New small molecule inhibitors of histone methyl transferase DOT1L with a nitrile as a non-traditional replacement for heavy halogen atoms

Article abstract of DOI:10.1016/j.bmcl.2016.07.041

A number of new nucleoside derivatives are disclosed as inhibitors of DOT1L activity. SARs established that DOT1L inhibition could be achieved through incorporation of polar groups and small heterocycles at the 5-position (5, 6, 12) or by the application of alternative nitrogenous bases (18). Based on these results, CN-SAH (19) was identified as a potent and selective inhibitor of DOT1L activity where the polar 5-nitrile group was shown by crystallography to bind in the hydrophobic pocket of DOT1L. In addition, we show that a polar nitrile group can be used as a non-traditional replacement for heavy halogen atoms.

Full text of DOI:10.1016/j.bmcl.2016.07.041

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
New small molecule inhibitors of histone methyl transferase DOT1L
with a nitrile as a non-traditional replacement for heavy halogen
atoms
Sophie S. Spurr a, Elliott D. Bayle a, Wenyu Yu b, Fengling Li b, Wolfram Tempel b, Masoud Vedadi b,c
,
Matthieu Schapira b,c, Paul V. Fish a,
⇑
a UCL School of Pharmacy, University College London, 29-39 Brunswick Square, London WC1N 1AX, UK
b Structural Genomics Consortium, University of Toronto, 101 College Street, MaRS Centre, South Tower, Toronto MG5 1L7, Canada
c Department of Pharmacology and Toxicology, University of Toronto, Toronto MG5 1A8, Canada
a r t i c l e i n f o
a b s t r a c t
Article history:
A number of new nucleoside derivatives are disclosed as inhibitors of DOT1L activity. SARs established
that DOT1L inhibition could be achieved through incorporation of polar groups and small heterocycles
at the 5-position (5, 6, 12) or by the application of alternative nitrogenous bases (18). Based on these
results, CN-SAH (19) was identified as a potent and selective inhibitor of DOT1L activity where the polar
5-nitrile group was shown by crystallography to bind in the hydrophobic pocket of DOT1L. In addition,
we show that a polar nitrile group can be used as a non-traditional replacement for heavy halogen atoms.
Ó 2016 Elsevier Ltd. All rights reserved.
Received 29 June 2016
Revised 18 July 2016
Accepted 19 July 2016
Available online xxxx
Keywords:
Protein methyltransferase
DOT1L inhibitors
Toyocamycin
Bioisostere
Cyano-deaza-SAH
DOT1L is a protein methyltransferase known to methylate
lysine 79 of histone 3 (H3K79), an epigenetic mark associated with
transcriptionally active genes.1 Chromosomal translocations recur-
rent in leukemia result in oncogenic fusions between MLL and a
variety of genes, such as AF4 or AF9, that recruit DOT1L to aberrant
genomic loci, and transcriptional activation of leukemic genes.2
Small molecule inhibitors of DOT1L activity can reverse the onco-
genic reprogramming of MLL-rearranged leukemia,3,4 and the
first-in-class DOT1L inhibitor pinometostat (EPZ-5676; Epizyme)
has progressed to Phase I clinical trials for the treatment of pedi-
atric patients with relapsed/refractory leukemias bearing a rear-
rangement of the MLL gene.5
tion of DOT1L can also be achieved by functionalizing the adeno-
sine ring of S-adenosyl-L-homocysteine (SAH), the reaction
product of SAM (Fig. 1). Bromo-deaza-SAH (Br-SAH) is a potent
and selective inhibitor of DOT1L activity with a bromine atom that
occupies a hydrophobic cavity specific to DOT1L, but this com-
pound is inactive in cell based systems, probably due to poor cell
penetration.10 Modification of EPZ004777 by introduction of a bro-
mine atom on the adenosine ring gave SGC0946 and both com-
pounds selectively kill human cord blood cells transformed with
an MLL-AF9 fusion oncogene.7 However, the poor pharmacokinetic
properties of EPZ004777 precluded conventional dosing methods
for efficacy studies in a mouse xenograft model of MLL.3 Nonclini-
cal in vivo pharmacokinetics of EPZ-5676 in mouse, rat and dog
showed moderate to high clearance and low oral bioavailability,11
and so EPZ-5676 was administered by continuous intravenous
infusion in the Phase I clinical trial.5,11
As part of our research efforts to discover new inhibitors of
DOT1L activity with improved pharmacokinetic properties, we
adopted a strategy to identify novel functional groups that would
favourably exploit the hydrophobic cavity adjacent to the adenine
binding site of SAH in DOT1L. In addition, we wished to avoid the
use of heavy halogen atoms as they confer higher lipophilicity to
drug molecules which tends to be detrimental to drug-like proper-
ties. In this Letter, we disclose cyano-deaza-SAH (19, CN-SAH) as a
Most current DOT1L inhibitors, such as EPZ004777 and EPZ-
5676, compete with the methyl donating cofactor S-adenosyl-L-
methionine (SAM). These SAM analogues lock a conformationally
dynamic loop of the enzyme into an inactive state.6,7 In a variation
on this theme, recently reported structurally novel inhibitors of
DOT1L also antagonize the formation of an enzymatically active
state, but only partially occupy the cofactor site.8,9 Selective inhibi-
⇑
Corresponding author at present address: Alzheimer’s Research UK UCL Drug
Discovery Institute, The Cruciform Building, University College London, Gower
Street, London WC1E 6BT, UK. Tel.: +44 (0)20 7679 6971.
