Optimisation of a 5-[3-phenyl-(2-cyclic-ether)-methyl-ether]-4-aminopyrrolopyrimidine series of IGF-1R inhibitors

Article abstract of DOI:10.1016/j.bmcl.2016.02.075

Taking the pyrrolopyrimidine derived IGF-1R inhibitor NVP-AEW541 as the starting point, the benzyl ether back-pocket binding moiety was replaced with a series of 2-cyclic ether methyl ethers leading to the identification of novel achiral [2.2.1]-bicyclic ether methyl ether containing analogues with improved IGF-1R activities and kinase selectivities. Further exploration of the series, including a fluorine scan of the 5-phenyl substituent, and optimisation of the sugar-pocket binding moiety identified compound 33 containing (S)-2-tetrahydrofuran methyl ether 6-fluorophenyl ether back-pocket, and cis-N-Ac-Pip sugar-pocket binding groups. Compound 33 showed improved selectivity and pharmacokinetics compared to NVP-AEW541, and produced comparable in vivo efficacy to linsitinib in inhibiting the growth of an IGF-1R dependent tumour xenograft model in the mouse.

Full text of DOI:10.1016/j.bmcl.2016.02.075

Bioorganic & Medicinal Chemistry Letters xxx (2016) xxx–xxx
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Optimisation of a 5-[3-phenyl-(2-cyclic-ether)-methyl-ether]-4-
aminopyrrolopyrimidine series of IGF-1R inhibitors
b
a
a
a
b
Robin A. Fairhurst a, , Thomas H. Marsilje , Stefan Stutz , Andreas Boos , Michel Niklaus , Bei Chen ,
Songchun Jiang b, Wenshuo Lu b, Pascal Furet a, Clive McCarthy a, Frédéric Stauffer a, Vito Guagnano a,
Andrea Vaupel a, Pierre-Yves Michellys b, Christian Schnell a, Sébastien Jeay a
⇑
a Novartis Institutes for BioMedical Research, CH-4002 Basel, Switzerland
b Genomics Institute of the Novartis Research Foundation, 10675 John Jay Hopkins Drive, San Diego, CA 92121, USA
a r t i c l e i n f o
a b s t r a c t
Article history:
Taking the pyrrolopyrimidine derived IGF-1R inhibitor NVP-AEW541 as the starting point, the benzyl
ether back-pocket binding moiety was replaced with a series of 2-cyclic ether methyl ethers leading to
the identification of novel achiral [2.2.1]-bicyclic ether methyl ether containing analogues with improved
IGF-1R activities and kinase selectivities. Further exploration of the series, including a fluorine scan of the
5-phenyl substituent, and optimisation of the sugar-pocket binding moiety identified compound 33 con-
taining (S)-2-tetrahydrofuran methyl ether 6-fluorophenyl ether back-pocket, and cis-N-Ac-Pip sugar-
pocket binding groups. Compound 33 showed improved selectivity and pharmacokinetics compared to
NVP-AEW541, and produced comparable in vivo efficacy to linsitinib in inhibiting the growth of an
IGF-1R dependent tumour xenograft model in the mouse.
Received 21 January 2016
Revised 22 February 2016
Accepted 24 February 2016
Available online xxxx
Keywords:
Oncology
Kinase inhibitor
Insulin-like growth factor receptor-1
Ó 2016 Elsevier Ltd. All rights reserved.
The IGF-1/insulin family of growth factors is an evolutionally
conserved system which plays a crucial role in the growth and
development of many tissues and in the regulation of metabolism.
This system comprises three receptors of the receptor tyrosine
kinase family: the insulin-like growth factor-1 receptor (IGF-1R);
the insulin receptor (InsR) and the IGF-2/mannose 6-phosphate
receptor (M-6-PR). These receptors interact with three ligands
(insulin, IGF-1, and IGF-2), which also interact with six known
types of circulating IGF-binding proteins (IGFBP1–6).1 The IGF-1R
is widely expressed in many human tissues and cell types and is
highly homologous to the InsR. However, these two receptors have
distinct functions, since the IGF-1R controls apoptosis, cell growth,
and differentiation, while the InsR regulates primarily physiologi-
cal processes. Additionally, the ability of the highly homologous
IGF-1R and InsR to form hybrid receptors through hetero dimeriza-
tion further increases the complexity of the signalling system.2
Mitogenic activity of IGF-1 was first established in breast carci-
noma and the anti-tumour activity of an IGF-1R specific antibody
was first demonstrated now over a quarter of a century ago.3 Since
then, substantial population and preclinical data have all pointed
towards the IGF-1R pathway as an important regulator of tumour
cell biology, and as a result many groups have identified antibody
and low molecular weight inhibitors which have entered into mul-
tiple clinical studies exploring the safety and anti-tumour activity
of these agents. Although the results of these clinical trials have
been mixed, in particular when tested as a single agent, a growing
effort is indicating the potential for inhibiting the IGF-1R pathway
in combination with cytotoxic chemotherapy and various targeted
agents.4 Therefore, assessing optimal combination therapies could
represent new opportunities for the development of IGF-1R inhibi-
tors in a range of common cancer indications.
Of the numerous IGF-1R inhibitors that have been developed
since the link to cancer was established those targeting the kinase
activity, through binding within the adenosine triphosphate (ATP)
binding site, represent the most common approach taken.5 Of
these linsitinib is the most advanced compound which is currently
in multiple phase II studies, in a range of cancers, in combination
with other treatment modalities.6 Prior to the disclosure of linsi-
tinib workers at Novartis had described the identification of the
structurally related 5-phenyl-4-aminopyrrolopyrimidine deriva-
tive NVP-AEW541, Figure 1.7 This molecule has subsequently been
used extensively as an IGF-1R inhibitor tool compound in a num-
ber of preclinical investigations.8 In this letter we describe the fur-
ther optimisation of the 5-phenyl-4-aminopyrrolopyrimidine
series, to improve upon NVP-AEW541, utilising the observation
⇑
Corresponding author at present address: Novartis Institutes for Biomedical
Research, Global Discovery Chemistry, Novartis Pharma AG, Werk Klybeck,
Postfach, CH-4002 Basel, Switzerland. Tel.: +41 61 6964294; fax: +41 61
6966246.
