General route for the preparation of diverse 17-membered macrocycles based on RCM and examination of the E/Z selectivity

Article abstract of DOI:10.1080/00397911.2011.570891

(Chemical Equation Presented) A convergent, general synthetic route to 17-membered macrocycles was developed to support biological evaluation and structure-activity relationship (SAR) studies during phenotypic screening for immunology targets. A series of amide coupling reactions led to a ring-closing metathesis (RCM) precursor that was cyclized using Grubbs' catalysts. It was found that the reaction formed the macrocyclic products in a 3:1 ratio of E/Z isomers. Moreover, it was shown that a number of similarly substituted RCM precursors undergo cyclization to produce the geometric E/Z isomers in roughly the same 3:1 ratio. The remarkable independence of the E/Z outcome from the substitution pattern of the RCM precursor makes this synthetic approach generally applicable. Separation of the E/Z isomers was achieved by preparative high-performance liquid chromatography and allowed biological profiling of the geometric isomers. Reactive groups in the macrocycle were utilized for late-stage modifications in the fashion of diversity-orientated synthesis (DOS), yielding analogs for SAR studies. Copyright Rigel Pharmaceuticals, Inc.

Full text of DOI:10.1080/00397911.2011.570891

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General Route for the Preparation of
Diverse 17-Membered Macrocycles Based
on RCM and Examination of the E/Z
Selectivity
Thilo J. Heckrodt a & Rajinder Singh a
a Department of Medicinal Chemistry , Rigel Pharmaceuticals , South
San Francisco , California , USA
Accepted author version posted online: 17 Nov 2011.Published
online: 29 May 2012.
To cite this article: Thilo J. Heckrodt & Rajinder Singh (2012) General Route for the Preparation of
Diverse 17-Membered Macrocycles Based on RCM and Examination of the E/Z Selectivity, Synthetic
Communications: An International Journal for Rapid Communication of Synthetic Organic Chemistry,
42:19, 2854-2865, DOI: 10.1080/00397911.2011.570891
To link to this article: http://dx.doi.org/10.1080/00397911.2011.570891
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Synthetic Communications1, 42: 2854–2865, 2012
Copyright # Rigel Pharmaceuticals, Inc.
ISSN: 0039-7911 print=1532-2432 online
DOI: 10.1080/00397911.2011.570891
GENERAL ROUTE FOR THE PREPARATION OF DIVERSE
17-MEMBERED MACROCYCLES BASED ON RCM AND
EXAMINATION OF THE E/Z SELECTIVITY
Thilo J. Heckrodt and Rajinder Singh
Department of Medicinal Chemistry, Rigel Pharmaceuticals, South San
Francisco, California, USA
GRAPHICAL ABSTRACT
Abstract A convergent, general synthetic route to 17-membered macrocycles was developed
to support biological evaluation and structure–activity relationship (SAR) studies during
phenotypic screening for immunology targets. A series of amide coupling reactions led to
a ring-closing metathesis (RCM) precursor that was cyclized using Grubbs’ catalysts. It
was found that the reaction formed the macrocyclic products in a 3:1 ratio of E/Z isomers.
Moreover, it was shown that a number of similarly substituted RCM precursors undergo
cyclization to produce the geometric E/Z isomers in roughly the same 3:1 ratio. The
remarkable independence of the E/Z outcome from the substitution pattern of the RCM
precursor makes this synthetic approach generally applicable. Separation of the E/Z iso-
mers was achieved by preparative high-performance liquid chromatography and allowed
biological profiling of the geometric isomers. Reactive groups in the macrocycle were uti-
lized for late-stage modifications in the fashion of diversity-orientated synthesis (DOS),
yielding analogs for SAR studies.
Keywords Diversity-orientated synthesis (DOS); E=Z isomers; Grubbs’ catalyst;
macrocycles; ring-closing metathesis (RCM)
INTRODUCTION
Drugs based on macrocyclic compounds play important roles in modern medi-
cine. The inherent features of macrocycles often make them particularly well-suited
as pharmaceuticals (e.g., increased stability, decreased conformational flexibility, low
number of rotatable bonds can favor high oral bioavailability). Current macrocyclic
Received February 15, 2011.
