Synthesis of the Deacetoxytubuvaline Fragment of Pretubulysin and its Lipophilic Analogues for Enhanced Permeability in Cancer Cell Lines

Article abstract of DOI:10.1055/s-0037-1611359

In the last two decades, tubulysins have emerged as alternatives to microtubule depolymerizing agents such as colchicine and vinblastine, which are well-established anticancer agents. However, the complex structure of tubulysins has always posed a challen

Full text of DOI:10.1055/s-0037-1611359

SYNLETT
0
9
3
6
-
5
2
1
4
1
4
3
7
-
2
0
9
6
© Georg Thieme Verlag Stuttgart · New York
2018, 29, A–E
letter
A
en
R. B. Reddy et al.
Letter
Synlett
Synthesis of the Deacetoxytubuvaline Fragment of Pretubulysin
and its Lipophilic Analogues for Enhanced Permeability in Cancer
Cell Lines
Ramesh B. Reddya
Premansh Dudhea
Venkatesh Chelvam*a,b
O
OH
N
BocN
Me
dTuv and lipophilic analogues
BocHN
N
X
O
OMe
Ts
R
a Discipline of Chemistry, Indian Institute of Technology Indore,
Khandwa Road, Simrol, Indore-453 552, India
b Discipline of Biosciences and Biomedical Engineering, Indian
Institute of Technology Indore, Khandwa Road, Simrol, Indore-
453 552, India
aziridine
g-amino acid
cvenkat@iiti.ac.in
Received: 18.09.2018
Accepted after revision: 26.10.2018
Published online: 06.12.2018
tubulysins.3 Structurally, tubulysins are linear tetrapeptides
containing four amino acid fragments: N-methylpipecolic
acid (D-Mep), L-isoleucine (Ile), and two unusual amino
acids, tubuvaline (Tuv) and tubutyrosine (Tut) or tubu-
phenylalanine (Tup). Moreover, during the biosynthesis of
tubulysins, D-Mep, L-Ile, deacetoxytubuvaline (dTuv), and
Tut or Tup are sequentially assembled to form pretubulysin
A or D, respectively, as precursor. Subsequently, a biocata-
lyzed oxidation/acylation reaction transforms the pretu-
bulysins into tubulysins through the introduction of an ace-
tate group at C-11.3
DOI: 10.1055/s-0037-1611359; Art ID: st-2018-d0603-l
Abstract In the last two decades, tubulysins have emerged as alterna-
tives to microtubule depolymerizing agents such as colchicine and vin-
blastine, which are well-established anticancer agents. However, the
complex structure of tubulysins has always posed a challenge for syn-
thetic chemists to scale up the production of these compounds. We re-
port a new strategy for the practical gram-scale synthesis of a (4R)-4-
[(tert-butoxycarbonyl)amino]-5-methylhexanoic acid through regiose-
lective cleavage of a chiral aziridine ring with a vinyl Grignard reagent to
afford tert-butyl [(1R)-1-isopropylbut-3-en-1-yl]carbamate, which was
subjected to regioselective hydroboration–oxidation with 9-BBN. The
resulting (4R)-4-[(tert-butoxycarbonyl)amino]-5-methylhexanoic acid
was successfully transformed into the deacetoxytubuvaline fragment of
pretubulysin or its highly lipophilic methyl-substituted thiazole and ox-
azole analogues for incorporation into pretubulysins. Increasing the li-
pophilicity of tubulysin or pretubulysin molecules should enhance their
cell permeability and cytotoxicity in cancer cell lines.
D-Mep
L-Ile
dTuv
Tup/Tut
R1
O
R2
O
H
N
N
1
OH
11
N
N
13
2
N
H
Me
O
R3
S
O
Key words amino acid, deacetoxytubuvaline, tubulysin, pretubulysin,
medicinal chemistry
R1 = OH, R2 = OAc, R3 = CH2OCOCH2CH(CH3)2, Tubulysin A
R1 = H, R2 = OAc, R3 = CH2OCOCH2CH(CH3)2, Tubulysin D
R1 = OH, R2 = H, R3 = Me, Pretubulysin A
R1 = R2 = H, R3 = Me, Pretubulysin D
Vinca alkaloids such as vinblastine, vincristine, or vi-
norelbine have been used to treat various types of cancer.
