First synthesis of nitrosporeusines, alkaloids with multiple biological activities

Article abstract of DOI:10.1016/j.tetlet.2015.01.143

Synthesis of nitrosporeusines A and B, thioester-bearing alkaloids from the Arctic Streptomyces nitrosporeus with exceptional biological activity is disclosed for the first time. In addition, we have prepared another biologically important natural product

Full text of DOI:10.1016/j.tetlet.2015.01.143

Tetrahedron Letters 56 (2015) 1252–1254
Contents lists available at ScienceDirect
Tetrahedron Letters
journal homepage: www.elsevier.com/locate/tetlet
First synthesis of nitrosporeusines, alkaloids with multiple biological
activities
Satish Chandra Philkhana, Gorakhnath R. Jachak, Vidya B. Gunjal, Nagsen M. Dhage , Ajay H. Bansode à,
⇑
D. Srinivasa Reddy
CSIR-National Chemical Laboratory, Division of Organic Chemistry, Dr. Homi Bhabha Road, Pune 411008, India
a r t i c l e i n f o
a b s t r a c t
Article history:
Synthesis of nitrosporeusines A and B, thioester-bearing alkaloids from the Arctic Streptomyces nitrospo-
reus with exceptional biological activity is disclosed for the first time. In addition, we have prepared
another biologically important natural product, maleimycin, in optically pure form using a gram-scale
enzymatic resolution method.
Received 3 December 2014
Revised 19 January 2015
Accepted 20 January 2015
Available online 24 January 2015
Ó 2015 Elsevier Ltd. All rights reserved.
Keywords:
Influenza H1N1 virus
Michael reaction
Enzymatic resolution
Nitrosporeusine
Maleimycin
- Nitrosporeusine A show
H
significant effects in various
experimental animal models of
disease:
One of the dreadful diseases rapidly spreading across the globe
is influenza commonly referred to as ‘the flu’. Influenza spreads
around the world in mostly seasonal epidemics, with an annual
attack rate estimated at 5–10% in adults and 20–30% in children.
This accounts to about 3–5 million cases of severe illness and about
250,000–500,000 deaths per annum.1 The currently existing drugs
in the market to treat influenza viruses are increasingly becoming
ineffective due to constant resistance being developed by viruses2
and hence the discovery of new inhibitors with a novel mode of
action is necessary.3 The natural products have also been used
for the identification of anti-viral compounds with novel scaffolds
so as to overcome the menace of drug resistance.4 One such natural
product family, with very good inhibitory activities against the
H1N1 virus, is the nitrosporeusines, reported by Lin and
co-workers,5 attracted our attention (Fig. 1). According to this
Letter, chemical examination from the sediments of the Arctic
Chukchi Sea actinomycete Streptomyces nitrosporeus resulted in
the isolation of two alkaloids, named as nitrosporeusines A (1)
and B (2), with an unprecedented skeleton containing benzenecar-
bo-thiocyclopenta[c]pyrrole-1,3-dione. Both the compounds 1 and
N
O
O
H
S
OH
i. Acute renal failure
ii. Rhinitis
HO
O
iii. Renal fibrosis
iv. Oral ulcers
nitrosporeusine A (1)
v. Chronic heart failure
H
- Nitrosporeusine B show
inhibitory activities against H1N1
virus in MDCK cells and hence it
has a potential to treat 'flu'.
- EC50 (inhibition of viral plaque
formation)
N
O
O
H
S
OH
HO
O
nitrosporeusine B (2)
Nitrosporeusine B = 113 μM
Oseltamivir = 67 μM
H
N
O
O
- Maleimycin inhibits Escherichia
coli, Staphylococcus aureus ,
Microbacterium phlei, and
leukemia L-1210 cells.
HO
unknown absolute
stereochemistry
maleimycin
⇑
Corresponding author. Tel.: +91 20 25902445.
E-mail address: ds.reddy@ncl.res.in (D.S. Reddy).
