Discovery of Macrocyclic Pyrimidines as MerTK-Specific Inhibitors

Article abstract of DOI:10.1002/cmdc.201600589

Macrocycles have attracted significant attention in drug discovery recently. In fact, a few de novo designed macrocyclic kinase inhibitors are currently in clinical trials with good potency and selectivity for their intended target. In this study, we successfully engaged a structure-based drug design approach to discover macrocyclic pyrimidines as potent Mer tyrosine kinase (MerTK)-specific inhibitors. An enzyme-linked immunosorbent assay (ELISA) in 384-well format was employed to evaluate the inhibitory activity of macrocycles in a cell-based assay assessing tyrosine phosphorylation of MerTK. Through structure–activity relationship (SAR) studies, analogue 11 [UNC2541; (S)-7-amino-N-(4-fluorobenzyl)-8-oxo-2,9,16-triaza-1(2,4)-pyrimidinacyclohexadecaphane-1-carboxamide] was identified as a potent and MerTK-specific inhibitor that exhibits sub-micromolar inhibitory activity in the cell-based ELISA. In addition, an X-ray structure of MerTK protein in complex with 11 was resolved to show that these macrocycles bind in the MerTK ATP pocket.

Full text of DOI:10.1002/cmdc.201600589

Accepted Article
Title: Discovery of Macrocyclic Pyrimidines as MerTK-specific Inhibitors
Authors: Andrew McIver, Weihe Zhang, Qingyang Liu, Xinpeng Jiang,
Michael Stashko, James Nichols, Michael Miley, Jacqueline
Norris-Drouin, Mischa Machius, Deborah DeRyckere, Edgar
Wood, Douglas Graham, H Shelton Earp, Dmitri Kireev,
Stephen Frye, and Xiaodong Wang
This manuscript has been accepted after peer review and appears as an
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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.201600589
Link to VoR: http://dx.doi.org/10.1002/cmdc.201600589
A Journal of
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10.1002/cmdc.201600589
ChemMedChem
Discovery of Macrocyclic Pyrimidines as MerTK-specific Inhibitors
Dr. Andrew L. McIver,†‡ Dr. Weihe Zhang,†‡ Dr. Qingyang Liu,† Dr. Xinpeng Jiang,† Michael A.
Stashko,† James Nichols,₸ Dr. Michael J Miley,§ Dr. Jacqueline Norris-Drouin,† Dr. Mischa Machius,§
Prof. Deborah DeRyckere,‖ Dr. Edgar Wood,₸ Prof. Douglas K. Graham,‖₸ Prof. H Shelton Earp,¶§₸ Prof.
Dmitri Kireev,† Prof. Stephen V. Frye,†¶ ₸ Prof. Xiaodong Wang†*
†Center for Integrative Chemical Biology and Drug Discovery, Division of Chemical Biology and Me-
dicinal Chemistry, Eshelman School of Pharmacy
₸Meryx, Inc., USA
§ Department of Pharmacology, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599,
USA
‖Aflac Cancer and Blood Disorders Center of Children’s Healthcare of Atlanta, Department of Pediat-
rics, School of Medicine, Emory University, Atlanta, GA 30322, USA
¶Lineberger Comprehensive Cancer Center, Department of Medicine, School of Medicine, University of
North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA
KEYWORDS. MerTK, MerTK-specific inhibitors, macrocycles, pyrimidines, intramolecular hydrogen bonds.
ABSTRACT: Macrocycles have attracted significant attention in drug discovery recently. In fact, a few
de novo designed macrocyclic kinases inhibitors are currently in clinical trials with good potency and
selectivity for their intended target. In this paper, we have successfully engaged a structure-based drug
design approach to discover macrocyclic pyrimidines as potent Mer tyrosine kinase (MerTK)-specific
inhibitors. An enzyme-linked immunosorbent assay (ELISA) in a 384-well format was employed to eval-
uate the inhibitory activity of macrocycles in a cell-based assay assessing tyrosine phosphorylation of
MerTK. Through SAR studies, analogue 11 (UNC2541) was identified as a potent and MerTK-specific
inhibitor and exhibits sub-micro molar inhibitory activity in the cell-based ELISA assay. In addition, an
X-ray structure of MerTK protein in complex with 11 was resolved to show that these macrocycles bind
in the MerTK ATP pocket.
