Novel 11-Substituted Ellipticines as Potent Anticancer Agents with Divergent Activity against Cancer Cells.

Ellipticines have well documented anticancer activity, in particular with substitution at the 1-, 2-, 6- and 9-positions. However, due to limitations in synthesis and coherent screening methodology the full SAR profile of this anticancer class has not yet been achieved. In order to address this shortfall, we have set out to explore the anticancer activity of this potent natural product by substitution. We currently describe the synthesis of novel 11-substituted ellipticines with two specific derivatives showing potency and diverging cellular growth effects.

Oxidation of sitosterol and transport of its 7-oxygenated products from different tissues in humans and ApoE knockout mice.

The most common phytosterols in the human diet are sitosterol and campesterol, which originate exclusively from plant derived food. These phytosterols are taken up by NPC1L1 transport from the intestine into the enterocytes together with cholesterol and other xenosterols. Phytosterols are selectively pumped back from the enterocytes into the intestinal lumen and on the liver site from hepatocytes into bile by heterodimeric ABCG5/G8 transporters. Like cholesterol, both phytosterols are prone to ring and side chain oxidation. It could be shown that oxyphytosterols, found in atherosclerotic tissue, are most likely of in situ oxidation (Schött et al.; Biochem. Biophys. Res. Commun. 2014 Apr 11;446(3):805-10). However, up to now, the entire mechanism of phytosterol oxidation is not clearly understood. Here, we provide further information about the oxidation of sitosterol and the transport of its oxidation products out of tissue. Our survey includes data of 104 severe aortic stenosis patients that underwent an elective aortic valve cusp replacement. We studied their phytosterol concentrations, as well as absolute and substrate corrected oxyphytosterol levels in plasma and valve cusp tissue. In addition, we also examined phytosterol and oxyphytosterol concentrations in plasma and tissues (from brain and liver) of 10 male ApoE knockout mice. The ratio of 7-oxygenated-sitosterol-to-sitosterol exceeds the ratio for 7-oxygenated-campesterol-to-campesterol in plasma and tissue of both humans and mice. This finding indicates that sitosterol is oxidized to a higher amount than campesterol and that a selective oxidative mechanism might exist which can differentiate between certain phytosterols. Secondly, the concentrations of oxyphytosterols found in plasma and tissue support the idea that oxysitosterols are preferably transported out of individual tissues. Selective oxidation of sitosterol and preferred transport of sitosterol oxidation products out of tissue seem to be a metabolic pathway of forced sitosterol clearance from tissue compartments.

Vascular effects of oxysterols and oxyphytosterols in apoE -/- mice.

OBJECTIVES: The aim of our study was to investigate vascular effects of oxysterols and oxyphytosterols on reactive oxygen species (ROS), endothelial progenitor cells, endothelial function and atherogenesis. METHODS: Male apoE-/-mice were treated with cholesterol, sitosterol, 7-ß-OH-cholesterol, 7-ß-OH-sitosterol, or cyclodextrin by daily intraperitoneal application. The respective concentrations in the plasma and in the arterial wall were determined by gas chromatography-flame ionization or mass spectrometry. ROS production was assessed by electron-spin resonance spectroscopy in the aorta, endothelial function of aortic rings and atherosclerosis in the aortic sinus was quantitated after 4 weeks. RESULTS: Compared to vehicle, there was no difference in plasma cholesterol levels and arterial wall concentrations after i.p. application of cholesterol. 7-ß-OH-cholesterol concentrations were increased in the plasma (33.7±31.5 vs. 574.57.2±244.92 ng/ml) but not in the arterial wall (60.1±60.1 vs. 59.3±18.2 ng/mg). Sitosterol (3.39±0.96 vs. 8.16±4.11 mg/dL; 0.08±0.04 vs. 0.16±0.07 µg/mg, respectively) and 7-ß-OH-sitosterol concentrations (405.1±151.8 vs. 7497±3223 ng/ml; 0.24±0.13 vs. 16.82±11.58 ng/mg, respectively) increased in the plasma and in the aorta. The i.p-application of the non-oxidized cholesterol or sitosterol did not induce an increase of plasma oxysterols or oxyphytosterols concentrations. Oxidative stress in the aorta was increased in 7-ß-OH-sitosterol treated mice, but not in mice treated with cholesterol, sitosterol, or 7-ß-OH-cholesterol. Moreover, cholesterol, sitosterol, 7-ß-OH-cholesterol, and 7-ß-OH-sitosterol did not affect endothelial-dependent vasodilation, or early atherosclerosis. CONCLUSION: Increased oxyphytosterol concentrations in plasma and arterial wall were associated with increased ROS production in aortic tissue, but did not affect endothelial progenitor cells, endothelial function, or early atherosclerosis.