E-mail address: p.fish@ucl.ac.uk (P.V. Fish).
http://dx.doi.org/10.1016/j.bmcl.2016.07.041
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Spurr, S. S.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.07.041

2
S. S. Spurr et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
NHR
NH2
N
Me
Me
X
N
X
N
N
H
N
H
N
HO2C
S
N
N
N
O
O
NH2
Me
Me
O
Me
HO OH
HO OH
SAH, X = N, R = H: DOT1L IC50 = 600 nM a
Me-SAH, X = N, R = Me: DOT1L Ki = 290 nM b
Br-SAH, X = C-Br, R = H: DOT1L IC50 = 77 nM a
EPZ004777, X = C-H: DOT1L Ki = 0.3 nM c
SGC0946, X = C-Br: DOT1L IC50 = 0.3 nM d
FED1, X = N: DOT1L IC50 = 2.9 nM d
Cmpd 10, X = C-Cl: DOT1L IC50 = 28.2 nM e
Figure 1. Structures of DOT1L inhibitors. aYu et al.10
;
bYao et al.12
;
cDaigle et al.3, Basavapathruni et al.6; dYu et al.7; eYi et al.13
.
potent and selective inhibitor of DOT1L activity that incorporates a
polar nitrile group which binds in this hydrophobic pocket.
Previously, a screen of a library of 3120 kinase inhibitors had
identified 5-iodotubercidin (3, 5-ITC) as an inhibitor of DOT1L
tubercidin template (2) was investigated as a model system with
the objective of transferring preferred structure–activity-relation-
ships (SARs) into more relevant templates for potential use as bio-
chemical and structural tools, chemical probes with cell activity,
and ultimately to chemical leads suitable for convenient dosing
in preclinical in vivo efficacy studies.
