Fused imidazoles as potential chemical scaffolds for inhibition of heat shock protein 70 and induction of apoptosis. Synthesis and biological evaluation of phenanthro[9,10-d] imidazoles and imidazo[4,5-f] [1,10]phenanthrolines

Article abstract of DOI:10.1039/c6ob00471g

The imidazole ring is widespread in biologically active compounds, and hence imidazole-containing scaffolds are useful starting points for drug discovery programmes. We report the synthesis of a series of novel imidazole-containing compounds fused with either phenanthrene or phenanthroline, which show enhanced growth inhibitory potency against human colon, breast and melanoma cancer cell lines, as well as evidence of inhibition of the molecular chaperone heat shock protein 70 (Hsp70) pathway in cells, as shown by depletion of downstream oncogenic client proteins of the Hsp90 chaperone pathway, and induction of apoptosis.

Full text of DOI:10.1039/c6ob00471g

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PAPER
Fused imidazoles as potential chemical scaffolds
for inhibition of heat shock protein 70 and
induction of apoptosis. Synthesis and biological
evaluation of phenanthro[9,10-d]imidazoles and
imidazo[4,5-f][1,10]phenanthrolines†
Cite this: DOI: 10.1039/c6ob00471g
Alpa Patel,a Swee Y. Sharp,b Katelan Hall,a William Lewis,a Malcolm F. G. Stevens,c
Paul Workmanb and Christopher J. Moody*a
The imidazole ring is widespread in biologically active compounds, and hence imidazole-containing
scaffolds are useful starting points for drug discovery programmes. We report the synthesis of a series of
novel imidazole-containing compounds fused with either phenanthrene or phenanthroline, which show
enhanced growth inhibitory potency against human colon, breast and melanoma cancer cell lines, as well
as evidence of inhibition of the molecular chaperone heat shock protein 70 (Hsp70) pathway in cells, as
shown by depletion of downstream oncogenic client proteins of the Hsp90 chaperone pathway, and
induction of apoptosis.
Received 1st March 2016,
Accepted 22nd March 2016
DOI: 10.1039/c6ob00471g
www.rsc.org/obc
Introduction
The imidazole ring is widely represented amongst biologically
active compounds, and is a component of the amino acid histi-
dine, the nucleic acid base adenine, a range of natural pro-
ducts and secondary metabolites,1 and a plethora of synthetic
pharmaceutical agents, many of which have had major
benefits for human health.2,3 Thus cimetidine 1 was an early
histamine H-2-receptor antagonist that inhibits gastric acid
secretion, losartan 2 was the first angiotensin II receptor antag-
onist to be marketed and is used to treat hypertension, whilst
fused imidazoles such as the proton pump inhibitor benzimi-
dazole omeprazole 3 and the imidazotetrazine temozolomide
4 are used to block stomach acid secretion and as a first line
treatment for glioblastoma mulltiforme respectively (Fig. 1).
More recent additions to the repertoire of bioactive imida-
zoles are the triarylimidazole p38α MAP-kinase inhibitor
SB203580 5, and the related imidazole apoptozole 6 (Fig. 1).
aSchool of Chemistry, University of Nottingham, University Park, Nottingham,
NG7 2RD, UK. E-mail: c.j.moody@nottingham.ac.uk
bCancer Research UK Cancer Therapeutics Unit, Division of Cancer Therapeutics,
The Institute of Cancer Research, Haddow Laboratories, London, SM2 5NG, UK
cSchool of Pharmacy, University of Nottingham, University Park, Nottingham,
NG7 2RD, UK
†Electronic supplementary information (ESI) available: Copies of NMR spectra.
CCDC 1456778–1456780. For ESI and crystallographic data in CIF or other elec-
tronic format see DOI: 10.1039/c6ob00471g
Fig. 1 Some biologically active imidazoles.
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In view of our interest in the heat shock protein (Hsp) mole- triarylimidazoles, starting from phenanthrene-9,10-dione to
cular chaperones,4,5 we became interested in the imidazole yield the phenanthro[9,10-d]imidazole scaffolds 7 with variable
scaffold exhibited by apoptozole, and its reported biological substitution of the C-2 aryl ring (Scheme 1).
activity. The compound 6 induced apoptosis in murine P19
Functionalization of the phenanthrene ring was achieved
embryonic carcinoma cells and was reported to bind to the by following MacLachlan and co-workers’ procedure,25 via
heat shock proteins Hsp72 and Hsc70 as determined by initial bromination of phenanthrene-9,10-dione under radical
surface plasmon resonance (SPR) using an apoptozole–biotin conditions to give 3,6-dibromophenanthrene-9,10-dione 8.
conjugate.6 Based on molecular modeling, Shin and co- Reduction of the ortho-quinone with sodium dithionite
workers suggested that apoptozole binds to the ATP-ase allowed Ullmann coupling of 3,6-dibromo-9,10-dimethoxy-
domain of Hsc70.7 More recently, the ability of apoptozole to phenanthrene 9 to produce tetramethoxyphenanthrene 10
induce apoptosis and to bind to Hsp70 was confirmed.8,9 (Scheme 1). Oxidation with ammonium cerium(IV) nitrate
However, a very recent study was unable to find any experi- afforded 3,6-dimethoxyphenanthrene-9,10-dione 11, an appro-
mental evidence that apoptozole binds to Hsp70 family priate precursor to imidazole synthesis using the multicompo-
members directly,10 suggesting an indirect effect in cells on nent conditions described above. Dione 11 was utilized in the
the Hsp70/Hsp90 chaperone pathway.
synthesis of the corresponding imidazoles with benzaldehyde
Heat shock proteins such as Hsp70 family members facili- to give 12 and 3,5-bis(trifluoromethyl)benzaldehyde for com-
tate the folding, activity, stability and cellular translocation of pound 13 (Scheme 1). For comparison and structure–activity
newly synthesized polypeptides, as well as preventing the relationship elaboration, the non-planar analogue of 7c, 2-(4-
aggregation of unfolded mature proteins. The frequent over- methoxyphenyl)-4,5-phenyl-1H-imidazole 14, was synthesized
expression of Hsps in many cancers supports the increasing from benzil in 61% yield. To investigate the influence of sub-
interest in Hsps as targets for new molecular cancer thera- stitution on the left hand side of the molecule, 2,4,5-tris-(4-tri-
peutics,11 and precedence has been established by the clinical methoxyphenyl)-1H-imidazole 15 was formed from 4,4′-
development and activity of inhibitors of the Hsp90 molecular
chaperone.12,13 There are at least eight Hsp70 family
members, of which the two main cytoplasmic isoforms are the
ubiquitously expressed Hsc70, and Hsp72, which is over-
expressed as a response to stress factors.14 Many human cancers
and tumour cell lines show increased levels of Hsp70 family
members, prompting the suggestion that this may contribute
to the ability of these cells to evade apoptosis.11 As a result,
inhibition of Hsp70 has emerged in recent years as a promis-
ing strategy to target cancer, both as an individual approach
and in conjunction with other heat shock proteins such as
Hsp90.15–21
Here, we report the synthesis and evaluation for anticancer
activity of a series of compounds based on a triarylimidazole
core, analogous to apoptozole, but with the incorporation of a
phenanthrene or phenanthroline ring system. We have shown
that these compounds potently reduce the viability of human
cancer cell lines, and deplete Hsp72 and Hsp27 as well as
Hsp90 client proteins. In addition, the novel imidazo[4,5-f]
[1,10]phenanthrolines have emerged as a highly promising
chemical scaffold for the induction of apoptosis.
Results and discussion
Phenanthro[9,10-d]imidazoles
Debus and Radziszewski independently reported the multi-
component synthesis of imidazoles in acidic conditions in the
19th century, using a diketone, ammonium acetate, aniline
and an aldehyde.22,23 In 2004, Wolkenberg and co-workers
Scheme 1 Synthesis of phenanthro[9,10-d]imidazoles. Reagents and
conditions: (i) ArCHO, AcOH, NH4OAc, μW, 150 °C, 15 min. (ii) bromine,
benzoyl peroxide, nitrobenzene, 110 °C, 16 h; (iii) Na2S2O4, Me2SO4,
Bu4NBr, THF, water, rt, 45 min; (iv) NaOMe (25%), CuBr, EtOAc, toluene,
applied microwave irradiation to synthesize polysubstituted
imidazoles using the same starting components, omitting an
aniline so that the 2,4,5-trisubstituted imidazole was formed.24
Therefore, these conditions were utilized to make several 2,4,5- 80 °C, 64 h; (v) Ce(NH4)2(NO3)6, MeCN, water, rt, 15 min.
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Scheme 3 Synthesis of imidazo[4,5-f][1,10]phenanthrolines. Reagents
t
and conditions: (i) MeI or 17, BuOK or K2CO3, DMF or MeCN, rt, 16 h;
(ii) 3,5-(CF3)2C6H3CHO, BnNH2, AcOH, NH4OAc, μW, 150 °C, 30 min.
Scheme 2 Alkylation of phenanthro[9,10-d]imidazoles. Reagents and
conditions: (i) BuOK, MeI or BnBr, THF, rt, 16 h to 3 days; (ii) BuOK, 17,
t
t
THF, rt, 16 h to 3 days.
imidazole synthesis through the multicomponent pathway in
acidic media (Scheme 3).
dimethoxybenzil, and 2,4,5-tris-(4-trifluorophenyl)-1H-imida-
zole 16 from 4,4′-difluorobenzil.
As with the previous series, alkylation of the imidazole
nitrogen was attempted. This proved particularly difficult with
compound 28, possibly due to its poor solubility and high
polar character, although methylation was possible to give 30.
Benzyl derivative 31 was synthesized through a multicompo-
nent imidazole formation with benzylamine. Alkylation of 29
with 4-(bromomethylbenzamide) 17 to give 32 was successful
as the bis(trifluoromethyl)phenyl ring provided improved solu-
bility in organic solvents (Scheme 3).
The structures of apoptozole 6, imidazole 14 and phena-
throimidazole 18 were confirmed by X-ray crystallography
(Fig. 2) that established that the aromatic rings are not co-
planar in the non-constrained imidazoles 6 and 14.
Next, we turned our attention to increasing the molecular
size in a bid to impart selectivity towards the target protein.
Methylation of 1H-imidazoles was achieved by reaction with
iodomethane with potassium tert-butoxide in THF. Analogous
benzylation was successful with benzyl bromide as the alkylat-
ing agent under the same reaction conditions. Alternatively,
compound 19 could also be made through a multicomponent
reaction with benzylamine. The final alkylation was proposed
due to the molecular modeling study cited by Shin et al.,
which indicated that the benzamide oxygen of apoptozole
could participate in a hydrogen bonding interaction with the
Glu376 residue of the ATP binding pocket of Hsc70.7 The com-
mercially available 4-(bromomethyl)benzoic acid could be
readily converted into 4-(bromomethyl)benzamide 17 to allow
alkylation of the imidazole for the incorporation of a methyl-
benzamide moiety at N-1, analogous to apoptozole (Scheme 2).
