Hydroxycamptothecin (HCPT) is used as a clinical chemotherapy regimen to treat bladder cancer, but more efficacious novel combination treatments are needed.

Cultured bladder cancer cell lines EJ and UMUC3 were treated with triptolide (TPL) and/or HCPT. A flow cytometry approach was used to detect cell cycle phase, apoptosis and reactive oxygen species. Western blotting was used to measure CDK4, CDK6, CyclinD1, catalase, Caspase8 and Bcl-xl protein levels in control, TPL treatment, HCPT treatment and TPL plus HCPT treatment bladder cancer cells. AKT pathway proteins, including AKT and p-AKT, were also detected by western blotting. UMUC3 cells treated with DMSO, HCPT, TPL and HCPT plus TPL were injected subcutaneously into mice (n = 3 per group).

The flow cytometry and western blot results indicated that compared to TPL or HCPT treatment alone, combination treatment of HCPT and TPL significantly induced cell cycle arrest at the G1 phase via suppressing CDK4, CDK6 and CyclinD1 in the EJ and UMUC3 bladder cancer cell lines. HCPT and TPL combination treatment also significantly increased the apoptosis rate and apoptosis-related protein levels (Caspase8 and Bcl-xl). Levels of the AKT pathway-related proteins AKT/p-AKT were significantly lower in EJ and UMUC3 cells treated with TPL and UMUC3 than in those cells treated with TPL or HCPT alone. TPL plus HCPT treatment significantly reduced bladder tumour sizes in vivo on the seventh and tenth days.

Compared to TPL or HCPT treatment alone, TPL plus HCPT combination treatment had significantly better anticancer effects.