A facile and efficient synthesis of d3-labelled Reyataz

Article abstract of DOI:10.1002/jlcr.1019

A facile and efficient synthesis of d₃-labelled Reyataz is described. The key step of synthesis involved the coupling of N-(d ₃-methoxycarbonyl)-L-tert-leucine 2b with an advanced chiral non-racemic building block 4. The latter was synthesized in 4 steps from the readily accessible hydrazine derivative 8. Copyright

Full text of DOI:10.1002/jlcr.1019

JOURNAL OF LABELLED COMPOUNDS AND RADIOPHARMACEUTICALS
J Label Compd Radiopharm 2005; 48: 1041–1047.
Published online in Wiley InterScience (www.interscience.wiley.com). DOI: 10.1002/jlcr.1019
Research Article
A facile and efficient synthesis of d -labelled ReyatazTM
3
Huiping Zhang*, Samuel J. Bonacorsi Jr, Bang-Chi Chen, Leslie W. Leith,
J. Kent Rinehart, Balu Balasubramanian and Joel C. Barrish
Discovery Chemistry, Pharmaceutical Research Institute, Bristol Myers Squibb Co.,
Princeton, NJ 08543-4000, USA
Summary
TM
A facile and efficient synthesis of d -labelled Reyataz
3
is described. The key step of
synthesis involved the coupling of N-(d -methoxycarbonyl)-l-tert-leucine 2b with an
3
advanced chiral non-racemic building block 4. The latter was synthesized in 4 steps
from the readily accessible hydrazine derivative 8. Copyright # 2005 John Wiley &
Sons, Ltd.
Key Words: ReyatazTM; HIV protease inhibitor; d -labelled; chiral non-racemic
3
Introduction
ReyatazTM (Atazanavir) is a potent human immunodeficiency virus (HIV)
protease inhibitor and is the first once-daily protease inhibitor approved by the
FDA to treat patients with HIV. For bioequivalence studies of ReyatazTM in
1
TM
humans, d -labelled Reyataz
3
is required because its physical and
chromatographic properties are identical to the unlabelled compound. Several
TM
efficient synthetic routes leading to Reyataz
have been published in the
2
,3
literature. Several of these use the symmetrical coupling of 2 equivalents of
N-methoxycarbony-l-tert-leucine 2a with a diamine derivative 3 (Scheme 1) as
a general approach. Retrosynthetic analysis suggested that a strategy involving
the unsymmetrical coupling of N-(d -methoxycarbonyl)-l-tert-leucine 2b with
3
TM
a key chiral non-racemic intermediate 4 would give d -labelled Reyataz
3
(Scheme 1). The chiral non-racemic building block 4 can be obtained by the
regioselective aminolysis of epoxide 5 with hydrazide 6a, which can be
synthesized by the coupling of phenyl hydrazine derivative 7 with N-
methoxycarbonyl-l-tert-leucine 2a. Herein we describe the design and
TM
synthesis of d -Reyataz
using this unsymmetrical coupling approach.
3
*Correspondence to: Huiping Zhang, Bristol Myers Squibb Co., Discovery Chemistry, MailStop 15-14,
Princeton, NJ 08543, USA. E-mail: huiping.zhang@bms.com
Copyright # 2005 John Wiley & Sons, Ltd.
Received 16 August 2005
Revised 29 September 2005
Accepted 30 September 2005