Apoptozole itself (6) was synthesized as a reference compound,
by multicomponent synthesis of the 2,4,5-substituted imida-
zole followed by alkylation with 17.
Biological evaluation
The imidazole library was evaluated for in vitro anticancer
activity in the human colon cancer cell line HCT116 using the
CellTiter-Blue® viability assay. In our hands, apoptozole 6 gave
a GI50 of 7.0 μM in this assay. All of the fused phenanthro[9,10-
d]-imidazoles tested showed potency equivalent to apoptozole,
ranging from 3.5 to 11.2 μM (Table 1), with compound 7c
being twice as potent as apoptozole. Interestingly, non-planar
compound 14 was found to be less active in the cell viability
Imidazo[4,5-f][1,10]phenanthrolines
Following the promising results in the phenanthro[9,10-d]imi- assay, whereas 15 and 16 both showed potency that was com-
dazole series (see below), we turned our attention to introdu- parable to apoptozole and the phenanthro[9,10-d]imidazoles.
cing heteroatoms within the tetracyclic ring system. 1,10- Selected compounds were also evaluated in the breast cancer
Phenanthroline is a readily available chemical that is com- cell line MCF7 with compound 7c showing similar activity in
monly used as a chelating ligand for metal-catalyzed reactions. MCF7 cells when compared with HCT116 (Table 1).
The ring system can be oxidized to phenanthroline-1,10-dione
Our previous studies showed that siRNA silencing of Hsp70
27, which in our case, provided a useful starting material for isoforms Hsp72 and Hsc70 resulted in depletion of Hsp90
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Imidazoles 7a, 7c and 16 showed varying degrees of
depletion of Hsp72 and Hsc70 as well as Hsp27 over the period
48 hours and at concentrations of 5 and 10 × GI50 (Fig. 3). The
greatest effects were seen for Hsp27 whereas no effects were
seen on Hsc70. Hsp72 showed evidence of depletion by all com-
pounds at 48 hours, particularly the most potent compound 7c.
Additionally, compounds 7a, 7c and 16 depleted oncogenic
client proteins of Hsp90, namely, ERBB2, CRAF and CDK4. In
addition, cleaved PARP was observed, indicative of apoptosis.
The inactive 4,5-diphenyl-1H-imidazole 14, which has a GI50 of
>20 μM, did not show any detectable effects on Hsps, Hsp90 or
cleaved PARP at the highest concentration tested.
The Hsp90 inhibitor 17-N-allylamino-17-demethoxygeldana-
mycin (17-AAG)27 was included as
a control compound
(Fig. 3A). As expected, 17-AAG caused induction of the heat
shock pathway as shown by elevated expression of Hsp72 and
Hsp27, whereas this was not seen with the imidazoles 7a, 7c
and 16. Both 17-AAG and compounds 7a, 7c and 16 caused
depletion of Hsp90 client proteins, indicating that they blocked
the function of the Hsp90 chaperone complex, consistent with
our published results with siRNA silencing of Hsp72 and
Hsc70.26 The precise effects of the active imidazole compounds
and 17-AAG varied between the Hsp90 client proteins studied.
Alkylation of the imidazole nitrogen gave compounds with
differing activities in the cell viability assay in HCT116 colon,
MCF7 breast and WM266.4 melanoma cells in the cell viability
assay (Table 2). Methylation or benzylation results in compar-
able activity to the non-alkylated imidazole. However, introduc-
tion of the methylbenzamide group resulted in loss of activity
for compounds 21 and 24. Interestingly, analogues 22 and 23
retained potency in comparison to their non-alkylated precur-
sors 7b and 7c, suggesting that functionalization of the C-2 aryl
ring with a para substituent that can participate in hydrogen
bonding may be important. Dimethoxyphenanthrene analogue
25 was found to be the most active benzamide derivative, with
a GI50 of 3.40 μM in HCT116 cells. Unfortunately, the direct
analogue of apoptozole 26, where the two methoxyphenyl rings
at the 4 and 5 positions of the imidazole are constrained into a
phenanthrene system, was insoluble in our assay conditions.
Interestingly, imidazo[4,5-f][1,10]phenanthroline 28 showed
almost an eighty-fold increase in potency when compared to
apoptozole, with a GI50 of 0.09 μM in HCT116 cells and submi-
cromolar values in WM266.4 and MCF7 cells (Table 2). Com-
pound 28 also showed the desired effects on client protein
depletion by Western blot (Fig. 4A). In addition, Hsp72
and Hsp27 protein levels were decreased over 48 hours, and
there was clear evidence of apoptosis, as shown by PARP
cleavage in HCT116 cells. Interestingly, analogues of the
imidazo[4,5-f][1,10]phenanthroline 28 that are substituted at
the C-2 aryl ring have recently been reported as DNA G-quadru-
Fig. 2 X-Ray crystal structures of (A) apoptozole 6 (CCDC 1456778),
(B) imidazole 14 (CCDC 1456780) and (C) phenathroimidazole 18
(CCDC 1456779).
client proteins and induction of apoptosis.26 Compounds were plex binders,28 although in the case of compound 28 itself we
therefore investigated for their effect on expression of the com- were unable to find evidence for such an interaction. Thus incu-
monly studied Hsp90 client proteins, together with effects on bation of 28 with the 22 base human telomeric sequence AGGG
the levels of Hsp72, Hsc70 and Hsp27, and also on cleaved (TTAGGG)3 showed no change in ellipticity in its CD spectrum.
PARP as a marker of apoptosis in HCT116 human colon carci-
noma cells, as used previously.26
Despite the evidence of desired on-pathway biomarker
changes, compound 28 was considered to have potential to be
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Table 1 Cell growth viability assay results of 4,5-disubstituted imidazoles and phenanthro[9,10-d]imidazoles series against HCT116 (colon) and
MCF7 (breast) human cancer cells, showing GI50 (μM) with standard error (SE)
HCT116
GI50/μM
7.0
MCF7
Compound
Structure
SE/ μM
GI50/μM
SE/ μM
6 (apoptozole)
0.5
7a
7b
7c
7d
12
7.6
6.7
3.0
7.3
7.8
0.3
0.4
0.3
1.2
0.3
>5
4.0
0.5
13
11.2
1.5
14
15
>20
4.7
0.6
7.8
16
9.5
0.6
>10
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Conclusion
We have identified a series of phenanthro[9,10-d]imidazoles
that show in vitro potency against HCT116 human cancer cell
lines that is equivalent to or greater than apoptozole, a
reported inhibitor of Hsp70 and inducer of apoptosis in
cancer cells. In addition, imidazo[4,5-f][1,10]phenanthrolines,
readily obtained in a single step from phenanthroline-1,10-
dione, have emerged as a highly promising chemical scaffold
for apoptosis induction, with at least a ten-fold increase in
potency when compared to apoptozole in a cell viability assay.
Both series of compounds have also demonstrated cellular bio-
marker activity consistent with inhibition of Hsp70 family
members Hsc70 and Hsp72, including depletion of Hsp90
client proteins together with induction of apoptosis,15
although we have no direct evidence of binding to Hsp70.
Nevertheless, irrespective of the precise molecular mechanism
of action, the novel imidazo[4,5-f][1,10]phenanthrolines reported
herein appear to have potential for further investigation as
therapeutic agents that induce apoptosis in cancer cells.
Experimental section
Chemistry
General information. Commercially available reagents were
used without further purification unless otherwise stated,
including anhydrous N,N-dimethylformamide, methanol and
ethyl acetate. Dichloromethane and tetrahydrofuran were
freshly distilled from calcium hydride and sodium-benzophe-
none ketyl, respectively, according to standard procedures.
Toluene was purged with nitrogen over activated alumina
towers. Water refers to deionized water, and ether refers to
diethyl ether. Light petroleum refers to the fraction with
boiling point range 40–60 °C.
Reactions were routinely carried out under a nitrogen or
argon atmosphere. Microwave reactions were conducted using
a focused (300 W) microwave reactor with an IR temperature
sensor. Analytical thin layer chromatography was carried out
on aluminum-backed plates coated with silica gel 60 F254 and
visualized under UV light at 254 nm and/or 360 nm. Flash
column chromatography was carried out using Aldrich 60 Å
silica gel with the eluent specified.
Melting points were measured using a digital melting point
apparatus, and are uncorrected. High and low resolution mass
spectra were recorded on a time-of-flight instrument using
Electrospray Ionization (ESI). Infrared spectra were recorded
over the range 4000–600 cm−1 using an FT-IR spectrometer,
either as a solution in chloroform or as a solid in Attenuated
Total Reflectance (ATR) mode. NMR spectra were recorded at
400 MHz for 1H NMR and corresponding 100 MHz for 13C
NMR. Chemical shifts are quoted in parts per million (ppm),
and are referenced to residual protonated solvent as an
internal standard. Multiplicity abbreviations used: s singlet d
doublet, t triple, q quartet, m multiplet. Coupling constants J
are quoted in Hertz (Hz). Assignments were made with the use
Fig. 3 Effects of imidazole compounds on expression of heat shock
proteins, Hsp90 client proteins and cleaved PARP in HCT116 human
colon cancer cells. Western blots of lysates of HCT116 human colon
cancer cells treated with (A) 5 × GI50 concentrations of compounds 7a,
7c, 16 (non-planar compound) and the inactive compound 14 or the
Hsp90 inhibitor 17-AAG or vehicle control for 8, 24 and 48 h or (B) mul-
tiples of GI50 of compounds for 24 h. GAPDH was used as loading
control.
a highly non-selective small molecule inhibitor due to its low
molecular weight, and it was thought that N-alkylation of this
series of compounds might be a step towards gaining selecti-
vity and reducing off-target effects when moving forward to
in vivo studies. Pleasingly, although methylation of the imidazole
nitrogen resulted in drop in potency (compound 30 vs. 28), the
N-methyl compound 30 is still more active than apoptozole.
Unfortunately, 3,5-bis(trifluoromethyl)phenyl derivative 29 was
too insoluble to test in our assay conditions, but the corres-
ponding alkylated derivatives 31 and 32 showed at least a
tenfold increase in activity in comparison to apoptozole in
HCT116 cells (Table 2 and Fig. 4B). Structurally, compound 32
shares the N-1 and C-2 imidazole substituents with apopto-
zole, but differs in that the fused phenanthroline at C-4/C-5
results in a more planar structure.
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Table 2 Cell growth viability assay results of alkylated phenanthro[9,10-d]imidazoles and of imidazo[4,5-f][1,10]phenanthrolines against HCT116
(colon), WM266.4 (melanoma) and MCF7 (breast) human cancer cells, showing GI50 (μM) with standard error (SE)
HCT116
GI50/μM
9.4
WM266.4
MCF7
Compound
Structure
SE/ μM
GI50/μM
SE/ μM
GI50/μM
SE/ μM
18
0.5
19
20
21
6.8
1.4
>20
>20
22
23
24
25
5.9
5.5
>20
3.4
1.1
1.0
>5
>5
>10
>10
1.0
>5
>10
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Table 2 (Contd.)
