-{3-[2-(methylsulfonyl)ethyl]phenyl}-1,3,5-
triazin-2-amine (150 mg), 4-fluoro-2-methoxyphenylboronic acid (122 mg,
0.72 mmol) and Pd(PPh3)4 (83 mg, 0.07 mmol) in 1,2-dimethoxyethane
(1.5 mL) and aq 2 M K2CO3 (0.5 mL) was degassed using argon. The
mixture was stirred under argon for 90 min at 100 °C. After cooling, the
mixture was diluted with EtOAc and washed with brine. The organic
phase was filtered using a Whatman filter and concentrated. The residue
was purified by preparative HPLC (acidic conditions) to give 6 as a light
beige solid (53 mg, 0.13 mmol): 1H NMR (400 MHz, CDCl3):  = 2.85 (s,
3H), 3.14–3.26 (m, 2H), 3.26–3.39 (m, 2H), 3.94 (s, 3H), 6.74–6.83 (m,
2H), 7.00 (br d, 1H), 7.29–7.39 (m, 1H), 7.44–7.60 (m, 2H), 7.65 (s, 1H),
7.95 (br s, 1H), 8.82 ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd for
C19H20FN4O3S: 403.1240, found: 403.1244.
DIPEA (3.7 mL, 21.3 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 1.60 g, 10.7 mmol) in THF/iPrOH (1:1, 20 mL)
at –40 °C. Then, a suspension of 3-[(methylsulfonyl)methyl]aniline (1.97
g, 10.7 mmol) in THF/iPrOH (1:1, 10 mL) was added at this temperature.
Under stirring, the temperature of the reaction mixture was slowly raised
over 3 h to 0 °C. The mixture was concentrated in vacuo to give the
crude product (5.2 g) that was used without further purification.
4-(4-Fluoro-2-methoxyphenyl)-N-{3-[(methylsulfonyl)methyl]phenyl}-
1,3,5-triazin-2-amine (4)
A mixture of crude 4-chloro-N-{3-[(methylsulfonyl)methyl]phenyl}-1,3,5-
triazin-2-amine (1000 mg), 4-fluoro-2-methoxyphenylboronic acid (569
mg, 3.35 mmol) and tetrakis(triphenylphosphine)palladium(0) [Pd(PPh3)4;
580 mg, 0.50 mmol] in 1,2-dimethoxyethane (10.3 mL) and aq 2 M K2CO3
(3.4 mL) was degassed using argon. The mixture was stirred under
argon for 90 min at 100 °C. After cooling, the mixture was diluted with
EtOAc and washed with brine. The organic phase was filtered using a
Whatman filter and concentrated. The residue was purified by flash
chromatography (DCM to DCM/EtOH, 95:5) to give 4 as a white solid
(402 mg, 1.03 mmol): 1H NMR (400 MHz, CDCl3):  = 2.79 (s, 3H), 3.94
(s, 3H), 4.26 (s, 2H), 6.75–6.81 (m, 2H), 7.16 (d, 1H), 7.38–7.46 (m, 2H),
7.69–7.87 (m, 2H), 7.96 (br s, 1H), 8.82 ppm (s, 1H); ESI-HRMS: m/z
[M + H]+ calcd for C18H18FN4O3S: 389.1084, found: 389.1080.
4-Chloro-N-(1,1-dioxido-2,3-dihydro-1-benzothien-6-yl)-1,3,5-triazin-
2-amine
DIPEA (0.88 mL, 5.07 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 400 mg, 2.53 mmol) in THF/iPrOH (1:1, 5 mL)
at –40 °C. Then, 2,3-dihydro-1-benzothiophen-6-amine 1,1-dioxide (464
mg, 2.53 mmol) was added at this temperature. Under stirring, the
temperature of the reaction mixture was slowly raised over 2 h to 0 °C.
The mixture was concentrated in vacuo to give the crude product (1432
mg) that was used without further purification.
N-(1,1-Dioxido-2,3-dihydro-1-benzothien-6-yl)-4-(4-fluoro-2-methoxy-
phenyl)-1,3,5-triazin-2-amine (7)
4-Chloro-N-[3-(methylsulfonyl)phenyl]-1,3,5-triazin-2-amine
A mixture of crude 4-chloro-N-(1,1-dioxido-2,3-dihydro-1-benzothien-6-
yl)-1,3,5-triazin-2-amine (200 mg), 4-fluoro-2-methoxyphenylboronic acid
(115 mg, 0.67 mmol) and Pd(PPh3)4 (117 mg, 0.10 mmol) in 1,2-
dimethoxyethane (2.1 mL) and aq 2 M K2CO3 (0.7 mL) was degassed
using argon. The mixture was stirred under argon for 2 h at 100 °C. After
cooling, the mixture was diluted with EtOAc and washed with brine. The
organic phase was filtered using a Whatman filter and concentrated. The
residue was purified by preparative HPLC (acidic conditions) to give 7 as
a white solid (40 mg, 0.10 mmol): 1H NMR (400 MHz, CDCl3):  = 3.32–
3.42 (m, 2H), 3.46–3.60 (m, 2H), 4.00 (s, 3H), 6.74–6.83 (m, 2H), 7.28–
7.38 (m, 1H), 7.45–7.69 (m, 2H), 8.01 (br s, 1H), 8.57 (br s, 1H), 8.84
ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd for C18H16FN4O3S: 387.0927,
found: 387.0929.
DIPEA (0.31 mL, 1.75 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 138 mg, 0.88 mmol) in THF/iPrOH (1:1, 1.8 mL)
at –40 °C. Then, a suspension of 3-(methylsulfonyl)aniline (150 mg, 0.88
mmol) in THF/iPrOH (1:1, 0.9 mL) was added at this temperature. Under
stirring, the temperature of the reaction mixture was slowly raised over 90
min to 0 °C. The mixture was concentrated in vacuo to give the crude
product (450 mg) that was used without further purification.
4-(4-Fluoro-2-methoxyphenyl)-N-[3-(methylsulfonyl)phenyl]-1,3,5-
triazin-2-amine (5)
11
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
4-Chloro-N-(2,2-dioxido-1,3-dihydro-2-benzothien-5-yl)-1,3,5-triazin-
2-amine
The residue was purified by flash chromatography (hexane to
hexane/EtOAc, 1:1) to give 1-[2-(methylsulfonyl)propan-2-yl]-3-
nitrobenzene (139 mg, 0.57 mmol, 25%): 1H NMR (400 MHz, CDCl3):  =
1.93 (s, 6H), 2.62 (s, 3H), 7.63 (t, 1H), 8.06 (dt, 1H), 8.26 (d, 1H), 8.47
ppm (s, 1H), and (rac)-1-[1-(methylsulfonyl)ethyl]-3-nitrobenzene (84 mg,
0.37 mmol, 16%): 1H NMR (400 MHz, CDCl3):  = 1.87 (d, 3H), 2.76 (s,
3H), 4.31 (q, 1H), 7.63 (t, 1H), 7.85 (d, 1H), 8.25–8.32 ppm (m, 2H).
DIPEA (0.33 mL, 1.90 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 150 mg, 0.95 mmol) in THF/iPrOH (1:1, 2 mL)
at –40 °C. Then, 1,3-dihydro-2-benzothiophen-5-amine 2,2-dioxide (174
mg, 0.95 mmol) was added at this temperature. Under stirring, the
temperature of the reaction mixture was slowly raised over 2 h to 0 °C.
The mixture was concentrated in vacuo to give the crude product (462
mg) that was used without further purification.
(rac)-3-[1-(Methylsulfonyl)ethyl]aniline
TiCl3 solution (ca. 15% in ca. 10% HCl, 2.4 mL) was added to a stirred
solution of (rac)-1-[1-(methylsulfonyl)ethyl]-3-nitrobenzene (80 mg, 0.35
mmol) in THF (3.7 mL) at RT, and the mixture was stirred for 16 h. The
pH was increased to 10 by the addition of 2 N NaOH, then the reaction
mixture was extracted with EtOAc (2 ×). The combined organic phases
were washed with brine, dried (Na2SO4), filtered and concentrated to give
the crude product (88 mg) that was used without further purification.
