Synthesis of the 26-Membered Core of Thiopeptide Natural Products by Scalable Thiazole-Forming Reactions of Cysteine Derivatives and Nitriles

Article abstract of DOI:10.1055/s-0040-1706478

The increased resistance of bacteria to clinical antibiotics is one of the major dilemmas facing human health and without solutions the problem will grow exponentially worse. Thiopeptide natural products have shown promising antibiotic activities and provide an opportunity for the development of a new class of antibiotics. Attempts to directly translate these compounds into human medicine have been limited due to poor physiochemical properties. The synthesis of the core structure of the 26-membered class of thiopeptide natural products is reported using chemistry that enables the synthesis of large quantities of synthetic intermediates and the common core structure. The use of cysteine/nitrile condensation reactions followed by oxidation to generate thiazoles has been key in enabling large academic scale reactions that in many instances avoided chromatography further aiding in accessing large amounts of key synthetic intermediates.

Full text of DOI:10.1055/s-0040-1706478

SYNTHESIS
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Georg Thieme Verlag KG Rüdigerstraße 14, 70469 Stuttgart
2
020, 52, A–K
feature
A
en
Synthesis
T. C. Johnson et al.
Feature
Synthesis of the 26-Membered Core of Thiopeptide Natural
Products by Scalable Thiazole-Forming Reactions of Cysteine
Derivatives and Nitriles
Trevor C. Johnsona
Mitchell P. Christya
Dionicio Siegel*b
Me
S
NH OH
N
O
Me
HN
Me
HO
O
NH
N
a
O
Department of Chemistry & Biochemistry, University of
chromatography-free
S
OH
S
California-San Diego, 9500 Gilman Drive, La Jolla,
California 92093, United States
Skaggs School of Pharmacy and Pharmaceutical Sciences,
MeO2C
L-threonine
N
O
N
Me
NH2•HCl
multi-decagrams
b
S
N
NH
S
University of California-San Diego, 9500 Gilman Drive,
La Jolla, California 92093-0934, United States
drsiegel@health.ucsd.edu
N
N
S
OH
CO2Me
>
200 mg
DSe Mki oa agn i Cgil csodi orSr rscSeiheis eopg goeo eln@l od fhi nPe gha laAt rhum. tu hac cos ydr .ae nd du Pharmaceutical Sciences, University of California-San Diego, 9500 Gilman Drive, La Jolla, California 92093-0934, United States
Received: 15.07.2020
Accepted after revision: 31.08.2020
Published online: 15.10.2020
opment of new chemical entities represent a significantly
better solution and is expected to have the largest impact
on the current resistance dilemma.
DOI: 10.1055/s-0040-1706478; Art ID: ss-2020-z0348-fa
An approach to develop a new class of antibiotics for use
in humans has been inspired by the natural product nosi-
heptide. Nosiheptide, initially named multhiomycin, was
first identified as a member of the thiazolyl peptide (thio-
peptide) class of antibiotics and reported to possess potent
Abstract The increased resistance of bacteria to clinical antibiotics is
one of the major dilemmas facing human health and without solutions
the problem will grow exponentially worse. Thiopeptide natural prod-
ucts have shown promising antibiotic activities and provide an opportu-
nity for the development of a new class of antibiotics. Attempts to di-
rectly translate these compounds into human medicine have been
limited due to poor physiochemical properties. The synthesis of the
core structure of the 26-membered class of thiopeptide natural prod-
ucts is reported using chemistry that enables the synthesis of large
quantities of synthetic intermediates and the common core structure.
The use of cysteine/nitrile condensation reactions followed by oxidation
to generate thiazoles has been key in enabling large academic scale re-
actions that in many instances avoided chromatography further aiding
in accessing large amounts of key synthetic intermediates.
2
antibiotic activity in 1970. The bactericidal activity occurs
through the inhibition of ribosomal protein synthesis in a
manner distinct from other clinical protein synthesis inhib-
3
itors. Nosiheptide has pronounced antibiotic effects
against a large number of clinically relevant strains includ-
4
ing those derived from patients. Potent activity is present
against numerous methicillin-resistant Staphylococcus au-
reus (MRSA) and other multidrug resistant strains. Single
digit, nanomolar activity is achieved against Clostridium dif-
ficile (C. difficile), a difficult bacterium to kill. Remarkably
no toxicity is seen when dosing animals at the extremely
Key words thiopeptide, natural product, thiazole, scalable, antibiotic,
pharmacophore
5
Resistance to clinical antibiotics has emerged as one of
the major challenges confronting global health. As a result,
this problem has become a top priority for international
governing bodies. The Centers for Disease Control and Pre-
vention estimates annually, in the United States, 2 million
adults infected with drug-resistant bacteria and, of those
high level of 2.5 g/kg. As a result, the compounds have
been used outside of the United States to increase feed con-
6
version in livestock. With desirable activity and no appar-
ent toxicity, chemistry is now needed to deliver derivatives
that possess improved solubility but maintain the potent
antibacterial effects. With the characterization of the po-
tential pharmacophore of nosiheptide and related thiazolyl
peptide natural products, total syntheses achieved for some
of the most structurally complex members, and paths for-
ward for development, now is the opportune time to ad-
1
infected, there is a 12% morbidity rate. These numbers will
increase yearly. Antibiotic resistance mechanisms, many of
which can be transferred between different bacteria, have
greatly outpaced drug development, as a result of this dis-
ease being deprioritized by the pharmaceutical industry.
Government efforts to incentivize the development of new
antibiotics have been approached in large part by rejuve-
nating existing clinically employed antibiotics providing a
lower barrier for entering the market. However, the devel-
7,8
vance this compound class through chemistry.
Toward the goal of developing new derivatives the
structural requirements for antibiotic activity are required.
Through significant effort by multiple research groups an
understanding of the pharmacophore of thiazolyl peptides
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K
Georg Thieme Verlag KG, Rüdigerstraße 14, 70469 Stuttgart, Germany

B
Synthesis
T. C. Johnson et al.
Feature
similar to nosiheptide has been developed. Key structural
similarities across this class are highlighted in red. In addi-
tion to the compounds shown in Figure 1, the related natu-
ral products thiostrepton, nocathiacins, and siomycins also
possess a similar core, a 26-membered macrocycle with
multiple thiazoles. Recently identified or prepared mem-
the macrocycle, however, as expected these compounds
1
2
were devoid of antibiotic activity. Evolved cross-resis-
tance to the parent natural products has provided informa-
tion with respect to interactions with the ribosome.13 Semi-
synthesis of active analogues, starting from QN3323A (YM-
266183), have undoubtedly built upon these findings.8
The total syntheses of thiazolyl peptide antibiotics have
been achieved with synthetic creativity and through these
studies numerous transformations have been developed
and adapted. To date, arguably, the most structurally chal-
lenging thiazolyl peptide natural product to be synthesized
,10
9
bers of this class include lactocillin from the human micro-
biome (absolute configuration yet to be reported) and
QN3323A (YM-266183),10 a compound undergoing devel-
opment.
Important structure-activity relationships have been
achieved accessing unnatural thiocillins and thiostrepton
derivatives through prepeptide gene replacement. These
compounds were tested for structural modifications to the
antibiotics and were achieved by exchanging amino acids
within the natural products precursors and determining
1
4
is thiostrepton by Nicolaou and co-workers. The total syn-
1
5
thesis of nosiheptide has also been achieved after signifi-
cant efforts were made in an attempt to prepare it through
1
6
total synthesis. In addition, many notable accomplish-
ments have been made through the total syntheses of other
1
1
17
the new, designed compounds’ activity. Remarkably the
use of prepeptide gene replacement techniques has even
yielded thiocillin derivatives with variations in the size of
thiazolyl peptide natural products. While a great deal of
chemistry has been learned from these syntheses the appli-
cation of these findings to drug development is still devel-
Biographical Sketches
Trevor Johnson was born in
Sacramento, California. He
completed his undergraduate
degree at the University of Cali-
fornia, Santa Cruz in Biochemis-
try and Molecular Biology and
completed research under the
mentorship of Professor Needhi
Bhalla (Molecular Biology). He
joined the labs of Professor Di-
onicio Siegel in 2012, where his
research focused on the total
synthesis of complex natural
products, bioactive small mole-
cules, and reaction methodolo-
gy and he received his Ph.D. in
chemistry in 2016. He joined
the Process Chemistry Depart-
ment at Gilead Sciences in 2016
where he is currently working
on the development of innova-
tive therapeutics for patients
worldwide.
