Stereochemical assignment of the C1-C6 fragment of psymberin by synthesis and natural product degradation

Article abstract of DOI:10.1021/ol051396s

(Chemical Equation Presented) Psymberin is a sponge-derived natural product that shows striking selectivity as a cytotoxic agent. Conformational mobility has precluded stereochemical assignment for the acyl fragment of this molecule (psymberic acid) by NM

Full text of DOI:10.1021/ol051396s

ORGANIC
LETTERS
2005
Vol. 7, No. 19
4117-4120
Stereochemical Assignment of the
C1 C6 Fragment of Psymberin by
−
Synthesis and Natural Product
Degradation
Michael E. Green, Jason C. Rech, and Paul E. Floreancig*
Department of Chemistry, UniVersity of Pittsburgh, Pittsburgh, PennsylVania 15260
florean@pitt.edu
Received June 15, 2005
ABSTRACT
Psymberin is a sponge-derived natural product that shows striking selectivity as a cytotoxic agent. Conformational mobility has precluded
stereochemical assignment for the acyl fragment of this molecule (psymberic acid) by NMR. Herein we report stereoselective syntheses of all
four stereoisomers of psymberic acid. A comparison of the acid-mediated cyclization products of these compounds to the product of psymberin’s
acidic methanolysis showed the stereochemical configuration of this fragment to be 4S,5S.
Despite impressive advances in the use of multidimensional
NMR techniques for determining natural product structures,1
stereochemical ambiguities are still common in structures
that have high conformational mobility. Synthesis is a
powerful tool that can provide precise structural information
in cases where spectral techniques fail to afford an unam-
biguous assignment of stereochemistry or connectivity.2
While structure determination via synthesis can be highly
labor intensive, rational natural product degradation can
significantly simplify the process by providing fragments of
lesser complexity.
Figure 1. Psymberin and pederin.
Recently, the isolation and structure of acyl aminal 1
(Figure 1) was reported by Pettit and co-workers, who named
it irciniastatin A,3 and by the Crews group, who named it
psymberin.4 This compound, isolated from the South Pacific
sponges Ircinia ramosa and Psammocinia sp., shows re-
markable cytostatic activity (GI50 < 1 nM in some cell lines)
(1) For a review, see: Reynolds, W. F.; Enr´ıquez, R. G. J. Nat. Prod.
2002, 65, 221-244.
(2) Nicolaou, K. C.; Snyder, S. A. Angew. Chem., Int. Ed. 2005, 44,
1012-1044.
10.1021/ol051396s CCC: $30.25
© 2005 American Chemical Society
Published on Web 08/18/2005

