Catharanthine C16 substituent effects on the biomimetic coupling with vindoline: Preparation and evaluation of a key series of vinblastine analogues

Article abstract of DOI:10.1016/j.bmcl.2010.09.091

The examination of the catharanthine C16 substituent effects on the Fe(III)-promoted biomimetic coupling reaction with vindoline is detailed, confirming the importance of the presence of a C16 electron-withdrawing substituent, and establishing an unanticipated unique role (>10-fold) that the C16 methyl ester plays in the expression of the natural product properties. Thus, replacement of the vinblastine C16′ methyl ester with an ethyl ester (10-fold), a cyano group (100-fold), an aldehyde (100-fold), a hydroxymethyl group (1000-fold) or a primary carboxamide (>1000-fold) led to surprisingly large reductions in cytotoxic activity.

Full text of DOI:10.1016/j.bmcl.2010.09.091

Bioorganic & Medicinal Chemistry Letters 20 (2010) 6408–6410
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Catharanthine C16 substituent effects on the biomimetic coupling
with vindoline: Preparation and evaluation of a key series of vinblastine
analogues
⇑
Annie Tam, Hiroaki Gotoh, William M. Robertson, Dale L. Boger
Department of Chemistry and The Skaggs Institute for Chemical Biology, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037, United States
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 30 July 2010
Revised 13 September 2010
Accepted 15 September 2010
Available online 19 September 2010
The examination of the catharanthine C16 substituent effects on the Fe(III)-promoted biomimetic cou-
pling reaction with vindoline is detailed, confirming the importance of the presence of a C16 electron-
withdrawing substituent, and establishing an unanticipated unique role (>10-fold) that the C16 methyl
ester plays in the expression of the natural product properties. Thus, replacement of the vinblastine
C160 methyl ester with an ethyl ester (10-fold), a cyano group (100-fold), an aldehyde (100-fold), a
hydroxymethyl group (1000-fold) or a primary carboxamide (>1000-fold) led to surprisingly large reduc-
tions in cytotoxic activity.
Dedicated to A. Eschenmoser in honor of his
85th birthday.
Ó 2010 Elsevier Ltd. All rights reserved.
Keywords:
Vinblastine
Catharanthine, Vindoline
Vinblastine (1) and vincristine (2) represent the most widely
recognized members of the vinca alkaloids as a result of their clin-
ical use as antitumor drugs (Fig. 1).1–3 Originally isolated in trace
quantities from Catharanthus roseus (L.) G. Don,1 their biological
properties were among the first to be shown to arise from pertur-
bations in microtubule dynamics and inhibition of mitosis that
today is still regarded as one of the more successful drug targets
for the treatment of cancer.3–5
We recently utilized a one-pot, two-step biomimetic Fe(III)-
promoted coupling of vindoline (3) with catharanthine (4) in the
total synthesis of vinblastine and reported its extension to the prep-
aration of a series of related natural products including vincristine
and key analogues.6,7 Although key mechanistic insights into this
coupling6–9 and subsequent olefin oxidation7,10 have been disclosed
in the studies to date, the unusual differences in the substrate scope
and diastereoselectivity of the Fe(III)-promoted coupling (single
natural C160 diastereomer at 25 °C in aqueous buffer) and the more
traditional Polonovski fragmentation11,12 (5:1 at ꢀ78 °C or 1:1 at
0 °C in CH2Cl2) or 3-chloroindolenine-based13 couplings that pro-
ceed through azabenzfulvene intermediates suggests that there
are mechanistic features of the former reaction that are not yet well
understood and that affect the resulting C160 stereochemistry
(Eq. 1). In the case of the Fe(III)-promoted coupling, the attack by
vindoline formally occurs with clean inversion of the stereochemis-
try at the reacting C16 center of the catharanthine C16–C21 bond
undergoing cleavage. It has been suggested that the reaction pro-
ceeds by two single electron oxidations of catharanthine with the
initial radical cation formation occurring at the basic tertiary amine
followedby a subsequentoxidative cleavage of the C16–C21bond.8,9
Herein, we disclose the first report of the examination of catharan-
thine C16 substituent effects on the biomimetic coupling reaction,
establishing and confirming the importance of the presence of a
C16 electron-withdrawing substituent, and further disclose the
unanticipated unique role that the C16 methyl ester plays in the
expression of the natural product properties.