activity (IC50 18.2 lM) which was subsequently optimised to Br-
SAH (IC50 0.077
l
M).10 We adopted a related approach where the
The nucleoside target compounds were either purchased from
commercial sources (1–6, 13–15) or prepared using short synthetic
sequences (7–12, 16–18) as described in detail in the Supplemen-
tary Material. The DOT1L inhibitory activity of target compounds
1–19 (Table 1) was evaluated by the assay conditions as reported
by Yu et al. with minor modifications (see: Supplementary
Material).10
The SARs were initially focused on exploring the substitution at
the 5-position (tubercidin numbering used throughout), which
presents this substituent towards the hydrophobic pocket adjacent
to the adenine binding site of SAH in DOT1L (PDB code 3QOX); this
interaction has been successfully exploited by 5-ITC (3: X = I) (PDB
code 3UPW) and Br-SAH (PDB code 3SX0).10 Adenosine (1) and
tubercidin (2: X = H) were very weakly active with 20–40% inhibi-
Table 1
Compound
DOT1L
% Inhibition at
200 or 250
Ma
IC50 (l
M)a
l
1
2
3
4
5
6
7
8
26 4% I @ 200
41 4% I @ 200
l
l
M
M
7
2c
<20% I @ 200 lM
44 10
26
7
ca. 52 5% I @ 200
<20% I @ 200
l
Mb
ca. 200b
41 12
tion at 200 lM respectively which supported the benefits of a
9
lM
group at this 5-position. Toyocamycin (5: X = CN), a bacterial
metabolite with a diverse range of activities,14 showed promising
activity as did sangivamycin (6: X = CONH2). These results were
10
11
12
13
14
15
16
17
18
19
44 5% I @ 200
42 1% I @ 250
l
l
M
M
43
8
<20% I @ 200
<20% I @ 200
<20% I @ 200
lM
lM
lM
somewhat unexpected as replacing a lipophilic iodine atom (
p
+1.12) with a polar nitrile (
p
ꢀ0.57) or primary carboxamide group
ꢀ1.49) is not a traditional bioisostere transformation.15,16 The
ca. 50 4% I @ 200
<20% I @ 200
l
Mb
ca. 200b
22
(p
lM
aromatic chloride to nitrile transformation has been reviewed by
Jones et al.16b whereas there are far fewer reports of the
corresponding aromatic iodide to nitrile substitution in medicinal
chemistry design.17,18 However, in the context of this work, these
results demonstrated the hydrophobic pocket could accommodate
both lipophilic and polar groups assuming the binding mode was
retained (vide infra).
2
0.026 0.005
Assay conditions: DOT1L (0.2 nM), [3H]-SAM (0.7
l
M), chicken nucleosome
a
(0.1
general, IC50s were determined side by side from 11-point dose response curves
with test concentrations from 0.20 to 200 M, except 11 (0.50–500 M) and 19
(0.01–10 M). Experiments were performed in triplicate with data presented as the
mean SD.
lM), buffer: 20 mM Tris pH8, 5 mM DTT, 10 mM MgCl2, 0.01% Triton X-100. In
l
l
l
Further SARs with synthetic analogues showed 5-fluorine 7, 5-
phenyl 10 or 5-benzyl 11 derivatives were similar in activity to
tubercidin. The 5-ethyne 8 was equipotent to toyocamycin (5)
b
Incomplete concentration-inhibition S-curve.
Data are presented for comparison in a common assay format. Literature values
for 5-ITC (3): DOT1L IC50 = 18.2
c
lM. See Ref. 10.
N
NH2
N
NH2
5
Br
N
6
HO2C
S
HO2C
S
N
5'
N
N
N
1
7
O
O
NH2
NH2
Δclog P = -1.4
HO OH
HO OH
19: CN-SAH
Br-SAH
Figure 2. Design of CN-SAH (19).
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S. S. Spurr et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
although extending the alkyne further with a 5-(2-cyclopropy-
lethyne) group 9 lost activity (<20% I @ 200 M). The use of an oxa-
these polar groups could then be applied to identify new DOT1L
chemical probes and would improve the ‘drug-like’ properties of
this template by significantly lowering the overall lipophilicity.19
The synthesis of CN-SAH (19) is described in Scheme 1.20
This 8-step sequence follows published procedures to related
compounds with minor modifications to improve yields/selec-
tivities or the substitution of highly toxic reagents for safer
alternatives.10,21 The preferential N7 glycosylation of pyrrolo
[2,3-d]pyrimidine 22 with furanose 23 was achieved with
1,1,1,3,3,3-hexamethyldisilazane (HMDS) as described recently
by Kim et al. to give 24.22 Reduction of the C6-Br by catalytic
transfer hydrogenation gave 25 and then deprotection of the
benzoates afforded toyocamycin (5).21 Conversion of the 50-OH
to the corresponding 50-chloride 26 activated the 50 position
of the sugar ready for the introduction of the homocysteine
moiety. This followed the procedure of Yu et al.10 except that
l
zole 12 as a surrogate for the p-carboxamide of 6 showed that a
small heterocyclic group could be accommodated with retention
of activity.
