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Background: In intermediate risk non-muscle-invasive bladder cancer (NMIBC) clinical guidelines suggest an adjuvant instillation with a chemotherapeutic agent. However, the agent and regimen are not clearly defined. Worldwide, less than 15% of patients receive this adjuvant chemotherapeutic instillation. We recently developed a pipeline for the generation of patient derived organoids (PDO) in NMIBC. In this phase II trial, we aim to use our in vitro pipeline to select the most effective drug for chemotherapeutic instillation in NMIBC patients. Methods: Patients with first diagnosis of intermediate risk NMIBC that are directed to TURBT are enrolled. During TURBT, tumor is sampled, and specimens are directed to generate PDOs. Once the PDOs are formed, drug screens on them for Epirubicin, Mitomycin C, Gemcitabine and Docetaxel are performed. The drug with the highest antitumor activity in vitro will then be selected for 6 adjuvant intravesical instillations once weekly. Thereafter, patients are followed according to clinical guidelines by cystoscopy. Discussion: The aim of this trial is to use drug screens in PDO to precise treatment selection for adjuvant instillation therapies in patients with intermediate risk NMIBC. The ultimate goal of this trail is to reduce the risk of cancer recurrence. In the future, we aim to conduct clinical multicenter trials with an increased sample size, a broader panel of compounds and a focus on the reduction of cancer recurrence by precision delivery of care. Trial registration: NCT05024734

Urothelial carcinomas (UC) arise from the urothelium that covers the proximal urethra, urinary bladder, and the upper urinary tract. In daily routine and clinical trials UC originating from different locations are often treated and investigated in the same manner. However, differences between the two locations seem to be apparent and may question in handling them as a single oncologic entity. In this review we discuss similarities and differences between bladder and upper urinary tract UC and consider their potential impact on treatment strategies. Despite similarities of UC in the bladder (BC) and the upper urinary tract (UTUC), clinicopathologic and molecular differences may question to generally assemble both as a single tumor entity. Treatment standards for UTUC are often adopted from BC. However, a specific investigation in the former may still be meaningful as shown by the example of adjuvant cisplatin based chemotherapy. In conclusion, future investigations should prioritize the understanding of the tumor biology of both BC and UTUC. This may reveal which UTUC can be treated according to treatment standards of BC and in which cases, a separate approach may be more appropriate.

Bladder Cancer (BLCa) inter-patient heterogeneity is considered the primary cause of tumor reoccurrence and treatment failure, suggesting that BLCa patients could benefit from a more personalized treatment approach. Patient-derived organoids (PDOs) have been successfully used as a functional model for predicting drug response in different cancer types. In our study, we established BLCa PDO cultures from different BLCa stages. BLCa PDOs preserve the histological and molecular heterogeneity of the parental tumors, including their multiclonal genetic landscapes. BLCa PDOs consistently share key genetic alterations detected in parental tumors, mirroring tumor evolution in longitudinal sampling. Our drug screening pipeline was implemented using BLCa PDOs, testing both standard-of-care and additional FDA-approved compounds for other solid tumors. Integrative analysis of drug response profiles with matched PDO genomic analysis was used to determine enrichment thresholds for candidate markers of therapy resistance and sensitivity. By assessing the clinical history of longitudinally sampled cases, the clonal evolution of the disease could be determined and matched with drug response profiles. In conclusion, we have developed a clinically relevant pipeline for drug response profile assessment and discovery of candidate markers of therapy resistance.

