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J. Nat. Prod. 1999, 62, 1346-1348
1′,2′,3′,4′-Tetradehydrotubulosine, a Cytotoxic Alkaloid from Pogonopus speciosus Aiko Ito,† You-Hui Lee,† Hee-Byung Chai,† Mahabir P. Gupta,‡ Norman R. Farnsworth,† Geoffrey A. Cordell,† John M. Pezzuto,† and A. Douglas Kinghorn*,† Program for Collaborative Research in the Pharmaceutical Sciences and Department of Medicinal Chemistry and Pharmacognosy, College of Pharmacy, University of Illinois at Chicago, Chicago, Illinois 60612, and CIFLORPAN, Universidad de Panama´ , Panama, Republic of Panama Received May 28, 1999
Bioassay-guided phytochemical investigation of the stems of Pogonopus speciosus, using human oral epidermoid carcinoma (KB) cells as a monitor, led to the isolation of a novel alkaloid, 1′,2′,3′,4′tetradehydrotubulosine (1), along with tubulosine (2) and psychotrine (3) as bioactive constituents. The structure of the novel compound was elucidated through 1D- and 2D-NMR spectroscopic methods. Alkaloids 1 and 3 showed weak cytotoxic activity against a panel of human cancer cell lines, with the potency of these compounds being markedly less than that of tubulosine (2). Pogonopus speciosus (Jacq.) K. Schum. (Rubiaceae), found in moist tropical rain forests throughout Central and South America, is a tree about 6 m high with thin obvate acuminate leaves and cymose flowers.1 Pogonopus is a small genus comprised of only three species,2 and P. speciosus is the sole species found in Panama.3 In previous phytochemical work on P. speciosus, an extract of the sap extract afforded the known alkaloids tubulosine (2) and psychotrine (3). The structure of 2 was confirmed by singlecrystal X-ray crystallography, and the cytotoxic activities of 2 and 3 were evaluated against a small panel of human tumor cell lines, with tubulosine (2) being broadly cytotoxic.4 A series of emetine derivatives including tubulosine was examined in the in vivo L1210 and P388 leukemia test systems; tubulosine (2) showed good activity, exhibiting 30% and 80% ILS (increase in life span) values, respectively, at a dose of 2 mg/kg in each case.5,6 Compounds 2 and 3 have been studied for various other biological activities, such as inhibition of protein biosynthesis,7-13 and have amebicidal,14,15 antimalarial,16 and HIV reversetranscriptase inhibitory activities.17 As a part of our ongoing program for the discovery of the anticancer agents from plants, supported by a National Cooperative Natural Products Drug Discovery Group grant, an alkaloidal fraction of the stems of P. speciosus was found to exhibit significant cytotoxic activity against a panel of human cancer cell lines. Bioassay-guided phytochemical investigation of this alkaloid fraction, using cytotoxicity with human oral epidermoid carcinoma (KB) cells in culture as a monitor, led to the isolation of the novel alkaloid 1, along with tubulosine (2) and psychotrine (3), as active constituents. Alkaloids 2 and 3 were identified on the basis of physical and spectral data comparison with literature values.4,18,19 The structure of compound 1 was elucidated using 1D- and 2D-NMR spectroscopic methods, and the three pure compounds were evaluated against a human cancer cell line panel. The HREIMS of alkaloid 1 showed a molecular ion peak at m/z 471.2503, indicating an elemental molecular formula of C29H33N3O3. The EIMS showed a molecular ion peak at m/z 471, along with major fragment ions at m/z 287, 273, * To whom correspondence should be addressed. Tel.: (312) 996-0914. Fax: (312) 996-7107. E-mail:
[email protected]. † University of Illinois at Chicago. ‡ Universidad de Panama ´.
10.1021/np990255u CCC: $18.00
272, 271, 258, 246, 244, 205, 198, and 191. The 1H NMR spectrum of compound 1 was similar to that of 2, except for the presence of signals consistent with the unsaturation of H-1′ through H-4′. In the 13C NMR spectrum of 1, when compared with that of 2, a quaternary carbon at δC 149.2 (C-1′) and double bond signals at δC 137.1 and δC 114.2 (C-3′ and C-4′) were observed, instead of a methine signal and two methylene signals of C-1′, C-3′, and C-4′, respectively. In the 1H NMR spectrum of 1, signals at δΗ 8.20 (1H, d, J ) 5.4 Hz) and δΗ 7.87 (1H, d, J ) 5.4 Hz) were assigned to the aromatic protons at C-3′ and C-4′, respectively. The structure of the side chain of 1 was confirmed by HMBC NMR spectral observations which showed crosspeaks between H-12 and C-1′; H-3′ and C-1′; H-4′ and C-4a′;
© 1999 American Chemical Society and American Society of Pharmacognosy Published on Web 08/27/1999
Notes
Journal of Natural Products, 1999, Vol. 62, No. 9 1347
Table 1. Cytotoxic Activity of Alkaloids 1-3a,b compound
BC1
Lu1
Col2
KB
KB-V+
KB-V-
LNCaP
SW626
SKNSH
M109
1 2 3
3.9 0.1 3.4
3.6