Mitchell P. Christy was born
in Houston, Texas and complet-
ed his undergraduate degree in
chemistry (B.S. 2014) from the
University of Texas at Austin
where he undertook research in
organometallic reaction meth-
od development under Dr.
Guangbin Dong. He then
moved to California to complete
his Ph.D. in organic chemistry at
UC San Diego under the direc-
tion of Dr. Dionicio Siegel where
he worked on the total synthesis
of natural products and drug
development in the areas of ma-
laria and cancer in collaboration
with the Winzeler and Ideker
labs in the school of medicine.
He completed his Ph.D. in 2019
and is currently a postdoc in the
Gerwick lab at the Scripps Insti-
tute of Oceanography in La Jolla
where he is pursuing the isola-
tion and synthesis of novel ma-
rine natural products and
medicinal chemistry develop-
ment of protease inhibitors
against SARS-CoV-2.
Dionicio Siegel, born in 1974
in Truchas New Mexico, studied
chemistry starting at Reed Col-
lege and earned his Ph.D. in
chemistry with Andy Myers at
Harvard University. Postdoctor-
al work with Samuel Danishef-
sky at the Memorial Sloan-
Kettering Cancer Center was
followed, from 2007 to 2014,
with a faculty position at The
University of Texas at Austin. In
2014 he transitioned to the Uni-
versity of California, San Diego
in the Skaggs School of Pharma-
cy and Pharmaceutical Sciences
where he is currently Chair of
the Division of Pharmaceutical
Chemistry. His research inter-
ests focus on synthetic organic
chemistry applied to natural
product-based drug discovery
and medicinal chemistry.
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

C
Synthesis
T. C. Johnson et al.
Feature
Me
MeO
S
Me
NH
S
O
O
NH
OH
Me
Me
OH
Me
N
Me
N
N
O
O
HN
O
NH
N
Me
O
HN
NH
N
O
O
S
O
O
O
S
N
O
OH
OH
O
N
O
O
S
N
OH
S
N
NH
O
N
O
O
NH
N
Me
N
N
S
S
N
H
O
S
S
N
H
NH2
N
S
NH2
N
H
O
O
O
OH
O
thiazomycin
nosiheptide
Me
MeO
S
NH
OH
N
Me
O
HN
Me
NH
OH
O
S
NH
N
O
S
O
OH
N
O
O
N
OH
O
Me
NH
S
N
O
MeO
HN
NH
O
N
O
O
HO
Me
O
HO
N
O
S
S
S
N
H
S
N
NH2
Me
N
N
HN
Me
H
O
S
N
O
O
Me
O
O
O
NH
N
N
O
OMe
N
MeO
Me
OH
S
S
S
O
S
HN
OMe
OH
O
philipimycin
lactocillin
Me
Me
NH
S
S
NH
OH
Me
OH
Me
N
N
Me
NH
Me
NH
O
O
HN
HN
Me
HO
Me
O
NH
N
O
Me
R
Me
O
NH
OH
O
O
S
S
O
N
O
N
N
O
O
S
N
S
N
N
H
NH
N
O
NH
N
O
N
N
N
H
Me
Me
N
N
Me
Me
S
S
S
S
S
S
OH
OR1
QN3323 A (YM-266183)
thiocillin I R = OH, R1 = H
thiocillin II R = OH, R1 = Me
micrococcin R = H, R1 = H
Figure 1 Structures of 26-membered thiazolyl peptides known or predicted to target the L11 protein/23S rRNA region of the ribosome. Related core
structures are highlighted in red.
oping. In our group, we have succeeded synthesizing micro-
coccin P1 implementing cysteine nitrile condensation reac-
tions to form thiazoles.18 This provided guidance in the
synthesis of the common core of the 26-membered thio-
peptides that we believe to be a fundamentally important
substrate to assess structure-activity relationships present
in this class of natural products as they are related to anti-
proliferative effects.
Our approach to synthesizing the fragments of thiazolyl
peptides, chiral carboxyaminothiazoles, used a general
strategy that has proven both reliable and scalable; cyste-
1
9
ine-nitrile condensation reactions followed by aromatiza-
20
tion. For comparison, the Hantzch thiazole synthesis has
proven useful in the past for the syntheses of this class of
natural products as the method is highly reliable, however,
conducting these thiazole forming reactions on the multi-
decagram scale proved difficult in our hands in maintaining
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

D
Synthesis
T. C. Johnson et al.
Feature
yields achieved at smaller scales. As the cysteine-nitrile
condensation links easily into the syntheses of all thiazoles
except for one we also utilized the addition of thiazole-
based Grignard reagents into chiral N-sulfinylimines, which
has similarly proven reliable and scalable.21
reagents. Simple acidic liberation of the amino alcohol pro-
vides 4 as the hydrochloride salt with the acid masked as
the methyl ester.
Coupling of the amino alcohol 4 to a compound generat-
ed earlier in the route, carboxylic acid 1, proceeded un-
eventfully using EDC and HOBt to generate amide 5
(Scheme 2). Dehydrative elimination to form alkene 6 on
large scale failed to provide good yields under multiple con-
ditions used for the syntheses dehydroamino acids, howev-
er, in situ formation of the tert-butyl carbonate followed by
reaction with DBU led to facile elimination and the reaction
scaled well, conducted as shown on 28 grams of material.22
Saponification of the ester of 6 generated carboxylic acid 7
that was combined with the previously synthesized amino
alcohol 4 using EDC/HOBt coupling to yield amide 8. The re-
sulting amide product 8 was then hydrolyzed at the thi-
azole methyl ester to yield carboxylic acid 9.
As an example, starting from threonine in a seven-step
sequence that does not require chromatography we accessed
3
0 grams of amino alcohol 4 as its hydrochloride salt
(
Scheme 1). The cysteine/nitrile condensation onto nitrile 2
followed by aromatization with trichlorobromomethane
and DBU was conducted on a 38 gram scale to generate
pure thiazole 3. The reaction was both clean and the prod-
uct readily separated from reactants and by-products. No-
tably, the purification is greatly simplified compared to the
Hantzch thiazole synthesis that employs Lawesson’s re-
agent and results in the generation of poorly behaved spent
1
. ethyl chloroformate, Et3N
THF, 23 °C then
aq NH4OH
Me
Me
1
. Boc2O, NaHCO3
MeOH/H2O, 23 °C
HO2C
BocN
NC
BocN
L-threonine
O
O
2
. PPTS, 2,2-DMP, 85 °C
2. cyanuric chloride
DMF, 23 °C
Me
Me
Me
Me
8
2
5% 2 steps
00 gram scale
74% 2 steps
58 gram scale
1
2
S
Me
S
OH
Me
1
2
. L-cysteine methyl ester•HCl
pH 7 buffer/2-PrOH, 50 °C
MeO2C
4 M HCl, H O
MeO2C
2
N
N
O
1
,4-dioxane, 23 °C
99%
. BrCCl3, DBU
CH2Cl2, 23 °C
BocN
NH2
•HCl
Me
Me
4
8
3
5% 2 steps
8 gram scale
3
30 gram scale
Scheme 1 Chromatography-free, multi-decagram synthesis of chiral carboxyaminothiazole 4
Me
HO2C
O
BocN
Me
HO
N
Me
Me
1
Me
O
Me
O
Me
S
OH
Boc2O, DMAP, DBU
MeCN, 23 °C
MeO2C
EDC•HCl, HOBt, DIPEA
DMF, 23 °C
S
S
N
H
N
H
N
Me
NH2•HCl
O
O
BocN
N
BocN
Me
Me
MeO2C
Me
4
85%
5 gram scale
MeO2C
Me
68%
28 gram scale
3
5
6
S
OH
Me
MeO2C
O
Me
N
Me
Me
S
O
Me
O
N
H
4
NH2•HCl
O
NaOH
THF/MeOH/H2O, 23 °C
S
N
N
BocN
EDC•HCl, HOBt, DIPEA
N
H
Me
Me
HO
HN
S
Me
BocN
DMF, 23 °C
76%
O
Me
HO2C
Me
99%
N
4
gram scale
1
0 gram scale
7
CO2R
NaOH
THF/MeOH/H2O, 23 °C
8%
8
9
R = Me
R = H
8
Scheme 2 Five step conversion of amino alcohol 4 into acid 9
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