HCT116
GI50/μM
Insoluble
WM266.4
MCF7
Compound
Structure
SE/ μM
GI50/μM
SE/ μM
GI50/μM
SE/ μM
26
28
29
30
31
0.09
0.01
0.32
0.10
0.69
0.11
Insoluble
2.6
0.04
0.2
0.5
2.9
2.5
0.3
2.8
2.3
0.8
0.5
32
0.7
0.3
0.07
Fig. 4 Western blot of lysates of HCT116 human colon cancer cells treated with (A) compound 28 at 5 × GI50 for 8, 24 and 48 h, and (B) with 5 ×
GI50 concentrations of compounds 31 or 32 for 8, 24 and 48 h. GAPDH was used as loading control.
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of DEPT spectra as well as COSY, HMQC and HMBC corre- (M + H+, 100%). Data are consistent with those reported in
lation techniques. LCMS was used to determine purity of literature.29
compounds, with purity ≥95% unless otherwise stated, as
measured by the area under curve (AUC) of the UV trace at
254 nm.
Experimental procedures
General procedure 1: synthesis of trisubstituted imidazoles.
2-(4-Fluorophenyl)-1H-phenanthro[9,10-d]imidazole 7b. Pre-
pared by general procedure 1 from phenanthrene-9,10-dione
(500 mg, 2.40 mmol) and 4-fluorobenzaldehyde (254 μL,
2.64 mmol). Purification by recrystallization from hexane–
dichloromethane (1 : 2) gave the title compound as a colourless
solid (251 mg, 34%); mp 214–215 °C; (Found: M + H+,
313.1143. C21H13FN2 + H+ requires 313.1136); νmax (CHCl3)/cm−1
3459, 3010, 1609, 1525, 1483, 1455, 1428, 1240, 1158, 1100, 842;
δH (400 MHz; DMSO-d6) 8.87 (1 H, d, J 8.0, ArH), 8.83 (1 H, d,
J 8.0, ArH), 8.59 (1 H, d, J 8.0, ArH), 8.53 (1 H, d, J 8.0, ArH), 8.37
(1 H, d, J 8.8, ArH), 8.35 (1 H, d, J 8.8, ArH), 7.76 (1 H, t, J 7.2,
ArH), 7.72 (1 H, t, J 7.2, ArH), 7.66–7.61 (2 H, m, ArH), 7.46 (2 H,
t, J 8.8, ArH); δC (100 MHz; DMSO-d6) 163.2 (d, J 245, CF), 148.7
(C), 137.4 (C), 128.8 (d, J 8, CH), 128.17 (C), 128.01 (C), 127.64
(CH), 127.56 (CH), 127.53 (C), 127.50 (C), 127.4 (C), 125.8 (CH),
125.64 (CH), 124.57 (CH), 124.2 (CH), 122.9 (C), 122.4 (CH),
122.3 (CH), 116.5 (d, J 21, CH); m/z (ESI) 313 (M + H+, 100%).
A 35 mL microwave vial was charged with 1,2-diketone
(0.5–5 mmol), aryl aldehyde (1.2 eq.), ammonium acetate
(10 eq.) and glacial acetic acid (6410 mL). The mixture was
heated at 150 °C for 15–30 min under microwave irradiation
(300 W). Following reaction completion by TLC, the mixture
was cooled to room temperature and added slowly to
ammonium hydroxide (35%; 30 mL). The resulting precipi-
tate was collected by filtration and purified by flash column
chromatography, recrystallization or trituration to give the
product.
General procedure 2a: alkylation of 1H-imidazoles. In an
oven-dried flask,
a solution of trisubstituted imidazole
(0.5–1 mmol) in THF or DMF (10 mL mmol−1) was treated
with potassium tert-butoxide (2 eq.) and the mixture stirred
for 5 min The alkylating agent (2–5 eq.) was added and the
mixture stirred at ambient temperature for 16 h to 3 days.
Once the reaction was complete, as judged by TLC, the
mixture was added to water (20–100 mL) and extracted with
ethyl acetate (3 × 10–50 mL). The combined organic extracts
were washed with saturated brine (20–100 mL), dried (MgSO4)
and concentrated onto silica. Purification by flash column
chromatography and trituration gave the product.
General procedure 2b: alkylation of 1H-imidazoles. General
procedure 2a was applied with the following changes: potassium
tert-butoxide was replaced with potassium carbonate (2 eq.),
and THF or DMF was replaced with acetonitrile (10 mL mmol−1).
2-(4-Methoxyphenyl)-1H-phenanthro[9,10-d]imidazole 7c. Pre-
pared by general procedure 1 from phenanthrene-9,10-dione
(300 mg, 1.44 mmol) and 4-methoxybenzaldehyde (193 μL,
1.58 mmol). Purification by recrystallization from ethanol gave
the title compound as a beige solid (196 mg, 42%); mp
265–266 °C (lit.,30 mp 265–266 °C); (Found: M + H+, 325.1331.
C22H16N2O + H+ requires 325.1335); νmax (CHCl3)/cm−1 3690,
3608, 1602, 1559, 1484, 1178, 839; δH (400 MHz; DMSO-d6)
13.30 (1 H, s, NH), 8.85 (1 H, d, J 8.4, ArH), 8.83 (1 H, d, J 8.4,
2-Phenyl-1H-phenanthro[9,10-d]imidazole 7a. Prepared by ArH), 8.60 (1 H, d, J 8.0, ArH), 8.54 (1 H, d, J 8.0, ArH), 8.27
general procedure 1 from phenanthrene-9,10-dione (300 mg, (2 H, d, J 8.4, ArH), 7.74 (1 H, t, J 7.4, ArH), 7.72 (1 H, t, J 7.4,
1.44 mmol) and benzaldehyde (175 μL, 1.73 mmol). Purifi- ArH), 7.62 (2 H, t, J 7.4, ArH), 7.17 (2 H, d, J 8.4, ArH), 3.87
cation by recrystallization from dichloromethane–methanol (3 H, s, Me); δC (100 MHz; DMSO-d6) 160.7 (C), 149.8 (C), 137.4
(1 : 1) gave the title compound as a beige solid (294 mg, 69%); (2 × C), 128.2 (CH), 128.0 (C), 127.93 (C), 127.87 (C), 127.52
mp 323–324 °C (lit.,1 mp 322–324 °C); (Found: M + H+, (CH), 127.48 (CH), 125.6 (CH), 125.5 (CH), 124.5 (CH), 124.2
295.1217. C21H14N2 + H+ requires 295.1230); νmax (CHCl3)/cm−1 (CH), 123.5 (C), 122.9 (C), 122.3 (2 × CH), 114.9 (CH), 55.8
3691, 3606, 3459, 3007, 1602, 1474, 1242, 824; δH (400 MHz; (Me); m/z (ESI) 325 (M + H+, 100%).
DMSO-d6) 8.86 (1 H, d, J 8.4, ArH), 8.83 (1 H, d, J 8.4, ArH),
8.61 (1 H, d, J 8.0, ArH), 8.57 (1 H, d, J 8.0, ArH), 8.33 (2 H, dd,
J 8.4, 1.2, ArH), 7.76 (1 H, t, J 7.4, ArH), 7.73 (1 H, t, J 7.4, ArH),
7.65–7.59 (4 H, m, ArH), 7.50 (1 H, t, J 7.4, ArH); δC (100 MHz;
DMSO-d6) 149.6 (C), 137.5 (C), 130.9 (C), 129.7 (CH), 129.4 (CH),
128.17 (C), 128.15 (C), 128.1 (C), 127.6 (CH), 127.54 (CH),
127.46 (C), 126.6 (CH), 125.8 (CH), 125.7 (CH), 124.6 (CH),
2-(3,5-Bis(trifluoromethyl)phenyl)-1H-phenanthro[9,10-d]imida-
124.2 (CH), 122.9 (C), 122.5 (CH), 122.4 (CH); m/z (ESI) 295 zole 7d. Prepared by general procedure 1 from phenanthrene-
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9,10-dione (1.00 g, 4.80 mmol) and 3,5-bis(trifluoromethyl)- acetate (3 × 15 mL). The combined organic extracts were
benzaldehyde (871 μL, 5.28 mmol). Purification by flash washed with water (30 mL) and saturated brine (30 mL), dried
chromatography, eluting with light petroleum–ethyl acetate (MgSO4) and concentrated under reduced pressure. Purifi-
(9 : 1), followed by trituration with ether (20 mL) gave the cation by flash chromatography, eluting with cyclohexane–
title compound as a colourless solid (1.08 g, 52% yield, 63% dichloromethane (1 : 1), gave the title compound as a cream
purity); mp 307–308 °C; (Found: M + H+, 431.0959. C23H12F6N2 solid (262 mg, 81%); mp 152–153 °C (lit.,25 mp 162–163 °C);
+ H+ requires 431.0977); νmax (CHCl3)/cm−1 3451, 3012, 1621, (Found: M + H+, 393.9191. C16H1279Br2O2 + H+ requires
1427, 1280, 1184, 1144, 1094, 1016, 896, 847; δH (400 MHz; 393.9199); νmax (CHCl3)/cm−1 3010, 2843, 1615, 1591, 1486,
acetone-d6) 12.98 (1 H, s, NH), 8.90 (3 H, br s, ArH), 8.85 (1 H, 1346, 1312, 1122, 1095, 986; δH (400 MHz; CDCl3) 8.65 (2 H, d,
d, J 8.0, ArH), 8.75 (1 H, d, J 8.0, ArH), 8.38 (1 H, d, J 7.4, ArH), J 1.8, ArH), 8.10 (2 H, d, J 8.8, ArH), 7.73 (2 H, dd, J 8.8, 1.8,
8.13 (1 H, s, ArH), 7.76 (1 H, t, J 7.4, ArH), 7.72–7.67 (3 H, m, ArH), 4.08 (6 H, s, Me); δC (100 MHz; CDCl3) 143.8 (C), 130.6
ArH); δC (100 MHz; acetone-d6) 146.8 (C), 138.9 (C), 134.0 (CH), 128.9 (C), 128.2 (C), 125.4 (CH), 124.0 (CH), 120.5 (C),
(2 × C), 132.7 (q, J 33, C), 129.8 (C), 129.3 (C), 129.2 (C), 128.3 61.0 (Me); m/z (ESI) 394/396/398 (M + H+, 8/16/8%). Data are
(C), 128.1 (CH), 127.9 (CH), 126.91 (CH), 126.87 (CH), 126.7 consistent with those reported in literature.25
(CH), 126.5 (CH), 124.9 (CH), 124.5 (q, J 270, CF3), 124.4 (CH),
123.3 (CH), 122.7 (CH), m/z (ESI) 431 (M + H+, 100%).
3,6,9,10-Tetramethoxyphenanthrene 10. Copper(I) bromide
was purified prior to use by stirring in glacial acetic acid for
3,6-Dibromophenanthrene-9,10-dione 8. Phenanthrene-9,10-
dione (5.00 g, 24.0 mmol) was dissolved in nitrobenzene
(48 mL) and treated with benzoyl peroxide (582 mg,
2.40 mmol) followed by the slow addition of bromine
(2.52 mL, 49.2 mmol). The mixture was heated at 110 °C for
16 h, cooled to room temperature and diluted with ethanol
(100 mL). The resulting precipitate was collected by filtration
to give the first crop, and the filtrate concentrated. The residue
was triturated with ethanol (50 mL) and filtered to give a
second crop, which was combined with the first to give the
title compound as a light brown solid (6.73 g, 77%); mp
283–285 °C (lit.,31 mp 281–284 °C); (Found: M + H+, 386.8639.