Figure 1. Structures of natural products along with their biological activities.
Summer project student from Department of Chemistry, NISER, Bhubaneswar,
India.
Summer project student from Department of Chemistry, Savitribai Phule Pune
Vidyapeeth (Pune University), Pune, India.
à
2 are reported to have shown very impressive multiple biological
activities (Fig. 1).5,6 In the light of the extensive and exceptional
http://dx.doi.org/10.1016/j.tetlet.2015.01.143
0040-4039/Ó 2015 Elsevier Ltd. All rights reserved.

S. C. Philkhana et al. / Tetrahedron Letters 56 (2015) 1252–1254
1253
O
HO
O
HO
O
AcO
O
O
O
Amano PS lipase,
THF
FG
FG
O
NH
NH
+
NH
NH
NH
NH
OAc
S
O
O
47% O
(+)-6
maleimycin
>98% ee (HPLC)
38%
(-)-9
O
O
O
6
"gram-scale resolution"
A
B
3
R
nitrosporeusines
and their analogs
[α]D = - 38.3°
(c, 0.77 in CHCl3)
α
[ ]D = +56.1°
O
Model reaction
(c, 1.07 in MeOH)
[α]D = +22.2°
H
S
O
NH
reported 16
aq. THF, 16 h, rt
70%
(c, 1.18 in water)
O
SH
,
y
+
NH
Amano PS
lipase,
phosphate
buffer, 92%
O
p
O
,
O
c 2
O
%
2
8
A
3
4
5
Scheme 1. Retrosynthesis and a model reaction.
AcO
O
HO
O
O
NH
NH
>99% ee (HPLC)
[α]D = -53.4°
[α]D = + 40.1°
(c, 0.54 in CHCl3)
multiple medical applications of nitrosporeusines,7 there is a dire
need to increase the availability of the natural products in
sufficient quantities by way of chemical synthesis, so as to
facilitate further research activity.
O
(+)-9
(-)-6
(c, 1.34 in MeOH)
ent-maleimycin
Scheme 3. Enzymatic resolution toward maleimycins.
Retrosynthetically, the target natural products and their analogs
A are imagined from the functionalized maleimide derivatives
B. The known compound 38 could be functionalized to obtain
the desired B using standard transformations known in the litera-
ture. To begin with, we have explored the Michael reaction on
5,6-dihydrocyclopenta[c]pyrrole-1,3(2H,4H)-dione 3 with a com-
mercially available thioacetic acid 4.9 As expected, the reaction
went smoothly to yield the desired Michael adduct 5 and a more
pleasing outcome was that the reaction could be carried out in
water at room temperature (Scheme 1).
With the success of model reaction, we began efforts toward
actual natural products. Compound 6, previously known in the lit-
erature,8 was prepared using allylic oxidation on 3 with the help of
SeO2 under microwave conditions in moderate yields.10 Alterna-
tively, the same compound can be prepared using the known pro-
tocol (bromination, trifluoroacetylation followed by hydrolysis) in
overall 44% yield.8 The other partner, thioacid 8, was synthesized
from the corresponding p-hydroxybenzoic acid 7 using Lawesson’s
reagent under microwave conditions.11 Having both partners in
hand, compounds 6 and 8 were mixed in water and stirred at
room temperature to furnish the desired racemic nitrosporeusines
A (1) and B (2) in ꢀ1:3 ratio, respectively (Scheme 2). Both the
compounds were cleanly separated on a silica gel column as pure
compounds. The spectral data (IR, 1H NMR, 13C NMR and HRMS)
of the synthesized compounds 1 and 2 were compared with those
of isolated nitrosporeusines5 and found that they are identical in
all respects. The next task was to make these compounds in opti-
cally pure form. For this purpose, we needed compound 6 in enan-
tiopure form which was not reported earlier in the literature.12 We
have considered a few options to make this desired compound, and
after a few attempts, we were successful in enzymatic resolution of
H
N
O
O
H
S
OH
OH
HO
O
(+)-1
HO
O
8
