Novel tricyclic pyrazole BRAF inhibitors with imidazole or furan central scaffolds

Bioorganic & Medicinal Chemistry 18 (2010) 6934–6952

Contents lists available at ScienceDirect

Bioorganic & Medicinal Chemistry

journal homepage: w ww.elsevier.com/locate/bmc

Novel tricyclic pyrazole BRAF inhibitors with imidazole or furan

central scaffolds

Dan Niculescu-Duvaz ^a, Ion Niculescu-Duvaz ^a, Bart M. J. M. Suijkerbuijk ^a, Delphine Ménard ^a,

Alfonso Zambon ^a, Arnaud Nourry ^a, Lawrence Davies ^a, Helen A. Manne ^a, Frank Friedlos ^a, Lesley Ogilvie ^a,

Douglas Hedley ^a, Andrew K. Takle ^c, David M. Wilson ^c, Jean-Francois Pons ^d, Tom Coulter ^d, Ruth Kirk ^b,

Neus Cantarino ^b, Steven Whittaker ^b, Richard Marais ^b, Caroline J. Springer ^a,

*

^aCancer Research UK Centre for Cancer Therapeutics, The Institute of Cancer Research, 15 Cotswold Road, Sutton, Surrey SM2 5NG, United Kingdom

^bThe Institute of Cancer Research, Cancer Research UK Centre for Cell and Molecular Biology, 237 Fulham Road, London SW3 6JB, United Kingdom

^cDepartment of Medicinal Chemistry, Neurology and GI Centre of Excellence for Drug Discovery, GlaxoSmithKline Pharmaceuticals, New Frontiers Science Park,

Third Avenue, Harlow, Essex CM19 5AW, United Kingdom

^dEvotec (UK) Ltd, 114 Milton Park, Abingdon, Oxfordshire OX14 4SA, United Kingdom

a r t i c l e i n f o

a b s t r a c t

Article history:

V-RAF murine sarcoma viral oncogene homolog B1 (BRAF) is a serine/threonine-speciﬁc protein kinase

that is mutated with high frequency in cutaneous melanoma, and many other cancers. Inhibition of

mutant BRAF is an attractive therapeutic approach for the treatment of melanoma. A triarylimidazole

BRAF inhibitor bearing a phenylpyrazole group (dimethyl-[2-(4-{5-[4-(1H-pyrazol-3-yl)-phenyl]-4-pyri-

din-4-yl-1H-imidazol-2-yl}-phenoxy)-ethyl]-amine, 1a) was identiﬁed as an active BRAF inhibitor. Based

on this starting point, we synthesized a series of analogues leading to the discovery of 6-{2-[4-(4-methyl-

piperazin-1-yl)-phenyl]-5-pyridin-4-yl-3H-imidazol-4-yl}-2,4-dihydro-indeno[1,2-c]pyrazole (1j), with

nanomolar activity in three assays: inhibition of puriﬁed mutant BRAF activity in vitro; inhibition of

oncogenic BRAF-driven extracellular regulated kinase (ERK) activation in BRAF mutant melanoma cell

lines; and inhibition of proliferation in these cells.

Received 9 April 2010

Revised 8 June 2010

Accepted 10 June 2010

Available online 15 June 2010

Keywords:

BRAF

Kinase inhibitors

Anticancer

Melanoma

Triarylimidazole

Dihydroindenopyrazole

1. Introduction

(V600E).²In cancer cells, ^V600EBRAF stimulates constitutive ERK

activity and drives proliferation and survival, thereby providing

The RAS-RAF-MEK-ERK pathway is hyper-activated in approxi-

mately 30% of human cancers,¹where it stimulates cell growth and

survival. This hyper-activation is caused in part by mutations in

receptor tyrosine kinases, the small G-proteins of the RAS family,

and the serine/threonine speciﬁc protein kinase BRAF. Mutations

in BRAF (one of the three rapidly growing ﬁbrosarcoma (RAF) genes

in humans; the other two being ARAF and CRAF) occur in ꢀ2% of

human cancers, and are particularly prevalent in cutaneous mela-

nomas (ꢀ50% of cases). The most common mutation (90% fre-

quency) is a glutamic acid for valine substitution at position 600

essential tumor growth and maintenance functions.³^V600EBRAF

also contributes to neoangiogenesis by stimulating vascular endo-

thelial growth factor secretion.⁴Consequently, BRAF is an attrac-

tive validated therapeutic target in melanoma, and several BRAF

inhibitors are currently under development and in clinical trials⁵

and we have previously described type II BRAF inhibitors, targeting

the inactive conformation of BRAF.^6,7

Triarylimidazoles have been reported as inhibitor scaffolds for

targeting the active kinase conformation kinase, for example

p38⁸and RAF.^9,10The general structure of these latter kinase inhib-

itors contains a hinge-binding group, usually pyridine or pyrimi-

dine (‘ring A’), a central imidazole scaffold occupying the ribose

position in the ATP-binding pocket (‘ring C’) and a substituted aro-

matic group interacting with the hydrophobic pocket (BPII accord-

ing to Liao’s nomenclature)¹¹next to the gatekeeper residue (‘ring

B’) (Fig 1). The third aryl ring (‘ring D’) extends outside the ATP

pocket towards the solvent. The co-crystal structure of BRAF with

SB590885, a triarylimidazole BRAF inhibitor, indicates binding to

Abbreviations: Boc, tert-butoxycarbonyl; BRAF, V-RAF murine sarcoma viral

oncogene homolog B1; DCM, dichloromethane; DMF, dimethylformamide; ERK,

extracellular regulated kinase; MAPK, mitogen-activated protein kinase; MEK,

MAPK/ERK kinase; MOM, methoxymethyl; PK, pharmacokinetics; RAF, rapidly

growing ﬁbrosarcoma; SAR, structure–activity relationship; TFA, triﬂuoroacetic

acid; THF, tetrahydrofuran.

* Corresponding author. Tel.: +44 20 8722 5013; fax: +44 20 8722 4046.

E-mail address: caroline.springer@icr.ac.uk (C.J. Springer).

doi:10.1016/j.bmc.2010.06.031

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6935

Figure 1. Examples of triarylimidazole BRAF inhibitors and their putative binding mode.

the active conformation of BRAF and supports the described bind-

ing mode.⁹Therefore, these compounds are BRAF type I inhibitors.

Two examples of reported active triarylimidazole BRAF inhibitors,

L-779450 and SB-590885, are also shown in Figure 1.^12,9These

BRAF inhibitors contain an H-bond donor on ‘ring B’ (phenol or

oxime, Fig. 1) that is essential for activity.

In order fully to exploit the BPII pocket of the active BRAF

conformation, we designed novel type I BRAF inhibitors based on

this scaffold, but with an H-bond donor pyrazole heterocycle

substituent on ‘ring B’, as a potential functional bioisostere of ind-

anoneoxime and chlorophenol. 4-(3-Pyrazolyl)phenyl ring B was

proposed to target the BPII pocket of BRAF and consequently com-

pound 1a was synthesized (Fig. 2). Docking of this compound with

the BRAF structure in the active conformation (pdb code 2FB8)

showed the expected orientation of the compound, similar to

SB590885, with the pyridyl ring A interacting with the hinge and

is dependent of the rest of the inhibitor: the co-crystal structure

of BRAF with an indanoneoxime BRAF inhibitor with a pyrazole

scaffold shows the oxime in E-geometry (pdb code 3D4Q),¹³

whereas SB590885 co-crystallises in an opposite Z-orientation.

Pleasingly, this compound was found to inhibit isolated full length

^V600EBRAF at low micromolar concentrations (IC₅₀BRAF = 1.6

l

M)

and also to inhibit ^V600EBRAF signalling in melanoma cells (GI₅₀

SRB = 7.4 M). Hit 1a was therefore used as starting point for the

l

structure–activity relationship (SAR) exploration and structure

optimization for new BRAF inhibitors.

2. Results and discussion

2.1. Chemistry

Two synthetic routes were designed to access the desired tri-

substituted imidazoles:

the 4-(3-pyrazolyl)phenyl ring

(Fig. 2). The geometry of the oxime in indanoneoxime inhibitors

B occupying the BPII pocket

Figure 2. Structure of phenylpyrazole hit 1a, and docking in BRAF structure.

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D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

(A) Starting from the 2,4,5-tribromo-imidazole core, 2, and

1e–g. MOM deprotection of intermediate 6 led to imidazole 7,

which could be coupled directly with boronic acid 33, to afford

compound 1d after 4-methoxybenzyl removal (Scheme 1).

introducing the desired substituents in sequence (Scheme 1).

(B) Using the protected indanone oxime (ring B) and building

the trisubstituted imidazole core (Scheme 2).

The alternative route (B) is a modiﬁcation of the synthesis re-

ported by Takle et al.¹⁰The starting material 5-bromo-2,3-dihy-

dro-1H-inden-1-one O-methyl oxime 10, was converted to the

corresponding aldehyde 11 with nBuLi and (dimethylformamide)

DMF, followed by coupling with 4-pyridylmethyleneoxy-t-butyldi-

methylsilane to afford intermediate 12. After deprotection with

tetrabutylammonium ﬂuoride, the corresponding diol, 13, was sub-

mitted to a Swern oxidation affording the dione 14. The coupling of

14 with selected aldehydes afforded the imidazole core linked to a

suitable ring D. The ﬁnal deprotection of the N-methyl indanone

oxime system yielded the ketones 16a–c, which were converted in

In the ﬁrst approach, the 2,4,5-tribromo-imidazole was

protected as a methoxymethyl (MOM) derivative 3, and converted

using a two-step procedure to the 2-(4-dimethylaminoethoxyphe-

nyl)-4,5-dibromoimidazole, 5. The Suzuki coupling of this interme-

diate with 1 equiv of 4-pyridylboronic acid led to the key

intermediate 6. A second Suzuki coupling with 4-methoxybenzyl-

protected boronic acids 26, 29a–b, 37, 40 or 43 (see Schemes 5

and 6) followed by acidic deprotection of the MOM and 4-

methoxybenzyl groups generated the ﬁnal compounds 1a–c,

Scheme 1. Synthesis of triarylimidazole pyrazoles by route A. Reagents and conditions: (i) MOMBr, NaH, THF; (ii) 4-HOC₆H₄B(OH)₂, K₂CO₃, Pd(PPh₃)₄, toluene, MeOH, reﬂux;

(iii) Cl(CH₂)₂N(CH₃)₂, Cs₂CO₃, DMF, 40 °C; (iv) pyridyl boronic ester, K₂CO₃, Pd(OAc)₂, PPh₃, DME, H₂O, 90 °C; (v) boronic esters, K₂CO₃, Pd(OAC)₂, PPh₃, DME, H₂O, 110 °C; (vi)

5 M HCl, 60 °C; (vii) TFA, 130 °C.

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6937

Scheme 2. Synthesis of triarylimidazole tricyclic pyrazoles by route B. Reagents: (i) nBuLi, THF, DMF; (ii) 4-pyridyl-CH₂OTBDMS, LDA, THF; (iii) TBAF, THF; (iv) oxalyl

chloride, DMSO, Et₃N, DMF; (v) RCHO, AcOH, AcONH₄; (vi) 5 M HCl, dioxane; (vii) HC(NMe₂)₃, DMF; (viii) AcOH, H₂NNH₂.

the ﬁnal tricyclic compounds 1h, 1j–k by condensation with tris-

dimethylamino-methane and hydrazine (see Scheme 2).

The triazole ﬁnal compound 1i was obtained from intermediate

16b by nitrosation followed by the addition of hydrazine

(Scheme 3).

zotriazole and triethylamine in DMF, produced the key intermedi-

ates 22a–d. The condensation of 22a–d with hydrazine or

methylhydrazine gave the desired ﬁnal compounds 1l–o and 1p,

respectively (Scheme 4).

Since most of the boronic acids used in the synthesis of the imi-

dazoles previously described in Scheme 1 are not commercially

available, their preparation is summarized in Schemes 5 and 6.

Diarylfurans with aliphatic, aromatic or carboxamide solubilis-

ing groups were previously shown to be alternative scaffolds for

the synthesis of BRAF inhibitors.¹⁴Consequently, we synthesized

several furan analogues with our most active tricyclic pyrazole ring

B. For the synthesis of the trisubstituted furan analogues, the start-

ing material was methyl-2,3-dibromofuran-5-yl carboxylate 17.

Using two consecutive Suzuki couplings with 4-pyridyl boronic

acid followed by 1-(methoxyimino)-2,3-dihydro-1H-inden-5-ylbo-

ronic acid, the intermediate 19 was obtained. Basic hydrolysis of

intermediate 19 with NaOH afforded the corresponding carboxylic

acid, 20, followed by acidic hydrolysis to remove the oxime ether

group and generate the ketone 21. Condensation with the desired

amines, in the presence of diisopropylcarbodiimide, hydroxyben-

2.2. SAR of ring B—phenylpyrazole

In order to identify new type I BRAF inhibitors with a triarylim-

idazole scaffold, compound 1a with 4-(3-pyrazolyl)phenyl as ring B

was synthesized and found to be effective. The SAR of ring B was

explored further, leading to the discovery of the more potent

1,4-dihydroindeno[1,2-c]pyrazole. Using this group, further SAR

exploration of rings C and D was performed.

The biological activities of compounds 1a–p were assessed in

three assays:

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Scheme 3. Synthesis of tricyclic triazole. Reagents and conditions: (i) tBuONO, HCl, methoxyethanol; (ii) ethylene glycol, KOH, H₂NNH₂, 190 °C.

Scheme 4. Synthesis of diarylfuranamides. Reagents: (i) 4-pyridylboronic acid, Cs₂CO₃, Ph₃As, Pd(PPh₃)Cl₂, DMF; (ii) 1-(methoxyimino)-2,3-dihydro-1H-inden-5-ylboronic

acid, Cs₂CO₃, Ph₃As, Pd(PPh₃)₂Cl₂, DMF; (iii) NaOH, THF, MeOH; (iv) 5 M HCl, dioxane–Me₂CO; (v) amine, DIC, HOBt, TEA, DMF; (vi) DMF–DMA, H₂NNH₂, AcOH; (vii) DMF–

DMA, MeHNNH₂, AcOH; (viii) 4 N HCl.

(1) Inhibition of the isolated ^V600EBRAF kinase activity (IC₅₀

BRAF), measuring MEK1/2 phosphorylation as the endpoint.¹⁵

An alternative BRAF ﬂuorescent ligand binding assay^10,14gave

much lower K_dvalues than the IC₅₀s for the ^V600EBRAF kinase

activity (for example: compound 1h, K_d= 6.1 nM whereas

IC₅₀BRAF = 230 nM). We measured the IC₅₀BRAF values in

this paper, since they represent the inhibition of BRAF func-

tion which we consider more meaningful than the binding

of inhibitor to BRAF.

(2) Inhibition of ^V600EBRAF-dependent ERK phosphorylation in

WM266.4 melanoma cells (IC₅₀pERK).

(3) Growth inhibition of WM266.4 melanoma cells measured by

sulforhodamine-B (GI₅₀SRB). The IC₅₀(BRAF), IC₅₀(pERK)

and GI₅₀(SRB) results are presented in Table 1.