More recently, the taxanes, especially paclitaxel and
docetaxel as lead compounds for this family of drugs, have
brought about a revolution in anticancer therapy. The Vinca
alkaloids and taxanes disrupt microtubule polymerization
dynamics in the cell by targeting tubulin proteins, conse-
quently inducing apoptosis. If we take into account the cur-
rent number of drugs of these classes in preclinical and
clinical treatments, microtubules undoubtedly emerge as
important targets for cancer chemotherapy.1
Figure 1 Structures of pretubulysins (Prt) A and D with the dTuv frag-
ment highlighted
Mechanistically, tubulysins interact with the eukaryotic
cytoskeleton and prevent the formation of microtubules by
binding with tubulin proteins, leading to cell death.4 Their
mode of action has been clinically validated, and their high
potency against multidrug-resistant tumors has made tu-
bulysins a first choice in the development of targeted drug-
delivery systems. Their subnanomolar cytotoxicity and re-
markable antiangiogenic properties have led to significant
research activity5 in developing amenable synthetic proce-
dures in the laboratory. However, the clinical development
of the tubulysin family of compounds into a marketable
Höfle and co-workers first identified a series of anti-
mitotic tetrapeptides called tubulysins from myxobacte-
rial culture broths.2 Later, Müller and co-workers discov-
ered the gene cluster responsible for biosynthesis of
© Georg Thieme Verlag Stuttgart · New York — Synlett 2018, 29, A–E

B
R. B. Reddy et al.
Letter
Synlett
drug has been hampered by their structural complexity and
by their instability during biological assays. Staben and co-
workers6 have recently demonstrated that during in vivo
metabolism of antibody–tubulysin conjugates, the labile C-
11 acetate group compromises activity. They therefore suc-
cessfully replaced the acetate moiety with a moderately
stable propyl ether functionality.
Despite these developments in methods for synthesiz-
ing pretubuvaline and its analogues, there are no proven
synthetic protocols for large-scale preparations of these bi-
ologically active entities. Here, we report a novel and prac-
tical route that permits gram-scale synthesis of the γ-ami-
no acid intermediate 2 by regioselective ring opening of
chiral N-tosylaziridine 4 with vinyl Grignard reagent fol-
lowed by hydroboration–oxidation to afford N-Boc-N-tosyl-
δ-amino alcohol 6. Derivative 6 is then oxidized and deto-
sylated to give N-Boc-γ-amino acid 2, which is converted
into dTuv (1) and two of its methyl-substituted analogues
in good yields. The methyl substituent in the heterocyclic
ring increases the lipophilicity and cell permeability of the
tubulysin and pretubulysin derivatives, thereby enhancing
their cytotoxic potency. In future, this large-scale synthesis
of 2 should permit the construction of a diverse set of pre-
tubulysin architectures for cytotoxic evaluation in various
cancer cell lines.
A retrosynthetic analysis of the close precursor, the N-
Boc-γ-amino acid 2, for the synthesis of dTuv (1) and its an-
alogues (R = Me, X = S, O) is depicted in Scheme 1. According
to this strategy, the key intermediate 2 might be obtained
from aminoalkene 3 through a regioselective hydrobora-
tion/oxidation sequence. Aminoalkene 3 might, in turn, be
prepared by regioselective ring opening of (S)-2-isopropyl-
1-tosylaziridine (4) with vinylmagnesium bromide.
