Total synthesis of a noricumazole A library and evaluation of HCV inhibition

Article abstract of DOI:10.1002/chem.201104042

The total synthesis of 16 new ion channel inhibitors derived from noricumazole A, a secondary metabolite from the myxobacterium Sorangium cellulosum, is reported. Particular focus of library design is put on stereochemical permutations in the central regi

Full text of DOI:10.1002/chem.201104042

FULL PAPER
DOI: 10.1002/chem.201104042
Total Synthesis of a Noricumazole A Library and Evaluation of HCV
Inhibition
Jenny Barbier,[a] Jens Wegner,[a] Stefan Benson,[a] Juliane Gentzsch,[b]
Thomas Pietschmann,[b] and Andreas Kirschning*[a]
Dedicated to Professor Dr. Dr. h.c. Ekkehard Winterfeldt on the occasion of his 80th birthday
Abstract: The total synthesis of 16 new
ion channel inhibitors derived from
noricumazole A, a secondary metabo-
lite from the myxobacterium Soran-
gium cellulosum, is reported. Particular
focus of library design is put on stereo-
chemical permutations in the central
region (C9 and C11), the oxazole
moiety and the side chain at C4 of the
isochromanone moiety. Noricumazo-
le A and all new noricumazole deriva-
tives were tested in an assay system
with inhibitory effect on the hepatitis C
virus (HCV) life cycle. Most of them
are moderate to strong HCV inhibitors
(350 nm–6 nm) but also exert pro-
nounced cytotoxicity. In contrast, the
thiazole analogue of noricumazole A is
a strong HCV inhibitor with only mod-
erate cytotoxic property. It may
become a lead structure with a good
therapeutic index (CC50/IC50) of greater
than 10.
Keywords: compound library
·
Hepatitis C virus · iron catalysis ·
natural products · total synthesis
Introduction
Recently, noricumazole A (1a) and the related icumazole A
(2a) were isolated from the myxobacterium Sorangium cel-
lulosum and the structures including all stereogenic centers
were elucidated by spectroscopic methods, chemical deriva-
tization, controlled degradation and total synthesis
(Figure 1).[1] Other members of these secondary metabolites,
noricumazole B (1b) and noricumazole C (1c) include gly-
cosylated derivatives at C11 and C18, respectively. Glycosy-
lated derivatives of icumazole A (2a), namely icumazole B1
(2b) and B2 (2c) are also known.[1] Remarkably, noricuma-
zole A (1a) is the first natural product reported that shows
a stabilizing effect on the tetrameric architecture of the po-
tassium channel KcsA thereby blocking it (1–4 mm).[1]
In addition, noricumazole A (1a) was found to interfere
with the complete life cycle of the hepatitis C virus (HCV)
in cell culture while exerting only moderate cytotoxicity to-
wards the host cell.[2] HCV is one of the primary causes of
Figure 1. Noricumazoles A–C (1a–c) and icumazoles A–C (2a–c); num-
bering does not relate to IUPAC; however, for clarity the depicted num-
bering was chosen in this report.
[a] Dr. J. Barbier, Dr. J. Wegner, Dr. S. Benson, Prof. Dr. A. Kirschning
Institut fꢀr Organische Chemie und
chronic liver disease worldwide. In fact, chronic HCV infec-
tion is associated with chronically active hepatitis, fibrosis,
and often results in end-stage liver cirrhosis and hepatocel-
lular carcinoma. Although treatment options have been con-
siderably improved over the past decades, and the first di-
rectly acting antivirals were licensed in 2011, side effects,
drug resistance and viral genotype-specific differences in ef-
ficacy of these therapies still limit current treatment options.
Thus, novel therapies are needed to efficiently treat the
about 160 million chronically infected HCV patients world-
wide.
Zentrum fꢀr Biomolekulare Wirkstoffe (BMWZ)
Leibniz Universitꢁt Hannover, Schneiderberg 1B
30167 Hannover (Germany)
Fax : (+49)511-762-3011
E-mail: andreas.kirschning@oci.uni-hannover.de
[b] Dr. J. Gentzsch, Prof. Dr. T. Pietschmann
TWINCORE, Zentrum fꢀr Experimentelle und
Klinische Infektionsforschung GmbH
Feodor-Lynen-Str. 7, 30625 Hannover (Germany)
Supporting information for this article is available on the WWW
under http://dx.doi.org/10.1002/chem.201104042.
