The evolutionarily important Hedgehog (HH) signaling pathway plays a critical role in the development and progression of multiple solid tumors, such as basal cell carcinoma, medulloblastoma, rhabdomyosarcoma, and various gastrointestinal, pulmonary, and brain tumors. The proteins of the Gli (glioma-associated oncogene homologue) family are key mediators of the HH pathway. In the present study, we have focused on triterpenoid derivatives, which have been shown to induce apoptosis and inhibit HH signaling in rhabdomyosarcoma. Utilizing a U-87MG glioblastoma-derived reporter cell line, we screened a structurally diverse library of triterpenoid derivatives to identify potential antagonists of Gli-mediated transcription. We revealed two derivatives that not only selectively inhibited Gli-mediated gene transactivation but also displayed greater potency than the known Gli1 inhibitor GANT61. These compounds also demonstrated dose- and time-dependent inhibition of U-87MG tumor cell proliferation in vitro. Further mechanistic studies provided genetic evidence for the inhibition of the downstream HH pathway by these compounds, via reduced expression of Gli1 and its transcription targets. However, these compounds did not affect the ciliary localization of Smoothened (Smo). Our findings suggest that the observed inhibitory effects are likely due to a direct interaction between our compounds and Gli1.
Keywords: Hedgehog signaling pathway, transcription; betulinic acid; glioblastoma, firefly luciferase; isothermal titration calorimetry (ITC); primary cilium; triterpenes.
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