E-mail address: robin.fairhurst@novartis.com (R.A. Fairhurst).
http://dx.doi.org/10.1016/j.bmcl.2016.02.075
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

2
R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
O
O
O
N
H2N
H2N
N
H2N
N
N
N
N
N
N
N
N
N
3
N
OH
NVP-AEW541
linsitinib
Figure 3. Structure of the 7-oxabicyclo[2.2.1]heptan-1-ylmethyl ether analogue 3.
Figure 1. Structures of NVP-AEW541 and linsitinib.
Phe1010 and Glu1050 in addition to the hydrogen bond with
Lys1033. Compound 3 docked into ATP-site of IGF-1R is shown in
Figure 4, and the model is based upon the X-ray structure of a cyclic
ether analogues with the closely related InsR (PDB code: 5HHW).9
To prepare analogues containing the 7-oxabicyclo[2.2.1]hep-
tan-1-ylmethylether moiety the convergent approach outlined in
Schemes 1 and 2 was developed in which the 5-phenyl pyrrolopy-
rimidine substituent was introduced via a Suzuki reaction in the
final phase of the synthesis.13 Preparation of the phenyl coupling
partner is shown in Scheme 1 and started from the mono-ketal
of cyclohexane-1,4-dione 4 which gave the vinyl alcohol 5 follow-
ing a sequence of methylenation, ketal deprotection and reduction.
Cyclisation of 5 through activation of the double bond with iodine
provided the iodide 6 which was then used to alkylate the phenol
leading to the key building block 7.
Scheme 2 shows the synthesis and reaction of the pyrrolopy-
rimidine coupling partner which starts with the annulation of 2-
(4,6-dichloropyrimidin-5-yl)acetaldehyde 8 upon reaction with
the benzoyl ester of cis-4-hydroxymethyl cyclobutylamine 914 to
give the pyrrolopyrimidine 10. Iodination of 10 produced the 5-
iodo analogue and subsequent displacement of the 4-chloro sub-
stituent with ammonia yielded 11 which readily underwent a
Suzuki reaction with 7 to give compound 12 with the key pharma-
cophore elements in place. Manipulation of the hydroxymethylcy-
clobutyl pyrrolopyrimidine 7-substituent via periodinane
oxidation followed by a reductive amination with the newly
formed aldehyde generated the targeted compound 3.
described in the preceding letter that 2-cyclic ether methyl ethers
can be used to replace the benzyl ether moiety.9
The switch to a 2-cyclic ether methyl ether sub-series, as
exemplified by the 2-tetrahydrofuran (THF) analogue 1 shown in
Figure 2, was envisioned as an opportunity to improve upon a
number of the limitations that had been identified for the benzyl
ether NVP-AEW541: cyclic ethers were anticipated to provide an
increase in aqueous solubility and enable an adequate level of sol-
ubility to be achieved without the need for a strongly basic group,
helping to reduce the cardiovascular risk associated with inhibition
of the human ether-à-go-go related gene (hERG) channel; in vitro
and in vivo metabolism studies indicated greatly reduced cleavage
of the aryl ether for the cyclic ethers,9,10 minimising the formation
of the phenol 2, which has been shown to be a multi-targeted
kinase inhibitor.11,12 Therefore, although NVP-AEW541 provided
satisfactory exposure to deliver efficacy in rodent xenograft models
and also showed a good kinase selectivity profile,7 at the onset of
the optimisation the 2-cyclic ether methyl ether sub-series was
seen as an opportunity to identify 5-phenyl-4-aminopyrrolopyrim-
idine analogues with improved pharmacokinetic (PK) and tolera-
bility/safety profiles.
To explore the 2-cyclic ether methyl ether structure activity
relationship (SAR) further, the observation that both enantiomers
of the 2-THF methyl ether showed similar high levels of activity
attracted our attention.9 In particular, for each enantiomer, mod-
elling indicated the cyclic ether oxygen atom makes the same
hydrogen bond with Lys1033. As a result, the remaining atoms of
each THF ring of the two enantiomers are positioned within the
IFG-1R ATP-pocket in orientations which suggested a 7-oxabicy-
clo[2.2.1]heptan-1-ylmethylether could be accommodated. Such
an achiral [2.2.1]-bicyclic ether moiety would then be postulated
to simultaneously make the hydrophobic contacts associated with
both 2-THF methyl ether enantiomers within the back-pocket
region. In addition to the potential for increasing potency, the fur-
ther filling of the relatively large back-pocket region of the IGF-1R
ATP binding site was anticipated to further improve the, already
high, kinase selectivity for the series.12 To explore this possibility
compound 3, the direct analogue of NVP-AEW541, was targeted
for synthesis and the structure is shown in Figure 3. Modelling sup-
ported the above hypothesis with the [2.2.1]-bicyclic ether making
hydrophobic contacts with residues Met1054, Phe1154, Phe1047,
The IGF-1R activities of the compounds were measured using a
biochemical assay and with cellular assays measuring p-IGF-1R in
HEK cells and IGF-1R dependent proliferation in Ba/F3 cells, data
are shown in Table 1.15 In line with the binding hypothesis, the
[2.2.1] bicyclic ether analogue 3 was found to be well tolerated
OH
O
O
H2N
N
H2N
N
N
N
N
N
N
N
1
2
Figure 4. Model of compound 3 docked into the ATP site of IGF-1R. Key hydrogen-
bonds are indicated with magenta lines: pyrrolopyrimidine N5 to M1082 NH 3.2 Å,
pyrrolopyrimidine 4-NH to E1080 C@O 2.9 Å, aryl ether to K1033 side-chain NH
3.0 Å, [2.2.1] bicyclic ether to K1033 side-chain NH 2.8 Å.
