Synthesis of farnesol analogues containing triazoles in place of isoprenes through 'click chemistry'

Article abstract of DOI:10.1055/s-0031-1290461

A solid-phase three-component Huisgen reaction has been used to generate polar farnesol and farnesyl diphosphate analogues. The Cu(I)-catalyzed 1,3-cycloadditions of various azides with solid supported (E)-3-methylhept-2-en- 6-yn-1-ol provided only the 1,4-disubstituted 1,2,3-triazole regioisomers. The organic azides were generated in situ to minimize handling of potentially explosive azides. We have employed this powerful click chemistry to make farnesol analogues where both β- and γ-isoprenes were replaced by triazole and substituted aromatic rings, respectively.

Full text of DOI:10.1055/s-0031-1290461

LETTER
▌2539
lSetyternthesis of Farnesol Analogues Containing Triazoles in Place of Isoprenes
through ‘Click Chemistry’
Synthesis of Farnesol Analogues
Thangaiah Subramanian,a Sean Parkin,b H. Peter Spielmann*a–d
a
Department of Cellular and Molecular Biochemistry, University of Kentucky, Lexington, KY, 40536, USA
E-mail: hps@uky.edu
b
Department of Chemistry, University of Kentucky, Lexington, KY, 40536, USA
c
Center for Structural Biology, University of Kentucky, Lexington, KY, 40536, USA
d
Markey Cancer Center, University of Kentucky, Lexington, KY, 40536, USA
Received: 12.07.2012; Accepted after revision: 17.08.2012
cilitates the removal of leftover reagents and soluble
Abstract: A solid-phase three-component Huisgen reaction has
byproducts.16 Cu(I)-catalyzed 1,3-cycloaddition of azides
and alkynes to form 1,4-triazoles is a robust way of pre-
been used to generate polar farnesol and farnesyl diphosphate ana-
logues. The Cu(I)-catalyzed 1,3-cycloadditions of various azides
with solid supported (E)-3-methylhept-2-en-6-yn-1-ol provided
paring small-molecule combinatorial libraries by
only the 1,4-disubstituted 1,2,3-triazole regioisomers. The organic SPOS.17–20 SPOS has been employed to prepare libraries
azides were generated in situ to minimize handling of potentially
of 1,4-triazoles where either the azide or alkyne is at-
tached to the solid support.21–30 Utilizing alkyne-function-
explosive azides. We have employed this powerful ‘click chemis-
try’ to make farnesol analogues where both β- and γ-isoprenes were
alized solid supports prevents formation of Cu(I)-
replaced by triazole and substituted aromatic rings, respectively.
catalyzed alkyne homocoupling side products. Recently,
Key words: farnesol, farnesyldiphosphate, cancer, cycloaddition,
triazoles, solid-phase synthesis, click chemistry
Gibbs et al. reported a library of S-farnesyl-thiopropionic
acid 1,4-triazoles prepared by solution-phase methods to
study their inhibitory activity against isoprenylcysteine
methyltransferase (ICMT).31 Herein, we report a concise
Farnesyl diphosphate (1, FPP) analogues have been used
SPOS of FPP analogues where the β-isoprene and γ-iso-
to study the mechanism of protein farnesyltransferase
prene were replaced by 1,4-triazole and substituted aryl
(FTase) and to examine the effect of analogue structure on
groups, respectively. The substitutions on the aryl rings
prenylated protein function.1–6 FTase catalyzes transfer of
were chosen to either increase or decrease the hydrophi-
the isoprenoid to the cysteine residue of proteins with C-
licity of the analogues (Scheme 1, Table 2).