Address correspondence to Thilo J. Heckrodt, Department of Medicinal Chemistry, Rigel
Pharmaceuticals, Inc., 1180 Veterans Boulevard, South San Francisco, CA 94080, USA. E-mail:
theckrodt@rigel.com
2854

PREPARATION OF 17-MEMBERED MACROCYCLES
2855
Figure 1. Seventeen-membered macrocycle 1.
drugs are almost exclusively derived from natural sources and are either identical
(e.g., rapamycin) or closely related (e.g., temsirolimus) to naturally occurring macro-
cycles.[1] However, several synthetic macrocycles unrelated to natural products are
now under active preclinical and clinical development.[2] In particular, several macro-
cyclic hepatitis C virus (HCV) protease inhibitors have entered clinical trails.[2b–e]
During a high-throughput screening (HTS) campaign we became interested in
fully synthetic macrocycles that showed activity against immunology targets. To gain
access to larger quantities for biological evaluation and initial structure–activity
relationship (SAR) studies, we developed a synthetic route to this compound class.
Compound 1 is an example of the macrocycles that were targeted for further
biologic evaluation (Fig. 1). The scaffold is a 17-membered triamide featuring a sali-
cylic acid–based aromatic moiety on the left-hand side, an amino acid at the bottom,
a glutamic methyl ester on the right-hand side, and an olefinic linker at the top.
RESULTS AND DISCUSSION
This article describes our synthetic approach to 17-membered macrocycles uti-
lizing ring-closing metathesis and studies the E=Z selectivity of the ring-closing
metathesis (RCM) reaction for this system.
Retrosynthetic disconnection of the macrocyclic scaffold 2 leads to the diolefi-
nic RCM precursor 3 (Scheme 1). Intermediate 3 can be obtained by a sequence of
Scheme 1. Retrosynthetic analysis.

2856
T. J. HECKRODT AND R. SINGH
˚
Scheme 2. Synthesis of glutamic ester derivative 10. Reagents and conditions: (a) CsOH(H2O (0.1 eq), 4 A
MS, DMF, ꢁ20 ꢂC to rT, 12 h, 52%; (b) EDC (1.5 eq), HOBt (1.5 eq), Et3N (2.0 eq), DCM, rT, 10 h; (c)
HCl (4 M in dioxane), MeOH, rT, 1 d, 92% over two steps, (EDC ¼ N-ethyl-N0-(3-dimethylaminopropyl)-
carbodiimide, HOBt ¼ 1-hydroxybenzotriazole).
amidations first coupling glutamic acid derivative 6 to protected amino acid 5 and
subsequently forming an amide bond with salicylic acid derivative 4.
The synthesis of glutamic ester derivative 10 starts with the monoallylation of
phenylethylamine 7. This highly exothermic monoallylation reaction proved to be
capricious, and most reactions evaluated also gave significant amounts of the dially-
lated by-product. Using CsOH ꢀ H2O as base[3] in combination with low temperature
gave satisfactory yields of the desired monoallylated phenylethyl amine product after
column chromatography. Secondary amine 8 was then coupled with Boc-protected
(S)-glutamic amino methyl ester 9 under standard N-ethyl-N0-(3-dimethylaminopropyl)-
carbodiimide (EDC) conditions. Subsequent deprotection with 4 M HCl furnished
the desired intermediate 10 (Scheme 2).
Salicylic acid derivative 13 was obtained by a two-step sequence. Starting
material 11 was diallylated to give intermediate 12, which was subsequently hydro-
lyzed to the acid 13. This procedure turned out to be more efficient than selective
monoallylation with 1 equivalent of allyl bromide (Scheme 3).
The free primary amine of glutamic ester 10 was coupled with Boc-protected
(R)-phenylalanine and subsequently deprotected under acidic conditions to yield amide
14. Final assembly of the RCM precursor 15 was achieved using O-(7-azabenzo-triazol-
1-yl)-N,N,N0,N0-tetramethyluronium hexafluorophosphate (HATU) to connect salicylic
acid 13 with fragment 14. Although the employment of first-generation Grubbs’ catalyst
produced the desired macrocycle 1, it was found that faster conversion and greater yields
were obtained with the use of second-generation Grubbs’ catalyst.[4] Special addition
protocols such as slow addition via syringe pump were not required (Scheme 4).
Scheme 3. Monoallylation of salicylic acid 11. Reagents and conditions: (a) allyl bromide (4.0 eq), K2CO3
(3.0 eq), Cs2CO3 (0.1 eq), acetone, reflux, 8 h; (b) NaOH (1.2 eq) H2O=EtOH 1:1, rT, 10 h, 94% over two
steps.