E
Synthesis
T. C. Johnson et al.
Feature
2
4
Synthesis of the double thiazole-substituted pyridyl
core was also greatly simplified using the cysteine/nitrile
condensation-aromatization sequence (Scheme 3). Starting
from inexpensive 2-chloro-3-pyridinecarbonitrile (10) sev-
en steps arrived at the appropriately modified pyridine
with the differentially protected esters, compound 14. Con-
densation of cysteine and 2-chloro-3-pyridinecarbonitrile
from 2,4-dibromothiazole (Turbo Grignard exchange )
provided a 4:1 ratio of diastereomers and a 66% isolated
yield of the major diastereomer.
Coupling of the three fragments chloropyridine 14,
stannane 18, and acid 9 followed by macrocyclization gen-
erated the core structure triol ester 21 (Scheme 5). Stille
coupling of chloropyridine 14 and stannane 18 was optimal
using Pd (dba) with Cy-JohnPhos, cleanly generating the
(
10) and oxidation of the thiazoline generated thiazole 11.
2
3
Protection of the free acid as the tert-butyl ester and oxida-
tion of the pyridine nitrogen to the N-oxide proceeded
tri-thiazole bearing pyridine 19. Selective cleavage of the
tert-butylsulfinyl group (and TBS) with hydrochloric acid
provided an amine that was directly coupled to carboxylic
acid 9 using HATU to provide the cyclization precursor in
protected form, compound 20. Deprotection of both Boc
groups and acetonide was followed by cyclization, again us-
ing HATU, providing the desired triol compound 21 in 27%
yield over two steps, conducted on the gram scale.
23
smoothly setting up a modified Reissert reaction with
trimethylsilyl cyanide and diethylcarbamoyl chloride to
provide nitrile 13 as a crystalline, off-white solid. The se-
quence was scalable with all intermediates crystallized
from the reaction mixture or purified by trituration. A sec-
ond cysteine/nitrile condensation followed by MnO oxida-
2
tion (proved optimal for this system) generated the second
appended thiazole ester.
With access to ample quantities of core compound 21,
which we envision to be the key pharmacophore for the 26-
membered thiopeptide natural products functionalization
can be achieved through modifications to the ester position
and the primary alcohol. This follows from previous efforts
to increase the water solubility of thiazolyl peptides, which
succeeded in developing an antibiotic that reached Phase II
clinical trials for the treatment of C. difficile, LFF571.25 The
compound, developed by Norvartis, function through a
The last required fragment possessed a thiazole with an
alternative substitution pattern. Although a Hunsdiecker
reaction could, in theory, generate this compound following
a cysteine/nitrile condensation-aromatization the broad
utility of the Ellman auxiliary21 led us to the route shown in
Scheme 4 for the synthesis of the coupling fragment, stan-
nane 18. Diastereoselective delivery of the Grignard derived
2
t-BuO C
2
HO C
1
. cysteine•HCl
pH 7 buffer/2-PrOH, 50 °C
1. TsCl, pyridine
N
NC
Cl
N
t-BuOH, 23 °C
S
S
N
2. BrCCl
3
, DBU
2. UHP, TFAA
+
N
DMF, 50 °C
CH
2 2
Cl , 23 °C
Cl
1
0
Cl
N
O
8
5
7% 2 steps
0 gram scale
8
6% 2 steps
1
1
45 gram scale
12
O
2
t-BuO C
2
t-BuO C
N
Cl
NEt
2
1. cysteine methyl ester•HCl
pH 7 buffer/2-PrOH, 50 °C
N
TMSCN
S
S
MeCN, 80 °C
2. MnO2
S
Cl
N
CH
2
Cl
2
, 23 °C
53%
Cl
N
CN
N
3
5 gram scale
64% 2 steps
13
2
CO Me
1
1 gram scale
1
4
Scheme 3 Tri-substituted thiazole-pyridine 14 synthesis from commercial pyridine 10
1
. i-PrMgCl•LiCl
THF, 23 °C
Me Sn2
Pd(PPh3)4
O
6
O
S
N
SnMe3
Br
S NH
S
NH
N
S
N
S
Br
t-Bu
t-Bu
2.
O
PhMe, 110 °C
60%
Br
TBSO
TBSO
S
15
N
t-Bu
1
7
18
2
gram scale
TBSO
H
16
CH2Cl2, –50 to 23 °C
6
6%
2
9 gram scale
Scheme 4 Diastereoselective Grignard addition into chiral N-sulfinylimine 16 and stannylation of bromide 17 generating 18
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

F
Synthesis
T. C. Johnson et al.
Feature
SnMe3
O
H
N
N
S
t-BuO2C
t-BuO2C
S
t-Bu
TBSO
18
N
N
1
2
. HCl
S
Pd2(dba)3, Cy-JohnPhos
O
S
N
1,4-dioxane/MeOH, 23 °C
S
NH
S
S
Cl
N
PhMe, 100 °C
95%
t-Bu
TBSO
N
Me
.
O
Me
O
N
N
S
432 mg scale
S
N
H
CO2Me
2
CO Me
1
4
19
N
BocN
Me
Me
HO
HN
S
Me
O
N
9
CO2H
Me
NH
Me
HATU, DIPEA
DMF, 23 °C
O
Me
O
S
79% 2 steps
910 mg scale
S
OH
Me
N
N
H
N
O
O
Me
HO
H
N
BocN
1
. TFA
H O, 23 °C
2
HN
t-BuO2C
Me
HO
O
NH
N
Me
Me
N
O
S
2
. HATU, DIPEA
DMF, 23 °C
N
S
N
O
S
N
27% 2 steps
S
N
NH
S
1 gram scale
NH
N
S
O
N
N
HO
S
N
S
CO2Me
OH
CO2Me
20
2
1
Scheme 5 Fragment coupling and completion of the synthesis of the 26-membered macrocycle 21
different mechanism of ribosome inhibition and like most
of the thiazolyl peptides was initially too insoluble for de-
velopment.
tium or carbon of the solvent; CHCl (H = 7.26) and CDCl ( = 77.0).
3 3 C
For DMSO-d
enced to residual protium or carbon of the solvents; (CD )(CHD )SO
solutions the chemical shifts are reported as ppm refer-
6
3
2
(
H = 2.50) or (CD ) SO ( = 39.5). For CD OD solutions the chemical
3 2 C 3
shifts are reported as ppm referenced to residual protium or carbon of
the solvents; CHD OD (H = 3.31) or CD OD ( = 49.0). Coupling con-
2
3
C
All reactions were performed in flame-dried round-bottomed flasks
1
stants are reported in hertz (Hz). Data for H NMR spectra are report-
ed as follows: chemical shift [ppm, referenced to protium; multiplici-
ty (standard abbreviations), coupling constant (Hz), and integration].
