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Cytotoxic effects of the cardenolide convallatoxin and its Na,K-ATPase regulation

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Abstract

Cardenolides are cardiac glycosides, mostly obtained from natural sources. They are well known for their inhibitory action on the Na,K-ATPase, an effect that regulates cardiovascular alterations such as congestive heart failure and atrial arrhythmias. In recent years, they have also sparked new interest in their anticancer potential. In the present study, the cytotoxic effects of the natural cardenolide convallatoxin (CON) were evaluated on non-small cell lung cancer (A549 cells). It was found that CON induced cytostatic and cytotoxic effects in A549 cells, showing essentially apoptotic cell death, as detected by annexin V-propidium iodide double-staining, as well as changes in cell form. In addition, it prompted cell cycle arrest in G2/M and reduced cyclin B1 expression. This compound also increased the number of cells in subG1 in a concentration- and time-dependent manner. At a long term, the reduction of cumulative population doubling was shown along with an increase of β-galactosidase positive cells and larger nucleus, indicative of senescence. Subsequently, CON inhibited the Na,K-ATPase in A549 cells at nM concentrations. Interestingly, at the same concentrations, CON was unable to directly inhibit the Na,K-ATPase, either in pig kidney or in red blood cells. Additionally, results of docking calculations showed that CON binds with high efficiency to the Na,K-ATPase. Taken together, our data highlight the potent anticancer effects of CON in A549 cells, and their possible link with non-classical inhibition of Na,K-ATPase.

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Abbreviations

CON:

Convallatoxin

NSCLC:

Non-small cell lung cancer

PAC:

Paclitaxel

FBS:

Fetal bovine serum

TB:

Trypan blue

NII:

Nuclear irregularity index

SDS:

Sodium dodecyl sulfate

DOC:

Deoxycholate

EDTA:

Ethylenediaminetetraacetic acid

RIPA:

Radioimmunoprecipitation assay buffer

PVDF:

Polyvinylidene fluoride

DAPI:

4′,6-Diamidino-2-phenylindole

EGTA:

Ethylene glycol tetraacetic acid

FACS:

Fluorescence-activated cell sorting.

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Acknowledgements

The Brazilian authors would like to thank the funding agencies CAPES /MEC (Ministry of Education) and CNPq/MCTI (Ministry of Science, Technology and Innovation) for their research scholarships. This work was also supported by the CNPq [Grants 472544/2013-6 and 490057/2011-0], the Marie Curie Foundation—IRSES/European Community [Grant 295251], and CAPES [Grant PNPD 2257/2011].

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Authors and Affiliations

  1. Departamento de Ciências Farmacêuticas, Centro de Ciências da Saúde, Universidade Federal de Santa Catarina (UFSC), LVA, CIF, CCS, Florianópolis,, SC, 88040-900, Brazil

    Naira Fernanda Zanchett Schneider, Izabella Thais Silva, Lara Persich, Annelise de Carvalho, Lucas Marostica & Cláudia M. O. Simões

  2. Laboratório de Bioquímica Celular, Campus Centro-Oeste Dona Lindu, Universidade Federal de São João del Rei, Divinopolis, MG, Brazil

    Sayonarah C. Rocha, Ana Carolina Pacheco Ramos & Leandro A. Barbosa

  3. Laboratório de Bioinformática, Campus Centro Oeste Dona Lindu, Universidade Federal de São João Del Rei, Divinópolis, MG, Brazil

    Alex G. Taranto

  4. Departamento de Produtos Farmacêuticos, Faculdade de Farmácia, Universidade Federal de Minas Gerais, Belo Horizonte, MG, Brazil

    Rodrigo M. Pádua & Fernão C. Braga

  5. Department of Biology, Friedrich-Alexander Universität, Erlangen, Germany

    Wolfgang Kreis

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  1. Naira Fernanda Zanchett Schneider
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  2. Izabella Thais Silva
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Schneider, N.F.Z., Silva, I.T., Persich, L. et al. Cytotoxic effects of the cardenolide convallatoxin and its Na,K-ATPase regulation. Mol Cell Biochem 428, 23–39 (2017). https://doi.org/10.1007/s11010-016-2914-8

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