terminal CAAX motifs, including the oncoprotein Ras.7,8
Activating mutations in Ras are found in 30% of all hu-
γ
β
α
man cancers, and Ras prenylation is obligatory for Ras
function.9–11 Ras function can be inhibited by FTase-cata-
lyzed transfer of FPP analogues with reduced hydropho-
bicity, such as 2a,b and 3.12 The FTase-catalyzed transfer
of FPP analogues depends on both analogue structure and
CAAX sequence.13 However, there is no correlation be-
tween FPP analogue hydrophobicity and FTase-catalyzed
isoprenoid transfer to protein. We have previously shown
that an aniline or phenoxy group is isosteric with the ter-
minal isoprene of FPP and that analogues with a range of
substituents on the aryl group are FTase-transferable sub-
strates.14 These observations open the door to FTase-
transferable FPP analogues that act as prenyl-function in-
hibitors (PFI) by interfering with a prenylated protein’s
function.12
OPP
1 farnesylpyrophosphate (FPP)
H
N
OPP
OPP
R
2a R = 4-NO2, 2b R = 4-CN
H
N
N
O
3
N
N
N
R1
OR2
We have previously used solid-phase organic synthesis
(SPOS) to generate a library of FTase-transferrable FPP
analogues where the γ-isoprene was replaced by substitut-
ed aryl groups14,15 (Figure 1). SPOS is particularly useful
for many synthetic transformations because excess re-
agents can be used to drive reactions to completion and fa-
4a–k R2 = H, for R1 see Table 2
3–
5 R1 = H, R2 = P2O6
3–
PP = P2O6
Figure 1 Structure of FPP analogues
Treatment of previously reported1 alcohol 6 with Ph3PBr2
and Hünig’s base (DIPEA) gave the pure E-isomer of al-
lylic bromide 7. Propargyl Grignard prepared with a cata-
lytic amount of HgCl was coupled to bromide 7 to give
SYNLETT 2012, 23, 2539–2543
Advanced online publication: 28.09.2012
DOI: 10.1055/s-0031-1290461; Art ID: ST-2012-S0587-L
© Georg Thieme Verlag Stuttgart · New York
0
9
3
6
-
5
2
1
4
1
4
3
7
-
2
0
9
6

2540
T. Subramanian et al.
LETTER
alkyne 8 in 72% yield. Treatment of alkyne 8 with PPTS
in MeOH furnished alcohol 9 in quantitative yield.32 Cou-
pling alcohol 9 to DHP-resin 10 generated the alkyne-
functionalized resin 11 with a traceless, acid-labile THP
linker.15
Figure 2 Structure and ellipsoid plot of compound 4l41
OTHP
HO
6
Table 1 Conditions Attempted for Conversion of 7a into 8a Using
Ph3PBr2
DIPEA
CuI as Catalyst
CH2Cl2
Entry
Solvent system
Yield (%)a
OTHP
Br
1
2
3
4
5
DMF–t-BuOH (3:1), DIPEA
DMF–t-BuOH–H2O (3:1:1), DIPEA
DMF–t-BuOH (1:1), DIPEA
DMF, DIPEA
28
0
7
MgBr
Et2O
H
32
53
42
OTHP
8
DMF, TMED
PPTS, MeOH
a Yields were calculated after cleaving the resin (solvent MeOH–DCE
= 1:1).
H
OH
9
DHP resin
PPTS, DCE
10
Heating the resin in MeOH–DCE (1:1) with catalytic
PPTS over night provided analogues 4a–k in moderate
yield (Scheme 2, Table 2). Prolonged heating of the cleav-
age reactions resulted in side products formed by 1,3-mi-
gration of the hydroxyl group. Exclusive formation of the
1,4-disubstituted 1,2,3-triazole regioisomer was con-
H
O
11
1
firmed by H-NOE experiments, as expected for the
Scheme 1 Synthesis of resin-linked alkyne 11
Cu(I)-catalyzed Huisgen reaction.39
Triazole diphosphate 5 (19% yield in three steps) was pre-
pared by Ph3PBr2 cleavage of THP resin 12a, followed by
trapping the resulting allylic bromide with 10 equivalents
of tris(tetra-n-butylammonium)hydrogen diphosphate.
The crude diphosphate was purified by ion-exchange
chromatography followed by RP-HPLC. All compounds
Resin loading of 82% was determined by recovering alco-
hol 9 from resin 11 treated with PPTS in MeOH and DCE.