Our assessment of substitution of the pyrazole ring focused on

the 4-position, since no further space is available for position 5

based on modelling studies of 1a with the BRAF structure reported

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6939

M)

Table 1

Biological activities of BRAF inhibitors

No

Compound

IC₅₀BRAF (

lM)

IC₅₀ppERK (lM)

GI₅₀SRB (l

1a

1.60

5.8

7.4

1b

1.20

2.02

3.9

1c

>10

>100

11

1d

1e

1f

3.7

10

5.8

25

4.1

1.84

—

1.71

3.3

(continued on next page)

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D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

Table 1 (continued)

No

Compound

IC₅₀BRAF (

lM)

IC₅₀ppERK (lM)

GI₅₀SRB (lM)

1g

>10

100

8.9

1h

0.23

1.09

1.15

1i

>10

4.4

—

1j

0.24

0.58

0.87

1k

1.33

4.9

36

1l

3.6

0.74

1.61

1m

2.15

0.84

0.94

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6941

M)

Table 1 (continued)

No

Compound

IC₅₀BRAF (

lM)

IC₅₀ppERK (lM)

GI₅₀SRB (l

1n

2.78

0.97

1.59

1o

7.5

0.73

1.91

1p

>10

—

in the co-crystal structure with SB590885 (Fig. 2).⁹Substitution in

the 4-position of the pyrazole with methyl (compound 1b) is toler-

ated and beneﬁcial. Larger substituents such as cyano or chlorine

lead to loss of activity against both isolated BRAF and in cellular as-

says (1c, 1d). This effect could be due to a steric clash with the

BRAF binding pocket or an unfavourable electrostatic interaction.

In order to ﬁx the position of the pyrazole ring, more rigid tricyclic

systems were synthesized: 1e–h.

The ﬁve-membered ring (1,4-dihydroindeno[1,2-c]pyrazole 1h)

increases the potency against BRAF ﬁvefold compared to the

non-cyclic analogue 1b. This tricyclic system mimics the ind-

anoneoxime moiety, with the oxime group in the E-geometry. Sat-

urated or aromatic six-membered rings (1e, 1f) are not as effective

as the ﬁve-membered ring. Methylation of the pyrazole leads to a

loss in activity (1p vs 1l) consistent with the hypothesis that an

H-donor group is beneﬁcial for potent BRAF inhibition. Interest-

ingly, the tricyclic triazole analogue 1i is inactive, possibly due to

a polarity mismatch between the polar 3-position of the triazole

(corresponding to the 5-position of the pyrazole) and a non-polar

area of the BPII binding pocket. Similar results were observed

with the tricyclic pyridazinone 1g, where the inactivity can be

attributed to either polarity mismatch or steric clash. Since the

1,4-dihydroindeno[1,2-c]pyrazole 1h is the most active ring B,

further optimization of rings C and D were performed keeping this

ring B constant.

but surprisingly good cellular activity (for example compound

1m has submicromolar activity in both IC₅₀ppERK and GI₅₀SRB

assays).

Modiﬁcation of the substituents on the phenyl ‘group D’ (solu-

bilizing group) of the imidazole inhibitors is expected to have a

minimal effect on the BRAF potency but could modulate the cellu-

lar activity or the physicochemical properties of this series. Indeed,

the replacement of the 2-dimethylaminoethoxy side chain of ring

D (in 1h) with N-methylpiperazine lead to compound 1j, which

is equipotent on BRAF IC₅₀with 1h, but has improved cellular

activity. Compound 1j has submicromolar activity in all 3 assays.

Replacing the whole aryl ring D with a piperidine ring (1k) has a

negative effect on BRAF potency, with a sixfold potency loss com-

pared to 1h or 1j.

2.4. Pharmacokinetic (PK) data

The apparent clearance, half-life (t_1/2) and maximal plasma con-

centration of a representative tricyclic pyrazole, compound 1h,

were determined in vivo in CD1 nu/nu female mice following an

intraperitoneal administration of 1h. This compound exhibits a good

PK proﬁle, with a low plasma clearance (CL = 11 mL/min/kg), long

half-life (T_1/2= 3.9 h) and reaches plasma concentrations above its

SRB cellular IC₅₀(C_max= 5.8 lM; IC₅₀(SRB) = 1.1 lM).

3. Conclusion

2.3. SAR of rings C and D

A triarylimidazole BRAF inhibitor bearing a phenylpyrazole

group (1a) was identiﬁed as an active BRAF inhibitor. Starting from

this lead, optimization and SAR investigations led to the identiﬁca-

tion of the more potent tricyclic 1,4-dihydroindeno[1,2-c]pyrazole

BRAF inhibitor 1h, with a favourable PK proﬁle. Replacement of the

imidazole core scaffold with furanamide led to compounds with

weaker activity against isolated mutant BRAF, but with comparable

Furan-based analogues of SB-590885, where the core imidazole

ring C was replaced by furan, with a variety of solubilizing D

groups have been reported as potent BRAF inhibitors.¹⁴We have

synthesized a number of furan-amides with our tricyclic pyrazole

‘ring B’ as BRAF inhibitors (1l–p), which were found to possess

weaker activity against BRAF than their imidazole counterparts,

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D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

cellular activity. The triarylimidazole 1j with a phenylpiperazine

ring D is the most promising compound, with nanomolar activity

in the mutant BRAF inhibition assay, the cellular pERK inhibition

and the mutant BRAF melanoma WM266.4 growth inhibition.

5:1 mixture of toluene/MeOH (120 mL). The suspension was stir-

red vigorously whilst de-gassing with nitrogen for 20 min.

Pd(PPh₃)₄(0.658 g, 0.57 mmol) was added and the mixture was re-

ﬂuxed overnight. The reaction was cooled to room temperature, di-

luted with H₂O (20 mL) and EtOAc (10 mL). The phases were

separated. The organic layer was then extracted with 1 M NaOH

(3 ꢁ 15 mL). Both aqueous layers were separately acidiﬁed with

1 M HCl until a white precipitate appeared (pH 6–7). The precipi-

tates were ﬁltered, washed with water, and dried under vacuum

to give a total of 0.800 g of 4 as a white solid. (Yield: 39%). ¹H

NMR (250 MHz, DMSO-d₆) d ppm: 7.37 (2H, d, J = 8.60 Hz), 6.70

(2H, d, J = 8.6 Hz), 5.21 (2H, s), 3.28 (3H, s). LCMS: t_R= 1.82 min,

360, (M+H)⁺calcd for C₁₁H₁₀Br₂N₂O₂. HRMS: (M+H)⁺calcd for

C₁₁H₁₀Br₂N₂O₂: 360.9187, found: 360.9179.

4. Experimental

4.1. Chemistry

4.1.1. Materials and methods

All starting materials, reagents and solvents for reactions were

reagent grade and used as purchased. Chromatography solvents

were HPLC grade and were used without further puriﬁcation. Reac-

tions were monitored by thin layer chromatography (TLC) analysis

using Merck Silica Gel 60 F-254 thin layer plates. Flash column

chromatography was carried out on Merck Silica Gel 60 (0.015–

0.040 mm) or in disposable Isolute Flash Si and Si II silica gel col-

umns. LC-MS analyses were performed on a Micromass LCT/Water’s

4.1.2.3. {2-[4-(4,5-Dibromo-1-methoxymethyl-1H-imidazol-2-yl)-

phenoxy]-ethyl}-dimethyl-amine (5). 4-(4,5-Dibromo-1-meth-

oxymethyl-1H-imidazol-2-yl)-phenol

4

(300 mg, 0.83 mmol),

Cs₂CO₃(649 mg, 1.99 mmol) and 2-dimethylaminoethyl chloride

hydrochloride (143 mg, 0.99 mmol) were dissolved in DMF

(4 mL). The solution was stirred at 40 °C overnight, then diluted

with water (20 mL) and extracted with EtOAc (3 ꢁ 10 mL). The or-

ganic layer was washed with brine (10 mL), dried (MgSO₄) and the

solvent removed under vacuum to yield 259 mg of 5 as a pale yel-

low oil. (Yield: 72%). ¹H NMR (250 MHz, CDCl₃) d ppm: 7.69 (2H, d,

J = 8.9 Hz), 7.00 (2H, d, J = 9.0 Hz), 5.25 (2H, s), 4.12 (2H, t,

J = 5.7 Hz), 3.46 (3H, s), 2.76 (2H, t, J = 5.7 Hz), 2.36 (6H, s). LCMS:

t_R= 1.37 min, 432 (M+H)⁺calcd for C₁₅H₁₉Br₂N₃O₂.

Alliance 2795 HPLC system using 5

l

m Atlantis C18, 50 mm ꢁ 2.1

mm columns at 22 °C with the following solvent system: aqueous:

water + 0.1% formic acid; organic: 0.1% formic + acetonitrile, at a

ﬂow rate of 1 mL/min. Method A: gradient starting with 100% aque-

ous to 100% organic in 2.5 min at room temperature and a ﬂow rate

of 0.6 mL/min or method B gradient starting with 100% aqueous to

100% organic in 5 min at 40 °C (column temperature) at a ﬂow rate

of 0.6 mL/min. UV detection was at 215 nm and ionisation was po-

sitive or negative ion electrospray. The molecular weight scan range

was 50–1000. Samples were injected at 3 lL on a partial loop ﬁll. All

automated HPLC puriﬁcation were performed on Gilson Prep LC

modules running on software version 1.71 or 3.0 and on Hyper-

4.1.2.4. {2-[4-(4-Bromo-1-methoxymethyl-5-pyridin-4-yl-1H-

imidazol-2-yl)-phenoxy]-ethyl}-dimethyl-amine (6). {2-[4-(4,5-

Dibromo-1-methoxymethyl-1H-imidazol-2-yl)-phenoxy]-ethyl}-

dimethyl-amine 5 (208 mg, 0.480 mmol), 4-pyridyl-boronic ester

(59 mg, 0.480 mmol), PPh₃(12.6 mg, 0.048 mmol), and K₂CO₃

(530 mg, 3.840 mmol) were suspended in a 2:1 mixture of DME/

H₂O (9 mL). The suspension was stirred vigorously whilst de-gas-

sing with N₂for 20 min before adding Pd(OAc)₂(5.4 mg,

0.024 mmol). The mixture was then reﬂuxed for 2 h, cooled to

room temperature, acidiﬁed to pH 1 with 1 M HCl and washed with

EtOAc (3 ꢁ 5 mL). The aqueous layer was basiﬁed with 2 M NaOH

to pH 14 and extracted with EtOAc (3 ꢁ 5 mL). This organic layer

was dried (MgSO₄) and the solvent removed under vacuum. The

residue was puriﬁed by chromatography using a stepped gradient

of 5–10% NEt₃in EtOAc, then 1:10:89 MeOH/NEt₃/EtOAc to give

43 mg of 6 as a yellow oil. (Yield: 21%). ¹H NMR (360 MHz, CDCl₃)

d ppm: 8.70–8.80 (2H, m), 7.75 (2H, d, J = 8.9 Hz), 7.56–7.64 (2H,

m), 6.97–7.09 (2H, m), 4.96 (2H, s), 4.14 (2H, t, J = 5.7 Hz), 3.34

(3H, s), 2.78 (2H, t, J = 5.7 Hz), 2.37 (6H, s). LCMS: t_R= 1.06 min,

431 (M+H)⁺calcd for C₂₀H₂₃BrN₄O₂. HRMS: (M+H)⁺calcd for

prepHSC18 100 mm ꢁ 21.2 mm columns, 12

lm, at a ﬂow rate of

30 mL/min at room temperature using as aqueous phase:

water + 0.1% (triﬂuoroacetic acid) TFA and as organic phase: aceto-

nitrile + 0.1% TFA (UV detector, at 215 and 254 nm). The purity of

the ﬁnal compounds was determined by HPLC as described above

and is 95% or higher unless speciﬁed otherwise. ¹H NMR spectra

were recorded on either a 250 MHz or a 400 MHz Bruker NMR ma-

chine. Accurate Mass Measurement was performed with a Waters

Micromass LCT Premier Orthogonal Acceleration Time-of-Flight

Mass Spectrometer 4 GHz TDC with LockSpray™ enable mass mea-

surements of 5 ppm or better for m/z of 400 or greater and 2 mDa or

better for m/z of 400 or less. Calibration reference: Wpos_150208.-

cal or Wneg_150208a.cal. MassLynx v4.1 SCN 633 was the operat-

ing software using the in-built elemental composition to report

data. Minimum 10 scans are combined across a MS peak.

4.1.2. Synthesis and characterization of intermediates 3–9

(Scheme 1)

C₂₀H₂₃BrN₄O₂: 431.1082, found: 431.1081.

4.1.2.1. 2,4,5-Tribromo-1-methoxymethyl-1H-imidazole (3). Tri-

bromo imidazole 2 (6.1 g, 20.0 mmol) was dissolved in dry THF

(25 mL) and NaH (60% in mineral oil, 960 mg, 24 mmol) was added

portionwise. The solution was cooled to 0 °C under nitrogen atmo-

sphere and MOMBr (3.33 g, 24.0 mmol) was added dropwise over

approximately 10 min. The reaction was stirred at room tempera-

ture for 2 h or until complete by LC/MS. The solution was diluted

with water (100 mL), extracted with EtOAc (3 ꢁ 25 mL) and dried

(MgSO₄). The solvent was removed under vacuum to yield 6.8 g of

3 as a white solid. (Yield: 97%). ¹H NMR (250 MHz, CDCl₃) d ppm:

5.34 (2H, s), 3.40 (3H, s). LCMS: t_R= 1.87 min, 346 (M+H)⁺calcd

for C₅H₅Br₃N₂O.

4.1.2.5. {2-[4-(4-Bromo-5-pyridin-4-yl-1H-imidazol-2-yl)-phen-

oxy]-ethyl}-dimethyl-amine (7). {2-[4-(4-Bromo-1-methoxy-

methyl-5-pyridin-4-yl-1H-imidazol-2-yl)-phenoxy]-ethyl}-di-

methyl-amine 6 (70 mg, 0.16 mmol) was dissolved in 1 mL 5 M

HCl and the reaction heated at 60 °C for 1 h. The solvents were re-

moved under vacuum to yield 70 mg of imidazole 7 as HCl salt.

(Yield: quantitative). ¹H NMR (360 MHz, MeOH) d ppm 8.82 (2H,

d, J = 6.8 Hz,), 8.65 (2H, d, J = 6.8 Hz,), 8.06 (2H, d, J = 9.1 Hz), 7.23

(2H, d, J = 9.1 Hz,), 4.39–4.55 (2H, m), 3.57–3.75 (2H, m), 3.02

(7H, s). LCMS: t_R= 0.94 min, 387 (M+H)⁺calcd for C₁₈H₁₉BrN₄O.

4.1.3. General method for the synthesis of {2-[4-(4-aryl-5-pyri-

din-4-yl-1H-imidazol-2-yl)-phenoxy]-ethyl}-dimethyl-amines

Imidazole 6 or 7 (1 equiv), boronic ester (1.2–2 equiv), PPh₃

(0.1 equiv), and K₂CO₃(8–10 equiv) were suspended in a 2:1 mix-

ture of DME/H₂O (3–9 mL). The suspension was stirred vigorously

4.1.2.2. 4-(4,5-Dibromo-1-methoxymethyl-1H-imidazol-2-yl)-

phenol (4). 2,4,5-Tribromo-1-methoxymethyl-1H-imidazole

3

(2.0 g, 5.72 mmol), 4-hydroxyphenyl-boronic acid (0.821 g,

5.95 mmol), 2 M K₂CO₃solution (6.4 mL) were suspended in a

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6943

whilst de-gassing with N₂for 20 min before adding Pd(OAc)₂

(0.01 equiv). The mixture was then reﬂuxed for 2 h to overnight,

cooled to room temperature, acidiﬁed to pH1 with 1 M HCl and

washed with EtOAc (3 ꢁ 5 mL). The aqueous layer was basiﬁed

with 2 M NaOH to pH14 and extracted with EtOAc (3 ꢁ 5 mL). This

organic layer was dried (MgSO₄) and the solvent removed under

vacuum. The residue was puriﬁed by preparative HPLC to yield

the desired product as TFA salt.