Macrocycles, especially de novo designed macrocycles, have recently gained attention in drug
discovery due to the new physicochemical properties and broader intellectual property (IP) that they may
provide.1-3 In macrocycles, cyclization leads to a structural preorganization which can increase both bind-
ing affinity and selectivity by engaging targets through numerous and spatially distributed binding inter-
actions.1,4 A few macrocyclic kinase inhibitors are currently in clinical trials with good potency and se-
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10.1002/cmdc.201600589
ChemMedChem
lectivity for their intended target.5-7 We have been interested in Mer tyrosine kinase (MerTK) as a thera-
peutic target8,9 and have developed several MerTK inhibitors with varying selectivity profiles.10-14 The
most advanced compound among these inhibitors is UNC2025 which is a potent and highly orally bioa-
vailable MerTK inhibitor.14 It is also equally potent against FMS-like tyrosine kinase (Flt3). This dual
inhibitory activity of UNC2025 is desirable for certain diseases such as acute myeloid leukemia (AML),
however, inhibition of Flt3 has been associated with hematopoietic toxicity15,16 and is therefore inadvisa-
ble for other applications of MerTK inhibitors. The new pyrrolopyrimidine macrocycles that we have
developed recently share the same undesired selectivity profile.17 In this paper, we will present a new type
of MerTK-specific inhibitor—macrocyclic pyrimidines.
We have recently discovered substituted-pyrimidines as novel MerTK specific inhibitors via a
structure-based drug design approach.12 Based on our published X-ray crystal structure of MerTK in com-
plex with 1 (Figures 1a and 1b), the butyl side chain and the cyclohexyl alcohol are close to each other
and well-positioned to form a macrocycle. One example of this design is compound 2 (Figure 1c), which
has a hydrogen donor, an amino group, at the same position as the hydroxyl group in 1. The macrocycle
is connected by an amide bond and the cyclohexyl ring has been opened to eliminate the chance to intro-
duce new stereogenic centers. Compound 2 fits the MerTK docking model and is predicted to retain three
key hydrogen bonds with MerTK protein (Figure 1d, two with the hinge area (F673 and P672) and one
with either D741, R727, or N728). Since substituted-pyrimidines show some selectivity for MerTK over
Flt3 (e.g. compound 1 is 14-fold more active against MerTK versus Flt3),12 we were interested to see if
macrocycles based on this scaffold could improve MerTK inhibitory activity and/or the selectivity profile
over other TAM family members and Flt3.
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Figure 1. a. Structure of 1; b. X-ray crystal structure of 1 in complex with MerTK (kinase domain) (PDB
ID code 4MHA); c. Structure of 2; d. Docking model of macrocyclic pyrimidine 2.
The syntheses of the designed macrocyclic compounds are straight forward. A general synthetic
route is shown in Scheme 1 (see Supporting Information for details). Commercially available 2,4-dichlo-
ropyrimidine-5-carbonyl chloride reacted with an amine or alcohol to form the amide/ester I. In a one-pot
reaction Boc protected amino acids with differing length alkyl chains and various diamines were attached
to the pyrimidine core to provide intermediate II. The macrocycle was closed using an intramolecular
amide coupling reaction, followed by cleavage of the Boc protecting group to yield the desired macrocy-
cles III. To explore SAR at the R´ position, the starting acid chloride was first converted into an ethyl
ester. After the formation of the desired macrocycle, the ethyl ester was hydrolyzed under basic conditions
and the resulting free acid was converted to the final amide IV via an amide bond coupling reaction and
cleavage of the Boc protecting group.
