Introduction: Despite effective anti-inflammatory therapies, ~70% of CD patients require surgery in their lifetime, due to fibrostenotic strictures linked to excess extracellular matrix (ECM) deposition. This process is mainly driven by activated mesenchymal cells (human intestinal myofibroblasts, HIMFs). No approved anti-fibrotic drugs targeting strictures exist. Integrins (INTs) are cell surface heterodimers that mediate cell-ECM interactions, and profibrotic cellular processes, but their effect on intestinal fibrosis remains to be investigated. We evaluated experimental anti-INT agents on ECM deposition (collagen, COL; fibronectin, FN) and cellular migration in vitro, and experimental intestinal fibrosis in vivo.

Methods: Four different INT inhibitors were evaluated: Cpd7 (FN-binding), cpd16 (pan αV/α5), TA1 (TGFβ-activating), and Alk5-inhibitor (Alk5i). HIMFs isolated from freshly resected intestinal tissues procured from subjects with CD or non-CD controls were allowed to produce ECM in the presence/absence of test agents and deposited ECM was quantified. Boyden chamber was used to evaluate the effect of test agents on migration of HIMFs. Male BALB/c mice (6-8 weeks old) with or without two cycles of 3% DSS were treated with test agents or vehicle provided in feed through study duration. Inflammation, fibrosis and wall thickness were assessed by H&E, Masson’s trichrome, and picrosirius red (PSR) staining; COL/FN levels were assessed by immunolabeling.

Results: ECM deposition in TGFβ–stimulated HIMFs was reduced 2.5-, 3.0-, 3.75-, and 7.5-fold by cpd7, cpd16, TA1, and Alk5i, respectively, in a dose-dependent manner. Serum-induced migration of HIMFs was reduced 6.6-, 2.6-, 2.6-, and 2.8-fold in the presence of cpd7, cpd16, TA1, and Alk5i, respectively. Treatment with cpd7 and cpd16 reduced the clinical score in mice throughout the course of DSS administration; no effect was detected with TA1 or Alk5i. Inflammation and fibrosis scores were reduced in mice treated with cpd7/16 compared to untreated mice. Thickness of muscularis propria was significantly reduced in mice treated with cpd7 (83.87 ± 16.15 µm), cpd16 (53.92 ± 9.44 µm), or Alk5i (50.04 ± 12.07 µm) compared to untreated mice (147.90 ± 35.37 µm, P < .05 for all comparisons), while TA1 had no effect (137.14 ± 13.12 µm, P = 0.661). Reduced PSR, COL1, and FN staining were noted for cpd7/16-treated mice.

Conclusions: Our results reveal potent target-dependent activity of select anti-INT agents that inhibit profibrotic processes in vitro, and influence inflammation and fibrosis in vivo. These studies may lead to successful development of selective INT-targeted anti-fibrotic therapies for IBD.

William Massey

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