Limited SAR of the ribose showed that the 20-hydroxy was
important for binding (13 v 3: X = I) as 13 was inactive (<20% I @
200
activity as 14 was also inactive (<20% I @ 200
l
M). The addition of a 20b-Me (14 v 2: X = H) failed to improve
lM) albeit with only
the inferior 5-H substituent. Modification of the nitrogenous base
proved to be more successful. Although 3-deazaadenosine (15)
was inactive (<20% I @ 200 lM) confirming the preference of the
N atom at this position (15 v 1), modification of the base at the
6-position by substitution of the 6-CH for a 6-N demonstrated that
the pyrazolo[3,4-d]pyrimidine could be of comparable potency to
the pyrrolo[2,3-d]pyrimidine (18 v 3: X = I cf. 17 v 2: X = H). DOT1L
inhibitory activity was again superior when combined with the 5-
iodo substituent as 18 was one of the most potent inhibitors from
this model system.
t-butyl Boc-L-homocysteinate (27) was used which greatly
improved the efficiency of the reactions even with the addi-
NH2
NH2
N
X
5
N
N
6
HO
HO
5'
N
N
N
N
1
O
O
2'
OH
OH
OH
OH
X =
H
1
adenosine
N
2
3
tubercidin
I
5-iodotubercidin
toyocamycin
sangivamycin
Me
N
5
CN
H2N
N
6
CONH2
F
7
8
C≡CH
C≡CcPr
Ph
HO
N
N
9
O
10
11
12
4-PMB
OHOH
2-oxazole
4
triciribine
NH2
NH2
N
X
N
N
N
HO
HO
O
N
N
O
R'
OH
R
OH
OH
13 X = I; R = R’ = H
15 3-deazaadenosine
14 X = H; R = OH; R’ = Me
N
NH2
N
X
NH2
N
N
HO
Br
N
HO
N
O
N
N
O
OH
OH
OH
OH
16
17 X = H
18 X = I
Based on these results, we selected toyocamycin (5) with the 5-
nitrile as the starting point for the design of the biochemical tool
cyano-deaza-SAH (19, CN-SAH) as proof-of-principle to deliver
potent and selective inhibitors of DOT1L that incorporate polar
groups that bind in the hydrophobic pocket (Fig. 2). We anticipated
tional step to simultaneously deprotect the homocysteine acid
and amine groups.
CN-SAH (19) was shown to be a potent inhibitor of DOT1L with
an IC50 value of 26 nM (Table 1). To establish the binding mode, we
solved the structure of CN-SAH in complex with DOT1L (2.3 Å, PDB
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4
S. S. Spurr et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
N
N
N
N
N
N
N
N
NH2
N
NH2
N
a
b
c
Br
Br
Br
NH2
BzO
N
H
N
H
N
N
N
O
20
21
22
BzO OBz
24
N
N
N
NH2
NH2
NH2
N
d
e
f
N
N
N
N
BzO
HO
Cl
N
N
N
O
N
O
O
BzO OBz
HO OH
HO OH
25
5
26
N
N
NH2
NH2
N
5
g
h
O
N
6
S
HO2C
S
5'
tBuO
N
N
N
N
7
O
O
1
NHBoc
NH2
HO OH
HO OH
28
19: CN-SAH
Scheme 1. Synthesis of CN-SAH (19). Reagents and conditions: (a) 33% HBr in AcOH, acetone, EtOAc, 0 °C–rt, 3 h, 60%; (b) formamidine acetate, 2-ethoxyethanol, reflux, 24 h,
36%; (c) (i) HMDS, TMSCl, reflux 18 h, then (ii) evaporate and add MeCN, then (iii) b- -Ribofuranose-1-acetate-2,3,5-tribenzoate (23), TMSOTf, reflux, 3 h, 49%; (d) ammonium
D
formate, 5 mol % Pd/C, EtOH, reflux, 3 h, 65%; (e) 7 M NH3 in MeOH, 0 °C–rt, 12 h, 60%; (f) SOCl2, DMPU, 0 °C, 5 h, 50%; (g) (i) KI, K2CO3, DMF, rt, 5 min, then (ii) t-Butyl Boc-
homocysteinate (27), 80 °C, 5 h, 75%; (h) 4 M HCl in dioxane, 0 °C, 7 h, 82%.
L-
Figure 3. Co-crystal structures of CN-SAH (19) (PDB code 5JUW, left) and Br-SAH (PDB code 3SX0, right) in DOT1L. Images generated with PyMOL.
code 5JUW) (Fig. 3). As expected, the structure is nearly identical to
DOT1L in complex with SAM (PDB code 1NW3) and Br-SAH (PDB
code 3SX0). The activation loop of DOT1L is folded on the cofactor
mimetic, in a conformationally active state. The nitrogen atom of
the 5-nitrile of CN-SAH occupies the hydrophobic pocket com-
posed of V249, L224, P133, and F245 as previously exploited by
the bromine of Br-SAH.
