Abstract 5300: Spatially resolved integrated omics profiling for ADC targets in urothelial carcinoma

Abstract Background: Urothelial Carcinoma (UC) is the tenth most common cancer worldwide, accounting for more than 200 000 deaths annually (Ferlay, 2020, Sung, 2021). Despite advances over the past decade, particularly with the introduction of immunotherapy (ICI) in various disease settings alongside conventional platinum-based chemotherapy, the prognosis for metastatic UC (mUC) remains poor. Antibody drug-conjugates (ADC) have emerged as a novel treatment option for mUC, offering selective and targeted delivery of cytotoxic agents to cancer cells. ADCs targeting tumor-associated surface proteins including Nectin-4, trophoblastic cell surface antigen 2 (Trop-2) and tyrosine kinase receptor ERBB-2 (HER2), have demonstrated promising efficacy and are all approved for mUC (Wong, 2021, Goodson, 2023). However, the response rate to these ADCs in mUC is limited 25-40% and with a growing portfolio of accessible ADCs, identifying predictive biomarkers to optimize treatment selection remains an unmet medical need. The use of multiplexed immunofluorescence (mIF) offers deeper insights into tissue microenvironment and can potentially be used to better stratify patients for the various ADC targets in cancer. Recently, an integrated assay combining the sequential IF for protein detection with RNAscope was developed for Lunaphore’s COMET platform. This allows for simultaneous detection of several RNA targets and their corresponding proteins to be analyzed within the same cells. While the ADC target proteins have been shown to be highly expressed in many UC patients, there are limited studies characterizing the spatial expression of these protein targets, especially in combination with their corresponding transcripts. Methods: The integrated assay of the COMET platform offers a fully automated workflow that includes iterative cycles of staining, imaging and elution facilitating the simultaneous detection of RNA and proteins at single cell resolution. We used custom designed RNAscope probes (ACDBio), a SPYRE antibody panel (Lunaphore) and additional antibodies to detect the ADC-targets Nectin-4, Trop-2 and HER-2, and to spatially map the tumor microenvironment of human FFPE bladder cancer tissues from a cohort of mUC patients. Results & Discussion: We have optimized the assay for bladder cancer tissues on the COMET platform to detect multiple ADC targets at transcript and protein levels. We aim to employ the integrated multi-omics setup to explore the expression of the ADC targets in sections from a retrospective cohort of mUC patients treated by monotherapy with Enfortumab Vedotin (EV) in the third line. We have already observed differences across patients with regard to Nectin-4, Trop-2 and HER-2 protein expression and aim to better characterize the spatial distribution and expression of these tumor-associated antigens with the inclusion of target transcripts and the association to treatment benefit from EV. Citation Format: Pranauti Panshikar, Eleanor O´Roberts, David Krantz, Tony Ullman, Carolina Oses Sepulveda, Fernanda Svedman, Päivi Östling, Anders Ullén, Charlotte Stadler. Spatially resolved integrated omics profiling for ADC targets in urothelial carcinoma [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5300.