Total synthesis, structure revision, and absolute configuration of (+)-yatakemycin

Article abstract of DOI:10.1021/ja0472735

The total synthesis of the reported structure 2 for yatakemycin, an exceptionally potent, naturally occurring antitumor agent disclosed in 2003, and its lack of correlation with the natural product are detailed. On the basis of spectroscopic distinctions

Full text of DOI:10.1021/ja0472735

Published on Web 06/16/2004
Total Synthesis, Structure Revision, and Absolute Configuration of
(+)-Yatakemycin
Mark S. Tichenor, David B. Kastrinsky, and Dale L. Boger*
Department of Chemistry and the Skaggs Institute for Chemical Biology, The Scripps Research Institute,
10550 North Torrey Pines Road, La Jolla, California 92037
Received May 10, 2004; E-mail: boger@scripps.edu
Yatakemycin (1),1 isolated from the culture broth of Streptomyces
sp. TP-AO356, represents the newest and most potent member of
a class of antitumor compounds that includes CC-1065,2 duocar-
mycin A,3 and duocarmycin SA4 (Figure 1). Each derives its
biological properties through a characteristic and distinguishable
DNA alkylation reaction.5-9 The structure of (+)-yatakemycin was
disclosed in 2003 as 2 on the basis of extensive spectroscopic
studies.1 As such, it represented a remarkable hybrid of the
preceding natural products containing a central alkylation subunit
identical to that found in duocarmycin SA, a right-hand subunit
similar to that of the duocarmycins and a left-hand subunit similar
to the central and right-hand subunits of CC-1065. Distinct from
the preceding natural products, it represents the first naturally
occurring member of this class that contains DNA-binding subunits
flanking each side of the alkylation subunit. Since earlier studies
established that this “sandwiched” arrangement conferred remark-
able properties (enhanced DNA alkylation rate, uniquely altered
alkylation selectivity, potent cytotoxic activity) unto a series of
duocarmycin SA analogues,10 we became especially interested in
yatakemycin. With a sample of natural material provided by
Igarashi, we recently reported yatakemycin’s DNA alkylation
properties, which proved to be characteristic of such a “sandwiched”
compound.5
Figure 1. Natural products.
Herein we report the total synthesis of 2 and its lack of correlation
with the natural product (Figure 2).11 On the basis of spectroscopic
distinctions between 2 and yatakemycin, the natural product
structure was reformulated as 3, now bearing a left-hand subunit
essentially identical to the central and right-hand subunits of CC-
1065. Total synthesis of this alternative structure (thiomethyl ester
vs thioacetate) provided a compound nearly identical to, but still
subtly distinct from, the natural product. A further reformulation
of the yatakemycin structure as 1, incorporating the alternatively
substituted right-hand subunit as well as the thiomethyl ester, was
confirmed by total synthesis of (+)- and ent-(-)-1 in studies that
also unambiguously established the absolute configuration of the
natural product.
Total Synthesis of 2. Key to the synthesis of the putative
yatakemycin structure 2 was the preparation of the left-hand subunit
21 bearing the indole thioacetate. The remaining central subunit
was obtained following protocols first developed in or adopted from
our total synthesis of duocarmycin SA,12,13 and the 6-hydroxy-5-
methoxyindole-2-carboxylic acid was readily available in one step
from commercially available material.14
Synthesis of the key left-hand subunit began with 4, which was
readily available from o-vanillin.15 Phenol protection as the benzyl
ether (1.5 equiv of BnBr, 2.0 equiv of K2CO3, 25 °C, 14 h, 74%)
followed by aldehyde oxidation of 5 (1.2 equiv of NaClO2, 25 °C,
2 h, 97%) provided 6 (Scheme 1). Subsequent Curtius rearrange-
ment (1.05 equiv of DPPA, 3 equiv of Et3N, 25 °C, 3 h) with water
trap of the intermediate isocyanate (78% overall) provided amine
Figure 2. Original and first alternative structure.
