Diterpenoid honatisine overcomes temozolomide resistance in glioblastoma by inducing mitonuclear protein imbalance through disruption of TFAM-mediated mtDNA transcription.

BACKGROUND: Glioblastoma (GBM) represents as the most formidable intracranial malignancy. The systematic exploration of natural compounds for their potential applications in GBM therapy has emerged as a pivotal and fruitful avenue of research. PURPOSE: In the present study, a panel of 96 diterpenoids was systematically evaluated as a repository of potential antitumour agents. The primary objective was to discern their potency in overcoming resistance to temozolomide (TMZ). Through an extensive screening process, honatisine, a heptacyclic diterpenoid alkaloid, emerged as the most robust candidate. Notably, honatisine exhibited remarkable efficacy in patient-derived primary and recurrent GBM strains. Subsequently, we subjected this compound to comprehensive scrutiny, encompassing GBM cultured spheres, GBM organoids (GBOs), TMZ-resistant GBM cell lines, and orthotopic xenograft mouse models of GBM cells. RESULTS: Our investigative efforts delved into the mechanistic underpinnings of honatisine's impact. It was discerned that honatisine prompted mitonuclear protein imbalance and elicited the mitochondrial unfolded protein response (UPRmt). This effect was mediated through the selective depletion of mitochondrial DNA (mtDNA)-encoded subunits, with a particular emphasis on the diminution of mitochondrial transcription factor A (TFAM). The ultimate outcome was the instigation of deleterious mitochondrial dysfunction, culminating in apoptosis. Molecular docking and surface plasmon resonance (SPR) experiments validated honatisine's binding affinity to TFAM within its HMG-box B domain. This binding may promote phosphorylation of TFAM and obstruct the interaction of TFAM bound to heavy strand promoter 1 (HSP1), thereby enhancing Lon-mediated TFAM degradation. Finally, in vivo experiments confirmed honatisine's antiglioma properties. Our comprehensive toxicological assessments underscored its mild toxicity profile, emphasizing the necessity for a thorough evaluation of honatisine as a novel antiglioma agent. CONCLUSION: In summary, our data provide new insights into the therapeutic mechanisms underlying honatisine's selective inducetion of apoptosis and its ability to overcome chemotherapy resistance in GBM. These actions are mediated through the disruption of mitochondrial proteostasis and function, achieved by the inhibition of TFAM-mediated mtDNA transcription. This study highlights honatisine's potential as a promising agent for glioblastoma therapy, underscoring the need for further exploration and investigation.

Triptolide synergistically enhances temozolomide-induced apoptosis and potentiates inhibition of NF-κB signaling in glioma initiating cells.

Glioblastoma multiforme (GBM) is a lethal solid cancer in adults. Temozolomide (TMZ) is a first-line chemotherapeutic agent but the efficacy is limited by intrinsic and acquired resistance in GBM. Triptolide (TPL), a derivative from traditional Chinese medicine, demonstrated anti-tumor activity. In this study, we explored the interaction of TPL and TMZ in glioma-initiating cells (GICs) and the potential mechanism. A GIC line (GIC-1) was successfully established. Cell viability of GIC-1 after treatment was measured using a CCK-8 assay. The interaction between TPL and TMZ was calculated from Chou-Talalay equations and isobologram. Self-renewal was evaluated with tumor sphere formation assay. Apoptosis was assessed with flow cytometry and western blot. Luciferase assay was employed to measure NF-κB transcriptional activity. The expression of NF-κB downstream genes, NF-κB nuclear translocalization and phoshorylation of IκBα and p65 were evaluated using western blot. We found that GIC-1 cells were resistant to TMZ, with the expected IC50 of 705.7 µmol/L. Co-treatment with TPL yielded a more than three-fold dose reduction of TMZ. TPL significantly increased the percentage of apoptotic cells and suppressed the tumor sphere formation when combined with TMZ. Phosphorylation of IκBα and p65 coupled with NF-κB nuclear translocalization were notably inhibited after a combined treatment. Co-incubation synergistically repressed NF-κB transcriptional activity and downstream gene expression. TPL sensitizes GICs to TMZ by synergistically enhancing apoptosis, which is likely resulting from the augmented repression of NF-κB signaling. TPL is therefore a potential chemosensitizer in the treatment of GBM.