Figure 2. Structures of the (S)-tetrahdrofuran-2-yl methyl ether analogue 1 and the
metabolite 2 formed upon cleavage of the phenyl ether.
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
3
OH
O
O
i), ii), iii)
iv)
v)
O
O
B
O
O
O
I
O
4
5
6
7
Scheme 1. Synthesis of the 7-oxabicyclo[2.2.1]heptan-1-ylmethyl ether intermediate 7. Reagents and conditions: (i) methyltriphenylphosphonium bromide, n-BuLi, THF,
À10 to 25 °C, 17 h (92%); (ii) (CO2H)2, acetone, H2O, 25 °C, 3.5 h (87%); (iii) NaBH4, THF, 3 h, 25 °C (100%); (iv) 2.5 equiv I2, Na2CO3, AcCN, 22 h, 25 °C (42%); (v) 3-
hydroxyphenyl boronic acid pinacol ester, NaH, Et4N+IÀ, DMF, 100 °C, 6 h (66%).
I
Cl
N
H2N
N
Cl
O
i)
ii), iii)
O
NH2
N
N
N
N
O
+
Ph
O
H
N
N
Cl
Ph
OH
O
8
9
10
11
O
O
v), vi)
iv)
H2N
3
N
N
N
OH
12
Scheme 2. Synthesis of the 7-oxabicyclo[2.2.1]heptan-1-ylmethyl ether containing analogues 12 and 3. Reagents and conditions: (i) Hünig’s base, EtOH, 6 h, 80 °C (67%); (ii)
N-iodosuccinimide (NIS), DMF, 23 h, 25 °C (92%); (iii) NH4OH, dioxane, 23 h, 110 °C (58%); (iv) 7, (Ph3P)4Pd, Na2CO3, H2O, DMF, 16 h, 80 °C (80%); (v) 2-iodoxybenzoic acid,
AcCN, 1 h, 80 °C (25%); (vi) azetidine, Na(OAc)3BH, AcOH, 1,2-DCE, 25 °C, 1 h, then reversed-phase chromatography (22%).
within the ATP-binding site and led to a similar level of IGF-1R
activity compared to NVP-AEW541 and the (S)-2-THF analogue 1
at both the biochemical and cellular level. However, there is an
overall trend in Table 1 for the [2.2.1] bicyclic ethers to be slightly
more potent compared to the (S)-2-THF analogues, vide infra, sup-
porting the original hypothesis of increasing favourable interac-
tions within the ATP site of IGF-1R. From the initial study the
most potent THF analogues incorporated 5,5-dimethyl substitution
in the THF moiety,9 and to explore this possibility in the [2.2.1]
bicyclic ether series the 4-methyl analogue of 12, compound 13,
was prepared as outlined in Scheme 3. Following an analogous
approach to the sequence shown in Scheme 1, but replacing the
ketone reduction step by a methyl Grignard addition, the key inter-
mediate 14 was readily prepared and converted to the boronate
ester 15 in 3 steps. Suzuki coupling between 15 and 11 then gave
the targeted compound 13.
Comparison of the IGF-1R activity of 13 with 12 indicated no
increase in IGF-1R activity upon the introduction of the 4-methyl
group into the 7-oxabicyclo[2.2.1]heptan-1-ylmethyl ether moiety
in contrast to the SAR seen for the 2-THF analogues. With no added
benefit from the 4-methyl group these analogues were not pursued
further.
Table 1
Biochemical and cellular IGF-1R activities for the cyclic ether analogues, the phenol 2 and the reference compounds NVP-AEW541 and linsitinib
Compound
Ether moiety
Bn
Phenyl substituent
H
Cyclobutyl 3-substituent
IGF-1R IC50 (nM)
HEK IC50 (nM)
Ba/F3 IC50 (nM)
NVP-AEW541
Linsitinib
1
2
3
cis-CH2-azetidine
trans-Me-cis-OH
cis-CH2-azetidine
cis-CH2-azetidine
cis-CH2-azetidine
cis-CH2OH
cis-CH2OH
cis-CH2-TMSO
cis-N-Ac-Pip
61
14
84
420
46
65
20
7.6
12
454
14
11
29
21
16
92
n.d.
<4.6
15
7.3
27
251
7.8
16
See Figure 1
2.8
n.d.
n.d.
53
128
183
64
(S)-2-THF-CH2
H
[2.2.1]-CH2
[2.2.1]-CH2
4-Me-[2.2.1]-CH2
[2.2.1]-CH2
[2.2.1]-CH2
(S)-2-THF-CH2
[2.2.1]-CH2
[2.2.1]-CH2
(S)-2-THF-CH2
[2.2.1]-CH2
(S)-2-THF-CH2
(R,S)-2-Oxetanyl-CH2
[2.2.1]-CH2
(S)-2-THF-CH2
(R)-2-THF-CH2
(S)-2-THF-CH2
(S)-2-THF-CH2
(R)-2-THF-CH2
[2.2.1]-CH2
H
H
H
H
H
H
H
H
2-F
2-F
2-F
6-F
6-F
6-F
6-F
6-F
6-F
6-F
5-F
5-F
4-F
12
13
16
17
25
26
27
28
29
30
31
32
33
34
35
36
37
38
33
47
n.d.
9.4
80
4.3
9.6
16
8.3
16
200
13
12
64
54
260
1100
12
10
cis-CH2-TMSO
cis-CH2-TMSO
cis-N-Ac-Pip
232
n.d.
