Simultaneous and dose dependent melanoma cytotoxic and immune stimulatory activity of betulin

PLoS One. 2015 Mar 10;10(3):e0118802. doi: 10.1371/journal.pone.0118802. eCollection 2015.

Abstract

Conventional cytostatic cancer treatments rarely result in the complete eradication of tumor cells. Therefore, new therapeutic strategies focus on antagonizing the immunosuppressive activity of established tumors. In particular, recent studies of antigen-loaded dendritic cells (DCs) eliciting a specific antitumor immune response has raised the hopes of achieving the complete elimination of tumor tissue. Genistein, fingolimod and betulin have already been described as active compounds in different types of cancer. Herein, we applied an integrated screening approach to characterize both their cytostatic and their immune-modulating properties side-by-side. As will be described in detail, our data confirmed that all three compounds exerted proapoptotic and antiproliferative activity in different B16 melanoma cell lines to a given extent, as revealed by an MTT assay, CFSE and DAPI staining. However, while genistein and fingolimod also affected the survival of primary bone marrow (BM) derived DCs of C57BL/6 mice, betulin exhibited a lower cytotoxicity for BMDCs in comparison to the melanoma cells. Moreover, we could show for the first time, that only betulin caused a simultaneous, highly specific immune-stimulating activity, as measured by the IL-12p70 release of Toll-like receptor 4-stimulated BMDCs by ELISA, which was due to increased IL-12p35 mRNA expression. Interestingly, the activation of DCs resulted in enhanced T lymphocyte stimulation, indicated by increased IL-2 and IFN-γ production of cytotoxic T cells in spleen cell co-culture assays which led to a decreased viability of B16 cells in an antigen specific model system. This may overcome the immunosuppressive environment of a tumor and destroy tumor cells more effectively in vivo if the immune response is specific targeted against the tumor tissue by antigen-loaded dendritic cells. In summary, cytostatic agents, such as betulin, that simultaneously exhibit immune stimulatory activity may serve as lead compounds and hold great promise as a novel approach for an integrated cancer therapy.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Adjuvants, Immunologic / pharmacology*
  • Animals
  • Antineoplastic Agents / pharmacology*
  • Apoptosis / drug effects
  • Cell Proliferation / drug effects
  • Cell Survival / drug effects
  • Dose-Response Relationship, Drug
  • Drug Screening Assays, Antitumor
  • Female
  • Fingolimod Hydrochloride / pharmacology
  • Genistein / pharmacology
  • Melanoma, Experimental / drug therapy*
  • Melanoma, Experimental / immunology
  • Mice, Inbred C57BL
  • Neoplasm Transplantation
  • T-Lymphocytes / drug effects
  • T-Lymphocytes / immunology
  • Triterpenes / pharmacology*

Substances

  • Adjuvants, Immunologic
  • Antineoplastic Agents
  • Triterpenes
  • betulin
  • Genistein
  • Fingolimod Hydrochloride

Grant support

This project was made possible by a grant to HHR of Merck KGaA to the DFG Graduate School 1172 in support of the PhD thesis of KP. CD was supported by the European Social Fund, Human Resources Development Operational Programme 2007-2013, project no. POSDRU/159/1.5/S/136893. Both funding institutions had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.