4.96 (2H, s), 4.15 (2H, t, J = 5.7 Hz), 3.78 (3H, s), 3.29 (3H, s),

2.65–2.90 (6H, m), 2.37 (6H, s). LCMS: t_R= 1.41 min, 641 (M+H)⁺

calcd for C₃₉H₄₀N₆O₃.

4.1.3.5. [2-(4-{4-[1-(4-Methoxy-benzyl)-1H-benzo[g]indazol-7-

yl]-1-methoxymethyl-5-pyridin-4-yl-1H-imidazol-2-yl}-phen-

oxy)-ethyl]-dimethyl-amine (8f). The standard coupling proce-

dure using 1-(4-methoxy-benzyl)-7-(4,4,5,5-tetramethyl-[1,3,2]

dioxaborolan-2-yl)-1H-benzo[g]indazole 43 (53 mg, 0.13 mmol)

and imidazole 6 (71 mg, 0.16 mmol) resulted in 8f (33 mg, crude).

Taken into next step without further puriﬁcation. LCMS: t_R= 1.47

min, 639 (M+H)⁺calcd for C₃₉H₃₈N₆O₃.

4.1.3.1. {2-[4-(5-{4-[1-(4-Methoxy-benzyl)-1H-pyrazol-3-yl]-phe-

nyl}-1-methoxymethyl-4-pyridin-4-yl-1H-imidazol-2-yl)-phen-

oxy]-ethyl}-dimethyl-amine (8a). Using the general coupling

procedure with 1-(4-methoxy-benzyl)-3-[4-(4,4,5,5-tetramethyl-

[1,3,2]dioxaborolan-2-yl)-phenyl]-1H-pyrazole 29a (77 mg, 0.20

mmol) and MOM protected imidazole 6 (71 mg, 0.16 mmol) gave

the title compound which was puriﬁed by chromatography on sil-

ica gel. Yield = 47 mg (48%). ¹H NMR (360 MHz, MeOH) d ppm: 8.63

(2H, d, J = 5.9 Hz), 7.79 (2H, d, J = 8.6 Hz), 7.73 (2H, d, J = 8.2 Hz),

7.61 (1H, d, J = 2.3 Hz), 7.54 (2H, d, J = 5.9 Hz), 7.46 (2H, d,

J = 8.6 Hz), 7.24 (2H, d, J = 8.6 Hz), 7.17 (2H, d, J = 8.6 Hz), 6.90

(2H, d, J = 8.6 Hz), 6.64 (1H, d, J = 2.3 Hz), 5.29 (2H, s), 5.03 (2H,

s), 4.22 (2H, t, J = 5.4 Hz), 3.78 (3H, s), 3.26 (3H, s), 2.86 (2H, t,

J = 5.2 Hz), 2.41 (6H, s). LCMS: t_R= 1.40 min, 615 (M+H)⁺calcd for

4.1.3.6. 7-{2-[4-(2-Dimethylamino-ethoxy)-phenyl]-1-methoxy-

methyl-5-pyridin-4-yl-1H-imidazol-4-yl}-2,4,4a,5-tetrahydro-

indeno[1,2-c]pyridazin-3-one (8g). The standard coupling pro-

cedure using 7-(4,4,5,5-tetramethyl-[1,3,2]dioxaborolan-2-yl)-2,4,

4a,5-tetrahydro-indeno[1,2-c]pyridazin-3-one 26 (60 mg, 0.19

mmol) and imidazole

6 (99 mg, 0.23 mmol) resulted in 8g

(65 mg, 63%). ¹H NMR (360 MHz, CDCl₃) d ppm: 8.72 (2H, d,

J = 6.1 Hz), 8.48 (1H, s), 7.79 (2H, d, J = 8.9 Hz), 7.67 (1H, br s),

7.56 (1H, d, J = 7.9 Hz), 7.47 (2H, d, J = 6.1 Hz), 7.40 (1H, d,

J = 8.4 Hz), 7.06 (2H, d, J = 8.9 Hz), 4.95 (2H, s), 4.16 (2H, t,

J = 5.7 Hz), 3.10–3.43 (5H, m), 2.91 (1H, dd, J = 16.6, 7.3 Hz), 2.81

(2H, t, J = 5.7 Hz), 2.70 (1H, dd, J = 16.3, 5.9 Hz), 2.29–2.42 (7H,

m). LCMS: t_R= 1.11 min, 537 (M+H)⁺calcd for C₃₁H₃₂N₆O₃.

C₃₇H₃₈N₆O₃.

4.1.3.2. {2-[4-(5-{4-[1-(4-Methoxy-benzyl)-4-methyl-1H-pyrazol-

3-yl]-phenyl}-1-methoxy methyl-4-pyridin-4-yl-1H-imidazol-2-

yl)-phenoxy]-ethyl}-dimethyl-amine (8b). Standard coupling

procedure using 1-(4-methoxy-benzyl)-4-methyl-3-[4-(4,4,5,5-

tetramethyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-1H-pyrazole 29b

4.1.3.7. {2-[4-(5-{4-[4-Chloro-1-(4-methoxy-benzyl)-1H-pyra-

zol-3-yl]-phenyl}-4-pyridin-4-yl-1H-imidazol-2-yl)-phenoxy]-

ethyl}-dimethyl-amine (9d). Standard coupling procedure using

4-chloro-1-(4-methoxy-benzyl)-3-[4-(4,4,5,5-tetramethyl-[1,3,2]-

dioxaborolan-2-yl)-phenyl]-1H-pyrazole 33 (30 mg, 0.071 mmol)

and deprotected imidazole hydrochloride 7 (27 mg, 0.059 mmol)

gave the title compound 9d which was puriﬁed by chromatogra-

phy on silica gel. Yield = 18 mg (51%). ¹H NMR (360 MHz, MeOH)

d ppm: 8.41 (2H, d, J = 5.9 Hz), 7.90–7.96 (4H, m), 7.77 (1H, s),

7.51–7.58 (4H, m), 7.26 (2H, d, J = 9.1 Hz), 7.06 (2H, d, J = 9.1 Hz),

6.90 (2H, d, J = 8.6 Hz), 5.24 (2H, s), 4.16 (2H, t, J = 5.4 Hz), 3.76

(3H, s), 2.83 (2H, t, J = 5.4 Hz), 2.39 (6H, s). LCMS: t_R= 1.41 min,

(63 mg, 0.16 mol) and MOM protected imidazole

6 (57 mg,

0.13 mmol) gave the title compound which was puriﬁed by chro-

matography on silica gel. Yield = 60 mg (73%). ¹H NMR (250 MHz,

CDCl₃) d ppm; 8.66 (2H, d, J = 5.9 Hz), 7.79 (2H, d, J = 8.7 Hz), 7.61

(2H, d, J = 11.5 Hz), 7.55 (2H, d, J = 11.2 Hz), 7.47 (2H, d,

J = 6.1 Hz), 7.21 (2H, d, J = 8.5 Hz), 7.11 (1H, s), 7.05 (2H, d,

J = 8.7 Hz), 6.87 (2H, d, J = 8.7 Hz), 5.20 (2H, s), 4.95 (2H, s), 4.14

(2H, t, J = 5.7 Hz), 3.78 (3H, s), 3.28 (3H, s), 2.78 (2H, t, J = 5.6 Hz),

2.36 (6H, s), 2.17 (3H, s). LCMS: t_R= 1.39 min, 629 (M+H)⁺calcd

for C₃₈H₄₀N₆O₃.

605 (M+H)⁺calcd for C₃₅

H ₃₂ClN₅O₂.

4.1.4. Synthesis and characterization of intermediates 11–16

(Scheme 2)

4.1.3.3. 3-(4-{2-[4-(2-Dimethylamino-ethoxy)-phenyl]-3-meth-

oxymethyl-5-pyridin-4-yl-3H-imidazol-4-yl}-phenyl)-1-(4-met-

hoxy-benzyl)-1H-pyrazole-4-carbonitrile (8c). Standard

coupling procedure using 1-(4-methoxy-benzyl)-3-[4-(4,4,5,5-tet-

ramethyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-1H-pyrazole-4-carbo-

nitrile 37 (65 mg, 0.16 mol) and MOM protected imidazole 6

(57 mg, 0.13 mmol) gave the title compound which was puriﬁed

by chromatography on silica gel. Yield = 61 mg (73%). ¹H NMR

(250 MHz, CDCl₃) d ppm: 8.62 (2H, d, J = 6.1 Hz), 7.82 (2H, d,

J = 8.5 Hz), 7.72 (2H, d, J = 8.8 Hz), 7.63 (1H, s), 7.54 (2H, d,

J = 8.5 Hz), 7.39 (2H, d, J = 6.1 Hz), 7.17 (2H, d, J = 8.8 Hz), 6.98

(2H, d, J = 8.8 Hz), 6.83 (2H, d, J = 8.8 Hz), 5.18 (2H, s), 4.94 (2H,

s), 4.07 (3H, t, J = 5.7 Hz), 3.73 (3H, s), 3.21 (3H, s), 2.70 (2H, t,

J = 5.6 Hz), 2.29 (6H, s). LCMS: t_R= 1.50 min, 640 (M+H)⁺calcd for

4.1.4.1. 1-Methoxyimino-indan-5-carbaldehyde (11). 5-Bromo-

2,3-dihydro-1H-inden-1-one O-methyl oxime 10 (3.71 g, 15.45

mmol) was dissolved in dry THF (75 mL). Solution was de-gassed

over N₂and then cooled to ꢂ78 °C. nBuLi (1.6 M in THF, 10.63 mL,

17.0 mmol) was added dropwise. Stirring at ꢂ78 °C was continued

for a further 10 min, before adding dry DMF (1.32 mL). Reaction

was allowed to gradually warm to room temperature over 3 h. Reac-

tion was then quenched with saturated NaHCO₃and THF was evap-

orated under vacuum. Residue was dissolved in EtOAc and washed

with water, brine and dried (MgSO₄), and solvent removed to leave

a brown oil. Puriﬁcation was achieved by Flash Column Chromatog-

raphy on Silica gel, eluting in 5–10% EtOAc/heptane. Yield: 2.27 g

(78%). ¹H NMR (250 MHz, CDCl₃) d ppm: 9.95 (1H, s), 7.64–7.80

(3H, m), 3.95 (3H, s), 2.98–3.09 (2H, m), 2.81–2.93 (2H, m). LCMS:

t_R= 1.84 min, 190 (M+H)⁺calcd for C₁₁H₁₁NO₂.

C₃₈H₃₇N₇O₃.

4.1.3.4. [2-(4-{4-[1-(4-Methoxy-benzyl)-4,5-dihydro-1H-benzo

[g]indazol-7-yl]-1-methoxy methyl-5-pyridin-4-yl-1H-imidazol-

2-yl}-phenoxy)-ethyl]-dimethyl-amine

(8e). The

standard

4.1.4.2. 5-(1-Hydroxy-2-pyridin-4-yl-2-TBDMS-hydroxy-propyl)-

indan-1-one O-methyl-oxime (12). 4-(tert-Butyl-dimethyl-sila-

nyloxymethyl)-pyridine (2.80 g, 12.5 mmol) was dissolved in dry

THF (20 mL) and the solution was de-gassed over N₂. Reaction

was cooled to ꢂ40 °C (MeCN:Cardice) and the LDA (2 M sol,

7.43 mL, 11.4 mmol) was added dropwise. Reaction was stirred at

ꢂ40 °C under N₂for 30 min, whereupon a de-gassed solution of

coupling procedure using 1-(4-methoxybenzyl)-7-(4,4,5,5-tetra-

methyl-1,3,2-dioxaborolan-2-yl)-4,5-dihydro-1H-benzo[g]indazole

40 (37 mg, 0.09 mmol) and imidazole 6 (31 mg, 0.07 mmol) re-

sulted in 8e (13 mg, 29%). ¹H NMR (250 MHz, CDCl₃) d ppm: 8.68

(2H, br s), 7.67–7.86 (3H, m), 7.36–7.58 (3H, m), 7.16–7.34 (5H,

m), 7.06 (2H, d, J = 10.1 Hz), 6.88 (1H, d, J = 8.7 Hz), 5.25 (2H, s),

6944

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

1-methoxyimino-indan-5-carbaldehyde 11 (2.16 g, 11.4 mmol) in

THF (20 mL), was added dropwise and the reaction mixture gradu-

ally allowed to warm to room temperature over 3 h. Reaction mix-

ture was quenched with satd Na₂HCO₃and the THF was removed

under vacuum. Residue was taken up in EtOAc, washed with satd

Na₂HCO₃, water, brine and dried (MgSO₄). Solvent was removed

under vacuum to leave a yellow oil which was puriﬁed by Flash

Column Chromatography on silica gel, eluting the desired product

in 50–100% EtOAc/heptane. Yield: 4.52 g (96%) as a yellow oil. ¹H

NMR (250 MHz, CDCl₃) d ppm: 8.60–8.72 (2H, m), 7.71–7.82 (1H,

m), 7.10–7.36 (4H, m), 4.69–5.00 (2H, m), 4.13–4.20 (3H, m),

3.00–3.21 (4H, m), 0.96–1.10 (9H, m), ꢂ0.13 (6H, m). LCMS:

t_R= 1.85 min 413 (M+H)⁺calcd for C₂₃H₃₀N₂O₃Si.

compound 15a to give O-methyl oxime 15b (197 mg, 29%) as a yel-

low oil. ¹H NMR (250 MHz, MeOD) d ppm: 8.42 (2H, dd, J = 4.8,

1.29 Hz), 7.90–7.97 (2H, m), 7.68 (1H, d, J = 8.1 Hz), 7.45–7.57

(3H, m), 7.37 (1H, dd, J = 8.1, 1.2 Hz), 7.07 (2H, d, J = 9.0 Hz), 4.16

(2H, t, J = 5.4 Hz), 3.96 (3H, s), 2.99–3.10 (2H, m), 2.85–2.95 (2H,

m), 2.80 (2H, t, J = 5.5 Hz), 2.36 (s, 6H). LCMS: t_R= 1.19 min, 468

(M+H)⁺calcd for C₂₈H₂₉N₅O₂.

4.1.4.7. (E)-tert-Butyl 4-(4-(1-(methoxyimino)-2,3-dihydro-1H-

inden-5-yl)-5-(pyridin-4-yl)-1H-imidazol-2-yl)piperidine-1-car-

boxylate (15c). 4-Formyl-piperidine-1-carboxylic acid tert-butyl

ester (141 mg 0.66 mmol) was reacted with diketone 14 (150 mg,

0.51 mmol) as described for 15a to give O-methyl oxime 15c

(229 mg, 92%). ¹H NMR (250 MHz, MeOD) d ppm: 8.41 (2H, d,

J = 5.94 Hz), 7.67 (1H, d, J = 8.22 Hz), 7.45 (3H, d, J = 12.03 Hz),

7.32 (1H, d, J = 8.22 Hz), 4.22 (2H, d, J = 12.94 Hz), 3.96 (3H, s),

2.83–3.11 (7H, m), 1.94–2.07 (2H, m), 1.71–1.92 (2H, m), 1.48 (s,

9H). LCMS: t_R= 1.55 min, 488 (M+H)⁺calcd for C₂₈H₃₃N₅O₃.