CN-SAH (19) was confirmed to be a potent inhibitor of DOT1L
(IC50 = 13 nM) by independent evaluation (Table 2 and Supplemen-
tary Material). In addition, CN-SAH was 17-times more potent than
SAH against DOT1L which, like the bromine atom of Br-SAH, shows
the nitrile group of CN-SAH better exploits the hydrophobic pocket
of DOT1L. CN-SAH was then screened against a panel of represen-
tative protein lysine (PKMT), protein arginine (PRMT) and DNA
(DNMT) methyltransferases to establish the selectivity profile
(Table 2). CN-SAH was inactive at concentrations up to 100
lM
against the PKMTs EZH2 (as part of PRC2 complex), SUV39H2
and SETD7 although there was very weak activity for G9a.
CN-SAH showed inhibitory activity for both PRMT3 and the PRMT5
complex but with acceptable selectivity at 66- and 180-fold
respectively. The selectivity of CN-SAH over these PRMTs is
modestly improved when compared to BrSAH (both 30-fold)
which reflects improved affinity for DOT1L rather than weaker
activity for the PRMTs. CN-SAH was shown to be an inhibitor of
DNMT1 with only 11-fold selectivity and would need to be
improved.
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S. S. Spurr et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
5
Table 2
References and notes
Inhibition of DOT1L activity and MTase selectivity profiles of CN-SAH (19), Br-SAH
and SAH
1. For a review, see: (a) Nguyen, A. T.; Zhang, Y. Genes Dev. 2011, 25, 1345; While
DOT1L is believed to be the main enzyme to write this epigenetic mark
associated with transcriptionally active genes, an isoform of WHSC1 was
recently reported to also methylate H3K79; see (b) Woo Park, J.; Kim, K. B.;
Kim, J. Y.; Chae, Y. C.; Jeong, O. S.; Seo, S. B. Sci. Rep. 2015, 5, 12485.
2. Bernt, K. M.; Zhu, N.; Sinha, A. U.; Vempati, S.; Faber, J.; Krivtsov, A. V.; Feng, Z.;
Punt, N.; Daigle, A.; Bullinger, L.; Pollock, R. M.; Richon, V. M.; Kung, A. L.;
Armstrong, S. A. Cancer Cell 2011, 20, 66.
3. Daigle, S. R.; Olhava, E. J.; Therkelsen, C. A.; Majer, C. R.; Sneeringer, C. J.; Song,
J.; Johnston, L. D.; Scott, M. P.; Smith, J. J.; Xiao, Y. H.; Jin, L.; Kuntz, K. W.;
Chesworth, R.; Moyer, M. P.; Bernt, K. M.; Tseng, J. C.; Kung, A. L.; Armstrong, S.
A.; Copeland, R. A.; Richon, V. M.; Pollock, R. M. Cancer Cell 2011, 20, 53.
4. Daigle, S. R.; Olhava, E. J.; Therkelsen, C. A.; Basavapathruni, A.; Jin, L.; Boriack-
Sjodin, P. A.; Allain, C. J.; Klaus, C. R.; Raimondi, A.; Porter Scott, M.; Waters, N.
J.; Chesworth, R.; Moyer, M. P.; Copeland, R. A.; Richon, V. M.; Pollock, R. M.
Blood 2013, 122, 1017.
MTase (h)
CN-SAH (19)
IC50
M)a
Br-SAH
IC50
M)b
SAH
IC50
(
l
(
l
(lM)
DOT1L
DNMT1
PRMT3
PRMT5
G9a
SETD7
SUV39H2
EZH2
0.013
0.15
0.87
2.4
0.077
1.9
2.3
0.22
0.20
2.0
0.20
4.1
47
2.3
99
No inhibition
No inhibition
No inhibition
No inhibition
No inhibition
No inhibition
No inhibition
22
65
a
Screened at Eurofins Cerep (France). In general, IC50s were determined from 8-
point dose response curves (test concentrations: 30 nM to 100 M) except DOT1L
which required a lower dynamic range (0.1 nM to 1 M). Data is the mean of two
l
l
5. Phase
1 Dose Escalation and Expanded Cohort Study of EPZ-5676 in the
replicates. SAH was used as a reference compound throughout and data are pre-
sented for comparison in a common assay format. See: Supplementary Material.