N-(2,2-Dioxido-1,3-dihydro-2-benzothien-5-yl)-4-(4-fluoro-2-methoxy-
phenyl)-1,3,5-triazin-2-amine (8)
A mixture of crude 4-chloro-N-(2,2-dioxido-1,3-dihydro-2-benzothien-5-
yl)-1,3,5-triazin-2-amine (75 mg), 4-fluoro-2-methoxyphenylboronic acid
(64 mg, 0.38 mmol) and K3PO4 (107 mg, 0.51 mmol) in dioxane/water
(20:1, 2.5 mL) was degassed with a stream of argon for 15 min.
Pd(dppf)Cl2·DCM (21 mg, 0.025 mmol) was added and the reaction
mixture was heated for 60 min at 140 °C in a microwave oven. The
mixture was diluted with dioxane, filtered and concentrated. The residue
was purified by preparative HPLC (acidic conditions) to give 8 as a white
solid (9 mg, 0.02 mmol): 1H NMR (400 MHz, CDCl3):  = 3.93 (s, 3H),
4.38 (d, 4H), 6.74–6.83 (m, 2H), 7.28–7.33 (m, 1H), 7.46 (s, 1H), 7.59 (br
d, 1H), 7.86 (s, 1H), 7.90–8.01 (m, 1H), 8.83 ppm (s, 1H); ESI-HRMS:
m/z [M + H]+ calcd for C18H16FN4O3S: 387.0927, found: 387.0926.
(rac)-4-Chloro-N-{3-[1-(methylsulfonyl)ethyl]phenyl}-1,3,5-triazin-2-
amine
DIPEA (0.15 mL, 0.86 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 65 mg, 0.43 mmol) in THF/iPrOH (1:1, 0.8 mL)
at –40 °C. Then,
a solution of crude (rac)-3-[1-(methylsulfonyl)-
ethyl]aniline (86 mg) in THF/iPrOH (1:1, 0.4 mL) was added at this
temperature. Under stirring, the temperature of the reaction mixture was
slowly raised over 2 h to 0 °C. The mixture was concentrated in vacuo to
give the crude product (233 mg) that was used without further purification.
N-{3-[(tert-Butylsulfonyl)methyl]phenyl}-4-chloro-1,3,5-triazin-2-
amine
(rac)-4-(4-Fluoro-2-methoxyphenyl)-N-{3-[1-(methylsulfonyl)ethyl]-
DIPEA (0.46 mL, 2.64 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 198 mg, 1.32 mmol) in THF/iPrOH (1:1, 2.6 mL)
at –40 °C. Then, 3-[(tert-butylsulfonyl)methyl]aniline (300 mg, 1.32 mmol)
was added at this temperature. Under stirring, the temperature of the
reaction mixture was slowly raised over 2 h to 0 °C. The mixture was
concentrated in vacuo to give the crude product (654 mg) that was used
without further purification.
phenyl}-1,3,5-triazin-2-amine (10)
A mixture of crude (rac)-4-chloro-N-{3-[1-(methylsulfonyl)ethyl]phenyl}-
1,3,5-triazin-2-amine (233 mg), 4-fluoro-2-methoxyphenylboronic acid
(127 mg, 0.75 mmol) and Pd(PPh3)4 (129 mg, 0.11 mmol) in 1,2-
dimethoxyethane (2.3 mL) and aq 2 M K2CO3 (0.7 mL) was degassed
using argon. The mixture was stirred under argon for 90 min at 100 °C.
After cooling, the mixture was diluted with EtOAc and THF, and washed
with brine. The organic phase was filtered using a Whatman filter and
concentrated. The residue was purified by preparative HPLC (acidic
conditions) to give 10 as a light beige solid (34 mg, 0.08 mmol): 1H NMR
(400 MHz, CDCl3):  = 1.82 (d, 3H), 2.69 (s, 3H), 3.94 (s, 3H), 4.20 (q,
1H), 6.74–6.82 (m, 2H), 7.20 (br d, 1H), 7.39–7.52 (m, 2H), 7.80 (s, 2H),
7.97 (br s, 1H), 8.82 ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd for
C19H20FN4O3S: 403.1240, found: 403.1245.
N-{3-[(tert-Butylsulfonyl)methyl]phenyl}-4-(4-fluoro-2-methoxy-
phenyl)-1,3,5-triazin-2-amine (9)
A mixture of crude N-{3-[(tert-butylsulfonyl)methyl]phenyl}-4-chloro-1,3,5-
triazin-2-amine (150 mg), 4-fluoro-2-methoxyphenylboronic acid (75 mg,
0.44 mmol) and Pd(PPh3)4 (76 mg, 0.07 mmol) in 1,2-dimethoxyethane
(1.4 mL) and aq 2 M K2CO3 (0.4 mL) was degassed using argon. The
mixture was stirred under argon for 90 min at 100 °C. After cooling, the
mixture was diluted with EtOAc and THF, and washed with brine. The
organic phase was filtered using a Whatman filter and concentrated. The
residue was purified by preparative HPLC (acidic conditions) to give 9 as
a light beige solid (30 mg, 0.07 mmol): 1H NMR (400 MHz, CDCl3):  =
1.46 (s, 9H), 3.93 (s, 3H), 4.21 (s, 2H), 6.74–6.82 (m, 2H), 7.18–7.25 (m,
1H), 7.36–7.59 (m, 2H), 7.72 (s, 1H), 7.82 (br s, 1H), 7.99 (br s, 1H), 8.81
ppm (br s, 1H); ESI-HRMS: m/z [M + H]+ calcd for C21H24FN4O3S:
431.1553, found: 431.1552.
3-[2-(Methylsulfonyl)propan-2-yl]aniline
TiCl3 solution (ca. 15% in ca. 10% HCl, 3.8 mL) was added to a stirred
solution of 1-[2-(methylsulfonyl)propan-2-yl]-3-nitrobenzene (135 mg,
0.56 mmol) in THF (5.8 mL) at RT, and the mixture was stirred for 16 h.
The pH was increased to 10 by the addition of 2 N NaOH, then the
reaction mixture was extracted with EtOAc (2 ×). The combined organic
phases were washed with brine, dried (Na2SO4), filtered and
concentrated to give the crude product (148 mg) that was used without
further purification.
1-[2-(Methylsulfonyl)propan-2-yl]-3-nitrobenzene
(methylsulfonyl)ethyl]-3-nitrobenzene
and
(rac)-1-[1-
4-Chloro-N-{3-[2-(methylsulfonyl)propan-2-yl]phenyl}-1,3,5-triazin-2-
Under argon, 1 M sodium bis(trimethylsilyl)amide in THF (4.65 mL, 4.65
mmol) was added dropwise to stirred solution of 1-
amine
a
[(methylsulfonyl)methyl]-3-nitrobenzene (500 mg, 2.32 mmol) in THF (25
mL) at –78 °C. The reaction mixture was stirred for 30 min at this
temperature before iodomethane (725 mg, 5.11 mmol) was added. The
reaction mixture was stirred for 1 h at –78 °C, then the cold bath was
removed and the mixture was slowly warmed to RT. Aq NH4Cl solution
was added and the mixture was extracted with EtOAc (3 ×). The
combined organic phases were dried (Na2SO4), filtered and concentrated.
DIPEA (0.24 mL, 1.37 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 103 mg, 0.68 mmol) in THF/iPrOH (1:1, 1.3 mL)
at –40 °C. Then, a solution of crude 3-[2-(methylsulfonyl)propan-2-
yl]aniline (146 mg) in THF/iPrOH (1:1, 0.6 mL) was added at this
temperature. Under stirring, the temperature of the reaction mixture was
slowly raised over 2 h to 0 °C. The mixture was concentrated in vacuo to
give the crude product (354 mg) that was used without further purification.
12
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
4-(4-Fluoro-2-methoxyphenyl)-N-{3-[2-(methylsulfonyl)propan-2-yl]-
phenyl}-1,3,5-triazin-2-amine (11)
using argon. The mixture was stirred under argon for 90 min at 100 °C.
After cooling, the mixture was diluted with EtOAc and THF, and washed
with brine. The organic phase was filtered using a Whatman filter and
concentrated. The residue was purified by preparative HPLC (acidic
conditions) to give 12 as a light beige solid (5 mg, 0.01 mmol): 1H NMR
(400 MHz, CDCl3):  = 2.96 (s, 3H), 3.93 (s, 3H), 6.05 (d, 1H), 6.73–6.83
(m, 2H), 7.28–7.37 (m, 1H), 7.42–7.58 (m, 2H), 7.85 (br s, 1H), 7.94 (s,
2H), 8.83 ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd for
C18H17F2N4O3S: 407.0989, found: 407.0990.