4.1.4.3. 5-(1,2-Dihydroxy-2-pyridin-4-yl-ethyl)-indan-1-one

O-methyl-oxime (13). 5-(1-Hydroxy-2-pyridin-4-yl-2-TBDMS-

hydroxy-propyl)-indan-1-one O-methyl-oxime 12 (1.24 g, 3.01

mmol) was dissolved in dry THF (50 mL) and TBAF (1 M in THF)

(4.81 mL, 4.81 mmol) was added. Reaction mixture was stirred at

room temperature over night. Solvent was removed under vacuum

and residue was puriﬁed by Flash Column Chromatography on silica

gel, eluting the desired product in 5% MeOH/DCM as a colourless so-

lid. Yield: 797 mg (89%). ¹H NMR (250 MHz, CDCl₃) d ppm: 8.39 (1H,

dt, J = 4.30, 1.81 Hz), 7.54 (1H, d, J = 8.07 Hz), 6.94–7.13 (4H, m),

4.93–5.17 (1H, m), 4.62–4.78 (1H, m), 3.98 (3H, s), 2.77–3.02 (4H,

m). LCMS: t_R= 1.06 min, 299 (M+H)⁺calcd for C₁₇H₁₈N₂O₃.

4.1.4.8. 5-(2-(4-(4-Methylpiperazin-1-yl)phenyl)-5-(pyridin-4-

yl)-1H-imidazol-4-yl)-2,3-dihydro-1H-inden-1-one (16a). 5-{2-

[2-(4-(4-Methylpiperazin-1-yl)-5-(pyridine-4-yl)-1H-imidazol-4-

yl)]}-1H-inden-1-one-O-methyl-oxime 15a (93.5 mg, 0.20 mmol)

was dissolved in dioxane (4 mL) and 5 M HCl (2 mL) was added.

The reaction was heated to 80 °C overnight. Solvents were evapo-

rated under vacuum and residue was puriﬁed by Flash column

chromatography on silica gel, eluting the desired product in

5–50% MeOH/DCM to give ketone 16a. Yield: 87 mg (99%). ¹H

NMR (250 MHz, MeOD) d ppm: 8.83 (2H, br s), 8.05–8.27 (4H,

m), 7.49–7.96 (3H, m), 7.30 (2H, d, J = 7.3 Hz), 4.17 (2H, br s),

3.55–3.78 (5H, m), 3.34–3.46 (3H, m), 2.91–3.06 (4H, m),

2.68–2.84 (1H, m). LCMS: t_R= 1.08 min, 450 (M+H)⁺calcd for

4.1.4.4. 1-(1-Methoxyimino-indan-5-yl)-2-pyridin-4-yl-ethane-

1,2-dione (14). DMSO (705

DCM (6.4 mL) and cooled to ꢂ78 °C under N₂. Oxalyl chloride

(653 L, 7.50 mmol) was then added dropwise and the solution

was stirred for 20 min. A solution of diol 13 (671 mg, 2.25 mmol)

in dry DCM (6.4 mL) and DMSO (959 L, 6.0 mmol) was added

ll, 9.95 mmol) was dissolved in dry

l

dropwise at ꢂ78 °C and the solution was stirred for 30 min. Trieth-

ylamine (1.94 mL, 14.0 mmol) was then added dropwise and the

reaction mixture was gradually allowed to warm to room temper-

ature over 3 h. Reaction mixture was diluted with water and ex-

tracted with DCM (3 ꢁ 20 mL), washed with brine, dried (MgSO₄)

and evaporated to dryness under vacuum to leave a yellow solid.

Puriﬁed by Flash column chromatography on silica gel, eluting

the desired product in 50–100% EtOAc/heptane. Yield: 1.15 g

(quantitative) as a yellow solid. ¹H NMR (250 MHz, CDCl₃) d

ppm: 8.78–9.06 (2H, m), 7.71–7.97 (5H, m), 4.04 (3H, s), 3.04–

3.18 (2H, m), 2.81–3.02 (2H, m). LCMS: t_R= 2.07 min, 295 (M+H)⁺

calcd for C₁₇H₁₄N₂O₃.

C

₂₈H₂₈N₆O.

4.1.4.9. {2-[4-(5-Indanyl-1-one)-5-pyridin-4-yl-1H-imidazol-2-

yl]-phenoxy]-ethyl}-dimethyl-amine (16b). 5-{2-[4-(2-Dimeth-

ylamino-ethoxy)-phenyl]-5-pyridin-4-yl-1H-imidazol-4-yl}-indan-

1-one O-methyl-oxime 15b (164 mg, 0.35 mmol) was treated as

described for compound 16a to give ketone 16b (36 mg, 23%) as

a yellow oil. ¹H NMR (250 MHz, MeOD) d ppm: 8.47 (2H, d,

J = 6.1 Hz), 7.97 (2H, d, J = 8.8 Hz), 7.72 (2H, s), 7.55 (3H, br s),

7.10 (2H, d, J = 8.8 Hz), 4.19 (2H, t, J = 5.4 Hz), 3.15–3.40 (2H, m),

2.62–2.91 (4H, m), 2.38 (6H, s). LCMS: t_R= 1.08 min, 439 (M+H)⁺

calcd for C₂₇H₂₆N₄O₂.

4.1.4.5. 5-{2-[2-(4-(4-methylpiperazin-1-yl)-5-(pyridine-4-yl)-

1H-imidazol-4-yl)]}-1H-inden-1-one-O-methyl-oxime (15a). 4-

(4⁰-N-Methyl-pyrazinyl)-benzaldehyde (135 mg, 0.66 mmol) was

dissolved in acetic acid (3 mL), diketone 14 (150 mg, 0.51 mmol)

and ammonium acetate (393 mg, 5.10 mmol) was added. The reac-

tion mixture was heated to 100 °C and stirred for 1 h. Reaction mix-

ture was basiﬁed with NH₃, and diluted with water. Aqueous was

extracted with DCM (3 ꢁ 10 mL), dried (MgSO₄) ﬁltered and evapo-

rated under vacuum. Puriﬁcation was carried out by Flash column

chromatography on silica gel, eluting the desired product in

5–10% MeOH/DCM, to yield imidazole 15a. Yield: 93.5 mg (38%).

¹H NMR (250 MHz, MeOD) d ppm: 8.43 (2H, dd, J = 4.7, 1.5 Hz),

7.89 (2H, d, J = 9.0 Hz), 7.69 (1H, d, J = 8.1 Hz), 7.48–7.58 (3H, m),

7.38 (1H, s), 7.08 (2H, d, J = 9.0 Hz), 3.97 (3H, s), 3.25–3.35 (4H,

m), 3.02–3.08 (2H, m), 2.88–2.96 (2H, m), 2.68–2.78 (4H, m), 2.44

(3H, s). LCMS: t_R= 1.21 min, 479 (M+H)⁺calcd for C₂₉H₃₀N₆O.

4.1.4.10. tert-Butyl 4-(4-(1-oxo-2,3-dihydro-1H-inden-5-yl)-5-

(pyridin-4-yl)-1H-imidazol-2-yl)piperidine-1-carboxylate (16c).

tert-Butyl 4-(4-(1-(methoxyimino)-2,3-dihydro-1H-inden-5-yl)-5-

(pyridin-4-yl)-1H-imidazol-2-yl)piperidine-1-carboxylate 15c

(229 mg, 0.47 mmol) was dissolved in 5 M HCl (2 mL) and dioxane

(4 mL) and heated at 80 °C overnight. All solvents were removed

under vacuum to leave a brown oil (LCMS: t_R= 0.86 min, 359

(M+H)⁺calcd for C₂₂H₂₂N₄O). This was dissolved in THF (15 mL)

and water (2 mL) before adding DIPEA (90 lL, 0.52 mmol) and

(BOC)₂O (256 mg, 1.17 mmol). After stirring at room temperature

overnight, the THF was evaporated, further water added and ex-

tracted with DCM (ꢁ3). The combined organic phases were dried

(MgSO₄), ﬁltered and evaporated to leave an oil which was puriﬁed

by Flash column chromatography on silica gel, eluting the desired

compound with 0–10% MeOH/DCM. Yield = 122 mg (57%). ¹H NMR

(250 MHz, MeOD) d ppm: 8.45 (2H, br s), 7.57–7.79 (2H, m), 7.34–

7.54 (3H, m), 4.22 (2H, d, J = 12.6 Hz), 3.15–3.40 (2H, m), 2.86–3.08

(3H, m), 2.72 (2H, t), 1.95–2.09 (2H, m), 1.84 (2H, t, J = 12.0 Hz),

1.48 (9H, s). LCMS: t_R= 1.44 min, 459 (M+H)⁺calcd for C₂₇H₃₀N₄O₃.

4.1.4.6. 5-{2-[4-(2-Dimethylamino-ethoxy)-phenyl]-5-pyridin-

4-yl-1H-imidazol-4-yl}-indan-1-one O-methyl-oxime (15b). 4-

(2-Dimethylamino-ethoxy)-benzaldehyde (367 mg, 1.90 mmol) was

reacted with diketone 14 (428 mg, 1.45 mmol) as described for

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6945

4.1.5. Synthesis and characterization of intermediates 17–22

(Scheme 4)

m), 3.42–3.50 (2H, m). HRMS: (M+H)⁺calcd for C₁₉H₁₃NO₄:

320.0923, found: 320.0915.

4.1.5.1. Methyl, 2,3-dibromofuranyl-carboxylate (17). To a solu-

tion of the 2,3-dibromofuran-5-yl carboxylic acid (4 g, 14.8 mmol)

in DCM (60 mL) was added oxalyl chloride (3.88 mL, 44.5 mmol)

under nitrogen. The reaction mixture was cooled to 0 °C and

0.2 mL of DMF added. The reaction was stirred at room tempera-

ture for 4 h, solvent was removed under vacuum and azeotroped

with DCM. The crude was dissolved in DCM (40 mL) and MeOH

added at 0 °C. The reaction was stirred at room temperature for

1 h and solvent removed under vacuum. Yielded product was ob-

tained as a pure solid (3.85 g) in 91% yield. ¹H NMR (250 MHz,

CDCl₃-d) d ppm: 7.11 (1H, s), 3.83 (3H, s).

4.1.5.6. 2-(4-Pyridyl)-3-(5-indanyl-oxime)-furanyl-5-carbonyl-

4-N-methyl-piperidine (22a). To a solution of acid 21 (300 mg,

0.95 mmol) in DMF was added DIC (13 mg, 1.05 mmol), methyl

piperazine (105 mg, 1.05 mmol), HOBt (141 mg, 1.05 mmol), and

TEA (146 lL, 1.05 mmol). The reaction was heated to 60 °C for

4 h. Solvent was removed under vacuum and the residue was puri-

ﬁed by chromatography on silica gel eluting with DCM/MeOH. This

yielded 150 mg (40%) of pure product as a white powder. LCMS:

t_R= 0.97 min, 402 (M+H)⁺calcd for C₂₄H₂₃N₃O₃.

4.1.5.7. 5-{5-[4-(2-Dimethylamino-ethyl)-piperidine-1-carbon-

yl]-2-pyridin-4-yl-furan-3-yl}-indan-1-one (22b). Acid 21 (250

mg, 0.78 mmol) was coupled with dimethyl-(2-piperazin-1-yl-

ethyl)-amine (135 mg, 0.86 mmol) as described for 22a to give

22b. Yield = 176 mg (49%). ¹H NMR (360 MHz, CDCl₃) d ppm:

8.58 (2H, d, J = 6.4 Hz), 7.81 (1H, d, J = 7.9 Hz), 7.52 (1H, s), 7.41

(1H, d, J = 7.9 Hz), 7.37 (2H, d, J = 6.1 Hz), 7.14 (1H, s), 3.91 (4H,

br s), 3.12–3.25 (2H, m), 2.69–2.85 (2H, m), 2.40–2.69 (8H, m),

2.28 (6H, s). LCMS: t_R= 1.05 min, 459 (M+H)⁺calcd for C₂₇H₃₀N₄O₃.

4.1.5.2. Methyl, 2-(4-pyridyl),3-bromofuranyl-carboxylate (18).

Dry DMF (150 mL) was added to a dry mixture of the dibromofuran

17 (5.68 g, 20.0 mmol), 4-pyridyl boronic acid (2.7 g, 22.0 mmol),

Cs₂CO₃(19 g, 60 mmol), AsPh₃(0.610 g, 2 mmol), and (PPh₃)₂PdCl₂

(1.12 g, 1.6 mmol). This solution was de-gassed with nitrogen for

20 min before being heated to 90 °C overnight. The majority of

DMF was then removed under vacuum and the crude diluted with

EtOAc. The crude organic was washed with NaHCO₃(1% aqueous)

and dried (MgSO₄). Puriﬁcation followed using ﬂash column chro-

matography on silica gel eluting with EtOAc/heptane to yield

2.12 g of product (37%) as an off-white solid. ¹H NMR (250 MHz,

CDCl₃-d) d ppm: 8.73 (2H, d, J = 6.2 Hz), 7.93–7.99 (2H, m), 7.30

(1H, s), 3.95 (3H, s). LCMS: t_R= 1.35 min, 282 (M+H)⁺calcd for

4.1.5.8. 5-{5-[4-Morpholine-1-carbonyl]-2-pyridin-4-yl-furan-

3-yl}-indan-1-one (22c). Acid 21 (766 mg; 2.4 mmol) was cou-

pled with morpholine (0.23 mL; 2.6 mmol) as described for 22a

to give 22c. Yield = 520 mg (56%). ¹H NMR (250 MHz, CDCl₃) d

ppm: 8.58 (2H, dd, J = 4.6, 1.6 Hz), 7.81 (1H, d, J = 7.9 Hz), 7.52

(1H, s), 7.33–7.44 (3H, m), 7.18 (1H, s), 3.75–4.07 (8H, m), 3.13–

3.23 (2H, m), 2.72–2.81 (2H, m). LCMS: t_R= 1.29 min, 389 (M+H)⁺

calcd for C₂₃H₂₀N₂O₄.

C₁₁H₈BrNO₃.

4.1.5.3. Methyl 4-(1-(methoxyimino)-2,3-dihydro-1H-inden-5-

yl)-5-(pyridin-4-yl)furan-2-carboxylate (19). Dry DMF (150 mL)

was added to a dry mixture of 18 (0.590 g, 2.1 mmol), 1-(methoxy-

imino)-2,3-dihydro-1H-inden-5-ylboronic acid (0.512 g, 2.5 mmol),

Cs₂CO₃(2.05 g, 6.3 mmol), AsPh₃(0.640 g, 2.1 mmol), and (PPh₃)₂

PdCl₂(1.05 g, 1.5 mmol). This solution was de-gassed with nitrogen

for 20 min before being heated to 90 °C overnight. The majority of

DMF was then removed under vacuum and the crude diluted with

EtOAc. The crude organic was washed with 1% aqueous NaHCO₃

and dried (MgSO₄). Puriﬁcation followed using ﬂash column chro-

matography on silica gel eluting with EtOAc/heptane to yield

0.519 g of product (69%) as an off-white solid. ¹H NMR (250 MHz,

CDCl₃-d) d ppm: 8.48 (2H, d, J = 4.8 Hz), 7.66 (1H, d, J = 8.1 Hz),

7.37–7.42 (2H, m), 7.15–7.23 (3H, m), 3.93 (3H, s), 3.87 (3H, s),

2.94–3.02 (2H, m), 2.81–2.90 (2H, m). LCMS: t_R= 1.84 min, 363

(M+H)⁺calcd for C₂₁H₁₈N₂O₄.

