Hendrik van den Berg

Reduced expression of endothelial and inducible nitric oxide synthase in a human breast cancer cell line which has acquired estrogen independence.

We have recently reported the presence of inducible nitric oxide synthase (iNOS) in the human breast cancer cell line ZR-75-1. The purpose of the present study was to examine differences in expression of endothelial (eNOS) and inducible nitric oxide synthase in normal human mammary epithelial cells (HMEC) compared with two variants of the ZR-75-1 cell line. One variant has acquired estrogen independence, the other has acquired resistance to tamoxifen. Immunohistochemical investigations demonstrated that 100% of HMEC cells staining positive for both eNOS and iNOS. ZR-75-1 cells showed 100% staining for eNOS and 52% positive staining for iNOS. There was no difference in staining between the parent cell line and cells which had acquired resistance to tamoxifen (ZR-75-9a1). However, in the breast cancer cell line which had acquired estrogen independence (ZR-PR-LT), less than 5% of cells exhibited positive staining for eNOS and staining for iNOS was undetectable. L-Arginine increased NO production in both ZR-75-9a1 and ZR-PR-LT cells. Progesterone was able to down regulate NO production in both ZR-75-1 and ZR-75-9a1 cells and this effect was reversible by RU486. These results support the suggestion that loss of NOS expression may be associated with the progression of breast cancers.

Benzothiazole bipyridine complexes of ruthenium(II) with cytotoxic activity

A series of benzothiazole-substituted trisbipyridine ruthenium(II) analogues {[Ru(bpy)2(4,5′-bbtb)]2+, [Ru(bpy)2(5,5′-bbtb)]2+ and [Ru(bpy)2(5-mbtb)]2+ [bpy is 2,2′-bipyridine, bbtb is bis(benzothiazol-2-yl)-2,2′-bipyridine, 5-mbtb is 5-(benzothiazol-2-yl),5′-methyl-2,2′-bipyridine]} have been prepared and compared with the complex [Ru(bpy)2(4,4′-bbtb)]2+ reported previously. From the UV–vis spectral studies, substitution at the 5-position of the bpy causes the ligand-centred transitions to occur at considerably lower energy than for those with the functionality at the 4-position, while at the same time causing the emission to be effectively quenched. However, substitution at the 4-position causes the metal-to-ligand charge transfer to occur at lower energies. Fluorescent intercalator displacement studies indicate that the doubly substituted complexes displace ethidium bromide from a range of oligonucleotides, with the greater preference shown for bulge and hairpin sequences by the Λ enantiomer. Since the complexes only show small variation in the UV–vis spectra on the introduction of calf thymus DNA and a small increase in fluorescence they do not appear to be intercalators, but appear to associate within one of the grooves. All of the reported bisbenzothiazole complexes show reasonable cytotoxicity against a range of human cancer cell lines.

Synthesis and in Vitro and in Vivo Evaluation of a Series of Dihydroisocoumarin Derivatives Conjugated with Fatty Acids, Alcohols, and Amines as Potential Anticancer Agents

In this paper, we report the synthesis and biological activity of a series of dihydroisocoumarin analogues conjugated with fatty acids, alcohols, or amines, of varying hydrocarbon chain length and degree of unsaturation, to the dihydroisocoumarins, kigelin and mellein, at the C-7 and C-8 positions on the core dihydroisocoumarin structure. These compounds were evaluated for their antiproliferative activity against human breast cancer (MCF-7 and MDA-MB-468) and melanoma cells (SK-MEL-28 and Malme-3M) using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay. Two compounds conjugated with gamma-linolenyl alcohol (18:3 n-6) demonstrated potent antiproliferative activity in vitro with one of these 4-hydroxy-3-oxo-1,3-dihydro-isobenzofuran-5-carboxylic acid octadeca-6,9,12-trienyl ester, demonstrating significant antitumor activity in vivo in a number of human tumor xenograft models.