Structure–activity relationship studies of natural and
synthetic analogues of tubulysins have revealed that these
tetrapeptides are surprisingly tolerant of structural modifi-
cations in several regions. Chemical and stereochemical
changes are tolerated moderately at the C-11 hydroxy
group. Although, hydrolysis of the C11-acetyl group led to a
1000-fold decrease in biological activity, complete reduc-
tion of the secondary alcohol at C-11 to a methylene group
has been reported to restore the anticancer activity to
nanomolar concentrations.7 Investigations revealed that the
antiangiogenic and antiproliferative activities and the over-
all potency of pretubulysin are comparable to those of tu-
bulysin A (Tub-A).8 Furthermore, the mode of action of pre-
tubulysin was found to be similar to that of Tub-A in induc-
ing apoptosis through DNA laddering and nuclear
fragmentation.8,9 The interesting biological profile, stability,
and easier chemical accessibility make pretubulysin an at-
tractive target. In addition, several molecular conjugates
containing pretubulysin or its derivatives have been stud-
ied, for example, as photoaffinity probes for tubulin imag-
ing studies10 or as folate receptor-targeted oligoamides.11
Existing methods for synthesizing the deacetoxytubu-
valine (dTuv) and its analogues generally use protected L-
valine for the incorporation of the C-13 stereocenter of pre-
tubulysin. This protocol is typically carried out by a two-
carbon homologation of a protected L-valine ester, followed
by installation of the heterocyclic ring through either nucle-
ophilic substitution/heterocyclization or by standard C–C
coupling. For example, in the former approach, Ullrich and
co-workers elegantly transformed methyl L-valinate into its
corresponding nitrile homologue through a Wittig reaction,
and they constructed the thiazole ring by a Hantzsch thi-
azole methodology.12 In the latter case, Nicolaou and co-
workers reduced the C-terminus of L-valine to form the cor-
responding N-methyl bromo derivative, which underwent
C–C bond formation with lithiated methylthiazole.13 Simi-
larly, Brindisi and co-workers prepared a chloromethylthi-
azole derivative from L-serine methyl ester and coupled this
to L-valinal through a Wittig reaction.14 In contrast to these
methods, Colombo and co-workers used a tert-butylsulfin-
amide chiral auxiliary handle to install the C-13 stereocen-
ter of pretubulysin through stereoselective nucleophilic ad-
dition of a terminal alkyne onto a tert-butylsulfinimine, fol-
lowed by Sonogashira C–C coupling with a bromothiazole
and selective hydrogenation of the resultant heterocyclic
alkyne.15
Heterocyclization
O
N
OH
BocN
Me
BocHN
X
OMe
O
R
1, R = H, X = S
2
Analogues: R = Me, X = S
Hydroboration–
Oxidation
R = Me, X = O
BrMg
+
TsHN
N
Regioselective
Ring Opening
Ts
4
3
Scheme 1 Retrosynthetic analysis of deacetoxytubuvaline (1; dTuv)
and its analogues
γ-Amino acids are versatile building blocks in synthetic
organic chemistry. Their wide range of applications16 as
peptidomimetics, in the treatment of nervous-system dis-
orders, and as anticancer drugs have led to significant ef-
forts to achieve their stereoselective synthesis.17 In this
context, we have developed a novel method for preparing
the required N-Boc-γ-amino acid 2 by a two-carbon ho-
mologation of enantiomerically pure aziridine substrate 4
with a vinyl Grignard reagent (Scheme 2).
(S)-2-Isopropyl-1-tosylaziridine (4) was prepared on a
multigram scale (see Supporting Information)22 and treated
with vinylmagnesium bromide in the presence of catalytic
amount of CuCN. Tosylaziridine 4 underwent ring-opening
© Georg Thieme Verlag Stuttgart · New York — Synlett 2018, 29, A–E

C
R. B. Reddy et al.
Letter
Synlett
O
H
N
(i)
(i)
BocHN
BocHN
OMe
OMe
2
N
TsHN
O
8
Ts
OH
3
4
(ii)
(ii)
O
H
H
N
(iii)
OH
OH
BocTsN
BocTsN
O
O
5
6
8a
(iii)
(iv)
OH
(v)
BocHN
BocTsN
O
N
O
O
BocHN
2
7
S
OMe
Scheme 2 Synthesis of the N-Boc γ-substituted γ-amino acid 2. Re-
agents and conditions: (i) vinylmagnesium bromide, CuCN, THF, 0 °C to
r.t., 2 h, 72%; (ii) Boc2O, DMAP, CH2Cl2, r.t., 2 h, 87%; (iii) 9-BBN, THF,
r.t., 12 h, then H2O2, 2 N aq NaOH, 0 °C to r.t., 12 h, 93%; (iv) PDC, DMF,
r.t., 24 h, 85%; (v) Mg, NH4Cl, MeOH, reflux, 2 h, 81%.