4.1.5.9. tert-butyl 4-(4-(1-oxo-2,3-dihydro-1H-inden-5-yl)-5-

(pyridin-4-yl)furan-2-carbonyl)piperazine-1-carboxylate (22d).

Acid 21 (385 mg; 1.21 mmol) was coupled with N-Boc piperazine

(248 mg; 1.33 mmol) as described for 22a to give 22d and was

used in the next step without further puriﬁcation. LCMS: t_R=

1.71 min, 488 (M+H)⁺calcd for C₂₈H₂₉N₃O₅.

4.1.6. Synthesis and characterization of intermediates 24–33

(Scheme 5)

4.1.6.1. (5-Bromo-1-oxo-indan-2-yl)-acetic acid ethyl ester (24). LDA

(2 M in hexane, 5.21 mL, 10.42 mmol) was mixed with THF (5 mL)

and cooled to ꢂ78 °C. 5-Bromo-indan-1-one (2.0 g, 9.48 mmol) in

THF (20 mL) was added dropwise under a stream of nitrogen and

the solution was stirred for 30 min. Bromo-acetic acid ethyl ester

(1.74 g, 10.42 mmol) in HMPA (1.5 mL) and THF (5 mL) was added

dropwise to the reaction mixture, which was then allowed to

warm up to room temperature overnight. The reaction was

quenched with NH₄Cl saturated solution (50 mL) and extracted

with EtOAc (3 ꢁ 50 mL), the organic layer was dried (MgSO₄), ﬁl-

tered and the solvent removed under vacuum. The residue was

puriﬁed by chromatography using a stepped gradient of 0–8%

EtOAc in heptane to yield 24 as a white solid. Yield: 577 mg, 20%.

¹H NMR (360 MHz, CDCl₃) d ppm: 7.59–7.68 (2H, m), 7.53 (1H, d,

J = 9.1 Hz), 4.14 (2H, q, J = 7.0 Hz), 3.44 (1H, dd, J = 17.1, 7.8 Hz),

2.83–3.06 (3H, m), 2.62–2.73 (1H, m), 1.23 (3H, t, J = 7.0 Hz). LCMS:

t_R= 2.07 min, 297 (M+H)⁺calcd for C₁₃H₁₃BrO₃. HRMS: (M+H)⁺

calcd for C₁₃H₁₃BrO₃: 297.0126, found: 297.0119.

4.1.5.4. 2-(4-Pyridyl)-3-(5-indanyl-N-O-methyl-oxime)-furanyl-

5-carboxylic acid (20). To a solution of 19 (335 mg, 0.93 mmol)

in 5 mL THF/MeOH (4:1) was added 2 M aqueous NaOH (0.95 mL,

1.90 mmol). The reaction was stirred vigorously at room tempera-

ture for 90 min and solvent was removed under vacuum to yield

220 mg product (68%) as a white solid. ¹H NMR (250 MHz,

DMSO-d₆) d ppm: 8.26 (2H, d, J = 6.8 Hz), 7.97 (2H, d, J = 6.5 Hz),

7.65 (1H, d, J = 8.1 Hz), 7.53 (2H, s), 7.38 (1H, d, J = 7.8 Hz), 3.88

(3H, s), 2.96–3.08 (2H, m), 2.76–2.88 (2H, m). LCMS: t_R= 1.40 min,

349 (M+H)⁺calcd for C₂₀H₁₆N₂O₄.

4.1.5.5.4-(1-Oxo-2,3-dihydro-1H-inden-5-yl)-5-(pyridin-4-yl)fu-

ran-2-carboxylic acid (21). To a solution of starting acid 20

(835 mg, 2.41 mmol) dioxane/acetone (8 mL:1.5 mL) was added

5 mL aqueous hydrochloric acid. The reaction was heated at 90 °C

for 90 min. Solvent was removed under vacuum to give 1.41 g of

pure product (quantitative, some NaCl present). ¹H NMR

(250 MHz, MeOD) d ppm: 9.48 (2H, d, J = 7.8 Hz), 8.70 (2H, d,

J = 7.7 Hz), 8.45–8.54 (2H, m), 8.24–8.33 (2H, m), 3.90–3.95 (2H,

4.1.6.2. 7-Bromo-2,4,4a,5-tetrahydro-indeno[1,2-c]pyridazin-3-

one (25). Intermediate 24 (250 mg, 0.84 mmol) was dissolved in

EtOH (3 mL) and NH₂NH₂ꢃH₂O (0.15 mL, 4.21 mmol) was added

in EtOH (1 mL) and stirred at room temperature overnight. A pre-

cipitate appeared which was ﬁltered and washed with EtOH

(10 mL) to yield compound 25 as a white solid. Yield: 134 mg,

6946

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

Scheme 5. Synthesis of boronic acids 26, 29a, 29b and 33. Reagents and conditions: (i) 2M LDA, THF, BrCH₂CO₂Et; (ii) H₂NNH₂, EtOH; (iii) bis pinacolato-diboron, KOAc,

Pd(dppf)Cl₂, DMF, 90 °C; (iv) 4-methoxy benzyl chloride, NaH, DMF; (v) NCS, DCM; (vi) (BOC)₂O, DMAP, CH₃CN.

60%. ¹H NMR (360 MHz, CDCl₃) d ppm: 8.54 (1H, br s), 7.53–7.60

(2H, m), 7.47–7.51 (1H, m), 3.37–3.47 (1H, m), 3.15–3.27 (1H,

m), 2.93 (1H, dd, J = 16.6, 7.5 Hz), 2.75 (1H, dd, J = 16.6, 6.1 Hz),

2.37 (1H, t, J = 16.3 Hz). LCMS: t_R= 1.65 min, 265 (M+H)⁺calcd

for C₁₁H₈BrN₂O₃.

1.37 (12H, s). LCMS: t_R= 1.89 min, 313 (M+H)⁺calcd for C₁₇H₂₂

BN₂O₃. HRMS: (M)⁺calcd for C₁₇H₂₁BN₂O₃: 312.1760, found:

312.1747.

4.1.6.4. 3-(4-Bromo-phenyl)-1-(4-methoxy-benzyl)-1H-pyrazole

(28a). 3-(4-Bromophenyl)pyrazole 27a (223 mg, 1 mmol) was

dissolved in anhydrous DMF (2.2 mL) and added to 60% NaH in

mineral oil (44 mg, 1.1 mmol) previously washed with diethyl

ether. After stirring for 20 min at room temperature, 4-methoxy-

benzyl chloride (0.15 mL; 1.1 mmol) was added and stirring con-

tinued at room temperature for a further 4 h. Water (5 mL) was

then added and the emulsion extracted with EtOAc (2 ꢁ 3 mL).

The combined organic phases were washed with saturated sodium

bicarbonate (3 ꢁ 2 mL) and dried (MgSO₄). After ﬁltration and

evaporation, the oil crystallised to give the product (352 mg,

100%). ¹H NMR (250 MHz, CDCl₃) d ppm: 7.66–7.73 (2H, m), 7.51

(2H, d, J = 8.7 Hz), 7.33 (1H, d, J = 2.3 Hz), 7.23 (2H, d, J = 8.8 Hz),

6.89 (2H, d, J = 8.7 Hz), 6.53 (1H, d, J = 2.3 Hz), 5.29 (2H, s), 3.81

(3H, s). The product was used in the next step without further

puriﬁcation.

4.1.6.3. 7-(4,4,5,5-Tetramethyl-[1,3,2]dioxaborolan-2-yl)-2,4,4a,

5-tetrahydro-indeno[1,2-c]pyridazin-3-one (26). Bromo inter-

mediate 25 (145 mg, 0.55 mmol), potassium acetate (160 mg,

1.64 mmol), bis(pinacolato)diboron (208 mg, 0.82 mmol) and 1,1-

bis(diphenylphosphino)ferrocene-palladium(II)dichloride (20 mg,

0.05 mmol) were dissolved in dry DMF (3 mL). The reaction was

heated to 80 °C in a sealed tube for 2 h, quenched with H₂O

(15 mL), extracted into EtOAc (3 ꢁ 10 mL) and dried (Na₂SO₄). Or-

ganic solvent was removed in vacuo. The residue was puriﬁed by

chromatography using a stepped gradient of 10–50% EtOAc in hep-

tane to yield 26 as a white solid (88 mg, 51%). ¹H NMR (360 MHz,

CDCl₃) d ppm: 8.47 (1H, s), 7.83 (1H, s), 7.66–7.82 (2H, m), 3.44

(1H, dd, J = 16.3, 8.6 Hz), 3.11–3.26 (1H, m), 2.93 (1H, dd, J = 16.3,

7.3 Hz), 2.75 (1H, dd, J = 16.3, 5.9 Hz), 2.37 (1H, t, J = 16.3 Hz),

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6947

4.1.6.5. 3-(4-Bromo-phenyl)-1-(4-methoxy-benzyl)-4-methyl-

1H-pyrazole (28b). 4-Methyl-3-(4-bromophenyl)pyrazole 27b

(474 mg, 2.0 mmol) was dissolved in anhydrous DMF (5 mL) and

added to 60% NaH in mineral oil (88 mg, 2.2 mmol) previously

washed with heptane. After stirring for 20 min at room tempera-

ture, 4-methoxybenzyl chloride (0.3 mL, 2.2 mmol) was added

and stirring continued at room temperature for a further 3.5 h.

Water (10 mL) was then added and the emulsion extracted with

EtOAc (2 ꢁ 10 mL). The combined organic phases were washed

with saturated sodium bicarbonate (3 ꢁ 20 mL) and dried (MgSO₄).

After ﬁltration and evaporation, the oil crystallised to give the

product (697 mg, 98%). ¹H NMR (360 MHz, CDCl₃) d ppm: 7.58–

7.63 (2H, m), 7.56 (2H, d), 7.26 (2H, d, J = 8.6 Hz), 7.17 (1H, br s),

6.92 (2H, d, J = 8.6 Hz), 5.25 (2H, s), 3.84 (3H, s), 2.21 (3H, s). LCMS:

t_R= 2.48 min, 357 (M+H)⁺calcd for C₁₈H₁₇BrN₂O. HRMS: (M+H)⁺

calcd for C₁₈H₁₇BrN₂O: 357.0602, found: 357.0613.

0.68 mmol) caused the loss of the BOC group. A 1:1 mix of

unprotected 4-chloro-3-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxa boro-

lan-2-yl)-phenyl]-1H-pyrazole 32 and unprotected 4-chloro-3-(4⁰-

bromophenyl)-pyrazole 30 was isolated after chromatography on

silica gel (116 mg, 61%). ¹H NMR (360 MHz, CDCl₃) d ppm: 7.91

(2H, d, J = 8.7 Hz), 7.79 (2H, d, J = 8.2 Hz), 7.60 (1H, s), 1.38 (12H, s).

LCMS: t_R= 2.05 min, 257/259/261 (M+H)⁺calcd for C₉H₆BrClN₂

and 2.23 min, 305/307 (M+H)⁺calcd for C₁₅H₁₈BClN₂O₂.

4.1.6.11. 4-Chloro-1-(4-methoxy-benzyl)-3-[4-(4,4,5,5-tetramet

hyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-1H-pyrazole (33). The

(1:1) mixture of 30 and 32 resulted from the previous reaction

(116 mg, 0.41 mmol) was dissolved in anhydrous DMF (1 mL)

and added to 60% NaH in mineral oil (18 mg, 0.45 mmol) previ-

ously washed with heptane. After stirring for 10 min at room tem-

perature, 4-methoxybenzyl chloride (0.06 mL, 0.45 mmol) was

added and stirring continued at room temperature overnight.

Water (2.5 mL) was then added and the oil extracted with EtOAc

(2 ꢁ 2 mL). The combined organic phases were washed with satu-

rated NaHCO₃(2 ꢁ 2 mL), dried (MgSO₄), ﬁltered and evaporated

to leave a 2:1 mix of boronyl and bromophenyl PMB protected pyr-

azoles (112 mg, ꢀ50% Br; 75%). ¹H NMR (360 MHz, CDCl₃) d ppm:

7.82 (2H, d, J = 8.6 Hz), 7.56 (2H, d, J = 8.6 Hz), 7.34 (1H, s), 7.25

(2H, dd, J = 8.6, 2.3 Hz), 6.91 (2H, d, J = 8.6 Hz), 5.22 (2H, s), 3.82

(3H, s), 1.37 (12H, s).

The crude mixture was subjected to standard boronylation pro-

cedure (as described for compound 26) affording the desired com-

pound (45 mg, 38%) after chromatography and crystallisation from

heptane. ¹H NMR (360 MHz, CDCl₃) d ppm: 7.85 (2H, d, J = 8.3 Hz),

7.79 (2H, d, J = 8.3 Hz), 7.26 (1H, s), 7.16 (2H, d, J = 8.5 Hz), 6.83

(2H, d, J = 8.6 Hz), 5.16 (2H, s), 3.74 (3H, s), 1.29 (12H, s). LCMS:

t_R= 2.73 min, 425, 4 (M+H)⁺calcd for C₂₃H₂₉BClN₂O₃.

4.1.6.6. 1-(4-Methoxy-benzyl)-3-[4-(4,4,5,5-tetramethyl-[1,3,2]

dioxaborolan-2-yl)-phenyl]-1H-pyrazole (29a). Using the same

boronylation procedure as described for 26, from 3-(4-bromo-phe-

nyl)-1-(4-methoxy-benzyl)-1H-pyrazole 28a (350 mg, 1.0 mmol)

the title compound was obtained (349 mg: 88%). ¹H NMR

(360 MHz, CDCl₃) d ppm: 7.83 (4H, s), 7.32 (1H, d, J = 2.7 Hz),

7.24 (2H, d, J = 8.6 Hz), 6.89 (2H, d, J = 8.6 Hz), 6.60 (1H, d,

J = 2.7 Hz), 5.31 (2H, s), 3.81 (3H, s), 1.37 (12H, s). LCMS:

t_R= 2.49 min, 391 (M+H)⁺calcd for C₂₃H₂₈BN₂O₃. HRMS: (M)⁺calcd

for C₂₃H₂₇BN₂O₃: 390.2229, found: 390.2234.

4.1.6.7. 1-(4-Methoxy-benzyl)-4-methyl-3-[4-(4,4,5,5-tetramet-

hyl-[1,3,2]dioxaborolan-2-yl)-phenyl]-1H-pyrazole (29b). Stan-

dard boronylation procedure using 3-(4-bromo-phenyl)-1-(4-

methoxy-benzyl)-4-methyl-1H-pyrazole 28b (357 mg, 1.0 mmol)

gave the desired compound (284 mg, 70%). ¹H NMR (250 MHz,

CDCl₃) d ppm: 7.86 (2H, d, J = 7.5 Hz), 7.72 (2H, d, J = 7.9 Hz), 7.24

(2H, d, J = 8.7 Hz), 7.14 (1H, s), 6.89 (2H, d, J = 8.7 Hz), 5.24 (2H,

s), 3.81 (3H, s), 2.21 (3H, s), 1.37 (12H, s). HRMS: (M+H)⁺calcd

for C₂₄H₂₉BN₂O₃: 404.2386, found: 404.2390.

4.1.7. Synthesis and characterization of intermediates 34–43

(Scheme 6)

4.1.7.1. 2-(4-Bromo-benzoyl)-3-dimethylamino-acrylonitrile (34).