Melting points were measured on a MEL-TEMP device without cor-
rections.
fitted with rubber septa under a positive pressure of argon or N , un-
2
less otherwise indicated. Air and moisture sensitive liquids and solu-
tions were transferred via syringe or cannula. Organic solutions were
concentrated by rotary evaporation at 20 torr in a water bath heated
to 40 °C, unless otherwise noted. Et O, CH Cl , THF, and toluene
2
2
2
(PhMe) were purified using a Pure-Solv MD-5 Solvent Purification
(
4S,5R)-3-(tert-Butoxycarbonyl)-2,2,5-trimethyloxazolidine-4-
System (Innovative Technology). MeCN, DMF, and MeOH were pur-
chased from Acros (99.8%, anhyd) and EtOH was purchased from
Pharmco-Aaper (200 proof, absolute). The molarity of n-BuLi was de-
termined by titration against diphenylacetic acid. All other reagents
were used directly from the supplier without further purification, un-
less otherwise noted. Analytical TLC was carried out using 0.2 mm
commercial silica gel plates (silica gel 60, F254, EMD chemical) and
visualized using a UV lamp and/or aqueous ceric ammonium molyb-
date (CAM), aqueous KMnO4 stain, or ethanolic vanillin. IR spectra
were recorded on a Nicolet 380 FTIR using neat thin film technique.
High-resolution mass spectra (HRMS) were recorded on a Karatos
MS9 or Agilent Technologies 6530 Accurate-Mass Q-TOF LC/MS and
are reported as m/z (relative intensity). Accurate masses are reported
carboxylic Acid (1)
Solid L-threonine (100 g, 839 mmol, 1 equiv) was dissolved in 1.7:1
THF/2 M NaOH and cooled in an ice bath. Boc anhydride was added as
a solid portionwise (220g, 1.01 mol, 1.2 equiv) over 10 min and the
reaction mixture was stirred for 1 h. The reaction vessel was removed
from ice bath and stirred at 23 °C for 48 h. THF was removed under
vacuum, the residue taken up in aq 2 M HCl (2 L), and the mixture was
extracted with EtOAc (3 × 1 L). The combined organic layers were
washed with H O (750 mL) and brine (750 mL), dried (Na SO ) and
2
2
4
concentrated to provide a colorless oil. The crude material (175.4 g)
was used directly in the next reaction without further purification.
Crude Boc-L-threonine was dissolved in 2,2-dimethoxypropane (1.03
L) and PPTS (recrystallized from acetone) was added as a solid in one
portion (60.3 g, 0.3 equiv) and the reaction mixture was heated to re-
flux for 14 h. The mixture was cooled to 23 °C and then concentrated
and the residue was dissolved in EtOAc (2 L). The EtOAc solution was
+
+
1
13
for the molecular ion [M + Na] , [M + H] , [M], or [M – H]. H and
C
1
NMR) were recorded on a Varian Gemini spectrometer [(400 MHz, H
at 400 MHz, 13C at 100 MHz), (500 MHz, H at 500 MHz, C at 125
1
13
1
13
MHz), (600 MHz, H at 600 MHz, C at 150 MHz)]. For CDCl solutions
3
the chemical shifts are reported as ppm referenced to residual pro-
washed with H O (500 mL) and brine (500 mL), dried (Na SO ), and
2
2
4
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

G
Synthesis
T. C. Johnson et al.
Feature
concentrated. Recrystallization of the residue from hexanes yielded
13.1 g of product and an additional 58 g collected from a 2nd recrys-
tallization to provide 1 as a white solid (85% yield over 2 steps).
The reaction was allowed to warm to 23 °C as the ice bath melts. After
completion, the mixture was poured into aq 1 M HCl (500 mL) and
extracted with additional DCM (3 × 200 mL). The combined organic
1
1H NMR (600 MHz, MeOD):  (rotamers) = 4.21–4.13 (m, 1 H), 3.90–
layers were washed with H
2
O (250 mL) and brine (250 mL), dried
(
Na SO ), and concentrated to give 47.8 g of thiazole 3 as an off-white
3.80 (m, 1 H), 1.58 (br s, 3 H), 1.54 (br s, 3 H), 1.45 (br d, 9 H), 1.37 (d,
2
4
solid (134 mmol, 85% over 2 steps); mp 120–123 °C. The crude mate-
rial was of sufficient purity to be used directly in the next reaction
J = 6.1 Hz, 3 H).
without further purification.
tert-Butyl (4R,5R)-4-Cyano-2,2,5-trimethyloxazolidine-3-carbox-
ylate (2)
1
H NMR (600 MHz, CDCl ):  = 8.17 (br s, 1 H), 4.78 (m, 1 H), 4.16 (m,
3
1
H), 3.94 (s, 3 H), 1.69 (br s, 6 H), 1.42 (br s, 9 H), 1.18 (br s, 3 H).
13C NMR (150 MHz, CDCl
):  = 173.3, 161.3, 151.0, 146.1, 127.3, 95.0,
80.4, 77.6, 65.7, 52.1, 27.8, 26.2, 25.6, 17.6.
HRMS (ESI): m/z calcd for C16H24N O SNa [M + Na] : 379.1298; found:
Solid 1 (58 g, 224 mmol, 1 equiv) was dissolved in THF (447 mL, 0.5
M) and cooled in an ice bath. Ethyl chloroformate (25.8 mL, 269
3
mmol, 1.2 equiv) was added followed by dropwise addition of Et N
3
(37.5 mL, 269 mmol, 1.2 equiv) with vigorous stirring over 20 min to
+
2
5
ensure stirring was not hindered. After the addition of ethyl chloro-
formate, the reaction mixture was warmed to 23 °C and stirred for 4 h
when full consumption of starting material was observed by TLC
3
79.1295.
Methyl 2-[(1S,2R)-1-Amino-2-hydroxypropyl]thiazole-4-carboxyl-
ate Hydrochloride (4)
(
2
ninhydrin stain). The mixture was cooled again in an ice bath and
5% aq NH OH (48.8 mL, 1.4 equiv) was added in a single portion and
4
1
,4-Dioxane (1 mL/g) was added to thiazole 3 (30 g, 84 mmol, 1 equiv)
the mixture was stirred with slow warming over 12 h. The solvent
was removed under reduced pressure and the residue was dissolved
to solubilize the material. A 4 M solution of HCl in 1,4-dioxane was
added (105 mL, 5 equiv) followed by dropwise addition of distilled
H O (11 mL, 10% v/v). The reaction mixture was stirred at 23 °C for 2
h. An oil or solid might appear to precipitate from the reaction, which
was the desired HCl salt. The mixture was concentrated from 4:1 ben-
zene/MeOH (3 × 100 mL). The solid crude material was of sufficient
purity to be used directly in the next reaction without further purifi-
cation (99%).
1H NMR (600 MHz, MeOD):  = 8.54 (s, 1 H), 4.88 (s, 1 H), 4.76 (d, J =
6
13C NMR (150 MHz, MeOD):  = 165.2, 162.9, 147.3, 131.7, 68.5, 58.8,
in EtOAc (1 L). The combined extracts were washed with H O (2 × 350
2
2
mL) and brine (350mL), dried (Na SO ), and concentrated to provide
2
4
an amber oil (48.8 g). This crude material was used in the next reac-
tion without further purification.
Crude amide (51.4 g, 199 mmol, 1 equiv) was dissolved in DMF (200
mL, 1.0 M) and the reaction flask was immersed in a 23 °C water bath.
Solid cyanuric chloride (18.44 g, 100 mmol, 0.5 equiv) was added in
portions over 10 min and stirred for 1 h. The reaction mixture was
poured slowly into ice water (2 L) with vigorous stirring and the solids
were collected by filtration. The precipitate was washed with cold
.6 Hz, 1 H), 4.32–4.26 (m, 1 H), 3.92 (s, 3 H), 1.23 (d, J = 6.4 Hz, 3 H).
H O (3 × 250 mL) and dried in vacuo to give the nitrile 2 as a white
53.0, 19.9.
2
solid (35.4 g, 147 mmol, 74% over two steps); mp 41–43 °C.