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This preliminary finding initiated a total synthesis pro-
gram in our laboratories to generate a library of noricuma-
zole A derivatives and carry out a first study on the struc-
ture–activity relationship. The synthesis program is based on
our first total synthesis of noricumazole A (1)[1] that helped
to unequivocally elucidate the relative and absolute configu-
rations of all stereogenic centers. The program was aimed at
simplifying noricumazole A, specifically with respect to the
unusual 2-methylbutyl side chain bound at C4 of the aro-
matic moiety. Furthermore, we envisaged the permutation
of the stereogenic centers at C9 and C11, respectively, as
well as the exchange of the oxazole by a thiazole ring.
(Table 1). As aryl moieties we chose bromides 7 to 11 as
well as boronic acid 12 of which aryl bromides 7–9 com-
pletely failed to serve as organomagnesium or organozinc
intermediates in cross coupling reactions with alkyl halides 5
or 6, respectively.[5]
Table 1. Selected examples of sp2–sp3 cross coupling reactions.
Arene Conditions
Product
Yield
[%][a]
Results and Discussion
5, PdACTHNURGTEN(UGN OAc)2, KOtBu,
tBu2MeP, amyl alcohol, room
1
2
12
24[b]
Syntheses of western fragments: Our principal retrosynthesis
plan towards new derivatives of noricumazole A is summar-
ized in Scheme 1. It is based on the key fusion of vinyl io-
temperature
i. Mg, THF, D, then
ii. PEPPSI-IPrTM
, ZnCl2,
[c]
11
n.p.[d,e]
LiBr, 5, DMI, THF, room
temperature, 3 h
3
4
5
11
11
11
Mg, TMEDA, THF, FeCl3, 5 14
(1.2 equiv), 08C, 16 h, room
temperature
Mg, TMEDA, THF, FeCl3, 5 14
or 6 (1.2 equiv), 16 h
33
25 (for 5) n.p.
(for 6) 50
(for 5)[f]
i. Mg, THF, D, 1 h then
ii. Fe(acac)3, THF, TMEDA,
14
98[g]
AHCTUNGTRENNUNG
HMTA, 6 (1 equiv), 08C,
1.5 h
[a] Yields are based on isolated products; [b] crude yield; [c] for PEPPSI-
IPrTM refer to ref. [7]; [d] n.p.=no product; [e] only the protodebrominat-
ed starting material was isolated; [f] distilled bromide 5 was employed;
[g] freshly distilled iodide 6 and bromide 11 were employed; (DMI=1,3-
dimethyl-2-Imidazolidinone; TMEDA=tetramethylethylene diamine;
acac=acetylacetonate; HMTA=hexamethylphosphortriamide).
The reversed mode of coupling, that is, transformation of
the alkyl halides into the corresponding organometallic spe-
cies and coupling with aryl bromides 7–9 was also not suc-
cessful. Reactions were typically probed in the presence of
a catalytic amount of an iron source and TMEDA or
HMTA,[6] or in the presence of a palladium catalyst (e.g.,
PEPPSI-IPrTM) following Organꢃs[7] or Fuꢃs[8] procedures.
The first successful cross coupling reaction was achieved
between boronic acid 12 and alkyl halide 6 following Fu’s
protocol (Table 1, entry 1);[8] however, the yield of 24% for
13 was far from satisfactory. Only, when the phenol protec-
tion was changed from the methyl to the MOM group the
process could be improved. Employing the conditions de-
scribed by von Wangelin and co-workers, aryl bromide 11
was coupled in the presence of iron (entry 3) instead of pal-
ladium (entry 2) to furnish arene 14 in 33% yield.[9] Finally,
the iron-catalyzed version[7,9,10] of the Kumada reaction[11]
gave the coupling product 14 in excellent yield (Table 1,
Scheme 1. General retrosynthesis plan.
dides 3 and aldehydes 4 by a nucleophilic vinylation
(Scheme 1). The western fragment has a typical polyketide
structure and could be synthesized by a classical aldol ap-
proach employing a Masamune anti-aldol[3] and a Nagao
aldol[4] reaction for setting up the stereotriade. The synthe-
ses of the western fragments were planned to start from
a simple phenol precursor and, depending whether a sub-
stituent is present at C4, two to three successive organome-
tallic transformations, which included a sp3–sp2 cross cou-
pling reaction for R is unlike H.
