Large-scale asymmetric synthesis of the bioprotective agent JP4-039 and analogs

Article abstract of DOI:10.1021/ol200567p

Chemical equations presented. JP4-039 is a novel nitroxide conjugate capable of crossing lipid bilayer membranes and scavenging reactive oxygen species (ROS). An efficient and scalable one-pot hydrozirconation- transmetalation-imine addition methodology h

Full text of DOI:10.1021/ol200567p

ORGANIC
LETTERS
2011
Vol. 13, No. 9
2318–2321
Large-Scale Asymmetric Synthesis of the
Bioprotective Agent JP4-039 and Analogs
†
†
†
‡
ꢀ
Marie-Celine Frantz, Joshua G. Pierce, Joan M. Pierce, Li Kangying,
Wan Qingwei,‡ Matthew Johnson,‡ and Peter Wipf*,†
Department of Chemistry, University of Pittsburgh, Pittsburgh, Pennsylvania 15260,
United States, and Asymchem Inc., 600 Airport Boulevard #1000, Morrisville,
North Carolina 27560, United States
pwipf@pitt.edu
Received March 2, 2011
ABSTRACT
JP4-039 is a novel nitroxide conjugate capable of crossing lipid bilayer membranes and scavenging reactive oxygen species (ROS). An efficient
and scalable one-pot hydrozirconationꢀtransmetalationꢀimine addition methodology has been developed for its asymmetric preparation.
Furthermore, this versatile methodology allows for the synthesis of cyclopropyl and fluorinated analogs of the parent lead structure.
The 4-amino-Tempo (4-AT) derivative JP4-039 ((S)-6a,
Scheme 2) is a lead structure among a new generation of
peptidomimetic conjugates targeted to mitochondria and
effective at scavenging reactive oxygen species (ROS) such
as the superoxide radical anion.1,2 In particular, JP4-039
has been shown to protect cells from radiation damage2b
and to prolong the survival of mice subjected to high levels
of ionizing irradiation.2c Moreover, the ameliorating ef-
fects of JP4-039 in irradiation-induced delay of bone
wound healing were demonstrated in a murine model of
combined bone wound/irradiation injury.2d JP4-039 and
its larger congener, XJB-5-131, are found to enrich in
mitochondria by a factor of 30ꢀ600 times over the cyto-
solic concentration, at least in part due to their affinity to
the mitochondrial lipid, cardiolipin.1b,2a There, they serve
to scavenge ROS and prevent hydroperoxidation of car-
diolipin by cycling between nitroxide, hydroxylamine, and
nitroxonium redox states.3 Their design was based on the
structure of the antibiotic gramicidin S; the alkene peptide
isostere replaces a polar internal amide bond and thus in-
creases conformational rigidity and membrane permeabi-
lity.1a,g Inorder tofurther explore the therapeutic potential
of JP4-039, a robust synthetic route was required to
prepare multigram quantities of highly pure material.
JP4-039 contains a single asymmetric carbon atom as
part of an alkene isostere dipeptide moiety composed of
leucineandglycineresidues. Numerousmethodshavebeen
† University of Pittsburgh.
‡ Asymchem Inc.
(1) (a) Wipf, P.; Xiao, J.; Jiang, J.; Belikova, N. A.; Tyurin, V. A.;
Fink, M. P.; Kagan, V. E. J. Am. Chem. Soc. 2005, 127, 12460. (b) Jiang,
J.; Kurnikov, I.; Belikova, N. A.; Xiao, J.; Zhao, Q.; Amoscato, A. A.;
