Interference of ursolic acid treatment with glioma growth: An in vitro and in vivo study

Glioblastoma multiforme is the most devastating tumor in the brain. Ursolic acid (UA) is found in a variety of plants, and exhibits several pharmacological activities. In this study, we investigated the effects of UA in vitro, clarifying the mechanisms that mediate its toxicity and the long-lasting actions of UA in C6 glioma cells. We also evaluated the antitumor activity of UA in an in vivo orthotopic glioma model. Cell numbers were assessed using the Trypan blue exclusion test, and the cell cycle was characterized by flow cytometry using propidium iodide staining. Apoptosis was analyzed using an Annexin V kit and by examining caspase-3. Akt immunocontent was verified by Western blot and the long-lasting actions of UA were measured by cumulative population doubling (CPD). In vivo experiments were performed in rats to measure the effects on tumor size, malignant features and toxicological parameters. In vitro results showed that UA decreased glioma cell numbers, increased the sub-G1 fraction and induced apoptotic death, accompanied by increased active caspase-3 protein levels. Akt phosphorylation/activation in cells was also diminished by UA. With regard to CPD, cell proliferation was almost completely restored upon single UA treatments, but when the UA was added again, the majority of cells died, demonstrating the importance of re-treatment cycles with chemotherapeutic agents for abolishing tumor growth. In vivo, ursolic acid slightly reduced glioma tumor size but did not decrease malignant features. Ursolic acid may be a potential candidate as an adjuvant for glioblastoma therapy.