C14H679Br2O2 + H+ requires 386.8627); λmax (MeOH)/nm 331
(log ε 3.46); νmax (CHCl3)/cm−1 3012, 1681, 1587, 1548, 1289,
923, 831; δH (400 MHz; DMSO-d6) 8.63 (2 H, br s, ArH), 7.91
(2 H, d, J 8.4, ArH), 7.75 (2 H, dd, J 8.4, 1.4, ArH); δC (100 MHz;
DMSO-d6) 177.6 (C), 135.9 (C), 132.7 (CH), 130.8 (CH), 130.7
(C), 130.0 (C), 127.7 (CH); m/z (ESI) 388 (M + H+, 100%).
16 h, filtering and triturating with ethanol and ether. 3,6-
Dibromo-9,10-dimethoxyphenanthrene 9 (3.02 g, 7.62 mmol)
was suspended in ethyl acetate–toluene (1 : 1; 8 mL) and
treated slowly with a solution of sodium methoxide (25%;
64 mL, 305 mmol). Copper(I) bromide (218 mg, 1.52 mmol)
was added and the resulting mixture heated at 80 °C for 64 h.
The reaction mixture was added to water (100 mL) and
extracted with dichloromethane (3 × 75 mL). The combined
organic extracts were dried (MgSO4), concentrated onto silica
gel and purified by flash chromatography, eluting with light
petroleum–ethyl acetate (9 : 1) to give the title compound as a
cream solid (1.95 g, 86%); mp 73–74 °C (lit.,25 mp 74–76 °C);
(Found: M + Na+, 321.1103. C18H18O4 + Na+ requires 321.1097);
νmax (CHCl3)/cm−1 3011, 2939, 2838, 1608, 1509, 1449, 1431,
1370, 1356, 1319, 1258, 1240, 1173, 1116, 1073, 1036, 990;
δH (400 MHz; CDCl3) 8.14 (2 H, d, J 9.0, ArH), 7.93 (2 H, d,
J 2.2, ArH), 7.30–7.27 (2 H, m, ArH), 4.07 (6 H, s, Me), 4.02
(6 H, s, Me); δC (100 MHz; CDCl3) 157.7 (C), 142.3 (C),
129.3 (C), 124.0 (C), 123.8 (CH), 116.5 (CH), 104.7 (CH),
61.0 (Me), 55.6 (Me); m/z (ESI) 321 (M + Na+, 100%). Data are
consistent with those reported in literature.25
3,6-Dibromo-9,10-dimethoxyphenanthrene 9. 3,6-Dibromophe-
nanthrene-9,10-dione 8 (300 mg, 0.82 mmol) was suspended
in THF–water (1 : 1; 14 mL) and treated with tetrabutyl-
ammonium bromide (106 mg, 0.33 mmol) and sodium dithio-
3,6-Dimethoxyphenanthrene-9,10-dione 11. 3,6,9,10-Tetra-
nite (571 mg, 3.28 mmol). The reaction mixture was stirred at methoxyphenanthrene 10 (330 mg, 1.11 mmol) was dissolved
room temperature for 15 min, dimethyl sulfate (543 μL, in acetonitrile (6 mL) and treated with solution of
a
5.74 mmol) added slowly and the mixture stirred for an ammonium cerium(IV) nitrate (1.21 g, 2.21 mmol) in aceto-
additional 15 min. Ice (10 g) was added and the reaction nitrile (15 mL). Water (45 mL) was added slowly and the result-
mixture stirred for 15 min, and then extracted with ethyl ing precipitate collected by filtration and triturated with
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ethanol to give the title compound as a yellow solid (236 mg, 127.4 (C), 127.3 (C), 125.7(CH), 125.6 (CH), 123.5 (CH), 123.3
79%); mp 228–229 °C (lit.,25 mp 226–230 °C); (Found: M + Na+, (q, J 271, CF3), 121.6 (CH), 116.7 (C), 116.5 (CH), 116.4 (CH),
291.0640. C16H12O4 + Na+ requires 291.0628); λmax (MeOH)/nm 107.1 (CH), 106.5 (CH), 55.6 (Me); m/z (ESI) 491 (M + H+, 100%).
349 (log ε 5.35); νmax (CHCl3)/cm−1 3011, 2843, 1670, 1595,
1566, 1344, 1324, 1313, 1247; δH (400 MHz; CDCl3) 8.17 (2 H,
d, J 8.6, ArH), 7.35 (2 H, d, J 2.0, ArH), 6.95 (2 H, dd, J 8.6, 2.0,
ArH), 3.97 (6 H, s, Me); δC (100 MHz; CDCl3) 179.0 (C), 165.6
(C), 137.5 (C), 133.3 (CH), 125.1 (C), 114.3 (CH), 109.8 (CH),
55.9 (Me); m/z (ESI) 291 (M + Na+, 100%). Data are consistent
with those reported in literature.25
2-(4-Methoxyphenyl)-4,5-diphenyl-1H-imidazole 14. Prepared
by general procedure 1 from benzil (300 mg, 1.43 mmol) and
4-methoxybenzaldehyde (174 μL, 1.43 mmol). Purification by
recrystallization from hexane–ethanol (1 : 4) gave the title com-
pound as a colourless solid (285 mg, 61%); mp 233–234 °C
(lit.,32 mp 234 °C); (Found: M + H+, 327.1483. C22H18N2O + H+
requires 327.1492); νmax (CHCl3)/cm−1 3690, 3449, 3011, 2839,
1615, 1506, 1464, 1254, 1177, 1032, 835; δH (400 MHz; DMSO-
6,9-Dimethoxy-2-phenyl-1H-phenanthro[9,10-d]imidazole 12. Pre-
d6) 12.49 (1 H, s, NH), 8.01 (2 H, d, J 8.8, ArH), 7.54 (2 H, d,
pared by general procedure 1 from 3,6-dimethoxyphenan-
J 7.6, ArH), 7.49 (2 H, d, J 7.6, ArH), 7.43 (2 H, t, J 7.2, ArH),
threne-9,10-dione 11 (150 mg, 0.56 mmol) and benzaldehyde
7.36 (1 H, t, J 7.2, ArH), 7.30 (2 H, t, J 7.2, ArH), 7.21 (1 H, t,
(62 μL, 0.61 mmol). Purification by flash chromatography,
J 7.2, ArH), 7.04 (2 H, d, J 8.8, ArH), 3.82 (3 H, s, Me);
eluting with light petroleum–ethyl acetate (3 : 1), followed by
δC (100 MHz; DMSO-d6) 159.4 (C), 146.6 (C), 136.7 (C), 135.3
trituration with ether (15 mL) gave the title compound as a pale
(C), 131.2 (C), 128.6 (CH), 128.3 (CH), 128.1 (CH), 127.6 (CH),
yellow solid (100 mg, 51%); mp 273–274 °C; (Found: M + H+,
127.0 (CH), 126.7 (CH), 126.43 (C), 126.39 (CH), 123.1 (C),
355.1434. C23H18N2O2 + H+ requires 355.1441); νmax (CHCl3)/
114.1 (CH), 55.2 (Me); m/z (ESI) 327 (M + H+, 100%).
cm−1 3462, 3006, 1731, 1624, 1594, 1548, 1468, 1456, 1408,
1389, 1173, 1031; δH (400 MHz; DMSO-d6) 13.25 (1 H, s, NH),
8.48 (2 H, d, 8.4, ArH), 8.28 (2 H, dd, J 8.4, 1.2, ArH), 8.24 (1 H,
br s, ArH), 8.21 (1 H, br s, ArH), 7.59 (2 H, t, J 7.2, ArH), 7.48
(1 H, t, J 7.2, ArH), 7.44 (1 H, d, J 8.4, ArH), 7.39 (1 H, d, J 8.4,
ArH), 4.03 (6 H, s, Me); δC (100 MHz; DMSO-d6) 157.1 (C),
157.0 (C), 148.1 (C), 135.7 (C), 130.6 (C), 128.9 (2 × CH), 128.8
(C), 128.4 (C), 126.5 (C), 125.9 (CH), 123.5 (CH), 123.3 (CH),
2,4,5-Tri(4-methoxyphenyl)-1H-imidazole 15. Prepared by
from 4,4′-dimethoxybenzil (250 mg,
121.6 (C), 117.1 (C), 116.5 (CH), 116.4 (CH), 106.9 (CH), general procedure
1
106.4 (CH), 55.6 (Me), 55.5 (Me); m/z (ESI) 355 (M + H+, 100%). 0.92 mmol) and benzaldehyde (124 μL, 1.02 mmol). Purification
by flash chromatography, eluting with light petroleum–ethyl
acetate (2 : 3), followed by trituration with ether (10 ml) gave the
title compound as a cream solid (249 mg, 70%); mp 184–185 °C
(lit.,33 mp 183–184 °C); (Found: M + H+, 387.1702. C24H22N2O3 +
H+ requires 387.1703); νmax (CHCl3)/cm−1 3450, 3011, 2962,
2839, 1616, 1520, 1499, 1466, 1441, 1295, 1248, 1176, 1034, 836;
δH (400 MHz; DMSO-d6) 7.99 (2 H, d, J 8.6, ArH), 7.46 (2 H, d,
2-(3,5-Bis(trifluoromethyl)phenyl)-6,9-dimethoxy-1H-phenan-
J 8.6, ArH), 7.40 (2 H, d, J 8.6, ArH), 7.04–6.99 (4 H, m, ArH),
thro[9,10-d]imidazole 13. Prepared by general procedure 1 from
6.87 (2 H, d, J 8.6, ArH), 3.81 (3 H, s, Me), 3.80 (3 H, s, Me), 3.74
3,6-dimethoxyphenanthrene-9,10-dione 11 (150 mg, 0.56 mmol)
(3 H, s, Me); δC (100 MHz; DMSO-d6) 159.2 (C), 158.6 (C), 157.8
and 3,5-bis(trifluoromethyl)benzaldehyde (101 μL, 0.61 mmol).
(C), 144.9 (C), 136.0 (C), 129.6 (CH), 128.1 (CH), 128.0 (C), 126.5
Purification by flash chromatography, eluting with light pet-
(CH), 123.7 (C), 123.3 (2 × C), 114.0 (2 × CH), 113.6 (CH), 55.2
roleum–ethyl acetate (85 : 15), followed by trituration with
(2 × Me), 55.0 (Me); m/z (ESI) 357 (M + H+, 100%).
dichloromethane (10 mL) gave the title compound as a yellow
solid (221 mg, 56%); mp 256–257 °C; (Found: M + H+, 491.1199.