Treatment of Pediatric Patients With Relapsed/Refractory Leukemias Bearing a
Rearrangement of the MLL Gene. ClinicalTrials.gov identifier: NCT02141828.
6. Basavapathruni, A.; Jin, L.; Daigle, S. R.; Majer, C. R. A.; Therkelsen, C. A.; Wigle,
T. J.; Kuntz, K. W.; Chesworth, R.; Pollock, R. M.; Scott, M. P.; Moyer, M. P.;
Richon, V. M.; Copeland, R. A.; Olhava, E. J. Chem. Biol. Drug Des. 2012, 80, 971.
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Engen, J. R.; Griffin, C.; Aman, A.; Wienholds, E.; Li, F. L.; Pineda, J.; Estiu, G.;
Shatseva, T.; Hajian, T.; Al-awar, R.; Dick, J. E.; Vedadi, M.; Brown, P. J.;
Arrowsmith, C. H.; Bradner, J. E.; Schapira, M. Nat. Commun. 2012, 3, 1218.
8. Chen, C.; Zhu, H.; Stauffer, F.; Caravatti, G.; Vollmer, S.; Machauer, R.; Holzer, P.;
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b
Ref. 10.
In summary, we have designed and evaluated a number of new
nucleoside derivatives as inhibitors of DOT1L activity. SARs estab-
lished that DOT1L inhibition could be achieved through incorpora-
tion of polar groups and small heterocycles at the 5-position (5, 6,
12) or by the application of alternative nitrogenous bases (18).
Based on these results, CN-SAH (19) was identified as a potent
and selective inhibitor of DOT1L activity where the polar 5-nitrile
group was shown by crystallography to bind in the hydrophobic
pocket of DOT1L. The next phase of these studies will be to develop
CN-SAH into a chemical probe that is active in cell-based models
and with good oral activity in rodent models of disease.
11. Nonclinical pharmacokinetics and metabolism of EPZ-5676: (a)
Basavapathruni, A.; Olhava, E. J.; Daigle, S. R.; Therkelsen, C. A.; Jin, L.;
Boriack-Sjodin, P. A.; Allain, C. J.; Klaus, C. R.; Raimondi, A.; Porter Scott, M.;
Dovletoglou, A.; Richon, V. M.; Pollock, Roy M.; Copeland, R. A.; Moyer, M. P.;
Chesworth, R.; Pearson, P. G.; Waters, N. J. Biopharm. Drug Dispos. 2014, 35,
237; However, the plasma clearance of EPZ-5676 was recently shown to be
markedly lower in human compared to preclinical species, see: (b) Smith, S. A.;
Gagnon, S.; Waters, N. J. Xenobiotica 2016. http://dx.doi.org/10.3109/
00498254.2016.1173265.
12. Yao, Y.; Chen, P.; Diao, J.; Cheng, G.; Deng, L.; Anglin, J. L.; Prasad, B. V. V.; Song,
Y. J. Am. Chem. Soc. 2012, 134, 17834.
13. Yi, J. S.; Federation, A. J.; Qi, J.; Dhe-Paganon, S.; Hadler, M.; Xu, X.; St Pierre, R.;
Varca, A. C.; Wu, L.; Marineau, J. J.; Smith, W. B.; Souza, A.; Chory, E. J.;
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14. Tolman, R. L.; Robons, R. K.; Townsend, L. B. J. Am. Chem. Soc. 1969, 91, 2102.
15. Hydrophobic fragmental constants (p) are used as a measure of lipophilicity.
16. For reviews on bioisosteres, see: (a) Meanwell, N. A. J. Med. Chem. 2011, 54,
2529; (b) Jones, L.; Summerhill, N.; Swain, N.; Mills, J. Med. Chem. Commun.
2010, 1, 309.