A mixture of crude 4-chloro-N-{3-[2-(methylsulfonyl)propan-2-yl]phenyl}-
1,3,5-triazin-2-amine (125 mg), 4-fluoro-2-methoxyphenylboronic acid (65
mg, 0.38 mmol) and Pd(PPh3)4 (66 mg, 0.06 mmol) in 1,2-
dimethoxyethane (1.2 mL) and aq 2 M K2CO3 (0.4 mL) was degassed
using argon. The mixture was stirred under argon for 90 min at 100 °C.
After cooling, the mixture was diluted with EtOAc and THF, and washed
with brine. The organic phase was filtered using a Whatman filter and
concentrated. The residue was purified by preparative HPLC (acidic
conditions) to give 11 as a light beige solid (29 mg, 0.07 mmol): 1H NMR
(400 MHz, CDCl3):  = 1.87 (s, 6H), 2.56 (s, 3H), 3.93 (s, 3H), 6.74–6.81
(m, 2H), 7.28–7.45 (m, 3H), 7.80 (br s, 1H), 7.95 (br s, 2H), 8.82 ppm (s,
1H); ESI-HRMS: m/z [M + H]+ calcd for C20H22FN4O3S: 417.1397, found:
417.1396.
3-[Difluoro(methylsulfonyl)methyl]aniline
TiCl3 solution (ca. 15% in ca. 10% HCl, 0.4 mL) was added to a stirred
solution of 1-[difluoro(methylsulfonyl)methyl]-3-nitrobenzene (16 mg, 0.06
mmol) in THF (0.7 mL) at RT, and the mixture was stirred for 16 h. The
pH was increased to 9–10 by the addition of an aq NaHCO3 solution,
then the reaction mixture was extracted with EtOAc/THF (1:1, 3 ×). The
combined organic phases were washed with brine, dried (Na2SO4),
filtered and concentrated to give the crude product (18 mg) that was used
without further purification.
1-[Difluoro(methylsulfonyl)methyl]-3-nitrobenzene
[fluoro(methylsulfonyl)methyl]-3-nitrobenzene
and
(rac)-1-
Under argon, 1 M sodium bis(trimethylsilyl)amide in THF (10.22 mL,
10.22 mmol) was added dropwise to stirred solution of 1-
a
4-Chloro-N-{3-[difluoro(methylsulfonyl)methyl]phenyl}-1,3,5-triazin-
[(methylsulfonyl)methyl]-3-nitrobenzene (1000 mg, 4.65 mmol) in THF
(50 mL) at –78 °C. The reaction mixture was stirred for 30 min at this
temperature before N-fluorobenzenesulfonimide (3662 mg, 11.61 mmol)
was added. The reaction mixture was stirred for 5 h at –78 °C. Aq NH4Cl
solution was added and the cold bath was removed. At RT, the mixture
was extracted with EtOAc (3 ×). The combined organic phases were
washed with aq NaHCO3 solution, dried (Na2SO4), filtered and
concentrated. The residue was purified by preparative HPLC (acidic
conditions) to give 1-[difluoro(methylsulfonyl)methyl]-3-nitrobenzene (173
mg, 0.69 mmol, 15%): 1H NMR (600 MHz, CDCl3):  = 3.20 (s, 3H), 7.76
(t, 1H), 8.01–8.05 (m, 1H), 8.49 (dd, 1H), 8.56 ppm (s, 1H), and (rac)-1-
[fluoro(methylsulfonyl)methyl]-3-nitrobenzene (235 mg, 1.01 mmol, 22%):
1H NMR (600 MHz, CDCl3):  = 3.07 (d, 3H), 6.16 (d, 1H), 7.70 (t, 1H),
7.91 (dd, 1H), 8.37–8.41 (m, 1H), 8.43–8.45 ppm (m, 1H).
2-amine
DIPEA (0.03 mL, 0.16 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 12 mg, 0.08 mmol) in THF/iPrOH (1:1, 0.2 mL)
at –40 °C. Then, a solution of crude 3-[difluoro(methylsulfonyl)methyl]-
aniline (17 mg) in THF/iPrOH (1:1, 0.2 mL) was added at this
temperature. Under stirring, the temperature of the reaction mixture was
slowly raised over 90 min to 0 °C. The mixture was concentrated in
vacuo to give the crude product (40 mg) that was used without further
purification.
N-{3-[Difluoro(methylsulfonyl)methyl]phenyl}-4-(4-fluoro-2-methoxy-
phenyl)-1,3,5-triazin-2-amine (13)
A mixture of crude 4-chloro-N-{3-[difluoro(methylsulfonyl)methyl]phenyl}-
1,3,5-triazin-2-amine (40 mg), 4-fluoro-2-methoxyphenylboronic acid (20
mg, 0.12 mmol) and Pd(PPh3)4 (21 mg, 0.02 mmol) in 1,2-
dimethoxyethane (0.4 mL) and aq 2 M K2CO3 (0.1 mL) was degassed
using argon. The mixture was stirred under argon for 90 min at 100 °C.
After cooling, the mixture was diluted with EtOAc and THF, and washed
with brine. The organic phase was filtered using a Whatman filter and
concentrated. The residue was purified by preparative HPLC (acidic
conditions) to give 13 as a white solid (6 mg, 0.01 mmol): 1H NMR
(400 MHz, CDCl3):  = 3.14 (s, 3H), 3.93 (s, 3H), 6.74–6.81 (m, 2H),
7.43–7.56 (m, 3H), 7.95–8.09 (m, 3H), 8.84 ppm (s, 1H); ESI-HRMS: m/z
[M + H]+ calcd for C18H16F3N4O3S: 425.0895, found: 425.0899.
(rac)-3-[Fluoro(methylsulfonyl)methyl]aniline
TiCl3 solution (ca. 15% in ca. 10% HCl, 0.4 mL) was added to a stirred
solution of (rac)-1-[fluoro(methylsulfonyl)methyl]-3-nitrobenzene (14 mg,
0.06 mmol) in THF (0.6 mL) at RT, and the mixture was stirred for 16 h.
The pH was increased to 9–10 by the addition of an aq NaHCO3 solution,
then the reaction mixture was extracted with EtOAc/THF (1:1, 3 ×). The
combined organic phases were washed with brine, dried (Na2SO4),
filtered and concentrated to give the crude product (20 mg) that was used
without further purification.
(rac)-4-Chloro-N-{3-[fluoro(methylsulfonyl)methyl]phenyl}-1,3,5-
triazin-2-amine
1-(3-{[4-(2-Methoxyphenyl)-1,3,5-triazin-2-yl]amino}phenyl)-
methanesulfonamide (14)
DIPEA (0.03 mL, 0.16 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 12 mg, 0.08 mmol) in THF/iPrOH (1:1, 0.2 mL)
at –40 °C. Then, a solution of crude (rac)-3-[fluoro(methylsulfonyl)-
methyl]aniline (16 mg) in THF/iPrOH (1:1, 0.2 mL) was added at this
temperature. Under stirring, the temperature of the reaction mixture was
slowly raised over 90 min to 0 °C. The mixture was concentrated in
vacuo to give the crude product (38 mg) that was used without further
purification.
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (103 mg), 2-methoxyphenylboronic acid (68 mg,
0.45 mmol) and K3PO4 (218 mg, 1.03 mmol) in dioxane/water (10:1, 8
mL) was degassed with
a
stream of nitrogen for 15 min.
Pd(dppf)Cl2·DCM (25 mg, 0.03 mmol) was added and the reaction
mixture was heated for 90 min at 145 °C in a microwave oven. The
mixture was diluted with EtOAc and washed with sat. aq NaHCO3. The
organic phase was dried (Na2SO4), filtered and concentrated. The
residue was purified by preparative HPLC (acidic conditions) to give 14
(rac)-4-(4-Fluoro-2-methoxyphenyl)-N-{3-[fluoro(methylsulfonyl)-
methyl]phenyl}-1,3,5-triazin-2-amine (12)
as a
light beige solid (36 mg, 0.10 mmol): 1H NMR (400 MHz,
[D6]DMSO):  = 3.87 (s, 3H), 4.25 (s, 2H), 6.87 (br s, 2H), 7.06–7.13 (m,
2H), 7.21 (d, 1H), 7.37 (t, 1H), 7.55 (dt, 1H), 7.73 (t, 1H), 7.84 (m, 2H),
8.82 (s, 1H), 10.52 ppm (s, 1H); MS (ESI) m/z: 372 [M + H]+.