For the key sp3–sp2 coupling reaction that introduces the
aliphatic side chain various strategies were pursued, that
rely on copper, palladium or iron catalysis either by using
organometallics based on magnesium, zinc or boronates
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entry 5). The key to success was that besides TMEDA also
freshly distilled alkyl halide and aryl bromide were em-
ployed.
Next, the second ortho-position in 14 was functionalized
by first deprotonating with nBuLi followed by treatment
with tert-butyl isocyanate (Scheme 2). Amide 15 was ob-
tained and then utilized to direct a second deprotonation
and the subsequent ortho lithiated species was then inde-
pendently treated with both enantiomeric oxiranes
25a,b[12,13] to ring opening products 16 and 17, respectively.
These alcohols could only be transformed into the d-lac-
tones after removal of the MOM as well as the silyl protect-
ing groups. Then, cyclization was achieved under acidic con-
ditions in refluxing toluene to afford both diastereomeric
fragments 18 and 19.
In a similar manner, we prepared the simplified western
fragment 24 with an n-butyl side chain by following the syn-
thesis protocols described for western fragments 18 and 19
(Scheme 2). The cross coupling reaction of butyl iodide 20
with aryl bromide 11 gave butylarene 21 in good yield.
Additionally, also the western fragment that is not alkylat-
ed at C4 was prepared starting from arylether 26
(Scheme 3). Following the synthesis sequence described in
Scheme 2 the isochromanone derivative 28a and the O-me-
thylated analogue 28b were straightforwardly prepared.
Scheme 3. Preparation of western fragment derivative 28a and 28b. Re-
agents and conditions: a) i. nBuLi, TMEDA, Et2O, À308C, 2 h, then 3 h
at À58C; ii. tBuNCO, room temperature, 14 h, 89%; b) nBuLi, TMEDA,
Et2O, À788C to À408C, 2 h, then addition of (R)-25a, À788C to À408C,
14 h [44% (99% b.o.r.s.m.) for 27a and 76% (99% b.o.r.s.m.) for 27b];
c) HClconc, EtOH, 508C, 50 min (70% from 27a and 99% from 27b);
d) pTsOH, toluene, D, 45 min (98% for 28a and 92% for 28b).
Syntheses of eastern fragments: Based on the published syn-
thesis,[1] we prepared one new eastern fragment that only
differs in the heterocylic moiety compared to noricumazo-
le A (1a). The natural oxazole ring was exchanged by thia-
zole moiety. The synthesis commenced with a Masamune
anti-aldol reaction[2] between propionate 29 and propanal
(Scheme 4). Aldehyde 30 was obtained after O-silylation,
DIBAL-mediated reduction and Dess–Martin oxidation
(Scheme 4).
Scheme 2. Preparation of western fragments 18 and 19 as well as 2’-nor-
derivative 24. Reagents and conditions: a) i. (S)-6, Mg, THF, D, 1 h; ii. 11,
HMTA, TMEDA, FeACHTUNGTRENNUNG(acac)3, 08C, 1.5 h (14, 98% and 21, 76% from 20);
The Nagao aldol reaction[3] served to establish the third
stereogenic center by using thioxothiazolidine 35. Second,
O-silylation and hydrolytic removal of the chiral auxiliary
gave the carboxylic acid 31, which was coupled with serine
methylester 36 in the presence of the condensation cocktail
of HOBt and TBTU. In the presence of DAST[14] ring clo-
sure to the oxazolidine ring was initiated, which upon oxida-
tion by using Williamsꢃ procedure led to the corresponding
oxazole 32 in good overall yield.[15] In contrast, thiazole 34
was prepared via thioamide 33, which was accessed from the
b) i. nBuLi, TMEDA, Et2O, À308C, 2 h, then 3 h at À58C; ii. tBuNCO,
room temperature, 14 h, (94% for 15 and 67% for 22); c) nBuLi,
TMEDA, Et2O, À788C to À408C, 2 h, then addition of (R)-25a, À788C
to À408C, 14 h, [73% (99% b.o.r.s.m.) for 16 and 57% (95% b.o.r.s.m.)
for 23], alternatively, addition of (S)-25b, À788C to À408C, 14 h, (81%
for 17); d) HClconc, EtOH, 508C, 2.5 h, (89% from 16, 93% from 23 and
94% from 17); e) pTsOH, toluene, D, 30 min, (89% for 18, 77% for 24
and 95% for 19); (TBS=tert-butyldimethylsilyl; MOM=methoxymeth-
yl).
