Phytochemical Analysis of Selected Plants in the Leguminosae and Moraceae Families for Anticancer Principles

Abstract

Cancer is an increasing public health problem in the world with about 17.5 million new cases and 8.7 million cancer deaths reported in 2015 alone. The development of drug resistant cancer cells represents one of the major challenges in cancer chemotherapy. The ever growing incidences of cancer, and the rapid development of drug resistance, has made it necessary to discover novel drugs to tackle this menace. A survey of current pharmaceutical drugs revealed that 60% of cancer therapeutics are derived from natural products. Due to drug resistance, the search for new anticancer agents has continued, especially among plant metabolites. Members of the family Leguminosae and Moraceae produce a broad variety of heterocycles compounds with a wide range of biological activities, including anticancer activities, indicating that these families could be sources of anticancer agents. Chromatographic separation of the CH2Cl2/MeOH (1:1) extracts of six plants belonging to the Leguminosae (Ormocarpum kirkii S x Moore, Derris trifoliata Lour, Lonchocarpus bussei Harms, Lonchocarpus eriocalyx Harms) and Moraceae (Dorstenia kameruniana Engl and Streblus usambarensis (Engl) x CC x Berg) families resulted in the isolation of forty one compounds (1-41), of which nine are new. Thus a new rotenoid derivative, 7a-O-methyl-12a-hydroxyelliptonol (16), along with eight known compounds (a pterocarpan, five rotenoid derivatives and two isoflavones) were isolated from the roots of Derris trifoliata. Phytochemical investigation of the leaves of Lonchocarpus bussei resulted in the identification of a new isoflavone, 4'-prenyloxyvigvexin A (24) along with four known isoflavones. The stem bark of Lonchocarpus eriocalyx afforded a new pterocarpan, (6aR,11aR)-3,8-dimethoxybitucarpin B (33) along with a known pterocarpan. The roots and twigs of Dorstenia kameruniana yielded three new benzylbenzofuran derivatives, 2-(p-hydroxybenzyl)-6-hydroxybenzofuran (34), 2-(p-hydroxy-benzyl)-6-hydroxy-7-methoxybenzofuran (35) and 2-(p-hydroxy-benzyl)-6-hydroxy-4ˈ-prenylbenzofuran (36) (named dorsmerunin A, B and C, respectively), along with a known coumarin known and a chalcone. Similar investigation of the roots and stems of Streblus usambarensis yielded three new unusual naphthobenzofuran derivatives, 2,3-(5'-hydroxy-naphtalene)-6,7-chromene-5-hydroxybenzofuran (39), 2,3-(5'-methoxy-naphthalene)-6,7-chromene-5-hydroxybenzofuran (40) and 2,3-(5'-methoxy-naphthalene)-6-methoxy-5-hydroxy-7-prenylbenzofuran (41) named usambarin A, B and C, respectively. The structures of the isolated compounds were elucidated by mass spectrometry and NMR (1H NMR, 13C NMR, COSY, NOESY, HSQC and HMBC) spectroscopy. The absolute configuration of some of the chiral compounds was determined by ECD spectroscopy. The cytotoxicity of the isolated compounds was determined based on the resazurin assay using drug-sensitive and multidrug-resistant cancer cell lines. Among the tested compounds, 5,7-dihydroxy-4'-methoxy-6,8-diprenylisoflavone (2), osajin (3) and 7,7"-di-O-methylchamaejasmin (4) displayed IC50 values below 20 μM in both CCRF-CEM and CEM/ADR5000 cells, while 3',6,7-trimethoxyl-4',5'-methylenedioxyisoflavone (27) and durmillone (28) were active against leukemia CCRF-CEM cells; 4-hydroxylonchocarpin (29) and durmillone (28) against its resistant counterpart CEM/ADR5000 cells. Bergapten (37) and licoagrochalcone A (38) showed good activities (IC50 values of 7.17 μM and 5.16 μM, respectively) against CCRF-CEM leukemia cells. Usambarin B (40) had significant effects towards CEM/ADR5000 leukemia cells with IC50 value of 6.13 μM. Osajin (3) and 7,7"-di-O-methylchamaejasmin (4) had significant cytotoxic effects with IC50 values below or around 10 μM against 7 carcinoma cells and against the normal AML12 hepatocytes (4/7, 5/7 and 7/7). Durmillone (28) showed IC50 values below 10 μM against the resistant breast adenocarcinoma MDA-MB231/BCRP cells and resistant gliobastoma U87MG.ΔEGFR cells. Licoagrochalcone A (38) also showed cytotoxicity against 7 sensitive or drug-resistant solid tumor cell lines (breast carcinoma, colon carcinoma, glioblastoma) with IC50 values below 50 μM, whilst bergapten (37) showed selective activity. Usambarin B (40) and usambarin C (41) had cytotoxic effects against the 7 tested carcinoma cell lines with IC50 values below 63 μM. Cytotoxicity of some isolated compounds was also assessed against human embryonic kidney cells (HEK293). The highest activity was observed for rotenone (10) an IC50 value of 0.82 ± 0.02 μM while the rotenoloids 7a-O-methyldeguelol (12) and 7-a-O-methylelliptonol (18) showed cytotoxicity with an IC50 values of 9.4 ± 0.25 μM and 7.1 ± 0.50 μM, respectively. The isoflavones, 5,7-dihydroxy-4'-methoxy-6,8-diprenylisoflavone (2) and osajin (3) showed comparable activity (IC50 27.1 and 27.3 ± 2.0 μM, respectively) against this cell line. Further studies were conducted to determine the modes of action of osajin (3) and 7,7"-di-O-methylchamaejasmin (4). The result showed osajin (3) and 7,7"-di-O-methylchamaejasmin (4) caused cell cycle arrest in G0/G1 phase as well as apoptosis with significant increase of cells in sub-G0/G1 phase. The activity of caspases in CCRF-CEM cells showed that the two compounds did not increase the activity of caspases 3/7, 8 and 9. These compounds (3, 4) induced apoptosis in CCRF-CEM cells mediated by MMP alteration and increased ROS production. Overall, the study has demonstrated that the plants investigated here elaborate diverse range of phenolics with some having unique structural features. These plants are also good sources of cytotoxic compounds with potential use in cancer therapy.