Interrogating Matrix Stiffness and Metabolomics in Pancreatic Ductal Carcinoma Using an Openable Microfluidic Tumor-on-a-Chip

Pancreatic ductal adenocarcinoma (PDAC) is characterized by a dense fibrotic stroma that contributes to aggressive tumor biology and therapeutic resistance. Current in vitro PDAC models lack sufficient optical and physical access for fibrous network visualization, in situ mechanical stiffness measurement, and metabolomic profiling. Here, we describe an openable multilayer microfluidic PDAC-on-a-chip platform that consists of pancreatic tumor cells (PTCs) and pancreatic stellate cells (PSCs) embedded in a 3D collagen matrix that mimics the stroma. Our system allows fibrous network visualization via reflected light confocal (RLC) microscopy, in situ mechanical stiffness testing using atomic force microscopy (AFM), and compartmentalized hydrogel extraction for PSC metabolomic profiling via mass spectrometry (MS) analysis. In comparing cocultures of gel-embedded PSCs and PTCs with PSC-only monocultures, RLC microscopy identified a significant decrease in pore size and corresponding increase in fiber density. In situ AFM indicated significant increases in stiffness, and hallmark characteristics of PSC activation were observed using fluorescence microscopy. PSCs in coculture also demonstrated localized fiber alignment and densification as well as increased collagen production. Finally, an untargeted MS study putatively identified metabolic contributions consistent with in vivo PDAC studies. Taken together, this platform can potentially advance our understanding of tumor–stromal interactions toward the discovery of novel therapies.