Et
N
21'
3
catharanthine
1) FeCl
3
H
N
16'
25 ºC, 2 h
N
N
H
OH
MeO C
2
MeO
N
H
2) NaBH
4
Et
N
OAc
Me CO Me
CO Me
2
ð1Þ
2
•+
N
+
N
−
−e
21
+
16
N
H
N
H
−
Et
Et
CO Me
2
CO Me
2
vindoline
The electron-withdrawing properties of the C16 methyl ester
group were anticipated to be key to the coupling of catharanthine
with vindoline. Consequently, only a small series of alternative
⇑
Corresponding author. Tel.: +1 858 784 7522; fax: +1 858 784 7550.
E-mail address: boger@scripps.edu (D.L. Boger).
0960-894X/$ - see front matter Ó 2010 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2010.09.091

A. Tam et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6408–6410
6409
9 failed to couple with 3, although this was not investigated in
sufficient detail to define the origin of this observation, and the cath-
aranthine analogues 10–13 that lack a C16 electron-withdrawing
substituent also, but expectedly, did not couple with 3.
OH
20'
N
N
N
H
16'
MeO2C
N
15
Since this vinblastine site has not been probed beyond reduction
or methyl ester hydrolysis and subsequent decarboxylation,3,15 each
derivative was also converted to the corresponding vinblastine
analogue either by exposing the anhydrovinblastine analogue to
the conditions developed for oxidation of the C150–C200 double
bond with installation of the C200 tertiary alcohol (Fe2(ox)3–NaBH4,
air, 0 °C, 30 min),7,10 or more directly using the one-pot, two-step
procedure of coupling and in situ oxidation.6,7 In turn, the cell
growth inhibition or cytotoxic activity of both the anhydrovinblas-
tine and vinblastine analogues were examined in three cytotoxic as-
says (72 h exposure) including the matched colon cancer cell lines
HCT116 and HCT116/VM46, the latter of which is multidrug resis-
tant by virtue of Pgp overexpression (Fig. 2).16 Whereas the impor-
tant role of the C16 methyl ester group in the coupling reaction
could be anticipated, the remarkable sensitivity of the properties
of the resulting anhydrovinblastine and vinblastine analogues to
modifications at this site was unexpected. Each series exhibited
the same striking sensitivity to the presence and nature of the
C160 substituent. Simply replacing the C160 methyl ester group with
the corresponding ethyl ester group resulted in a 10-fold loss in
activity, a nitrile or aldehyde substitution resulted in a 100-fold loss
in activity, incorporation of a hydroxymethyl group led to a 1000-
fold loss in activity, and the primary carboxamide replacement
produced a >1000-fold loss in activity. Although the importance of
this site has been reported previously with more significant modifi-
cations (R = H, CH2OH and COCH2CO2tBu),15 it is remarkable that
even the more subtle changes detailed herein have such a profound
impact on the biological properties of the natural products. Clearly,
the role of the C160 methyl ester extends well beyond facilitating the
coupling reaction in the biosynthesis of the natural product; rather
it plays an integral role in establishing the biological properties of
the natural product presumably stabilizing the interaction of vin-
blastine with tubulin.17
MeO
OH
1, Vinblastine R = CH3
2, Vincristine R = CHO
OAc
R
CO2Me
N
10
N
MeO 16
OH
N
H
16
N
Me
OAc
CO2Me
Et
3
CO2Me
4, Catharanthine
3, Vindoline
Figure 1. Natural products.
electron-withdrawing substituents were examined (R = CO2Et,
CONH2, CN, CHO, CO2H) along with derivatives where it was
removed (R = H), or replaced with an alkyl (R = CH2OH, CH3) or hy-
droxy (R = OH) group (Fig. 2).14 As anticipated, the coupling (5 equiv
FeCl3, 0.05 N aq HCl–CF3CH2OH 10:1, 25 °C, 2 h) required the pres-
ence of a C16 electron-withdrawing substituent with 4–7 providing
comparable superb conversions (79–95% vs 90% for 4) to the corre-
sponding anhydrovinblastine analogue, and the aldehyde 8 provid-
ing a perceptibly lower yield for the generation of product (49%)
reflecting some competitive aldehyde reduction upon iminium ion
reduction with NaBH4. Interestingly, the carboxylic acid derivative
Et
N
single, natural
C16' diastereomer
16'
N
N
N
H
3
1) FeCl3, 25 ºC
R
OH
16
Et
N
H
Et
R
2) NaBH4, 0 ºC
MeO
N
OAc
Me
CO2Me
requires electron-withdrawing
substituent for coupling
IC50 (nM)
L1210 HCT116 HCT116/VM46
Product
Compd
Acknowledgements
(% yield)
4, R = CO2Me
5, R = CO2Et
6, R = CN
7, R = CONH2
8, R = CHO
9, R = CO2H
10, R = H
11, R = CH2OH 11a (0%)
12, R = Me
13, R = OH
4a (90%)
5a (82%)
6a (95%)
65
620
6500
75
640
6100
600
4100
6800
10-fold
We gratefully acknowledge the financial support of the National
Institutes of Health (CA115526 and CA042056). We wish to thank
Dr. P. Hellier (P. Fabre) for the gift of catharanthine and vindoline,
Professor A. Eschenmoser for insightful discussions and NIH
(GM087948, A.T.) and JSPS (H.G.) for fellowship support. W.M.R.