and intriguingly selective cytotoxicity against several cancer
cell lines. While its interesting chemical structure, biological
activity, and relative scarcity from natural sources make
psymberin5 an attractive target for total synthesis, a firm
stereochemical assignment for this molecule has proven to
be elusive due to conformational mobility in the C1-C6
region. The C5 stereocenter was tentatively assigned as
having an (S)-configuration on the basis of psymberin’s
structural homology to the pederin (2) and mycalamide class
of molecules and its potent biological activity, but no
assignment was made for the C4 stereocenter. Recently, Kiren
and Williams reported6 an analysis of the C1-C6 fragment
using 1H and 13C NMR chemical shift homology and
postulated that the stereocenters at C4 and C5 have an anti
relationship. On the basis of the potential for acid-mediated
cleavage of the acylaminal group, we speculated that the
stereochemistry of the C1-C6 region of psymberin, which
we have named psymberic acid, could be assigned through
natural product degradation and fragment comparison with
synthetic material. Herein we report stereoselective syntheses
of all four stereoisomers of psymberic acid derivatives, a
method for effecting acid-catalyzed methanolysis of acyl
aminals, the application of this method to psymberin
degradation, and studies that provided the relative and
absolute stereochemical assignments of C4 and C5.
Scheme 2. Synthesis of the Anti Series
(R)-stereoisomer through optical rotation analysis, was
selectively protected to provide 7. Reduction of 7 provided
aldehyde 8,7 which serves as a common intermediate for the
synthesis of both the anti and syn diastereomers. Adding
methallyl trimethylsilane to 8 in the presence of BF3‚THF
proceeded with reasonable (4:1) stereocontrol9 in the Felkin-
Anh sense to provide the expected homoallylic alcohol,
which was methylated to yield ether 9. Cleaving the silyl
ether and oxidizing the primary alcohol in a two-step
sequence produced protected psymberic acid derivative 10.
While the anti and syn diastereomers could be separated after
the methylation reaction, material throughput was improved
when separation was postponed until the silyl ether was
cleaved. The opposite enantiomer (ent-10) was readily
accessed from L-serine through the same sequence.
To minimize the number of operations required to
synthesize all stereoisomers of psymberic acid derivatives
(3), we devised an approach (Scheme 1) that proceeded
Scheme 1. Stereochemically Versatile Approach to Psymberic
Acid
We postulated that changing the reaction conditions in the
methallylation step to promote a chelation-controlled addition
would allow us to access greater quantities of the syn
stereoisomers. Toward this goal, ester 7 was subjected
(Scheme 3) to a one-pot reduction with DIBAL-H followed
through stereochemically divergent methallyl group additions
into aldehyde 4. This aldehyde can be accessed from methyl
glycerate (5), which is readily available in either enantiomeric
form either from the diazotization of serine7 or through the
hydrolytic kinetic resolution of methyl glycidate.8
Scheme 3. Synthesis of the Syn Series
The synthesis of the anti diastereomer is shown in Scheme
2. Diol 6, prepared from D-serine and confirmed to be the
(3) Pettit, G. R.; Xu, J.-P.; Champuis, J. C.; Pettit, R. K.; Tackett, L. P.;
Doubek, D. L.; Hooper, J. N. A.; Schmidt, J. M. J. Med. Chem. 2004, 47,
1149-1152.
(4) Cichewicz, R. H.; Valeriote, F. A.; Crews, P. Org. Lett. 2004, 6,
1951-1954.
(5) We prefer the name psymberin for 1 because it describes the likely
biosynthesis of the molecule by symbiotic bacteria rather than sponges.
For a review on the biogenetic origins of this class of molecules, see: Piel,
J.; Butzke, D.; Fusetani, N.; Hui, D. Q.; Platzer, M.; Wen, G. P.; Matsunaga,
S. J. Nat. Prod. 2005, 68, 472-479.
(6) Kiren, S.; Williams, L. J. Org. Lett. 2005, 7, 2905-2908.
(7) Mukaiyama, T.; Shiina, I.; Iwadare, H.; Saitoh, M.; Nishimura, T.;
Ohkawa, N.; Sakoh H.; Nishimura, K.; Tani, Y.; Hasegawa, M.; Yamada,
K.; Saitoh, K. Chem. Eur. J. 1999, 5, 121-161.
(8) Schaus, S. E.; Brandes, B. D.; Larrow, J. F.; Tokunga, M.; Hansen,
K. B.; Gould, A. E.; Furrow, M. E.; Jacobsen, E. N. J. Am. Chem. Soc.
2002, 124, 1307-1315.
by the addition of methallylmagnesium chloride to provide
11 in 91% overall yield as a 1.8:1 ratio of diastereomers.
Superior diastereoselectivity in reactions of this type has been
(9) (a) Reetz, M. T.; Kesseler, K. J. Org. Chem. 1985, 50, 5434-5436.
(b) Morimoto, Y.; Mikami, A.; Kuwabe, S.-i.; Shirahama, H. Tetrahedron:
Asymmetry 1996, 7, 3371-3390.
4118
Org. Lett., Vol. 7, No. 19, 2005

