A 3D fluid-dynamic tumor microenvironment-on-chip to model tumor-stroma-vascular interactions in esophageal adenocarcinoma

Tumor progression and therapeutic resistance in esophageal adenocarcinoma (EAC) are influenced by the tumor microenvironment (TME), which is composed of cancer cells, stromal cells including cancer-associated fibroblasts, endothelial cells, immune cells, and extracellular matrix [1]. To better understand interactions within TME and overcome the limitations of traditional 2D cultures, we propose a 3D, fluid-dynamic model for EAC using the MIVO® milli-fluidic platform (Figure 1). In this model, spheroids composed of OE33 esophageal cancer cells and CCD-1132Sk dermal fibroblasts are cultured on the apical side of a 24-well insert, while endothelial cells (HMEC-1) form a monolayer on the underside of the insert membrane and this insert is integrated within the MIVO® chamber (Figure 1). Culture conditions were optimized to develop OE33 spheroids with diameters ranging from 200 - 600 μm. Spheroid viability was tested under static and dynamic conditions (0.12 to 0.38 mm/s). While smaller spheroids remained viable under both conditions for 48 h, necrotic cores were observed in larger spheroids. Furthermore, stable HMEC-1 endothelial monolayers were obtained on the underside of the insert across flow velocities 0.02 to 0.12 mm/s and were maintained for up to 72 h. Going forward, we will upgrade these spheroids by coculturing with fibroblasts and characterize them under static and dynamic conditions using biomarkers. This model enables precise control over nutrient gradients and mechanical cues, making it a unique platform to investigate tumor-stroma-vascular interactions under physiologically relevant flow conditions. Designed to recapitulate the in vivo TME accurately, this model has potential for both mechanistic studies as well as preclinical testing.