2.8
16
8.9
14
n.d.
12
13
159
n.d.
1291
5602
31
cis-N-Ac-Pip
cis-CH2-TMSO
cis-CH2-TMSO
cis-CH2-TMSO
cis-N-Ac-Pip
cis-N-Ac-Pip
cis-N-Ac-Pip
trans-N-Ac-Pip
cis-CH2-TMSO
cis-CH2-TMSO
cis-CH2-TMSO
47
n.d.
n.d.
n.d.
n.d.
n.d. not determined.
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

4
R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
O
O
OH
i), ii), iii), iv), v)
vi), vii), viii)
ix)
O
4
H2N
N
B
O
O
O
N
15
N
14
OH
13
Scheme 3. Synthesis of the 4-methyl-7-oxabicyclo[2.2.1]heptan-1-ylmethyl ether containing analogue 13. Reagents and conditions: (i) slow addition MeMgBr, Et2O, 25 °C
(85%); (ii) 1 N HCl(aq), 25 °C, 24 h (87%); (iii) 3,4-dihydro-2H-pyran, cat. pTSA, 25 °C with cooling, 45 min (81%); iv) methyltriphenylphosphonium bromide, n-BuLi, THF, À10
to 25 °C, 4 h (88%); (v) cat. pTSA, MeOH, 25 °C, 24 h (83%); (vi) 2.5 equiv I2, Na2CO3, AcCN, 1 h, 25 °C (37%); (vii) 3-bromophenol, K2CO3, AcCN, sealed-vial, 150 °C, 48 h (82%);
(viii) bis(pinacolato)diboron, Pd(dppf)Cl2ÁCH2Cl2, KOAc, DMF, 80 °C, 15 h (53%); (ix) 11, (Ph3P)4Pd, Na2CO3, H2O, DMF, 16 h, 80 °C (43%).
O
O
O
O
i)
ii)
H2N
N
H2N
N
12
N
N
N
N
O
S
H
N
18
16
O
Scheme 4. Synthesis of the cis-methyl-(N-thiomorpholine sulfoxide) aminocyclobutyl analogue 16. Reagents and conditions: (i) 2-iodoxybenzoic acid, AcCN, 1 h, 80 °C; (ii)
i
thiomorpholine sulfoxide hydrochloride, Na(OAc)3BH, Pr2NEt, CH2Cl2, 25 °C, 1 h, (2 steps 50%).
O
O
I
I
NH2
H2N
N
Cl
H2N
N
v), vi)
iii), iv)
i), ii)
N
N
N
N
N
N
N
HO
OH
O
OH
N
HO
19
20
21
N
O
17
Scheme 5. Synthesis of the cis-(N-acylpiperazine) aminocyclobutyl analogue 17. Reagents and conditions: (i) 8, Hünig’s base, EtOH, 4.5 h, reflux, then CF3CO2H, 1 h, reflux
(91%); (ii) N-iodosuccinimide, DMF, 7 h, 60 °C (62%); (iii) NH3, 1,4-dioxan, 15 h, 100 °C, sealed tube (71%); (iv) NaIO4, THF/H2O, 18 h, 25 °C (83%); (v) N-acetylpiperazine, Na
(OAc)3BH, AcOH, 1,2-DCE, 25 °C, 4 h (63%); (vi) 7, (PPh3)4Pd, K3PO4, Na2CO3, DMF, H2O, 3 h, 100 °C, then normal-phase separation of a 4:1 cis:trans isomer mixture (51%).
methyl-(N-thiomorpholine sulfoxide) 16 (cis-CH2-TMSO) and cis-
4-(N-acetylpiperazine) 17 (cis-N-Ac-Pip). The routes to these two
3-cyclobutyl substituted analogues are exemplified as outlined
below for the cis-CH2-TMSO 16 in Scheme 4 and the cis-N-Ac-Pip
17 in Scheme 5.
Scheme 4 shows the synthesis of the cis-CH2-TMSO analogue 16
starting from the cis-hydroxymethyl cyclobutyl analogue 12: peri-
odinane oxidation to the aldehyde 18 was followed by a reductive
amination with thiomorpholine sulfoxide to give the desired com-
pound 16.
Table 2
Tertiary amine pKa values and hERG-channel binding-affinities for NVP-AEW541 and
the cyclic ether analogues 3, 16 and 17
Compound
pKa
hERG IC50 (lM)
NVP-AEW541
3
16
17
9.5
n.d.
5.7
5.9
0.13
1.1
>30
15
With additional phenyl ether opportunities identified the
attention turned to the pyrrolopyrimidine 7-substituent SAR
interacting with the sugar-pocket region of the ATP-binding site.
Extensive exploration of the series in this region identified two
3-cyclobutyl substituted variants which exhibited favourable
profiles compared to NVP-AEW541 based upon: improved levels
of IGF-1R activity and kinase selectivity; an adequate level of
solubility with a tertiary amine of reduced pKa, leading to lower
levels of hERG activity; increased metabolic stability resulting in
improved overall PK profiles. As a result these two pyrrolopyrim-
idine 7-substituents were extensively explored and are exemplified
by the 7-oxabicyclo[2.2.1]heptan-1-ylmethylether analogues:
Scheme 5 shows the synthesis of the cis-N-Ac-Pip analogue 17,
starting from 3-hydroxy-3-hydroxymethylaminoazetidine 19.16
Cyclisation with 2-(4,6-dichloropyrimidin-5-yl)acetaldehyde
8
and iodination gave the pyrrolopyrimidine 20 which was reacted
with ammonia, to displace the 4-chloro substituent, followed by
oxidative cleavage of the diol to give the ketone 21. Reductive ami-
nation of 21 with N-acetylpiperazine gave predominantly the
trans-substituted cyclobutyl isomer which could be most readily
separated from the cis-isomer following the Suzuki reaction with
the boronate ester 7 to give the desired compound 17.