C25H16F6N2O2 + H+ requires 491.1189); λmax (CH2Cl2)/nm 390
(log ε 5.58), 369 (5.53); νmax (CHCl3)/cm−1 3454, 1624, 1600,
1467, 1361, 1280, 1175, 1143; δH (400 MHz; DMSO-d6) 13.55
(1 H, br s, NH), 8.83 (2 H, s, ArH), 8.39 (2 H, br s, ArH), 8.19
(2 H, br s, ArH), 8.15 (1 H, s, ArH), 7.39 (2 H, d, J 7.6, ArH), 4.01
(6 H, s, Me); δC (100 MHz; DMSO-d6) 157.5 (C), 157.4 (C), 144.8
2,4,5-Tri(4-fluorophenyl)-1H-imidazole 16. Prepared by
(C), 136.0 (C), 132.9 (C), 131.0 (q, J 33, C), 129.4 (C), 128.8 (C), general procedure 1 from 4,4′-difluorobenzil (300 mg,
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1.22 mmol) and 4-fluorobenzaldehyde (117 μL, 1.22 mmol). 309.1386); νmax (CHCl3)/cm−1 3069, 3008, 2960, 1613, 1576,
Purification by recrystallization from hexane–ethanol (1 : 2) 1532, 1473, 1462, 1422, 1380, 1329, 1028; δH (400 MHz;
gave the title compound as a colourless solid (183 mg, 43%); DMSO-d6) 8.97 (1 H, d, J 8.0, ArH), 8.86 (1 H, d, J 8.0, ArH),
mp 245–246 °C (lit.,34 mp 162–164 °C); (Found: C, 72.1; H, 3.7; 8.60 (2 H, t, J 7.2, ArH), 7.88 (2 H, d, J 6.0, ArH), 7.76–7.62
N, 8.0. C21H13F3N2 requires C, 72.0; H, 3.7; N, 8.0%); (Found: (7 H, m, ArH), 4.30 (3 H, s, Me); δC (100 MHz; DMSO-d6)
M + H+, 351.1097. C21H13F3N2 + H+ requires 351.1104); νmax 152.1 (C), 136.6 (C), 130.2 (C), 129.7 (CH), 129.4 (CH),
(CHCl3)/cm−1 3447, 2974, 1602, 1518, 1497, 1439, 1240, 1157, 128.7 (CH), 128.2 (C), 127.5 (C), 127.4 (C), 127.3 (CH),
1094, 841; δH (400 MHz; DMSO-d6) 12.73 (1 H, br s, NH), 8.10 127.1 (CH), 126.9 (C), 125.4 (CH), 125.1 (CH), 124.5 (CH),
(1 H, d, J 9.0, ArH), 8.09 (1 H, d, J 9.0, ArH), 7.53 (2 H, d, J 8.8, 123.6 (CH), 123.2 (C), 121.9 (CH), 121.3 (CH), 36.0 (Me); m/z
ArH), 7.52 (2 H, d, J 8.8, ArH), 7.33 (2 H, t, J 9.0, ArH), (ESI) 309 (M + H+, 100%).
7.32–7.12 (4 H, m, ArH); δC (100 MHz; DMSO-d6) 162.2 (d, J
244, CF), 161.7 (d, J 244, CF), 161.1 (d, J 242, CF), 144.7 (C),
136.2 (C), 131.4 (d, J 3, C), 130.6 (d, J 7, CH), 128.9 (d, J 9, CH),
127.31 (d, J 3, C), 127.30 (d, J 8, CH), 127.1 (C), 126.9 (d, J 3, C),
115.73 (d, J 22, CH), 115.68 (d, J, 21, CH), 115.1 (d, J 21, CH);
m/z (ESI) 351 (M + H+, 100%).
1-Benzyl-2-phenyl-1H-phenanthro[9,10-d]imidazole
19. Phe-
nanthrene-9,10-dione (250 mg, 1.20 mmol) was suspended in
glacial acetic acid (6 mL) and ammonium acetate (463 mg,
6.00 mmol), benzylamine (144 μL, 1.32 mmol) and benz-
aldehyde (133 µL, 1.32 mmol) were added. The mixture was
heated at 150 °C for 15 min under microwave irradiation (300
W), cooled to room temperature and added to water (20 mL).
The aqueous solution was extracted with ethyl acetate (3 ×
10 mL), and the combined organic extracts were washed with
saturated brine (20 mL), dried (MgSO4) and concentrated onto
silica. Purification by flash chromatography, eluting with light
petroleum–ethyl acetate (3 : 1), followed by trituration with
ether (10 mL) gave the title compound as a colourless solid
(96 mg, 21%); mp 239–240 °C (lit.,38 mp 241 °C); (Found: M +
H+, 385.1698. C28H20N2 + H+ requires 385.1699); νmax (CHCl3)/
cm−1 3068, 3011, 2968, 1605, 1531, 1512, 1498, 1474, 1455,
1426, 1391, 1265; δH (400 MHz; DMSO-d6) 8.92 (1 H, d, J 8.2,
ArH), 8.86 (1 H, d, J 8.2, ArH), 8.66 (1 H, dd, J 8.2, 1.2, ArH),
8.10 (1 H, dd, J 8.2, 1.2, ArH), 7.78–7.71 (3 H, m, ArH), 7.67
(1 H, td, J 7.6, 1.2, ArH), 7.59–7.54 (4 H, m, ArH), 7.49 (1 H, td,
J 7.2, 1.2, ArH), 7.34 (2 H, t, J 7.6, ArH), 7,26 (1 H, t, J 7.2, ArH),
7.13 (2 H, d, J 7.6, ArH), 5.96 (2 H, s, CH2); δC (100 MHz;
DMSO-d6) 152.6 (C), 137.14 (C), 137.08 (C), 130.2 (C),
129.7 (CH), 129.3 (CH), 129.1 (CH), 128.8 (CH), 128.3 (C), 127.7
(C), 127.5 (CH), 127.4 (CH), 126.9 (CH), 126.8 (C), 126.4 (C),
125.7 (CH), 125.4 (CH), 125.1 (CH), 124.3 (CH), 123.6 (CH),
122.4 (C), 121.9 (CH), 121.2 (CH), 50.0 (CH2); m/z (ESI) 385
(M + H+, 100%).
4-(Bromomethyl)benzamide 17. 4-(Bromomethyl)benzoic acid
(2.50 g, 11.6 mmol) was suspended in ethyl acetate (45 mL)
and treated with thionyl chloride (1.27 mL, 17.4 mmol). The
mixture was heated at 75 °C for 15 h, cooled and concentrated
under reduced pressure. The residue was dissolved in dichloro-
methane (45 mL), cooled to 0 °C and ammonium hydroxide
(35%; 1.13 mL, 23.3 mmol) added slowly. The mixture was
stirred at 0 °C for 15 min and then quenched with water
(45 mL). The resulting precipitate was filtered, washed with
water (50 mL) and dried under vacuum to give the title com-
pound as a colourless solid (2.22 g, 89%); mp 181–182 °C
(lit.,35 mp 185–187 °C); (Found:
M +
Na+, 235.9684.
C8H879BrNO + Na+ requires 235.9681); νmax (CHCl3)/cm−1 3691,
3521, 1734, 1697, 1611, 1421, 1371, 1315, 1286, 1171, 1089;
δH (400 MHz; DMSO-d6) 7.99 (1 H, br s, NH), 7.84 (2 H, d,
J 8.4, ArH), 7.51 (2 H, d, J 8.4, ArH), 7.40 (1 H, br s, NH), 4.73
(2 H, s, CH2); δC (100 MHz; DMSO-d6) 167.3 (C), 141.1 (C),
134.1 (C), 129.1 (CH), 127.8 (CH), 33.5 (CH2); m/z (ESI) 235/237
(M + Na+, 98/100%). Data are consistent with those reported in
literature.36
1-Methyl-2-phenyl-1H-phenanthro[9,10-d]imidazole
18. Pre-
pared by general procedure 2 from 2-phenyl-1H-phenanthro
[9,10-d]imidazole 7a (200 mg, 0.68 mmol) and iodomethane
(85 μL, 1.36 mmol) in THF (7 mL). Purification by flash chrom-
1-Benzyl-2-(3,5-bis(trifluoromethyl)phenyl)-1H-phenanthro-
atography, eluting with light petroleum–ethyl acetate (4 : 1), fol- [9,10-d]imidazole 20. Prepared by general procedure 2 from
lowed by trituration with ether (15 mL) gave the title compound 2-(3,5-bis(trifluoromethyl)phenyl)-1H-phenanthro[9,10-d]imida-
as a colourless solid (135 mg, 64%); mp 184–185 °C (lit.,37 mp zole 7d (200 mg, 0.46 mmol) and benzyl bromide (111 µL,
185–186 °C); (Found: M + H+, 309.1388. C22H16N2 + H+ requires 0.93 mmol) in THF (5 mL). Purification by flash chromato-
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graphy, eluting with light petroleum–ethyl acetate (9 : 1), fol- 0.64 mmol) and 4-(bromomethyl)benzamide 17 (206 mg,
lowed by trituration with ether gave the title compound as a col- 0.96 mmol) in THF (6 mL). The mixture was added to water
ourless solid (108 mg, 45%); mp 215–216 °C; (Found: M + H+, (20 mL), and the resulting precipitate filtered to give the first
521.1433. C30H18F6N2 + H+ requires 521.1447); νmax (CHCl3)/ crop of product. The filtrate was extracted with ethyl acetate
cm−1 2361, 1455, 1396, 1353, 1280, 1186, 1144, 1038, 905, 850; (3 × 10 mL) and the combined organic extracts washed with
δH (400 MHz; acetone-d6) 8.96 (1 H, d, J 8.8, ArH), 8.87 (1 H, d, saturated brine (20 mL), dried (Na2SO4) and concentrated
J 8.0, ArH), 8.79 (1 H, dd, J 8.0, 1.4, ArH), 8.36 (2 H, br s, ArH), under reduced pressure. The residue was adsorbed onto silica
8.21 (1 H, d, J 8.8, ArH), 8.19 (1 H, s, ArH), 7.78 (1 H, t, J 7.2, and purified by flash chromatography, eluting with light pet-
ArH), 7.71 (1 H, t, J 7.6, ArH), 7.63 (1 H, t, J 7.6, ArH), 7.52 roleum–ethyl acetate (1 : 4), followed by trituration with ether
(1 H, t, J 7.6, ArH), 7.45 (2 H, t, J 7.6, ArH), 7.39–7.34 (3 H, m, to give a second crop. Both crops were combined to give the
ArH), 6.11 (2 H, s, CH2); δC (100 MHz; acetone-d6) 150.7 (C), title compound as a colourless solid (124 mg, 43% yield, 91%
139.0 (C), 137.9 (C), 134.2 (C), 132.6 (q, J 34, C), 130.6 (CH), purity); mp 300–301 °C; (Found:
M +
H+, 446.1673.
