Decoding immune heterogeneity in bladder cancer : From cellular cartography to ex vivo models and antibody-based targeting

Bladder cancer is the ninth most common type of cancer globally, and the projected number of deaths is estimated to almost double by the year 2040. Immunotherapy with checkpoint inhibitors has revolutionized treatment of bladder cancer and can achieve durable responses; however, only a minority of patients respond. By identifying additional targets and treatment combinations, more patients could benefit from immunomodulatory treatments. To achieve this, an improved understanding of the bladder tumor microenvironment is necessary. There is also a significant challenge for transitioning novel drugs from preclinical research to clinical implementation, highlighting the need for improved translational models.In paper I, we found that immune infiltration increased with the invasiveness of the disease, but the composition of the T cell compartment did not differ. In contrast, we found differences in both immune infiltration and T cell composition when comparing different molecular subtypes, indicating that these shape different immune responses and may require different immunomodulatory interventions. In paper II, we defined the immune cell populations that infiltrate bladder tumors based on single-cell RNA-sequencing and further investigated their spatial distribution. We identified cellular niches in the tumor enriched for specific immune cell subsets. In particular, we identified SPP1 expressing macrophages in the tumor nest and their relative abundance among macrophages was associated with worse cancer-specific survival in patients receiving radical cystectomy, independently of other clinical parameters. Mature regulatory dendritic cells were preferentially located in an immune cell niche, and their interactions with the other immune cells present in this niche are potentially important for regulating the anti-tumor immune response. In paper III, we developed a protocol for ex vivo culturing of human tumor explants with intact tissue architecture using precision-cut bladder tumor slices and demonstrated that it is a relevant model for studying response to antibody treatments. In paper IV, we identified CD177 as a putative target on regulatory T cells based on analysis of single-cell RNA-sequencing data of bladder tumors, and our in vitro studies support that CD177 suppresses T cell activity. We furthermore developed human antibodies against CD177 and initial functional evaluation of a set of antibodies support that they stimulate T cell activation in the tumor microenvironment.In conclusion, the work in this thesis contributes to the bladder cancer corpus of knowledge, particularly regarding the immune system and its role in the disease. Our work has potential clinical implications in the form of identification of a putative prognostic macrophage population, as well as a novel potential cancer treatment strategy based on the antibodies developed. Further, we describe a pre-clinical model for evaluation of such antibodies.