17. Bell, A. S.; Campbell, S. F.; Morris, D. S.; Roberts, D. A.; Stefaniak, M. H. J. Med.
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Acknowledgments
We are grateful to the UCL Drug Discovery PhD Studentship
Programme (to S.S.S.) for financial support. HRMS were recorded
at the EPSRC UK National Mass Spectrometry Facility (NMSF) at
Swansea University, UK. We also acknowledge the contribution
of Prima Patel and Adil Butt for their assistance, John R. Walker
for suggestions toward crystallographic model refinement and
Peter J. Brown for his support.
The SGC is a registered charity (number 1097737) that receives
funds from AbbVie, Bayer Pharma AG, Boehringer Ingelheim,
Canada Foundation for Innovation, Eshelman Institute for Innova-
tion, Genome Canada through Ontario Genomics Institute, Innova-
tive Medicines Initiative (EU/EFPIA) [ULTRA-DD grant no. 115766],
Janssen, Merck & Co., Novartis Pharma AG, Ontario Ministry of Eco-
nomic Development and Innovation, Pfizer, São Paulo Research
Foundation-FAPESP, Takeda, and the Wellcome Trust. Results
shown in this report are derived from work performed at Argonne
National Laboratory, Structural Biology Center at the Advanced
Photon Source. Argonne is operated by UChicago Argonne, LLC,
for the U.S. Department of Energy, Office of Biological and Environ-
mental Research under contract DE-AC02-06CH11357.
18. For a review of aromatic iodide to nitrile substitution in synthetic chemistry,
see: Ellis, G. P.; Romney-Alexander, T. M. Chem. Rev. 1987, 87, 779.
19. For a review on the benefits of reducing lipophilicity of drug candidates, see:
Leeson, P. D.; Springthorpe, B. Nat. Rev. Drug Disc. 2007, 6, 881.
20. (S)-2-amino-4-((((2S,3S,4R,5R)-5-(4-amino-5-cyano-7H-pyrrolo[2,3-d]pyrimidin-
7-yl)-3,4-dihydroxytetrahydrofuran-2-yl)methyl)thio)butanoic acid dihydrochloride
salt was prepared as an off-white amorphous solid (45 mg) (19: CN-SAH): m.p.
25
130–5 °C (dec.); [
a]
D
+16 (c 0.5, MeOH); IR mmax (neat) 2922.36 (br), 2232.60
(s), 1665.52 (s), 1518.04 (s), 1427.84 (s), 1047.29 (s) cmꢀ1 1H NMR (500 MHz,
;
CD3OD) d: 8.51 (1H, s, 2/6-CH); 8.48 (1H, s, 2/6-CH); 6.28 (1H, d, J = 4.4 Hz, 10-
CH); 4.52 (1H, t, J = 4.6 Hz, 20-CH); 4.28–4.23 (2H, m, 30-CH, 40-CH); 4.15 (1H, t,
Supplementary data
J = 6.1 Hz, Cys-a c-
-CH); 3.07–2.99 (m, 2H, 50-CH2); 2.81 (2H, t, J = 7.4 Hz, Cys-
CH2); 2.32–2.25 (1H, m, Cys-b-CH); 2.20–2.13 (1H, m, Cys-b-CH); 13C NMR
(125 MHz, CD3OD) d: 171.4 (C@O), 152.5, 149.7, 145.8, 135.7, 113.9 (CN),
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2016.07.
041. These data include: Sources of compounds 1–6, 13–15 and
synthetic schemes for compounds 7–12, 16–18; 1H and 13C NMR
data for CN-SAH (19); materials and methods for the DOT1L IC50
determinations; crystal structure data and model statistics.
103.0, 90.7 (C10), 88.7, 85.6 (C40), 76.0 (C20), 73.8 (C30), 52.7 (Cys-
a-CH), 35.0
(C50), 31.6 (Cys-b-CH2), 29.0 (Cys- -CH2); LCMS m/z: 409.2 ([M+H]+; 92.5%
c
pure; RT 0.85 min); HRMS (m/z HNESP) for C16H21N6O5S [M+H]+ calc. 409.1289,
observed 409.1284.
21. Leonard, P.; Ingale, S. A.; Ding, P.; Ming, X.; Seela, F. Nucleosides, Nucleotides
Nucleic Acids 2009, 28, 678.
22. Kim, Y.; Kwon, S.; Bae, I.; Kim, B. Tetrahedron Lett. 2013, 54, 5484.
Please cite this article in press as: Spurr, S. S.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.07.041