A phenotypic screen identifies xanthohumol and other flavonoids as killers of bladder cancer

Bladder cancer accounts for 4 % of cancer diagnoses in the US. Current treatments primarily involve trans-urethral resection of bladder tumors (TURBT) and immunotherapy with Bacille Calmette-Guerin (BCG). Despite the efficacy of TURBT, issues with residual tumors persist. We chose to utilize Cell Painting to screen a set of 244 flavonoid compounds for bladder cancer cell toxicity. Using multiparametric high content analysis termed SPACe, we discover promising candidates underscoring the potential of flavonoids in targeting bladder cancer cells and elucidating their mechanisms of action. Notably, compounds such as xanthohumol show promise in reducing cancer cell viability by altering lipid metabolism. We further show the effectiveness of hit compounds in subsequent spheroid and chorioallantoic membrane systems. Overall, this research emphasizes the role of innovative screening methods in drug discovery and potential synergistic effects of combining flavonoids with existing therapies like BCG for improved bladder cancer treatment outcomes. • Phenotypic screening of a 224-compound flavonoid library identifies 10 flavonoids toxic to bladder cancer cell lines. • Phenotypic screening allowed determination of both DNA damaging and mitochondrial dysfunction-inducing compounds. • Xanthohumol, a flavonoid found in hops ( Humulus lupulus ), reduced lipid droplets within the cancer cell lines. • Xanthohumol could decrease bladder tumor growth in a chorioallantoic membrane model.