Introduction:

Current preclinical disease models for bladder cancer, including immortalized tumor cell lines and murine xeno- and allograft tumor models, fail to recapitulate the disease heterogeneity of urothelial carcinoma. Additionally, viral vector-based gene therapeutics, such as nadofaragene firadenovec, have species-specific tropisms that limit the ability of traditional murine preclinical disease models to predict clinical efficacy. Therefore, we evaluated and optimized patient-derived tumor organoids as a preclinical disease model for characterizing nadofaragene firadenovec direct cytotoxicity in urothelial carcinoma of the bladder.

Methods:

Patient-derived tumor organoids were established from cold-cup bladder tumor biopsies within six hours of tissue harvest. Tissue processing included enzymatic digestion followed by suspension in a solid Matrigel matrix submerged in hepatocyte growth media supplemented with human epidermal growth factor. Organoids were treated with nadofaragene firadenovec and evaluated for cell viability by Cell Titer Glo assay five days after treatment. Organoid transduction was confirmed by ELISA for IFN transgene expression in the supernatant of treated cells. Organoid growth was evaluated in suspension culture after eliminating the solid Matrigel matrix. 

Results:

From 15 unique patient tumor samples for which adequate tissue was obtained by cold cup biopsy, 9 tumor organoids were successfully established in vitro and expanded for at least one passage. Five organoids were subsequently expanded adequately for cytotoxicity studies, three from Ta disease and two from T1 disease.  All organoids showed decreased viability ranging from 20% to 60% after nadofaragene firadenovec treatment compared to organoids treated with a control adenoviral vector, and IFN transgene expression was detected in the supernatant of all nadofaragene firadenovec-treated organoids confirming viral transduction. Most recently, two organoids have been successfully established and expanded in suspension culture without the need for embedding in Matrigel, improving scalability and decreasing organoid doubling time.

Conclusion:

Patient-derived tumor organoids represent a novel preclinical model for evaluating the direct cytotoxicity of nadofaragene firadenovec, and optimization of organoid growth in suspension culture improves capacity for real-time evaluation of individual tumor sensitivities to novel therapeutics. Current studies are evaluating baseline and post-treatment organoid gene expression profiles and developing organoid-based orthotopic murine xenograft disease models.   

Funding: N/A