11a
Scheme 3 Unsuccessful attempt at the synthesis of deacetoxytubu-
valine derivative 11a. Reagents and conditions: (i) Methyl L-serinate,
HOBt, EDC, DIPEA, CH2Cl2, 0 °C to r.t., 12 h, 73%; (ii) IBX, EtOAC, reflux,
12 h; (iii) Lawesson’s reagent, THF, reflux, 12 h.
nucleophilic attack by the Grignard reagent on the less-hin-
dered site of aziridine to give the 1-tosylamino alkene 3 in
72% yield. N-Boc protection of 3 was carried out by treat-
ment with Boc2O in the presence of DMAP to afford N-Boc-
1-(tosylamino)alkene 5 in 93% yield.
Regioselective hydroboration of 5 with 9-BBN, followed
by alkaline oxidation of the resulting alkyl borane with
H2O2, resulted in the formation of alcohol 6 in 93% yield.
Pyridinium dichromate (PDC) oxidation of 6 afforded the N-
Boc-N-tosyl-γ-amino acid 7 in 85% yield. This was deto-
sylated with magnesium metal in the presence of ammoni-
um chloride to provide the key intermediate N-Boc-γ-ami-
no acid 2 (Scheme 2). Intermediate 2 is a critical intermedi-
ate in the proposed synthesis of dTuv (1) and its methyl-
substituted thiazole and oxazole analogues, as outlined in
Schemes 4–6 below.
Initially, the N-Boc-γ-amino acid 2 was coupled with L-
serine by using N-ethyl-N′-(3-dimethylaminopropyl)carbo-
diimide (EDC) as a coupling agent in the presence of ben-
zotriazol-1-ol (HOBt) to obtain dipeptide 8, which was
transformed into dTuv (1) by heterocyclization of interme-
diate 8a. However, all our attempts to convert the 2-iodoxy-
benzoic acid (IBX)-oxidation product of 8 into 1 in situ via
the unstable intermediate 8a by using Lawesson’s reagent
were unsuccessful, giving no trace of the required deriva-
tive 11a (Scheme 3).
Consequently, we chose to prepare the N-Boc-γ-amino
carboxamide 9, which could be converted into N-Boc-γ-
amino thioamide 10 under standard reaction conditions.
The thioamide 10 should subsequently be readily trans-
formed into N-Boc-protected dTuv through heterocycliza-
tion by treatment with ethyl bromopyruvate (Scheme 4).
We therefore treated the N-Boc-γ-amino acid 2 with am-
monium chloride in the presence of a base with 3-[bis(di-
methylamino)methyliumyl]-3H-benzotriazole
1-oxide
hexafluorophosphate (HBTU) as the coupling agent, to fur-
nish N-Boc-γ-aminocarboxamide 9 in 95% yield. Intermedi-
ate 9 was then converted into thioamide 10 in 89% yield by
treatment with Lawesson’s reagent in refluxing THF. The
thioamide 10 was subsequently converted into the 2-alkyl-
substituted thiazole ethyl ester 11 in an excellent yield of
86% by following the procedure reported by Menche and
co-workers.18 Finally, N-methylation of 11 by methyl iodide
in the presence of sodium hydride in N,N-dimethylforma-
mide (DMF) as solvent gave dTuv (1) in 87% yield (Scheme
4). Note that during the N-methylation reaction of 11,23
parallel and complete conversion of the ethyl ester of 11
into methyl ester 1 took place.
a
Studies by Stark and Assaraf19 have shown that increas-
ing the lipophilicity of tubulysin derivatives can enhance
their cell diffusion, leading to lower IC50 values. We are cur-
rently focusing on synthesizing more-lipophilic derivatives
of tubulysins and pretubulysins. Pretubulysins are naturally
more lipophilic than the corresponding tubulysins due to
the absence of the C-11 acetoxy group. Furthermore, we
were interested in designing novel dTuv analogues and en-
hancing the lipophilicity of the heterocyclic ring by the in-
troduction of alkyl substituents as simple as a methyl
group. Subsequently, we intended to incorporate the meth-
yl-substituted dTuv analogues into a series of modern pre-
tubulysin architectures.
