P1.105. Epithelial–Mesenchymal Plasticity Underlies Adaptive Resistance to FLOT Chemotherapy in Esophageal Adenocarcinoma

Epithelial–mesenchymal transition in carcinoma: navigating phenotypic states to target resistance and metastasis

The clinical translation of EMT-directed interventions faces steep methodological hurdles, primarily driven by unverified causal dynamics and unresolved therapeutic safety profiles, and will require dynamic, multi-pronged strategies tailored to specific carcinoma cell states and spatial niches.

TNIK maintains a MYC-driven partial EMT state that supports proliferation and evasion of senescence in lung squamous cell carcinoma

Abstract A040: Microphysiological models uncover invasive and proteomic shifts in Ewing sarcoma cell subpopulations

Epithelial-to-mesenchymal transition (EMT) drives morphological and molecular changes that promote cancer progression. Ewing sarcoma cells, driven by fluctuations in the master oncogene EWS FLI1, can shift along the EMT spectrum. Yet the proteomic changes underlying EMT, including alterations in cell–cell and cell–matrix interactions, remain poorly understood. We use microphysiological systems to identify metastatic drivers and examine how 3D tumor organization and external EMT induction (via Wnt-5a and TGF-β1) reshape cell subpopulation protein expression and invasiveness. TC71 GFP cells were treated with StemXVivo EMT Inducing Supplement for 5 days and assessed by wound healing, viability assays, western blot, and single-cell proteomics. We developed a 3000-cell tumor spheroid micromodel (500–1000 µm diameter) and used it to evaluate invasion and chemoresistance with greater physiological relevance. EMT induction was chemoprotective in 2D monolayers (Doxorubicin IC50: control 8.88±1.57nM (±95%Confidence Interval) vs EMT 12.6±1.67nM) but not in spheroids (control 114.2±28.16nM vs EMT 72.73±16.88nM). To assess invasive potential, spheroids were embedded in Cultrex basement membrane matrix with 100 ng/ml IGF-1—an ignition signal for bone sarcomas that mimics adolescent hormonal conditions. IGF-1 significantly increased the furthest invasive distance from the spheroid centroid (n=15/group, control ± IGF-1 p=0.02; EMT ± IGF-1 p=0.0123). While EMT enhanced wound-healing and migration in 2D (p<0.001), control spheroids showed greater invasion (p=0.0063), indicating that increased motility does not necessarily translate into greater invasiveness. To investigate the structural changes underlying these behaviors, we examined F-actin—a cytoskeletal regulator —and ezrin—a membrane–cytoskeleton crosslinker and EMT regulator. We found a statistically significant increase in bulk ezrin expression by western blot, both with the addition of IGF-1 from control and after EMT induction (n=3 replicates, p=0.015 and p=0.0086, respectively). To investigate the heterogeneity of actin expression and shifts induced by EMT stimulation, we used Single Cell Protein Interaction Fractionation Through Electrophoresis and Immunoassay Readout (SIFTER), a single-cell proteomics tool for quantifying multimeric protein complexes. EMT stimulation significantly increased median F actin levels in monolayers (control median 6.76E4 Arbitrary Fluorescence Units (AFU) (n=760 cells, coefficient of quartile variation CQV=0.74) vs EMT median 8.41E4 AFU (n=324, CQV=1.06) p<0.0001), with a further rise in spheroids (control spheroid median 10.55E4 AFU (n=214, CQV=0.80) vs spheroid EMT median 22.12E3 AFU (n=244, CQV=0.74) p<0.0001). This work represents the first use of a spheroid model for single-cell proteomic analysis in Ewing sarcoma and reveals that EMT stimulation increased invasion, elevated F-actin levels, and broadened single-cell heterogeneity, with stronger shifts in 3D spheroids. Neica Ivens. Joseph, Julea Vlassakis, PhD. Microphysiological models uncover invasive and proteomic shifts in Ewing sarcoma cell subpopulations [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Breaking Barriers in the Fight against Rare Cancers; 2026 Jul 18-20; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2026;86(14_Suppl):Abstract nr A040.

Epithelial interferon-stimulated gene programs modulate chemotherapy sensitivity in colorectal cancer via EPSTI1.

Although chemotherapy in colorectal cancer (CRC) primarily targets epithelial tumor cells, the epithelial cell-intrinsic mechanisms underlying heterogeneous treatment responses remain poorly understood. We integrated the genomic, transcriptomic, and in vitro drug response profiles of CRC patient-derived tumor organoids (PDTOs) with functional assays to identify epithelial cell-intrinsic determinants of chemotherapy response. An epithelial interferon-stimulated gene (ISG) program, reflecting JAK-STAT pathway activity, was associated with reduced chemosensitivity in PDTOs. Single-cell RNA sequencing data of CRC tumors before and after neoadjuvant chemotherapy showed persistent epithelial ISG expression, with EPSTI1 notably enriched in tumors with incomplete pathological responses. Functional studies demonstrated that EPSTI1 knockdown reduced CRC cell viability and enhanced chemosensitivity. Pharmacologic JAK-STAT inhibition with ruxolitinib suppressed ISG expression and attenuated chemotherapy-induced ISG upregulation. These findings highlight the utility of PDTOs for studying tumor-intrinsic mechanisms of treatment response and identify EPSTI1 as an epithelial modulator of chemosensitivity.

Hypoxia and epithelial to mesenchymal transition pathways are enriched in bladder tumor epithelium adjacent to tertiary lymphoid structures

P1.106. Proteomic Signatures of Chemoresistance in Oesophageal Adenocarcinoma Cell Models

Quantitative proteomic profiling of acquired chemoresistant oesophageal adenocarcinoma cell models identifies broad adaptive changes with both heterogeneous and recurrent components, and suggests a platinum-class component supported by overlap between oxaliplatin- and carboplatin-resistant models.