Braslau, R.; Studer, A.; Fink, M. P.; Greenberger, J. S.; Wipf, P.; Kagan,
V. E. J. Pharmacol. Exp. Ther. 2007, 320, 1050. (c) Kanai, A.; Zabbarova,
I.; Amoscato, A.; Epperly, M.; Xiao, J.; Wipf, P. Org. Biomol. Chem. 2007,
5, 307. (d) Macias, C. A.; Chiao, J. W.; Xiao, J.; Arora, D. S.; Tyurina,
Y. Y.; Delude, R. L.; Wipf, P.; Kagan, V. E.; Fink, M. P. Ann. Surg. 2007,
245, 305. (e) Jiang, J.; Belikova, N. A.; Hoye, A. T.; Zhao, Q.; Epperly,
M. W.;Greenberger, J. S.; Wipf, P.;Kagan, V. E.Int. J. Radiat. Oncol. Biol.
Phys. 2008, 70, 816. For reviews on mitochondria targeting agents, see: (f)
Hoye, A. T.;Davoren, J. E.;Wipf, P.;Fink, M. P.;Kagan, V. E. Acc. Chem.
Res. 2008, 41, 87. (g) Frantz, M.-C.; Wipf, P. Environ. Mol. Mutagen. 2010,
51, 462.
(2) (a) Kagan, V. E.; Wipf, P.; Stoyanovsky, D.; Greenberger, J. S.;
Borisenko, G.; Belikova, N. A.; Yanamala, N.; Samhan Arias, A. K.;
Tungekar, M. A.; Jiang, J.; Tyurina, Y. Y.; Ji, J.; Klein-Seetharaman, J.;
Pitt, B. R.; Shvedova, A. A.; Bayir, H. Adv. Drug Delivery Rev. 2009, 61,
1375. (b) Rajagopalan, M. S.; Gupta, K.; Epperly, M. W.; Franicola, D.;
Zhang, X.; Wang, H.; Zhao, H.; Tyurin, V. A.; Pierce, J. G.; Kagan,
V. E.; Wipf, P.; Kanai, A. J.; Greenberger, J. S. In Vivo 2009, 23, 717. (c)
Epperly, M. W.; Goff, J. P.; Li, S.; Gao, X.; Wipf, P.; Dixon, T.; Wang,
H.; Franicola, D.; Shen, H.; Rwigema, J.-C. M.; Kagan, V.; Bernard,
M.; Greenberger, J. S. In Vivo 2010, 24, 811. (d) Gokhale, A.; Rwigema,
J.-C.; Epperly, M. W.; Glowacki, J.; Wang, H.; Wipf, P.; Goff, J. P.;
Dixon, T.; Patrene, K.; Greenberger, J. S. In Vivo 2010, 24, 377.
(3) Tyurina, Y. Y.; Tyurin, V. A.; Kaynar, A. M.; Kapralova, V. I.;
Wasserloos, K.; Li, J.; Mosher, M.; Wright, L.; Wipf, P.; Watkins, S.;
Pitt, B. R.; Kagan, V. E. Am. J. Physiol. Lung Cell Mol. Physiol. 2010,
299, L73.
r
10.1021/ol200567p
Published on Web 03/31/2011
2011 American Chemical Society

developed to generate these R-chiral allylic amines.4 For
example, Overman developed the rearrangement of allylic
trichloroacetimidates,4b Berkowitz described an approach
which relies on a Ni(0)-mediated allylic amination,4c and
Krische reported a CꢀC bond-forming hydrogenation for
this purpose.4k Overall, vinyl addition to imines remains the
most commonly used strategy to prepare chiral allylic
amines.4dꢀk Highly effective protocols for this route are
based on the reductive coupling of alkynes,4dꢀf the diaster-
eoselective addition of vinylorganometallic species,4gꢀi and
the acylvinyl anion addition.4j
Scheme 2. Synthesis of the (E)-Alkene Isosteres (S)-6aꢀc
Scheme 1. Synthesis of the Common Amine Intermediate (S)-3
Figure 1. X-ray structure of JP4-039.
diastereomerically pure (>20:1 dr) allylic amine according to
1H NMR analysis of the crude reaction mixture (Scheme 1).
A four-membered chelate model has been proposed to
account for this excellent diastereoselectivity.4a,c Treat-
ment of the crude reaction mixture with HCl in diethyl
ether yielded the amine hydrochloride (S)-3 on multigram
scale and in 81% yield over three steps from 3-butyn-1-ol.