A
mixture of crude (rac)-4-chloro-N-{3-[fluoro(methylsulfonyl)methyl]-
phenyl}-1,3,5-triazin-2-amine (38 mg), 4-fluoro-2-methoxyphenylboronic
acid (20 mg, 0.12 mmol) and Pd(PPh3)4 (21 mg, 0.02 mmol) in 1,2-
dimethoxyethane (0.4 mL) and aq 2 M K2CO3 (0.1 mL) was degassed
13
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
1-(3-{[4-(4-Chloro-2-methoxyphenyl)-1,3,5-triazin-2-yl]amino}-
phenyl)methanesulfonamide (15)
1-[3-({4-[2-(Trifluoromethoxy)phenyl]-1,3,5-triazin-2-yl}amino)-
phenyl]methanesulfonamide (19)
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (174 mg), 4-chloro-2-methoxyphenylboronic acid
(116 mg, 0.62 mmol) and K3PO4 (217 mg, 1.02 mmol) in dioxane/water
(10:1, 10 mL) was degassed with a stream of nitrogen for 15 min.
Pd(dppf)Cl2·DCM (28 mg, 0.03 mmol) was added and the reaction
mixture was heated for 90 min at 145 °C in a microwave oven. The
mixture was diluted with EtOAc and washed with sat. aq NaHCO3. The
organic phase was dried (Na2SO4), filtered and concentrated. The
residue was purified by preparative HPLC (acidic conditions) to give 15
methanesulfonamide (300 mg), 2-(trifluoromethoxy)phenylboronic acid
(309 mg, 1.50 mmol) and K3PO4 (425 mg, 2.00 mmol) in dioxane/water
(20:1, 10 mL) was degassed with a stream of argon for 15 min.
Pd(dppf)Cl2·DCM (82 mg, 0.10 mmol) was added and the reaction
mixture was heated for 1 h at 140 °C in a microwave oven. The mixture
was diluted with an aq NaCl solution and extracted with EtOAc. The
combined organic phases were dried (Na2SO4), filtered and concentrated.
The residue was purified by preparative HPLC (basic conditions) to give
19 (10 mg, 0.02 mmol): 1H NMR (400 MHz, [D6]DMSO):  = 4.25 (s, 2H),
6.88 (s, 2H), 7.10 (d, 1H), 7.36 (t, 1H), 7.54 (d, 1H), 7.59 (t, 1H), 7.71 (td,
2H), 7.81 (br s, 1H), 8.06 (br s, 1H), 8.88 (s, 1H), 10.51 ppm (s, 1H); ESI-
HRMS: m/z [M + H]+ calcd for C17H15F3N5O3S: 426.0848, found:
426.0844.
as
a
light beige solid (13 mg, 0.03 mmol): 1H NMR (400 MHz,
[D6]DMSO):  = 3.90 (s, 3H), 4.24 (s, 2H), 6.86 (br s, 2H), 7.08 (d, 1H),
7.15 (d, 1H), 7.28 (d, 1H), 7.36 (t, 1H), 7.74 (s, 1H), 7.88 (m, 2H), 8.81 (s,
1H), 10.44 ppm (s, 1H); MS (ESI) m/z: 406 [M + H]+.
1-(3-{[4-(3-Methoxypyridin-4-yl)-1,3,5-triazin-2-yl]amino}phenyl)-
methanesulfonamide (16)
1-[3-({4-[2-(Methoxymethyl)phenyl]-1,3,5-triazin-2-yl}amino)phenyl]-
methanesulfonamide (20)
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (141 mg), 3-methoxypyridine-4-boronic acid (104
mg, 0.68 mmol) and K3PO4 (213 mg, 1.00 mmol) in dioxane/water (10:1,
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (400 mg), 2-(methoxymethyl)phenylboronic acid
(221 mg, 1.33 mmol) and K3PO4 (377 mg, 1.80 mmol) in dioxane/water
(50:1, 9 mL) was degassed with a stream of nitrogen for 15 min.
Pd(dppf)Cl2·DCM (70 mg, 0.09 mmol) was added and the reaction
mixture was heated for 15 h at 140 °C in a sealed tube. The mixture was
poured onto ice and extracted with EtOAc. The organic phase was dried
(Na2SO4), filtered and concentrated. The residue was purified by
preparative TLC (CHCl3/MeOH, 29:1) yielding 20 as a white solid (6 mg,
0.02 mmol): 1H NMR (400 MHz, CDCl3):  = 3.32 (s, 3H), 4.32 (s, 2H),
4.82 (s, 2H), 5.27 (br s, 2H), 7.11 (d, 1H), 7.26 (m, 1H), 7.41 (t, 1H), 7.52
(t, 1H), 7.59–7.76 (m, 4H), 7.93 (br s, 1H), 8.67 ppm (s, 1H); MS (ESI)
m/z: 386 [M + H]+.
10 mL) was degassed with
a stream of nitrogen for 15 min.
Pd(dppf)Cl2·DCM (38 mg, 0.05 mmol) was added and the reaction
mixture was heated for 90 min at 145 °C in a microwave oven. The
mixture was diluted with EtOAc and washed with sat. aq NaHCO3. The
organic phase was dried (Na2SO4), filtered and concentrated. The
residue was purified by preparative HPLC (acidic conditions) to give 16
as a white solid (51 mg, 0.14 mmol): 1H NMR (400 MHz, [D6]DMSO):  =
3.99 (s, 3H), 4.25 (s, 2H), 6.87 (s, 2H), 7.11 (d, 1H), 7.36 (dt, 1H), 7.74 (s,
1H), 7.83 (m, 2H), 8.39 (m, 1H), 8.62 (m, 1H), 8.87 (m, 1H), 10.52 ppm (s,
1H); MS (ESI) m/z: 373 [M + H]+.
1-(3-{[4-(4-Fluoro-2-hydroxyphenyl)-1,3,5-triazin-2-yl]amino}phenyl)-
methanesulfonamide (17)
N-[5-Fluoro-2-(4-{[3-(sulfamoylmethyl)phenyl]amino}-1,3,5-triazin-2-
yl)phenyl]acetamide (21)
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (300 mg), 4-fluoro-2-hydroxyphenylboronic acid
(234 mg, 1.50 mmol) and K3PO4 (425 mg, 2.00 mmol) in dioxane/water
(20:1, 10 mL) was degassed with a stream of argon for 15 min.
Pd(dppf)Cl2·DCM (82 mg, 0.10 mmol) was added and the reaction
mixture was heated for 1 h at 140 °C in a microwave oven. The mixture
was diluted with an aq NaCl solution and extracted with EtOAc. The
combined organic phases were dried (Na2SO4), filtered and concentrated.
The residue was purified by preparative HPLC (basic conditions) to give
17 as a white solid (91 mg, 0.24 mmol): 1H NMR (400 MHz, [D6]DMSO):
 = 4.29 (s, 2H), 6.64 (s, 2H), 6.73 (dd, 1H), 6.75–6.82 (m, 1H), 7.18 (d,
1H), 7.38 (t, 1H), 7.66–7.71 (m, 1H), 7.75 (s, 1H), 8.41 (dd, 1H), 8.78 (s,
1H), 10.41 (br s, 1H), 12.95–13.35 ppm (m, 1H); ESI-HRMS: m/z
[M + H]+ calcd for C16H15FN5O3S: 376.0880, found: 376.0883.
