Antiproliferative Activity of Derivatives of Ouabain, Digoxin and Proscillaridin A in Human MCF-7 and MDA-MB-231 Breast Cancer Cells

Abstract

Three derivatives of ouabain, digoxin and proscillaridin A containing the carboxylic group instead of the lactone moiety were synthesized and examined for cytotoxicity in human breast cancer cells. Evaluation of the cytotoxicity of these compounds employing an MTT assay and inhibition of [ 3 H]thymidine incorporation into DNA in both MCF-7 and MDA-MB-231 breast cancer cells demonstrated that compound 3, the most active of the series, proved to be only slightly less potent than proscillaridin A. We evaluated the effects of these compounds 1-3 on change in intracellular Ca 2؉ , appearance of apoptosis, inhibition of DNA topoisomerase I and II, and the activity of caspase-3 in breast cancer cells. These studies indicate that the increase in potency for 3 may be related, in part, to an activation of caspase-3, increasing free calcium concentration and topoisomerase II inhibition. All these data emphasize the potential usefulness of these derivatives of cardiac glycosides as anticancer agents.Key words cardiac glycoside; cytotoxicity; breast cancer cell; DNA topoisomerase Biol. Pharm. Bull. 31(6) 1131-1140 (2008) © 2008 Pharmaceutical Society of Japan * To whom correspondence should be addressed. e-mail: kwin@amb.edu.pl mine, penicillin and streptomycin were obtained from Quality Biologicals (U.S.A.). [ 3 H]Thymidine (6.7 Ci/mmol) was purchased from NEN (U.S.A.), and Scintillation Coctail "Ultima Gold XR" from Packard (U.S.A.). Nitrocellulose membrane (0.2 mm), sodium dodecylsulfate (SDS), polyacrylamide and molecular weight standards were received from Bio-Rad Laboratories (U.S.A.). Topoisomerase I and II, supercoiled pHOT1 DNA, supercoiled pRYG DNA, etoposide, camptothecin were purchased from TopoGEN (U.S.A.).Chemistry The structures of all the compounds were confirmed by ). Melting points were determined on Büchi 535 melting-point (Germany) apparatus and were uncorrected. Elemental analysis of C, H, was performed on a Perkin Elmer 240 analyser (U.S.A.) and satisfactory results within Ϯ0.4% of calculated values were obtained. Chemical shifts are expressed in d value (ppm). Multiplicity of resonance peaks are indicated as singlet (s), doublet (d), triplet (t), quarter (q), and multiplet (m).(Z)-4-Hydroxy-3-((1R,3S,5S,8R,9S,10R,11R,13R,14S, 17R)-1,5,11,14-tetrahydroxy-10-(hydroxymethyl)-13-methyl-3-((2R,3R,4R,5R,6S)-3,4,5-trihydroxy-6-methyltetrahydro-2H-pyran-2-yloxy)hexadecahydro-1H-cyclopenta-[a]phenanthren-17-yl)but-2-enoic Acid (Compound 1) Ouabain (0.57 g, 0.78 mmol) was suspended in a mixture of 2 ml of THF and 2 ml of 1 M LiOH. The reaction was allowed to stir at room temperature until no starting ouabain could be detected by TLC (CHCl 3 : MeOH 5 : 1). The basic mixture was acidified to pH 4 with 2 M HCl and diluted with acetone. The precipated acid was filtred and dried in vacuo at room temperature to afford 0. 17.11, 17.62, 22.96, 26.07, 32.30, 33.20, 34.91, 35.37, 38.94, 46.58, 47.37, 48.28, 48.76, 49.45, 62.57, 63.61, 66.70, 68.17, 69.22, 70.56, 70.68, 72.18, 73.54, 83.30, 97.08, 115.96, 172.3...

Docosahexaenoic acid intake decreases proliferation, increases apoptosis and decreases the invasive potential of the human breast carcinoma cell line MDA-MB-231

“…However, the dramatic fall in this activity over longer-term suggests that there could be two populations within the cell line, only one of which is sensitive to the caspase-3 pathway. As previously described, MDA-MB-231 cells can activate the caspase-3 pathway via polyunsaturated fatty acids (19). Thus, early activation of this enzymatic activity, resulting in the rapid death of this population, would be compatible with the fall in cell number observed in Fig.…”

The antiproliferative effects of ouabain and everolimus on adrenocortical tumor cells

“…Other investigators found that ouabain and digoxin selectively induced apoptosis in a human acute T-cell lymphoblastic leukemia cell line [2]. In the estrogen dependent/independent breast cancer cell lines, CS similarly exerted antiproliferative effects, suggesting their potential use as anticancer agents [7].…”

Targeting the Na⁺/K⁺-ATPase α1 Subunit of Hepatoma HepG2 Cell Line to Induce Apoptosis and Cell Cycle Arresting

“…Ouabain could induce HepG2 cell apoptosis and generate S phase arrest, and siRNA could enhance the anti-cancer effect of ouabain that induced HepG2 cells apoptosis via an intracellular Ca 2؉ ؉ and ROS increase-mediated, and generated cell cycle S phase arresting by decreasing the CyclinA1/cyclin-dependent kinase 2 (CDK2)/proliferating cell nuclear antigen (PCNA) complex product and increasing the expression of cyclin-dependent kinase inhibitor 1A (P21 tive oxygen species (ROS) by mitochondria. 12) We believe that ouabain can induce DNA double strand breaks by increasing the intracellular free Ca 2ϩ and ROS concentrations. In China, HCC ranks 3rd in terms of the mortality rate of malignant tumors.…”

“…Previous research has shown the Na ϩ /K ϩ -ATPase a1 subunit was highly expressed in malignant cells, including human prostate cancer PC-3, DU-145 cells, human non-small cell lung cancer (NSCLC) A549 cells, 10) glioblastomas Hs683, U373-MG, U87-MG, T98G cells, 11) and breast cancer MCF-7 cells. 12) Mijatovic found that cell invasion and proliferation were significantly reduced after knockdown of the Na ϩ /K ϩ -ATPase a1 subunit expression in NSCLC (A549) cells. 13) These findings indicated that the Na , which provides the driving force for the transport of other solutes, notably amino acids, sugars, and phosphate.…”