Abstract 5311: Characterization of ADC and TCE targets in patient tumors using high-parameter spatial proteomics

Abstract Target heterogeneity and intensity of expression in patient tumors are two of many critical factors in understanding the clinical potential of antibody drug conjugates (ADCs) and T cell engagers (TCEs) in oncology.1, 2 Despite extensive clinical usage, conventional immunohistochemistry (IHC) is often restricted to a handful of markers per section, limiting its capacity to identify complex patterns of antigen expression in patient samples. In contrast, high-parameter, spatially resolved, proteomic analysis has the potential to not only provide widespread characterization of tumor-associated antigens (TAAs) but also to reveal impactful cell-cell and protein-protein neighborhoods for bispecific pairings and/or novel biomarkers. Here, we utilized imaging mass cytometry (IMC) complemented by multiplex immunofluorescence (mIF) to spatially evaluate the expression of greater than 30 protein markers simultaneously in 35 tumor specimens from 7 indications. Using whole slide imaging and a bespoke analysis pipeline, coupled with statistical modeling, we profiled multiple TAAs, including HER2, TROP2, B7-H3, and B7-H4 for expression in tumor and adjacent non-tumor tissues.3 Notably, whole-slide mIF was used in tandem with IMC on the same tissue section to enable both region of interest (ROI) selection and image co-registration. With this approach, we observed a high level of concordance between the two modalities for markers of interest such as B7-H3 and B7-H4, helping to further validate our findings. Stromal and immune markers such as SMA (cancer-associated fibroblasts [CAFs, ]) CD3/CD8/FoxP3 (T cells, ) and CD206 (macrophages) were also included to help characterize the tumor microenvironment and cellular distribution of each TAA. The panel was also incorporated with markers of sensitivity to common ADC payloads, such as SLFN11 and TOP1. In addition to differences in target prevalence and level of expression, we describe the spatial relationship between various TAAs across tumors including such parameters as the average distance between cytotoxic T cells and a given TAA+ cell. Altogether, these results provide a framework to apply dual-modality imaging of mIF and IMC on the same tumor samples for comprehensive characterization of TCE and ADC targets. In the future, the total number of markers which can be characterized by both mIF and IMC are expected to increase, allowing for an even deeper characterization of tumor biology which may inform bispecific pairings and more precise treatment strategies for ADCs and TCEs. References: 1 - Cancers(Basel). 2023 Mar 19; 15(6):1845. doi: 10.3390/cancers15061845 2 - Cancers(Basel). 2020 Apr 26; 12(5):1075. doi: 10.3390/cancers12051075 3 - Cell Rep Methods. 2023 Oct 23;3(10):100595. doi: 10.1016/j.crmeth.2023.100595 Citation Format: Michael Alan Conner, Fang Xie, Lijun Zhou, Eliot McKinley, Jimmy Tarrant, Takahiro Sato, Ya-Fang Chang, Derek Poore, Shannon McKearnan, Ken Hance, Jeremy Waight. Characterization of ADC and TCE targets in patient tumors using high-parameter spatial proteomics [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 5311.