Scheme 1. The synthetic route for macrocyclic compounds
Compound 2 was synthesized using the route presented in Scheme 1 and was tested using in-house
microfluidic capillary electrophoresis (MCE) assays at the ATP Kms (details see supporting information
Table S1).18-20 As shown in Table 1, compound 2 demonstrated exceptional selectivity over Flt3 (110-fold
vs 14-fold for compound 1) along with weaker activity against MerTK (12-fold lower IC50 than compound
1). To improve the MerTK activity of compound 2, we initially explored the ring size of the macrocycle
since this would simultaneously vary the position of the hydrogen-bond donor, the amino group, and the
flexibility of the ring. As shown in Table 1, when m = 1, the inhibitory activity of macrocycles varied
depending on the ring size. Compound 2 (n = 2) was 3-fold more potent than compound 3 (n = 1),
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10.1002/cmdc.201600589
ChemMedChem
however, compound 4 (n = 3) was 12-fold less active than compound 2. When n ≥ 4, the potency of the
macrocycles was improved as the macrocyclic ring size was increased (compounds 4-7). The trend was
also evident when m = 2. Compound 8 (n = 1) was a weak MerTK inhibitor with micromolar potency
while compound 11 (UNC2541) (n = 4) was very potent with a low nanomolar IC50 against MerTK. The
MerTK inhibitory activities of compound 9 (n = 2) and 10 (n = 3) were intermediate. When the macrocy-
clic ring became larger (n = 5 or 6), the MerTK activity of the corresponding analogues 12 and 13 was
retained, however, the selectivity over Flt3 decreased (73-fold for 11 vs 59-fold for 12 & 19-fold for 13).
In general, most of the analogues had excellent selectivity for MerTK over Flt3 and good selectivity over
Axl and Tyro3. To evaluate the inhibitory activity of these compounds in a cell-based assay of tyrosine
phosphorylated MerTK (pMerTK), we used a 384-well format enzyme-linked immunosorbent assay
(ELISA),17 in which HEK293 cells were transfected with a cDNA encoding chimeric protein consisting
of the extracellular and transmembrane domains from the epidermal growth factor receptor (EGFR) and
the MerTK intracellular kinase domain. Activity was then stimulated with EGF. A chimeric protein was
utilized because consistent stimulation of native MerTK kinase with the complex, natural ligand (GAS6
plus PtdSer) was not practical. Compound 11 proved to be the most active in this pMerTK ELISA assay
with an EC50 510 nM. The 100-fold IC50 shift from the MCE assay to the pMerTK ELISA assay isn’t
unexpected due to the ATP concentration difference (22 µM in the MCE assay vs low millimolar range in
the pMerTK ELISA assay).
Table 1. Preliminary SAR of macrocycle ring size
IC50 (nM)a
EC50 (nM)a
Compound
n
m
MerTK
Axl
Tyro3
Flt3
pMerTK ELISA
2
3
4
5
6
7
8
2
1
3
4
5
6
1
1
1
1
1
1
1
2
61 ± 46
200 ± 140
760 ± 330
230 ± 54
91 ± 28
1600 ± 370
1800 ± 970
6700 ± 6300
inactive
>1000
>1000
>1000
>1000
>1000
>1000
>1000
21000 ± 7300 8200 ± 7000
28000 ± 2900
7800 ± 2800
2700 ± 1000
1100 ± 340
inactive
inactive
inactive
8500 ± 5300 14000 ± 14000
3400 ± 1500
2700 ± 2000
24000 ± 3300
10000 ± 1800
2100 ± 560
inactive
62 ± 25
3200 ± 2500
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9
2
3
4
5
6
2
2
2
2
2
140 ± 96
36 ± 14
4000 ± 1400
1700 ± 350
120 ± 60
6900 ± 5800
1600 ± 910
220 ± 82
inactive
3200 ± 610
320 ± 11
240b
>1000
>1000
10
11
12
13
4.4 ± 0.5
4.1 ± 0.2
6.2 ± 3.3
510 ± 300
ND
130 ± 16
150 ± 4.4
170 ± 13
160 ± 110
120 ± 36
790 ± 110
a Values are the mean of two or more independent assays ± SD. b Only test once.