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (75 mg), 2-(acetylamino)-4-fluorophenylboronic
acid (74 mg, 0.38 mmol) and K3PO4 (107 mg, 0.50 mmol) in
dioxane/water (20:1, 2.5 mL) was degassed with a stream of argon for 15
min. Pd(dppf)Cl2·DCM (21 mg, 0.025 mmol) was added and the reaction
mixture was heated for 60 min at 140 °C in a microwave oven. The
mixture was diluted with dioxane, filtered and concentrated. The residue
was purified by preparative HPLC (acidic conditions) to give 21 as a
white solid (15 mg, 0.04 mmol): 1H NMR (500 MHz, [D6]DMSO, 80 °C): 
= 2.08 (br s, 3H), 4.30 (s, 2H), 6.68 (s, 2H), 7.03 (ddd, 1H), 7.18 (d, 1H),
7.40 (t, 1H), 7.67 (br d, 1H), 7.77 (s, 1H), 8.38 (dd, 1H), 8.56 (dd, 1H),
8.87 (s, 1H), 10.30 (s, 1H), 12.33 ppm (br s, 1H); ESI-HRMS: m/z
[M + H]+ calcd for C18H18FN6O3S: 417.1145, found: 417.1146.
1-(3-{[4-(2,4-Difluorophenyl)-1,3,5-triazin-2-yl]amino}phenyl)-
methanesulfonamide (18)
1-[3-({4-[4-Fluoro-2-(trifluoromethyl)phenyl]-1,3,5-triazin-2-yl}amino)-
phenyl]methanesulfonamide (22)
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (300 mg), 2,4-difluorophenylboronic acid (237 mg,
1.50 mmol) and K3PO4 (425 mg, 2.00 mmol) in dioxane/water (20:1, 10
mL) was degassed with a stream of argon for 15 min. Pd(dppf)Cl2·DCM
(82 mg, 0.10 mmol) was added and the reaction mixture was heated for 1
h at 140 °C in a microwave oven. The mixture was diluted with an aq
NaCl solution and extracted with EtOAc. The combined organic phases
were dried (Na2SO4), filtered and concentrated. The residue was purified
by preparative HPLC (basic conditions) to give 18 as a light pink solid
(141 mg, 0.37 mmol): 1H NMR (400 MHz, [D6]DMSO):  = 4.25 (s, 2H),
6.89 (s, 2H), 7.10 (d, 1H), 7.24–7.47 (m, 3H), 7.80 (br s, 2H), 8.30 (br s,
1H), 8.86 (s, 1H), 10.47 ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd for
C16H14F2N5O2S: 378.0836, found: 378.0834.
methanesulfonamide (100 mg), 4-fluoro-2-(trifluoromethyl)phenylboronic
acid (104 mg, 0.50 mmol) and K3PO4 (142 mg, 0.67 mmol) in
dioxane/water (20:1, 3.3 mL) was degassed with a stream of argon for 15
min. Pd(dppf)Cl2·DCM (27 mg, 0.033 mmol) was added and the reaction
mixture was heated for 60 min at 140 °C in a microwave oven. The
mixture was diluted with dioxane, filtered and concentrated. The residue
was purified by preparative HPLC (acidic conditions) to give 22 as a
white solid (18 mg, 0.04 mmol): 1H NMR (400 MHz, [D6]DMSO):  = 4.23
(br s, 2H), 6.88 (s, 2H), 7.10 (d, 1H), 7.28–7.38 (m, 1H), 7.66–7.77 (m,
3H), 7.81 (dd, 1H), 7.91 (br s, 1H), 8.85 (s, 1H), 10.52 ppm (s, 1H); ESI-
HRMS: m/z [M + H]+ calcd for C17H14F4N5O2S: 428.0804, found:
428.0813.
14
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
1-[3-({4-[2-(Benzyloxy)-4-fluorophenyl]-1,3,5-triazin-2-yl}amino)-
phenyl]methanesulfonamide (23)
purification: 1H NMR (400 MHz, CDCl3):  = 1.31 (t, 3H), 3.07 (s, 3H),
4.18 (q, 2H), 4.79 (d, 1H), 4.88 (d, 1H), 7.64 (m, 1H), 7.81 (m, 1H), 8.30
ppm (m, 2H).
A
mixture of crude 3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzene-
methanesulfonamide (300 mg), 2-(benzyloxy)-4-fluorophenylboronic acid
(369 mg, 1.50 mmol) and K3PO4 (425 mg, 2.00 mmol) in dioxane/water
(20:1, 10 mL) was degassed with a stream of argon for 15 min.
Pd(dppf)Cl2·DCM (82 mg, 0.10 mmol) was added and the reaction
mixture was heated for 1 h at 140 °C in a microwave oven. The mixture
was diluted with an aq NaCl solution and extracted with EtOAc. The
combined organic phases were dried (Na2SO4), filtered and concentrated.
The residue was purified by flash chromatography (hexane/EtOAc, 2:1 to
EtOAc) to give 23 (32 mg, 0.07 mmol): 1H NMR (400 MHz, [D6]DMSO): 
= 4.21 (br s, 2H), 5.27 (s, 2H), 6.82–6.95 (m, 3H), 7.06 (br s, 1H), 7.14
(dd, 1H), 7.22–7.49 (m, 6H), 7.69 (br s, 1H), 7.84 (br s, 2H), 8.84 (s, 1H),
10.36 ppm (s, 1H); ESI-HRMS: m/z [M + H]+ calcd for C23H21FN5O3S:
466.1349, found: 466.1351.
(rac)-Ethyl
carbamate (29)
[(3-aminobenzyl)(methyl)oxido-6-sulfanylidene]-
TiCl3 solution (ca. 15% in ca. 10% HCl, 118 mL) was added to a stirred
solution of (rac)-28 (5.0 g, ca. 17.5 mmol) in THF (220 mL) at RT, and
the mixture was stirred for 18 h. The pH was increased to 8 by the
addition of 2 N NaOH to the reaction mixture, which was cooled with an
ice bath. The mixture was saturated with solid NaCl and extracted with
EtOAc (3 ×). The combined organic phases were washed with brine,
dried (Na2SO4), filtered and concentrated to give (rac)-29 (4.2 g, 16.5
mmol, 94%) that was used without further purification: 1H NMR (400 MHz,
CDCl3):  = 1.13 (t, 3H), 3.08 (s, 3H), 3.95 (m, 2H), 4.62 (s, 2H), 5.18 (br,
2H), 6.53 (m, 3H), 7.00 ppm (m, 1H).
(rac)-1-[(Methylsulfinyl)methyl]-3-nitrobenzene (26)
(rac)-Ethyl
[{3-[(4-chloro-1,3,5-triazin-2-yl)amino]benzyl}(methyl)-
oxido-6-sulfanylidene]carbamate (30)
FeCl3 (0.55 g, 3.4 mmol) was added to
a
solution of 1-
[(methylsulfanyl)methyl]-3-nitrobenzene (25; 21.6 g, 117.9 mmol) in
MeCN (280 mL), and the mixture was stirred at RT for 10 min. Periodic
acid (28.8 g, 126.1 mmol) was added under stirring in one portion and
the temperature was kept below 30 °C by cooling. The mixture was
stirred at RT for 90 min, then added to a stirred solution of sodium
thiosulfate pentahydrate (163 g, 660 mmol) in ice–water (1500 mL). This
mixture was saturated with solid NaCl and extracted with THF (2 ×). The
combined organic phases were washed with brine, dried (Na2SO4),
filtered and concentrated. The residue was purified by flash
chromatography (DCM/EtOH, 95:5) to give 26 (16.6 g, 83.1 mmol, 70%):
1H NMR (400 MHz, CDCl3):  = 2.53 (s, 3H), 3.97 (d, 1H), 4.10 (d, 1H),
7.58 (m, 1H), 7.67 (m, 1H), 8.17 (m, 1H), 8.21 ppm (m, 1H).
DIPEA (3.1 mL, 17.8 mmol) was added to a stirred solution of 2,4-
dichloro-1,3,5-triazine (1; 1.34 g, 8.9 mmol) in THF/iPrOH (1:1, 18 mL) at
–40 °C. Then, a solution of (rac)-29 (2.29 g, ca. 8.9 mmol) in THF/iPrOH
(1:1,
9 mL) was added at this temperature. Under stirring, the
temperature of the reaction mixture was slowly raised over 3 h to 0 °C.
The mixture was concentrated to give (rac)-30 (4.9 g) that was used
without further purification.