The N-tert-butylsulfinimine (R)-1 could be easily obtained
from isovaleraldehyde in 90% yield and proved remark-
ably stable to storage.
We were able to use the allylic sulfinylamine (S)-3 as a
linchpin intermediate for the preparation of JP4-039 as well
as a first series of analogs. Thus, acylation of (S)-3 with
either a Boc, Cbz, or diphenylphosphinoyl group, followed
by TBAF-mediated deprotection of the silyl ether, afforded
the alcohols (S)-5aꢀc (Scheme 2). Jones oxidation to the
corresponding carboxylic acids and final coupling with
4-AT using the EDCI/HOBt/DMAP protocol provided the
desired alkene isosteres (S)-6aꢀc in moderate to good yields.
This methodology was further adapted for a >150 g
scale preparation of JP4-039 ((S)-6a), obtained in 32%
overall yield, 99% purity, and >99% ee based on HPLC
and chiral SFC analysis. At this scale, the target compound
was isolated from a 20:3 mixture of n-hexane/EtOAc as a
crystalline solid. The X-ray structure of (S)-6a is in good
agreement with a type II0 β-turn (Figure 1). The distance of
Our group has previously developed an efficient Zr-
based method for the asymmetric synthesis of allylic
amines.5 Hydrozirconation of alkyne 26 with Cp2ZrHCl,7
followed by transmetalation to trimethylaluminum and
subsequent addition to chiral sulfinimine8 (R)-1, provided a
(4) For a review on allylic amination, see: (a) Johannsen, M.; Jorgensen,
K. A. Chem. Rev. 1998, 98, 1689. For selected examples, see: (b) Anderson,
C. E.; Overman, L. E. J. Am. Chem. Soc. 2003, 125, 12412. (c) Berkowitz,
D. B.; Maiti, G. Org. Lett. 2004, 6, 2661. (d) Grossman, R. B.; Davis, W. M.;
Buchwald, S. L. J. Am. Chem. Soc. 1991, 113, 2321. (e) Patel, S. J.; Jamison,
T. F. Angew. Chem., Int. Ed. 2004, 43, 3941. (f) Ngai, M.-Y.; Barchuk, A.;
Krische, M. J. J. Am. Chem. Soc. 2007, 129, 12644 and references cited
therein. (g) Denmark, S. E.; Weber, T.; Piotrowski, D. W. J. Am. Chem. Soc.
1987, 109, 2224. (h) Cogan, D. A.; Liu, G.; Ellman, J. Tetrahedron 1999, 55,
8883. (i) Brak, K.; Ellman, J. A. J. Am. Chem. Soc. 2009, 131, 3850. (j)
Reynolds, T. E.; Binkley, M. S.; Scheidt, K. A. Org. Lett. 2008, 10, 5227. (k)
Skucas, E.; Ngai, M.-Y.; Komanduri, V.; Krische, M. J. Acc. Chem. Res.
2007, 40, 1394.
(5) (a) Wipf, P.; Nunes, R. L.; Ribe, S. Helv. Chim. Acta 2002, 85,
3478. (b) Wipf, P.; Kendall, C.; Stephenson, C. R. J. J. Am. Chem. Soc.
2003, 125, 761. (c) Wipf, P.; Pierce, J. G. Org. Lett. 2006, 8, 3375. For a
review, see: (d) Wipf, P.; Kendall, C. Topics Organomet. Chem. 2004, 8, 1.
(6) Nicolaou, K. C.; Lizos, D. E.; Kim, D. W.; Schlawe, D.; de
Noronha, R. G.; Longbottom, D. A.; Rodriquez, M.; Bucci, M.; Cirino,
G. J. Am. Chem. Soc. 2006, 128, 4460.
(7) (a) Buchwald, S. L.; LaMaire, S. J.; Nielsen, R. B.; Watson, B. T.;
King, S. M. Org. Synth. 1998, Coll. Vol. 9, 162. (b) Wipf, P.; Jahn, H.
Tetrahedron 1996, 52, 12853 and references cited therein.
