pathway is initiated by the binding of the SHH ligand to the
cellular membrane receptor Patched (PTC), which relieves
the PTC-mediated inhibition of the transmembrane protein
Smoothened (SMO) (Figure 2).5,6 Activated SMO transduces
spirotetrahydrofuran ring, followed by aromatization of the
D ring (Scheme 1).14 Unlike cyclopamine, 2 does not act as
Scheme 1
.
Acid-Mediated Conversion of Cyclopamine 1 to
Veratramine 2
an SHH antagonist and causes hemolysis by targeting other
receptors.15,16
Two strategies have been reported to address the issues
of water solubility and acid lability of 1: (1) covalent
modification of naturally occurring cyclopamine 1 to generate
structurally related and metabolically stable lead structures,9
as pioneered by Adams and co-workers at Infinity Pharma-
ceuticals, and (2) identification of small-molecule SHH
antagonists through high-throughput screening.17 The first
approach, however, relies on the availability of 1, which is
prohibitively expensive ($1990/gram), while preliminary data
suggests limited success with the Curis and Genentech
structures.18 Consequently, there is an urgent need to identify
readily available potent inhibitors of SHH as lead structures
for the development of new cancer chemotherapies.
We have opted to explore a third approach, which is not
dependent on the availability of 1 and yet generates new
lead compounds that closely resemble 1 in both structure
and function. The advantage of cyclopamine-like structures
is underscored by the observation that cyclopamine crosses
the blood-brain barrier, a critical property for the develop-
ment of clinical candidates for the treatment of brain
malignancies.12
Figure 2. Hedgehog signaling pathway and the structure of
cyclopamine 1.
the signal to the nucleus to regulate gene expression via Gli
transcription factors. Beachy and co-workers have established
that cyclopamine 1 disrupts this pathway by inhibition of
SMO.7 The teratogenicity described above has not hampered
interest in the development of this class of compounds.8,9
Treatment of cancer cells with cyclopamine 1 induces a
decrease in proliferation, an increase of apoptosis, and/or a
decrease of metastasis.10-13 Therefore, cyclopamine and its
related structures hold great promise in cancer chemotherapy.
Despite the attractive pharmacological profile against a
number of cancer xenografts, in vivo evaluation of cyclo-
pamine has been hampered by its poor aqueous solubility
(ca. 5 µg/mL) and acid lability. Cyclopamine readily converts
to veratramine 2 via acid-catalyzed ring-opening of the
(7) Chen, J. K.; Taipale, J.; Cooper, M. K.; Beachy, P. A. Inhibition of
Hedgehog signaling by direct binding of cyclopamine to Smoothened. Genes
DeV. 2002, 16, 2743–2748.
The difference in teratogenicity between cyclopamine 1
and the close structural analogue tomatidine 3 (Figure 3;
nonteratogenic) has been attributed to the difference in the
orientation of the nitrogen atom relative to the steroid plane
in 1 and 3. The C-nor-D-homo framework of 1 can thus be
viewed as a scaffold that orients the E/F heterobicyclic
moiety orthogonal to the steroidal ring system, with the
F-ring nitrogen atom on the R-face of the steroid plane
(8) Janardanannair, S.; Adams, J.; Ripka, A. S.; Hospital, M. R.;
Tremblay, M. Methods for preparation cyclopamine analogs and use thereof
in treating cancers. 2005-US30406; 2006026430, 20050826, 2006
.
(9) Tremblay, M. R.; Nevalainen, M.; Nair, S. J.; Porter, J. R.; Castro,
A. C.; Behnke, M. L.; Yu, L.-C.; Hagel, M.; White, K.; Faia, K.; Grenier,
L.; Campbell, M. J.; Cushing, J.; Woodward, C. N.; Hoyt, J.; Foley, M. A.;
Read, M. A.; Sydor, J. R.; Tong, J. K.; Palombella, V. J.; McGovern, K.;
Adams, J. Semisynthetic Cyclopamine Analogues as Potent and Orally
Bioavailable Hedgehog Pathway Antagonists. J. Med. Chem. 2008, 51,
6646–6649
.
(10) Berman, D. M.; Karhadkar, S. S.; Hallahan, A. R.; Pritchard, J. I.;
Eberhart, C. G.; Watkins, D. N.; Chen, J. K.; Cooper, M. K.; Taipale, J.;
Olson, J. M.; Beachy, P. A. Medulloblastoma growth inhibition by hedgehog
(15) Nagata, R.; Izumi, K. Veratramine-induced behavior associated with
serotonergic hyperfunction in mice. Jpn. J. Pharmacol. 1991, 55, 129–
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pathway blockade. Science 2002, 297, 1559–1561
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(16) Thron, C. D.; McCann, F. V. Pharmacological tests of the
mechanism of the periodic rhythm caused by veratramine in the sinoatrial
(11) Dahmane, N.; Sanchez, P.; Gitton, Y.; Palma, V.; Sun, T.; Beyna,
M.; Weiner, H.; Ruiz i Altaba, A. The Sonic Hedgehog-Gli pathway
regulates dorsal brain growth and tumorigenesis. DeVelopment 2001, 128,
node of the guinea pig. Gen. Pharmacol. 1998, 32, 81–89
.
(17) Williams, J. A.; Guicherit, O. M.; Zaharian, B. I.; Xu, Y.; Chai,
L.; Wichterle, H.; Kon, C.; Gatchalian, C.; Porter, J. A.; Rubin, L. L.; Wang,
F. Y. Identification of a small molecule inhibitor of the hedgehog signaling
pathway: effects on basal cell carcinoma-like lesions. Proc. Natl. Acad.
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(12) Sanchez, P.; Ruiz i Altaba, A. In vivo inhibition of endogenous
brain tumors through systemic interference of Hedgehog signaling in mice.
Mech. DeV. 2005, 122, 223–230
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(13) Stecca, B.; Mas, C.; Clement, V.; Zbinden, M.; Correa, R.; Piguet,
V.; Beermann, F.; Ruiz, I. A. A. Melanomas require HEDGEHOG-GLI
signaling regulated by interactions between GLI1 and the RAS-MEK/AKT
(18) Hosoya, T.; Arai, M. A.; Koyano, T.; Kowithayakorn, T.; Ishibashi,
M. Naturally occurring small-molecule inhibitors of hedgehog/GLI-mediated
transcription. ChemBioChem 2008, 9, 1082–1092.
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(19) Keeler, R. F. Cyclopamine and related steroidal alkaloid teratogens:
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