1
042
H. ZHANG ET AL.
N
OH
O
N
O
H N
N
R=H
OH
OH
2
+
NH2
H CO
3
N
H
Symmetrical
coupling
CH Ph 3
2
O
O
O
OH
O
2a
H
N
H
N
N
OCH3
N
R CO
N
H
N
H
3
O
CH Ph
2
O
OH
O
R=D
H
N
H N
2
N
OCH3
a:R=H, ReyatazTM
D CO
3
+
1
1
N
H
N
H
TM
b:R=D,d - Reyataz
Unsymmetrical
coupling
3
CH Ph
2
O
4
2b
N
O
N
O
O
H
+
OH
O
+
O
N
H CO
3
N
H
O
H
HN
N
OCH3
O
CH Ph
2
HN
N
H
6a
NH2
2a
5
7
Scheme 1. Retrosynthesis of ReyatazTM and d -labelled ReyatazTM
3
O
N
N
OH
H
3
CO
N
H
N
N
2
a
O
O
O
HCl/THF
100%
HOBT, WSC, DCM
H
N
N
HN
OCH
3
+ H
3
CO
N
H
NH
2
O
N
H
HN
O
HN
NH
2
O
N
O
6b
6a/6b=10/1
H
8
7
6a
7
8%
O
H
N
O
N
O
N
O
2
CH Ph
OH
O
H
OH
O
H
N
HCl/THF
100%
H
IPA
H
N
N
N
OCH
3
O
N
N
OCH
3
2
N
H
N
H
2
5%
O
CH
2
Ph
O
2
CH Ph
9
4
O
N
O
OH
H
3
CO
N
H
2
a
O
O
OH
O
HOBT, WSC, DCM
H
N
H
N
N
OCH
3
H
3
CO
N
H
N
H
9
5%
O
2
CH Ph
TM
a: Reyataz
1
Scheme 2. Synthesis of intermediate 4 and ReyatazTM
Results and discussion
The key amino intermediate 4 was synthesized as outlined in Scheme 2. Acid
hydrolysis of hydrazinocarbamate 8 gave the hydrochloride salt of phenyl
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 1041–1047

FACILE AND EFFICIENT SYNTHESIS OF d
3
-LABELLED REYATAZTM
1043
OH
H N
2
1
0
O
O
9
0% D CO
Cl
3
Dioxane/2N NaOH
N
O
N
O
O
OH
N
D CO
3
OH
O
O
OH
O
H
H
N
2b
H
N
H
N
O
H N
N
OCH3
N
OCH3
2
HOBT, WSC, DCM
78%
N
H
D CO
3
N
H
N
H
4
CH Ph
O
CH Ph
2
2
TM
1
b:d - Reyataz
3
TM
Scheme 3. Synthesis of d -labelled Reyataz
3
hydrazine derivative 7. Compound 7 was treated with diisopropylethylamine
and added to a mixture of N-methoxycarbonyl-l-tert-leucine 2a, water-soluble
carbodiimide [WSC, 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydro-
chloride] and 1-hydroxybenzotriazole (HOBT) in dichloromethane to yield a
crude solid containing the desired hydrazide 6a and undesired hydrazide 6b in
1
0:1 HPLC area ratio. Subsequent recrystallization from EtOH gave the
desired hydrazide 6a in 78% yield. Treatment of compound 6a with epoxide 5
in refluxing IPA for 24 h gave compound 9. Removal of the protecting group
using HCl in THF gave the requisite precursor 4 in quantitative yield. Since
compound 4 was a novel intermediate, it was further converted to the known
free base of ReyatazTM to confirm the structure assignment. Treatment of
compound 4 with N-methoxycarbonyl-l-tert-leucine, WSC and HOBT in
TM
dichloromethane yielded the free base of Reyataz
1a in 95% yield. The
TM
spectroscopic and physical data for synthetic free base of Reyataz
3
identical to that reported previously.
were
As shown in Scheme 3, the incorporation of deuterium into N-
methoxycarbonyl-l-tert-leucine was achieved by the acylation of l-tert-leucine
4
with d -methyl chloroformate to give compound 2b in 90% yield. Coupling of
3
compound 4 directly with d -methyl chloroformate 2b in the presence of WSC
3
TM
and HOBT gave d -labelled Reyataz
3
in 78% yield.
Experimental
All reagents were obtained from Aldrich Chemical Company and used without
further purification unless otherwise stated. Tetrahydrofuran ultra-low water
was obtained from J.T. Baker and used without further purification.
d3-Methyl chloroformate was obtained from Cambridge Isotope Laboratories
with a chemical and isotopic abundance of greater than 98%. The hydrazine
derivative 8, carboxylic acid 2a and epoxide 5 were obtained from Process
Research & Development Department of Bristol Myers Squibb Company.
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 1041–1047