PREPARATION OF 17-MEMBERED MACROCYCLES
2857
Scheme 4. Synthesis of the RCM precursor and ring closure. Reagents and conditions: (a) Boc-
(R)-phenylalanine (1.0 eq), EDC (1.5 eq), HOBt (1.5 eq), Et3N (2.0 eq), DCM, rT, 12 h; (b) HCl (4 M in
dioxane), MeOH, rT, 8 h, 71% over two steps; (c) 13 (1.0 eq), HATU (1.2 eq), TEA (2.2 eq), DMF, rT,
14 h, 73%; (d) Grubbs’ catalyst 2 (0.08 eq), DCM, rT, 8 h, 89% (mixture of E=Z isomers) [HATU ¼ O-
(7-azabenzo-triazol-1-yl)-N,N,N0,N0-tetramethyluronium hexafluorophosphate, TEA ¼ triethylamine].
The product of the RCM reaction was observed as a single spot on thin-layer
chromatography (TLC) and isolated by flash chromatography. However, analysis of
the purified product by (high performance liquid chromatography) (HPLC) revealed
that a 78=22 mixture of E=Z isomers had been formed in the RCM reaction.[5] The
E=Z isomers were readily separated by preparative HPLC and further analyzed by
1H NMR. The main isomer was assigned as the E-isomer based on the large coupling
constant (J ¼ 15.3 Hz) indicative for a trans double-bond geometry. We subsequently
subjected a few more similarly substituted RCM precursors (Table 1, compounds 17,
20, and 23; prepared using the synthetic route as described) to the same metathesis
conditions and found that the E=Z ratio of the products was fairly independent of
the substitution pattern (Table 1). In all four examples, the E=Z ratio was in the same
range (E=Z ꢃ 3=1). It was possible to separate the E=Z isomers by preparative HPLC
in all cases. Some of the E-isomers (compounds 18, 21, and 24) were commercially
available from AnalytiCon (Potsdam, Germany) as part of a screening library. The
main isomers observed in the RCM reaction gave an exact match in retention time
on HPLC compared to the commercial E-isomers purchased from AnalytiCon [e.g.,
for compound 18, E-isomer: t R(Rigel) ¼ 10.70 min vs. t R(AnalytiCon) ¼10.70 min].
(HPLC was performed on an Agilent Zorbax SB-C18.2.1 ꢄ 50-mm (5mm) column.)
These findings further confirmed the assignment of the double-bond geometry.
To further take advantage of the products obtained in the complexity-generating
macrocyclization process, we envisioned postmacrocyclization functionalizations in
the fashion of diversity-orientated synthesis (DOS). Hence, the HPLC-purified macro-
cyclic E-isomer 1 was subjected to a number of late-stage manipulations to gain access

2858
T. J. HECKRODT AND R. SINGH
Table 1. E=Z ratio of RCM products
E=Z
ratio
Entry
RCM precursor
E-isomer
Z-isomer
A
78=22
B
68=32
C
72=28
D
73=27
to analogs for SAR studies. Functional groups and sites of reactivity were exploited
for the following transformations: (a) the double bond was hydrogenated using pal-
ladium on charcoal as catalyst, (b) the olefin was dihydroxylated with Sharpless’
AD-mix-a,[6] (c) the aromatic chloride was used as a handle for a microwave-assisted

PREPARATION OF 17-MEMBERED MACROCYCLES
2859
Scheme 5. Late-stage modifications to access analogs for SAR studies. Reagents and conditions: (a) H2,
t
Pd=C (10 wt%); (b) AD-mix-a (1.4 g=mmol), Bu-OH, H2O, 0 ꢂC to rT, 58%; (c) p-Pyr-B(OH)2 (2.0 eq),
PdCl2(PPh3)2 (0.1 eq), Na2CO3 (1.2 eq), DME=EtOH=H2O 7:3:2, 15 min, microwave, 150 ꢂC, 19%; (d)
NaOH (1.5 eq), MeOH=H2O 1:1, rT, 12 h, 94%.
Suzuki–Miyaura coupling[7] to generate a biaryl moiety, and (d) the methyl ester was
hydrolyzed to the carboxylic acid (Scheme 5).
In summary, we developed a synthetic entry into the described class of
17-membered macrocycles using a sequence of amide couplings followed by a
RCM reaction to obtain the final product. In all four cases, the RCM reaction
yielded the products as a mixture of E=Z isomers in a ratio of about three to one.
Reactive groups in the macrocycle were used for a number of late-stage modifica-
tions to yield analogs for SAR studies.
EXPERIMENTAL
All moisture-sensitive reactions were carried out under nitrogen. Anhydrous sol-
vents were purchased from Aldrich. All other solvents were HPLC grade. Column
chromatography was performed with EMD Merck silica gel (0.040–0.63 mm,
240–400 mesh) under the low pressure of 5–10 psi. Flash chromatography was carried

2860
T. J. HECKRODT AND R. SINGH
out on a Teledyne Isco CombiFlash Rf flash system with variable solvent gradients.