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corresponding amide by thionation with Lawessonꢃs reagent
37.[16] Next, the Hantzsch thiazole synthesis by using bromo-
pyruvate 38 gave the desired eastern fragment 34 in very
good yield.[17]
With methyl ester 32 and ethyl ester 34 in hand, further
elaboration to the corresponding vinyl iodides 39 and 40 (re-
sembling 3; Scheme 1), respectively, was pursued. This was
achieved by first a reduction step yielding the corresponding
aldehyde (Scheme 5) followed by Seyferth–Gilbert homolo-
gation[18] by using the Ohira–Bestmann reagent 51.[19]
The resulting alkyne was subjected to a syn-hydro-zirco-
nation[20] followed by iodination, which furnished vinyl io-
dides 39 and 40, respectively. Noteworthy, we also tested the
Takai olefination[21] of the intermediate aldehyde because
this transformation would yield the desired vinyl iodides 39
and 40 in one step. However, we discovered that it was im-
possible to quantitatively isolate the vinyl iodide in the pres-
ence of excess iodoform.
The key step, the vinylation of aldehyde 41 with metalat-
ed vinyl iodide 39, required substantial optimization. First of
all, oxidation of the primary hydroxyl groups in 18 and its
epimer 19 posed a problem because the b-center underwent
facile epimerization for which we propose a b-elimination/
oxo-Michael addition mechanism; this might be accelerated
by hydrogen bonding between the phenol moiety and the
lacton carbonyl group (see substructure A, Scheme 5). Epi-
merization could be suppressed when oxidation was carried
out under strictly dry and acid-free conditions and the alde-
hyde was immediately employed in the next step.
Subsequently, vinylation was achieved after lithiation of
vinyl iodide 39 and transmetalation to the corresponding or-
ganozinc species. Nucleophilic addition to aldehyde 41[22]
furnished the desired product 43a and its 11-epimer 43b as
a 1:1 mixture.[1] Asymmetric coupling variants, like the use
of chiral ligands[23] known to be applicable in the zinc-medi-
ated vinylation or the chromium(II)-mediated Nozaki–Kishi
reaction,[24] failed. Alternatively, we pursued the diastereose-
lective alkynylation of aldehyde 41 with lithiated alkyne col-
lected en route to vinyl iodide 39 in the presence of various
chiral ligands,[25] such as (À)-N-methylephedrine and (R)-
(+)-1,1’-bi-2-naphthol. All these attempts were not success-
ful. Thus, after chromatographic separation of both diaste-
reomers the undesired 11-epi-allyl alcohol 43b was directly
epimerized to 43a utilizing the Mitsunobu reaction[26] fol-
lowed by ester hydrolysis. Following this route, we improved
the yield for the desired coupling product 43a to 68%. The
mixture of epimeric alcohols 43a and 43b could also be oxi-
dized to the enone 44. However, all attempts to achieve
a facial-selective reduction by using (S)-2-methyl-CBS-oxa-
zaborolidine, alpine–borane, Ipc2BCl, L-Selectride or (R)-
BINAL-H as reductants were not successful with respect to
overall yield and selectivity.[27–29] Thus, completion of the
synthesis was finally achieved by Lewis acid-mediated TBS-
deprotection (BF3·Et2O, CH3CN, 08C, 12 h, 90%) to furnish
noricumazole A (1a) in 14% overall yield over 15 steps
(longest linear sequence; by including the Mitsunobu epime-
rization protocol the synthesis proceeded in 17 linear steps
Scheme 4. Synthesis of the eastern thiazole fragment 33. Reagents and
conditions: a) c-HexBOTf, NEt3, propanal, CH2Cl2, À788C, 2 h, 91%,
d.r.>20:1; b) TBSOTf, 2,6-lutidine, CH2Cl2, À788C to room temperature,
50 min, quant.; c) DIBAL-H, CH2Cl2, À788C, 30 min, 91%; d) Dess–
Martin periodinane, CH2Cl2, room temperature, 1 h; e) 35, TiCl4,
iPr2EtN, À408C to À788C, 1.5 h; f) TBSOTf, 2,6-lutidine, CH2Cl2,
À788C, 30 min, 91% over three steps; g) 1m LiOH, 30% H2O2, THF/
H2O 4:1, 08C to room temperature, 12 h, 99%; h) 36, HOBt, TBTU,
iPr2EtN, CH2Cl2, room temperature, 12 h, 97%; i) DAST, CH2Cl2,
À788C, 3 h, 78% (88% b.o.r.s.m.); j) DBU, BrCCl3, CH2Cl2, 08C to room
temperature, 6 h, 85%; k) i. (COCl)2, DMF, Et2O, room temperature,
1 h; ii. NH4OH, MeOH, 08C, 1 h, 99%; l) 37, THF, room temperature,
4 h, 65%; m) i. 38, acetone, À108C, 2 h; ii. pyridine, (CF3CO)2O, CH2Cl2,
À308C to room temperature over 4 h, 14 h, 88%; (DAST=diethylamino-
sulfur trifluoride; DBU=1,8-diazabicycloACTHNUTRGNEU[GN 5.4.0]undec-7-en; DMF=N,N-
dimethylformamid; DIBAL-H=diisobutylaluminium hydride; Hex=n-
hexyl; HOBt=hydroxybenzotriazole; TBSOTf=tert-butyldimethylsilyl
triflate; TBTU=O-benzotriazol-yl-N,N,N’,N’-tetramethyluronium tetra-
fluoroborate).
