Communications to the Editor
J ournal of Medicinal Chemistry, 1999, Vol. 42, No. 16 3017
(2) Bazzoli, A. S.; Manson, J .; Scott, W. J .; Wilson, J . G. The effects
of thalidomide and two analogues on the regenerating forelimb
of the newt. J . Embryol. Exp. Morph. 1977, 41, 125-135.
(3) Sampaio, E. P.; Sarno, E. N.; Gallily, R.; Cohn, Z. A.; Kaplan,
genesis) and thus is an ideal assay for identifying
antimetastatic agents.19
These structure-activity relationship studies on tha-
lidomide’s ability to inhibit B16BL6 metastasis revealed
that the glutarimide moiety of thalidomide is not
essential for antimetastatic activity, since a completely
hydrolyzed glutarimide group in 5c, 7, 10a , and 10b
does not render the molecule inactive. Conversely, data
in this paper suggest that the antimetastatic action of
thalidomide resides in the phthalimide group. Partial
reduction of the phthalimide moiety results in a thali-
domide analogue with enhanced activity compared to
thalidomide, presumably by increasing the in vivo t1/2
of the drug by removing one of the known sites of in
vivo hydrolysis. Also, aromatic substitution (hydroxy-
lation) on phthalimide gives rise to inactive derivatives.
Whether this is due to changing the spatial or electronic
requirements for binding to the cellular target or
whether this is due to increased secretion following
sulfate or glucuronidation is not known. This will be
assessed in future studies by synthesis of ethers of 5a
and 5b. The data indicate that hepatic hydroxylation
of thalidomide is not required for inhibition of metasta-
sis, as was at one time proposed.18
Previous reports on enantioselectivity of thalidomide
and analogues such as N-(2′,6′-dioxopiperiden-3′-yl)-
phthalimidine are questionable, since preparations of
unstable enantiomers, e.g., nonmethylated chiral car-
bon, racemized in vitro and in vivo.14 Heger et al.
suggested that preparations of derivatives locked in
each optical configuration would be required to confirm
selective effects of optical isomers of thalidomide ana-
logues.16 We report here the preparation and purifica-
tion of 10a and 10b, which are the stabilized enanti-
omers of 5c. As the activity of racemic 5c is indistin-
guishable from that of racemic 7, the presence of the
methyl group does not affect the antitumor activity,
enabling us to ignore the alkyl group’s contributions to
lipophilicity, volume, and access to and binding to the
cellular target. The clear difference between the activi-
ties of 10a and 10b indicates that the cellular target of
this series of molecules is enantioselective and prefers
the S(-)-enantiomer.
G. Thalidomide selectively inhibits tumor necrosis factor
R
production by stimulated human monocytes. J . Exp. Med. 1991,
173, 699-703.
(4) D’Amato, R. J .; Loughnan, M. S.; Flynn, E.; Folkman, J .
Thalidomide is an inhibitor of angiogenesis. Proc. Natl. Acad.
Sci. U.S.A. 1994, 91, 4082-4085.
(5) Minchinton, A. I.; Fryer, K. H.; Wendt, K. R.; Clow, K. A.; Hayes,
M. M. The effect of thalidomide on experimental tumors and
metastases. Anti-Cancer Drugs 1996, 7, 339-343.
(6) Helm, F.-Ch.; Frankus, E.; Graudums, I.; Flohe, L. Comparative
teratological investigation of compounds structurally and phar-
macologically related to thalidomide. Drugs Res. 1981, 31, 941-
949.
(7) Miyachi, H.; Azuma, A.; Ogasawara, A.; Uchimura, E.; Wa-
tanabe, N.; Kobayashi, Y.; Kato, F.; Kato, M.; Hashimoto, Y.
Novel biological response modifiers: Phthalimides with tumor
necrosis factor-R production-regulating activity. J . Med. Chem.
1997, 40 (18), 2858-2865.
(8) Corral, L. G.; Kaplan, G.; Stirling, D.; Wu, M.; Chen, Y.; Moriera,
A. L.; Muller, G. W. Selection of novel analogs of thalidomide
with enhanced tumor necrosis factor-R inhibitory activity. Mol.
Med. 1996, 2 (4), 506-515.