(rac)-Ethyl
[(3-{[4-(4-fluoro-2-methoxyphenyl)-1,3,5-triazin-2-yl]-
amino}benzyl)(methyl)oxido-6-sulfanylidene]carbamate (31)
A mixture of crude (rac)-30 (400 mg), 4-fluoro-2-methoxyphenylboronic
acid (276 mg, 1.62 mmol) and Pd(PPh3)4 (187 mg, 0.16 mmol) in 1,2-
dimethoxyethane (5.0 mL) and aq 2 M K2CO3 (1.1 mL) was degassed
using argon. The mixture was stirred under argon for 80 min at 100 °C.
After cooling, the mixture was diluted with EtOAc and washed with brine.
The organic phase was filtered using a Whatman filter and concentrated.
The residue was purified by flash chromatography (DCM/EtOH, 95:5) to
give (rac)-31 (178 mg, 0.39 mmol, 36%): 1H NMR (400 MHz, CDCl3):  =
1.30 (t, 3H), 3.00 (s, 3H), 3.93 (s, 3H), 4.17 (q, 2H), 4.74 (m, 2H), 6.77 (m,
2H), 7.16 (m, 1H), 7.42 (m, 1H), 7.53 (s, 1H), 7.74 (br, 1H), 7.84 (s, 1H),
7.94 (m, 1H), 8.82 ppm (s, 1H).
(rac)-2,2,2-Trifluoro-N-[methyl(3-nitrobenzyl)oxido-6-sulfanylidene]-
acetamide
To a suspension of (rac)-26 (16.6 g, 83.1 mmol), trifluoroacetamide (18.8
g, 166.1 mmol), MgO (13.4 g, 332.3 mmol) and rhodium(II) acetate dimer
(1.7 g, 3.9 mmol) in DCM (2290 mL) was added iodobenzene diacetate
(40.1 g, 124.6 mmol) at RT. The mixture was stirred for 16 h at RT, then
filtered and concentrated. The residue was purified by flash
chromatography (DCM/EtOH, 97:3) to give the desired product (25.6 g),
still containing minor impurities: 1H NMR (400 MHz, CDCl3):  = 3.28 (s,
3H), 4.79 (d, 1H), 4.91 (d, 1H), 7.69 (m, 1H), 7.80 (m, 1H), 8.31 (m, 1H),
8.36 ppm (m, 1H).
(rac)-4-(4-Fluoro-2-methoxyphenyl)-N-{3-[(S-methylsulfonimidoyl)-
methyl]phenyl}-1,3,5-triazin-2-amine (32 and BAY 1143572)
(rac)-1-[(S-Methylsulfonimidoyl)methyl]-3-nitrobenzene (27)
Freshly prepared 1.5 M sodium ethanolate in EtOH (2.9 mL, 4.35 mmol)
was added under argon to a solution of (rac)-31 (500 mg, 1.09 mmol) in
EtOH (18.5 mL), and the mixture was stirred at 60 °C for 2 h. Further
1.5 M sodium ethanolate in EtOH (2.9 mL, 4.35 mmol) was added, and
the mixture was stirred for an additional 5 h at 60 °C. After cooling, the
mixture was diluted with brine and extracted with EtOAc (3 ×). The
combined organic phases were filtered using a Whatman filter and
concentrated. The residue was purified by flash chromatography
(DCM/EtOH, 9:1) to give the desired racemic product as a white solid
(378 mg, 0.98 mmol, 90%): 1H NMR (400 MHz, CDCl3):  = 2.71 (s, 1H),
2.96 (s, 3H), 3.92 (s, 3H), 4.26 (d, 1H), 4.39 (d, 1H), 6.75 (m, 2H), 7.15
(m, 1H), 7.40 (m, 1H), 7.55 (s, 1H), 7.77 (m, 2H), 7.95 (m, 1H), 8.80 ppm
(s, 1H); ESI-HRMS: m/z [M + H]+ calcd for C18H19FN5O2S: 388.1243,
found: 388.1239.
K2CO3 (56.9 g, 411.8 mmol) was added to a solution of (rac)-2,2,2-
trifluoro-N-[methyl(3-nitrobenzyl)oxido-6-sulfanylidene]acetamide (25.6 g,
ca. 82.4 mmol) in MeOH (1768 mL) at RT. The mixture was stirred for 1 h
at RT, then diluted with EtOAc and brine, and extracted with EtOAc (2 ×).
The combined organic phases were dried (Na2SO4), filtered and
concentrated to give (rac)-27 (13.9 g, 65.1 mmol, 79%): 1H NMR
(400 MHz, CDCl3):  = 2.66 (br, 1H), 2.99 (s, 3H), 4.34 (d, 1H), 4.47 (d,
1H), 7.63 (m, 1H), 7.79 (m, 1H), 8.29 ppm (m, 2H).
(rac)-Ethyl [methyl(3-nitrobenzyl)oxido-6-sulfanylidene]carbamate
(28)
Ethyl chlorocarbonate (8.1 mL, 84.6 mmol) was added dropwise to a
stirred solution of (rac)-27 (13.9 g, 65.1 mmol) in pyridine (615 mL) at
0 °C. The mixture was slowly warmed to RT. After 24 h, the mixture was
concentrated and the residue was dissolved in EtOAc and washed with
brine. The organic phase was filtered using a Whatman filter and
concentrated to give (rac)-28 (19.7 g) that was used without further
The racemate was separated into the single enantiomers by preparative
HPLC [system: Dionex P580 pump, Gilson 215 Liquid Handler, Knauer
K-2501 UV detector; column: Chiralpak IC 5 m, 250 × 20 mm; eluent:
hexane/EtOH (6:4) + 0.1% diethylamine; flow: 40 mL/min; solution: 2600
mg/44 mL EtOH/DMSO (2:1); injection: 55 × 0.8 mL; temperature: RT;
15
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
detection: UV 254 nm]: tR (min): 13.4–15.6 (enantiomer 1: compound 32;
ESI-HRMS: m/z [M + H]+ calcd for C18H19FN5O2S: 388.1243, found:
388.1241), 15.6–18.5 (enantiomer 2: BAY 1143572).
physicochemical properties. M. Bergmann and K. Greenfield are
acknowledged for valuable technical support with the manuscript.
The activities of the LDC have been co-funded by the Max-
Planck Foundation, on behalf of the Max-Planck Society, as well
as by a grant from the Ministry for Research and Technology
(BMBF, grant number 0315326).
(R)-4-(4-Fluoro-2-methoxyphenyl)-N-{3-[(S-methylsulfonimidoyl)-
methyl]phenyl}-1,3,5-triazin-2-amine (BAY 1143572)
Mp: 165 °C; []58920 = –14.0 ± 0.40 (c = 1 in DMSO); 1H NMR (600 MHz,
[D6]DMSO):  = 2.81 (br s, 3H), 3.57 (s, 1H), 3.88 (br s, 3H), 4.29–4.40
(m, 2H), 6.90 (td, 1H), 7.09 (dd, 1H), 7.13 (d, 1H), 7.36 (t, 1H), 7.62–8.09
(m, 3H), 8.75–8.83 (m, 1H), 10.34 ppm (s, 1H); 13C NMR (125 MHz,
[D6]DMSO):  = 40.9, 56.2, 62.3, 100.4, 106.7, 120.2, 122.8, 122.9, 125.8,
128.4, 130.9, 133.2, 138.5, 160.0, 163.1, 164.5, 166.2, 171.1 ppm; IR
Keywords: PTEFb • CDK • antitumor agents • sulfoximines •
drug design
References:
(KBr):  = 3325–3246, 3013, 2960–2918, 1616–1403, 1284–1103 cm–1
;
ESI-HRMS: m/z [M + H]+ calcd for C18H19FN5O2S: 388.1243, found:
[1]
a) J. Wesierska-Gadek, I. Chamrad, V. Krystof, Future Med. Chem.
2009, 1, 1561; b) G. Mariaule, P. Belmont, Molecules 2014, 19, 14366;
c) J. Cicenas, K. Kalyan, A. Sorokinas, A. Jatulyte, D. Valiunas, A.
Kaupinis, M. Valius, Cancers 2014, 6, 2224; d) L. Santo, K. T. Siu, N.
Raje, Semin. Oncol. 2015, 42, 788; e) C. Sanchez-Martinez, L. M.
Gelbert, M. Jose Lallena, A. de Dios, Bioorg. Med. Chem. Lett. 2015,
25, 3420; f) U. Asqhar, A. K. Witkiewicz, N. C. Turner, E. S. Knudsen,
Nat. Rev. Drug Discovery 2015, 14, 130.