A new approach to combretastatin D2

A concise and convergent route to combretastatin D2 is described together with some preliminary biological data.

Growth Inhibitory Activity of Extracted Material and Isolated Compounds from the Fruits of Kigelia pinnata

A study of the components of the fruits of Kigelia pinnata was undertaken to identify compounds with potential growth inhibitory activity against human melanoma cells, since extracts from the fruits of this plant have been described in traditional medicine to have application in the treatment of skin cancer and other skin ailments. A bioactivity-guided fractionation process yielded a number of crude fractions, which demonstrated cytotoxicity in vitro against human melanoma cells. Compounds isolated and identified included the isocoumarins, demethylkigelin (1) and kigelin (2), fatty acids, oleic (3) and heneicosanoic acids (4), the furonaphthoquinone, 2-(1-hydroxyethyl)-naphtho[2,3-b]furan-4,9-dione (5), and ferulic acid (6). A number of structurally related synthetic compounds were also tested using the MTT assay. The most potent series of these compounds, the furonaphthoquinones, also demonstrated a cytotoxic effect in two human breast cancer cell lines tested.

Reduced levels of cathepsin D associated with tamoxifen resistance and estrogen independence in the ZR-75-1 human breast cancer cell line

The expression and secretion of cathepsin D by ZR-75-1 human breast cancer cells, and tamoxifen-resistant (ZR-75-9a1) and estrogen-independent (ZR-PR-LT) variants was examined by electrophoresis of labeled proteins and Western blotting. Secreted proteins of 160 kDa, 52 kDa and 34 kDa were identified, and in ZR-75-1 cells, they were shown to be estrogen-inducible. Treatment of ZR-75-9a1 cells with 17 beta-estradiol (E2) and the progestin ORG 2058 increased secretion of the 52 kDa protein; ZR-PR-LT cells were unaffected. Western blotting showed that each cell line expressed high levels of the 52 kDa and 34 kDa forms of cathepsin D but that relatively little was being secreted. Each cell line secreted 52 kDa procathepsin D, but 34 kDa mature-cathepsin D was not detected as a secreted protein. ZR-75-1 cells expressed and secreted the highest levels of cathepsin D while ZR-75-9a1 cells expressed and secreted the least.

Abstract A175: Isolation and identification of compounds from the fruits of Kigelia pinnata with anti‐proliferative activity towards human cancer cells

Kigelia pinnata DC. (Bignoniaceae also referred to as K. Africana or sausage tree), is a tree which is found widely in Africa and the tropics in general. Traditionally hot aqueous extracts of fruit pods from Kigelia pinnata have been claimed to have widespread ethnomedicinal importance. A preliminary study of the components of the aqueous extracts from the fruits of Kigelia pinnata was undertaken to identify fractions with potential anti‐cancer activity. A bioactivity guided fractionation process of extracts from fruits of the tree was carried out yielding a number of crude fractions, which demonstrated cytotoxicity in vitro against human melanoma cells. A number of compounds were isolated and identified within purified fractions of several fruit extracts including the dihydroisocoumarins, demethylkigelin and kigelin, a number of fatty acids, and a furonaphthoquinone, as compounds at least in part responsible for the anti‐proliferative activity observed in extracted material from the fruits. We tested a number of structurally related dihydroisocoumarins for their anti‐proliferative activity in order to conduct a preliminary structure‐activity relationship (SAR) study. The preliminary medicinal chemistry programme clearly identified structures with enhanced anti‐proliferative activity relative to the isolated compounds. In conclusion, using a bioassay fractionation approach, we have isolated and characterized several classes of compounds that exhibit biological activity against human cancer cell lines. Structurally‐related derivatives of these compounds have been synthesized to develop knowledge of the structure‐activity relationship for these classes of compounds. Our data to date suggest that this class of molecule has structural features that appear to be of importance in determining its activity against human cancer cells. Citation Information: Mol Cancer Ther 2009;8(12 Suppl):A175.