effected through the addition of MgBr2 prior to Grignard
addition,10 but the modest control that we obtained was
sufficient to meet the objectives of this study. Completing
the synthesis of acid 12 proceeded through a sequence that
parallels the synthesis of 10. As before, ent-12 was prepared
through the same sequence from L-serine.
Due to the small amount of natural psymberin that was
available to us,14 our analytical techniques for the degradation
reaction were limited to GC and mass spectrometry. Psym-
berin (100-200 µg) was exposed to methanolic H2SO4 at
60 °C for 12 h (Scheme 5). Following neutralization and
In anticipation of psymberin degradation, we needed a
model system to establish conditions for acyl aminal cleav-
age. Therefore, we converted 12 to its acid chloride (Scheme
4) and coupled it to methyl imidate 13 to yield acyl imidate
Scheme 5. Psymberin Methanolysis
Scheme 4. Acyl Aminal Cleavage
concentration, the solution was analyzed by GC using a
Chiraldex G-TA column (Figure 3). A peak was observed
11
14. Reducing 14 with NaBH4 followed by cleaving the
PMB ether with DDQ provided acyl aminal 15 as a relevant
psymberin model.12 Exposing 15 to 0.1 M H2SO4 in MeOH
at 60 °C for 12 h resulted in the formation of tetrahydrofuran
16 through a sequence of acyl aminal cleavage, amide
methanolysis, and hydroxyl group addition into the tertiary
carbocation that arises from alkene protonation. The syn
relationship between the methoxy and carbomethoxy groups
was established through a NOESY experiment.13 Of note,
no epimerization at C5 was observed in this sequence.
In a similar manner, 10, ent-10, and ent-12 were converted
to their methyl esters, deprotected at C5 with ceric ammonium
nitrate, and subjected to methanolic H2SO4 at 60 °C to
provide 17, ent-17, and ent-16, respectively.13 Again no
epimerization was observed at C5. Moreover, all stereoiso-
mers were readily separable by GC using a Chiraldex G-TA
column (Figure 2).
Figure 3. GC-MS studies of psymberin degradation: (A) dilute
17; (B) psymberin degradation mixture; (C) co-injection of 17 and
psymberin degradation mixture.
that was coincident with 17 at two different temperature
gradients. Co-injection of the mixture with 17 strongly
suggested that these materials were identical. This was
confirmed by analyzing the psymberin degradation mixture
by GC-MS. The fragmentation pattern observed for the
relevant peak was identical to the fragmentation pattern of
17.15 Thus, we conclude that the stereochemistry of the acyl
fragment of psymberin can be assigned as 4S,5S, as predicted
through chemical shift analogy.6
(10) Galch, T.; Mulzer, J. Org. Lett. 2005, 7, 1311-1313.
(11) Matsuda, F.; Tomiyoshi, N.; Yanagiya, M.; Matsumoto, T. Tetra-
hedron 1988, 44, 7063-7080.
(12) This reaction provided an approximately 2:1 ratio of diastereomers.
No attempt at stereochemical assignment was made.
(13) For structural analysis and experimental details, see Supporting
Information.
(14) The psymberin that was used in these studies was generously
provided by Professor Phil Crews and his group at the University of
California, Santa Cruz.
(15) Relevant fragments result from the loss of a methyl group, the loss
of a methyl group and methanol, and the loss of the carbomethoxy group.
Figure 2. GC trace of all tetrahydrofuran stereoisomers.
Org. Lett., Vol. 7, No. 19, 2005
4119

This stereochemical assignment provides further evidence
that psymberin and pederin, as well as members from the
mycalamide,16 theopederin,17 and onnamide18 class of mol-
ecules, express their cytotoxic activity by interacting with
the same biological receptor. Strong evidence has recently
been presented that this target is the 60S ribosomal subunit.19
In addition to having the same configuration adjacent to the
carbonyl group, the C4 methoxy group and the isobutenyl
group of psymberic acid and its amides (18) spatially map
onto the anomeric methoxy group and the alkyl portion of
the tetrahydropyran ring in pederic acid and its amides (19)
(Figure 4).
mediated methanolysis showed that all psymberic acid
stereoisomers undergo cyclization reactions to form tetrahy-
drofuranyl products that are readily separable by GC.
Exposing psymberin to identical conditions resulted in the
formation of a single tetrahydrofuran, allowing the config-
uration of psymberic acid to be defined as 4S,5S. In addition
to providing valuable information for efforts in total syn-
thesis, this work supports hypotheses regarding a common
biological target for psymberin and the pederin family of
molecules.20
Acknowledgment. This work was supported by generous
funding from the National Institutes of Health (GM-62924).
We thank Professor Phil Crews and his group (UCSC) for
the sample of psymberin and Dr. Kasi Somayajula for
assistance with GC-MS experiments.
Supporting Information Available: Experimental pro-
cedures and characterization for all reactions and GC traces
for psymberin degradation reactions. This material is avail-
able free of charge via the Internet at http://pubs.acs.org.
Figure 4. Structural similarity between amides of psymberic (18)
and pederic (19)13a acids.
OL051396S
(17) Fusetani, N.; Sugawara, T.; Matsunaga, S. J. Org. Chem. 1992, 57,
3828-3832.
We have developed a versatile synthetic sequence to all
stereoisomers of protected derivatives of psymberic acid, the
C1-C6 subunit of the potent cytotoxin psymberin. Acid-
(18) Sakemi, S.; Ichiba, T.; Kohmoto, S.; Saucy, G.; Higa, T. J. Am.
Chem. Soc. 1988, 110, 4851-4853.
(19) Nishimura, S.; Matsunaga, S.; Yoshida, M.; Hirota, H.; Yokoyama,
S.; Fusetani, N. Bioorg. Med. Chem. 2005, 13, 449-454.
(20) During the review of this manuscript, the De Brabander group
reported the total synthesis of psymberin. Jiang, X.; Garcia-Fortanet, J.;
De Brabander, J. K. J. Am. Chem. Soc. 2005, 127, ASAP.
(16) (a) Perry, N. B.; Blunt, J. W.; Munro, M. H. G.; Thompson, A. M.
J. Org. Chem. 1990, 55, 223-227. (b) Perry, N. B.; Blunt, J. W.; Munro,
M. H. G.; Pannell, L. K. J. Am. Chem. Soc. 1988, 110, 4850-4851.
4120
Org. Lett., Vol. 7, No. 19, 2005