Abstract 1716: Acquired cell adhesion molecule 1 (CADM1) ADC resistance model for optimizing payload efficacy and toxicity in osteosarcoma preclinical testing

Abstract Instruction: Antibody-drug conjugates (ADCs) are a rapidly advancing therapeutic approach. A proof of principle CADM1 ADC has shown significant efficacy in osteosarcoma preclinical studies; however, its activity may be compromised by inherent or acquired resistance during prolonged treatment. In this study we generated osteosarcoma models resistant to a CADM1 ADC to define the underlying molecular mechanisms. Methods: The antitumor efficacy of CADM1 ADC was validated in vivo in six osteosarcoma patient-derived xenografts (PDXs) at a 3 mg/kg dose. To model acquired resistance, osteosarcoma PDXs were subjected to three cycles of treatment with a low-dose (0.5 mg/kg) CADM1 ADC. Tumor samples were harvested from pre-treated controls and post-treatment resistant PDXs. These samples were analyzed using immunohistochemistry (IHC), flow cytometry, whole-genome sequencing (WGS), and bulk RNA sequencing (RNA-seq). Resistant PDXs were further tested with two alternative ADCs: one using the same payload but targeting a different antigen and another using the same target antigen but with a different payload. IC50 values were determined in pre-treated and resistant PDX-derived cells to evaluate cytotoxicity. Results: After three cycles of low-dose CADM1 ADC treatment, PDX models exhibited a gradual decline in tumor regression, indicating the development of resistance. IHC and flow cytometry analyses demonstrated variable CADM1 expression between pre-treatment and resistant samples, with only some tumors exhibiting decreased target expression. Whole-genome sequencing (WGS) revealed increased gene copy number gains in resistant tumors compared to controls. RNA-seq analysis identified significant changes in gene expression, including reduced CADM1 expression in some samples and upregulation of resistance-associated pathways. Notably, the multidrug resistance-associated protein (MRP) transporter ABCC3 was significantly overexpressed in resistant tumors compared to pre-treatment samples. Cytotoxicity assays further confirmed resistance, with resistant PDX-derived cells exhibiting higher IC50 values compared to pre-treated cells, indicating reduced sensitivity to CADM1 ADC. Conclusions: CADM1 ADC demonstrates potent antitumor activity in osteosarcoma PDX models but resistance can develop. Future studies will delve deeper into protein localization and post-translational modifications as well as single cell genomics to better understand the mechanisms of resistance. Citation Format: Zhongting Zhang, Wendong Zhang, Xin Zhou, Caterina Longo, Yifei Wang, Dongxue Mao, Yanhua Yi, Zhaohui Xu, Zhao Li, Simon Olivares, Sylvester Jusu, Harjeet Singh, Michael Roth, Jonathan Gill, Bissan Al-Lazikani, Richard Gorlick. Acquired cell adhesion molecule 1 (CADM1) ADC resistance model for optimizing payload efficacy and toxicity in osteosarcoma preclinical testing [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 1716.