Introduction: Ex-vivo high-throughput drug screening has the potential to improve risk-stratification, treatment personalisation, and novel drug development within clinically relevant timeframes. Bladder cancer (BC) is particularly challenging cancer to treat, with high morbidity and mortality rates, stagnant clinical outcomes, and poor patient reported experiences. The primary aim of this thesis was to explore the feasibility of ex-vivo drug screening using fresh patient BCs, its ability to identify differential standard and novel treatment responses, and whether whole exome sequencing (WES) could provide further molecular insight into response phenotypes.
Patients and methods: Bladder tumours were collected fresh from surgery and dissociated into BC patient-derived ex-vivo cultures (PDCs) (ethical approval STH15574/STH20854). Cells were incubated on bespoke, pre-loaded drug plates for four days. Endpoint metabolic (CellTiterGlo) assessment generated area dose-response curves (AUC). Tumours with vehicle-control (n=28) coefficient of variation >0.25 were excluded as methodological failures. AUC scores were normalised (modified-Z) across the tumour cohort and positive drug hits explored to determine differential ex-vivo phenotypic signatures. Whole exome sequencing (WES) was performed by MacrogenEurope and data used to identify candidate response biomarkers.
Results: There was high methodological success using the optimised ex-vivo methodological protocol, where 75.9% (41/54) of tumours screened passed quality control. In total, 39 (39/54, 72.2%) were malignant BCs. Diverging drug and tumour clusters and ex-vivo response phenotypes (EVP) were identified. Resistant-EVP was significantly associated with more aggressive clinical features (grade 3, p=0.0002, stage ≥T1, p=0.025, and carcinoma-in-situ, p=0.041. Resistant-EVP tumours had a higher median number of mutations per tumour (11.4 versus 7.5, p=0.036) and differential genotype to sensitive-EVP tumours. Ex-vivo determined cisplatin-resistant tumours dichotomised into those with alternative sensitivities and multi-drug resistant phenotypes, with differential genetic enrichment (Figure.1B&C).
Conclusions: Ex-vivo screening of BCs is feasible and can identify differential phenotypic behaviours. Resistant-EVP tumours displayed more aggressive clinical, drug phenotypic, and genotypic features. Identification of specific drug resistant cohorts may aid in clinical triage to more effective therapies or radical surgery.