TLC analysis was performed with E. Merck silica-gel 60-F254 plates. NMR spectra
were recorded on a Varian Mercury VX 300-MHz spectrometer. NMR spectra mea-
sured in CDCl3 solutions were referenced to the residual CHCl3 signal (1H, d ¼ 7.26;
13C, d ¼ 77.0); spectra measured in dimethylsulfoxide (DMSO-d6) were referenced to
1
the residual DMSO signal (1H, d ¼ 2.50; 13C, d ¼ 39.50). H and 13C shifts are given
in ppm (s ¼ singlet, d ¼ doublet, t ¼ triplet, q ¼ quadruplet, quin¼ quintet,
m ¼ multiplet, br s ¼ broad signal). Coupling constants, J, are given in hertz (Hz).
Mass spectra were measured on a Waters Micromass ZQ or ZMD instrument. Accu-
rate mass measurements (HRMS) were obtained using an Acquity ultra performance
liquid chromatography (UPLC) system with PDA, ELSD, and LCT Premier XE mass
spectrometer (time of flight). Monitored wavelength was 254 nm. Mass analysis was
performed in extended W-mode using leucine enkephalin as reference lock mass,
556.2771 Da for positive ion, and 554.2615 Da for negative ion. Samples were initially
prepared in methanol at 1 mg=mL and diluted as necessary to remove dead time arti-
facts. Column: Acuity UPLC, BEH18, 1.7 mm, 2.1 ꢄ 50 mm, solvents: A ¼ water
0.05% formic acid, B ¼ acetonitrile 0.05% formic acid, flow rate: 0.735 mL=min, and
gradient: 95% A, 5% B to 100% B in 4 min.
Amine 8
This compound was synthesized using a modified procedure described by Sal-
vatore et al.[3] Phenylethylamine (50.0 g, 413 mmol), CsOH ꢀ H2O (6.93 g, 41.3 mmol),
˚
and activated 4-A molecular sieves (20 g) were combined with dry dimethylforma-
mide (DMF, 300 ml) and stirred at room temperature for 30 min. The mixture was
cooled to ꢁ20 ꢂC using a NaCl=ice bath. Then allyl bromide (50.0 g, 423 mmol)
was added very slowly dropwise. The reaction mixture was allowed to warm to room
temperature overnight. Most of the solvent was removed in vacuo, and saturated
NaHCO3 (400 ml) was added to the crude product. The mixture was extracted with
Et2O (3 ꢄ), the combined organic layers were dried over MgSO4, and solvents were
evaporated under reduced pressure. Analysis by TLC (3% MeOH [2 M NH3] in
CHCl3] and HPLC showed that the mono- and di-allylated products had been
formed in roughly the same amounts. The desired product was isolated by flash
chromatography eluting with 3% MeOH [2 M NH3] in CHCl3. Monoallylated com-
pound 8 was obtained in 52% yield (34.5 g) in form of a pale yellow oil.
1H NMR (300 MHz, DMSO) d 7.28–7.10 (m, 5H), 5.96–5.65 (m, 1H), 5.11 (d,
J ¼ 17.2 Hz, 1H), 5.00 (d, J ¼ 10.2 Hz, 1H), 3.15 (d, J ¼ 5.8 Hz, 2H), 2.70 (s, 4H), 1.60
(s br, 1H) ppm; MS (ESI) (m=z): 162 [M þ H]þ, 146, 119.
Amide 10
Boc-protected (S)-glutamic methyl ester 9 (2.73 g, 10.4 mmol), ethylene dichlor-
ide (EDC; 3.00 g, 15.7 mmol), HOBt (2.11 g, 15.7 mmol), and Et3N (2.9 ml,
20.8 mmol) were dissolved in dry DCM (150 ml). The mixture was stirred for
15 min, and allyl amine 8 was subsequently added to the reaction mixture. Stirring
was continued overnight at room temperature. The reaction mixture was then con-
centrated in vacuo and further purified by flash chromatography eluting with

PREPARATION OF 17-MEMBERED MACROCYCLES
2861
chloroform=methanol (20=1). The Boc-protected amide was obtained in form of a
clear yellowish oil in 94% yield (3.95 g).
1H NMR (300 MHz, DMSO) d 7.35–7.12 (m, 5H), 5.87–5.59 (m, 1H), 5.11 (d,
J ¼ 11.5 Hz, 1H), 5.04 (d, J ¼ 10.5 Hz, 1H), 4.06–3.91 (m, 1H), 3.85 (d, J ¼ 19.7 Hz,
2H), 3.60 (s, 3H), 3.44–3.29 (m, 2H), 2.75 (m, 2H), 2.43–2.20 (m, 2H), 1.90 (m, 1H),
1.72 (m, 1H), 1.34 (d, J ¼ 7.2 Hz, 9H) ppm; MS (ESI) (m=z): 405 [M þ H]þ, 349, 305.