First total synthesis of the marine natural products clavulolactones II and III.

The first total synthesis of the marine prostanoids clavulolactones II and III is presented from an easily accessible chiral, non-racemic cyclopentenone intermediate. Key steps involve selective TBDMS deprotection, selective reduction of the ß-side chain and aldol condensation. Clavulolactones II and III were successfully prepared from (S)-4-((tert-butyldimethylsilyl)oxy) cyclopent-2-en-1-one over nine steps, in overall yields of 21 and 7% respectively.

7ß-Hydroxysitosterol crosses the blood-brain barrier more favored than its substrate sitosterol in ApoE-/- mice.

In this study, we compare the distribution of intraperitoneally injected sitosterol, 7ß-hydroxysitosterol or vehicle only (control) for 28days in male ApoE-/- mice. Furthermore we examine its impact on surrogate markers of cholesterol biosynthesis and sterol absorption rate in plasma, brain and liver tissues from these animals. Injection of sitosterol revealed a 32.1% (P=0.013) lower plasma total cholesterol compared with control. Cholesterol corrected plasma and absolute brain and liver levels of sitosterol are 4.1-, 1.7-, and 7.2-fold (P<0.001 for all) higher, respectively. This is in accordance with a reduced plasma campesterol to cholesterol ratio (-16.2%; P=0.018) together with a 24.1% (P=0.047) lower concentration of hepatic lathosterol. After injection of 7ß-hydroxysitosterol the concentrations of 7ß-hydroxysitosterol in plasma, brain and liver are 21.0-, 65.8- and 42.7-fold (P<0.001 for all) higher, respectively, compared with control. Injection of 7ß-hydroxysitosterol revealed significantly lower plasma cholesterol corrected cholestanol and campesterol (-44.2%; P=0.001 and -24.5; P=0.004) as well as lower absolute liver cholestanol levels (-31.9%; P<0.001) compared with control. Intraperitoneally injected sitosterol and 7ß-hydroxysitosterol differently influence cholesterol metabolism in plasma and liver. We conclude that the polar 7ß-hydroxysitosterol compound can pass the blood brain barrier with higher efficacy than its substrate, sitosterol. Though present in higher amounts in the brain, both, sitosterol and 7ß-hydroxysitosterol do not influence cholesterol metabolism in the brain as proven by our surrogate markers.

Synthesis and biological evaluation of novel isoellipticine derivatives and salts.

Synthesis of novel 7-substituted isoellipticines and isoellipticinium salts is described, with optimisation of routes, representing a new class of anti-cancer agent. Initial assessment of biological activity using a topoisomerase II decatenation assay and NCI screening highlighted strong anti-cancer activity, further developed in a panel of isoellipticinium salts. Interestingly, low correlation between results of the topoisomerase II decatenation assay and NCI screen throughout the panel suggest that topo II is not the most important biological target with respect to anti-cancer activity in this new class of compounds. Results also suggest that solubility is not the limiting factor in activity of the isoellipticinium salts. Overall, 20 novel ellipticine analogues were prepared and full anti-cancer profiling was completed for 13 isoellipticine derivatives and salts. Two compounds display significant specificity towards CNS cancer cell lines and are lead compounds for future development.