1
044
H. ZHANG ET AL.
All glassware was dried and purged with nitrogen or argon before use. All
reactions were monitored by HPLC using a Shimadzu system (model SCL-
0A) equipped with a Shimadzu PDA detector (model SPD-10AV) or a
Rainin Dynamax system (model SD-200) equipped with a Varian ProStar
1
1
13
PDA detector (model 330). All H NMR and C NMR spectra were recorded
on a Bruker 400 MHz spectrometer using CDCl or DMSO-d as the solvent.
3
6
Preparation of 1-(4-(pyridin-2-yl)benzyl)hydrazine (7)
To a stirring solution of compound 8 (15 g, 50 mmol) in THF (70 ml) was
added HCl (12 M, 10 ml) under nitrogen at room temperature. The reaction
mixture was heated to 508C, stirred at this temperature for 3 h, then cooled to
room temperature. THF was decanted from the mixture and the resulting
yellow solid washed with THF (2 ꢀ 20 ml) and dried in vacuo to give the
product (7) (10 g, 100%). The product was used for the next step without
further purification.
0
Preparation of {(S)-2,2-dimethyl-1-[N -(4-pyridin-2-yl-benzyl)-hydrazinocar-
bonyl]-propyl}-carbamic acid methyl ester(6a)
To a stirring solution of compound 2a (10.4 g, 50 mmol) in dichloromethane
(250 ml) was added HOBT (9.44 g, 70 mmol) and WSC (9.74 g, 50 mmol). After
the mixture was stirred at room temperature under nitrogen for 20 min,
compound 7 (9.95 g, 50 mmol) in dichloromethane (60 ml) and diisopropy-
lethylamine (19.2 g, 150 mmol) was added. The reaction was carried out for
1
.5 h and was quenched with H O (125 ml). The organic layer was separated
2
and the aqueous layer was extracted with dichloromethane (2 ꢀ 150 ml). The
combined organic layers were dried over anhydrous Na SO , filtered, and
2
4
concentrated to give crude product (25 g) with a HPLC area ratio of
a=6b ¼ 10=1. The crude product was recrystallized from 150 ml of EtOH/
hexane (50/50) to give pure compound (6a) (14.43 g, 78%) as a white solid.
6
1
HNMR (CDCl ): 8.67(d, J ¼ 6:2 Hz, 1H), 8.28 (s, 1H), 7.93(d, J ¼ 8:15 Hz,
3
2
H), 7.72(m, 2H), 7.43(d, J ¼ 8:18 Hz, 2H), 7.21(m, 1H), 3.97(m, 3H), 3.18
s, 3H), 0.96 (s, 9H). CNMR (CDCl ): 170.7, 157.5, 157.4, 150.0, 139.0,
13
(
3
1
38.6, 137.1, 129.6, 127.7, 127.3, 122.5, 120.8, 61.5, 55.9, 52.7, 34.9, 26.8.
Analytically calculated for C H N O : C, 64.84; H, 7.07; N, 15.12; found: C,
26
20
4
3
6
4.86; H, 7.16; N, 15.25.
0
Preparation of {(1S,2S)-1-benzyl-2-hydroxy-3-[N -((S)-2-methoxycarbonyl-
amino-3,3-dimethyl-butyryl)-N-(4-pyridin-2-yl-benzyl)-hydrazino]-propyl}-
carbamic acid tert-butyl ester (9)
The stirring solution of compound 6a (11.3 g, 30 mmol) in IPA (80 ml) was
heated to reflux and compound 5 (8.67 g, 33 mmol) was added. The mixture
was refluxed for 24 h, cooled to room temperature and concentrated in vacuo
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 1041–1047