(+)-nitrosporeusine A
H2O, 12 h, rt
(natural product)
NH
O
+
65%
H
°
O
SeO2 , 1,4-dioxane, MW, 110 C
30 min, 28% (61%, brsm)
HO
O
N
O
O
(-)-6
NH
H
S
NH
or
HO
NBS, hν, CHCl3, 20min.;
AgOCOCF3.citrate buffer, pH 4
44% (for two steps)
O
O
O
3
6
(-)-2
(−)-nitrosporeusine B
(natural product)
O
OH
O
SH
H
N
O
Lawesson's reagent
acetonitrile, MW
O
H2O, 12 h
rt, 63%
H
S
°
OH
OH
7
8
100 C, 15 min
HO
OH
OH
O
53%
HO
O
NH
(-)-1
same
as above
(−)-nitrosporeusine A
H
N
H
N
+
O
H
N
O
O
O
O
O
O
H
S
(+)-6
H
S
H
S
OH
+
OH
HO
HO
O
HO
O
O
1
2
(+)-2
( )-nitrosporeusine A
( )-nitrosporeusine B
(+)-nitrosporeusine B
~ 1:3 ratio
Scheme 2. Synthesis of racemic nitrosporeusines A and B.
Scheme 4. Synthesis of enantiopure nitrosporeusines.

1254
S. C. Philkhana et al. / Tetrahedron Letters 56 (2015) 1252–1254
racemic alcohol 6.13 The Amano PS lipase mediated transesterifica-
tion using vinylacetate in THF produced enantiopure alcohol (+)-6
and corresponding acetate (À)-9 in almost equal quantities. The
enantiomeric excess (>98% ee) was determined by using the chiral
HPLC method.14 Also, we have cross checked the enantiopurity by
converting (À)-9 to (À)-6 and (+)-6 to (+)-9 (Scheme 3).15 It is
worth highlighting that the biologically active natural product
maleimycin (+)-6 and its enantiomer were prepared in enantiopure
form for the first time in good scale.
Supplementary data
Supplementary data (Detailed experimental procedures, data
comparison tables of synthetic and natural nitrosporeusines charac-
terization data, and copies of NMR spectra for all the new compounds
are provided.) associated with this article can be found, in the online
version, at http://dx.doi.org/10.1016/j.tetlet.2015.01.143.
References and notes
Maleimycin was reported to have shown interesting antibacte-
rial activities like the inhibition of Escherichia coli, Staphylococcus
aureus, and Mycobacterium phlei and anticancer activities in leuke-
mia L-1210 cells.16 Now that we have made this interesting mole-
cule and its enantiomer available through chemical synthesis, they
can be used for further biological evaluation. In addition, we have
assigned the absolute configuration (indirectly) to the natural
maleimycin (+)-6 as (R)-configuration, since we have synthesized
the natural nitrosporeusines of known absolute stereochemistry
starting from (À)-6. Maleimycin (+)-6 synthesized above was taken
toward the natural products in a way similar to that followed for
racemic compounds. To our surprise, we isolated the unnatural
enantiomers of the reported nitrosporeusines A and B.17 So, to
obtain the desired natural nitrosporeusines, compound (À)-6 was
subjected to Michael addition with p-hydroxy thiobenzoic acid 8
to give nitrosporeusines A and B. The spectral data and optical
rotation of synthetically prepared nitrosporeusines A (1) and B
(2) were in agreement with the spectral data and optical rotation
values reported by Lin and co-workers5,18 Thus, we have prepared
all the four enantiomers of nitrosporeusines (Scheme 4). As the
absolute configuration of the hydroxyl group in the two natural
products is known, we could indirectly assign their absolute ste-
reochemistry of (À)-maleimycin as S-configuration and that of
(+)-maleimycin as R-configuration, which were previously
unknown.17 Although nothing much is known about the biogenesis
of nitrosporeusines, based on our findings, it can be surmised that
natural (+)-maleimycin may not be the biogenetic precursor for the
natural nitrosporeusines A and B.