4-Bromophenyl-2 oxopropionitrile (448 mg, 2.0 mmol) was dis-

solved in DMF (2 mL) and tris(dimethylamino)methane (1.04 mL,

6.0 mmol) was added. The reaction was heated at 70 °C for 4 h before

cooling on ice and adding water (7 mL). The desired product was ﬁl-

tered off, washed with water and dried. Yield = 339 mg (61%). ¹H

NMR (250 MHz, CDCl₃) d ppm: 7.97 (1H, s), 7.68 (2H, d, J = 10.2 Hz),

7.57 (2H, d, J = 9.9 Hz), 3.50 (3H, s), 3.32 (3H, s). The product was used

in the next step without further puriﬁcation.

4.1.6.8. 3-(4-Bromo-phenyl)-4-chloro-1H-pyrazole (30). 3-(4-

Br- omophenyl)-pyrazole 27a (223 mg, 1.0 mmol) and N-chloro-

succinimide (140 mg, 1.05 mmol) were dissolved in DCM (30 mL)

and stirred at room temperature for 84 h. The solvent was evapo-

rated and puriﬁed on a silica gel column with 10–25% EtOAc in

heptane to yield the desired product 30, 190 mg (74%). ¹H NMR

(360 MHz, CDCl₃) d ppm: 7.69 (2H, d, J = 5.8 Hz), 7.62 (1H, s),

7.58 (2H, d, J = 5.4 Hz). LCMS: t_R= 2.04 min, 257 (M+H)⁺calcd for

C₉H₆BrClN₂.

4.1.7.2. 3-(4-Bromo-phenyl)-1H-pyrazole-4-carbonitrile (35). 2-

(4-Bromo-benzoyl)-3-dimethylamino-acrylonitrile 34 (339 mg, 1.22

mmol) was suspended in ethanol (3.4 mL) and hydrazine hydrate

(77 mL, 1.58 mmol) was added. The starting material dissolved and

the reaction was stirred at room temperature for 3 h. The solvent

was evaporated and water (5 mL) added. The solid was ﬁltered off

and chromatographed on silica gel with 60:40 EtOAc/heptane to give

the desired product (233 mg; 77%). ¹H NMR (250 MHz, acetone-d₆): d

ppm: 8.44 (1H, s), 7.92 (2H, d, J = 8.5 Hz), 7.68–7.74 (3H, m). LCMS:

t_R= 1.85 min, 248 (M+H)⁺calcd for C₁₀H₆BrN₃.

4.1.6.9. 3-(4-Bromo-phenyl)-4-chloro-pyrazole-1-carboxylic acid

tert-butyl ester (31). 4-Chloro-3-(4⁰-bromophenyl)-pyrazole 30

(190 mg, 0.74 mmol) was dissolved in acetonitrile (2.3 mL) with

heating. After cooling on ice, BOC anhydride (193 mg, 0.89 mmol)

and DMAP (9 mg; 0.1 mmol) were added and stirring continued

at 0 °C for 2 h and room temperature for 1 h. The reaction was di-

luted with EtOAc (2 mL) and washed with 1.2 N HCl (2 ꢁ 2 mL),

saturated sodium bicarbonate (2 mL) and brine (2 mL) before dry-

ing (Na₂SO₄), ﬁltering and evaporating to leave the product as an

oil. Yield = 244 mg (92%). ¹H NMR (250 MHz, CDCl₃) d ppm: 8.11

(1H, s), 7.87 (2H, d, J = 8.7 Hz), 7.58 (2H, d, J = 8.8 Hz), 1.66 (9H,

s). LCMS: t_R= 2.05 min, 298/300/302 (M+H-56 Boc fragmentation)⁺

calcd for C₁₄H₁₄BrClN₂O₂.

4.1.7.3. 3-(4-Bromo-phenyl)-1-(4-methoxy-benzyl)-1H-pyrazole-

4-carbonitrile (36). 3-(4-Bromo-phenyl)-1H-pyrazole-4-carboni-

trile 35 (233 mg, 0.94 mmol) was dissolved in anhydrous DMF

(2.3 mL)andaddedto60%NaHinmineral oil(41 mg, 1.03 mmol) pre-

viously washed with heptane. After stirring for 10 min at room tem-

perature, 4-methoxybenzyl chloride (0.14 mL; 1.03 mmol) was

added and stirring continued at room temperature for a further 4 h.

Water (5 mL) was then added and an oil separated which crystallised

with stirring. This was ﬁltered and washed with water to leave the

product (309 mg, 89%). ¹H NMR (360 MHz, CDCl₃) d ppm: 7.87 (2H,

4.1.6.10. 4-Chloro-3-[4-(4,4,5,5-tetramethyl-[1,3,2]dioxaboro-

lan-2-yl)-phenyl]-1H-pyrazole (32). Using the same boronylation

procedure as described for compound 26, with 3-(4-bromo-phenyl)-

4-chloro-pyrazole-1-carboxylic acid tert-butyl ester 31 (244 mg,

6948

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

Scheme 6. Synthesis of boronic acids 37, 40 and 43. Reagents and conditions: (i) HC(NMe₂)₃, DMF; (ii) H₂NNN₂ꢃH₂O, EtOH; (iii) 4-methoxy benzyl chloride, NaH, DMF; (iv) bis

pinacolato-diboron, KOAc, Pd(dppf)Cl₂, DMF, 90 °C; (v) AcOH, H₂NNH₂; (vi) DDQ, dioxane.

d, J = 8.6 Hz), 7.72 (1H, s), 7.59 (2H, d, J = 8.6 Hz), 7.27 (2H, t,

J = 4.3 Hz), 6.94 (2H, d, J = 8.6 Hz), 5.28 (2H, s), 3.83 (3H, s). The prod-

uct was used in the next step without further puriﬁcation.

4.53 mmol) was added. The reaction was stirred for 1 h, then fur-

ther tris-dimethylamino methane (657 mg, 4.53 mmol) was added,

and the reaction stirred overnight. The solvents were removed un-

der vacuum and the residue azeothroped with heptane. The crude

was then dissolved in AcOH (2 mL) and treated with NH₂NH₂ꢃH₂O

(227 mg, 4.53 mmol). The reaction was stirred for 15 min, upon

which concd NH₄OH solution was added to pH 11. NaHCO₃satu-

rated solution (10 mL) was added and the aqueous layer was ex-

tracted with DCM (3 ꢁ 10 mL), dried (MgSO₄) and the solvent

removed in vacuo to yield 38 as a red brick solid. Yield: 224 mg,

99%. ¹H NMR (360 MHz, CDCl₃) d ppm: 7.89 (1H, d, J = 8.2 Hz),

7.73 (1H, s), 7.43 (1H, dd, J = 8.4, 2.0 Hz), 7.34 (1H, d, J = 1.8 Hz),

2.88–2.97(2H, m), 2.79–2.85 (2H, m). LCMS: t_R= 1.91 min, 249

(M+H)⁺calcd for C₁₁H₉BrN₂.

4.1.7.4. 1-(4-Methoxy-benzyl)-3-[4-(4,4,5,5-tetramethyl-[1,3,2]

dioxaborolan-2-yl)-phenyl]-1H-pyrazole-4-carbonitrile (37).

Standard boronylation procedure (as described for compound 26)

using 3-(4-bromo-phenyl)-1-(4-methoxy-benzyl)-1H-pyrazole-4-

carbonitrile 36 (309 mg, 0.84 mmol) gave the desired compound

(237 mg, 68%). ¹H NMR (250 MHz, CDCl₃) d ppm: 7.99 (2H, d,

J = 8.8 Hz), 7.90 (2H, d, J = 8.8 Hz), 7.72 (1H, s), 7.28 (2H, d,

J = 8.7 Hz), 6.94 (2H, d, J = 8.7 Hz), 5.29 (2H, s), 3.83 (3H, s), 1.37

(12H, s). LCMS: t_R= 2.54 min, 416 (M+H)⁺calcd for C₂₄H₂₇BN₃O₃.

HRMS: (M)⁺calcd for C₂₄H₂₆BN₃O₃: 415.2182, found: 415.2175.

4.1.7.5. 7-Bromo-4,5-dihydro-1H-benzo[g]indazole (38). 6-Bro-

mo-3,4-dihydro-2H-naphthalen-1-one (204 mg, 0.91 mmol) was

dissolved in DMF (2 mL) and tris-dimethylamino methane (657 mg,

4.1.7.6. 7-Bromo-1-(4-methoxy-benzyl)-4,5-dihydro-1H-benzo-

[g]indazole (39). NaH (60% in mineral oil, 15.6 mg, 0.39 mmol)

was suspended in DMF (3.5 mL) and 7-bromo-4,5-dihydro-1H-

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6949

benzo[g]indazole 38 (90 mg, 0.36 mmol) was added in DMF (1 mL).

The suspension was stirred for 10 min and PMBCl (61 mg,

0.39 mmol) was added dropwise. The reaction was stirred at room

temperature for 1 h. Water (10 mL) was added and the aqueous

layer was extracted with EtOAc (3 ꢁ 10 mL). The organic layer

was washed with water (2 ꢁ 10 mL) and brine (10 mL), dried

(MgSO₄), ﬁltered, and the solvent removed under vacuum. The res-

idue was puriﬁed by chromatography using 5% EtOAc in heptane to

yield 39 as a yellow semi-solid. Yield: 62 mg, 47%. ¹H NMR

(360 MHz, CDCl₃) d ppm: 7.64 (1H, d, J = 8.2 Hz), 7.27–7.33 (2H,

m), 7.15 (2H, d, J = 8.6 Hz), 7.01 (1H, s), 6.81 (2H, d, J = 8.6 Hz),

5.17 (2H, s), 3.72 (3H, s), 2.77–2.84 (2H, m), 2.60–2.67 (2H, m).

LCMS: t_R= 2.50 min, 369 (M+H)⁺calcd for C₁₉H₁₇BrN₂O.

4.1.8. Synthesis of ﬁnal compounds 1a–1p

4.1.8.1. Dimethyl-[2-(4-{5-[4-(1H-pyrazol-3-yl)-phenyl]-4-pyri-

din-4-yl-1H-imidazol-2-yl}-phenoxy)-ethyl]-amine (1a). {2-[4-

(5-{4-[1-(4-Methoxy-benzyl)-1H-pyrazol-3-yl]-phenyl}-1-methoxy-

methyl-4-pyridin-4-yl-1H-imidazol-2-yl)-phenoxy]-ethyl}-dimeth-

yl-amine 8a (47 mg; 0.077 mmol) and anisole (17 lL; 0.153 mmol)

were dissolved in TFA (1.5 mL) and heated at 130 °C for 4 h then

cooled to room temperature overnight. The solvent was evapo-

rated, the residue washed with TBME and puriﬁed by preparative

LC to give the desired compound (11 mg; 32%). ¹H NMR

(360 MHz, MeOH) d ppm: 8.65 (2H, d, J = 5.4 Hz), 8.20 (2H, d,

J = 6.4 Hz), 8.12 (2H, d, J = 9.1 Hz), 8.05 (2H, d, J = 8.2 Hz), 7.80

(1H, s), 7.72 (2H, d, J = 8.2 Hz), 7.26 (2H, d, J = 9.1 Hz), 6.85 (1H,

s), 4.47–4.54 (2H, m), 3.67–3.73 (2H, m), 3.06 (6H, s). LCMS:

t_R= 2.36 min, 451 (M+H)⁺calcd for C₂₇H₂₆N₆O. HRMS: (M+H)⁺

calcd for C₂₇H₂₆N₆O: 451.2246, found: 451.2235.

4.1.7.7. 1-(4-Methoxy-benzyl)-7-(4,4,5,5-tetramethyl-[1,3,2]dio-

xaborolan-2-yl)-4,5-dihydro-1H-benzo[g]indazole (40). Stan-

dard boronylation procedure (as described for compound 26) using

intermediate 39 (62 mg, 0.17 mmol) resulted in the desired boro-

nic acid 40 (contaminated with 30% of the reduced material, 70%

corrected yield). This mixture was used in the next step without

further puriﬁcation. ¹H NMR (250 MHz, CDCl₃) d ppm: 7.82–7.90

(1H, m), 7.65–7.76 (2H, m), 7.24 (2H, d, J = 8.8 Hz), 7.08 (1H, s),

6.89 (2H, d, J = 8.7 Hz), 5.27 (2H, s), 3.81 (5H, s), 2.93 (2H, t,

J = 7.1 Hz), 2.72 (2H, t, J = 7.2 Hz), 1.36 (12H, s). LCMS:

t_R= 2.62 min, 417 (M+H)⁺calcd for C₂₅H₂₉BN₂O.

4.1.8.2. Dimethyl-[2-(4-{5-[4-(4-methyl-1H-pyrazol-3-yl)-phe-

nyl]-4-pyridin-4-yl-1H-imidazol-2-yl}-phenoxy)-ethyl]-amine

(1b). {2-[4-(5-{4-[1-(4-Methoxy-benzyl)-4-methyl-1H-pyrazol-3-

yl]-phenyl}-1-methoxy methyl-4-pyridin-4-yl-1H-imidazol-2-yl)-

phenoxy]-ethyl}-dimethyl-amine 8b (60 mg; 0.096 mmol) and

anisole (21 ll; 0.192 mmol) were dissolved in TFA (1.5 mL) and

heated at 130 °C for 4 h then cooled to room temperature over-

night. The solvent was evaporated, the residue washed with TBME

and puriﬁed by preparative LC to give the desired compound

(8 mg; 18%). ¹H NMR (250 MHz, acetone) d ppm: 11.80 (1H, br s),

8.56 (1H, d, J = 5.2 Hz), 8.46 (1H, d, J = 6.2 Hz), 8.08 (2H, d,

J = 8.8 Hz), 7.83 (1H, d, J = 8.4 Hz), 7.73 (1H, s), 7.63–7.66 (2H, m),

7.61 (1H, s), 7.56 (1H, s), 7.52 (1H, d, J = 5.9 Hz), 7.06 (2H, d,

J = 9.0 Hz), 4.16 (2H, t, J = 5.9 Hz), 2.70 (2H, t, J = 5.9 Hz), 2.30 (3H,

s), 2.28 (6H, s). LCMS: t_R= 2.45 min, 465 (M+H)⁺calcd for

4.1.7.8. 7-Bromo-1H-benzo[g]indazole (41). Compound 38

(170 mg, 0.68 mmol) was dissolved in dioxane (6 mL) and

DDQ (310 mg, 1.36 mmol) was added. The solution was re-

ﬂuxed for 3 h and the solvent was removed under vacuum.

2 M NaOH (10 mL) was added, and the aqueous layer was ex-

tracted with TBME (3 ꢁ 10 mL), dried (MgSO₄) and the solvent

removed under vacuum. The residue was puriﬁed by chroma-

tography using a stepped gradient of 20–40% EtOAc in heptane

to yield 41 as red brick solid. Yield: 81 mg, 48%. ¹H NMR

(250 MHz, CDCl₃) d ppm: 8.10–8.18 (2H, m), 8.07 (1H, d,

J = 8.5 Hz), 7.67–7.78 (2H, m), 7.44 (1H, d, J = 8.5 Hz). LCMS:

t_R= 1.96 min, 247 (M+H)⁺calcd for C₁₁H₇BrN₂.

C

₂₈H₂₉N₆O. HRMS: (M+H)⁺calcd for C₂₈H₂₉N₆O: 465.2403, found:

465.2400.