FACILE AND EFFICIENT SYNTHESIS OF d
3
-LABELLED REYATAZTM
1045
to give a syrup which was purified by column chromatography on silica gel
(
elution with 40% EtOAc/hexanes, TLC R ¼ 0:2) to give the desired product
f
1
(9) (4.97 g, 25%) as a white solid. H NMR (CDCl ): 8.67(d, J ¼ 4:8 Hz, 1H),
3
7
.92(d, J ¼ 8:1 Hz, 2H), 7.74(m, 2H), 7.42(d, J ¼ 8:0 Hz, 2H), 7.21(m, 6H),
.3(bs, 1H), 5.1(bs, 1H), 4.8(bs, 1H), 4.1(d, J ¼ 13:9 Hz, 1H), 3.91
5
(
d, J ¼ 13:8 Hz, 1H), 3.62(m, 6H), 2.90(m, 3H), 2.63(d, J ¼ 11:6 Hz, 1H),
.10(bs, 1H), 1.34(s, 9H), 0.72 (s, 9H). C NMR (CDCl ): 171.3, 157.3, 157.2,
3
13
2
1
1
56.3, 150.0, 139.3, 138.8, 137.1, 136.9, 129.8, 129.6, 128.6, 127.4, 126.5, 122.5,
20.8, 79.4, 67.3, 62.8, 61.9, 61.7, 53.5, 52.8, 39.6, 34.6, 28.7, 26.5. Analytically
calculated for C H N O .0.39 H O: C, 65.60; H, 7.52; N, 10.93; found: C,
3
5
47
5
6
2
6
5.60; H, 7.57; N, 10.84.
0
0
Preparation of {(S)-1-[N -((2S,3S)-3-amino-2-hydroxy-4-phenyl-butyl)-N -
4-pyridin-2-yl-benzyl)-hydrazinocarbonyl]-2,2-dimethyl-propyl}-carbamic acid
methyl ester (4)
(
To a stirring solution of compound 9 (2.20 g, 3.47 mmol) in THF (20 ml) was
added HCl (12 M, 1.0 ml) under nitrogen at room temperature. The reaction
mixture was heated to 508C, stirred for 5 h at that temperature, and cooled to
room temperature. The THF was decanted from the mixture and the resulting
yellow solid was washed with THF (2 ꢀ 20 ml) and dried in vacuo to give the
1
product (4) (1.85 g, 100%). HNMR (DMSO-d ): 8.61(d, J ¼ 5:5 Hz, 1H),
6
8
.32(t, J ¼ 8:2 Hz, 1H), 8.15(m, 1H), 7.86(d, J ¼ 8:2 Hz, 3H), 7.71
(t, J ¼ 6:6 Hz, 1H), 7.31(d, J ¼ 8:2 Hz, 2H), 7.04(m, 3H), 6.81(m, 1H),
3
1
5
.93(bs, 1H), 3.70(d, J ¼ 3:7 Hz, 1H), 3.38(m, 4H), 3.16(s, 3H), 2.76(m, 3H),
þ
.5(bs, 3H), 0.37(s, 9H) ESIMS m/z 534.3 (½M þ H ꢁ, C H N O requires
3
0
39
5
4
33.3.
Preparation of N-(d -methoxycarbonyl)-l-tert-leucine (2b)
3
A stirred solution of l-tert-leucine 10 (11.4 g, 87.2 mmol) in dioxane (40 ml)
and sodium hydroxide (2 M, 145 ml) was cooled in ice and d -methyl
3
chloroformate (17 g, 174.3 mmol) was added dropwise over 45 min, maintain-
ing the internal temperature between 15 and 188C. The reaction flask was
warmed to 608C, stirred at this temperature for 18 h, and extracted with
dichloromethane (2 ꢀ 50 ml). The organic solution was acidified to pH ¼ 2 by
aqueous HCl (6 M) and extracted with ethyl acetate (3 ꢀ 50 ml). The extracts
were dried over Na SO , filtered and concentrated in vacuo. The residue was
2
4
crystallized from hexanes (80 ml) to give the product (2b) (14.2 g, 85%) as a
1
white solid. H NMR (300 MHz, CDCl ): d 9.9 (s, 1H), 5.2 (bs, 1H), 4.1 (bs,
3
1
3
1
H), 0.9 (s, 9H); C NMR (100 MHz, CDCl ): d 176.5, 156.9, 62.1, 34.5, 26.4;
3
þ
ESIMS m/z 193.1 (½M þ H ꢁ, C H D NO requires 193.12.
8
13
3
4
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 1041–1047