With the optimal ring size and linker of macrocycle 11, the SAR at the R´ position was explored
using the synthetic route presented in Scheme 1 (Table 2). Introducing a larger and basic dimethyl amine
group in the para-position of the benzyl group at the R´ position didn’t affect the activity or selectivity of
the corresponding analogue 14 in either the MCE or pMerTK ELISA assays. However, 3,5-dimethoxyl
benzyl group at the R´ position led to a 6-fold less active analogue 15. A para-substituted phenyl group
at the same position (16‒18) yielded analogues with dramatically diminished selectivity for MerTK over
Flt3 although their MerTK enzymatic activities were similar to 11. No MerTK cellular activity was ob-
served for these derivatives at a concentration of 1.0 µM in the pMerTK ELISA assay. When placing
heterocycles such as 3-pyridyl and 4-pyridyl at the R´ position, the resulting analogues 19 and 20 had
similar inhibitory enzymatic activity and selectivity for MerTK over Flt3 compared to 11, and again, no
cellular activity was observed at a concentration of 1.0 µM. 2-Pyridyl at the same position led to a 5-fold
less active analogue 21. However, when 3-pyridyl at the R´ site was further substituted at the para-position
with 1-pyrazole, the resulting analogue 22 was 4-fold more active than the parental analogue 19 and ex-
hibited improved Flt3 selectivity (193-fold for 22 vs 152-fold for 19) and cellular activity (IC50 420 nM).
Saturated heterocycles such as 4-piperidine (23) and 4-tetrahydropyran (24) were also tolerated at this
position. Analogues 23 and 24 were equally potent to 19 in either the MCE or pMerTK ELISA assays.
Similarly, further substitution on the nitrogen of 4-piperidine with a benzyl group at the R´ position in-
creased the activity of the corresponding analogue 25, especially in the pMerTK ELISA assay (IC50 250
nM).
Table 2. SAR Study of the R´ position
Compound
R'
IC50 (nM)a
Tyro3
EC50 (nM)a
MerTK
Axl
Flt3
pMerTK ELISA
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14
6.4 ± 1.3 180 ± 98 330 ± 110 740 ± 210
470 ± 79
>1000
15
25 ± 21
500 ± 30 920 ± 470 5900 ± 470
16
17
6.0 ± 2.8 100 ± 34 340 ± 210
9.5 ± 2.9 300 ± 130 800 ± 340
49 ± 11
38 ± 43
>1000
>1000
18
19
4.3 ± 1.1
83 ± 54
150 ± 27
22 ± 3.1
>1000
>1000
6.3 ± 3.3 330 ± 270 340 ± 230 960 ± 410
20
21
22
4.4 ± 4.1 110 ± 41 360 ± 230 780 ± 110
22 ± 16 540 ± 170 680 ± 470 890 ± 260
>1000
>1000
420b
1.5 ± 0.4
72 ± 31
86 ± 2.1
290 ± 25
23
24
25
6.5 ± 2.0 150 ± 66 290 ± 220 850 ± 280
>1000
>1000
14 ± 8.5
2.9 ± 1.9
300 ± 89 970 ± 680 860 ± 220
50 ± 22 150 ± 200 730 ± 260
250 ± 28
a Values are the mean of two or more independent assays ± SD. b Only test once.
The weak cellular activity of these analogues may be due to the polar α-amino amide linker. There-
fore, we varied the linker of the macrocycle to explore if any other linker would improve the cellular
activity (Table 3). When the primary amine was protected by a Boc group (26) or removed (27), the
activity against MerTK was dramatically reduced (over 10-fold in the MCE assay compared to compound
11) due to lack of the key hydrogen bond with either D741 or R727 or N728. On the other hand, a similarly
active compound 28 was obtained by substitution of the primary amine to yield a secondary amino group.
Unfortunately, the cellular activity of 28 wasn’t improved by the reducing the polarity and removing a
hydrogen bond donor of the amino group because these changes decreased the MerTK inhibitory activity
of 28 (3-fold) as well. In addition, replacement of the amide group in 11 with a less polar ester group such
as in 29, preserved activity in either the MerTK MCE or pMerTK ELISA assays. Another attempt to
reduce the polarity while retaining the hydrogen bond donor property of the α-amino amide linker was to
replace it with a secondary hydroxyl group. Three analogues were prepared (30‒32). The ring size and
the position of the hydrogen bond donor of analogue 30 was the same as analogue 10. Both analogues had
similar MerTK activity, however, the Flt3 selectivity of 30 was diminished from 89-fold (10) to 24-fold
(30). The activity and selectivity were worse for analogue 31. Although the ring size and the position of
the hydrogen donor of 31 was the same as 12, analogue 31 was 34-fold less active than 12 in the MerTK
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10.1002/cmdc.201600589
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MCE assay and only had 5-fold selectivity over Flt3. Interestingly, analogue 32 had the same ring size
and the same position of its hydrogen bond donor as 3, was 10-fold more active than 3 in the MCE assay.