The product from this reaction was dissolved in methanol (100 ml), and HCl
(20 ml, 4 M in dioxane) was added via syringe. The mixture was stirred at room tem-
perature until analysis by TLC showed complete conversion (1 d). After neutralizing
with 1 M NaOH (80 ml), the mixture was extracted with Et2O (2 ꢄ) and EtOAc (2 ꢄ).
The combined organic layers were dried over Na2SO4, and solvents were removed
under reduced pressure to give 2.91 g (98%) amide 10.
Acid 13
Salicylic acid 11 (30.0 g, 174 mmol) and allyl bromide (84.1 g, 696 mmol) were
dissolved in acetone (500 ml). K2CO3 (72.1 g, 522 mmol) and Cs2CO3 (5.67 g,
17.4 mmol) were added, and the reaction mixture was refluxed for 8 h. Salts were fil-
tered off, and the acetone was removed under reduced pressure. Excess allyl bromide
was removed by applying high vacuum to the rotary evaporator. The yellowish crude
oil was dissolved in ethanol (250 ml), and 2 M NaOH (104 ml, 208 mmol) was added.
The mixture was stirred overnight at room temperature and then acidified (ice-bath
cooling) using concentrated HCl. Acid 13 was precipitated by adding ice water and
keeping the flask in the refrigerator for several hours. The product was obtained in
form of a solid and collected by filtration (34.7 g, 94%).
1H NMR (300 MHz, DMSO) d 7.65 (d, J ¼ 8.3 Hz, 1H), 7.16 (s, 1H), 7.03 (d,
J ¼ 8.3 Hz, 1H), 6.12–5.88 (m, 1H), 5.46 (d, J ¼ 17.3 Hz, 1H), 5.23 (d, J ¼ 10.6 Hz,
1H), 4.64 (d, J ¼ 4.7 Hz, 2H) ppm; 13C NMR (75 MHz, DMSO) d 167.13, 158.52,
137.90, 133.56, 132.94, 120.87, 117.81, 114.64, 69.59 ppm; MS (ESI) (m=z): 213
[M þ H]þ, 195, 167, 157.
Amide 14
Boc-(R)-phenylalanine (2.48 g, 9.35 mmol), EDC (2.69 g, 14.0 mmol), HOBt
(1.89 g, 14.0 mmol), and Et3N (2.60 ml, 18.7 mmol) were dissolved in dry DCM
(150 ml). The mixture was stirred for 30 min, and glutamic amino methyl ester 10
(2.84 g, 9.35 mmol) was subsequently added to the reaction mixture. Stirring was
continued overnight at room temperature. The reaction mixture was then concen-
trated in vacuo and further purified by flash chromatography eluting with
chloroform=methanol (40=1). The Boc-protected amide was obtained in form of a
pale yellow oil in 78% yield (4.02 g).
1H NMR (300 MHz, DMSO) d 8.35 (d, J ¼ 7.6 Hz, 1H), 7.33–7.07 (m, 10H),
6.87 (d, J ¼ 8.7 Hz, 1H), 5.73 (tdt, J ¼ 15.8, 10.4, 5.3 Hz, 1H), 5.09 (m, 2H), 4.25
(m, 2H), 3.89 (d, J ¼ 5.2 Hz, 1H), 3.80 (d, J ¼ 4.0 Hz, 1H), 3.61 (s, 3H), 3.37 (d,
J ¼ 6.5 Hz, 2H), 2.99–2.84 (m, 1H), 2.83–2.61 (m, 3H), 2.24 (m, 2H), 1.92 (td,
J ¼ 13.8, 6.3 Hz, 1H), 1.84–1.66 (m, 1H), 1.27 (s, 9H) ppm; 13C NMR (75 MHz,
DMSO) d 172.83, 172.34, 171.53, 171.11, 139.84, 139.18, 129.82, 129.41, 129.24,

2862
T. J. HECKRODT AND R. SINGH
129.01, 128.57, 126.79, 117.18, 116.69, 79.89, 78.66, 56.25, 52.62, 50.65, 47.88, 38.67,
35.09, 34.31, 28.91, 27.54 ppm; MS (ESI) (m=z): 552 [M þ H]þ, 496, 452.