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Likewise, the metallated
vinyl iodide 40 that contains
a thiazole instead of an oxazole
heterocycle was coupled with
aldehyde 41. The two separable
diastereomers 45a and 45b
were also formed in a 1:1 ratio.
Both diastereomeric thiazole
derivatives were then depro-
tected to yield thia-noricumazo-
le A (49a; 51%) and 11-epi-
thia-noricumazole A
(49b;
38%), respectively. Further-
more, we generated the 12,13-
hydro-noricumazole A deriva-
tive 52 from noricumazole A
(1a).
At this point it is necessary
to briefly discuss how the ste-
reochemistry at C11 with re-
spect to C9 was elucidated, be-
cause the vinylation always pro-
vided two diastereomers. We
addressed this issue in all cases
as is exemplified for the 9,11-
bis-epi derivative 46b in
Scheme 6.[30] First, the lactone
moiety was reduced to yield the
tetraol 53 followed by forma-
tion of bisacetonide 54. Accord-
ing to Rychnovsky et al.[31] the
13C d values for the acetonide
carbon atoms spanning C9 and
C11 were diagnostic to un-
equivocally prove the 1,3-anti
relationship. In case of the n-
butyl side-chain-containing de-
rivative 59a and methyl pro-
tected phenol derivative 60a
the anti monoacetonides were
Scheme 5. Syntheses of noricumazole A (1a) and derivatives 47–50. Reagents and conditions: a) Dess–Martin
periodinane, NaHCO3, CH2Cl2, 08C, 45 min, [85% for 41 and 44% (62% b.o.r.s.m.) for 42]; b) DIBAL-H, formed.[30]
CH2Cl2, À788C, 1 h, (quant. for both esters 32 and 34); c) K2CO3, Ohira–Bestmann reagent 51, MeOH, 08C to
room temperature, (86% for oxazole and 96% for thiazole); d) Cp2ZrHCl, THF, 08C, 1 h, then NIS, À788C,
45 min, (90% for 39 and 89% for 40); e) 39, tBuLi, Et2O, À788C, 1 h, then Me2Zn, À788C, 15 min, then 41,
Et2O, À788C, 2.5 h, 74%, d.r. 1:1 (43a/43b from 41) and 74%, d.r. 1:1 (46a/46b from 42); f) Dess–Martin peri-
odinane, NaHCO3, CH2Cl2, 08C, 1 h, 65% for 44; g) 43b, PPh3, p-nitrobenzoic acid, DEAD, THF, 08C to
room temperature, 3 h, 90%; h) NaOH, H2O, THF/MeOH 2:1, 08C, 12 h, 92%; i) 40, tBuLi, Et2O, À788C, 1 h,
then Me2Zn, À788C, 15 min, then 41, Et2O, À788C, 2.5 h, 74%, d.r. 1:1 (45a/45b); j) BF3·Et2O, CH3CN, 08C,
12 h (1a: 90%, 47: 90%, 48: 83%, 49a: 51%, 49b: 38%, 50a: 83% and 50b: 86%); k) Pd/C (10%), H2
(1 bar), MeOH, room temperature, 2 h, 50%; (Cp=cyclopentadienyl; DEAD=diethyl azodicarboxylate;
NIS=N-iodosuccinimide).