1. See Factsheet at WHO site: (accessed on 30th September, 2014) http://www.
who.int/mediacentre/factsheets/fs211/en/index.html.
2. Selected references and related internet sources on drug resistance in influenza
viruses (a) Hayden, F. G. N. Engl. J. Med. 2006, 354, 785; (b) Gubareva, L. Morb.
Mortal. Wkly Rep. 2009, 58, 433; (c) Deyde, V. M.; Xu, X.; Bright, R. A.; Shaw, M.;
Smith, C. B.; Zhang, Y.; Klimov, A. I. J. Infect. Dis. 2007, 196, 249; (d) Samson, M.;
Pizzorno, A.; Abed, Y.; Boivin, G. Antiviral Res. 2013, 98, 174; (e) http://www.
who.int/mediacentre/factsheets/fs194/en/ (accessed on 30th September,
2014).; (f) http://www.who.int/csr/disease/swineflu/frequently_asked_questions/
antivirals/resistance/en.
3. (a) Rossignol, J.-F. Antiviral Res. 2014, 110, 94; (b) Clercq, E. D. Nat. Rev. Drug
Disc. 2006, 5, 1015; (c) Meijer, A.; Rebelo-de-Andrade, H.; Correia, V.; Besselaar,
T.; Drager-Dayal, R.; Fry, A.; Gregory, V.; Gubareva, L.; Kageyama, T.; Lackenby,
A.; Lo, J.; Odagiri, T.; Pereyaslov, D.; Siqueira, M. M.; Takashita, E.; Tashiro, M.;
Wang, D.; Wong, S.; Zhang, W.; Daniels, R. S.; Hurt, A. C. Antiviral Res. 2014, 110,
31; (d) Tomassini, E. J.; Davies, M. E.; Hastings, J. C.; Lingham, R.; Mojena, M.;
Raghoobar, S. L.; Singh, S. B.; Tkacz, J. S.; Goetz, M. A. Antimicrob. Agents
Chemother. 1996, 40, 1189; (e) Hao, L.-H.; Li, Y.-P.; He, W.-Y.; Wang, H.-Q.;
Shan, G.-Z.; Jiang, J.-D.; Li, Y.-H.; Li, Z.-R. Eur. J. Med. Chem. 2012, 55, 117.
4. Selected reviews (a) Grienke, U.; Schmidtke, M.; von Grafenstein, S.; Kirchmair,
J.; Liedl, K. R.; Rollinger, J. M. Nat. Prod. Rep. 2012, 29, 11; (b) Lin, L.-T.; Hsu, W.-
C.; Lin, C.-C. J. Tradit. Complement. Med. 2014, 4, 24; (c) Kurokawa, M.; Shimizu,
T.; Watanabe, W.; Shiraki, K. Open Antimicrob. Agents J. 2010, 2, 49. and
references cited therein.
5. Yang, A.; Si, L.; Shi, Z.; Tian, Li; Liu, D.; Zhou, D.; Prokch, P.; Lin, W. Org. Lett.
2013, 15, 5366.
6. Patents filed on nitrosporeusines: (a) Chen, B. Chinese patent CN 103599102 A,
2014.; (b) Chen, B. Chinese patent CN 103585150 A, 2014.; (c) Chen, B. Chinese
patent CN 103585149 A, 2014.; (d) Chen, B. Chinese patent CN 103585147 A,
2014.; (e) Chen, B. Chinese patent CN 103585148 A, 2014.
7. It is also worth noting that nitrosporeusines A and B were highlighted as part of
the article ‘Hot off the press’ in the natural products reports published in
January, 2014. Hill, R. A.; Sutherland, A. Nat. Prod. Rep. 2014, 31, 148.
8. (a) Lee, C.-J.; Mundy, B. P. Synth. Commun. 1992, 22, 803; (b) Singh, P.;
Weinereb, S. M. Tetrahedron 1976, 32, 2379.
