Antithrombin-binding heparan sulfate is ubiquitously expressed in epithelial cells and suppresses pancreatic tumorigenesis
Thomas Mandel Clausen, Ryan J. Weiss, Jacob R. Tremblay, Benjamin P. Kellman, Joanna Coker, Leo A. Dworkin, Jessica P. Rodriguez, Ivy M. Chang, Timothy Chen, Vikram Padala, Richard Karlsson, Hyemin Song, Kristina L. Peck, Satoshi Ogawa, Daniel R. Sandoval, Hiren J. Joshi, Gaowei Wang, L. Paige Ferguson, Nikita Bhalerao, Allison Moores, Tannishtha Reya, Maike Sander, Thomas C. Caffrey, Jean L. Grem, Alexandra Aicher, Christopher Heeschen, Dzung Le, Nathan E. Lewis, Michael A. Hollingsworth, Paul M. Grandgenett, Susan L. Bellis, Rebecca L. Miller, Mark M. Fuster, David W. Dawson, Dannielle D. Engle, Jeffrey D. Esko
Thomas Mandel Clausen, Ryan J. Weiss, Jacob R. Tremblay, Benjamin P. Kellman, Joanna Coker, Leo A. Dworkin, Jessica P. Rodriguez, Ivy M. Chang, Timothy Chen, Vikram Padala, Richard Karlsson, Hyemin Song, Kristina L. Peck, Satoshi Ogawa, Daniel R. Sandoval, Hiren J. Joshi, Gaowei Wang, L. Paige Ferguson, Nikita Bhalerao, Allison Moores, Tannishtha Reya, Maike Sander, Thomas C. Caffrey, Jean L. Grem, Alexandra Aicher, Christopher Heeschen, Dzung Le, Nathan E. Lewis, Michael A. Hollingsworth, Paul M. Grandgenett, Susan L. Bellis, Rebecca L. Miller, Mark M. Fuster, David W. Dawson, Dannielle D. Engle, Jeffrey D. Esko
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Research Article Cell biology Oncology

Antithrombin-binding heparan sulfate is ubiquitously expressed in epithelial cells and suppresses pancreatic tumorigenesis

Abstract

3-O-sulfation of heparan sulfate (HS) is the key determinant for binding and activation of antithrombin III (AT). This interaction is the basis of heparin treatment to prevent thrombotic events and excess coagulation. Antithrombin-binding HS (HSAT) is expressed in human tissues but is thought to be expressed in the subendothelial space, mast cells, and follicular fluid. Here, we show that HSAT is ubiquitously expressed in the basement membranes of epithelial cells in multiple tissues. In the pancreas, HSAT is expressed by healthy ductal cells, and its expression is increased in premalignant pancreatic intraepithelial neoplasia lesions but not in pancreatic ductal adenocarcinoma (PDAC). Inactivation of HS3ST1, a key enzyme in HSAT synthesis, in PDAC cells eliminated HSAT expression, induced an inflammatory phenotype, suppressed markers of apoptosis, and increased metastasis in an experimental mouse PDAC model. HSAT-positive PDAC cells bind AT, which inhibits the generation of active thrombin by tissue factor and factor VIIa. Furthermore, plasma from patients with PDAC showed accumulation of HSAT, suggesting its potential as a marker of tumor formation. These findings suggest that HSAT exerts a tumor-suppressing function through recruitment of AT and that the decrease in HSAT during progression of pancreatic tumorigenesis increases inflammation and metastatic potential.

Authors

Thomas Mandel Clausen, Ryan J. Weiss, Jacob R. Tremblay, Benjamin P. Kellman, Joanna Coker, Leo A. Dworkin, Jessica P. Rodriguez, Ivy M. Chang, Timothy Chen, Vikram Padala, Richard Karlsson, Hyemin Song, Kristina L. Peck, Satoshi Ogawa, Daniel R. Sandoval, Hiren J. Joshi, Gaowei Wang, L. Paige Ferguson, Nikita Bhalerao, Allison Moores, Tannishtha Reya, Maike Sander, Thomas C. Caffrey, Jean L. Grem, Alexandra Aicher, Christopher Heeschen, Dzung Le, Nathan E. Lewis, Michael A. Hollingsworth, Paul M. Grandgenett, Susan L. Bellis, Rebecca L. Miller, Mark M. Fuster, David W. Dawson, Dannielle D. Engle, Jeffrey D. Esko

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Figure 7

Loss of HSAT increases tumor inflammation and decreases apoptosis.

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Loss of HSAT increases tumor inflammation and decreases apoptosis. (A) S...
(A) Schematic representation of the orthotopic transplantation model performed with WT and HS3ST1–/– F1242 and FC1245 lines. Graphic illustration created in BioRender (https://BioRender.com/x05fpme). (B) Tumor size at 2 weeks after implantation, measured by ultrasound. No significant difference in tumor size observed. (C) RNA was extracted from the tumors (n = 5 per group) and subjected to RNA-Seq analysis. Data represented as a volcano plot with cutoffs of adjusted P value < 0.05 and log2 fold change ≥ 0.58. (D) Gene ontology analysis was analyzed in the upregulated genes in HS3ST1–/– line (log2 fold change > 0.58, adjusted P < 0.05) using Metascape (54). (E) Gene Set Enrichment Analysis was performed on the same data set, revealing enrichment for pathways involved in inflammation, EMT, and KRAS signaling. (F and G) Staining for Ki67 in FFPE tissue sections derived from the tumors (F) and for cleaved caspase-3 (CC3) (G). Graphs show the mean ± SD. Statistical analysis by 2-tailed t test in B, F, and G. *P ≤ 0.05; **P ≤ 0.01; NS, not significant.