Having established a general
route to new noricumazole de-
rivatives we extended our ef-
forts to a simplified n-butyl side
chain at C3 as well as deriva-
tives altogether without an
alkyl side chain. These deriva-
tives would provide information
on the biological or pharma-
ceutical importance of this bio-
and 25% overall yield). In a similar manner we also ob-
tained the 11-epi noricumazole A (47; 90%), 11-dehydro-
noricumazole A (48; 83%), 9-epi-noricumazole A (50a;
83%) and 9,11-bis-epi-noricumazole A (50b; 86%) from the
corresponding silyl-protected noricumazole derivatives 43b,
44, 46a and 46b, respectively.
synthetically unusual aryl substituent. For this purpose alco-
hols 24, 28a and 28b were oxidized to the corresponding al-
dehydes 55–57 and directly coupled with metallated vinyl
iodide 39 to yield coupling products 58a,b–60a,b as pair of
diastereomers (1:1 ratio) after desilylation (Scheme 7).
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A. Kirschning et al.
diagnostic when epimerization at C9 has taken place during
or after oxidation. However, for determining the stereogenic
integrity at C9 of butyl-substituted derivatives, we pursued
a strategy that is exemplified for derivative 61 (Scheme 8).
Scheme 8. Determination of the diastereomeric purity of 61. Reagents
and conditions: a) Pd/C (10%), H2 (1 bar), MeOH, room temperature,
1 h, 93%; b) (R)-(À)-a-methoxyphenylacetic acid, EDC·HCl, DMAP,
CH2Cl2, 408C, 40 min, 78%; (DMAP=4-dimethylaminopyridine; EDC=
1-(3-dimethylamino-propyl)-3-ethylcarbodiimide).
Scheme 6. Determination of the relative stereochemistry at C9 to C11 in
46b. Reagents and conditions: a) 46b, LiBH4, THF, room temperature to
408C, 5.5 h; b) 2,2-dimethoxypropane, pTsOH, room temperature, 1.5 h,
37% (for two steps; Ts=tosyl).
In order to prevent epimerization at C11,[1] we first reduced
the double bond at C12=C13. The saturated derivative was
transformed into the mandelate derivative 62 for proving
the diastereomeric purity of 61 by NMR spectroscopy.[32]
Choosing the mandelic ester turned out to be advantageous
compared to the Mosher ester[33,34] because the preparation
of the latter derivative was always accompanied with the
formation of the bisacylated products as well as with partial
epimerization. When the stereochemical purity of deriva-
tives 58a,b and 60a,b, that are devoid of the alkyl side
chain, were studied in this way, unexpectedly the formation
of two diastereomers was detected.[30] Again, we assume
that epimerization took place at C9 this time during deriva-
tization with O-methyl mandelic acid.[35] However, at this
point we cannot fully exclude that epimerization at C9 takes
place during formation of aldehydes 55 and 57.
Thus, the alkyl side chain at C4 in noricumazole A (1a)
and 2’-nor noricumazole A (59a/b) appears to have a stabi-
lizing effect on the stereochemical integrity at the b-position
of aldehydes 55 to 57 as well as the corresponding coupling
products. Surprisingly, after HPLC purification of the reac-
tion mixture that resulted from desilylation of 58a, 58b, 60a
and 60b devoid of the aliphatic side, the 11-O-methylated
byproducts 63a/b and 64a/b were isolated in small amounts
(63a: 8%; 63b: 16%; 64a, 9%; 64b: 7%; Scheme 7).[36] It
can be assumed that the HPLC solvent MeOH served as
methylating agent.
Scheme 7. Syntheses of noricumazole A derivatives 58–60 and byproducts
63 and 64. Reagents and conditions: a) Dess–Martin periodinane,
NaHCO3, CH2Cl2, 08C, [49% (60% b.o.r.s.m.) for 56 (from 24), 53% for
55 (from 28a) and 70% for 57 (from 28b)]; b) tBuLi, Et2O, À788C, 1 h,
then Me2Zn, À788C, 15 min, then 39, Et2O, À788C, 3 h, 81%, d.r. 1:1
[58a,b from 55; 64%, d.r. 1:1 (59a,b from 56) and 31%, d.r. 1:1 (60a,b
from 58)]; c) BF3·Et2O, CH3CN, 08C, 12 h, 72% for 58a, 40% for 58b,
81% for 59a, 90% for 59b, 53% for 60a, 29% for 60b.