To achieve the synthesis of the 5-methylthiazole-sub-
unit-containing dTuv analogues, the N-Boc-γ-amino acid 2
was coupled with L-threonine by using EDC as a coupling
agent in the presence of HOBt, with DIPEA as a base, to fur-
nish dipeptide 12 in 70% yield. The secondary alcohol func-
tionality in dipeptide 12 was oxidized to a ketone by using
© Georg Thieme Verlag Stuttgart · New York — Synlett 2018, 29, A–E

D
R. B. Reddy et al.
Letter
Synlett
The oxazole analogue of the Tuv fragment in the tu-
bulysin series was first introduced by Wipf et al.20 and, sub-
sequently, Shankar and co-workers21 found that oxazole-
containing tubulysin U analogues are even more cytotoxic
than their thiazole counterparts.
NH2
(i)
BocHN
2
O
9
(ii)
With this in mind, we also synthesized the dTuv 5-
methyloxazole analogue 16. The keto amide ester 13 was
treated with triphenylphosphine (PPh3) and iodine to form
the 5-methyloxazole subunit through an oxidative cycliza-
tion reaction. Without further purification of the interme-
diate and after workup, N-methylation gave the final dTuv
oxazole analogue 16 in 80% yield (Scheme 6).
(iii)
NH2
BocHN
O
S
10
N
(iv)
BocHN
S
OEt
11
O
O
H
N
N
BocN
Me
BocHN
OMe
S
OMe
O
O
1
13
Scheme 4 Synthesis of deacetoxytubuvaline (dTuv; 1). Reagents and
conditions: (i) HOBt, HBTU, NH4Cl, DIPEA, CH2Cl2, 0 °C to r.t., 4 h, 95%;
(ii) Lawesson’s reagent, THF, 0 °C to r.t., 12 h, 89%; (iii) ethyl bromopy-
ruvate, EtOH, 65 °C, 1 h, 86%; (iv) MeI, NaH, DMF, r.t., 12 h, 87%.
(i)
O
N
BocN
Me
O
16
OMe
Dess–Martin periodinane (DMP) to afford keto amide ester
13 in 82% yield. Intermediate 13 was treated with Lawesson’s
reagent for in situ thionation–cyclization to obtain the N-
Boc-protected 5-methylthiazole ester 14. N-Methylation of
14 with MeI in the presence of NaH provided the target
dTuv analogue 15 in 80% yield (Scheme 5).
Scheme 6 Synthesis of 5-methyloxazole analogue of deacetoxytubu-
valine 16. Reagents and conditions: (i) PPh3, I2, THF, –40 °C, 3 h, then
MeI, NaH, DMF, r.t., 12 h, 80%.
In summary, we have developed a simple protocol for
the synthesis of the deacetoxytubuvaline (dTuv) fragment 1
of pretubulysin and its methyl-substituted thiazole and ox-
azole analogues, 15 and 16, respectively, in high yields. The
synthetic design is suitable for scaled-up synthesis of these
highly desirable fragments. Total syntheses of new pretu-
bulysins, containing these lipophilic fragments, and an in
vitro evaluation of their cytotoxicity in a range of cancer
cell lines is currently underway in our laboratory and will
be reported in due course.
O
H
N
(i)
BocHN
OMe
2
O
HO
12
(ii)
O
H
N
BocHN
OMe
O
O
13
Funding Information
(iii)
O
We would like to extend our sincere gratitude to the Science and En-
gineering Research Board, Department of Science and Technology,
Government of India, for providing funding under the grant number
EMR/2015/001764.()
N
BocHN
S
OMe
14
(iv)
O
Acknowledgements
N
BocN
Me
S
OMe
The authors are grateful to the Ministry of Human Resource Develop-
ment (MHRD), Government of India, Indian Institute of Technology
(IIT) Indore, and the Council of Scientific and Industrial Research
(CSIR), India for research fellowships. We also thank the Sophisticated
Instrumentation Centre (SIC), IIT Indore, for providing compound-
characterization facilities.
15
Scheme 5 Synthesis of the 5-methylthiazole analogue of deacetoxytu-
buvaline 15. Reagents and conditions: (i) L-threonine methyl ester hydro-
chloride, EDC, HOBt, DIPEA, CH2Cl2, 0 °C to r.t., 12 h, 70%; (ii) DMP, CH2Cl2,
r.t., 4 h, 82%; (iii) Lawesson’s reagent, THF, reflux, 6 h, 70%; (iv) MeI,
NaH, DMF, r.t., 12 h, 80%.