4.1.8.3. 3-(4-{2-[4-(2-Dimethylamino-ethoxy)-phenyl]-5-pyri-

din-4-yl-3H-imidazol-4-yl}-phenyl)-1H-pyrazole-4-carbonitrile

(1c). 3-(4-{2-[4-(2-Dimethylamino-ethoxy)-phenyl]-3-methoxy-

methyl-5-pyridin-4-yl-3H-imidazol-4-yl}-phenyl)-1-(4-methoxy-

benzyl)-1H-pyrazole-4-carbonitrile 8c (61 mg; 0.096 mmol) was

dissolved in TFA (1.2 mL) and heated at 70 °C for 4 h then cooled

to room temperature overnight. The solvent was evaporated, the

residue washed with TBME and puriﬁed by preparative LC to give

the desired compound (10 mg; 22%). ¹H NMR (250 MHz, acetone) d

ppm: 8.51 (2H, dd, J = 4.6, 1.5 Hz), 8.43 (1H, s), 8.07 (4H, dd), 7.75

(2H, d, J = 8.5 Hz), 7.61 (2H, dd, J = 4.6, 1.5 Hz), 7.06 (2H, d,

J = 9.0 Hz), 4.16 (2H, t, J = 5.9 Hz), 2.70 (2H, t, J = 5.9 Hz), 2.28 (6H,

s). LCMS: t_R= 2.42 min, 476 (M+H)⁺calcd for C₂₈H₂₅N₇O. HRMS:

(M+H)⁺calcd for C₂₈H₂₅N₇O: 476.2199, found: 476.2194.

4.1.7.9.

7-Bromo-1-(4-methoxy-benzyl)-1H-benzo[g]indazole

(42). NaH (60% in mineral oil, 8.6 mg, 0.36 mmol) was suspended

in DMF (2 mL) and 41 (81 mg, 0.33 mmol) was added in DMF

(1 mL). The suspension was stirred for 10 min and PMBCl (56 mg,

0.36 mmol) was added dropwise. The reaction was stirred at room

temperature for 1 h. Water (10 mL) was added and the aqueous

layer was extracted with TBME (3 ꢁ 10 mL). The organic layer

was washed with water (2 ꢁ 10 mL) and brine (10 mL), dried

(MgSO₄), ﬁltered, and the solvent removed under vacuum. The res-

idue was puriﬁed by chromatography using 10% EtOAc in heptane

to yield 42 as a yellow solid. Yield: 93 mg, 77%. ¹H NMR (250 MHz,

CDCl₃) d ppm: 8.47 (1H, d, J = 8.7 Hz), 7.96 (1H, d, J = 1.8 Hz), 7.81

(1H, s), 7.67 (1H, dd, J = 8.5, 2.0 Hz), 7.51 (1H, d, J = 9.0 Hz), 7.31

(1H, s), 6.91 (2H, d, J = 8.7 Hz), 5.57 (2H, s), 3.81 (3H, s). LCMS:

t_R= 2.47 min, 367 (M+H)⁺calcd for C₁₉H₁₅BrN₂O.

4.1.8.4. [2-(4-{5-[4-(4-Chloro-1H-pyrazol-3-yl)-phenyl]-4-pyri-

din-4-yl-1H-imidazol-2-yl}-phenoxy)-ethyl]-dimethyl-amine

(1d). {2-[4-(5-{4-[4-Chloro-1-(4-methoxy-benzyl)-1H-pyrazol-3-

yl]-phenyl}-4-pyridin-4-yl-1H-imidazol-2-yl)-phenoxy]-ethyl}-di-

methyl-amine 9d (18 mg; 0.03 mmol) was dissolved in TFA

(0.5 mL) and heated at 65 °C for 11 h then cooled to room temper-

ature overnight. The solvent was evaporated, the residue washed

with TBME to give the desired compound (19 mg; 76%). ¹H NMR

(360 MHz, MeOH/ CDCl₃) d ppm: 8.54 (2H, br s), 8.11 (2H, d,

J = 5.4 Hz), 8.04 (4 H, dd, J = 8.6, 2.7 Hz), 7.72 (1H, s), 7.66 (2H, d,

J = 8.2 Hz), 7.14 (2H, d, J = 9.1 Hz), 4.42 (2H, t), 3.61 (2H, t),

2.99 (6H, s). LCMS: t_R= 2.55 min, 485 (M+H)⁺calcd for C₂₇H₂₅Cl-

N₆O. HRMS: (M+H)⁺calcd for C₂₇H₂₅ClN₆O: 485.1856, found:

485.1873.

4.1.7.10. 1-(4-Methoxy-benzyl)-7-(4,4,5,5-tetramethyl-[1,3,2]di-

oxaborolan-2-yl)-1H-benzo[g]indazole (43). Standard boronyla-

tion procedure (as described for compound 26) using intermediate

42 (93 mg, 0.25 mmol) resulted in the desired boronic acid 43

(53 mg, 51%). ¹H NMR (250 MHz, CDCl₃) d ppm: 8.59 (1H, d,

J = 8.1 Hz), 8.31 (1H, s), 7.99 (1H, d, J = 6.9 Hz), 7.79 (1H, s), 7.36–

7.51 (2H, m), 7.31(2H, d), 6.91 (2H, d, J = 8.7 Hz), 5.59 (2H, s),

3.81 (3H, s), 1.41 (12H, s). LCMS: t_R= 2.59 min, 415 (M+H)⁺calcd

for C₂₅H₂₇BN₂O.

6950

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

4.1.8.5. (2-{4-[5-(4,5-Dihydro-2H-benzo[g]indazol-7-yl)-4-pyri-

din-4-yl-1H-imidazol-2-yl]-phenoxy}-ethyl)-dimethyl-amine (1e).

[2-(4-{4-[1-(4-Methoxy-benzyl)-4,5-dihydro-1H-benzo[g]indazol-

7-yl]-1-methoxy methyl-5-pyridin-4-yl-1H-imidazol-2-yl}-phe-

noxy)-ethyl]-dimethyl-amine 8e (13 mg; 0.02 mmol) was dis-

solved in TFA (1.5 mL), and anisole (4.4 mg; 0.04 mmol) was added.

The solution was heated in a sealed vessel for 24 h. The solvent was

removed under vacuum and the residue puriﬁed by preparative

HPLC to yield the desired compound 1e. TFA salt as a yellow glue

(1 mg, 6%). ¹H NMR (360 MHz, MeOD) d ppm: 8.58 (2H, d,

J = 6.4 Hz), 8.16 (2H, d, J = 6.8 Hz), 8.07 (2H, d, J = 9.1 Hz), 7.91

(1H, d, J = 8.2 Hz), 7.54 (2H, d, J = 5.0 Hz), 7.50 (1H, d, J =

7.3 Hz), 7.21 (2H, d, J = 9.1 Hz), 4.44–4.48 (2H, m), 3.63–3.68

(2H, m), 3.49–3.52 (1H, m), 3.10–3.13 (1H, m), 3.02 (6H, s),

2.83–2.89 (2H, m). LCMS: t_R= 2.50 min, 477 (M+H)⁺calcd for

4.1.8.10. 6-(2-Piperidin-4-yl-5-pyridin-4-yl-3H-imidazol-4-yl)-

2,4-dihydro-indeno[1,2-c]pyrazole (1k). Using the same proce-

dure as for 1j from 16c (33 mg; 0.069 mmol), the tert-butoxycar-

bonyl (Boc)-protected version of 1k was obtained. The residue

was dissolved in DCM (5 mL) and 4 N HCl (0.09 mL) added. After

stirring at rt for 4 h, the solvents were evaporated under vacuum

and the residue triturated with diethyl ether to leave the desired

product as the HCl salt of 1k (33 mg; 100%). ¹H NMR (250 MHz,

MeOD) d ppm: 8.83 (2H, br s), 8.20 (2H, br s), 8.07 (1H, s), 7.64–

8.02 (3H, m), 3.94 (2H, br s), 3.63 (4H, d, J = 14.3 Hz), 2.20–2.57

(5H, m). LCMS: t_R= 0.93 min, 383 (M+H)⁺calcd for C₂₃H₂₂N₆.

HRMS: (M+H)⁺calcd for C₂₃H₂₂N₆: 383.1984, found: 383.1077.

4.1.8.11. (2-{4-[5-(3,8-Dihydro-indeno[1,2-d] [1,2,3]triazol-6-

yl)-4-pyridin-4-yl-1H-imidazol-2-yl]-phenoxy}-ethyl)-dimethyl-

amine (1i). To a solution of ketone 16b (55 mg; 0.144 mmol) in

1 mL of methoxyethanol/HCl concd (3:1) at 0 °C was added tBuON-

C

₂₉H₂₉N₆O. HRMS: (M+H)⁺calcd for C₂₉H₂₉N₆O: 477.2403, found:

477.2400.

O (30

lL; 0.29 mmol). The reaction was stirred at 0 °C for 1 min

4.1.8.6. (2-{4-[4-(1H-Benzo[g]indazol-7-yl)-5-pyridin-4-yl-1H-imi-

dazol-2-yl]-phenoxy}-ethyl)-dimethyl-amine (1f). Using the same

procedure as for 1e, 1f was obtained from 8f (33 mg; crude). Yield:

5 mg (8% over two steps). ¹H NMR (250 MHz, MeOD) d ppm: 8.52–

8.62 (3H, m), 8.27 (1H, d, J = 1.37 Hz), 8.22 (1H, s), 8.07–8.16 (4H,

m), 7.79–7.89 (2H, m), 7.60 (1H, d, J = 8.98 Hz), 7.23 (2H, d,

J = 8.98 Hz), 4.47 (2H, t), 3.66 (2H, t), 3.02 (s, 6H). LCMS:

t_R= 2.33 min, 475 (M+H)⁺calcd for C₂₉H₂₇N₆O. HRMS: (M+H)⁺

calcd for C₂₉H₂₇N₆O: 475.2246, found: 475.2244.

then more tBuONO (30

l

L; 0.29 mmol) were added. The solution

was stirred for 30 min at 0 °C. Slowly 2 mL of NaOH (2 M) were

added. The solution was extracted with EtOAc (5 mL). The solvent

was removed under vacuo and the oily residue was dissolved in

8 mL of ethylene glycol/KOH (85%) (10:1). Hydrazine hydrate

(800 mg; 16 mmol) was added and the solution was heated at

190 °C for 2 h in a boiling tube. The reaction was allowed to cool

to room temperature, diluted with 5 mL of water and extracted

with DCM (10 mL). The aqueous layer was evaporated under vac-

uum. The residue was loaded on Ambersep (sulfonic acid) resin

(10 mL). The resin was washed with MeOH (3 ꢁ 10 mL), H₂O

(10 mL), HCl (1 M, 5 ꢁ 10 mL) then cleaved with HCl (5 M,

5 ꢁ 10 mL). The fractions were evaporated under vacuum. The res-

idue was suspended in 10 mL IPA/EtOH (1:1), ﬁltered. The solvent

was removed under vacuum, the residue was puriﬁed by reverse

phase column chromatography (C18) eluting with MeOH to afford

1.9 mg of 1i (3%). ¹H NMR (360 MHz, MeOD) d ppm: 8.56 (2H, br s),

8.03–8.10 (3H, m), 7.92 (1H, d, J = 7.7 Hz), 7.84 (1H, s), 7.65 (1H, d,

J = 7.7 Hz), 7.21 (2H, d, J = 9.1 Hz), 4.46 (2H, t, J = 10.0 Hz), 3.91 (2H,

s), 3.66 (2H, t, J = 10.0 Hz), 3.02 (6H, s) LCMS: t_R= 2.30 min, 464

(M+H)⁺calcd for C₂₇H₂₆N₇O. HRMS: (M+H)⁺calcd for C₂₇H₂₆N₇O:

464.2199, found: 464.2193.

4.1.8.7. 7-{2-[4-(2-Dimethylamino-ethoxy)-phenyl]-5-pyridin-

4-yl-1H-imidazol-4-yl}-2,4,4a,5-tetrahydro-indeno[1,2-c]pyri-

dazin-3-one (1g). Using the same procedure as for 1e, 1g (2.4 mg,

47%) was obtained from 8g. ¹H NMR (360 MHz, MeOD) d ppm: 8.57

(2H, br s), 8.05 (2H, d, J = 8.9 Hz), 8.01 (2H, br s), 7.85 (1H, d,

J = 7.9 Hz), 7.66 (1H, s), 7.56 (1H, d, J = 7.9 Hz), 7.20 (2H, d,

J = 8.9 Hz), 4.41–4.50 (2H, m), 3.61–3.69 (2H, m), 3.44–3.55 (1H,

m), 3.01 (6H, s), 2.79–2.94 (2H, m), 2.45 (1H, t, J = 16.3 Hz). LCMS:

t_R= 2.19 min, 493 (M+H)⁺calcd for C₂₉H₂₉N₆O₂. HRMS: (M+H)⁺

calcd for C₂₉H₂₉N₆O₂: 493.2352, found: 493.2347.

4.1.8.8. 6-{2-[4-(4-Methyl-piperazin-1-yl)-phenyl]-5-pyridin-

4-yl-3H-imidazol-4-yl}-2,4-dihydro-indeno[1,2-c]pyrazole (1j).

Ketone 16a (37.1 mg; 0.083 mmol) was dissolved in dry DMF

(0.5 mL) and tris(dimethylamine)methane (86 lL; 0.50 mmol)

was added, reaction was heated to 85 °C, under N₂for 3 h. Solvents

were removed under vacuum and residue was dissolved in AcOH

(0.5 mL). Hydrazine hydrate (12 lL; 0.25 mmol) was added to the

reaction mixture and was then stirred overnight. Solvent was re-

moved under vacuum and residue was puriﬁed by Flash column

chromatography on silica gel, eluting the desired product in 10–

50% EtOH/dichloromethane (DCM) (6 mg; 15%). ¹H NMR (250

4.1.8.12. [4-(2,4-Dihydro-indeno[1,2-c]pyrazol-6-yl)-5-pyridin-

4-yl-furan-2-yl]-[4-(2-dimethylamino-ethyl)-piperazin-1-yl]-

methanone (1l). Ketone 22b (86 mg; 0.19 mmol) was dissolved

in dry DMF (1 mL) and DMF–DMA (112 mg; 0.94 mmol) was added.

The reaction was heated to 85 °C for 3 h. Solvent was removed under

vacuum and remaining DMF was azeothroped with EtOH/heptane.

The residue was dissolved in AcOH (1 mL) and the hydrazinehydrate

(45 lL; 0.94 mmol) was added. Reaction was stirred at room tem-

perature overnight. The reaction mixture was basiﬁed to pH 11 with

NH₄OH (concd), extracted with EtOAc (4 ꢁ 6 mL), dried (MgSO₄) ﬁl-

tered and evaporated to dryness to leave a yellow solid. This solid

was dissolved in 1 M HCl (10 mL) and washed with EtOAc

(2 ꢁ 5 mL). Aqueous was basiﬁed with 2 M NaOH to pH 11 and ex-

tracted again with EtOAc, dried (MgSO₄), ﬁltered and evaporated

to dryness to leave a cream solid. Yield: 20 mg (22%). ¹H NMR

(360 MHz, MeOH) d ppm: 8.49 (2H, d, J = 7.5 Hz), 7.79 (1H, d,

J = 7.8 Hz), 7.55–7.66 (4H, m), 7.45 (1H, d, J = 7.8 Hz), 7.24 (1H, s),

3.94 (4H, br s), 3.70 (2H, s), 2.53–2.68 (8H, m), 2.32 (6H, s). LCMS:

t_R= 1.16 min, 483(M+H)⁺calcd for C₂₈H₃₀N₆O₂. HRMS:(M+H)⁺calcd

for C₂₈H₃₀N₆O₂: 483.2508, found: 483.2514.