The key reaction was the 1,4-disubstituted 1,2,3-triazole
formation between the resin-linked alkyne and various
benzyl azides to allow introduction of diversity in the ter-
minal aryl group.18,33,34 The required benzyl azides were
prepared in situ from commercially available bromides
and NaN3 in DMF. CAUTION! Due to risk of explo-
sion,35–38 it is crucially important not to use halogenated
solvents at this stage and to ensure that all azide ion is re-
moved by thorough washing of the resin before introduc-
ing halogenated solvents in any subsequent steps.
H
O
11
RBr, NaN3
Cu(I)
DIPEA
DMF
Agitating resin 11 with catalytic Cu(I)I and 10 equivalents
each of BnBr, NaN3, and Hünig’s base in DMF provided
triazole resin 7a–k. A large excess of CuI (>5 equiv) leads
to the formation of a side product where iodine is incorpo-
rated into the triazole ring 4l. The structure of (E)-5-(1-
benzyl-5-iodo-1H-1,2,3-triazol-4-yl)-3-methylpent-2-en-
1-ol (4l) was confirmed by X-ray crystallography (Figure
2).
N
N
R
N
O
12a–k
PPTS
(i) Ph3PBr2
MeOH–DCE
MeCN
(ii) [(n-Bu)4N]3HP2O7
N
N
N
R1
The in situ generation of Cu(I) from CuSO4 and ascorbic
acid in the presence of water inhibited resin swelling and
reduced the yield of triazole to almost nil. Triazole yield
was also significantly reduced when other protic and non-
protic solvent mixtures drawn from the literature were
used. Optimum triazole yields were obtained using
Hünig’s base and DMF as solvent (Table 1).
OR2
4a–k R2 = H, for R1 see Table 2
3–
5 R1 = H, R2 = P2O6
Scheme 2 Synthesis of Triazole-Containing Farnesols 4a–k and di-
phosphate 5
Synlett 2012, 23, 2539–2543
© Georg Thieme Verlag Stuttgart · New York

LETTER
Synthesis of Farnesol Analogues
2541
in Table 1 were fully characterized by 1H NMR spectros-
copy and low- and high-resolution mass spectrometry.40
Acknowledgment
This work was supported in part by the National Institutes of Health
GM66152 (to H.P.S.) and the Kentucky Lung Cancer Research Pro-
gram (to H.P.S). The NMR instruments used in this work were ob-
tained with support from NSF CRIF Grant CHE- 9974810. We
acknowledge University of Kentucky Mass Spectroscopy Facility
for mass data.
In summary, we have developed a short synthetic route to
triazole-containing farnesol analogues and their diphos-
phates through solid-phase ‘click chemistry’. The bio-
chemical characterization of these compounds is ongoing
and will be published elsewhere.
Table 2 1,2,3-Triazole-Containing Farnesol Analogues
Entry
1
Compound
Structure
Yield (%)a
53
Purity LC–MS (%)b
N
N
N
4a
96
OH
N
N
N
N
N
N
N
2
3
4b
4c
4d
4e
4f
55
43
36
26
57
41
32
28
38
47
89
96
88c
94c
90
94
98
98
94
98
OH
OH
OH
O2N
N
N
N
4
N
N
N
5
OH
F
F
N
N
6
OH
N
N
N
7
4g
4h
4i
OH
F
N
F3CO
N
N
N
8
OH
OH
N
N
9
CF3O
NC
N
N
N
OH
10
11
4j
N
N
N
4k
OH
NC
a Yield (%) for 4a–k in two steps from 11. Dehydrated side products account for the bulk of the balance except for 4d and 4e.
b The percentage was calculated by integrating HPLC chromatogram peaks at 230 nm.
c Unidentified oxidized compounds [M + 16] account for the balance.
