Engineered hydrogel reveals contribution of matrix mechanics to esophageal adenocarcinoma and identifies matrix-activated therapeutic targets
Ricardo Cruz-Acuña, … , Jason A. Burdick, Anil K. Rustgi
Ricardo Cruz-Acuña, … , Jason A. Burdick, Anil K. Rustgi
Published October 3, 2023
Citation Information: J Clin Invest. 2023;133(23):e168146. https://doi.org/10.1172/JCI168146.
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Research Article Cell biology Oncology

Engineered hydrogel reveals contribution of matrix mechanics to esophageal adenocarcinoma and identifies matrix-activated therapeutic targets

Abstract

Increased extracellular matrix (ECM) stiffness has been implicated in esophageal adenocarcinoma (EAC) progression, metastasis, and resistance to therapy. However, the underlying protumorigenic pathways are yet to be defined. Additional work is needed to develop physiologically relevant in vitro 3D culture models that better recapitulate the human tumor microenvironment and can be used to dissect the contributions of matrix stiffness to EAC pathogenesis. Here, we describe a modular, tumor ECM–mimetic hydrogel platform with tunable mechanical properties, defined presentation of cell-adhesive ligands, and protease-dependent degradation that supports robust in vitro growth and expansion of patient-derived EAC 3D organoids (EAC PDOs). Hydrogel mechanical properties control EAC PDO formation, growth, proliferation, and activation of tumor-associated pathways that elicit stem-like properties in the cancer cells, as highlighted through in vitro and in vivo environments. We also demonstrate that the engineered hydrogel serves as a platform for identifying potential therapeutic targets to disrupt the contribution of protumorigenic matrix mechanics in EAC. Together, these studies show that an engineered PDO culture platform can be used to elucidate underlying matrix-mediated mechanisms of EAC and inform the development of therapeutics that target ECM stiffness in EAC.

Authors

Ricardo Cruz-Acuña, Secunda W. Kariuki, Kensuke Sugiura, Spyros Karaiskos, Eleanor M. Plaster, Claudia Loebel, Gizem Efe, Tatiana Karakasheva, Joel T. Gabre, Jianhua Hu, Jason A. Burdick, Anil K. Rustgi

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Figure 1

Engineered hydrogel supports EAC PDO development.

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Engineered hydrogel supports EAC PDO development. (A) Images of patient ...
(A) Images of patient tissue sections from normal and EAC biopsies stained for HA (HABP), CD44, or fibronectin (FN1). Scale bar: 100 μm. Original magnification, ×5 (insets). (B) Bulk RNA-Seq analysis of ESCA and normal pancreatic tissue samples for fibronectin and hyaluronan-associated genes (HAS1, HAS2, UGDH). n = 286 for ESCA; n = 283 for normal. (C) Relationship between crosslinker density (mg/mL) and storage modulus, G′ (mean ± SEM; n = 3 independently prepared hydrogels per condition). (D) Quantification of PDO viability as assessed by calcein-AM labeling at 7 days after encapsulation. Viability is quantified as the percentage of PDOs that stained positive for calcein-AM (mean ± SEM; n = average number of calcein-AM+ organoids per hydrogel; at least 20 organoids per hydrogel were analyzed). Welch’s t test with 2-tailed comparison showed no significant differences between groups. NS = P > 0.05. (E) Representative transmitted light and fluorescence microscopy images of EAC PDOs cultured in NorHA hydrogels or Matrigel. Scale bar: 200 μm. (F) Representative fluorescence microcopy images of EAC PDOs within NorHA hydrogels stained for MUC5ac, E-cad, and CK8 at 14 days after encapsulation. Scale bar: 50 μm. Three independent experiments were performed, and data are presented for 1 of the experiments. Every independent experiment was performed with 4 gel samples per experimental group.