(9) Shannon, E. J .; Sandoval, F.; Krahenbuhl, J . L. Hydrolysis of
thalidomide abrogates its ability to enhance mononuclear cell
synthesis of IL-2 as well as its ability to suppress the synthesis
of TNF-R. Immunopharmacology 1996, 990.
(10) Nicolson, G. L.; Brunson, K. W.; Fidler, I. J . Specificity of arrest,
survival, and growth of selected metastatic variant cell lines.
Cancer Res. 1978, 38, 4105-4111.
(11) Hart, I. R. The selection and characterization of an invasive
variant of the B16 melanoma. Am. J . Pathol. 1979, 97, 587-
600.
(12) Braun, A. G.; Weinreb, S. L. Teratogen metabolism: spontaneous
decay products of thalidomide and thalidomide analogues are
not bioactivated by liver microsomes. Teratog. Carcinog. Mu-
tagen. 1985, 5, 149-158.
(13) Bauer, K. S.; Dixon, S. C.; Figg, W. D. Inhibition of angiogenesis
by thalidomide requires metabolic activation, which is species-
dependent. Biochem. Pharmacol. 1998, 55 (11), 1827-1834.
(14) Schmahl, H.-J .; Heger, W.; Nau, H. The enantiomers of the
teratogenic analog EM12. Toxicol. Lett. 1989, 45, 23-33.
(15) Kenyon, B. M.; Browne, F.; D’Amato, R. J . Effects of thalidomide
and related metabolites in a mouse cornea model of neovascu-
larization. Exp. Eye Res. 1997, 64 (6), 971-978.
(16) Heger, W.; Schmahl, H. J .; Klug, S.; Felies, A.; Nau, H.; Merker,
H. J .; Neubert, D. Embryotoxic effects of thalidomide derivatives
in the non-human primate callithrix jacchus. IV. Teratogenicity
of micrograms/kg doses of the EM12 enantiomers. Teratog.
Carcinog. Mutagen. 1994, 14, 115-122.
(17) Muller, G. W.; Corral, L. G.; Shire, M. G.; Wang, H.; Moreira,
A.; Kaplan, G.; Stirling, D. I. Structural modifications of thali-
domide produce analogs with enhanced tumor necrosis factor
inhibitory activity. J . Med. Chem. 1996, 39, 3238-3240.
(18) Gordon, G. B.; Spielberg, S. P.; Blake, D. A.; Balassubramanian,
V. Thalidomide teratogenesis: evidence for a toxic arene oxide
metabolite. Proc. Natl. Acad. Sci. U.S.A. 1981, 78, 2545-2548.
(19) Post, G.; Doll, J .; Hart, I. R.; Fidler, I. J . In vitro selection of
murine B16 melanoma varients with enhanced tissue-invasive
properties. Cancer Res. 1980, 40, 1636-44.
Collectively, this is the first study on the structural
requirements for antimetastatic activity of thalidomide
and thalidomide analogues. Further, this report clearly
shows that a specific property of these analogues is
enantioselective. This property, as well as the increased
stability of the PGA series of analogues compared to the
parent, thalidomide, may also be helpful in identifying
the cellular target of these molecules, and so defining
the mechanism of the anticancer action of thalidomide.
(20) Kenyon, B. M.; Browne, F.; D’Amato, R. J . Effects of thalidomide
and related metabolites in a mouse corneal model of neovascu-
larization. Exp. Eye Res. 1997, 64, 971-978.
(21) Shimazawa, R.; Miyachi, H.; Takayama, H.; Kuroda, K.; Kato,
F.; Kato, M.; Hashimoto, Y. Antiangiogenic activity of tumor
necrosis factor-a production regulators derived from thalidomide.
Biol. Pharm. Bull. 1999, 22, 224-226.
Su p p or tin g In for m a tion Ava ila ble: Experimental de-
tails. This material is available free of charge via the Internet
at http://pubs.acs.org.
(22) Stirling, D.; Sherman, M.; Strauss, S. Thlidomide. A surprising
recovery. J . Am. Pharm. Assoc. 1997, 37, 306-13.
Refer en ces
(1) Cahen, R. L. Evaluation of the teratogenicity of drugs. Clin.
Pharmacol. Ther. 1964, 5, 480-514.
J M990083Y