© Georg Thieme Verlag Stuttgart · New York — Synlett 2018, 29, A–E

E
R. B. Reddy et al.
Letter
Synlett
Supporting Information
(16) Trabocchi, A.; Guarna, F.; Guarna, A. Curr. Org. Chem. 2005, 9,
1127.
(17) Ordóñez, M.; Cativiela, C. Tetrahedron: Asymmetry 2007, 18, 3.
(18) Menche, D.; Hassfeld, J.; Li, J.; Rudolph, S. J. Am. Chem. Soc. 2007,
129, 6100.
Supporting information for this article is available online at
https://doi.org/10.1055/s-0037-1611359.
S
u
p
p
orti
n
gInformati
o
n
S
u
p
p
orit
n
gInformati
o
n
(19) Stark, M.; Assaraf, Y. G. Oncotarget 2017, 8, 49973; DOI:
10.18632/oncotarget.18385.
References and Notes
(20) Wipf, P.; Takada, T.; Rishel, M. J. Org. Lett. 2004, 6, 4057.
(21) Shankar, P. S.; Sani, M.; Saunders, F. R.; Wallace, H. M.; Zanda,
M. Synlett 2011, 1673.
(22) (a) Kitir, B.; Baldry, M.; Ingmer, H.; Olsen, C. A. Tetrahedron
2014, 70, 7721. (b) Ye, W.; Leow, D.; Goh, S. L. M.; Tan, C.-T.;
Chian, C.-H.; Tan, C.-H. Tetrahedron Lett. 2006, 47, 1007.
(23) Methyl 2-{(3R)-3-[(tert-Butoxycarbonyl)(methyl)amino]-4-
methylpentyl}-1,3-thiazole-4-carboxylate (1)
(1) Mukhtar, E.; Adhami, V. M.; Mukhtar, H. Mol. Cancer Ther. 2014,
13, 275.
(2) Sasse, F.; Steinmetz, H.; Heil, J.; Höfle, G.; Reichenbach, H.
J. Antibiot. 2000, 53, 879.
(3) Sandmann, A.; Sasse, F.; Müller, R. Chem. Biol. 2004, 11, 1071.
(4) Wang, Z.; McPherson, P. A.; Raccor, B. S.; Balachandran, R.; Zhu,
G.; Day, B. W.; Vogt, A.; Wipf, P. Chem. Biol. Drug Des. 2007, 70,
75.
Anhyd DMF (2 mL) was added to the thiazole ethyl ester 11
(5) Rath, S.; Liebl, J.; Fürst, R.; Ullrich, A.; Burkhart, J. L.; Kazmaier,
U.; Herrmann, J.; Müller, R.; Günther, M.; Schreiner, L.; Wagner,
E.; Vollmar, A. M.; Zahler, S. Br. J. Pharmacol. 2012, 167, 1048.
(6) Staben, L. R.; Yu, S.-F.; Chen, J.; Yan, G.; Xu, Z.; Del Rosario, G.;
Lau, J. T.; Liu, L.; Guo, J.; Zheng, B.; dela Cruz-Chuh, J.; Lee, B. C.;
Ohri, R.; Cai, W.; Zhou, H.; Kozak, K. R.; Xu, K.; Phillips, G. D. L.;
Lu, J.; Wai, J.; Polson, A. G.; Pillow, T. H. ACS Med. Chem. Lett.
2017, 8, 1037.
(7) Murray, B. C.; Peterson, M. T.; Fecik, R. A. Nat. Prod. Rep. 2015,
32, 654.
(8) Herrmann, J.; Elnakady, Y. A.; Wiedmann, R. M.; Ullrich, A.;
Rohde, M.; Kazmaier, U.; Vollmar, A. M.; Müller, R. PLoS One
2012, 7, e37416.
(9) Braig, S.; Wiedmann, R. M.; Liebl, J.; Singer, M.; Kubisch, R.;
Schreiner, L.; Abhari, B. A.; Wagner, E.; Kazmaier, U.; Fulda, S.;
Vollmar, A. M. Cell Death Dis. 2014, 5, e1001.