9. Selected refs. for related Michael addition reactions: (a) Sobhani, S.; Rezazadeh,
S. Phosphorus, Sulfur Silicon 2010, 185, 2076; (b) Marjani, K.; Khalesi, M.;
Ashouri, A.; Jalali, A.; Ziyaei-Halimehjani, A. Synth. Commun. 2011, 41, 451; (c)
Li, H.; Wang, J.; Zu, L.; Wang, W. Tetrahedron Lett. 2006, 47, 2585; (d) Kim, B. H.;
Lee, H. B.; Hwang, J. K.; Kim, Y. G. Tetrahedron: Asymmetry 2005, 16, 1215.
10. (a) Zou, Y.; Chen, C. H.; Taylor, C. D.; Foxman, B. M.; Snider, B. B. Org. Lett. 2007,
9, 1825; (b) Kad, G.; Chhabra, B. J. Chem. Res., Synop. 1997, 7, 264.
11. Rao, Y.; Lia, X.; Nagornya, P.; Hayashidaa, J.; Danishefsky, S. J. Tetrahedron Lett.
2009, 50, 6684.
The highlights of the present disclosure include (i) the first
chemical synthesis of nitrosporeusines A and B in both racemic
and enantiopure forms, (ii) scalable route under mild and green
reaction conditions, (iii) gram-scale enzymatic resolution to access
optically pure materials, (iv) synthesis of maleimycin in both enan-
tiomeric forms and (v) determination of absolute stereochemistry
of maleimycin. As the natural products are showing multiple
interesting biological activities, it is expected that the analogs of
nitrosporeusines will be highly useful. Efforts along these lines
are underway and it will be the subject of future publications.
12. The optical rotation data of Maleimycin was reported as +22.2°, but the
absolute stereochemistry was not assigned. Also no enantiopure synthesis was
reported in the literature.
13. (a) Adam, W.; Groer, P.; Humpf, H.-U.; Saha-Möller, C. R. J. Org. Chem. 2000, 65,
4919; (b) Mehta, G.; Reddy, D. S. Tetrahedron Lett. 1999, 40, 991.
14. HPLC performed on Chiralpak IB column, pet ether/2-propanol (95:5), flow
rate = 1 mL/min, 230 nm UV detector, t1 = 47.3 min and t2 = 52.6 min.
15. Optical rotation data obtained for both the acetates were in opposite sign with
Acknowledgments
same magnitude. (À)-9: [a]D À38.3 (c 0.77 in CHCl3); (+)-9: [a]D +40.1 (c 0.54 in
We thank the CSIR, New Delhi for the support through XII Five
Year Plan programs: CSC0108 (ORGIN) and CSC0109 (NICE);
Dr. Yanping Yan (Shantani Proteome Analytics Pvt. Ltd, Pune) for
her help in translating Chinese patents; Professor Subrata Ghosh,
Indian Association for the Cultivation of Science, Kolkata for helpful
discussions in preparing starting materials for the project. SCP
thanks the UGC, GRJ and VBG thank the CSIR, for the award of
research fellowships.
CHCl3).
16. Elstner, E. F.; Carnes, D. M.; Suhadolnik, R. J.; Kreishman, G. P.; Schweizer, M. P.;
Robins, R. K. Biochemistry 1973, 12, 4992.
17. We anticipated that absolute stereochemistry of natural (+)-maleimycin and
natural nitrosporeusines will be the same. However, we have isolated
unnatural (À)-nitrosporeusines from natural (+)-maleimycin and natural
isomers of nitrosporeusines from unnatural enantiomer (À)-maleimycin.
18. In the case of (À)-nitrosporeusine B, we have observed some discrepancy in the
magnitude of optical rotation. We have recorded [
a
]
D À121.8 (c 0.61 in MeOH)
and the reported value for the same is [
a
]
D
À59.3, (c 0.1 in MeOH).