388.1246.
(R)-N-[(3-{[4-(4-Fluoro-2-methoxyphenyl)-1,3,5-triazin-2-yl]amino}-
benzyl)(methyl)oxido-6-sulfanylidene]acetamide (33)
Acetyl chloride (22 mg, 0.28 mmol) was added dropwise to a solution of
BAY 1143572 (100 mg, 0.26 mmol) in TEA (31 mg, 0.31 mmol) and DCM
(3.0 mL) at 0 °C. The ice bath was removed and the mixture was stirred
at RT for 3 h. Then, the mixture was diluted with water and extracted with
DCM (3 ×). The combined organic phases were filtered using a Whatman
filter and concentrated. The residue was purified by preparative HPLC
(acidic conditions) to give 33 as a white solid (63 mg, 0.15 mmol, 57%):
1H NMR (600 MHz, [D6]DMSO):  = 2.14 (s, 3H), 3.05 (s, 3H), 3.94 (s,
3H), 4.67 (d, 1H), 4.80 (d, 1H), 6.74–6.83 (m, 2H), 7.17 (d, 1H), 7.42 (t,
1H), 7.82 (br s, 2H), 7.97 (br s, 1H), 8.03 (br s, 1H), 8.83 ppm (s, 1H);
ESI-HRMS: m/z [M + H]+ calcd for C20H21FN5O3S: 430.1349, found:
430.1346.
[2]
a) M. E. Law, P. E. Corsino, S. Narayan, B. K. Law, Mol. Pharmacol.
2015, 88, 846; b) B. O’Leary, R. S. Finn, N. C. Turner, Nat. Rev. Clin.
Oncol. 2016, 13, 417.
[3]
[4]
[5]
S. Cho, S. Schroeder, M. Ott, Cell Cycle 2010, 9, 1697.
V. Krystof, S. Baumli, R. Fürst, Curr. Pharm. Des. 2012, 18, 2883.
See, for example: a) R. E. J. Beckwith, D. T. Curtis, E. Harrington, J.
H.-H. Hinrichs, J. A. Tallarico (Novartis AG), WO 2008079933, 2008; b)
M. Augustin, L. Zeitlmann, G. Stumm, M. A. Pleiss, P. Wabnitz, H.
Allgeier (Ingenium Pharmaceuticals AG), WO 2008049856, 2008; c) H.
Allgeier, M. Augustin, A. Mueller, L. Zeitlmann, A. Marquardt, M. A.
Pleiss, U. Heiser, A. J. Niestroj (Ingenium Pharmaceuticals AG), WO
2009047359, 2009; d) P. Barsanti, C. Hu, J. Jin, R. Keyes, R. Kucejko,
X. Lin, Y. Pan, K. B. Pfister, M. Sendzik, J. Sutton, L. Wan (Novartis
AG), WO 2011012661, 2011; e) M. W. Nowicki, M. D. Walkinshaw,
Chem. Biol. 2010, 17, 1047; f) X. Liu, S. Shi, F. Lam, C. Pepper, P. M.
Fischer, S. Wang, Int. J. Cancer 2012, 130, 1216; g) H. Shao, S. Shi, S.
Huang, A. J. Hole, A. Y. Abbas, S. Baumli, X. Liu, F. Lam, D. W. Foley,
P. M. Fischer, M. Noble, J. A. Endicott, C. Pepper, S. Wang, J. Med.
Chem. 2013, 56, 640; h) http://www.in-pharmatechnologist.com/Drug-
Delivery/AstraZeneca-boosts-oncology-arsenal-with-CDK9-inhibitor-
platform (accessed July, 2017); i) J. Gong, Z.-F. Tao, Y. Tong, G. Zhu,
T. Penning, A. J. Souers (Abbvie Inc.), WO 2014151444, 2014; j) T. Yin,
M. J. Lallena, E. L. Kreklau, K. R. Fales, S. Carballares, R. Torrres, G.
N. Wishart, R. T. Ajamie, D. M. Cronier, P. W. Iversen, T. I. Meier, R. T.
Foreman, D. Zeckner, S. E. Sissons, B. W. Halstead, A. B. Lin, G. P.
Donoho, Y. Qian, S. Li, S. Wu, A. Aggarwal, X. S. Ye, J. J. Starling, R.
B. Gaynor, A. de Dios, J. Diu, Mol. Cancer Ther. 2014, 13, 1442; k) F.
Morales, A. Giordano, Cell Cycle 2016, 15, 519.
X-ray structure determination of 33
Single crystals of 33 were obtained by slow evaporation of an ethyl
acetate solution of 33 at RT. A single crystal was mounted on a
CryoLoop using inert oil. Data collection was carried out using a Bruker
diffractometer equipped with an APEX II CCD area detector, an IS
microsource with Cu K radiation, mirrors as monochromator and a
Kryoflex low temperature device (T = 110 K). The program APEX II
v.2011.8-0 was used for data collection and reduction.[29] Absorption
correction and scaling was performed using SADABS.[30] The crystal
structure solution was achieved using direct methods as implemented in
SHELXTL version 6.14[31] and visualized using the XP program[31]
.
Missing atoms were subsequently located from difference Fourier
synthesis and added to the atom list. Least-squares refinement on F2
using all measured intensities was carried out using the program
SHELXTL version 6.14.[31] All non-hydrogen atoms were refined including
anisotropic displacement parameters. Final data collection and structure
refinement parameters:  = 1.54178 Å, T = 110 K, space group = P2(1),
a = 10.3041(8) Å, b = 7.9030(9) Å, c = 12.8716(11) Å ,  = 108.990(5)°, Z
[6]
[7]
[8]
P. L. Toogood, P. J. Harvey, J. T. Repine, D. J. Sheehan, S. N.
VanderWel, H. Zhou, P. R. Keller, D. J. McNamara, D. Sherry, T. Zhu, J.
Brodfuehrer, C. Choi, M. R. Barvian, D. W. Fry, J. Med. Chem. 2005,
48, 2388.
= 2, reflections collected = 6794, independent reflections = 1261 (Rint
=
0.0848), completeness = 98.2%, data-to-parameter ratio = 11.5, R1 (I >
2) = 0.061, wR2 (all data) = 0.1420, GOF = 0.988, Flack parameter =
0.05(4), largest difference peak and hole = 0.297/–0.262 e·Å–3. The
crystallographic data for 33 have been deposited with the Cambridge
Crystallographic Data Centre with the code CCDC 1565910.
D. Parry, T. Guzi, F. Shanahan, N. Davis, D. Prabhavalkar, D. Wiswell,
W. Seghezzi, K. Paruch, M. P. Dwyer, R. Doll, A. Nomeir, W. Windsor,
T. Fischmann, Y. Wang, M. Oft, T. Chen, P. Kirschmeier, E. M. Lees,
Mol. Cancer Ther. 2010, 9, 2344.
a) G. Siemeister, U. Lücking, A. M. Wengner, P. Lienau, W. Steinke, C.
Schatz, D. Mumberg, K. Ziegelbauer, Mol. Cancer Ther. 2012, 11,
2265; b) U. Lücking, R. Jautelat, M. Krüger, T. Brumby, P. Lienau, M.
Schäfer, H. Briem, J. Schulze, A. Hillisch, A. Reichel, A. M. Wengner, G.
Siemeister, ChemMedChem 2013, 8, 1067.
Acknowledgements
[9]
M. Herbrink, J. H. M. Schellens, J. H. Beijnen, B. Nuijen, J. Controlled
Release 2016, 239, 118.
We thank K. Sauvageot-Witzku, R. Golde, A. Glowczewski, C.
Pakebusch, N. Gallus and R. Droschinski for technical support,
O. Schenk for HPLC separations, S. Gründemann and G. Depke
for analytical support, and U. Ganzer for the measurement of
[10] C. Bissantz, B. Kuhn, M. Stahl, J. Med. Chem. 2010, 53, 5061.
[11] H. R. Bentley, E. E. McDermott, J. Pace, J. K. Whitehead, T. Moran,
Nature 1949, 163, 675.
16
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
[12] a) C. R. Johnson, Aldrichimica Acta 1985, 18, 3; b) M. Reggelin, C. Zur,
Synthesis 2000, 1.