is a Skaggs Fellow.
100-fold
>100-fold
100-fold
7a (79%) >10000 >10000 >10000
8a (49%)
9a (0%)
10a (0%)
6300
7000 >10000
−
−
−
−
−
−
−
−
−
−
−
−
−
−
−
12a (0%)
13a (0%)
Supplementary data
OH
Et
Supplementary data (full experimental details are provided)
associated with this article can be found, in the online version, at
doi:10.1016/j.bmcl.2010.09.091.
N
20'
Fe2(ox)3−NaBH4
air, 0 ºC, 30 min
N
16'
N
H
R
OH
Et
OAc
remarkable, unanticipated
impact on biological activity
References and notes
MeO
N
Me
CO2Me
1. (a) Noble, R. L.; Beer, C. T.; Cutts, J. H. Ann. N.Y. Acad. Sci. 1958, 76, 882; (b)
Noble, R. L. Lloydia 1964, 27, 280; (c) Svoboda, G. H.; Nuess, N.; Gorman, M. J.
Am. Pharm. Assoc. Sci. Ed. 1959, 48, 659.
2. Moncrief, J. W.; Lipscomb, W. N. J. Am. Chem. Soc. 1965, 84, 4963.
3. Review: Neuss, N.; Neuss, M. N. In The Alkaloids; Brossi, A., Suffness, M., Eds.;
Academic: San Diego, 1990; Vol. 37, p 229.
4. Reviews: (a) Pearce, H. L. In The Alkaloids; Brossi, A., Suffness, M., Eds.;
Academic: San Diego, 1990; Vol. 37, p 145; (b) Borman, L. S.; Kuehne, M. E. In
The Alkaloids; Brossi, A., Suffness, M., Eds.; Academic: San Diego, 1990; Vol. 37,
p 133; (c) Fahy, J. Curr. Pharm. Des. 2001, 7, 1181.
5. Reviews: (a) Kuehne, M. E.; Marko, I. In The Alkaloids; Brossi, A., Suffness, M.,
Eds.; Academic: San Diego, 1990; Vol. 37, p 77; (b) Potier, P. J. Nat. Prod. 1980,
43, 72; (c) Kutney, J. P. Nat. Prod. Rep. 1990, 7, 85; (d) Kutney, J. P. Synlett 1991,
IC50 (nM)
L1210 HCT116 HCT116/VM46
Product
Compd
R = CO2Me
R = CO2Et
R = CN
R = CONH2
R = CH2OH
1
6.0
60
630
6.8
70
670
600
830
7400
10-fold
100-fold
>1000-fold
1000-fold
5b
6b
7b
11ba
>10000 >10000 >10000
6500 5800 >10000
aObtained from the coupling and oxidation of 8.
Figure 2. C16 substituent effects.

6410
A. Tam et al. / Bioorg. Med. Chem. Lett. 20 (2010) 6408–6410
11; (e) Kutney, J. P. Acc. Chem. Res. 1993, 26, 559; For recent studies, see: (f)
9. For additional seminal studies on the Fe(III)-coupling to provide
anhydrovinblastine, see: (a) Szantay, C., Jr.; Balazs, J.; Bolcskei, J.; Szantay, C.
Tetrahedron 1991, 47, 1265; (b) Sundberg, R. J.; Hong, J.; Smith, S. Q.; Sabato,
M.; Tabakovic, I. Tetrahedron 1998, 54, 6259.