The product from this reaction was dissolved in methanol (120 ml), and HCl
(20 ml, 4 M in dioxane) was added via syringe. The mixture was stirred at room
temperature until analysis by TLC showed complete conversion (1 d). After neutra-
lization with 1 M NaOH (80 ml), the mixture extracted with Et2O (2 ꢄ) and EtOAc
(2 ꢄ). The combined organic layers were passed though a plug of MgSO4 to remove
residual water, and solvents were removed under reduced pressure to furnish 2.99 g
(91%) amide 14.
RCM Precursor 15
Salicylic acid 13 (0.735 g, 3.45 mmol), HATU (1.57 g, 4.14 mmol), and Et3N
(1.05 ml, 7.59 mmol) were dissolved in dry DMF (50 ml). The mixture was stirred
for 30 min, and compound 14 (1.56 g, 3.45 mmol) was subsequently added to the
reaction mixture. Stirring was continued overnight at room temperature. The reac-
tion mixture was then concentrated in vacuo and further purified by flash chroma-
tography, eluting with chloroform=methanol (30=1). The desired amide 15 was
obtained in form of a pale yellow oil in 73% yield (1.63 g).
1H NMR (300 MHz, CDCl3) d 8.25 (d, J ¼ 7.8 Hz, 1H), 8.08 (dd, J ¼ 7.8,
3.8 Hz, 1H), 7.35–7.04 (m, 11H), 6.99 (d, J ¼ 8.8 Hz, 1H), 6.87 (s, 1H), 5.92 (m,
1H), 5.62 (m, 1H), 5.33 (dd, J ¼ 17.2, 9.8 Hz, 2H), 5.12 (dd, J ¼ 18.4, 10.6 Hz, 2H),
4.95 (m, 1H), 4.55 (m, 4H), 4.32 (m, 1H), 3.67 (s, 3H), 3.51–3.33 (m, 3H), 3.21 (d,
J ¼ 6.8 Hz, 2H), 2.78 (m, 3H), 2.38–2.07 (m, 1H), 2.06–1.83 (m, 1H) ppm; 13C
NMR (75 MHz, CDCl3) d 172.19, 171.79, 171.05, 167.44, 164.42, 157.22, 139.24,
138.73, 138.25, 136.93, 133.54, 132.86, 131.64, 129.51, 128.95, 128.71, 127.00,
126.50, 121.83, 119.77, 116.94, 113.68, 70.65, 55.17, 52.63, 51.12, 48.69, 38.26,
35.20, 34.39, 29.30, 27.17 ppm; MS (ESI) (m=z): 646 [M þ H]þ, 305.
Macrocycles 1 and 16
The product 15 (758 mg, 1.17 mmol) from this reaction was dissolved in dry
DCM (200 ml, c ꢃ 0.006 mol=L); the solution was degassed and flushed with N2
(3 ꢄ). Grubbs’ II catalyst (80.0 mg, 0.091 mmol) dissolved in dry DCM (20 ml) was
added dropwise over a period of 5 min. Stirring was continued for 8 h at room tem-
perature. The clear, peach-colored solution turned blackish within a couple of hours.
Water (30 ml) was added to deactivate the catalyst; the organic layer was separated
and filtered through a plug of MgSO4. Solvents were removed under reduced press-
ure, and the crude product was purified by column chromatography eluting with
chloroform=methanol (30=1). The product (dark oil, 644 mg, 89%) was obtained
as a mixture of E=Z-isomers in a ratio of 78=22 (determined by LCMS). The
geometric isomers were separated by preparative HPLC to allow spectroscopic
characterization. The final compounds were obtained in the form of white solids.
Macrocycle 1 (E-Isomer). 1H NMR (300 MHz, DMSO) d 8.30 (d, J ¼ 8.8 Hz,
1H), 7.76 (d, J ¼ 8.4 Hz, 1H), 7.32–7.05 (m, 11H), 5.63 (d, J ¼ 15.3 Hz, 1H), 5.44 (d,
J ¼ 15.3 Hz, 1H), 4.84 (d, J ¼ 3.7 Hz, 1H), 4.59 (s, 1H), 4.55–4.46 (m, 1H), 4.45–4.25

PREPARATION OF 17-MEMBERED MACROCYCLES
2863
(m, 2H), 4.09–3.95 (m, 1H), 3.66 (d, J ¼ 7.3 Hz, 1H), 3.62 (s, 3H), 3.24 (dd, J ¼ 13.7,
5.3 Hz, 1H), 3.05 (dd, J ¼ 13.0, 6.6 Hz, 1H), 2.96 (dd, J ¼ 13.3, 4.5 Hz, 1H), 2.70
(s, 2H), 2.14 (d, J ¼ 13.2 Hz, 2H), 1.94–1.69 (m, 2H) ppm; 13C NMR (75 MHz,
CDCl3) d 172.49, 171.59, 170.24, 164.12, 157.26, 139.24, 139.14, 136.56, 133.44,
130.51, 130.37, 128.96, 128.71, 128.60, 127.15, 126.56, 124.18, 121.93, 119.62,
112.83, 69.18, 53.92, 53.01, 52.29, 49.06, 48.66, 37.30, 34.20, 28.49, 28.07 ppm; MS
(ESI) (m=z): 618 [M þ H]þ; HRMS (TOF MS ESþ) m=z calcd. for C34H36ClN3O6:
618.2371; found: 618.2319.