MHz, MeOD)

d ppm: 8.35 (2H, d, J = 8.1 Hz), 7.88 (2H, d,

J = 8.8 Hz), 7.70 (1H, d, J = 7.9 Hz), 7.59 (2H, s), 7.51 (1H, s), 7.43

(1H, d, J = 7.6 Hz), 7.07 (2H, d, J = 9.0 Hz), 3.25–3.67 (10H, m),

2.87 (3H, s). LCMS: t_R= 1.11 min 474 (M+H)⁺calcd for C₂₉H₂₈N₇.

HRMS: (M+H)⁺calcd for C₂₉H₂₈N₇: 474.2406, found: 474.2399.

4.1.8.9. (2-{4-[5-(2,4-Dihydro-indeno[1,2-c]pyrazol-6-yl)-4-pyri-

din-4-yl-1H-imidazol-2-yl]-phenoxy}-ethyl)-dimethyl-amine

(1h). Using the same procedure as for 1j from 16b (38 mg;

0.087 mmol) the desired 1h was obtained (26 mg; 65%). ¹H NMR

(250 MHz, MeOD) d ppm: 8.32 (2H, br s), 7.84 (2H, d, J = 8.7 Hz),

7.33–7.71 (6H, m), 6.97 (2H, d, J = 8.8 Hz), 4.06 (2H, t, J = 5.4 Hz),

3.58 (2H, s), 2.70 (2H, br s), 2.26 (6H, s). LCMS: t_R= 1.10 min, 463

(M+H)⁺calcd for C₂₈H₂₇N₆O. HRMS: (M+H)⁺calcd for C₂₈H₂₇N₆O:

463.2246, found: 463.2242.

4.1.8.13. [4-(2,4-Dihydro-indeno[1,2-c]pyrazol-6-yl)-5-pyridin-

4-yl-furan-2-yl]-morpholin-4-yl-methanone (1m). Starting from

ketone 22c (96 mg; 0.25 mmol) and using the same procedure as

for compound 1l the desired compound was obtained (18 mg;

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

6951

11%). ¹H NMR (250 MHz, MeOD) d ppm: 8.48 (2H, br s), 7.79 (2H, d,

J = 7.8 Hz), 7.54–7.63 (4H, m), 7.44 (1H, dd, J = 7.9, 1.4 Hz), 7.26

(1H, s), 3.91 (4H, br s), 3.75–3.84 (4H, m), 3.71 (2H, s). LCMS:

t_R= 3.03 min, 413 (M+H)⁺calcd for C₂₄H₂₁N₄O_3.HRMS: (M+H)⁺

calcd for C₂₄H₂₁N₄O₃: 413.1614, found: 413.1611.

Partial charges of the ligand were derived using the Charge-2 CORI-

NA 3D package in TSAR 3.3, and its geometry optimized using the

COSMIC module of TSAR. The calculations were terminated if the en-

ergy difference or the energy gradientwere smallerthan 1E-005. Ten

docking solutions were generated, and the best three stored for

analysis.

4.1.8.14. 4-[4-(2,4-Dihydro-indeno[1,2-c]pyrazol-6-yl)-5-pyri-

din-4-yl-furan-2-carbonyl]-piperazine-1-carboxylic acid tert-

butyl ester (23d). Starting from ketone 22d (106 mg; 0.22 mmol)

and using the same procedure as for compound 1l the desired com-

pound was obtained (70 mg; 63%). ¹H NMR (250 MHz, CDCl₃-d) d

ppm: 8.53 (2H, br s), 7.81 (1H, d, J = 7.8 Hz), 7.45–7.57 (2H, m),

7.32–7.45 (3H, m), 7.18 (1H, s), 3.87 (4H, br s), 3.65–3.72 (2H,

m), 3.49–3.64 (4H, m), 1.48 (9H, s). LCMS: t_R= 3.62 min, (M+H)⁺

calcd for C₂₉H₂₉N₅O_4.

4.3. Biology

4.3.1. ^V600EBRAF kinase assay and SRB IC₅₀for BRAF inhibitors

These assays have been described by Niculescu-Duvaz et al.¹⁵

4.3.2. Phospho-ERK IC₅₀assay

To determine the effect of compounds on BRAF activity in

cells, WM266.4 cells were seeded at a density of 3 ꢁ 10⁴cells

per well of a 96 well plate. The following day, test compounds

were diluted into growth medium to 2ꢁ the desired ﬁnal con-

centration and then added directly to the cells. After a 6 h incu-

bation, the medium was removed and cells were ﬁxed and

permeabilized in 4% formaldehyde, 0.1% triton X-100 in PBS for

30 min. The wells were then blocked with 5% milk in PBS for

30 min at room temperature, followed by the addition of an anti-

body for phospho-ERK1/2 (Sigma, Dorset, UK) at 3 mg/ml in

blocking solution. Plates were incubated for 3 h with shaking.

Plates were washed three times with 0.1% Tween 20 using an

ELx50 plate washer (BioTek, Winooski, USA). 0.5 mg/ml of a

Europium-labelled anti-mouse secondary antibody (Perkin Elmer,

Turku, Finland) was added to the wells in DELFIA assay buffer for

1 h. Plates were washed again and Enhancement solution was

added to the wells and time resolved ﬂuorescence was measured

as instructed by the manufacturer after 20 min using a Spectra-

max M5 plate reader (Molecular Devices, Berkshire, UK). The

plates were washed again and BCA protein assay reagent (Sigma,

Dorset, UK) was added to the wells and incubated for 30 min at

37 °C. Absorbance at 570 nm was measured using a plate reader

and used to normalize the ﬂuorescence data. Inhibition of ERK

phosphorylation was determined as a percentage of DMSO-trea-

ted cells and IC₅₀values were calculated using Prism (GraphPad

Software, San Diego, USA).

4.1.8.15. [4-(2,4-Dihydro-indeno[1,2-c]pyrazol-6-yl)-5-pyridin-

4-yl-furan-2-yl]-piperazin-1-yl-methanone (1n). BOC amide

23d was dissolved in DCM (0.5 mL) and dioxane (0.3 ml) and 4 N

HCl in dioxane (0.3 mL) was added. After stirring at room temper-

ature for 2 h, the solvents were evaporated under vacuum to leave

the desired 1n (30 mg; 100%). ¹H NMR (250 MHz, MeOD) d ppm:

8.74 (2H, d, J = 6.1 Hz), 8.16 (2H, d, J = 6.3 Hz), 8.08 (1H, s), 7.96

(1H, d, J = 7.8 Hz), 7.87 (1H, s), 7.68 (1H, d, J = 7.9 Hz), 7.47 (1H,

s), 4.00–4.37 (4H, m), 3.86–3.98 (2H, m), 3.36–3.52 (4H, m). LCMS:

t_R= 2.45 min, 412 (M+H)⁺calcd for C₂₄H₂₂N₅O₂. HRMS: (M+H)⁺

calcd for C₂₄H₂₂N₅O₂: 412.1774, found: 412.1769.

4.1.8.16. [4-(2,4-Dihydro-indeno[1,2-c]pyrazol-6-yl)-5-pyridin-

4-yl-furan-2-yl]-(4-methyl-piperazin-1-yl)-methanone (1o). Start-

ing from ketone 22a (79 mg; 0.20 mmol) and using the same pro-

cedure the desired compound was obtained (50 mg; 60%). ¹H NMR

(250 MHz, MeOD) d ppm: 8.47 (2H, br s), 7.77 (1H, d, J = 7.8 Hz),

7.48–7.67 (4H, m), 7.43 (1H, d, J = 7.6 Hz), 7.22 (1H, s), 3.73–4.11

(4H, m), 3.68 (2H, s), 2.45–2.70 (4H, m), 2.36 (3H, s). LCMS:

t_R= 2.41 min, 426 (M+H)⁺calcd for C₂₅H₂₃N₅O₂. HRMS: (M+H)⁺

calcd for C₂₅H₂₃N₅O₂: 426.1930, found: 426.1934.

4.1.8.17. [4-(2-Dimethylamino-ethyl)-piperazin-1-yl]-[4-(1-met

hyl-1,4-dihydro-indeno [1,2-c]pyrazol-6-yl)-5-pyridin-4-yl-fur

an-2-yl]-methanone (1p). The starting ketone 22b (34 mg,

0.07 mmol) was dissolved in dry DMF (0.5 mL) and DMFꢃDMA

(44 mg; 0.37 mmol) was added. The reaction was stirred for

90 min. The solvents were removed under vacuum and the residue

azeotroped with heptane. The crude was then dissolved in AcOH

(0.5 mL) and treated with methylhydrazine (51 mg; 1.11 mmol).

The reaction was stirred at room temperature for two days, upon

which concentrated NH₄OH solution was added to pH 11. NaHCO₃

saturated solution (10 mL) was added and the aqueous layer was

extracted with EtOAc (3 ꢁ 10 mL), dried (MgSO₄) and the solvent

removed under vacuum. The residue was puriﬁed by chromatogra-

phy using a stepped gradient of 0–15% MeOH in DCM. The product

was re-suspended in EtOAc (2 mL) and washed with water

(5 ꢁ 1 mL). The organic layer was dried (MgSO₄) and the solvent

removed in vacuo to yield a pale yellow glue (3 mg; 8%). ¹H NMR

(250 MHz, MeOD) d ppm: 8.46–8.52 (2H, m), 7.80 (1H, d), 7.64

(1H, s), 7.54–7.60 (2H, m), 7.45–7.52 (1H, m), 7.43 (1H, s), 7.25

(1H, s), 4.15 (3H, s), 3.91 (4H, br s), 3.64 (2H, s), 2.58–2.67 (8H,

m), 2.34 (6H, s). LCMS: t_R= 2.73 min, 497 (M+H)⁺calcd for

C₂₉H₃₃N₆O₂. HRMS: (M+H)⁺calcd for C₂₉H₃₃N₆O₂: 497.2665,

found: 497.2660.

4.3.3. PK assessment

Female Crl:CD1-Foxn1nu mice at least six weeks of age bearing

mutant BRAF WM266.4 human tumor xenografts were used for the

PK analyses. The mice were dosed intraperitoneally (4 mg/kg,

equivalent to ca.

8 lmol/kg, 10 ml/kg, in 10% ethanol:40%

PEG:50% Gelofusine v/v) in a cassette with 4 other compounds.

Samples were taken at 8 time-points between 15 min and 24 h.

Three mice were utilized per time-point. They were placed under

isoﬂurane anaesthesia and blood for plasma preparation was taken

by terminal cardiac puncture into heparinized syringes. Plasma

samples were snap frozen in liquid nitrogen and then stored at

ꢂ70 °C prior to analysis. All procedures involving animals were

performed in accordance with national Home Ofﬁce regulations

under the Animals (Scientiﬁc Procedures) Act 1986 and within

guidelines set out by the Institute’s Animal Ethics Committee and

the United Kingdom Coordinating Committee for Cancer Research’s

ad hoc Committee on the Welfare of Animals in Experimental

Neoplasia.

5. Disclosure statement

This work was carried out as part of a research collaboration be-

tween the Institute of Cancer Research, The Wellcome Trust, GSK

and Cancer Research UK. Please note that all authors who are, or

have been, employed by The Institute of Cancer Research are sub-

4.2. Docking and modelling

Inhibitor 1a was docked using GOLD version 3.1.1[5] on the

crystal structure of BRAF in complex with SB590885 [PDB 2FB8].

6952

D. Niculescu-Duvaz et al. / Bioorg. Med. Chem. 18 (2010) 6934–6952

3. Gray-Schopfer, V. C.; da Rocha Dias, S.; Marais, R. Cancer Metastasis Rev. 2005,

24, 165.

4. Sharma, A.; Trivedi, N. R.; Zimmerman, M. A.; Tuveson, D. A.; Smith, C. D.;

Robertson, G. P. Cancer Res. 2005, 65, 1223.

ject to a ‘Rewards to Inventors Scheme’ that may reward contribu-

tors to a programme that is subsequently licensed.

Andrew K. Takle and David M. Wilson were employees of

GlaxoSmithKline at the time of the aforementioned research

collaboration.

5. www.clinicaltrials.gov.

6. Niculescu-Duvaz, D.; Gaulon, C.; Dijkstra, H. P.; Niculescu-Duvaz, I.; Zambon,

A.; Menard, D.; Suijkerbuijk, B. M. J. M.; Nourry, A.; Davies, L.; Manne, H.;

Friedlos, F.; Ogilvie, L.; Hedley, D.; Whittaker, S.; Kirk, R.; Gill, A.; Taylor, R. D.;

Raynaud, F. I.; Moreno-Farre, J.; Marais, R.; Springer, C. J. J. Med. Chem. 2009, 52,

2255.

Acknowledgments

7. Menard, D.; Niculescu-Duvaz, I.; Dijkstra, H. P.; Niculescu-Duvaz, D.;

Suijkerbuijk, B. M. J. M.; Zambon, A.; Nourry, A.; Roman, E.; Davies, L.;

Manne, H. A.; Friedlos, F.; Kirk, R.; Whittaker, S.; Gill, A.; Taylor, R. D.; Marais,

R.; Springer, C. J. J. Med. Chem. 2009, 52, 3881.

8. Young, P. R.; McLaughlin, M. M.; Kumar, S.; Kassis, S.; Doyle, M. L.; McNulty, D.;

Gallagher, T. F.; Fisher, S.; McDonnell, P. C.; Carr, S. A.; Huddleston, M. J.; Seibel,

G.; Porter, T. G.; Livi, G. P.; Adams, J. L.; Lee, J. C. J. Biol. Chem. 1997, 272, 12116.

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R.; Rusnak, D. W.; Takle, A. K.; Wilson, D. M.; Hugger, E.; Wang, L.; Karreth, F.;

Lougheed, J. C.; Lee, J.; Chau, D.; Stout, T. J.; May, E. W.; Rominger, C. M.;

Schaber, M. D.; Luo, L.; Lakdawala, A. S.; Adams, J. L.; Contractor, R. G.; Smalley,

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This work is supported by the Wellcome Trust (ref: 080333/Z/

06/Z), Cancer Research UK (refs: C309/A8274 and C107/A10433),

the Isle of Man Anti-Cancer Association and The Institute of Cancer

Research. We acknowledge NHS funding to the NIHR Biomedical

Research Centre. We thank Professors Paul Workman and Julian

Blagg for their advice and support. We thank Dr. Raffaella Di Lucre-

zia (Evotec) for her contribution to the synthesis.

Supplementary data

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W.; King, F. D.; Lovell, P. J.; Naylor, A.; Reith, A. D.; Steadman, J. G.; Wilson, D.

M. Bioorg. Med. Chem. Lett. 2006, 16, 378.

Supplementary data associated with this article can be found, in

the online version, at doi:10.1016/j.bmc.2010.06.031.

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MOL ﬁle

The following ZIP ﬁle contains the MOL ﬁle of the most impor-

tant compounds referred to in this article.

14. Takle, A. K.; Bamford, M. J.; Davies, S.; Davis, R. P.; Dean, D. K.; Gaiba, A.; Irving,

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References and notes

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Article Doi

Novel tricyclic pyrazole BRAF inhibitors with imidazole or furan central scaffolds

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