10. (a) Sakamoto, N.; Tan, H.; Hata, E.; Kihara, N. JP 04,164,087, 1992; Chem. Abstr.
1992, 117, 192139.; (b) Tan, H.; Sakamoto, N.; Hata, E.; Ishitoku, T.; Kihara, N.
US 5,037,977, 1990; Chem. Abstr. 1990, 113, 6663.
11. (a) Potier, P.; Langlois, N.; Langlois, Y.; Gueritte, F. J. Chem. Soc., Chem. Commun.
1975, 670; (b) Langlois, N.; Gueritte, F.; Langlois, Y.; Potier, P. J. Am. Chem. Soc.
1976, 98, 7017; (c) Sundberg, R. J.; Gadamasetti, K. G.; Hunt, P. J. Tetrahedron
1992, 48, 277.
12. (a) Kutney, J. P.; Ratcliffe, A. H.; Treasurywala, A. M.; Wunderly, S. Heterocycles
1975, 3, 639; (b) Kutney, J. P.; Hibino, T.; Jahngen, E.; Okutani, T.; Ratcliffe, A.
H.; Treasurywala, A. M.; Wunderly, S. Helv. Chim. Acta 1976, 59, 2858.
13. For additional approaches to effecting analogous couplings, see: (a) Magnus, P.;
Stamford, A.; Ladlow, M. J. Am. Chem. Soc. 1990, 112, 8210; (b) Magnus, P.;
Mendoza, J. S.; Stamford, A.; Ladlow, M.; Willis, P. J. Am. Chem. Soc. 1992, 114,
10232; (c) Kuehne, M. E.; Matson, P. A.; Bornmann, W. G. J. Org. Chem. 1991, 56,
513; (d) Bornmann, W. G.; Kuehne, M. E. J. Org. Chem. 1992, 57, 1752; (e) Kuehne,
M. E.; Zebovitz, T. C.; Bornmann, W. G.; Marko, I. J. Org. Chem. 1987, 52, 4340; (f)
Schill, G.; Priester, C. U.; Windhovel, U. F.; Fritz, H. Tetrahedron 1987, 43, 3765;
(g) Yokoshima, S.; Ueda, T.; Kobayashi, S.; Sato, A.; Kuboyama, T.; Tokuyama, H.;
Fukuyama, T. J. Am. Chem. Soc. 2002, 124, 2137; (h) Kuboyama, T.; Yokoshima, S.;
Tokuyama, H.; Fukuyama, T. Proc. Natl. Acad. Sci. U.S.A. 2004, 101, 11966.
14. Details of the substrate preparations are provided in the Supplementary data.
15. (a) Potier, P.; Guenard, D.; Zavala, F. Comp. Rend. 1979, 173, 414; (b) Barnett, C.
J.; Cullinan, G. J.; Gerzon, K.; Hoying, B. C.; Jones, W. E.; Newlon, W. M.; Poore,
G. A.; Robison, R. L.; Sweeney, M. J.; Todd, G. C.; Dyke, R. W.; Nelson, R. L. J. Med.
Chem. 1978, 21, 88.
Kuehne, M. E.; Bornmann, W. G.; Marko, I.; Qin, Y.; Le Boulluec, K. L.; Frasier, D.
A.; Xu, F.; Malamba, T.; Ensinger, C. L.; Borman, L. S.; Huot, A. E.; Exon, C.;
Bizzarro, F. T.; Cheung, J. B.; Bane, S. L. Org. Biomol. Chem. 2003, 1, 2120; (g)
Miyazaki, T.; Yokoshima, S.; Simizu, S.; Osada, H.; Tokuyama, H.; Fukuyama, T.
Org. Lett. 2007, 9, 4737; (h) Voss, M. E.; Ralph, J. M.; Xie, D.; Manning, D. D.;
Chen, X.; Frank, A. J.; Leyhane, A. J.; Liu, L.; Stevens, J. M.; Budde, C.; Surman, M.
D.; Friedrich, T.; Peace, D.; Scott, I. L.; Wolf, M.; Johnson, R. Bioorg. Med. Chem.
Lett. 2009, 19, 1245; (i) Shao, Y.; Zhang, H.; Ding, H.; Quan, H.; Lou, L.; Hu, L. J.
Nat. Prod. 2009, 72, 1170; (j) Sheng, L. X.; Da, Y. X.; Long, Y.; Hong, L. Z.; Cho, T.