Comparison of 16 and 17 with NVP-AEW541 and 3 showed
comparable, or improved, IGF-1R activity, in particular for the
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
5
of IGF-1R. The incorporation of fluorine into the 2-position was
anticipated to provide an additional hydrogen bond with Lys1033,
and to make a hydrophobic contact with Val1013. The incorporation
of fluorine into the 6-position was designed to create additional
hydrophobic contacts with Met1142 and Val1063, and Figure 5
shows the 6-fluoro analogue 33 modelled into the ATP-pocket of
IGF-1R highlighting these interactions. To fully explore the above
possibility a fluorine scan was conducted across all four available
positions within the pyrrolopyrimidine 5-phenyl substituent with
the 7-oxabicyclo[2.2.1]heptan-1-ylmethylether and the 2-THF cyc-
lic ethers combined with the optimal pyrrolopyrimidine 7-sub-
stituents, analogues 26–38 in Table 1. The fluorination of the
linsitinib scaffold in the 2-position of the equivalent phenyl ring
has also been shown to lead to improvements in activity, in particu-
lar increased residence time within the ATP-binding site of IGF-1R.18
To prepare the fluorinated analogues 26–38 the key boronate
ester intermediates 22 were prepared as shown in Scheme 6. To
begin the sequence the appropriately fluorinated 3-bromophenol
derivatives 23 were either alkylated under Mitsunobu conditions
to give the ether derivatives 24 which were then converted to the
desired boronate ester intermediates. Alternatively, the fluorinated
7-oxabicyclo[2.2.1]heptan-1-ylmethylether boronate ester inter-
mediates were prepared by the alkylation of the 3-bromophenols
23 with the iodide intermediate 6 in the first step, as outlined in
Scheme 1. The boronate ester intermediates 22 were then coupled
following the sequences shown in Schemes 2, 4 and 5 to prepare the
analogues 26–38, structures and IGF-1R data are shown in Table 1.
Consistent with the hypothesised impact of introducing fluorine
into the 5-phenyl moiety, the 2- and 6-fluoro analogues exhibited
comparable or slightly improved IGF-1R activities as exemplified
by the cis-N-Ac-Pip analogues 27 and 32 compared to the unsubsti-
tuted compound 17. The introduction of fluorine into the 5-position
of the phenyl moiety led to a 4-fold decrease in IGF-1R activity,
based upon a comparison between the cis-CH2-TMSO analogues
Figure 5. Model of compound 33 docked into the ATP site of IGF-1R. Hydrogen
bonds are represented with magenta lines: pyrrolopyrimidine N5 to M1082 NH
3.2 Å, pyrrolopyrimidine 4-NH to E1080 C@O 3.0 Å, aryl ether to K1033 side-chain
NH 3.1 Å, THF ether to K1033 side-chain NH 2.9 Å. Hydrophobic contacts between
the 6-fluoro substituent and residues V1063 and M1142 are indicated by yellow
arrows, van der Waals distances: M1142 to 6-fluorophenyl 3.4 Å, V1063 to 6-
fluorophenyl 3.4 Å.
cis-N-Ac-Pip analogue 17. Additionally, the lower pKa values for
both of the pyrrolopyrimidine 7-substituents were associated with
decreased affinity for the hERG channel compared to the parent
compounds, as shown in Table 2.17
To further explore the SAR of the series, molecular modelling
indicated the potential for the introduction of fluorine into the
2- and 6-positions of the pyrrolopyrimidine 5-phenyl substituent
to make additional favourable interactions within the ATP-pocket
5
X
X
X = H or F
X
4
6
i)
ii)
O
B
O
()
Br
O
n
2
()n
n = 0, 1
Br
OH
O
O
O
23
24
22
Scheme 6. Synthesis of the phenyl and fluorophenyl boronate ester 2-methyltetrahydrofuran ether and 2-methyloxetane ether building blocks 22 (X = H or F). Reagents and
conditions: (i) diisopropyl azodicarboxylate, PPh3, (R)- or (S)-(tetrahydrofuran-2-yl)methanol or (R,S)-(oxetan-2-yl)-methanol, THF, 24 h, 25 °C (58–82%); (ii) bis(pinacolato)
diboron, 3 mol % PdCl2(dppf), KOAc, DMF, 18 h, 100 °C (73–91%).
Table 3
Physicochemical and in vitro PK data for the 2-cyclic ether methyl ether analogues and the reference compounds NVP-AEW541 and linsitinib
Compound
cLogP/PSA (Å2)
HDM FA (%)
Caco-2 A–B/B–A (10À6 cm sÀ1
)
Rat microsome ER (%)
Rat PPB (%)
NVP-AEW541
Linsitinib
1
12
13
16
17
25
26
27
28
29
30
31
32
33
34
4.4/72
4.0/92
3.0/81
2.3/98
2.8/98
2.3/98
2.0/101
2.0/98
2.4/98
2.1/101
1.7/101
2.6/98
2.3/98
1.6/98
2.3/101
1.9/101
1.9/101
1.9/101
99
99
86
93
99
26
49
30
18
40
24
17
22
19
65
31
31
27
5.4/10.1
15.1/13.2
n.d.
12.0/18.9
n.d.
4.4/21.2
5.6/15.5
n.d.
4.3/20.1
n.d.
74
85
n.d.
80
70
72
61
59
69
46
34
65
76
20
46
43
46
85
95
>99
n.d.
n.d.
n.d.
85
89
89
n.d.
n.d.
91
n.d.
n.d.
n.d.
n.d.
78
n.d.
4.8/20.6
2.9/17.8
n.d.
n.d.
11.7/13.3
n.d.
n.d.
n.d.