7, which was N-tosylated (1.5 equiv of p-TsCl, 25 °C, 2 h, 79%)
to give 8. Nitro reduction (5 equiv of SnCl2‚2H2O, 85 °C, 0.5 h,
92%), Boc protection of amine 9 (1.2 equiv of Boc2O, 65 °C, 14
h, 89%), and subsequent oxidation of 10 (1.05 equiv of Pb(OAc)4,
25 °C, 2 h, 96%) provided the selectively activated p-quinonedi-
imine 11. The key Diels-Alder reaction of 11 with 2-methoxy-
1,3-butadiene (20 equiv, 40 °C, 48 h) provided a single cycloadduct
regioisomer that was treated with Et3N in situ to effect aromatization
to 12 (57% overall).16 The cycloaddition regioselectivity is domi-
9
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10.1021/ja0472735 CCC: $27.50 © 2004 American Chemical Society

C O M M U N I C A T I O N S
Scheme 1
Scheme 2
precludes effective coupling of the left-hand subunit.10 N-Boc
deprotection of 21 was achieved by treatment with rigorously dry
10% TFA-CH2Cl2 (25 °C, 0.5 h) under conditions that avoided
thioacetate cleavage. Following deliberate chloride for trifluoro-
acetate exchange (saturated NaCl 1:1 MeOH-H2O, evaporation,
and then trituration with THF), the corresponding hydrochloride
salt coupled smoothly with 26 to give 27. From a variety of
conditions screened, the most effective spirocyclization of 27
occurred upon treatment with saturated NaHCO3 in 1:1:1 CH3CN-
THF-H2O (25 °C, 1 h, 40%, unoptimized) providing each
enantiomer of 2. Alternative methods commonly employed resulted
in significant (1:1:1 Et3N-THF-H2O) or quantitative (NaH/DMF,
DBU/DMF) cleavage of the sensitive thioacetate.
1
The H NMR of 2 did not match that of natural yatakemycin,
nated by the stronger electron-withdrawing character of the N-
tosylimine and set the stage for cleavage of the enol ether with
release of two differentially oxidized side chains suitable for closure
to an appropriately functionalized dihydropyrroloindole.16 Thus,
ozonolysis of 12 (O3, -78 °C) followed by reductive workup (10
equiv of Me2S, 71%) provided the labile 2-hydroxyindoline 13,
which in turn was treated with 4 N HCl-EtOAc (25 °C, 0.6 h,
96%) to induce aromatization and N-Boc deprotection. Further
cyclization of 14 to lactam 15 was achieved upon treatment with
HOAc (25 °C, 24 h, 91%). At this stage, all the necessary
functionality was in place for straightforward synthesis of 21. The
indole N-tosyl group was reductively cleaved using Mg in MeOH
(20 equiv of Mg, sonication, 25 °C, 1.5 h, 96%). Subsequent indole
reduction (2.0 equiv of NaCNBH3, 25 °C, 2 h, 96%) afforded 17,
which was N-Boc protected (2.0 equiv of Boc2O, 65 °C, 1.5 h,
93%) and followed by benzyl ether hydrogenolysis (1 atm H2, 10%
Pd/C, 25 °C, 2 h, 97%) to afford 19. Treatment of 19 with
Lawesson’s reagent (1.0 equiv, 25 °C, 1.5 h, 96%) gave the
thiolactam 20, which was selectively acylated (3 equiv of Ac2O, 5
equiv of pyridine, 25 °C, 7 h, 77%) to provide 21.
After considerable experimentation, a sequence for the as-
semblage of the subunits was developed as shown in Scheme 2.