[23] W. Kueng, E. Silber, U. Eppenberger, Anal. Biochem. 1989, 182, 16.
[24] See, for example: E. H. Kerns, L. Di, Drug-like Properties: Concepts,
Structure Design and Methods, Elsevier Inc./Academic Press,
Burlington, MA, USA, 2008, pp. 276–286.
[13] a) C. R. Johnson, Acc. Chem. Res. 1973, 6, 341; b) D. Craig, F.
Grellepois, A. J. P. White, J. Org. Chem. 2005, 70, 6827; c) H.-J. Gais,
G. S. Babu, M. Günter, P. Das, Eur. J. Org. Chem. 2004, 1464; d) M.
Harmata, X. Hong, J. Am. Chem. Soc. 2003, 125, 5754; e) M. Harmata,
X. Hong, C. L. Barnes, Tetrahedron Lett. 2003, 44, 7261; f) M. Harmata,
N. Pavri, Angew. Chem. Int. Ed. 1999, 38, 2419; g) S. Koep, H.-J. Gais,
G. Raabe, J. Am. Chem. Soc. 2003, 125, 13243; h) X. Shen, W. Miao,
C. Ni, J. Hu, Angew. Chem. Int. Ed. 2014, 53, 775; i) X. Shen, Q. Liu, W.
Zhang, J. Hu, Eur. J. Org. Chem. 2016, 906.
[25] A. B. Ahmad, P. N. Bennett, M. Rowland, J. Pharm. Pharmacol. 1983,
35, 219.
[26] Glide, version 5.7, Schrödinger, LLC, New York, NY, 2011.
[27] S. Baumli, J. A. Endicott, L. N. Johnson, Chem. Biol. 2010, 17, 931.
[28] LigPrep, version 2.5, Schrödinger, LLC, New York, NY, 2011.
[29] APEX II, version 2011.8-0, Bruker AXS Inc., Madison, WI, 2011.
[30] L. Krause, R. Herbst-Irmer, G. M. Sheldrick, D. Stalke, J. Appl. Cryst.
2015, 48, 3
[14] a) C. Bolm, O. Simic, J. Am. Chem. Soc. 2001, 123, 3830; b) M.
Harmata, S. K. Ghosh, Org. Lett. 2001, 3, 3321; c) C. Bolm, M. Martin,
O. Simic, M. Verrucci, Org. Lett. 2003, 5, 427; d) C. Bolm, M. Felder, J.
Müller, Synlett 1992, 439; e) C. Bolm, M. Verrucci, O. Simic, P. G.
Cozzi, G. Raabe, H. Okamura, Chem. Commun. 2003, 2826; f) M.
Langner, C. Bolm, Angew. Chem. Int. Ed. 2004, 43, 5984, g) M.
Langner, P. Remy, C. Bolm, Chem. – Eur. J. 2005, 11, 6254; h) M. T.
Reetz, O. G. Bondarev, H.-J. Gais, C. Bolm, Tetrahedron Lett. 2005, 46,
5643.
[31] SHELXTL, version 6.14, Bruker AXS Inc., Madison, WI, 2003.
[15] U. Lücking, Angew. Chem. Int. Ed. 2013, 52, 9399.
[16] a) M. Frings, C. Bolm, A. Blum, C. Gnamm, Eur. J. Med. Chem. 2017,
126, 225; b) J. A. Sirvent, U. Lücking, ChemMedChem 2017, 12, 487.
[17] See, for example: a) U. Lücking, M. Krueger, R. Jautelat, G. Siemeister
(Schering AG), WO 2005037800, 2005; b) U. Lücking, G. Siemeister, R.
Jautelat (Schering Aktiengesellschaft), WO 2006099974, 2006; c) U.
Lücking (Schering Aktiengesellschaft), EP 1710246, 2006; d) U.
Lücking, G. Kettschau, H. Briem, W. Schwede, M. Schäfer, K.-H.
Thierauch, M. Husemann (Schering Aktiengesellschaft), WO
2006108695, 2006; e) U. Luecking, D. Nguyen, A. von Bonin, O. von
Ahsen, M. Krueger, H. Briem, G. Kettschau, O. Prien, A. Mengel, K.
Krolikiewicz, U. Boemer, U. Bothe, I. Hartung (Schering
Aktiengesellschaft), WO 2007071455, 2007; f) U. Lücking, G.
Siemeister, B. Bader (Schering Aktiengesellschaft), WO 2007079982,
2007; g) U. Luecking, G. Siemeister, R. Jautelat (Bayer Schering
Pharma Aktiengesellschaft), WO 2008025556, 2008; h) O. Prien, K. Eis,
U. Lücking, J. Guenther, D. Zopf (Bayer Schering Pharma
Aktiengesellschaft), DE 102007024470, 2008; i) I. Hartung, U. Bothe, G.
Kettschau, U. Luecking, A. Mengel, M. Krueger, K.-H. Thierauch, P.
Lienau, U. Boemer (Bayer Schering Pharma Aktiengesellschaft), WO
2008155140, 2008; j) D. Nguyen, A. Von Bonin, M. Haerter, H. Schirok,
A. Mengel, O. Von Ahsen (Bayer Schering Pharma Aktiengesellschaft),
WO 2009089851, 2009; k) M. Härter, H. Beck, P. Ellinghaus, K.
Berhoerster, S. Greschat, K.-H. Thierauch, F. Süssmeier (Bayer
Schering Pharma Aktiengesellschaft), WO 2010054763, 2010; l) F. von
Nussbaum, D. Karthaus, S. Anlauf, M. Delbeck, V. M.-J. Li, D. Meibom,
K. Lustig, D. Schneider (Bayer Schering Pharma Aktiengesellschaft),
WO 2010115548, 2010; m) W. Schwede, U. Klar, C. Möller, A. Rotgeri,
W. Bone (Bayer Schering Pharma Aktiengesellschaft), WO
2011009531, 2011.
[18] H. Okamura, C. Bolm, Org. Lett. 2004, 6, 1305.
[19] For an alternative synthesis based on a palladium-catalyzed -arylation,
see ref. 16b.
[20] P. Ertl, B. Rohde, P. Selzer, J. Med. Chem. 2000, 43, 3714.
[21] J. E. Bradner, D. Hnisz, R. A. Young, Cell 2017, 168, 629.
[22] a) A. Scholz, T. Oellerich, A. Hussain, S. Lindner, U. Luecking, A. O.
Walter, P. Ellinghaus, R. Valencia, F. von Nussbaum, D. Mumberg, M.
Brands, S. Ince, H. Serve, K. Ziegelbauer, Cancer Res. 2016, 76 (14
Suppl), Abstract nr 3022; DOI: 10.1158/1538-7445.AM2016-3022; b) A.
Scholz, U. Luecking, G. Siemeister, P. Lienau, U. Boemer, P.
Ellinghaus, A. O. Walter, R. Valencia, S. Ince, F. von Nussbaum, D.
Mumberg, M. Brands, K. Ziegelbauer, Cancer Res. 2015, 75 (15 Suppl),
Abstract nr DDT02-02; DOI: 10.1158/1538-7445.AM2015-DDT02-02; c)
U. Luecking, A. Scholz, P. Lienau, G. Siemeister, D. Kosemund, R.
Bohlmann, K. Eis, M. Gnoth, I. Terebesi, K. Meyer, K. Prelle, R.
Valencia, S. Ince, F. von Nussbaum, D. Mumberg, K. Ziegelbauer, B.
Klebl, A. Choidas, P. Nussbaumer, M. Baumann, C. Schultz-
Fademrecht, G. Ruehter, J. Eickhoff, M. Brands, Cancer Res. 2015, 75
(15 Suppl), Abstract nr 2828; DOI: 10.1158/1538-7445.AM2015-2828.
17
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201700447
ChemMedChem
FULL PAPER
Entry for the Table of Contents
atuveciclib (BAY1143572)
The benzyl sulfoximine atuveciclib (BAY 1143572), a potent and highly selective, oral PTEFb/CDK9 inhibitor, exhibited the most
promising overall profile with respect to potency, selectivity, physicochemical properties, and in vivo PK as well as in vivo potency in
animal models during lead optimization. BAY 1143572 is the first selective PTEFb/CDK9 inhibitor that entered clinical evaluation for
the treatment of cancer.
18
This article is protected by copyright. All rights reserved.