35
10.1/22.5
PSA = polar surface area; HDM FA = hexadecane membrane permeability assay, predicting fraction absorbed from the gastrointestinal tract following oral dosing;
PPB = plasma protein binding.20
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

6
R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Table 5
Biochemical and cellular InsR activities, and fold selectivities versus the IGF-1R for
selected cyclic ether analogues, the phenol 2 and the reference compounds
Compound
IC50 (nM), (IGF-1R fold selectivity)
InsR
HEK
Ba/F3
NVP-AEW541
Linsitinib
1
2
3
28
33
1018, (17)
34, (2.4)
930, (11)
3800, (9.0)
510, (11)
94, (5.9)
892, (14)
16, (5.7)
n.d.
244, (12)
126, (17)
64, (5.3)
638, (2.1)
65, (4.6)
36, (2.4)
93, (5.8)
n.d.
Figure 6. Extent of IGF-1R and AKT phosphorylation in NIH3T3:IFG-1R:IGF2
xenografts measured by Western blot analysis 2 and 8 h post treatment with
compounds 28, 30 and 33.
471, (8.9)
102, (6.4)
166, (13)
175, (15)
36 and 25. This decrease in activity with the 5-fluoro substituent
was anticipated based upon a steric clash with the side chain of
residue Val1063 in the modelled interaction. Introduction of fluo-
rine into the 4-position proved to be more complex, and for the
example 38 a comparable level of activity is retained compared to
the non-fluorinated analogue 16. However, a decrease in activity
of 5-fold, or greater, was seen for other 4-fluoro analogues. Based
upon the above observations, the further optimisation of the series
was focused upon the 2- and 6-fluorinated analogues.
established maintaining a trough inhibition of IGF-1R phosphory-
lation to be minimally required to deliver the maximum efficacy
upon chronic dosing in a murine IGF-1R dependent human-
xenograft model (NIH3T3:IFG-1R:IGF2).19 Our goal was to achieve
the maximum level of regression in the above model with a
compound that could be administered twice-daily, ideally with a
dose below 50 mg/kg, with a favourable prediction for translating
the PK profile into man, and good overall drug properties.
Therefore, to further differentiate between the analogues of
interest with high IGF-1R activities, and with higher levels of
microsome stability: the extent of pathway inhibition following a
single dose was assessed using the NIH3T3:IFG-1R:IGF2 xenograft
model in the nude mouse. Compounds were administered orally as
a single dose of 50 mg/kg and the extent of PD modulation was
determined 2 h and 8 h after dosing. The PD response was assessed
by western blot analysis to measure directly the level of IGF-1R
phosphorylation, and further downstream in the pathway the level
of AKT-phosphorylation was also measured. From this assessment,
compounds 28 and 33 were identified as producing strong PD
modulation at both time points, which was comparable to the ref-
erence compounds NVP-AEW541 and linsitinib when administered
at an efficacious dose level, at the same time points. In contrast, a
number of compounds only produced strong inhibition at the 2 h
time point which was greatly diminished by the 8 h time point,
as exemplified by compound 30, data are shown in Figure 6.
From the above PD-modulation screening, the cis-N-Ac-Pip ana-
logues 28 and 33 were identified for further profiling. Both com-
pounds exhibited reasonable levels of solubility (>1 mM at pH
Exploring the series further, one trend that became apparent
during the optimisation of the 2-cyclic ether methyl ethers was
that higher levels of lipophilicity resulted in increased microsomal
clearance values, and clogP values higher than 2.0 would likely be
associated with high microsome extraction ratios (ER). Measured
logP values were found to be in good agreement with the calcu-
lated values for the 2-cyclic ether methyl ether analogues. Physic-
ochemical and in vitro PK parameters are shown for selected
compounds in Table 3. Additionally, a reasonable in vitro/in vivo
correlation was observed for the rat using microsomal clearance,
at least to the extent that high in vitro microsomal clearance
proved to be a very good predictor of unacceptably high in vivo
clearance. For a number of the most interesting analogues the
lipophilicity threshold for acceptable microsome clearance was
often exceeded when comparing the [2.2.1]-bicyclic ether with
the corresponding 2-THF ether analogues, for example comparing
16, 27, 29 and 32 with 25, 28, 30 and 33. This increased metabolic
stability for the 2-THF methyl ethers was sufficient to overcome
the slightly lower levels of IGF-1R potency compared to the
[2.2.1]-bicyclic ethers when considered in terms of the compounds
overall profiles. Looking to further lower the level of lipophilicity
the introduction of a 2-oxetanyl methyl ether was also investi-
gated, as exemplified by compounds 31, which led to a further
increase in metabolic stability. However, the greater than 10-fold
decrease in IGF-1R potency for the 2-oxetanyl methyl ethers was
again observed and sufficient to concentrate the optimisation on
the 2-THF and the [2.2.1]-bicyclic ethers.9 Additionally, the nature
of the cyclobutyl 3-substituent can also be seen to impact upon the
metabolic stability, the higher lipophilicity of the cis-CH2-TMSO
analogues tending towards the highest levels of in vitro clearance
for the analogues in Table 3.
