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Betulinic Acid-induced Mcl-1 Expression in Human Melanoma — Mode of Action and Functional Significance

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Abstract

Background

Currently there is no information on the regulation of expression and physiological role of the anti-apoptotic protein Mcl-1 in cells of the melanocytic lineage. This study investigates the regulation and expression of Mcl-1 in human melanoma cells, which was recently found to be induced by betulinic acid, a compound with anti-melanoma and apoptosis-inducing potential.

Materials and Methods

Mcl-1 phosphorthioate anti-sense oligonucleotides were used to investigate the effect of downregulating the expression of Mcl-1. Regulation of Mcl-1 expression was analyzed with the specific PI3-kinase inhibitors LY294002 and wortmannin and the inhibitor of MAP-kinase activation, PD98059. Western blot analysis was performed with anti ERK1/2, Mcl-1, Bak, Bcl-x and Bax antibodies. Activation status of PI-3 kinase and MAP-kinase pathways was investigated using phospho-Akt and phosphorylation-state independent Akt as well as phospho-MAP kinase, phospho-MEK and phospho-GSK-3α/β antibodies.

Results

Upregulation of Mcl-1 in human melanoma cells by betulinic acid is mediated via a signal-transduction pathway that is inhibited by LY294002 and wortmannin. Betulinic acid-induced phosphorylation and activation of the Akt protein kinase was inhibited by LY294002. The inhibitor PD98059 reduced expression levels of Mcl-1 in melanoma cells and this effect was counteracted by betulinic acid. Downregulation of Mcl-1 by antisense oligodeoxynucleotides in combination with betulinic treatment led to a synergistic effect regarding growth inhibition.

Conclusions

These results suggest that in human melanoma cells Mcl-1 is (i) of functional relevance for survival and (ii) subject to dual regulation by the MAP-kinase pathway and a pathway involving protein kinase B/Akt, the latter of which is modulated in response to betulinic acid. This study provides an experimental foundation for future therapeutic strategies using anti-Mcl-1 antisense oligonucleotides in human melanoma.

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Acknowledgments

We thank R. Haslinger for technical assistance. The work in B.J.’s laboratory was supported by grants from: Austrian Science Fund (FWF), Austrian National Bank, “Kommission Onkologie”, “Kamillo Eisner Stiftung”, “Hirtl-Buss Stiftung”, “Hans and Blanca Moser Stiftung”, “Anton Dreher Stiftung”, “Hygienefonds”, “Virologiefonds”.

The work in E.S.’s Laboratory was supported by a “Fonds des Wiener Bürgermeisters” (grant no. 1690).

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Authors and Affiliations

  1. Department of Clinical Pharmacology 18–20, Section of Experimental Oncology and Molecular Pharmacology, University Hospital Vienna, 1090, Vienna, Austria

    Edgar Selzer, Christiane Thallinger, Philipp Oberkleiner, Volker Wacheck & Burkhard Jansen

  2. Department of Radiotherapy and Radiobiology, University Hospital Vienna, Vienna, Austria

    Edgar Selzer

  3. Department of Dermatology, Division of General Dermatology, University Hospital Vienna, Vienna, Austria

    Christiane Thallinger, Christoph Hoeller, Hubert Pehamberger & Burkhard Jansen

  4. Center of Excellence for Clinical and Experimental Oncology (CLEXO), University Hospital Vienna, Vienna, Austria

    Edgar Selzer & Hubert Pehamberger

  5. Ludwig Boltzmann Institute for Clinical and Experimental Oncology, University Hospital Vienna, Vienna, Austria

    Hubert Pehamberger

Authors
  1. Edgar Selzer
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  2. Christiane Thallinger
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  3. Christoph Hoeller
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  4. Philipp Oberkleiner
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  5. Volker Wacheck
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  6. Hubert Pehamberger
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  7. Burkhard Jansen
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Correspondence to Edgar Selzer.

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Selzer, E., Thallinger, C., Hoeller, C. et al. Betulinic Acid-induced Mcl-1 Expression in Human Melanoma — Mode of Action and Functional Significance. Mol Med 8, 877–884 (2002). https://doi.org/10.1007/BF03402094

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