© Georg Thieme Verlag Stuttgart · New York
Synlett 2012, 23, 2539–2543

2542
T. Subramanian et al.
LETTER
(31) Bergman, J. A.; Hahne, K.; Song, J.; Hrycyna, C. A.; Gibbs,
R. A. ACS Med. Chem. Lett. 2012, 3, 15.
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Supporting Information for this article is available online at
http://www.thieme-connect.com/ejournals/toc/synlett. Included are
the detailed experimental procedure and spectroscopic data of 4a–k
1
and 5; 1H spectra and HRMS data sheet of 4a–k; and H, 31P, and
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Chem. Soc. 2010, 132, 14570.
LRMS spectra of 5.SuInopigfmot
riratSuInoopigf
n
m
iortartn
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(40) Typical Procedure for Cycloaddition Adducts 12a–k
Reaction vessels were charged with resin 11 (100 mg) and
NaN3 (57.6 mg, 0.99 mmol, 10 equiv) followed by DMF (5
mL) and then CuI (3 mg) and agitate was initiated. Hünig’s
base (114 mg, 0.99 mmol, 10 equiv) was added, and, finally,
the bromide (0.99 mmol, 10 equiv) was added. The resultant
mixture was agitated for 1 d at r.t., heated to 50 °C for 3 h,
then cooled. Then the resin was filtered and washed with
DMF (2×), DMF–H2O (1:1, 2×), DMF (2×), CH2Cl2 (3×),
Et2O (2×) and dried in vacuo.
References and Notes
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General Procedure for the Synthesis of Alcohols 4a–k
Resin (100 mg) was heated at 80 °C with DCE–anhyd
MeOH (1:1, 8 mL) in the presence of PPTS (5 mg) for 8 h.
The liquid phase was collected, the resin was rinsed with
CH2Cl2, and the washings were combined and sequentially
washed with sat. aq NaHCO3, H2O, brine, dried over
MgSO4, concentrated, and purified by silica gel column
chromatography to give 4a–k.
Compound 4a: 1H NMR (400 MHz, CDCl3): δ = 1.68 (s, 3
H), 2.35 (t, J = 8.4 Hz, 2 H), 2.81–2.85 (m, 2 H), 4.10 (d,
J = 7.6 Hz, 2 H), 5.34–5.38 (m, 1 H), 5.48 (s, 2 H), 7.20 (s,
1 H), 7.21–7.27 (m, 2 H), 7.32–7.40 (m, 3 H) ppm. LRMS
[M + H+]: m/z = 258.2. HRMS [M+]: m/z calcd for
C15H19N3O: 257.1528; found: 257.1529.
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Sorba, G.; Genazzani, A. A. Med. Res. Rev. 2008, 28, 278.
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K. B. Angew. Chem. Int. Ed. Engl. 2002, 41, 2596.
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2003, 5, 1753.
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Med. Chem. Commun. 2012, 3, 506.
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G. W. Bioorg. Med. Chem. 2012, 20, 2638.
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Compound 4b: 1H NMR (400 MHz, CDCl3): δ = 1.63 (s, 3
H), 2.31 (t, J = 8.0 Hz, 2 H), 2.81 (t, J = 8.0 Hz, 2 H), 4.10
(d, J = 6.8 Hz, 2 H), 5.33 (t, J = 6.8 Hz, 1 H), 5.58 (s, 2 H),
7.32–7.35 (m, 3 H), 8.14 (d, J = 10.8 Hz, 2 H) ppm. LRMS
[M + H+]: m/z = 302.2. HRMS [M+]: m/z calcd for
C15H18N4O3: 302.1379; found: 302.1375.
Compound 4c: 1H NMR (400 MHz, CDCl3): δ = 1.29 (s, 3
H), 1.66 (s, 3 H), 2.33 (t, J = 8.0 Hz, 2 H), 2.80 (t, J = 7.6 Hz,
2 H), 4.10 (d, J = 6.8 Hz, 2 H), 5.37 (t, J = 6.8 Hz, 1 H), 5.43
(s, 2 H), 7.16–7.20 (m, 3 H), 7.36 (d, J = 8.4 Hz, 2 H) ppm.