Resolution of 22 on a Chiralcel OD column provided both
enantiomers cleanly (2 × 25 cm, 2% i-PrOH-hexanes, 7 mL/min,
R ) 1.22). Boc deprotection of 22 (4 N HCl-EtOAc, 70 °C, 1 h)
followed by coupling of the indoline hydrochloride salt with 2314
(4 equiv of EDCI, 3 equiv of NaHCO3, DMF, 25 °C, 14 h, 78%)
provided 24 (Scheme 2, only one enantiomer shown). Hydrolysis
of the methyl ester (4 equiv of LiOH, THF-MeOH-H2O, 25 °C,
14 h, 78%) and benzyl ether deprotection (1 atm H2, 10% Pd/C,
THF, 2 h, 85%) afforded 26. This order of deprotections avoided
competitive spirocyclization of the alkylation subunit, which in turn
and the greatest discrepancies occurred in the left-hand subunit.
The 1H NMR chemical shift of the indole C3-H of natural
yatakemycin is found at δ 7.52, significantly downfield of the
corresponding proton in 2 at δ 6.85, and the thioacetate methyl
group of 2 occurred 0.16 δ upfield of the corresponding protons in
the natural material.1 Model substrates and computer 1H NMR
predictions supported a reassignment of the C2 substituent as a
thiomethyl ester versus thioacetate. The electron-withdrawing
character of a C2-carbonyl would account for the downfield shift
in the indole C3-H, and this reformulated structure (3) is more
consistent with other members of the natural product family. Most
notably, the left-hand subunit would now be identical to the central
and right-hand subunits of CC-1065, albeit capped as a thiomethyl
ester. Most diagnostically, aryl thiomethyl esters17 have a methyl
13C NMR chemical shift of roughly δ 11, matching the correspond-
ing chemical shift reported for yatakemycin (δ 11.14), whereas aryl
thioacetates18 occur at δ 30. Thus, yatakemycin was reformulated
as 3 and targeted for synthesis.
Total Synthesis of Revised Structure 3. The revised left-hand
subunit was prepared enlisting a late-stage intermediate in the
synthesis of the central and right-hand subunits of CC-1065.19
Selective reduction of the more reactive indole of 28 (10 equiv of
NaCNBH3, HOAc, 1 h, 83%) followed by N-Boc protection (3
equiv of Boc2O, THF, 65 °C, 2 h, 94%) and benzyl ether
hydrogenolysis (1 atm H2, 10% Pd/C, THF, 2 h, 92%) provided
31 (Scheme 3). Methyl ester hydrolysis (4 equiv of LiOH, THF-
MeOH-H2O, 25 °C, 14 h, 95%) provided the carboxylic acid 32
that was coupled with CH3SH (excess, 4 equiv of EDCI, DMF, 0
°C, 2 h, 82%) to provide the key thiomethyl ester 33. N-Boc
deprotection (4 N HCl-EtOAc, 25 °C, 0.5 h) followed by coupling
with 26 (0.7 equiv, 4 equiv of EDCI, 25 °C, 16 h, 35%,
unoptimized) and spirocyclization of 34 (saturated NaHCO3 1:1:1
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C O M M U N I C A T I O N S
Scheme 3
H2O, 25 °C, 0.7 h, 100%) provided (+)- and ent-(-)-1, which
proved to be indistinguishable from natural yatakemycin (1H NMR,
IR, MS, TLC, HPLC). Moreover, the enantiomer depicted in
Scheme 4 exhibited a strong dextrorotatory [R]D (+100) matching
that of the natural product (+99), establishing the absolute
stereochemistry and confirming an earlier tentative assignment made
in the DNA alkylation studies of 1.5
Thus, the first total synthesis of yatakemycin is described in
efforts that served to revise the natural product structure as 1 and
to establish the absolute stereochemistry. In addition to constituting
the first naturally occurring “sandwiched” member of this class of
DNA alkylating compounds, its structure now incorporates an
unusual thiomethyl ester, suggesting that protein conjugation may
contribute to its biological properties. Such studies on 1 and its
key analogues are in progress and will be reported in due course.21
Acknowledgment. We gratefully acknowledge the financial
support of the NIH (CA41986 and CA42056) and the Skaggs
Institute for Chemical Biology. We thank Professor Igarashi of the
Toyama Perfectural University for authentic samples of yatake-
mycin. D.B.K. and M.S.T. are Skaggs Fellows.