(10) Eirich, J.; Burkhart, J. L.; Ullrich, A.; Rudolf, G. C.; Vollmar, A.;
Zahler, S.; Kazmaier, U.; Sieber, S. A. Mol. BioSyst. 2012, 8, 2067.
(11) Truebenbach, I.; Gorges, J.; Kuhn, J.; Kern, S.; Baratti, E.;
Kazmaier, U.; Wagner, E.; Lächelt, U. Macromol. Biosci. 2017, 17,
1600520.
(12) Ullrich, A.; Herrmann, J.; Müller, R.; Kazmaier, U. Eur. J. Org.
Chem. 2009, 6367.
(13) Nicolaou, K. C.; Erande, R. D.; Yin, J.; Vourloumis, D.; Aujay, M.;
Sandoval, J.; Munneke, S.; Gavrilyuk, J. J. Am. Chem. Soc. 2018,
140, 3690.
(0.080 g, 0.22 mmol) in
a 10 mL round-bottomed flask
equipped with a magnetic stirrer bar. MeI (0.05 mL, 0.89 mmol)
was then added, and the mixture was cooled to 0 °C. NaH (0.013
g, 0.55 mmol) was added portionwise over 15 min with con-
stant stirring, and the mixture was stirred at 25 °C under N2 for
a further 12 h. When 11 was completely consumed (TLC), the
reaction was quenched with sat. aq NH4Cl (10 mL). The aqueous
layer was extracted with EtOAc (3 × 25 mL), and the combined
organic extracts were washed with brine (3 × 10 mL), dried
(Na2SO4), filtered, and concentrated under reduced pressure.
The resulting crude residue was purified by column chromatog-
raphy [silica gel, hexane–EtOAc (80:20)] to give a colorless
liquid; yield: 70.0 mg (87%); TLC: Rf = 0.25 (hexane–EtOAc,
70:30). [α]D20 –20.4 (c 0.1, CH2Cl2).
IR (CH2Cl2): 2963, 2920 (C–H), 1722, 1694 (C=O), 1685 (C=N),
1484, 1392 (C–H), 1170 (C–O), 1155 (S–O), 867 (=C–H), 775,
720 (C–H) cm–1. 1H NMR (400 MHz, CDCl3, 25 °C): δ = 8.08 (s, 1
H, rotamer 1), 8.06 (s, 1 H, rotamer 2), 3.95 (s, 3 H, rotamer 1),
3.94 (s, 3 H, rotamer 2), 2.99–2.95 (m, 4 H, rotamers 1 + 2), 2.70
(s, 3 H, rotamer 1), 2.65 (s, 3 H, rotamer 2), 2.18–2.11 (m, 2 H,
rotamers 1 + 2), 1.89–1.81 (m, 2 H, rotamers 1 + 2), 1.70–1.67
(m, 4 H, rotamers 1 + 2), 1.46 (s, 9 H, rotamer 1), 1.43 (s, 9 H,
rotamer 2), 0.96 (t, J = 6.0 Hz, 6 H, rotamer 1), 0.85 (d, J = 6.5 Hz,
6 H, rotamer 2). 13C NMR (100 MHz, CDCl3, 25 °C): δ = 171.9,
162.0, 156.6, 146.5, 127.2, 79.6, 60.3, 52.5, 30.73, 30.5, 30.0,
29.7, 28.4, 20.2, 19.9. Diagnostic signals of minor rotamer 13C
NMR (100 MHz, CDCl3, 25 °C): δ = 171.5, 161.9, 146.4, 79.2,
52.4, 20.1, 19.6. HRMS (ESI): m/z [M + H]+ calcd for C17H29N2O4S:
379.1662; found: 379.1675.
(14) Brindisi, M.; Maramai, S.; Grillo, A.; Brogi, S.; Butini, S.;
Novellino, E.; Campiani, G.; Gemma, S. Tetrahedron Lett. 2016,
57, 920.
(15) Colombo, R.; Wang, Z.; Han, J.; Balachandran, R.; Daghestani, H.
N.; Camarco, D. P.; Vogt, A.; Day, B. W.; Mendel, D.; Wipf, P.
J. Org. Chem. 2016, 81, 10302.
© Georg Thieme Verlag Stuttgart · New York — Synlett 2018, 29, A–E