1
046
H. ZHANG ET AL.
TM
Preparation of d -labelled Reyataz
3
(1b)
A solution of N-(d -methoxycarbonyl)-l-tert-leucine (331 mg, 1.72 mmol) in
3
dichloromethane (10 ml) was added HOBT (253 mg, 1.87 mmol) and WSC
(343 mg, 1.79 mmol), stirred at room temperature for 2 h, and treated with
compound 4 (1.0 g, 1.56 mmol) in aqueous K HPO (1.14 M, 10 ml). The
2
4
mixture was stirred at this temperature for 12 h. The organic phase was
separated, washed with aqueous NaH PO (1 M, 2 ꢀ 10 ml, pH 1.5), stirred
2
4
with aqueous NaOH (0.5 M, 10 ml) for 2 h, dried over anhydrous Na SO , and
2
4
concentrated in vacuo. The residue (967 mg, 88%) was crystallized from
dichloromethane/toluene to give the desired product (1b) (849 mg, 78%) as a
1
white solid. H NMR (300 MHz, DMSO-d ): d 9.1 (s, 1H), 8.6 (d, J ¼ 4:6 Hz,
6
1
H), 8.0 (d, J ¼ 8:2 Hz, 2H), 7.9 (m, 2H), 7.5 (d, J ¼ 9:1 Hz, 1H), 7.4
(d, J ¼ 8:2 Hz, 2H), 7.3 (m, 1H), 7.2 (s, 4H), 7.1 (m, 1H), 6.9 (d, J ¼ 9:3 Hz,
1
1
H), 6.8 (d, J ¼ 9:3 Hz, 1H), 4.04 (m, 1H), 3.9 (m, 2H), 3.8 (d, J ¼ 9:3 Hz,
H), 3.6 (m, 2H), 3.5 (s, 3H), 2.7 (m, 4H), 0.8 (s, 9H), 0.6 (s, 9H); C NMR
1
3
(
100 MHz, DMSO-d ): d 170.1, 170, 156.4, 156, 149.4, 138.9, 138.6, 137.4,
6
1
5
37.1, 129, 128.7, 128.2, 127.9, 126.1, 125.8, 122.3, 119.9, 68.1, 63, 61.2, 60.8,
þ
1.6, 51.4, 37.7, 33.5, 33.3, 26.6, 26.2; ESIMS m/z 708.5 (½M þ H ꢁ,
C H D N O requires 708.41).
3
8
50
3
6
7
Conclusion
A versatile advanced chiral non-racemic building block 4 was synthesized in 4
steps from the readily accessible hydrazine derivative 8. The structure
assignment was established by the conversion into the free base of ReyatazTM
.
The synthetic utility of this chiral building block was illustrated in the high
yielding one step conversion into d -labelled ReyatazTM, which was a valuable
3
internal standard for an assay designed to quantify the plasma concentration
of ReyatazTM taken from subjects in clinical studies.
Appendix A
TM
HPCL of d -labelled Reyataz
3
(1b) is given in Figure A1.
TM
Figure A1. HPLC of d -labelled Reyataz
3
(1b)
Copyright # 2005 John Wiley & Sons, Ltd.
J Label Compd Radiopharm 2005; 48: 1041–1047

FACILE AND EFFICIENT SYNTHESIS OF d
3
-LABELLED REYATAZTM
1047
Method: Column: XTERRA C MS column, 3.5 mm (4:6 ꢀ 150 mm).
18
Mobile phase A: 90% water, 10% CH CN, 0.075% TFA.
3
Mobile phase B: 10% water, 90% CH CN, 0.075% TFA.
3
Program: Isocratic (100% A) 10 min, Gradient (100% B) 10 min. Isocratic
100% B) 10 min, Gradient (100% A) 5 min, flow rate: 1 ml/min, injection size
(
1
0 ml.
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J Label Compd Radiopharm 2005; 48: 1041–1047