130.5 (CH), 130.4 (C), 130.3 (CH), 129.4 (C), 129.0 (C), 128.9 C29H20FN3O + H+ requires 446.1663); νmax (CHCl3)/cm−1 3691,
(CH), 128.3 (CH), 128.0 (CH), 127.0 (q, J 270, CF3), 126.9 (CH), 3491, 3291, 3152, 1683, 1615, 1567, 1532, 1474, 1414, 1162;
126.8 (CH), 126.5 (CH), 125.3 (CH), 124.4 (CH), 123.83 (C), δH (400 MHz; DMSO-d6) 8.93 (1 H, d, J 8.0, ArH), 8.87 (1 H, d,
123.78 (C), 123.4 (CH), 122.5 (CH), 51.7 (CH2); m/z (ESI) 521 J 8.0, ArH), 8.66 (1 H, dd, J 8.0, 1.4, ArH), 8.05 (1 H, d, J 8.0,
(M + H+, 100%).
ArH), 7.91 (1 H, br s, NH), 7.80 (2 H, d, J 8.0, ArH), 7.77–7.74
(3 H, m, ArH), 7.68 (1 H, td, J 8.0, 1.4, ArH), 7.58 (1 H, td, J 8.0,
1.4, ArH), 7.49 (1 H, td, J 8.0, 1.4, ArH), 7.41 (2 H, t, J 8.0, ArH),
7.33 (1 H, br s, NH), 7.21 (2 H, d, J 8.0, ArH), 5.98 (2 H, s, CH2);
δC (100 MHz; DMSO-d6) 167.4 (C), 162.9 (d, J 245, CF), 151.7
(C), 140.2 (C), 137.1 (C), 133.7 (C), 131.4 (d, J 9, CH), 128.4
(CH), 127.7 (2 × C), 127.4 (CH), 127.0 (CH), 126.7 (C), 126.6
(d, J 4, C), 126.3 (C), 125.8 (CH), 125.4 (CH), 125.2 (CH),
124.4 (CH), 123.6 (CH), 122.3 (C), 122.0 (CH), 121.1 (CH), 115.9
(d, J 22, CH), 49.1 (CH2); m/z (ESI) 446 (M + H+, 91%).
1-(4-Carboxamidobenzyl)-2-phenyl-1H-phenanthro[9,10-d]imid-
azole 21. Prepared by general procedure 2b from 2-phenyl-1H-
phenanthro[9,10-d]imidazole 7a (200 mg, 0.68 mmol) and
4-(bromomethyl)benzamide 17 (175 mg, 0.82 mmol). Purifi-
cation by flash chromatography, eluting with light petroleum–
ethyl acetate (1 : 4) followed by recrystallization from dichloro-
methane–methanol (1 : 1) gave the title compound as a colour-
less solid (16 mg, 6%); mp 280–281 °C; (Found: M + H+,
428.1752. C29H21N3O + H+ requires 428.1757); νmax (CHCl3)/
cm−1 3690, 3607, 3012, 1682, 1602, 1540, 1474, 1458, 1368,
1239; δH (400 MHz; DMSO-d6) 8.92 (1 H, d, J 8.0, ArH), 8.86
(1 H, d, J 8.0, ArH), 8.67 (1 H, d, J 8.0, ArH), 8.05 (1 H, d, J 8.0,
ArH), 7.92 (1 H, br s, NH), 7.82 (2 H, d, J 8.2, ArH), 7.78–7.66
(4 H, m, ArH), 7.59–7.55 (4 H, m, ArH), 7.47 (1 H, t, J 8.0, ArH),
7.33 (1 H, br s, NH), 7.21 (2 H, d, J 8.2, ArH), 6.00 (2 H, s, CH2);
δC (100 MHz; DMSO-d6) 167.4 (C), 152.6 (C), 140.3 (C), 137.2
(C), 133.6 (C), 130.1 (2 × C), 129.7 (CH), 129.3 (CH), 128.8 (CH),
128.3 (CH), 127.7 (C), 127.4 (CH), 127.0 (C), 126.8 (CH), 126.3
(C), 125.8 (CH), 125.4 (CH), 125.1 (CH), 124.4 (CH), 123.6 (CH),
122.4 (C), 122.0 (CH), 121.1 (CH), 49.9 (CH2); m/z (ESI) 428
(M + H+, 65%).
1-(4-Carboxamidobenzyl)-2-(4-methoxyphenyl)-1H-phenanthro-
[9,10-d]imidazole 23. Prepared by general procedure 2 from
2-(4-methoxyphenyl)-1H-phenanthro[9,10-d]imidazole 7c
(200 mg, 0.62 mmol) and 4-(bromomethyl)benzamide 17
(198 mg, 0.92 mmol) in DMF (6 mL). Purification by flash
chromatography, eluting with dichloromethane–methanol
(9 : 1), followed by trituration with dichloromethane (10 mL)
gave the title compound as a colourless solid (42 mg, 15% yield,
67% purity); mp 297–298 °C; (Found: M + H+, 458.1850.
C30H22N3O2 + H+ requires 458.1863); νmax (CHCl3)/cm−1 3691,
3513, 3319, 3164, 3012, 1686, 1606, 1568, 1476, 1417, 1298,
1264, 1184, 950; δH (400 MHz; DMSO-d6) 8.93 (1 H, d, J 8.4,
ArH), 8.86 (1 H, d, J 8.4, ArH), 8.66 (1 H, d, J 8.4, ArH), 8.04
(1 H, d, J 8.4, ArH), 7.92 (1 H, br s, NH), 7.82 (1 H, d, J 8.0,
ArH), 7.75 (1 H, t, J 7.6, ArH), 7.69–7.63 (3 H, m, ArH), 7.56
(1 H, t, J 7.8, ArH), 7.48 (1 H, t, J 7.8, ArH), 7.33 (1 H, s, NH),
7.21 (2 H, d, J 8.4, ArH), 7.10 (2 H, d, J 8.4, ArH), 5.98 (2 H, s,
CH2), 3.82 (3 H, s, Me); δC (100 MHz; DMSO-d6) 167.5 (C),
160.4 (C), 152.7 (C), 140.5 (C), 137.1 (C), 133.7 (C), 130.8 (CH),
1-(4-Carboxamidobenzyl)-2-(4-fluorophenyl)-1H-phenanthro- 128.4 (CH), 128.3 (C), 127.7 (C), 127.4 (CH), 127.0 (CH),
[9,10-d]imidazole 22. Prepared by general procedure 2 from 126.8 (C), 126.2 (C), 125.7 (CH), 125.4 (CH), 125.1 (CH),
2-(4-fluorophenyl)-1H-phenanthro[9,10-d]imidazole 7b (200 mg, 124.4 (CH), 123.6 (CH), 122.5 (C), 122.3 (C), 122.1 (CH),
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121.1 (CH), 114.3 (CH), 55.4 (Me), 50.0 (CH2); m/z (ESI) 458 (1 H, dd, J 8.8, 2.4, ArH), 7.33 (1 H, br s, NH), 7.19 (2 H, d,
(M + H+, 100%).
J 8.0, ArH), 7.15 (1 H, dd, J 8.8, 2.4, ArH), 5.94 (2 H, s, CH2),
4.01 (3 H, s, Me), 3.94 (3 H, s, Me); δC (100 MHz; DMSO-d6)
167.4 (C), 157.4 (C), 156.6 (C), 151.6 (C), 140.4 (C), 135.9 (C),
133.6 (C), 130.3 (2 × C), 129.6 (C), 129.5 (CH), 129.2 (CH), 128.8
(CH), 128.6 (C), 128.3 (CH), 125.4 (CH), 125.2 (C), 123.5 (CH),
122.6 (CH), 121.5 (C), 117.0 (CH), 115.8 (CH), 107.5 (CH), 106.1
(CH), 55.53 (Me), 55.45 (Me), 49.8 (CH2); m/z (ESI) 488 (M + H+,
100%).
1-(4-Carboxamidobenzyl)-2-(3,5-bis(trifluoromethyl)phenyl)-
1H-phenanthro[9,10-d]-imidazole 24. Prepared by general pro-
cedure 2 from 2-(3,5-bis(trifluoromethyl)phenyl)-1H-phenan-
thro[9,10-d]imidazole 7d (200 mg, 0.46 mmol) and 4-
(bromomethyl)benzamide 17 (199 mg, 0.93 mmol) in DMF
(5 mL). Purification by flash chromatography, eluting with
dichloromethane–methanol (19 : 1), followed by trituration
with dichloromethane (10 mL) gave the title compound as a col-
ourless solid (22 mg, 8%); mp 312–313 °C; (Found: M + H+,
564.1492. C31H20F6N3O + H+ requires 564.1505); νmax (CHCl3)/
cm−1 3688, 3414, 3010, 1680, 1602, 1353, 1280, 1186, 1145,
904; δH (400 MHz; DMSO-d6) 8.93 (1 H, d, J 8.4, ArH), 8.86
(1 H, d, J 8.4, ArH), 8.66 (1 H, d, J 8.4, ArH), 8.04 (1 H, d, J 8.4,
ArH), 7.92 (1 H, br s, NH), 7.82 (1 H, d, J 8.0, ArH), 7.75 (1 H, t,
J 7.6, ArH), 7.69–7.63 (3 H, m, ArH), 7.56 (1 H, t, J 7.8, ArH),
7.48 (1 H, t, J 7.8, ArH), 7.33 (1 H, s, NH), 7.21 (2 H, d, J 8.4,
ArH), 7.10 (2 H, d, J 8.4, ArH), 5.98 (2 H, s, CH2), 3.82 (3 H, s,
Me); δC (100 MHz; DMSO-d6) 167.3 (C), 149.6 (C), 139.9 (C),
137.4 (C), 133.6 (C), 132.5 (C), 130.8 (q, J 32, C), 129.8 (CH),
128.7 (C), 128.4 (CH), 127.9 (C), 127.6 (CH), 127.3 (CH), 127.2
(C), 126.6 (C), 126.1 (CH), 125.9 (q, J 270, CF3), 125.6 (CH),
124.5 (CH), 123.7 (CH), 123.4 (CH), 123.3 (CH), 122.1 (CH),
121.7 (C), 121.3 (CH), 50.3 (CH2); m/z (ESI) 564 (M + H+,
100%).