(8) (a) Staas, D. D.; Savage, K. L.; Homnick, C. F.; Tsou, N. N.; Ball,
R. G. J. Org. Chem. 2002, 67, 8276. (b) For a review on the use of
sulfinimines, see: Robak, M. T.; Herbage, M. A.; Ellman, J. A. Chem.
Rev. 2010, 110, 3600.
Org. Lett., Vol. 13, No. 9, 2011
2319

˚
3.30 A for the i to i þ 3 intramolecular hydrogen bond
between the carbonyl oxygen of the Boc function and the
nitrogen atom of 4-AT is indicative of a weak covalent
component in this H-bond.
Scheme 5. Synthesis of 10
The (R)-enantiomer of JP4-039 (R)-6a was obtained in
52% overall yield according to an analogous synthetic
route starting from (S)-sulfinimine (S)-1 (Scheme 3). Chir-
al SFC on a Chiralpak-IC phase was used to determine an
ee of 96.6% for alcohol (S)-5a and 98.0% for its enantio-
mer (R)-5a.
Scheme 3. Synthesis of (R)-6a
Scheme 4. Synthesis of the Cyclopropyl Isosteres 8aꢀb
Figure 2. X-ray structure of 10.
an X-ray analysis of a suitable crystalline derivative. Hydro-
genolysis, coupling with p-bromobenzoyl chloride, TBAF-
desilylation, and treatment with 1-naphthylisocyanate af-
forded urea 10, which gave fine colorless needles suitable
for X-ray diffraction (Scheme 5). The X-ray analysis con-
firmed the anti-configuration of the cyclopropane vs the CꢀN
bond (Figure 2). This diastereoselectivity had been previously
noted by our group for the dimethylzinc-mediated addition
of alkenylzirconocenes to N-diphenylphosphinoyl imines,
which provided diastereomerically pure C-cyclopropylalkyl-
amines.5b,9f The high level of anti-selectivity is also consistent
with diastereoselectivities observed in the SimmonsꢀSmith
cyclopropanation of allylic ethers.12
Somewhat surprisingly, the Boc group on (S)-4a was not
compatible with the SimmonsꢀSmith conditions. The
desired intermediate 7a was therefore prepared by hydro-
genolysis of 7b followed by Boc protection of the resulting
amine (Scheme 4). Subsequent TBAF-desilylation of 7a
and 7b, Jones oxidation, and coupling with 4-AT afforded
the cyclopropyl isosteres 8a and 8b.
An isosteric replacement of the (E)-alkene bond with a
cyclopropyl moiety was also investigated as part of our medi-
cinal chemistry program.9 Charette-modified Simmonsꢀ
Smith cyclopropanation10 of the Cbz-protected allylic amine
(S)-4b with the Zn(CH2I)2•DME complex11 provided the
cyclopropyl analog 7b on gram scale in 54% yield (65%
based on recovered intermediate (S)-4b) over two steps
(Scheme 4). Only one diastereomer was isolated after chro-
matography on SiO2 (>20:1 dr by 1H NMR). Since 7b was
not crystalline and spectroscopic analysis did not allow an
unambiguous assignment of its configuration, we resorted to
(9) (a) Wipf, P.; Henninger, T. C.; Geib, S. J. J. Org. Chem. 1998, 63,
6088. (b) Xiao, J.; Weisblum, B.; Wipf, P. J. Am. Chem. Soc. 2005, 127,
5742. (c) Xiao, J.; Weisblum, B.; Wipf, P. Org. Lett. 2006, 8, 4731. (d)
Wipf, P.; Xiao, J. Org. Lett. 2005, 7, 103. (e) Wipf, P.; Werner, S.; Woo,
G. H. C.; Stephenson, C. R. J.; Walczak, M. A. A.; Coleman, C. M.;
Twining, L. A. Tetrahedron 2005, 61, 11488. (f) Wipf, P.; Stephenson,
C. R. J. Org. Lett. 2005, 7, 1137. For a review, see: (g) Wipf, P.; Xiao, J.;
Stephenson, C. R. J. Chimia 2009, 63, 764.