It also had weak activity in the pMerTK ELISA assay (IC50 600 nM). Linkers containing oxygen as the
hydrogen bond acceptor such as epoxide (33) or ether (34 & 35), showed much lower MerTK inhibitory
activity. More rigid macrocycles containing a phenyl ring resulted in analogues 36 and 37. Analogue 36
had the same-sized macrocycle with its hydrogen bond donor, an amino group, at the same position as 22.
However, it was 27-fold less active than 22 in the MCE assay. Similarly, analogue 37 was 7-fold weaker
than 25 with similar selectivity over Flt3. Overall, the amide linker was optimal in this series.
Table 3. Exploration of different macrocycle linkers
Com-
pound
Structure
IC50 (nM)a
Axl
EC50 (nM)a
Mer
Tyro3
Flt3
pMerTK
ELISA
>1000
26
54 ± 2.7
1300 ± 190
2200 ± 350
inactive
27b
28
59
670
2300
4500
750
11 ± 2.7
210 ± 92
850 ± 350
840 ± 350
410 ± 140
29
3.9 ± 0.1
220 ± 11
190 ± 49
65 ± 13
430 ± 42
30b
31b
68
1600
1300
1500
2500
1600
630
>1000
>1000
140
32
21b
920b
1100b
900b
600 ± 76
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33
480 ± 150
770 ± 600
4400 ± 1100 9200 ± 4100
3100 ± 1700
ND
34
35
36
20170
±17020
17350 ±
15500
20080 ±
17190
ND
14000 ±
10000
inactive
inactive
inactive
ND
40 ± 13
300 ± 150
670 ± 470
9700 ± 6500
>1000
37
21 ± 7.0
590 ± 340
1200 ± 1500
15140 ±
21010
>1000
a Values are the mean of two or more independent assays ± SD. b Only test once.
To confirm our initial design strategy and gain better insight into ligand-protein interactions, we
solved the X-ray structure of MerTK in complex with 11 at a resolution of 2.23 Å. Overall, the structure
corroborated the preliminary docking hypothesis (Figure 1). However, there was one notable difference.
In the crystal structure (Figure 2), both the carbonyl and primary amine groups of the macrocycle pointed
outward with respect to the residues R727, N728 and D741, while we had hypothesized that the main
interaction in this region would occur between only the primary amine and the above three residues. Our
docking hypothesis was based on previous non-macrocyclic amino-pyridines, such as 1, which all needed
either a hydroxyl or a primary amino group to gain potency through interactions with R727, N728 or
D741.
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Figure 2. a. Structure of 11; b and c. X-ray crystal structure of 11 in complex with MerTK (kinase domain)
(PDB ID code 5K0X).
In addition, we used a panel of 30 kinases which emphasized tyrosine kinases along with selected
serine/threonine kinases to assess the selectivity of promising compounds over other kinase families.8-9
Analogue 11 was evaluated in this panel at a concentration 100 fold above its Mer IC50 (Figure 3) (detailed
in supplemental materials). Similar to its open-chained analogue (substituted pyrimidines),8 five tyrosine
kinases were inhibited greater than 50% in the presence of 430 nM 11. However, three additional ser-
ine/threonine kinases were inhibited greater than 50% and were not inhibited by 1 under the same condi-
tions.
Figure 3. Kinase Tree
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In summary, novel and potent macrocyclic pyrimidines have been discovered as MerTK kinase
inhibitors via a structure-based drug design approach. The binding mode of these macrocycles was con-
firmed by an X-ray structure of MerTK protein complexed with analogue 11. A high-throughput ELISA
assay was also employed to evaluate the inhibitory activity of the compounds in a cell-based assay of
tyrosine phosphorylated MerTK. These macrocycles proved to be MerTK-selective within the TAM fam-
ily and over Flt3. However, the selectivity over other kinase families and the cellular activity of these
macrocycles need to be further improved to yield a lead compound worthy of further progression in this
series.