Scheme 4
Supporting Information Available: Full experimental details and
comparison 1H NMR spectra of natural and synthetic 1. This material
is available free of charge via the Internet at http://pubs.acs.org.
References
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THF-CH3CN-H2O, 25 °C, 2 h, 75%) provided both enantiomers
of 3 (only one enantiomer shown), which proved to be substantially
more stable and easy to handle relative to 2. Like 2, 3 did not
correlate with yatakemycin. However, the 1H NMR chemical shifts
in the left-hand and central subunits of 3 matched those of an
authentic sample, suggesting that this portion of the structure
incorporating the reformulated thiomethyl ester was now in place.
The remaining discrepancies rested with subtle perturbations in the
1H NMR chemical shifts in the right-hand subunit of 3. A
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1
reexamination of the H NMR and HMBC data disclosed in the
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structure identification established that the substituent locations,
but not their identity, had been defined. Computer simulations and
literature comparisons indicated that the reported chemical shifts
of C4-H and C7-H of the right-hand subunit may have been
switched and more closely matched those of a 5-hydroxy-6-
methoxyindole. Consequently, 1 was targeted for synthesis and
bears this right-hand subunit substituent reformulation as well as
the left-hand subunit thiomethyl ester.
(14) 6-Hydroxy-5-methoxyindole-2-carboxylic acid (23) was prepared by
benzyl deprotection (H2, 10% Pd/C, MeOH, 1 h, 99%) of 6-benzyloxy-
5-methoxyindole-2-carboxylic acid (Spectrum Chemical).
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Total Synthesis of (+)- and ent-(-)-Yatakemycin (1). N-Boc
deprotection of each enantiomer of 22 (4 N HCl-EtOAc, 70 °C, 1
h) and coupling with 5-hydroxy-6-methoxyindole-2-carboxylic acid
(35,20 4 equiv of EDCI, 3 equiv of NaHCO3, DMF, 25 °C, 14 h,
71%) followed by sequential methyl ester hydrolysis of 36 (4 equiv
of LiOH, THF-MeOH-H2O, 25 °C, 14 h, 74%) and benzyl ether
deprotection of 37 (1 atm H2, 10% Pd/C, THF, 2 h, 79%) provided
38 (Scheme 4, only the natural enantiomer is shown). N-Boc
deprotection of 33 (4 N HCl-EtOAc, 25 °C, 0.5 h), coupling with
38 (0.7 equiv, 4 equiv of EDCI, DMF, 25 °C, 16 h, 47%), and
subsequent spirocyclization of 39 (saturated NaHCO3 2:1 DMF-
(19) (a) Bolton, R. E.; Moody, C. J.; Rees, C. W.; Tojo, G. J. Chem. Soc.,
Perkin Trans. 1 1987, 931-935. (b) Boger, D. L.; Coleman, R. S. J. Am.
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Invergo, B. J.; Sakya, S. M.; Ishizaki, T.; Munk, S. M.; Zarrinmayeh, H.;
Kitos, P. A.; Thompson, S. C. J. Am. Chem. Soc. 1990, 112, 4623-4632.
(20) Synthesis of 5-hydroxy-6-methoxyindole-2-carboxylic acid (35): 3-ben-
zyloxy-4-methoxybenzaldehyde was condensed with N3CH2CO2Me (4
equiv, 4 equiv of NaOMe, MeOH, -15 °C, 3 h, then 0 °C, 24 h, 82%)
and then subjected to cyclization in xylenes (130 °C, 12 h, 66%). Methyl
ester hydrolysis (4 equiv of LiOH, 25 °C, 14 h, 99%) and benzyl ether
deprotection (1 atm H2, 10% Pd/C, 25 °C, 1 h, 83%) provided 35. See:
MacKenzie, A. R.; Moody, C. J.; Rees, C. W. J. Chem. Soc., Chem.
Commun. 1983, 22, 1372-1373.
(21) L-1210 IC50 ) 5 pM (indistinguishable) for both (+)- and ent-(-)-1.
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