The new derivatives lacking the stereogenic center at C2’,
posed a challenge to us with respect to the determination of
the degree of racemization at C9. For these derivatives that
contain a methyl branch at C2ꢃ (e.g., 1a and 47) epimeriza-
tion at C9 leads to two diastereomers that can be distin-
guished NMR spectroscopically by analysis of the chemical
shift difference at H1ꢃ (Dd=0.02 ppm).[1] This difference is
Biological evaluation: Recently, we disclosed a new dual re-
porter gene assay of the complete HCV life cycle that
allows to identify small molecules that interfere with differ-
ent steps of the HCV replication cycle.[2] The assay is able
to discriminate between antiviral activity and cytotoxicity
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Noricumazole A Library
FULL PAPER
(towards the host cell) and at
the same time distinguishes be-
tween inhibitory influence on
viral RNA translation and rep-
lication, and other steps of the
viral life cycle. The assay is
based on human hepatocarcino-
ma cells[37] and determines the
efficiency of virus propagation
in the presence or absence of
compounds to be tested.
We specifically focused our
attention to this bioassay since
noricumazole A (1a) was iden-
tified to show antiviral activity
against this important human
pathogen in a high throughput
screening.[2,38] Notably, thia-nor-
icumazole A (49a) revealed
strong anti-HCV activity with
50% inhibitory concentration
(IC50) of 16 nm (Figure 2). Im-
portantly, it showed only mod-
erate cytotoxicity with a 50%
cytotoxic concentration (CC50)
of 350 nm, and therefore dis-
plays a good therapeutic index
(i.e., [CC50]/ACHTUNGTRENNUNG[IC50]). Although
the parent natural product, nor-
icumazole A, shows enhanced
anti-HCV activity compared to
the thia-analogue 49a, its thera-
peutic index is much worse due
to pronounced cytotoxicity. In
both cases, epimerization at
C11 (e.g., 47 and 49b) or oxida-
tion at C11 (48) led to weak
HCV inhibitors. Epimerization
at C9 has a small deactivating
effect although still good activi-
ty was retained for 9-epi-noricu-
mazole A (50a). The trend
noted for 11-epimerization
holds in the case of the 9-epi
compound, 50b, too. When the
olefinic double bond at C12=
C13 is saturated (52) anti-HCV
activity remains in the same
range as determined for noricu-
Figure 2. Inhibitory concentration (IC50) on HCV life cycle, cytotoxic concentration (CC50) on host cell
(human hepatocarcinoma cells) and therapeutic index (TI). Structural changes with reference to noricumazo-
le A (1a) are marked in gray; values in parentheses refer to standard deviations.[a]
mazole A. When altering the isobutyl side chain to the n-
butyl group (59a,b) still anti-HCV inhibitory activity is re-
tained, however, it is reduced by a factor of approximately
12 for 59b and by about 2 for 59a compared to noricuma-
zole (1a). Finally, noricumazole A derivatives (58, 60, 63
and 64) devoid of the alkyl side chain only showed low ac-
tivity with IC50 values 70-fold or more of that determined
for noricumazole (1a).
In summary, these findings highlight thia-noricumazole A
(49a) to be a lead candidate for further development as an
antiviral compound against HCV. It is highly potent with an
already excellent therapeutic index. In future studies it will
be of interest to prepare a small set of thia-noricumazole de-
rivatives and find out whether 49a impedes HCV replication
by interfering with the HCV p7 ion channel.[39]
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Received: December 23, 2011
Published online: && &&, 0000
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Noricumazole A Library
FULL PAPER
Sulfur matters! A library of 16 noricu-
mazole A derivatives was prepared by
total synthesis. Noricumazole A is
a strong inhibitor of the hepatitis C
virus (HCV) life cycle but also shows
pronounced cytotoxicity. In contrast,
the new thiazole analogue (see struc-
ture) may be regarded as a lead struc-
ture with a good therapeutic index
(CC50/IC50) of greater than 10.
Natural Products
J. Barbier, J. Wegner, S. Benson,
J. Gentzsch, T. Pietschmann,
A. Kirschning* ................ &&&& — &&&&
Total Synthesis of a Noricumazole A
Library and Evaluation of HCV Inhib-
ition
Chem. Eur. J. 2012, 00, 0 – 0
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