Finally, a difluorinated analog of JP4-039 was envisioned
to enhance the bioavailability of the agent. The methyl ester
(10) (a) Simmons, H. E.; Smith, R. D. J. Am. Chem. Soc. 1959, 81, 4256.
(b) Charette, A. B.; Prescott, S.; Brochu, C. J. Org. Chem. 1995, 60, 1081.
(11) (a) Wittig, G.; Schwarzenbach, K. Angew. Chem. 1959, 71, 652.
(b) Furukawa, J.; Kawabata, N.; Nishimura, J. Tetrahedron 1968, 24, 53.
(12) (a) Charette, A.; Lebel, H.; Gagnon, A. Tetrahedron 1999, 55,
8845. (b) For a review on the SimmonsꢀSmith cyclopropanation, see:
Charette, A. B.; Beauchemin, A. Org. React. 2004, 58, 1.
2320
Org. Lett., Vol. 13, No. 9, 2011

tion of both enantiomers of a chiral allylic amine and the
corresponding dipeptide alkene isosteres. This straightfor-
ward approach enabled the synthesis of the bioprotective
Tempo-conjugate JP4-039 on a 160 g scale. Noteworthy is
also the ready access to diverse analogs of the parent
compound, including the β,γ-cyclopropylamine isosteres
and difluoromethylene derivatives. SAR studies of the
JP4-039 scaffold are ongoing, and the synthesis of further
derivatives will be reported in due course, along with their
antioxidant and biological activities.
Scheme 6. Synthesis of the Difluoro Analog (S)-13
Acknowledgment. This project was supported by a
BARDA contract (HHS0100200800062C), the NIH/
NIAID CMCR program (AI068021), and the NIH/
NIGMS CMLD program (GM067082). We thank the
NMR and MS facilities at the University of Pittsburgh
for their services, Dr. Steven J. Geib (University of
Pittsburgh) for X-ray analyses, Mr. David M. Arnold
(University of Pittsburgh) for SFC analyses, and Ms.
Kayla R. Lloyd (University of Pittsburgh) for LC-MS
analyses and for technical and administrative assistance.
M.C.F. wishes to thank Mr. Chris J. Rosenker (University
of Pittsburgh) and Dr. Gary C. Davis (University of
Pittsburgh) for helpful comments. P.W. thanks Profs.
Valerian Kagan, Laura Niedernhofer, Mike Epperly, Joel
Greenberger, and Paul Robbins (all at University of
Pittsburgh) for stimulating discussions and collaborative
studies on the bioprotective properties of the JP4 series.
(S)-11, prepared from alcohol (S)-5a by Jones oxidation
and esterification of the acid with TMS-diazomethane, was
treated with 3 equiv of the fluorinating agent N-fluoro-
N-(phenylsulfonyl)benzene-sulfonamide (accufluor, NFSi)13
and 2.3 equiv of NaHMDS in THF at ꢀ78 °C to afford the
desired R,R-difluoroester (S)-12 in 86% yield (Scheme 6).14
Saponification with tetra-butylammonium hydroxide
(TBAH)15 and condensation with 4-AT provided the di-
fluorinated JP4-039 analog (S)-13 in good yield. The biolo-
gical activities of (S)-13 as well as (R)-6a are currently under
investigation.
In summary, the hydrozirconationꢀtransmetalationꢀ
imine addition methodology was extended to the prepara-
Supporting Information Available. Experimental pro-
cedures, X-ray and complete spectroscopic data for new
compounds. This material is available free of charge via
the Internet at http://pubs.acs.org.
(13) Differding, E.; Ofner, H. Synlett 1991, 187.
(14) Liu, S.; Dockendorff, C.; Taylor, S. D. Org. Lett. 2001, 3, 1571.
(15) Abdel-Magid, A. F.; Cohen, J. H.; Maryanoff, C. A.; Shah,
R. D.; Villani, F. J.; Zhang, F. Tetrahedron Lett. 1998, 39, 3391.
Org. Lett., Vol. 13, No. 9, 2011
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