Betulinic acid induces apoptosis and inhibits hedgehog signalling in rhabdomyosarcoma.
BACKGROUND
Rhabdomyosarcoma (RMS) is the most common soft-tissue sarcoma in childhood with the ability to resist apoptosis by the activation of survival promoting and anti-apoptotic proteins.


METHODS
Efficacy of the apoptosis-inducing agent betulinic acid (BA) was determined in RMS cell cultures and in vivo by measuring cell viability, survival, apoptosis, hedgehog signalling activity, and neovascularisation.


RESULTS
Betulinic acid had a strong cytotoxic effect on RMS cells in a dose-dependent manner. The BA treatment caused a massive induction of apoptosis mediated by the intrinsic mitochondrial pathway, which could be inhibited by the broad-range caspase inhibitor zVAD.fmk. Exposure of hedgehog-activated RMS-13 cells to BA resulted in a strong decrease in GLI1, GLI2, PTCH1, and IGF2 expression as well as hedgehog-responsive luciferase activity. Intraperitoneal injection of 20 mg BA per kg per day significantly retarded growth of RMS-13 xenografts in association with markedly higher counts of apoptotic cells and down-regulation of GLI1 expression compared with control tumours, while leaving microvascular density, cell proliferation, and myogenic differentiation unaffected.


CONCLUSIONS
Our data show that induction of apoptosis and inhibition of hedgehog signalling are important features of the anti-tumourigenic effect of BA in RMS and advices this compound for the use in a multimodal therapy of this highly aggressive paediatric tumour.
Department of Paediatric Surgery, Dr. von Hauner Children's Hospital, Ludwig-Maximilians-University Munich, Lindwurmstrasse 4, Munich 80337, Federal Republic of Germany.
M
M
Eichenmüller
B
B
Hemmerlein
D
D
von Schweinitz
R
R
Kappler
D000818:Animals; D000972:Antineoplastic Agents, Phytogenic / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D005260:Female; D053823:Hedgehog Proteins / Q000037:antagonists & inhibitors* / Q000502:physiology; D006801:Humans; D051379:Mice; D008928:Mitochondria / Q000187:drug effects; D053978:Pentacyclic Triterpenes; D012208:Rhabdomyosarcoma / Q000188:drug therapy* / Q000378:metabolism / Q000473:pathology; D015398:Signal Transduction / Q000187:drug effects*; D014315:Triterpenes / Q000494:pharmacology*; D023041:Xenograft Model Antitumor Assays; D000094062:Betulinic Acid
D000972:Antineoplastic Agents, Phytogenic; D053823:Hedgehog Proteins; D053978:Pentacyclic Triterpenes; D014315:Triterpenes; D000094062:Betulinic Acid