AUTHOR INFORMATION
Corresponding Author
* Tel: 919-843-8456. Fax: 919-843-8465. E-mail: xiaodonw@unc.edu.
Present Addresses
† Andrew McIver: Chimerix. Inc., 2505 Meridian Pkwy, Suite 100, Durham, NC 27713;Weihe Zhang: BioCryst Pharmaceuti-
cals, Inc., 2100 Riverchase Center, Suite 200, Birmingham, AL 35244; Qingyang Liu: Department of chemistry, Montgomery
College, 51 Mannakee Street, Rockville, MD 20850; Xingpeng Jiang: School of Pharmacy, Zhejiang University of Technology,
Jiangshan, Zhejiang, China.
Author Contributions
This manuscript was written through contributions of all authors. / All authors have given approval to the final version of the manuscript. /
‡These authors contributed equally.
Funding Sources
This work was supported by the University Cancer Research Fund and Federal Funds from the National Cancer institute, National Institute
of Health, under Contract No. HHSN261200800001E. The content of this publication does not necessarily reflect the views or policies of
the Department of Health and Human Services, nor does mention of trade names, commercial products, or organizations imply endorsement
by the U.S. Government.
ACKNOWLEDGMENTS
We thank Dr. Brenda Temple for her help in depositing the X-ray crystallographic structure to the Protein Data Bank (PDB).
Supporting Information
Experimental details and characterization of all compounds and biological methods. The Supporting Information is available, in the online
version, at
ABBREVIATIONS
TAM, Tyro3-Axl-Mer; AML, acute myeloid leukemia; MCE, microfluidic capillary electrophoresis; IC50, half maximal inhibitory concen-
tration; pMerTK, phosphorylated Mer tyrosine kinase; ELISA, enzyme-linked immunosorbent assay; EC50, half maximal effective concen-
tration.
REFERENES
[1] Giordanetto, F.; Kihlberg, J. J Med Chem 2014, 57 (2), 278-295.
[2] Rosenquist, Å.; Samuelsson, B.; Johansson, P.-O.; Cummings, M. D.; Lenz, O.; Raboisson, P.; Simmen, K.; Vendeville, S.; de Kock, H.;
Nilsson, M. J Med Chem 2014, 57, 1673.
[3] De Rosa, M.; Unge, J.; Motwani, H. V.; Rosenquist, Å.; Vrang, L.; Wallberg, H.; Larhed, M. J Med Chem 2014, 57, 6444.
[4] Heinis, C. Nat Chem Biol 2014, 10, 696-698.
[5] Breslin, H. J.; Lane, B. M.; Ott, G. R.; Ghose, A. K.; Angeles, T. S.; Albom, M. S.; Cheng, M.; Wan, W.; Haltiwanger, R. C.; Wells-Knecht,
K. J.; Dorsey, B. D. J Med Chem 2012, 55 (1), 449-464.
[6] William, A. D.; Lee, A. C. H.; Goh, K. C.; Blanchard, S.; Poulsen, A.; Teo, E. L.; Nagaraj, H.; Lee, C. P.; Wang, H.; Williams, M.; Sun, E.
T.; Hu, C.; Jayaraman, R.; Pasha, M. K.; Ethirajulu, K.; Wood, J. M.; Dymock, B. W. J Med Chem 2012, 55 (1), 169-196.
[7] William, A. D.; Lee, A. C. H.; Poulsen, A.; Goh, K. C.; Madan, B.; Hart, S.; Tan, E.; Wang, H.; Nagaraj, H.; Chen, D.; Lee, C. P.; Sun, E. T.;
Jayaraman, R.; Pasha, M. K.; Ethirajulu, K.; Wood, J. M.; Dymock, B. W. J Med Chem 2012, 55 (6), 2623-2640.
[8] Graham, D. K.; Dawson, T. L.; Mullaney, D. L.; Snodgrass, H. R.; Earp, H. S. Cell Growth Differ 1994, 5 (6), 647-657.