+
HRMS (ESI): m/z calcd for C16H24N O SNa [M + Na] : 239.0461; found:
239.0463.
2
5
IR (film): 2358, 1715 cm–1
.
1
H NMR (600 MHz, CDCl ):  = 4.40 (pent, J = 6.2 Hz, 1 H), 3.99 (m, 1
3
tert-Butyl (4S,5R)-4-({(1S,2R)-2-Hydroxy-1-[4-(methoxycarbon-
yl)thiazol-2-yl]propyl}carbamoyl)-2,2,5-trimethyloxazolidine-3-
carboxylate (5)
H), 1.59 (br s, 3 H), 1.52 (br s, 4 H), 1.48 (s, 9 H), 1.40 (d, J = 6.1 Hz, 3
H).
1
3
C NMR (150 MHz, CDCl ):  = 150.4, 117.1, 95.7, 82.0, 74.1, 52.9,
3
Amine 4 (35.3 g, 140 mmol, 1 equiv) was dissolved in DMF (280 mL,
28.1, 26.4, 24.4, 18.2.
0
.5 M) along with protected threonine 1 (39.9 g, 154 mmol, 1.1
equiv), EDC hydrochloride (32.2 g, 168 mmol, 1.2 equiv), and HOBt
22.7 g, 168 mmol, 1.2 equiv, 80% w/w). DIPEA was added dropwise
+
HRMS (ESI): m/z calcd for C12H20N O Na [M + Na] : 263.1366; found:
2
3
263.1366.
(
over 5 min (73 mL, 420 mmol, 3 equiv) and the reaction mixture was
stirred to completion at 23 °C for 14 h. The mixture was diluted with
tert-Butyl (4S,5R)-4-[4-(Methoxycarbonyl)thiazol-2-yl]-2,2,5-
trimethyloxazolidine-3-carboxylate (3)
H O (2 L) and extracted with EtOAc (3 × 500 mL). The combined or-
2
Nitrile 2 (37.7 g, 157 mmol, 1.0 equiv) was dissolved in a 1.5:1 mix-
ture of i-PrOH/pH 7 phosphate buffer (785 mL, 0.2 M buffer, 0.1 M
concentraion) and solid cysteine methyl ester hydrochloride was add-
ed in a single portion (40.4 g, 236 mmol, 1.5 equiv). The reaction mix-
ture was stirred and heated to 50 °C for 14 h. The solvent was re-
moved under reduced pressure and the residue was partitioned be-
ganic layers were washed with aq 3 M LiCl (3 × 300 mL), dried
(Na SO ), and concentrated. The crude material was purified by col-
2
4
umn chromatography with 60 → 80% EtOAc/hexane on a short length
column; the desired product 5 was collected pure after chromatogra-
phy (54.5 g, 119 mmol, 85%); white foam.
1
H NMR (600 MHz, CDCl ):  = 7.98 (s, 1 H), 5.15 (br s, 1 H), 4.44 (m, 1
3
tween H O (1 L) and EtOAc (500 mL) and the aqueous layer was
2
H), 4.13 (br s, 1 H), 3.82 (d, J = 7.4 Hz, 1 H), 3.76 (s, 3 H), 1.48 (s, 3 H),
extracted with EtOAc (3 × 250 mL). The combined organic layers were
dried (Na SO ) and concentrated to give the thiazoline (50.9 g) as a
1
.46 (s, 3 H), 1.26 (br s, 9 H), 1.15 (d, J = 6.5 Hz, 3 H).
2
4
1
3
C NMR (150 MHz, CDCl ):  = 171.1, 170.1, 161.2, 151.9, 145.9,
3
colorless oil that solidifies upon standing. This crude material was
used directly in the next reaction without further purification.
1
1
27.6, 94.5, 80.7, 73.7, 68.7, 67.1, 55.8, 52.0, 28.0, 27.4, 25.1, 19.3,
8.7.
Crude thiazoline was dissolved in DCM (475 mL, 0.3 M) and cooled in
+
HRMS (ESI): m/z calcd for C20H31N O SNa [M + Na] : 480.1775; found:
3
7
an ice bath. BrCCl was added (21 mL, 188 mmol, 1.2 equiv) followed
3
4
80.1778.
by DBU (25.4 mL, 188 mmol, 1.2 equiv) dropwise over several min.
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

H
Synthesis
T. C. Johnson et al.
Feature
tert-Butyl (4S,5R)-4-({(Z)-1-[4-(Methoxycarbonyl)thiazol-
give the carboxylic acid 9 as a white solid. This crude material was an-
alytically pure and was used in the next reaction without further pu-
rification (>99%).
2
3
-yl]prop-1-en-1-yl}carbamoyl)-2,2,5-trimethyloxazolidine-
-carboxylate (6)
1H NMR (600 MHz, MeOD):  = 8.30 (s, 1 H), 8.19 (s, 1 H), 6.88–6.54
(m, 1 H), 5.35 (br s, 1 H), 4.55 (br s, 1 H), 4.27–4.22 (m, 1 H), 4.05 (d,
J = 8.0 Hz, 1 H), 1.91 (d, J = 7.2 Hz, 3 H), 1.59–1.39 (m, 18 H), 1.28 (d,
J = 6.4 Hz, 3 H).
Solid 5 (28.1 g, 61.4 mmol, 1 equiv) was dissolved in MeCN (205 mL,
.3 M). Boc anhydride (16.1 g, 73.7 mmol, 1.2 equiv) was added in a
0
single portion followed by DMAP (1.5 g, 12.3 mmol, 0.2 equiv) and the
reaction mixture was stirred until all starting materials were con-
sumed. After full conversion of starting materials by TLC, DBU (45.8
mL, 307.0 mmol, 5 equiv) was added dropwise at 23 °C and the mix-
ture was stirred for 14 h. The mixture was diluted with EtOAc (500
2-(2-Chloropyridin-3-yl)thiazole-4-carboxylic Acid (11)
2
-Chloronicotinonitrile (10; 50 g, 361 mmol, 1 equiv) was dissolved
mL) and washed with aq 1 M HCl (200 mL), H O (200 mL) and brine
2
in 1.5:1 i-PrOH/pH 7 phosphate buffer (722 mL, 0.5 M) and solid L-
cysteine hydrochloride (65.6 g, 542 mmol, 1.5 equiv) was added in
one portion. The reaction mixture was heated to 50 °C and stirred for
(
200 mL), dried (Na SO ), and concentrated. The crude material was
2 4
purified by column chromatography with 15 → 35% EtOAc/hexane on
a medium length column to give 18.38 g of intermediate 6 as a color-
less oil (41.8 mmol, 68%).
1
6 h. The reaction was then terminated by removing i-PrOH under re-
duced pressure and diluting with EtOAc (500 mL) and aq 2 M HCl un-
til the solution was acidic (pH >2). The mixture was extracted with
EtOAc (3 × 400 mL), the combined extracts were dried (Na SO ), and
1
H NMR (600 MHz, CDCl ):  = 7.99 (s, 1 H), 7.96 (br s, 1 H), 6.54 (br s,
3
1
H), 4.32 (br s, 1 H), 3.97 (d, J = 7.7 Hz, 1 H), 3.85 (s, 3 H), 1.82 (d, J =
2
4
6.6 Hz, 3 H), 1.61 (br s, 6 H), 1.44 (d, J = 6.1 Hz, 3 H), 1.40 (s, 9 H).
concentrated to give the intermediate thiazoline as a yellow solid
(76.9 g, 317 mmol, 88%); mp 156–158 °C. The crude material was of
sufficient purity to be used directly in the next reaction without fur-
ther purification.
1H NMR (600 MHz, MeOD):  = 8.49 (dd, J = 4.9, 1.8 Hz, 1 H), 8.10 (dd,
J = 7.7, 1.8 Hz, 1 H), 7.49 (dd, J = 7.7, 4.9 Hz, 1 H), 5.36 (t, J = 9.2 Hz, 1
H), 3.87–3.80 (m, 2 H).