LRMS [M + H+]: m/z = 314.2. HRMS [M+]: m/z calcd for
C19H27N3O: 313.2154; found: 313.2154.
Compound 4d: 1H NMR (400 MHz, CDCl3): δ = 1.23 (d,
J = 6.8 Hz, 6 H), 1.69 (s, 3 H), 2.35 (t, J = 8.4 Hz, 2 H), 2.83
(t, J = 8.4 Hz, 2 H), 2.87–2.94 (m, 1 H), 4.10 (d, J = 6.8 Hz,
2 H), 5.30 (t, J = 8.0 Hz, 1 H), 5.44 (s, 2 H), 7.16–7.26 (m, 4
H) ppm. LRMS [M + H+]: m/z = 300.2. HRMS [M+]: m/z
calcd for C18H25N3O: 299.1998; found: 299.2002.
Compound 4e: 1H NMR (400 MHz, CDCl3): δ = 1.69 (s, 3
H), 2.35 (t, J = 8.0 Hz, 2 H), 2.83 (t, J = 8.0 Hz, 2 H), 4.11
(d, J = 6.8 Hz, 2 H), 5.38 (t, J = 8.0 Hz, 1 H), 5.45 (s, 2 H),
7.02–7.09 (m, 2 H), 7.21–7.26 (m, 3 H) ppm. LRMS [M +
H+]: m/z = 276.2. HRMS [M+]: m/z calcd for C15H18FN3O:
275.1434; found: 275.1431.
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Compound 4f: 1H NMR (400 MHz, CDCl3): δ = 1.69 (s, 3
Synlett 2012, 23, 2539–2543
© Georg Thieme Verlag Stuttgart · New York

LETTER
H), 2.36 (t, J = 8.0 Hz, 2 H), 2.84 (t, J = 8.0 Hz, 2 H), 4.11
Synthesis of Farnesol Analogues
2543
(d, J = 12.0 Hz, 2 H) ppm. LRMS [M + H+]: m/z = 283.2.
HRMS [M+]: m/z calcd for C16H18N4O: 282.1480; found:
282.1487.
(d, J = 6.8 Hz, 2 H), 5.38 (t, J = 8.0 Hz, 1 H), 5.48 (s, 3 H),
6.91 (d, J = 9.2 Hz, 1 H), 7.02–7.05 (m, 2 H), 7.30–7.35 (m,
1 H) ppm. LRMS [M + H+]: m/z = 276.2. HRMS [M+]: m/z
calcd for C15H18FN3O: 275.1434; found: 275.1437.
Compound 4g: 1H NMR (400 MHz, CDCl3): δ = 1.69 (s, 3
H), 2.36 (t, J = 8.0 Hz, 2 H), 2.83 (t, J = 8.0 Hz, 2 H), 4.11
(d, J = 6.8 Hz, 2 H), 5.38 (t, J = 8.0 Hz, 1 H), 5.54 (s, 2 H),
7.08–7.37 (m, 4 H) ppm. LRMS [M + H+]: m/z = 276.2.
HRMS [M+]: m/z calcd for C15H18FN3O: 275.1434; found:
275.1429.
Procedure for the Synthesis of (E)-5-(1-Benzyl-1H-1,2,3-
triazol-4-yl)-3-methylpent-2-en-1-yl Diphosphate (5)
The vacuum-dried resin 12a (100 mg, 0.09 mmol) was
suspended in CH2Cl2 (2 mL), Ph3PBr2 (84 mg, 0.19 mmol)
was added, and the slurry was stirred at r.t. under N2 for 4 h.
