Effects of BCG-treatment on urinary bladder cancer with focus on nitric oxide

Intravesical Bacillus Calmette-Guérin is used as a single agent or adjuvant therapy in the treatment of non-muscle invasive bladder cancer. In spite being one of the most efficient immunological treatments in cancer, BCG for bladder cancer is afflicted with a number of unsolved problems. One third of the patients do not respond, and BCG treatment may delay alternative curative therapies. The exact mechanism of action of BCG on bladder cancer is still in question. Further research and investigation is required to understand how to identify the non-responders, how to optimize treatments and decrease side effects, and ultimately which patients are most suitable for treatment and will have the most favourable results.

In this thesis, we have examined the long-term effects and efficacy of BCG in bladder cancer, and in line with previous research in our group, investigated possible roles of nitric oxide in the BCG reaction. Our aim was to deepen the understanding of the BCG-reaction, and ultimately to identify a predictive marker for BCG-treatment of bladder cancer. We investigated the long-term efficacy of BCG at reducing recurrence, progression and cancer specific mortality (CSM); and more specifically if NO played a part in these processes.

The third paper of this thesis showed that BCG reduced the long-term risk of recurrence (HR 0.40 p<0.0001) and progression (HR 0.52 p=0.038) in high-risk NMIBC in a fixed cohort followed for 15 years. Concerning progression, the risk reduction was only statistically significant in patients without concomitant CIS. For CSM there was no statistically significant risk reduction. The other two studies suggest a role for NO in BCG treatment of bladder cancer. Firstly, we have identified possible predictive markers in BCG treatment. In the same cohort as above, promoter and intragenic polymorphisms in NOS2 and NOS3 were associated with altered outcomes after BCG treatment. Secondly, we have confirmed earlier findings that NO concentration is increased in the bladder during the induction course of BCG treatment and that macrophages stimulated with BCG express both NOS2 and NO. We have shown in vitro that urothelial bladder cancer cells express NOS2 and NO and, perhaps more importantly that this reaction and BCG induced cytotoxicity depend upon activated macrophages. Interestingly, macrophage dependent BCG induced cytotoxicity was independent of nitric oxide.

Future implications of these findings require further research, most importantly to verify our findings about NOS polymorphisms and BCG efficacy in a larger cohort, but also to investigate the association between NOS polymorphisms and functional outcome (NOS expression and NO formation) in bladder cancer.

List of scientific papers

I. Thiel, T., Ryk, C., Chatzakos, V., Hallén Grufman, K., Bavand-Chobot, N., Flygare, J., Wiklund, P., de Verdier, P. Secondary stimulation from Bacillus Calmette-Guérin induced macrophages induce nitric oxide independent celldeath in bladder cancer cells. Cancer Lett. 2014 Jun 28;348(1-2):119-25.
https://doi.org/10.1016/j.canlet.2014.03.014

II. Ryk, C., Renström-Koskela, L., Thiel, T., Wiklund, P., Steineck, G., Schumacher, M., de Verdier, P. Outcome after BCG treatment for urinary bladder cancer may be influenced by polymorphisms in the NOS2 and NOS3 genes. Redox Biology. 2015 Dec;6:272-7.
https://doi.org/10.1016/j.redox.2015.08.008

III. Thiel, T., Ryk, C., Renström-Koskela, L., Steineck, G., Schumacher, M., Wiklund, P., de Verdier, P. Intravesical BCG treatment causes a longlasting reduction of recurrence and progression in patients with high-risk non-muscle-invasive bladder cancer. World J Urol. 2019 Jan;37(1):155-163.
https://doi.org/10.1007/s00345-018-2375-7