TUMOR INHIBITORS .44. ISOLATION AND CHARACTERIZATION OF HELLEBRIGENIN 3-ACETATE AND HELLEBRIGENIN 3,5-DIACETATE, BUFADIENOLIDE TUMOR INHIBITORS FROM BERSAMA ABYSSINICA

An alcoholic extract of the stem bark of Bersama abyssinica Fresen, was found to show significant inhibitory activity against human carcinoma of the nasopharynx in cell culture (KB). Systematic fractionation led to the isolation and characterization of hellebrigenin 3-acetate (1) and hellebrigenin 3,5-diacetate (2) as the major cytotoxic principles. Hellebrigenin 3-acetate was characterized by comparison with a sample prepared from hellebrigenin. Preliminary characterization of the 3,5-diacetate on the basis of spectral evidence indicated the presence of the bufadienolide skeleton and a second (tertiary) acetate group. The compound was shown to be a hellebrigenin diacetate by synthesis from hellebrigenin by acetylation with isopropenyl acetate and p-toluenesulfonic acid. The 3,5-diacetate structure was indicated by comparative studies of base-catalyzed ester solvolysis of 1 and 2, and confirmed by conversion of 2 to the methyl isohellebrigeninate derivative (6). A new and efficient enzymatic hydrolysis of hellebrin to hellebrigenin was developed, and a minor hydrolytic transformation product was assigned the scilliglaucosidin-3-one (4) structure. Hellebrigenin 3-acetate was found to show significant inhibitory activity against the Walker intramuscular carcinosarcoma 256 in rats, and is thus the first cardiotonic steroid found to show significant activity against an in vivo tumor system. Speculative rationalization of this observation is discussed. © 1969, American Chemical Society. All rights reserved.