[9] Graham, D. K.; DeRyckere, D.; Davies, K. D.; Earp, H. Cancer 2014, 14 (12), 769-785.
This article is protected by copyright. All rights reserved.

10.1002/cmdc.201600589
ChemMedChem
[10] Liu, J.; Yang, C.; Simpson, C.; DeRyckere, D.; Van, D. A.; Miley, M. J.; Kireev, D.; Norris-Drouin, J.; Sather, S.; Hunter, D.; Korboukh, V.
K.; Patel, H. S.; Janzen, W. P.; Machius, M.; Johnson, G. L.; Earp, H. S.; Graham, D. K.; Frye, S. V.; Wang, X. ACS Med. Chem. Lett. 2012, 3, 129-
134.
[11] Liu, J.; Zhang, W.; Stashko, M. A.; Deryckere, D.; Cummings, C. T.; Hunter, D.; Yang, C.; Jayakody, C. N.; Cheng, N.; Simpson, C.; Norris-
Drouin, J.; Sather, S.; Kireev, D.; Janzen, W. P.; Earp, H. S.; Graham, D. K.; Frye, S. V.; Wang, X. Eur J Med Chem 2013, 65, 83-93.
[12] Zhang, W.; McIver, A. L.; Stashko, M. A.; Deryckere, D.; Branchford, B. R.; Hunter, D.; Kireev, D.; Miley, M. J.; Norris-Drouin, J.; Stewart,
W. M.; Lee, M.; Sather, S.; Zhou, Y.; Di Paola, J. A.; Machius, M.; Janzen, W. P.; Earp, H. S.; Graham, D. K.; Frye, S. V.; Wang, X. J Med Chem
2013, 56 (23), 9693-9700.
[13] Zhang, W.; Zhang, D.; Stashko, M. A.; DeRyckere, D.; Hunter, D.; Kireev, D.; Miley, M. J.; Cummings, C.; Lee, M.; Norris-Drouin, J.;
Stewart, W. M.; Sather, S.; Zhou, Y.; Kirkpatrick, G.; Machius, M.; Janzen, W. P.; Earp, H. S.; Graham, D. K.; Frye, S. V.; Wang, X. J Med Chem
2013, 56 (23), 9683-9692.
[14] Zhang, W.; DeRyckere, D.; Hunter, D.; Liu, J.; Stashko, M. A.; Minson, K. A.; Cummings, C. T.; Lee, M.; Glaros, T. G.; Newton, D. L.;
Sather, S.; Zhang, D.; Kireev, D.; Janzen, W. P.; Earp, H. S.; Graham, D. K.; Frye, S. V.; Wang, X. J Med Chem 2014, 57 (16), 7031-7041.
[15] Taylor, S. J.; Dagger, S. A.; Thien, C. B.; Wikstrom, M. E.; Langdon, W. Y. Blood 2012, 120 (19), 4049-4057.
[16] Knapper, S.; Burnett, A. K.; Littlewood, T.; Kell, W. J.; Agrawal, S.; Chopra, R.; Clark, R.; Levis, M. J.; Small, D. Blood 2006, 108 (10),
3262-3270.
[17] Wang, X.; Liu, J.; Zhang, W.; Stashko, M. A.; Nichols, J.; Miley, M. J.; Norris-Drouin, J.; Chen, Z.; Machius, M.; DeRyckere, D.; Wood,
E.; Graham, D. K.; Earp, H. S.; Kireev, D.; Frye, S. V., ACS Med Chem Lett 2016, 7 (12), 1044-1049.
[18] Pommereau, A.; Pap, E.; Kannt, A. J Biomol Screen 2004, 9 (5), 409-416.
[19] Dunne, J.; Reardon, H.; Trinh, V.; Li, E.; Farinas, J. Assay Drug Dev Technol 2004, 2 (2), 121-129.
[20] Bernasconi, P.; Chen, M.; Galasinski, S.; Popa-Burke, I.; Bobasheva, A.; Coudurier, L.; Birkos, S.; Hallam, R.; Janzen, W. P. J Biomol Screen
2007, 12 (7), 972-982.
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