Macrocycle 16 (Z-Isomer). 1H NMR (300 MHz, CDCl3) d 8.49 (d,
J ¼ 4.0 Hz, 1H), 8.31 (d, J ¼ 8.3 Hz, 1H), 8.18 (d, J ¼ 8.5 Hz, 1H), 7.36–7.17 (m,
10H), 7.04 (d, J ¼ 8.5 Hz, 1H), 6.98 (s, 1H), 5.62–5.47 (m, 2H), 5.04 (td, J ¼ 8.3,
4.8 Hz, 1H), 4.81 (t, J ¼ 9.5 Hz, 1H), 4.56 (dd, J ¼ 14.4, 9.8 Hz, 1H), 4.49–4.41 (m,
1H), 4.32 (s, 1H), 3.77 (s, 3H), 3.52 (m, 2H), 3.20 (dd, J ¼ 14.5, 4.6 Hz, 1H), 3.07
(m, 2H), 2.85 (t, J ¼ 7.3 Hz, 2H), 2.38–2.10 (m, 2H), 2.03 (dd, J ¼ 16.1, 6.0 Hz,
1H), 1.88 (dd, J ¼ 13.5, 5.5 Hz, 1H) ppm; 13C NMR (75 MHz, CDCl3) d 172.90,
172.61, 171.99, 164.55, 158.05, 138.91, 137.79, 137.40, 134.06, 130.33, 129.65,
129.16, 128.89, 128.69, 127.29, 126.90, 124.78, 122.00, 119.26, 113.51, 64.19, 55.11,
53.12, 52.27, 51.49, 45.01, 38.55, 35.57, 28.88, 23.23 ppm; MS (ESI) (m=z): 618
[M þ H]þ; HRMS (TOF MS ESþ) m=z calcd. for C34H36ClN3O6: 618.2371; found:
618.2315.
Additional Data for Macrocycles in Table 1
Macrocycle 18 (E-Isomer). 1H NMR (300 MHz, DMSO) d 8.34 (d,
J ¼ 7.7 Hz, 1H), 8.24 (d, J ¼ 7.7 Hz, 1H), 7.77 (d, J ¼ 8.3 Hz, 1H), 7.27 (s, 1H),
7.20–7.03 (m, 3H), 6.86 (d, J ¼ 8.7 Hz, 2H), 5.97–5.69 (m, 2H), 4.84–4.57 (m, 3H),
4.53 (dd, J ¼ 14.4, 7.1 Hz, 1H), 4.36 (t, J ¼ 9.3 Hz, 1H), 4.15–3.88 (m, J ¼ 37.3 Hz,
2H), 3.71 (s, 3H), 3.60 (s, 4H), 2.40–2.07 (m, 4H), 1.91–1.68 (m, 1H), 1.34 (d,
J ¼ 7.0 Hz, 3H) ppm; 13C NMR (75 MHz, DMSO) d 172.52, 171.78, 163.98,
159.04, 157.42, 137.42, 132.78, 130.61, 129.90, 129.78, 126.64, 122.19, 121.46,
114.44, 113.97, 100.15, 69.69, 55.69, 52.63, 51.98, 49.72, 48.11, 47.90, 28.36, 26.83,
19.15 ppm; MS (ESI) (m=z): 558 [M þ H]þ, 333; HRMS (TOF MS ESþ) m=z calcd.
for C28H32ClN3O7: 558.2007; found: 558.1954.