1
3
C NMR (150 MHz, CDCl ):  = 168.4, 167.3, 161.7, 152.3, 146.7,
3
127.9, 127.1, 95.1, 81.1, 74.2, 67.8, 52.3, 28.3, 27.7, 25.5, 19.0, 14.4.
+
+
HRMS (ESI) : m/z calcd for C20H29N O SNa [M + Na] : 462.1669;
3
6
found: 462.1665.
tert-Butyl (4S,5R)-4-({(Z)-1-[4-({(1S,2R)-2-Hydroxy-1-[4-(meth-
oxycarbonyl)thiazol-2-yl]propyl}carbamoyl)thiazol-2-yl]prop-1-
en-1-yl}carbamoyl)-2,2,5-trimethyloxazolidine-3-carboxylate (8)
Crude thiazoline (76.9 g, 317 mmol, 1 equiv) and BrCCl (46.9 mL, 476
3
mmol, 1.5 equiv) were dissolved in DMF (317 mL, 1 M) in a flask and
the flask was immersed in an ice bath. DBU (99 mL, 666 mmol, 2.1
equiv) was added dropwise via an addition funnel over 20 min. On
large scale this addition was significantly exothermic. After addition,
the reaction mixture was then brought to 50 °C and stirred for 3 h.
After completion, the mixture was slowly introduced dropwise into a
vigorously stirring aq 1 M HCl (2 L) at 0 °C to ensure a uniform precip-
itate. The mixture was stirred for 10 min and the fine brown precipi-
tate was filtered and dried under vacuum at 50 °C for 18 h to give 76.0
g (99%, 87% over 2 steps) of crude thiazole product 11 as a grey-brown
solid; mp > 200 °C. The crude material was of sufficient purity to be
used directly in the next reaction without further purification.
Methyl ester 6 (10.0 g, 22.8 mmol, 1 equiv) was dissolved in a mixture
of 3:1 THF and MeOH (11.4 mL, 0.2 M) and aq 10% NaOH was added
(22.8 mL, 2.28 g, 2.5 equiv). The reaction mixture was stirred for 1 h
and poured into aq 2 M HCl (100 mL), extracted with EtOAc (3 × 50
mL); the combined organic layers were dried (Na SO ) and concen-
2
4
trated. The crude acid 7 (clear oil was used directly in the next reac-
tion without further purification (99%).
Crude acid 7 (3.83 g, 9.0 mmol, 1 equiv) was dissolved in DMF (45 mL,
0.2 M) and solid amine hydrochloride 4 was added (2.50 g, 9.9 mmol,
1.1 equiv) followed by HATU (2.07 g, 10.8 mmol, 1.2 equiv). Neat
DIPEA was added dropwise (7.84 mL, 45.0 mmol, 5 equiv) and the re-
action mixture was stirred for 12 h. The mixture was poured into
brine (500 mL) and extracted with EtOAc (3 × 125 mL). The combined
organic layers were washed with aq 3 M LiCl (3 × 100 mL), dried
1H NMR (600 MHz, MeOD):  = 8.76 (dd, J = 7.9, 1.8 Hz, 1 H), 8.58 (s, 1
H), 8.50 (dd, J = 4.7, 1.9 Hz, 1 H), 7.58 (dd, J = 7.9, 4.7 Hz, 1 H).
1
3
C NMR (150 MHz, DMSO-d ):  = 162.4, 161.6, 151.3, 148.0, 147.6,
6
(
Na SO ), and concentrated. The crude material was purified by col-
140.2, 131.1, 128.3, 124.4.
2
4
umn chromatography with 55 → 75% EtOAc/hexane on a medium
length column to give 4.25 g of 8 as a white solid (6.81 mmol, 76%
over 2 steps).
–
HRMS (ESI): m/z calcd for C H ClN O S [M – H] : 238.9687; found:
9
5
2
2
2
38.9689.
1H NMR (600 MHz, MeOD):  (rotamers) = 8.34 (s, 1 H), 8.19 (s, 1 H),
3
-[4-(tert-Butoxycarbonyl)thiazol-2-yl]-2-chloropyridine 1-Oxide
6
(
.89–6.53 (m, 1 H), 5.34 (s, 1 H), 4.53 (s, 1 H), 4.25 (s, 1 H), 4.12–4.03
m, 1 H), 3.92 (s, 3 H), 1.91 (dd, J = 6.8, 2.2 Hz, 3 H), 1.59–1.26 (m, 21
H).
(
12)
Thiazolyl pyridine 11 (45.4 g, 189 mmol, 1 equiv) was dissolved in 3:1
t-BuOH/pyridine (756 mL, 0.25 M). TsCl (71.9 g, 2 equiv) was added
slowly at 23 °C and the reaction mixture was then stirred at 60 °C for
2
-[(1S,2R)-1-(2-{(Z)-1-[(4S,5R)-3-(tert-Butoxycarbonyl)-2,2,5-
4
h. The mixture was slowly poured into vigorously stirring distilled
trimethyloxazolidine-4-carboxamido]prop-1-en-1-yl}thiazole-4-
carboxamido)-2-hydroxypropyl]thiazole-4-carboxylic Acid (9)
H O (3 L) and stirred for 10 min. The precipitate was filtered, washed
with cold distilled H O (2 × 500 mL), and dried on the filter for 4 h to
2
2
Methyl ester 8 (462 mg, 1.17 mmol, 1 equiv) was dissolved in a 3:1
mixture of THF and MeOH (7.5 mL) and aq 2 M NaOH was added (0.74
mL, 2.93 mmol, 2.5 equiv) and the reaction mixture was stirred for 1
h. After full conversion of starting material, the mixture was acidified
with aq 2 M HCl (40 mL) and extracted with EtOAc (3 × 25 mL). The
combined organic layers were dried (Na SO ) and concentrated to
give the t-Bu ester as a brown solid (51.83 g, 93%); mp 115–117 °C.
Crude t-Bu ester (10.36 g, 34.9 mmol, 1 equiv) and urea-H O com-
2
2
plex (6.57 g, 69.8 mmol, 2 equiv) were dissolved in anhyd DCM (175
mL, 0.2 M) in a flame-dried flask equipped with a stir bar. Trifluoro-
acetic anhydride (TFAA; 9.71 mL, 69.8 mmol, 2 equiv) was added
dropwise over 30 min via an addition funnel to the reaction mixture
2
4
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

I
Synthesis
T. C. Johnson et al.
Feature
1
in an ice bath. The mixture was then warmed to 23 °C and stirred to
completion over 12 h. After completion, the reaction was diluted with
additional DCM (175 mL) and aq 10% K CO (175 mL). The organic lay-
H NMR (600 MHz, CDCl ):  = 8.97 (d, J = 8.2 Hz, 1 H), 8.38 (d, J = 8.2
3
Hz, 1 H), 8.35 (s, 1 H), 8.24 (s, 1 H), 4.01 (s, 3 H), 1.64 (s, 9 H).
13
2
3
C NMR (150 MHz, CDCl ):  = 167.1, 161.6, 161.0, 160.2, 150.7,
3
er was washed with H O (2 × 100 mL), sat. aq Na S O (2 × 100 mL)
2
2
2
3
148.3, 147.4, 140.8, 130.7, 129.1, 128.5, 119.1, 82.4, 52.6, 28.1.
and brine (100 mL), dried (Na SO ), and concentrated to give 10.1 g of
2
4
+
HRMS (ESI): m/z calcd for C18H16ClN O S Na [M + Na] : 438.0344;
found: 438.0346.
3
4 2
crude N-oxide as a yellow solid (32.1 mmol, 92%; 86% over 2 steps);
mp 134–136 °C. The crude material was of sufficient purity to be used
directly in the next reaction without further purification.