4.0 and >20 lM at pH 6.8) and rat PK studies showed both com-
pounds were able to deliver high levels of exposure following oral
dosing, data are shown in Table 4. In comparison to NVP-AEW541,
in vivo clearance was greatly reduced, in line with the in vitro
microsome data, leading to longer half-lives, even with signifi-
cantly reduced volumes of distribution (Vss) and higher unbound
fractions in plasma. Oral bioavailabilites (F) of 100% were deter-
mined for both 28 and 33, when administered as suspensions, indi-
cating good absorption from the gastro intestinal tract. This profile
being consistent with the Caco-2 permeability data shown in
Table 3, and highlighting the under prediction of the fraction
absorbed by the passive membrane permeability assay for these
two compounds.20
A further key requirement for the optimisation of the series was
to identify compounds with an adequate level of oral exposure to
be able to deliver the targeted pharmacodynamic (PD) response
at an acceptable dose level. Earlier PD/efficacy studies had
In addition, with the exception of the closely related InsR com-
pounds 28 and 33 showed high levels of kinase selectivity against
Table 4
Rat PK data following iv and po dosing of compounds 28 and 33 compared to NVP-AEW541
Compound
Clearance (ml minÀ1 kgÀ1
)
Vss (L/kg)
Terminal t (h)
AUC po d.n. (nmol h LÀ1
)
AUC iv d.n. (nmol h LÀ1
)
Oral F (%)
½
NVP-AEW541
28
33
123 13
40 12
19 2.3
6.5 1.1
5.0 0.6
2.0 0.5
3.1 1.8
3.1 2.4
311 33
862 52
1395 193
72 13
892 285
1210 184
23
97
115 16
4
6
28
5
d.n. = dose normalised to 1.0 mg kgÀ1
.
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
7
*
Figure 7. Efficacy study for compound 33, NVP-AEW541 and linsitinib in nude mice bearing NIH3T3 xenotransplants.
an internal panel of sixty kinases.12 Interestingly, and as previously
reported for NVP-AEW541, some degree of selectivity versus the
closely related InsR was observed for the 5-phenyl-4-aminopy-
rrolopyrimidine series, data are shown in Table 5. This trend was
evident throughout the series, at both the biochemical and cellular
level. To further explore this selectivity an in vivo comparison of
the time course of IGF-1R PD modulation in the murine NIH3T3:
IFG-1R:IGF2 tumour xenograft model compared to the inhibition
of pInsR in the liver was conducted with compound 33.19 In line
with the biochemical and cellular data this study also revealed evi-
dence for a modest degree of selectivity in vivo between the IGF-1R
and InsR receptors for this compound. However, the impact of any
selectivity upon efficacy and tolerability remains unclear based
upon the role IGF-1R/InsR hetero dimers may play in driving
tumour growth.2 Additional further off-target screening revealed
both compounds 28 and 33 to have acceptable profiles, including
led to approximately tumour stasis (18% regression) and was well
tolerated, as assessed by no change in body weigh compared to the
vehicle treated control group. Similarly, stasis with an equivalent
level of tolerability was also determined for NVP-AEW541 (19%
regression) and linsitinib (16% regression) at doses of 75 and
20 mg/kg respectively. Increasing the doses of compound 33 and
linsitinib to 40 and 30 mg/kg respectively led to comparable reduc-
tions in the tumour volumes (53% and 70% regression). However,
this degree of response was associated with a reduction in body
weight for both compounds, and represented the limit of tolerability
in the mouse. As a further measure of tolerability, the functionality
of the insulin pathway was assessed through an insulin-tolerance
test (ITT) in the mouse for all three compounds. Comparing the ITT
responses for compound 33 and linsitinib, the increases in circulat-
ing insulin and glucose levels supported these changes to be the
key factor most likely driving the tolerability limitations of both
compounds in this model.21
affinities for the hERG channel >30 l
M.17
Based upon their encouraging profiles, compounds 28 and 33
were investigated in efficacy studies using the NIH3T3:IGF-1R:
IGF2 xenograft model in the nude mouse. From these studies 33
showed the most encouraging profile, and an extensive in vivo data
set was generated for this compound and compared to NVP-
AEW541 and linsitinib, key data are shown in Figure 7.19 A clear
anti-tumour dose response was observed for compound 33 when
administered orally twice-daily at doses from 10 to 40 mg/kg for
1 week. When administered at a dose of 20 mg/kg compound 33
In conclusion, taking NVP-AEW541 as the starting point, the
optimisation of a 5-phenyl-4-aminopyrrolopyrimidine series of
IGF-1R inhibitors focused upon three areas: replacing the benzyl
ether back-pocket binding moiety with a series of 2-cyclic ether
methyl ethers; a fluorine scan of the 5-pheny group; and optimisa-
tion of the sugar-pocket binder. This approach resulted in the
identification of compound 33 containing (S)-2-THF methyl ether
6-fluorophenyl ether back-pocket and cis-N-Ac-Pip sugar-pocket
binding-moieties. Compared with NVP-AEW541, compound 33
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075

8
R. A. Fairhurst et al. / Bioorg. Med. Chem. Lett. xxx (2016) xxx–xxx
Receptors. In Accounts in Drug Discovery: Case Studies in Medicinal Chemistry;
Barrish, J. C., Carter, P. H., Cheng, P. T. W., Zahler, R., Eds.; Drug Discovery Series
No. 4; Royal Society of Chemistry, 2011.
exhibited increased IGF-1R potency, reduced hERG activity and an
improved PK profile. In a murine IGF-1R dependent xenograft
model compound 33 inhibited tumour growth to the level of stasis
at less than 30% of the dose required to produce the same effect
with NVP-AEW541, and at the highest tolerated dose produced a
comparable degree of tumour regression compared to linsitinib.
7. Garcia-Echeverria, C.; Pearson, M. A.; Marti, A.; Meyer, T.; Mestan, J.;
Zimmermann, J.; Gao, J.; Brueggen, J.; Capraro, H.-G.; Cozens, R.; Evans, D. B.;
Fabbro, D.; Furet, P.; Graus Porta, D.; Liebetanz, J.; Martiny-Baron, G.; Ruetz, S.;
Hofmann, F. Cancer Cell 2004, 5, 231.
8. (a) Weisberg, E.; Nonami, A.; Chen, Z.; Nelson, E.; Chen, Y.; Liu, F.; Cho, H. Y.;
Zhang, J.; Sattler, M.; Mitsiades, C.; Wong, K.-K.; Liu, Q.; Gray, N. S.; Griffin, J. D.