Introduction: Interpatient heterogeneity is one of the causes of treatment failure in bladder cancer (BLCa) patients, who would therefore profit from tailor-made therapies. Toward the direction of precision medicine, patient-derived organoids (PDOs), which retain parental tumor (PT) features, have been successfully tested in different cancer types and shown to predict patients’ responses. Therefore, our study aimed to explore the ability of BLCa PDOs to retain PT features and to determine patient drug sensitivity. We correlated PDO drug response profiles to their genomic profiles and the patient's clinical history. Methods: PDOs were derived from the suspension culture of single cells. Whole exome sequencing was performed on genomic DNA and scRNA sequencing on 3 PT/PDO pairs. Histological evaluation of PDO and PT marker expression and morphology was performed. PDOs were treated for 48h and drug response was evaluated by viability assay. Results: We derived PDOs from non-muscle-invasive and muscle-invasive BLCa. Genomic analysis of 15 representative PT/PDOs pairs showed high similarity in terms of copy number variations and single nucleotide variants (SNVs) with preservation of peculiar BLCa alterations. PT genomic heterogeneity was preserved in PDOs as demonstrated by the correlation between SNVs clonality profiles of PT/PDOs pairs. Moreover, marker analyses at the scRNA and protein levels supported that PDOs retained the main tumor phenotype. Interestingly, PDO grew accordingly to three morphologies linked to marker expression and PT stage.PDOs were used in drug screen assays for testing standard-of-care (SOC) and FDA-approved drugs. PDO responses were highly heterogeneous highlighting interpatient heterogeneity. In 2 case, PDO sensitivity mimicked the patient’s response to SOC and in a few cases correlated with the sample-specific genomic background. Due to the high relapse rate of BLCa, we performed 2 longitudinal studies. In the first one, PDOs from a relapse collected after epirubicin treatment showed less sensitivity to the drug compared to the baseline sample. Moreover, the SNVs' mean clonality of relapse PDOs was significantly higher than the baseline PDOs which were characterized by mutations in DNA damage repair genes (e.g., ATM, FANC1, ERCC2, and BRCA2). We hypothesize a drug-induced selection of a pre-existing epirubicin-resistant population. In the second study, PDOs from the cystectomy and the following relapse showed similar genomic and drug sensitivity profiles suggesting that the first sample could be informative for adjuvant therapy. Conclusion: We established PDOs from different BLCa stages and grades which preserve PT features and can be implemented in drug screening assays for personalized approaches. The power of PDOs in precision medicine was highlighted by the longitudinal studies that showed that PDOs mirrored patient response and tumor evolution in vitro. Citation Format: Martina Minoli, Thomas Cantore, Daniel Hanhart, Mirjam Kiener, Tarcisio Fedrizzi, Federico La Manna, Sofia Karkampouna, Panagiotis Chouvardas, Vera Genitsch, José Antonio Rodriguez-Calero, Eva Compérat, Irena Klima, Paola Gasperini, Bernhard Kiss, Roland Seiler-Blarer, Francesca Demichelis, George N. Thalmann, Marianna Kruithof-de Julio. Bladder cancer organoids as a functional system to model different disease stages and therapy response [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2023; Part 1 (Regular and Invited Abstracts); 2023 Apr 14-19; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2023;83(7_Suppl):Abstract nr 182.

INTRODUCTION AND OBJECTIVE:Bladder cancer (BLCa) is the 10th most frequent cancer worldwide [1]. Non-muscle invasive (NMBC) and muscle invasive BLCa (MIBC) are the two major categories used to defi...

Bladder cancer is a heterogeneous disease that is not depicted by current classification systems. It was originally classified into non-muscle invasive and muscle invasive. However, clinically and genetically variable tumors are summarized within both classes. A definition of three groups may better account for the divergence in prognosis and probably also choice of treatment. The first group represents mostly non-invasive tumors that reoccur but do not progress. Contrarily, the second group represent non-muscle invasive tumors that likely progress to the third group, the muscle invasive tumors. High throughput tumor profiling improved our understanding of the biology of bladder cancer. It allows the identification of molecular subtypes, at least three for non-muscle invasive bladder cancer (Class I, Class II and Class III) and six for muscle-invasive bladder cancer (luminal papillary, luminal non-specified, luminal unstable, stroma-rich, basal/squamous and neuroendocrine-like) with distinct clinical and molecular phenotypes. Molecular subtypes can be potentially used to predict the response to treatment (e.g., neoadjuvant chemotherapy and immune checkpoint inhibitors). Moreover, they may allow to characterize the evolution of bladder cancer through different pathways. However, to move towards precision medicine, the understanding of the biological meaning of these molecular subtypes and differences in the composition of cell subpopulations will be mandatory.