1H NMR (599 MHz, MeOD):  = 8.61 (dd, J = 6.5, 1.4 Hz, 1 H), 8.56 (s, 1
H), 8.36 (dd, J = 8.2, 1.4 Hz, 1 H), 7.60 (dd, J = 8.2, 6.5 Hz, 1 H), 1.63 (s,
(
R)-N-{(S)-1-(4-Bromothiazol-2-yl)-2-[(tert-butyldimethyl-
silyl)oxy]ethyl}-2-methylpropane-2-sulfinamide (17)
Solid 2,4-dibromothiazole 15 (28.9 g, 119 mmol, 1.5 equiv) was dis-
solved in THF (50 mL, ?2 mL/g) and cooled in an ice bath. A 1.3 M
solution of i-PrMgCl·LiCl in THF (98 mL, 127 mmol, 1.6 equiv) was
added dropwise over 10 min. The reaction mixture was warmed to 23
°C over 30 min. The resulting solution was added dropwise over 2 h to
a separate reaction vessel cooled to approximately –50 °C containing
a solution of chiral imine 16 (22 g, 79 mmol, 1.0 equiv) in DCM (793
mL, 0.1 M). The mixture was allowed to warm to 23 °C over 12 h, then
poured into brine (2 L), and the aqueous layer was extracted with
DCM (3 × 500 mL). The combined organic layers were dried (Na SO )
9
H).
1
3
C NMR (150 MHz, CDCl ):  = 160.0, 159.9, 148.8, 140.4, 131.6,
3
128.8, 126.8, 122.9, 82.6, 28.1.
+
HRMS (ESI): m/z calcd for C14H12ClN O S [M + Na] : 335.0228; found:
3
2
335.0221.
tert-Butyl 2-(2-Chloro-6-cyanopyridin-3-yl)thiazole-4-carboxyl-
ate (13)
The N-oxide 12 (35.1 g, 112 mmol, 1 equiv) was dissolved in anhyd
MeCN (374 mL, 0.3 M) and TMSCN (35.1 mL, 2.5 equiv) and diethyl-
carbamyl chloride (35.6 mL, 2.5 equiv) were added at 23 °C. The reac-
tion mixture was then brought to reflux and stirred for 14 h to com-
pletion. The reaction mixture was slowly poured into a vigorously
stirred ice cold solution of aq 10% K CO (1.5 L) and stirred for 10 min.
2
4
and concentrated to give an amber oil. The crude material was puri-
fied by flash chromatography with 10 → 30% EtOAc/hexane on a me-
dium length column to give 23.1 g of the chiral aminothiazole 17 as
an amber oil (52.3 mmol, 66%).
1
H NMR (600 MHz, CDCl ):  = 7.16 (s, 1 H), 4.83–4.77 (m, 1 H), 4.66
2
3
3
The brown precipitate was filtered and washed with additional cold
(d, J = 6.4 Hz, 1 H), 4.16 (dd, J = 9.8, 3.6 Hz, 1 H), 4.08 (dd, J = 9.9, 3.5
H O (3 × 200 mL). The collected solids were dissolved in EtOAc (500
Hz, 1 H), 1.30 (s, 9 H), 0.81 (s, 9 H), 0.03 (s, 3 H), –0.08 (s, 3 H).
2
mL), washed with brine (300 mL), dried (Na SO ), and concentrated.
13
2
4
C NMR (150 MHz, CDCl ):  = 173,7, 125.0, 117.3, 66.0, 59.0, 56.3,
3
The crude product was recrystallized from hexanes and EtOAc to give
9.13 g of 13 as a crystalline, slightly yellow solid. (59.5 mmol; 53%);
2
5.6, 22.5, 18.0, –5.5.
1
+
HRMS (ESI): m/z calcd for C15H29BrN O S SiNa [M + Na] : 463.0515;
found: 463.0512
2
2 2
mp 151–154 °C.
1
H NMR (600 MHz, CDCl ):  = 9.01 (d, J = 8.0 Hz, 1 H), 8.30 (s, 1 H),
3
7
.78 (d, J = 8.0 Hz, 1 H), 1.64 (s, 9 H).
(
R)-N-{(S)-2-[(tert-Butyldimethylsilyl)oxy]-1-[4-(trimethylstan-
1
3
C NMR (150 MHz, CDCl ):  = 160.0, 159.6, 149.2, 148.7, 140.4,
3
nyl)thiazol-2-yl]ethyl}-2-methylpropane-2-sulfinamide (18)
133.0, 131.6, 129.4, 127.2, 115.6, 82.8, 28.2.
Bromide 17 (2.2 g, 5.0 mmol, 1 equiv) was added to a flame-dried, N2-
filled flask and dissolved in toluene (25 mL, 0.2 M). Solid Pd(PPh3)4
(576 mg, 0.5 mmol, 0.1 equiv) and neat Me Sn (2.19 mL, 10 mmol, 2
+
HRMS (ESI): m/z calcd for C14H12ClN O SNa [M + Na] : 344.0231;
found: 344.0227.
3
2
6
2
equiv) were added and the reaction mixture was heated to 100 °C for
1 h. The mixture was then cooled to 23 °C and partially concentrated.
The remaining residue was loaded directly onto a column and puri-
fied by flash chromatography with 10 → 20% EtOAc/hexane to give
tert-Butyl 2-{2-Chloro-6-[4-(methoxycarbonyl)thiazol-2-yl]-
pyridin-3-yl}thiazole-4-carboxylate (14)
Cyanopyridine 13 (10.94 g, 34.0 mmol, 1 equiv) was dissolved in 1.5:1
i-PrOH/pH 7 phosphate buffer (340 mL, 0.2 M buffer, 0.1 M reaction)
followed by solid cysteine methyl ester hydrochloride (8.75 g, 51
mmol, 1.5 equiv) in one portion. The reaction mixture was heated to
1
.56 g of pure stannane 18 as a slightly yellow oil (2.97 mmol, 60%).
1
H NMR (600 MHz, CDCl ):  = 7.30–7.27 (m, 1 H), 4.89 (dd, J = 10.0,
3
4.2 Hz, 1 H), 4.70 (d, J = 6.1 Hz, 1 H), 4.15 (dd, J = 9.7, 4.4 Hz, 1 H), 4.07
(dd, J = 9.7, 3.9 Hz, 1 H), 1.29 (s, 9 H), 0.80 (s, 9 H), 0.39–0.28 (m, 9 H),
0.01 (s, 3 H), –0.11 (s, 3 H).
50 °C and stirred for 6 h. i-PrOH was removed under reduced pressure
and the residue was diluted with H O (300 mL) and this mixture was
2
extracted with EtOAc (3 × 100 mL). The combined organic layers were
washed with brine (100 mL), dried (Na SO ), and concentrated to give
2
4
tert-Butyl 2-[2-(2-{(1S)-2-[(tert-Butyldimethylsilyl)oxy]-1-[(tert-
butylsulfinyl)amino]ethyl}thiazol-4-yl)-6-[4-(methoxycarbon-
yl)thiazol-2-yl]pyridin-3-yl]thiazole-4-carboxylate (19)
7.2 g of crude thiazoline as a yellow solid (9.18 mmol, 65%). The crude
material was used directly in the next reaction without further purifi-
cation.
Solid chloropyridine 14 (432 mg, 0.99 mmol, 1 equiv), stannane 18
Crude thiazoline (7.2 g, 22.2 mmol, 1 equiv) was dissolved in anhyd
DCM (222 mL, 0.1 M) and activated MnO2 (42.4 g, 444 mmol, 20
equiv) was added (Alfa Aesar; tech. 90%; LOT: W08D050). The reac-
tion mixture was stirred rapidly for 18 h, then filtered through a pad
of silica gel with MeOH to give the desired product as a yellow solid
(518 mg, 0.99 mmol, 1 equiv), Pd (dba)3 (28.4 mg, 0.05 mmol, 0.05
equiv), and CycloJohnPhos (69.1 mg, 0.2 mmol, 0.2 equiv) were added
2
to a flame-dried flask and purged with N . Toluene was added (10 mL,
2
0.1 M) and the reaction mixture was heated to 100 °C and stirred for
18 h. The mixture was then cooled to 23 °C and partially concentrat-
ed. The crude residue was loaded directly onto a column and purified
by flash chromatography with 35 → 50% EtOAc/hexane on a medium
length column to give 716 mg of the coupled product 19 (0.94 mmol,
(>99%, 64% over 2 steps); mp > 200 °C. The crude material was of suf-
ficient purity to be used directly in the next reaction without further
purification.