P. Bioorg. Med. Chem. Lett. 2008, 18, 4602; (k) Passarella, D.; Giardini, A.; Peretto,
B.; Fontana, G.; Sacchetti, A.; Silvani, A.; Ronchi, C.; Cappelletti, G.; Cartelli, D.;
Borlak, J.; Daneili, B. Bioorg. Med. Chem. 2008, 16, 6269.
6. Ishikawa, H.; Colby, D. A.; Boger, D. L. J. Am. Chem. Soc. 2008, 130, 420.
7. For supporting synthetic studies, see: (a) Ishikawa, H.; Colby, D. A.; Seto, S.; Va,
P.; Tam, A.; Kakei, H.; Rayl, T. J.; Hwang, I.; Boger, D. L. J. Am. Chem. Soc. 2009,
131, 4904; (b) Va, P.; Campbell, E. L.; Robertson, W. M.; Boger, D. L. J. Am. Chem.
Soc. 2010, 132, 8489; (c) Sasaki, Y.; Kato, D.; Boger, D. L. J. Am. Chem. Soc. 2010,
132, 13533; For related studies, see: (d) Kato, D.; Sasaki, Y.; Boger, D. L. J. Am.
Chem. Soc. 2010, 132, 3685; (e) Ishikawa, H.; Elliott, G. I.; Velcicky, J.; Choi, Y.;
Boger, D. L. J. Am. Chem. Soc. 2006, 128, 10596; (f) Elliott, G. I.; Fuchs, J. R.; Blagg,
B. S. J.; Ishikawa, H.; Tao, H.; Yuan, Z.; Boger, D. L. J. Am. Chem. Soc. 2006, 128,
10589; (g) Elliott, G. I.; Velcicky, J.; Ishikawa, H.; Li, Y.; Boger, D. L. Angew.
Chem., Int. Ed. 2006, 45, 620; (h) Choi, Y.; Ishikawa, H.; Velcicky, J.; Elliott, G. I.;
Miller, M. M.; Boger, D. L. Org. Lett. 2005, 7, 4539; (i) Yuan, Z.; Ishikawa, H.;
Boger, D. L. Org. Lett. 2005, 7, 741; (j) Ishikawa, H.; Boger, D. L. Heterocycles
2007, 72, 95; (k) Wilkie, G. D.; Elliott, G. I.; Blagg, B. S. J.; Wolkenberg, S. E.;
Soenen, D. B.; Miller, M. M.; Pollack, S.; Boger, D. L. J. Am. Chem. Soc. 2002, 124,
11292; (l) Wolkenberg, S. E.; Boger, D. L. J. Org. Chem. 2002, 67, 7361; (m)
Campbell, E. L.; Zuhl, A. M.; Liu, C.; Boger, D. L. J. Am. Chem. Soc. 2010, 132, 3009;
(n) Boger, D. L. Tetrahedron 1983, 39, 2869; (o) Boger, D. L. Chem. Rev. 1986, 86,
781.
16. (a) Lampidis, T. J.; Kolonias, D.; Podona, T.; Israel, M.; Safa, A. R.; Lothstein, L.;
Savaraj, N.; Tapiero, H.; Priebe, W. Biochemistry 1997, 36, 2679; (b) Perego, P.;
De Cesare, M.; De Isabella, P.; Carenini, N.; Beggiolin, G.; Pezzoni, G.; Palumbo,
M.; Tartaglia, L.; Prtesi, G.; Pisano, C.; Carminati, P.; Scheffer, G. L.; Zunino, F.
Cancer Res. 2001, 61, 6034.
8. (a) Vukovic, J.; Goodbody, A. E.; Kutney, J. P.; Misawa, M. Tetrahedron 1988, 44,
325; For an analogous electrochemical coupling (0.6 V in buffer; NaBH4) to
provide anhydrovinblastine, see: (b) Gunic, E.; Tabakovic, I.; Gasic, M. J. J. Chem.
Soc., Chem. Commun. 1993, 1496; For an enzymatic coupling, see: (c) Sagui, F.;
Chirivi, C.; Fontana, G.; Nicotra, S.; Passarella, D.; Riva, S.; Danieli, B.
Tetrahedron 2009, 65, 312.
17. Gigant, B.; Wang, C.; Ravelli, R. B. G.; Roussi, F.; Steinmetz, M. O.; Curmi, P. A.;
Sobel, A.; Knossow, M. Nature 2005, 435, 519.