Macrocycle 19 (Z-Isomer). 1H NMR (300 MHz, DMSO) d 8.66–8.49 (m,
1H), 8.41 (d, J ¼ 7.9 Hz, 1H), 7.98 (t, J ¼ 8.1 Hz, 1H), 7.64 (d, J ¼ 8.3 Hz, 1H),
7.27 (s, 1H), 7.20–7.03 (m, 3H), 6.94–6.80 (m, 2H), 5.98–5.81 (m, 1H), 5.72 (dd,
J ¼ 11.2, 5.9 Hz, 1H), 5.45 (dd, J ¼ 11.5, 5.8 Hz, 1H), 4.72 (s, 2H), 4.54–4.36 (m,
3H), 4.19 (dd, J ¼ 16.1, 9.1 Hz, 1H), 4.09–4.00 (m, 1H), 3.71 (s, 3H), 3.59 (s, 3H),
2.41–2.25 (m, 2H), 2.13–1.92 (m, 2H), 1.27 (d, J ¼ 7.1 Hz, 3H) ppm; 13C NMR
(75 MHz, DMSO) d 173.51, 172.85, 172.49, 171.97, 164.92, 159.07, 157.19, 136.88,
132.08, 130.46, 129.95, 129.09, 124.89, 123.72, 121.49, 114.50, 65.25, 55.69, 52.98,
52.57, 50.13, 48.89, 45.20, 29.31, 25.98, 18.44 ppm; MS (ESI) (m=z): 558 [M þ H]þ;
HRMS (TOF MS ESþ) m=z calcd. for C28H32ClN3O7: 558.2007; found: 558.1947.
Macrocycle 21 (E-Isomer). 1H NMR (300 MHz, CDCl3) d 9.11 (s, 1H), 8.82
(d, J ¼ 6.9 Hz, 1H), 8.37 (d, J ¼ 9.1 Hz, 1H), 7.28–7.22 (m, J ¼ 3.1 Hz, 4H), 7.14

2864
T. J. HECKRODT AND R. SINGH
(d, J ¼ 8.6 Hz, 1H), 7.08 (d, J ¼ 9.2 Hz, 1H), 6.82 (d, J ¼ 8.7 Hz, 2H), 6.44 (d,
J ¼ 7.2 Hz, 1H), 5.82–5.65 (m, 2H), 5.21 (d, J ¼ 14.6 Hz, 1H), 5.05 (d, J ¼ 6.3 Hz,
1H), 4.77 (t, J ¼ 12.5 Hz, 2H), 4.55 (dd, J ¼ 10.4, 5.9 Hz, 1H), 3.79 (s, 3H), 3.78 (s,
3H), 3.74–3.64 (m, 1H), 3.54–3.37 (m, 2H), 3.33–3.22 (m, 1H), 2.60–2.45 (t,
J ¼ 11.7 Hz, 2H), 2.37–2.15 (m, 1H), 2.06 (d, J ¼ 17.8 Hz, 1H), 1.90–1.70 (m, 2H)
ppm; MS (ESI) (m=z): 645 [M þ H]þ, 333; HRMS (TOF MS ESþ) m=z calcd. for
C34H36N4O9: 645.2560; found: 645.2487.
Macrocycle 24 (E-Isomer). 1H NMR (300 MHz, CDCl3) d 8.50 (d,
J ¼ 7.8 Hz, 2H), 8.20 (d, J ¼ 8.8 Hz, 1H), 7.29–7.16 (m, 5H), 7.13 (d, J ¼ 8.5 Hz,
2H), 6.82 (d, J ¼ 8.6 Hz, 2H), 6.63 (dd, J ¼ 8.8, 2.1 Hz, 1H), 6.52 (d, J ¼ 7.1 Hz,
1H), 6.46 (s, 1H), 5.56 (d, J ¼ 14.1 Hz, 1H), 5.13 (d, J ¼ 14.6 Hz, 1H), 4.57 (dd,
J ¼ 10.8, 4.4 Hz, 1H), 4.43 (dd, J ¼ 15.6, 7.6 Hz, 1H), 4.36 (dd, J ¼ 10.7, 6.2 Hz,
1H), 3.86 (s, 3H), 3.77 (s, 3H), 3.75 (s, 3H), 3.70–3.56 (m, 1H), 3.55–3.42 (m, 1H),
3.39 (dd, J ¼ 14.3, 3.6 Hz, 1H), 3.24 (d, J ¼ 5.4 Hz, 1H), 2.90 (s br, 2H), 2.62–2.38
(m, 1H), 2.17–2.01 (m, 1H), 1.97–1.68 (m, 2H) ppm; MS (ESI) (m=z): 630 [M þ H]þ,
333; HRMS (TOF MS ESþ) m=z calcd. for C35H39N3O8: 630.2816; found: 630.2776.
ACKNOWLEDGMENTS
We are thankful to the Department of Analytical Chemistry (Van Nguyen,
Duayne Tokushige, and Mark Irving) of Rigel Pharmaceuticals for numerous pre-
parative HPLC separations. Thanks are also due to Taisei Kinoshita from the
Biology Department for various helpful discussions.
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