Bladder cancer (BC) is one of the most prevalent cancers worldwide. However, the recurrence rate and five-year survival rate have not been significantly improved in advanced BC, and new therapeutic strategies are urgently needed. The anticancer activity of stellettin B (SP-2), a triterpene isolated from the marine sponge Rhabdastrella sp., was evaluated with the MTT assay as well as PI and Annexin V/7-AAD staining. Detailed mechanisms were elucidated through an NGS analysis, protein arrays, and Western blotting. SP-2 suppressed the viability of BC cells without severe toxicity towards normal uroepithelial cells, and it increased apoptosis with the activation of caspase 3/8/9, PARP, and γH2AX. The phosphorylation of FGFR3 and its downstream targets were downregulated by SP-2. Meanwhile, it induced autophagy in BC cells as evidenced by LC3-II formation and p62 downregulation. The inhibition of autophagy using pharmacological inhibitors or through an ATG5-knockout protected RT-112 cells from SP-2-induced cell viability suppression and apoptosis. In addition, the upregulation of DAPK2 mRNA and protein expression also contributed to SP-2-induced cytotoxicity and apoptosis. In RT-112 cells, an FGFR3-TACC3-knockout caused the downregulation of DAPK2, autophagy, and apoptosis. In conclusion, this is the first study demonstrating that SP-2 exhibits potent anti-BC activity by suppressing the FGFR3-TACC3/Akt/mTOR pathway, which further activates a novel autophagy/DAPK2/apoptosis signaling cascade.

Bladder cancer is one of the most prevalent cancers worldwide. Although most of the patients have good prognosis and good life expectancy with standard treatments, recurrence rate and five-year survival rate have not been significantly improved in patients with bladder cancer, demonstrating there is an urgent need for development of a novel therapeutic strategy for the treatment of bladder cancer. Stellettin B (SP-2), a triterpene isolated from marine sponge Jaspis stellifera , suppresses cell viability in bladder cancer RT112 cells without severe toxicity toward normal uroepithelial SV-HUC-1 cells. Our results showed that SP-2 induces accumulation of cells at subG1 phase in parallel with increase of early apoptotic cells via annexin V/PI staining and activation of cell apoptotic death proteins. Moreover, non-cytotoxic concentrations of SP-2 induces LC3-II formation and p62 downregulation to activate autophagy in RT112 cells, suggesting autophagy may play a role in SP-2 -induced cell death. Inhibition of autophagy by pharmacological inhibitors (3-MA and chloroquine) or ATG5 knockout protects stellettin B-induced apoptosis and cell viability suppression, indicating SP-2 induces autophagy to promotes cell death. Taken together, stellettin B may be a promising therapeutic agent for bladder cancer treatment. Citation Format: Mei-Chuan Chen, Chun-Han Chang. Stellettin B induces apoptosis and autophagy in human bladder cancer RT112 cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 6236.