Hyaluronan network remodeling by ZEB1 and ITIH2 enhances the motility and invasiveness of cancer cells
Sieun Lee, … , Jonathan M. Kurie, Young-Ho Ahn
Sieun Lee, … , Jonathan M. Kurie, Young-Ho Ahn
Published April 3, 2025
Citation Information: J Clin Invest. 2025;135(11):e180570. https://doi.org/10.1172/JCI180570.
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Research Article Cell biology Oncology

Hyaluronan network remodeling by ZEB1 and ITIH2 enhances the motility and invasiveness of cancer cells

Abstract

Hyaluronan (HA) in the extracellular matrix promotes epithelial-mesenchymal transition (EMT) and metastasis; however, the mechanism by which the HA network constructed by cancer cells regulates cancer progression and metastasis in the tumor microenvironment (TME) remains largely unknown. In this study, inter-α-trypsin inhibitor heavy chain 2 (ITIH2), an HA-binding protein, was confirmed to be secreted from mesenchymal-like lung cancer cells when cocultured with cancer-associated fibroblasts. ITIH2 expression is transcriptionally upregulated by the EMT-inducing transcription factor ZEB1, along with HA synthase 2 (HAS2), which positively correlates with ZEB1 expression. Depletion of ITIH2 and HAS2 reduced HA matrix formation and the migration and invasion of lung cancer cells. Furthermore, ZEB1 facilitates alternative splicing and isoform expression of CD44, an HA receptor, and CD44 knockdown suppresses the motility and invasiveness of lung cancer cells. Using a deep learning–based drug-target interaction algorithm, we identified an ITIH2 inhibitor (sincalide) that inhibited HA matrix formation and migration of lung cancer cells, preventing metastatic colonization of lung cancer cells in mouse models. These findings suggest that ZEB1 remodels the HA network in the TME through the regulation of ITIH2, HAS2, and CD44, presenting a strategy for targeting this network to suppress lung cancer progression.

Authors

Sieun Lee, Jihye Park, Seongran Cho, Eun Ju Kim, Seonyeong Oh, Younseo Lee, Sungsoo Park, Keunsoo Kang, Dong Hoon Shin, Song Yi Ko, Jonathan M. Kurie, Young-Ho Ahn

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Figure 2

ITIH2 KD inhibits the migration and invasion of lung cancer cells.

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ITIH2 KD inhibits the migration and invasion of lung cancer cells. (A) q...
(A) qRT-PCR analysis of Itih2 mRNA levels in 344SQ cells transduced with Itih2 shRNAs (shC or shE). P values from 1-way ANOVA followed by Dunnett’s multiple-comparison test. (B) Boyden chamber migration assay in 344SQ-ITIH2-KD cells. After 24 hours, migrated cells were stained with crystal violet. P values determined by 1-way ANOVA followed by Dunnett’s multiple-comparison test. Scale bar: 200 μm. (C) Scratch migration assay in 344SQ-ITIH2-KD cells. Migration activity, (1 – [scratch area ratio of 24 hours to 0 hours]) × 100%, was measured using ImageJ (NIH). Mean ± SD (n = 12). P values determined by 1-way ANOVA followed by Dunnett’s multiple-comparison test. (D) Boyden chamber invasion assay in HCC827 cells overexpressing ITIH2. P value determined by 2-tailed Student’s t test. Scale bar: 200 μm. (E) Mouse subcutaneous injection of 344SQ-ITIH2-KD cells. After 6 weeks of injection, the primary tumor volume and lung metastatic tumor nodules were measured at necropsy (n = 9 or 10). P values determined by 1-way ANOVA followed by Dunnett’s multiple-comparison test. (F) Mouse orthotopic injection of 344SQ-ITIH2-KD cells. After a week of injection, the tumor nodules on mediastinal lymph nodes and lungs were measured at necropsy (n = 7 or 8). H&E staining results of the lung sections are described. P values determined by 1-way ANOVA followed by Dunnett’s multiple-comparison test. Scale bar: 5 mm. (G) Mouse tail-vein injection of 344SQ-ITIH2-KD cells. After 10 days of injection, tumor nodules colonized in the lungs were measured at necropsy under a fluorescent microscope (n = 5). P values determined by 2-tailed Student’s t test. Scale bars: 5 mm (left), 2 mm (right). (H) Kaplan-Meier curve for progression-free survival in patients with LUAD was generated from the Kaplan-Meier plotter (66). Data represent the mean ± SD from a single experiment with biological replicates (n = 3, unless otherwise specified) and are representative of at least 3 independent experiments.