9
5%) as a pale-yellow foam.
©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

J
Synthesis
T. C. Johnson et al.
Feature
1
H NMR (600 MHz, CDCl ):  = 8.41 (d, J = 8.2 Hz, 1 H), 8.38 (d, J = 8.2
matography with 2 → 8% MeOH/DCM on a long column, dry loaded
on silica gel. Macrocycle 21 (233 mg) was collected pure after chroma-
tography as a white solid (0.25 mmol, 27% over 2 steps); mp > 200 °C.
1
3
Hz, 1 H), 8.32 (s, 1 H), 8.07 (s, 1 H), 7.82 (s, 1 H), 4.73 (q, J = 5.0 Hz, 1
H), 4.59 (d, J = 5.6 Hz, 1 H), 4.01 (s, 3 H), 3.97–3.89 (m, 2 H), 1.62 (s, 9
H), 1.30 (s, 9 H), 0.86 (s, 9 H), 0.06 (s, 3 H), –0.02 (s, 3 H).
H NMR (600 MHz, CDCl ):  = 8.72 (s, 1 H), 8.62 (d, J = 8.1 Hz, 1 H),
3
1
3
C NMR (150 MHz, CDCl ):  = 171.6, 168.9, 164.8, 161.7, 160.1,
8.38 (d, J = 8.1 Hz, 1 H), 8.33 (s, 1 H), 8.24 (d, J = 9.2 Hz, 1 H), 8.17 (s, 1
H), 8.15 (s, 1 H), 8.03 (d, J = 8.1 Hz, 1 H), 8.00 (s, 1 H), 7.98 (s, 1 H), 7.92
(d, J = 7.7 Hz, 1 H), 6.43 (q, J = 7.0 Hz, 1 H), 5.46–5.39 (m, 2 H), 4.89
(dd, J = 7.8, 2.4 Hz, 1 H), 4.70–4.65 (m, 1 H), 4.39–4.35 (m, 1 H), 4.01
3
1
1
53.0, 150.8, 150.4, 148.3, 148.0, 140.2, 130.3, 129.4, 128.2, 121.5,
19.0, 82.1, 66.0, 56.2, 52.5, 28.1, 27.5, 22.5, 18.0.
+
HRMS (ESI): m/z calcd for C33H45N O S Si [M] : 764.2905; found:
7
5
6 4
(
s, 3 H), 3.96 (dd, J = 11.0, 2.9 Hz, 1 H), 1.82 (d, J = 7.0 Hz, 3 H), 1.48 (d,
64.2094.
J = 6.3 Hz, 3 H), 1.34 (d, J = 6.3 Hz, 3 H).
1
3
C NMR (150 MHz, CDCl ):  = 169.8, 168.9, 168.8, 168.7, 166.2,
tert-Butyl (4S,5R)-4-{[(Z)-1-(4-{[(1S,2R)-1-(4-{[(S)-1-(4-{3-[4-(tert-
Butoxycarbonyl)thiazol-2-yl]-6-[4-(methoxycarbonyl)thiazol-2-
yl]pyridin-2-yl}thiazol-2-yl)-2-hydroxyethyl]carbamoyl}thiazol-
3
1
1
1
65.8, 161.8, 161.2, 161.0, 160.4, 153.7, 150.8, 150.6, 149.8, 149.5,
48.5, 148.2, 140.3, 130.5, 128.9, 128.6, 128.3, 125.1, 124.9, 123.7,
21.5, 118.9, 69.0, 67.9, 63.7, 57.5, 54.6, 52.6, 51.4, 20.1, 19.0, 14.5.
+
2-yl)-2-hydroxypropyl]carbamoyl}thiazol-2-yl)prop-1-en-1-
yl]carbamoyl}-2,2,5-trimethyloxazolidine-3-carboxylate (20)
HRMS (ESI): m/z calcd for C37H34N10O S Na [M + Na] : 945.1006;
9 5
Trithiazolyl pyridine 19 (910 mg, 1.19 mmol, 1 equiv) was dissolved
in MeOH (6 mL, 0.2 M) and 4 N HCl in 1,4-dioxane (1.5 mL, 5.95
mmol, 5 equiv) was added and the reaction mixture was stirred at 23
found: 945.1011.
°
C for 2 h. After completion, the mixture was diluted with toluene and
Funding Information
concentrated (3 × 25 mL). The crude residue was used directly in the
next reaction without further purification.
We would like to thank the NIH/NCI Training Grant (T32 CA009523)
and UC San Diego for funding this project.Natio
n
al
I
nsitutes
o
f
H
ealht (T
3
2
C
A
0
0
9
5
2
3)U
n
iversiyt
o
f
California, San
D
iego ()
The crude amine was dissolved in DMF (11.9 mL, 0.1 M) and the frag-
ment 9 was added (800 mg, 1.31 mmol, 1.1 equiv) followed by HATU
(
498 mg, 1.31 mmol, 1.1 equiv) and DIPEA (0.62 mL, 3.57 mmol, 3
Acknowledgment
equiv). The reaction mixture was stirred for 14 h at 23 °C. The mix-
ture was diluted with H O (100 mL) and extracted with EtOAc (3 × 50
mL). The combined organic layers were washed with aq 3 M LiCl (3 ×
2
We would like to thank Dr. Brendan Duggan for NMR experiment as-
sistance and analysis. NMR spectra were collected at the UCSD Skaggs
School of Pharmacy and Pharmaceutical Sciences NMR Facility.
5
0 mL), dried (Na SO ), and concentrated. The crude material was dry
2 4
loaded on silica gel and purified by column chromatography with 2 →
% MeOH/DCM on a long column to give 1.07 g of fully assembled in-
6
termediate 20 as a yellow solid (1.07 g, 0.94 mmol; 79% over 2 steps);
mp > 200 °C.
Supporting Information
1
Supporting information for this article is available online at
H NMR (600 MHz, CDCl ):  = 8.38 (d, J = 8.1 Hz, 1 H), 8.32 (s, 1 H),
.28 (d, J = 8.1 Hz, 1 H), 8.13 (s, 1 H), 8.07 (s, 1 H), 8.04 (s, 1 H), 7.98 (s,
H), 5.41–5.37 (m, 1 H), 5.31 (dd, J = 8.8, 1.8 Hz, 1 H), 4.76–4.68 (m, 1
3
8
1
https://doi.org/10.1055/s-0040-1706478. Su
p
p
orting Inform atio
n
Su
p
p
orting Inform atio
n
H), 4.37 (s, 1 H), 4.05 (dd, J = 11.6, 2.9 Hz, 1 H), 3.85 (dd, J = 11.5, 4.1
Hz, 1 H), 1.85 (d, J = 6.7 Hz, 3 H), 1.60–1.58 (m, J = 4.5 Hz, 12 H), 1.44
References
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1
d, J = 6.1 Hz, 3 H), 1.32 (d, J = 6.4 Hz, 3 H).
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3
C NMR (150 MHz, CDCl ):  = 171.8, 168.9, 168.8, 168.3, 167.0,
3
(2) (a) Tanaka, T.; Endo, T.; Shimazu, A.; Yoshida, R.; Suzuki, Y.
1
1
9
65.1, 161.8, 161.3, 160.7, 160.6, 152.5, 150.7, 150.6, 149.1, 148.6,
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(
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2
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(
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500 mL) and washed with aq 3 M LiCl (3 × 200 mL), dried (Na SO ),
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©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K

K
Synthesis
T. C. Johnson et al.
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©
2020. Thieme. All rights reserved. Synthesis 2020, 52, A–K