Therapeutic action of BCG mediated by H₂O₂?

The clinical effects of BCG on bladder cancer might be mediated by the production of hydrogen peroxide (H₂O₂), according to new research published online in The Journal of Urology.

Despite BCG being an established treatment option for non-muscle-invasive bladder cancer, its mechanism of action remains unclear. Existing data largely suggest that BCG triggers an immune response in the bladder, leading to tumour cytotoxicity. However, BCG has also been shown to generate oxidative stress in the bladder. To investigate this association further, Gopitkumar Shah and colleagues at the Medical College of Wisconsin in Milwaukee, USA, performed a series of experiments to assess the relationship between cellular redox status and the antitumour effects of BCG.

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First, the investigators used fluorescent probes to profile the reactive oxygen and nitrogen species produced by BCG itself, and by urothelial cells that had been treated with BCG. They found that viable BCG generated H₂O₂ and superoxide (O₂⁻), but not nitric oxide (NO), and that heat-killed BCG produced twofold less H₂O₂ than viable BCG, suggesting that H₂O₂ might be involved in clinical activity. The urothelial cell lines 253J and T24 showed early increases in H₂O₂ and NO production after BCG treatment, with H₂O₂ and NO levels peaking in the first 12–18 h and 6 h, respectively.

Using the spindle toxin cytochalasin b to prevent BCG uptake into urothelial cells, Shah and colleagues demonstrated that H₂O₂ production was dependent on BCG internalization; urothelial cells treated with both BCG and cytochalasin b generated significantly lower levels of H₂O₂, O₂⁻ and NO than cells treated with BCG alone.

Similarly, pretreating cells with the potent H₂O₂ scavenger ebselen significantly reduced the production of H₂O₂, O₂⁻ and NO compared with BCG alone, highlighting the importance of H₂O₂ generation in this system.

To clarify the role of H₂O₂, researchers studied the effects of ebselen on BCG-induced gene expression, intracellular signalling and cell proliferation. Scavenging H₂O₂ with ebselen treatment significantly reduced the expression of eight BCG-responsive genes, and inhibited the activation of a number of signalling molecules known to be downstream of BCG (NFκB, CEBP and NRF2) in urothelial cancer cells. In addition, ebselen blocked the cytotoxic effect of BCG on urothelial cells, measured using the MTT assay.

These findings suggest that H₂O₂ is a source of oxidative stress and cell damage downstream of BCG in the bladder. The authors postulate that pharmacological manipulation of oxidative stress might be used to optimize the clinical benefits of BCG.