Tris[tetra(n-butyl)ammonium]hydrogen diphosphate (388
mg, 0.39 mmol) in anhyd MeCN (3 mL) was added and the
reaction mixture was stirred at r.t. for 4 h under N2. The
heterogeneous filtrate was concentrated, and the residue was
suspended in 25 mM NH4HCO3 (4 mL) and extracted with
Compound 4h: 1H NMR (400 MHz, CDCl3): δ = 1.71 (s, 3
H), 2.38 (t, J = 8.0 Hz, 2 H), 2.86 (t, J = 8.0 Hz, 2 H), 4.12
(t, J = 6.8 Hz, 2 H), 5.39 (t, J = 8.0 Hz, 1 H), 5.52 (s, 2 H),
7.10–7.25 (m, 4 H), 7.40 (t, J = 8.0 Hz, 1 H) ppm. LRMS [M
+ H+]: m/z = 341.2. HRMS [M+]: m/z calcd for
+
Et2O (5 mL, 3×). The aqueous phase was applied to an NH4
form DOWEX-AW-50 ion-exchange column (50 mL of
resin) and eluted with four-column volume 25 mM
NH4HCO3 buffer. The aqueous phase was lyophilized to
obtain the crude diphosphate 5 as a white solid, which was
dissolved in a minimum volume of 25 mM NH5CO3 buffer
and purified by RP-HPLC (Varian Dynamax, 10 μm, 300 Å,
C-4 (10 mm × 250 mm) column with a gradient mobile
phase: 25 mM NH5AcO and MeCN.15 The product collected
between 5.2–5.6 min was lyophilized to obtain compound 5
(9.0 mg, 19% in 2 steps). 1H NMR (400 MHz, D2O): δ = 1.44
(s, 3 H), 2.10 (t, J = 7.6 Hz, 2 H), 2.57 (t, J = 7.6 Hz, 2 H),
4.15 (t, J = 6.8 Hz, 2 H), 5.11 (t, 6.4 Hz, 1 H), 5.28 (s, 2 H),
7.03–7.05 (m, 1 H), 7.14–7.20 (m, 2 H), 7.50 (s, 1 H) ppm.
31P NMR (D2O, 161.8 MHz): δ = –6.80 (d, J = 21.36 Hz, 1
P), –10.40 (d, J = 21.36 Hz, 1 P). LRMS [M + H+]: m/z =
418.0.
C16H18F3N3O2: 341.1350; found: 341.1359.
Compound 4i: 1H NMR (400 MHz, CDCl3): δ = 1.67 (s, 3
H), 2.35 (t, J = 8.0 Hz, 2 H), 2.83 (t, J = 8.0 Hz, 2 H), 4.11
(d, J = 6.8 Hz, 2 H), 5.39 (t, J = 8.0 Hz, 1 H), 5.52 (s, 2 H),
7.18–7.27 (m, 4 H) ppm. LRMS [M + H+]: m/z = 341.2.
HRMS [M+]: m/z calcd for C16H18F3N3O2: 341.1350; found:
341.1357.
Compound 4j: 1H NMR (400 MHz, CDCl3) δ = 1.69 (s, 3H),
2.36 (t, J = 8.0Hz, 2H), 2.85 (t, J = 8.0Hz, 2H), 4.11 (d,
J = 6.8 Hz, 2 H), 5.38 (t, J = 8.0 Hz, 1 H), 5.53 (s, 2 H), 7.29
(s, 1 H), 7.47–7.50 (m, 2 H), 7.61–7.63 (m, 1 H) ppm. LRMS
[M + H+]: m/z = 283.2. HRMS [M+]: m/z calcd for
C16H18N4O: 282.1480; found: 282.1482.
Compound 4k: 1H NMR (400 MHz, CDCl3): δ = 1.70 (s, 3
H), 2.38 (t, J = 8.0 Hz, 2 H), 2.87 (t, J = 8.0 Hz, 2 H), 4.12
(d, J = 6.8 Hz, 2 H), 5.38 (t, J = 8.0 Hz, 1 H), 5.56 (s, 2 H),
5.56 (s, 2 H), 7.26 (s, 1 H), 7.31 (d, J = 12.0 Hz, 2 H), 7.65
(41) X-ray crystal data of the structure 4l have been deposited at
the Cambridge Crystallographic Data Center with the
deposition number CCDC 871186.
© Georg Thieme Verlag Stuttgart · New York
Synlett 2012, 23, 2539–2543

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