Clin. Cancer Res. 2014, 20, 5483; (b) Suda, K.; Mizuuchi, H.; Sato, K.; Takemoto,
T.; Iwasaki, T.; Mitsudomi, T. Int. J. Cancer 2014, 135, 1002; (c) Ioannou, N.;
Seddon, A. M.; Dalgleish, A.; Mackintosh, D.; Modjthedi, H. BMC Cancer 2013,
13, 41.
9. Stauffer, F.; Jacob, S.; Scheufler, C.; Furet, P. Bioorg. Med. Chem. Lett. 2016, 26.
http://dx.doi.org/10.1016/j.bmcl.2016.02.074.
10. In vitro and in vivo metabolism data for NVP-AEW541 and representative 2-
cyclic ether methylether derivatives are included in Supplementary data.
11. Widler, L.; Green, J.; Missbach, M.; Šuša, M.; Altmann, E. Bioorg. Med. Chem. Lett.
2001, 11, 849.
12. Kinase selectivity was assessed against a panel of biochemical assays: data for
compounds NVP-AEW541, 1, 2, 3, 28 and 33 are included in Supplementary
data.
13. (a) Experimental details of the syntheses of the final products and
intermediates are described. In: Chen, B.; Fairhurst, R. A.; Jiang, S.; Lu, W.;
Marsilje, T. H.; McCarthy, C.; Michellys, P-Y.; Stutz, S. WO 2012/120469, Sep 13,
2012.; (b) Chen, B.; Fairhurst, R. A.; Floersheimer, A.; Furet, P.; Guagnano, V.;
Jiang, S.; Lu, W.; Marsilje, T. H.; McCarthy, C.; Michellys, P-Y.; Stauffer, F.; Stutz,
S.; Vaupel, A. WO2011/029915, Mar 17, 2011.
Acknowledgements
The authors would like to thank Peter Drueckes and Joerg Trappe
for conducting biochemical assays; Markus Wartmann for conduct-
ing cellular assays; Rainer Aichholz, Francesca Blasco, Jerome Dayer,
Francis Risser and Peter Wipfli for conducting pharmacokinetic
studies; Rita Andraos-Rey for generating the NIH3T3:IGF-1R:IGF2
cell lines, Geneviève Albrecht for conducting in vitro and in vivo
PD analyses; Jasmin Wirth, Milen Todorov and Mickael Le Douget
for their excellent technical assistance.
Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2016.02.
075.
14. Slade, J.; Bajwa, J.; Liu, H.; Parker, D.; Vivelo, J.; Chen, G.-P.; Calienni, J.;
Villhauer, E.; Prasad, K.; Repic, O.; Blacklock, T. J. Org. Process Res. Dev. 2007, 11,
825.
15. Details of the biochemical IGF-1R and InsR, and the cellular HEK and Ba/F3
assays are described in Supplementary data.
References and notes
16. Jandu, K. S.; Barrett, V.; Brockwell, M.; Cambridge, D.; Farrant, D. R.; Foster, C.;
Giles, H.; Glen, R. C.; Hill, A. P.; Hobbs, H.; Honey, A.; Martin, G. R.; Salmon, J.;
Smith, D.; Woollard, P.; Selwood, D. L. J. Med. Chem. 2001, 44, 681.
17. Details of the hERG-channel binding assay are described in Supplementary
data.
1. Pollak, M. Nat. Rev. Cancer 2008, 8, 925.
2. (a) Gallagher, E. J.; LeRoith, D. Trends Endocrinol. Metab. 2010, 21, 610;
(b) Denley, A.; Wallace, J. C.; Cosgrove, L. J.; Forbes, B. E. Horm. Metab. Res. 2003,
35, 778.
18. Jin, M.; Petronella, B. A.; Cooke, A.; Kadalbajoo, M.; Siu, K. W.; Kleinberg, A.;
May, E. W.; Gokhale, P. C.; Schulz, R.; Kahler, J.; Bittner, M. A.; Foreman, K.;
Pachter, J. A.; Wild, R.; Epstein, D.; Mulvihill, M. J. Med. Chem. Let. 2013, 4, 627.
19. Details of the NIH3T3:IGF-1R:IGF2 xenograft model, including the comparison
of IGF-1R and InsR PD modulation for compound 33, are described in
Supplementary data.
20. Details of the in vitro permeability assays and plasma protein binding
measurements are described in Supplementary data.
21. Details of the insulin-tolerance test are described in Supplementary data.
3. Arteaga, C. L.; Kitten, L. J.; Coronado, E. B.; Jacobs, S.; Kull, F. C.; Allred, D. C.;
Osborne, C. K. J. Clin. Invest. 1989, 84, 1418.
4. Pollak, M. Nat. Rev. Cancer 2012, 12, 159.
5. (a) Negi, A.; Ramarao, P.; Kumar, R. Mini-Rev. Med. Chem. 2013, 13, 653; (b)
Sarma, P. K. S.; Tandon, R.; Gupta, P.; Dastidar, S. G.; Ray, A.; Das, B.; Cliffe, I. A.
Expert Opin. Ther. Patents 2007, 17, 25.
6. (a) Puzanov, I.; Lindasy, C. R.; Goff, L.; Sosman, J.; Gilbert, J.; Berlin, J.; Poondru,
S.; Simantov, R.; Gedrich, R.; Stephens, A.; Chan, E.; Ecans, T. R. J. Clin. Cancer
Res. 2015, 21, 701; (b) Mulvihill, M. J.; Buck, E. The Discovery of OSI-906, a
Small-Molecule Inhibitor of the Insulin-Like Growth Factor-1 and Insulin
Please cite this article in press as: Fairhurst, R. A.; et al. Bioorg. Med. Chem. Lett. (2016), http://dx.doi.org/10.1016/j.bmcl.2016.02.075