1-(4-Carboxamidobenzyl)-6,9-dimethoxy-2-(3,5-bis(trifluoro-
methyl)phenyl)-1H-phenanthro[9,10-d]imidazole 26. 2-(3,5-Bis-
(trifluoromethyl)phenyl)-6,9-dimethoxy-1H-phenanthro[9,10-d]-
imidazole 13 (250 mg, 0.51 mmol) was dissolved in THF
(5 mL) and potassium bis(trimethylsilyl)amide (0.7 M;
1.46 mL, 1.02 mmol) was added. The mixture was stirred for
5 min, treated with 4-(bromomethyl)benzamide 17 (546 mg,
2.55 mmol) and stirred at room temperature for 2 days. The
mixture was added to water (20 mL) and extracted with ethyl
acetate (3 × 15 mL). The combined organic extracts were
washed with saturated brine (20 mL), dried (Na2SO4) and con-
centrated onto silica. Purification by column chromatography,
eluting with light petroleum–ethyl acetate (2 : 3) followed by tri-
turation with ether (10 mL) gave the title compound as a colour-
less solid (36 mg, 11% yield, 94% purity); mp 319–320 °C;
(Found: M + H+, 624.1701. C33H23F6N3O3 + H+ requires
624.1716); νmax (CHCl3)/cm−1 3691, 3607, 3440, 3011, 1666,
1619, 1602, 1468, 1355, 1240, 1174, 1127, 1100, 1032; δH
(400 MHz; DMSO-d6) 8.59 (1 H, d, J 8.8, ArH), 8.30 (1 H, d,
J 2.4, ArH), 8.30 (3 H, br s, ArH), 8.23 (1 H, d, J 2.4, ArH), 8.01
(1 H, d, J 8.8, ArH), 7.,97 (1 H, br s, NH), 7.88 (2 H, d, J 8.8,
ArH), 7.43 (1 H, dd, J 8.8, 2.4, ArH), 7.37 (1 H, br s, NH), 7.29
(2 H, d, J 8.8, ArH), 7.20 (1 H, dd, J 8.8, 2.4, ArH), 5.99 (2 H, s,
CH2), 4.04 (3 H, s, Me), 3.97 (3 H, s, Me); δC (100 MHz; DMSO-
d6) 167.3 (C), 157.7 (C), 157.0 (C), 148.5 (C), 140.0 (C),
136.1 (C), 133.6 (C), 133.1 (CH), 132.6 (C), 130.8 (q, J 33, C),
130.1 (C), 129.5 (CH), 128.8 (C), 128.4 (CH), 126.3 (C),
125.6 (CH), 123.6 (CH), 123.0 (q, J 271, CF3), 122.7 (CH),
121.2 (C), 117.0 (CH), 116.7 (C), 115.9 (CH), 107.7 (CH),
106.2 (CH), 55.6 (Me), 55.5 (Me), 50.2 (CH2); m/z (ESI) 624
(M + H+, 40%).
1-(4-Carboxamidobenzyl)-6,9-dimethoxy-2-phenyl-1H-phenan-
thro[9,10-d]imidazole 25. Prepared by general procedure 2 from
6,9-dimethoxy-2-phenyl-1H-phenanthro[9,10-d]imidazole
12
(200 mg, 0.56 mmol) and 4-(bromomethyl)benzamide 17
(181 mg, 0.85 mmol) in THF (6 mL). The reaction mixture was
added to water (20 mL) and the resulting precipitate collected
by filtration and triturated with ethyl acetate (10 mL) followed
by ether (10 mL) to give the title compound as a colourless solid
(69 mg, 25% yield, 86% purity); mp 286–288 °C (decomp.);
(Found:
M + +
H+, 488.1966. C31H25N3O3 H+ requires
488.1969); νmax (CHCl3)/cm−1 3690, 3608, 1678, 1602, 1559,
1508, 1466, 1372, 1240; δH (400 MHz; DMSO-d6) 8.55 (1 H, d,
J 8.8, ArH), 8.27 (1 H, d, J 2.4, ArH), 8.20 (1 H, d, J 2.4, ArH),
7.94 (1 H, d, J 8.8, ArH), 7.92 (1 H, br s, NH), 7.82 (2 H, d, J 8.0,
1,10-Phenanthroline-5,6-dione 27. An ice-cold solution of
ArH), 7.69–7.67 (2 H, m, ArH), 7.54–7.52 (3 H, m, ArH), 7.41 concentrated sulfuric acid (5 mL) and concentrated nitric acid
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(2.5 mL) was added dropwise to 1,10-phenanthroline (500 mg, (2 H, s, ArH), 7.99 (1 H, s, ArH), 7.66 (2 H, br s, ArH); δC
2.77 mmol) and potassium bromide (495 mg, 4.16 mmol). The (100 MHz; acetone-d6) 149.1 (CH), 147.0 (C), 145.4 (C), 137.7
mixture was heated at 110 °C for 3 h, cooled to room tempera- (C), 133.9 (C), 132.7 (q, J 33, C), 131.4 (CH), 129.0 (C), 127.5
ture and added to ice-water (20 mL). The solution was neutral- (CH), 124.6 (CH), 124.3 (q, J 270, CF3), 123.2 (CH); m/z (ESI)
ized to pH 7 by the careful addition of saturated sodium 433 (M + H+, 100%).
hydroxide, and the resulting suspension was extracted with
dichloromethane (3 × 20 mL). The combined organic extracts
were dried (Na2SO4) and concentrated under reduced pressure,
and the residue triturated with ether (30 mL) to give the
title compound as a yellow solid (719 mg, 81%); mp 260–261 °C
(lit.,39 mp 259–260 °C); (Found: M + H+, 233.0326. C12H6N2O2
+ Na+ requires 233.0321); λmax (CH2Cl2)/nm 309 (log ε 3.12),
1-Methyl-2-phenyl-1H-imidazo[4,5-f][1,10]phenanthroline
375 (2.79); νmax (CHCl3)/cm−1 2981, 1704, 1690, 1580, 1566,
30. Prepared by general procedure
2 from 2-phenyl-1H-
1461, 1421, 1316, 1297, 1115, 1012, 927; δH (400 MHz; CDCl3)
9.14 (2 H, dd, J 4.8, 1.8, ArH), 8.53 (2 H, dd, J 7.8, 1.8, ArH),
7.61 (2 H, dd, J 7.8, 4.8, ArH); δC (100 MHz; CDCl3) 178.6 (C),
156.4 (CH), 152.9 (C), 137.3 (CH), 128.0 (C), 125.6 (CH); m/z
(ESI) 233 (M + Na+, 89%). Data are consistent with those
reported in literature.39
imidazo[4,5-f][1,10]phenanthroline 28 (200 mg, 0.67 mmol)
and iodomethane (84 µL, 1.35 mmol) in DMF (7 mL). Purifi-
cation by flash chromatography, eluting with dichloro-
methane–methanol (19 : 1), and recrystallization from
dichloromethane–methanol (1 : 1) gave the title compound as a
pale orange solid (43 mg, 21%); mp 246–247 °C; (Found:
M
+ +
H+, 311.1292. C20H14N4 H+ requires 311.1291);
λmax (MeOH)/nm 315 (log ε 5.08), 410 (4.49); νmax (CHCl3)/cm−1
3012, 2971, 1602, 1524, 1474, 1454, 1424, 1239, 924, 850;
δH (400 MHz; DMSO-d6) 9.68 (1H, dd, J 8.2, 1.2, ArH), 9.47
(1 H, d, J 6.0, ArH), 9.20 (1 H, dd, J 4.4, 1.8, ArH), 9.08 (1 H, dd,
J 8.2, 1.8, ArH), 8.43 (1 H, t, J 6.8, ArH), 8.26 (2 H, dt, J 6.8, 1.2,
ArH), 8.09 (1 H, dd, J 8.2, 4.4, ArH), 7.66 (2 H, t, J 6.8, ArH),
7.60 (1 H, tt, J 6.8, 1.2, ArH), 5.26 (3 H, s, Me); δC (100 MHz;
DMSO-d6) 152.7 (C), 149.3 (CH), 147.1 (CH), 139.2 (CH), 138.2
(2 × C), 134.8 (4 × C), 130.7 (CH), 130.5 (CH), 129.2 (CH), 129.1
(C), 126.5 (CH), 125.0 (CH), 124.4 (CH), 54.5 (Me); m/z (ESI)
311 (M + H+, 100%).
2-Phenyl-1H-imidazo[4,5-f][1,10]phenanthroline 28. Prepared
by general procedure 1 from 1,10-phenanthroline-5,6-dione 27
(150 mg, 0.71 mmol) and benzaldehyde (79 μL, 0.79 mmol).
Purification by trituration with ether (10 mL) followed by
dichloromethane (10 mL) gave the title compound as a pale
yellow solid (92 mg, 44%); mp 390–392 °C (decomp.) (lit.,40 mp
380 °C); (Found: M + H+, 297.1133. C19H12N4 + H+ requires
297.1135); νmax (CHCl3)/cm−1 3691, 3454, 3063, 2964, 1602,
1563, 1545, 1476, 1455, 1398, 1350, 1067, 1028; δH (400 MHz;
DMSO-d6) 13.77 (1 H, s, NH), 9.05 (2 H, dd, J 4.2, 1.4, ArH),
8.94 (2 H, dd, J 8.0, 1.4, ArH), 8.30 (2 H, d, J 8.0, ArH),
7.86–7.83 (2 H, m, ArH), 7.63 (2 H, t, J 8.0, ArH), 7.53 (1 H, t,
J 7.2, ArH); δC (100 MHz; DMSO-d6) 150.8 (C), 148.1 (CH),
143.6 (2 × C), 130.1 (2 × C), 129.9 (2 × CH), 129.2 (CH),
126.4 (CH), 123.5 (CH); m/z (ESI) 297 (M + H+, 100%).
1-Benzyl-2-(3,5-bis(trifluoromethyl)phenyl)-1H-imidazo[4,5-f]-
[1,10]phenanthroline 31. 1,10-Phenanthroline-5,6-dione 27
(200 mg, 0.95 mmol) was suspended in glacial acetic acid
(2 mL) and ammonium acetate (147 mg, 1.90 mmol), 3,5-bis
(trifluoromethyl)benzaldehyde (188 μL, 1.14 mmol) and ben-
zylamine (125 μL, 1.14 mmol) were added. The mixture was
heated at 100 °C for 18 h, cooled to room temperature and
added slowly to ammonium hydroxide (35%; 15 mL). The solu-
2-(3,5-Bis(Trifluoromethyl)phenyl)-1H-imidazo[4,5-f][1,10]- tion was extracted with dichloromethane (3 × 15 mL) and the
phenanthroline 29. Prepared by general procedure 1 from 1,10- combined organic extracts dried (Na2SO4) and concentrated
phenanthroline-5,6-dione 27 (500 mg, 2.38 mmol) and 3,5-bis under reduced pressure. Recrystallization from cyclohexane–
(trifluoromethyl)benzaldehyde (470 μL, 2.85 mmol). Purifi- ethyl acetate (1 : 1) followed by trituration with ether (10 mL)
cation by recrystallization from cyclohexane–ethyl acetate (1 : 2) gave the title compound as a colourless solid (93 mg, 19% yield,
gave the title compound as a cream solid (356 mg, 35% yield, 91% purity); mp 234–235 °C; (Found: M + H+, 523.1354.
