Abstract C108: The therapeutic candidate antibody PODO447 recognizes a glycopeptide on podocalyxin-expressing tumor cells that have undergone a partial epithelial-mesenchymal transition

Abstract In tumor cells, protein glycosylation patterns are often dramatically altered compared to normal cells. This leads to the emergence of novel neo-antigens on the tumour cell surface. Podocalyxin is a heavily glycosylated transmembrane sialomucin that is overexpressed in many solid tumors, with higher expression being significantly correlated with poor prognosis. We have developed a candidate therapeutic monoclonal antibody called PODO447 that binds to a tumor-associated altered glycoform of podocalyxin (Canals Hernaez et al., J Immunother Cancer. 2020 Nov;8(2):e001128). Clinically, the glycopeptide epitope recognized by PODO447 is present on a significant subset of podocalyxin-positive tumors and high expression of the epitope is associated with an aggressive ‘cold’ immune phenotype in patient tumors (Brassard et al., Front Oncol. 2023 Dec 21:13:1286754.). An antibody drug conjugate (ADC) that contains PODO447 as its targeting arm has shown promising results for tumor-killing efficacy in pre-clinical models of pancreatic and ovarian carcinoma (Canals Hernaez et al., Front Oncol. 2022 May 4:12:856424). As the expression of the PODO447 glycoepitope associates with aggressive tumor types, we assessed the phenotype of PODO447-expressing cells to better elucidate the potential role of the emergence of this novel epitope in tumor progression. The CFPAC-1 pancreatic ductal adenocarcinoma-derived cell line, while being universally podocalyxin-positive, contains subpopulations of cells that express either high or low levels of the PODO447 epitope. We were able to sort these two populations through fluorescent-activated cell sorting (FACS) and performed bulk RNA-seq, comparing expression patterns between these two populations. Interestingly, we found genes that are known markers of epithelial to mesenchymal transition (EMT) to be upregulated in PODO447High cells, including transcription factors (TWIST1, ZEB1, ZEB2) and extracellular matrix components and remodelling factors (FN1, ITGA5, COL1A1, MMP2, MMP13). CFPAC-1 PODO447High cells also have elevated N-cadherin levels compared to PODO447Low cells. In addition, while E-cadherin mRNA and total protein levels are unchanged, we found that the localization of E-cadherin is altered in PODO447High cells, being restricted to the intracellular compartment, compared to a junctional, cell surface localization in PODO447Low cells. Furthermore, the PODO447High cells have a more mesenchymal morphology and increased single-cell motility. Collectively, these data suggest that the emergence of the PODO447 epitope is associated with the acquisition of a partial EMT (p-EMT) phenotype that may functionally contribute to tumor progression. Targeting this tumor-specific podocalyxin epitope clinically with our antibody may prove effective in preventing disease progression. Citation Format: Pamela Austin Dean, Erin M. Bell, Stephane Flibotte, Anjali Parthasarathy, Lydia Liu, Aileen Liman, Julyanne Brassard, Kelly M. McNagny, Calvin D. Roskelley. The therapeutic candidate antibody PODO447 recognizes a glycopeptide on podocalyxin-expressing tumor cells that have undergone a partial epithelial-mesenchymal transition [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2025 Oct 22-26; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2025;24(10 Suppl):Abstract nr C108.