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P002 Inhibition of Axl signaling by BGB324 reduces fibrogenesis in human intestinal cells and human intestinal organoids

Steiner C.1, Rodansky E.2, Johnson L.A.2, Huang S.2, Spence J.1, Higgins P.D.*3

1University of Michigan, Internal Medicine - Gastroenterology, Ann Arbor, United States 2University of Michigan, Internal Medicine - Gastroenterology, Ann Arbor, Michigan, United States 3University of Michigan, Internal Medicine - Gastroenterology, Ann Arbor, MI, United States

Background

In Crohn's disease, fibrosis is the final common pathway to intestinal failure. Fibrostenotic disease is the primary cause of obstruction and reason for surgical intervention in these patients. Currently no medical therapies exist to treat intestinal fibrosis.

We found that Axl is induced in both in vitro and in vivo models of intestinal fibrosis. Axl is a tyrosine kinase targeted for the treatment of acute myeloid leukemia (AML). Inhibition of Axl signaling has been shown to reverse epithelial-mesenchymal transition, sensitize cells to apoptosis, and reduce liver fibrosis via hepatic stellate cell inactivation. BGB324 is an orally available tyrosine kinase inhibitor that is highly selective for Axl. We evaluated the effect of BGB324 on pro-fibrotic gene expression in three in vitro models.

Methods

We assessed Axl gene expression in strictured human intestine vs. unaffected margin from human Crohn's patients and in 5 models of Crohn's disease. These included the fibrotic vs. unaffected intestine in rat TNBS, mouse salmonella, CCD18Co substrate stiffness, CCD18Co TGF-β (fibrogenic cytokine), and human intestinal organoid (HIO) TGF-β models of fibrosis. We then treated our three in vitro models with BGB324, assessing for reduction in pro-fibrotic gene expression of collagen 1A1 (COL1A1), fibronectin 1 (FN1), myosin light chain kinase (MYLK), and smooth muscle actin (ACTA2) using real-time PCR.

Results

In strictured intestine vs. unaffected margin from Crohn's patients, Axl expression was increased approximately 5 fold. Axl induction was also observed in the fibrogenic state in all 5 models of intestinal fibrosis. In rat TNBS and mouse salmonella models, Axl expression increased 1.8 and 2 fold respectively. In CCD18Co cell cultures, pathological substrate stiffness and TGF-β models caused 1.4 and 1.7 fold increases respectively. In HIO, TGF-β increased Axl by 1.8 fold.

BGB324 abrogated expression of COL1A1 in the CCD stiffness model as well as the CCD TGF-β model. Similar results were obtained when analyzing FN1, MYLK, and ACTA2.

In the TGF-β HIO model, BGB324 reduced pro-fibrotic gene expression. Random-effects meta-analysis of multiple biologic replicates demonstrated reduction in pro-fibrotic gene. Individual fold reductions and 95% CI for MYLK, FN1, COL1A1, and ACTA2 were 3.87 [1.58. 6.17], 3.17 [1.55, 4.79], 2.24 [0.91, 3.57], and 1.37 [0.27, 2.46] respectively.

Conclusion

Axl signaling appears to be an important pathway in intestinal fibrosis. BGB324 is being actively investigated in clinical trials for AML, and thus far appears safe for human use. The use of potent Axl inhibitors including BGB324 represents a therapeutic avenue for the treatment of intestinal fibrosis that warrants further testing.