91% purity); mp 236–237 °C; (Found: M + H+, 433.0900. C28H16F6N4 + H+ requires 523.1352); νmax (CHCl3)/cm−1 2962,
C21H10F6N4 + H+ requires 433.0882); νmax (CHCl3)/cm−1 3115, 1603, 1565, 1499, 1455, 1397, 1352, 1280, 1186, 1145, 1056,
2972, 1603, 1568, 1509, 1402, 1367, 1280, 1184, 1142, 1071, 1027, 906, 848; δH (400 MHz; DMSO-d6) 9.08 (1 H, dd, J 4.2,
981, 901; δH (400 MHz; acetone-d6) 8.95 (4 H, br s, ArH), 8.89 1.6, ArH), 8.99–8.97 (2 H, m, ArH), 8.48 (1 H, dd, J 8.4, 1.6,
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ArH), 8.33 (1 H, s, ArH), 8.31 (2 H, s, ArH), 7.84 (1 H, dd, J 8.0, M + H+, 493.1360. C25H18F6N2O2 + H+ requires 493.1345);
4.2, ArH), 7.65 (1 H, dd, J 8.4, 4.2, ArH), 7.39–7.31 (3 H, m, νmax (CHCl3)/cm−1 3441, 3006, 2963, 2839, 1615, 1519, 1490,
ArH), 7.21 (2 H, d, J 7.6, ArH), 7.21 (2 H, d, J 7.6, ArH), 6.00 1356, 1280, 1250, 1182, 1143, 1033, 836; δH (400 MHz; DMSO-
(2 H, s, CH2); δC (100 MHz; DMSO-d6) 150.7 (C), 148.8 (CH), d6) 13.01 (1 H, br s, NH), 8.68 (2 H, s, ArH), 8.06 (1 H, br s,
147.8 (CH), 144.2 (C), 143.9 (C), 136.2 (C), 135.9 (C), 132.1 (C), ArH), 7.50–7.46 (4 H, m, ArH), 7.04 (2 H, d, J 8.0, ArH), 6.90
130.9 (q, J 33, C), 129.8 (2 × CH), 129.3 (CH), 129.0 (CH), (2 H, d, J 8.0, ArH), 3.80 (3 H, s, Me), 3.76 (3 H, s, Me);
127.9 (CH), 126.2 (C), 125.6 (CH), 123.8 (CH), 123.5 (CH), δC (100 MHz; DMSO-d6) 159.1 (C), 158.2 (C), 141.8 (C),
123.3 (C), 123.0 (q, J 272, CF3), 122.9 (CH), 119.2 (C), 137.3 (C), 132.7 (C), 130.9 (q, J 32, C), 129.7 (CH), 128.7 (C),
50.2 (CH2); m/z (ESI) 523 (M + H+, 100%).
128.3 (CH), 127.2 (C), 124.9 (CH), 123.3 (q, J 270, CF3),
122.9 (C), 120.7 (CH), 114.2 (CH), 113.7 (CH), 55.2 (Me),
55.0 (Me); m/z (ESI) 493 (M + H+, 100%).
(b)
1-(4-Carboxamidobenzyl)-2-(3,5-bis(trifluoromethyl)-
phenyl)-4,5-di(4-methoxyphenyl)-1H-imidazole (apoptozole) 6
was prepared by general procedure 2 from 2-(3,5-bis(trifluoro-
methyl)phenyl)-4,5-di(4-methoxyphenyl)-1H-imidazole (200 mg,
0.41 mmol) and 4-(bromomethyl)benzamide 17 (130 mg,
0.61 mmol) in THF (4 mL). Purification by flash chromato-
graphy, eluting with light petroleum–ethyl acetate (2 : 3) gave
the title compound as a colourless solid (143 mg, 56%); mp
191–192 °C; (Found: M + H+, 626.1851. C33H25F6N3O3 + H+
requires 626.1873); νmax (CHCl3)/cm−1 3531, 3415, 3003, 2964,
2840, 1680, 1615, 1588, 1520, 1491, 1358, 1280, 1250, 1183,
1143, 1033, 903, 836; δH (400 MHz; CD3OD) 8.17 (2 H, s, ArH),
8.01 (1 H, s, ArH), 7.75 (2 H, d, J 8.4, ArH), 7.42 (2 H, d, J 8.8,
ArH), 7.26 (2 H, d, J 8.8, ArH), 6.97 (2 H, d, J 8.8, ArH), 6.95
(2 H, d, J 8.4, ArH), 6.82 (2 H, d, J 8.8, ArH), 5.27 (2 H, s, CH2),
3.81 (3 H, s, Me), 3.76 (3 H, s, Me); δC (100 MHz; CD3OD)
171.6 (C), 162.1 (C), 160.5 (C), 146.0 (C), 142.5 (C), 140.2 (C),
134.5 (C), 134.4 (C), 133.7 (CH), 133.3 (q, J 33, C), 132.5 (C),
130.4 (CH), 129.7 (CH), 129.4 (CH), 127.7 (C), 127.3 (CH),
124.7 (q, J 270, CF3), 123.8 (CH), 123.2 (C), 115.8 (CH),
114.9 (CH), 56.0 (Me), 55.8 (Me), 49.4 (CH2); m/z (ESI) 626
(M + H+, 100%). Data are consistent with those published in
literature.6
1-(4-Carboxamidobenzyl)-2-(3,5-bis(trifluoromethyl)phenyl)-
1H-imidazo[4,5-f][1,10]-phenanthroline 32. Prepared by general
procedure 2b from 2-(3,5-bis(trifluoromethyl)phenyl)-1H-
imidazo[4,5-f][1,10]phenanthroline 29 (200 mg, 0.46 mmol)
and 4-(bromomethyl)benzamide 17 (119 mg, 0.56 mmol). Puri-
fication by flash chromatography, eluting with dichloro-
methane–methanol (9 : 1) and recrystallization from methanol
gave the title compound as a colourless solid (47 mg, 18% yield,
86% purity); mp 183–184 °C; (Found: M + H+, 566.1413.
C29H17F6N5O + H+ requires 566.1410); νmax (CHCl3)/cm−1 2964,
1682, 1590, 1398, 1352, 1280, 1266, 1186, 1146, 906;
δH (400 MHz; DMSO-d6) 9.08 (1 H, dd, J 4.4, 1.8, ArH), 8.99
(1 H, t, J 1.8, ArH), 8.98 (1 H, t, J 1.8, ArH), 8.45 (1 H, dd, J 8.4,
1.4, ArH), 8.33 (3 H, br s, ArH), 7.96 (1 H, br s, NH), 7.87–7.82
(3 H, m, ArH), 7.64 (1 H, dd, J 8.4, 4.4, ArH), 7.37 (1 H, br s,
NH), 7.30 (2 H, d, J 8.4, ArH), 6.05 (2 H, s, CH2); δC (100 MHz;
DMSO-d6) 167.3 (C), 150.8 (C), 148.8 (CH), 147.8 (CH), 144.2
(C), 143.9 (C), 139.4 (C), 136.0 (C), 133.7 (C), 132.0 (C), 130.9
(q, J 33, C), 129.9 (CH), 129.8 (CH), 129.0 (CH), 128.4 (CH),
126.1 (C), 125.6 (CH), 123.8 (CH), 123.6 (CH), 123.3 (C), 123.0
(q, J 272, CF3), 122.9 (CH), 119.1 (C), 50.1 (CH2); m/z (ESI) 566
(M + H+, 57%).
Biology
Cell lines. The human cancer cell lines HCT116 (colon),
WM266.4 (melanoma) and MCF7 (breast) were obtained from
the American Type Culture Collection (LGC Promochem, UK).
Cells were grown in DMEM/10% FCS, 2 mM glutamine and
non-essential amino acids in 5% CO2. Cells were free of Myco-
plasma contamination (Venor GeM kit, Minerva Biolabs,
Germany). HCT116 cells have been used in previous studies on
the Hsp70 pathway.26
In vitro cell viability assay. The CellTiter-Blue® viability
assay (Promega, USA) provides a homogenous, fluorometric
1-(4-Carboxamidobenzyl)-2-(3,5-bis(trifluoromethyl)phenyl)- method for estimating the number of viable cells. It uses the
4,5-di(4-methoxyphenyl)-1H-imidazole (apoptozole) 6. (a) 2-(3,5- dark blue indicator dye resazurin to measure the metabolic
Bis(trifluoromethyl)phenyl)-4,5-di(4-methoxyphenyl)-1H-imid- capacity of cells that is an indicator of cell viability. Viable cells
azole was prepared by general procedure 1 from 4,4′-dimethoxy- are able to reduce resazurin into resorufin (pink) that is highly
benzil (500 mg, 1.85 mmol) and 3,5-bis(trifluoromethyl)- fluorescent. Briefly, cells (6 × 103 cells per mL) were seeded
benzaldehyde (335 μL, 2.03 mmol). Purification by flash into 384-well plates and were incubated for 24 h. Compounds
chromatography, eluting with light petroleum–ethyl acetate (at a range of concentrations) were added using the ECHO
(4 : 1), followed by trituration with ether (20 mL) gave 2-(3,5-bis- liquid handler (Labcyte, USA) and then left at 37 °C for 96 h.
(trifluoromethyl)phenyl)-4,5-di(4-methoxyphenyl)-1H-imidazole Titer blue reagent was added to each well and left at 37 °C for
as a cream solid (641 mg, 70%); mp 219–220 °C; (Found: 3–4 h. Fluorescence was measured using the Envision Multi-
Org. Biomol. Chem.
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Organic & Biomolecular Chemistry
Paper
label Plate Reader (Perkin Elmer, UK). The GI50 was deter- 14 M. Daugaard, M. Rohde and M. Jaattela, FEBS Lett., 2007,
mined as the concentration required to cause 50% inhibition,
and data are reported as the mean SE of three replicates.
Western blotting. HCT116 cells were treated with either 5 ×
581, 3702–3710.
15 M. V. Powers, P. A. Clarke and P. Workman, Cell Cycle,
2009, 8, 518–526.
GI50 of compounds for 8, 24 and 48 h or multiples of GI50 for 16 S. Patury, Y. Miyata and J. E. Gestwicki, Curr. Top. Med.
24 h. Cell lysates were prepared and proteins were then immuno-
Chem., 2009, 9, 1337–1351.
blotted and detected by enhanced chemiluminescence.41 Anti- 17 A. J. Massey, J. Med. Chem., 2010, 53, 7280–7286.
bodies to Hsps were from Stressgen and those for ERBB2, CRAF, 18 M. V. Powers, K. Jones, C. Barillari, I. Westwood,
CDK4, cleaved PARP and GAPDH were from Abcam, BD Bio-
science, Santa Cruz, Cell Signaling and Chemicon, respectively.
R. L. M. van Montfort and P. Workman, Cell Cycle, 2010, 9,
1542–1550.
19 C. G. Evans, L. Chang and J. E. Gestwicki, J. Med. Chem.,
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20 A. R. Goloudina, O. N. Demidov and C. Garrido, Cancer
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Acknowledgements
We thank Marloes Technologies Ltd for support of this work 21 J. R. McConnell and S. R. McAlpine, Bioorg. Med. Chem.
and the University of Nottingham for funding (AP), Carroll Uni- Lett., 2013, 23, 1923–1928.
versity for funding (KH). PW and SYS were supported by a pro- 22 H. Debus, Liebigs Ann. Chem., 1858, 107, 199–208.
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is a Cancer Research UK Life Fellow. The authors declare no 24 S. E. Wolkenberg, D. D. Wisnoski, W. H. Leister, Y. Wang,
competing financial interests, although The Institute of Cancer
Z. J. Zhao and C. W. Lindsley, Org. Lett., 2004, 6, 1453–1456.
Research, London, has a commercial interest in Hsp90 inhibi- 25 B. N. Boden, K. J. Jardine, A. C. W. Leung and
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M. J. MacLachlan, Org. Lett., 2006, 8, 1855–1858.
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