Preparation of novel ring-A fused azole derivatives of betulin and evaluation of their cytotoxicity

Abstract An efficient scheme to synthesize novel ring-A fused heterocyclic derivatives of betulin was developed. The starting reaction of this synthesis was one-pot selective bacterial oxidation of betulin to betulone used as the key compound to synthesize the substituted azoles such as C(2)–C(3)-fused 1,2,3-triazoles, oxazoles and 1,2,4-triazine, as well as C(1)–C(2)-fused isoxazoles. The semi-synthetic compounds were screened for their cytotoxic activity against human cancer cell lines A549, HCT 116, HEp-2, MS and RD TE32 with use of the photometric MTT assays. Among the tested compounds, N -acetyltriazole of betulin ( 10 ) displayed impressive cytotoxic activity with IC 50 2.3–7.5 μM against HCT 116, HEp-2, MS and RD TE32 cell lines as well as 3-methyl-4-oxido-1,2,4-triazine-derivative of betulonic acid ( 12 ) that was active against HCT 116 and HEp-2 cell lines with IC 50 1.4 and 1.5 μM, respectively. Comparative experiments showed triazole ( 10 ) to have a lower cytotoxicity to normal epithelial cells, in comparison with compound ( 12 ). In accord with the in vivo acute toxicity test, the LD 50 of triazole ( 10 ) exceeded 600 mg/kg. The ability of the most potent active triazole ( 10 ) to trigger apoptotic cell death was explored in the Annexin V-FITC test and by analyzing of caspase activity and morphological alterations in mitochondria and nuclei of HCT 116 cells.