Cellular responses to morpholinated curcumin derivatives in human muscle-invasive bladder cancer cells

Curcumin is a well-known compound with anticancer properties, as demonstrated in preclinical studies. However, its clinical potential is limited by several drawbacks. Medicinal chemistry, therefore, offers the opportunity to develop curcumin derivatives through structural modifications. This study showed cytotoxic activity of curcumin derivatives in muscle-invasive bladder cancer cells (HT-1376). Cytotoxicity was assessed under normoxic and hypoxic conditions and compared with effects on human primary bladder epithelial cells (BdEC). The most promising compounds (2a and 2a-B) were further investigated in a 3D cell culture model. Mechanistic studies included the evaluation of lactate dehydrogenase release, cell cycle distribution, and induction of apoptosis and necrosis using luminescent-, colorimetric-, and fluorescence-based assays. Furthermore, the intracellular uptake was investigated using flow cytometry and confocal microscopy. Tested compounds showed greater cytotoxicity than curcumin, with enhanced effects under hypoxic conditions. The derivatives containing a keto-enol moiety showed greater selectivity for bladder cancer cells than curcumin. Compounds 2a and 2a-B also showed greater cytotoxicity in the 3D cell culture model than curcumin. Compounds 2a and 2a-B altered cell cycle progression, increasing the population of cells in the G2/M and S phases, and induced apoptosis in a time- and concentration-dependent manner. Comparative analysis with previously reported data identifies compound 2a as a promising candidate for further investigation due to its high selectivity for malignant versus normal bladder epithelial cells.