Nintedanib inhibits keloid fibroblast functions by blocking the phosphorylation of multiple kinases and enhancing receptor internalization

Abstract

Keloid is a benign skin tumor characterized by its cell hyperproliferative activity, invasion into normal skin, uncontrolled growth, overproduction and deposition of extracellular matrices and high recurrence rate after various therapies. Nintedanib is a receptor tyrosine kinase inhibitor targeting VEGF, PDGF, FGF, and TGF-β receptors with proved efficacy in anti-angiogenesis and in treating various types of cancers. In this study, we investigated the effects of nintedanib on keloid fibroblasts in both in vitro and ex vivo models. Keloid fibroblasts were prepared from 54 keloid scar samples in active stages collected from 49 patients. We found that nintedanib (1−4 μM) dose-dependently suppressed cell proliferation, induced G0/G1 cell cycle arrest, and inhibited migration and invasion of keloid fibroblasts. The drug also significantly inhibited the gene and protein expression of collagen I (COL-1) and III (COL-3), fibronectin (FN), and connective growth factor (CTGF), as well as the gene expression of other pathological factors, such as alpha smooth muscle actin (α-SMA), plasminogen activator inhibitor-1 (PAI-1), FK506-binding protein 10 (FKBP10), and heat shock protein 47 (HSP47) in keloid fibroblasts. Furthermore, nintedanib treatment significantly suppressed the phosphorylation of p38, JNK, ERK, STAT3, and Smad, enhanced endocytosis of various growth factor receptors. Using an ex vivo tissue explant model, we showed that nintedanib significantly suppressed cell proliferation, migration, and collagen production. The drug also significantly disrupted microvessel structure ex vivo. In summary, our results demonstrate that nintedanib is likely to become a potential targeted drug for keloid systemic therapy.

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Fig. 1: Nintedanib inhibits the proliferation of treated keloid fibroblasts.
Fig. 2: The inhibitory effect of nintedanib on the migration of keloid fibroblasts.
Fig. 3: The inhibitory effect of nintedanib on keloid fibroblast invasion.
Fig. 4: The antifibrotic effect of nintedanib on reducing ECM gene expression and inhibiting related protein production.
Fig. 5: Nintedanib inhibited collagen accumulation and disrupted microvessels in cultured keloid explants.
Fig. 6: Nintedanib inhibited intracellular signaling pathways in vitro and enhanced receptor internalization.

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Acknowledgements

This study was financially supported by the National Natural Science Foundation of China (No. 81671921). The authors also thank Prof. Jung Huang from the Biochemistry Department of St. Louis University School of Medicine (USA) for his valuable suggestion on part of the experimental design and valuable discussion on related scientific issues.

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BYZ and WBW performed research, analyzed the data and drafted the manuscript; XLW, WJZ and GDZ provided valuable insight into the experimental design, data review and critical advice on manuscript preparation; WL and ZG designed research, reviewed the experimental data; WL edited and finalized the manuscript.

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Correspondence to Zhen Gao or Wei Liu.

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The authors declare no competing interests.

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Zhou, By., Wang, Wb., Wu, Xl. et al. Nintedanib inhibits keloid fibroblast functions by blocking the phosphorylation of multiple kinases and enhancing receptor internalization. Acta Pharmacol Sin 41, 1234–1245 (2020). https://doi.org/10.1038/s41401-020-0381-y

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Keywords

  • nintedanib
  • anti-keloid activity
  • keloid fibroblasts
  • TGF-β/Smad signaling
  • MAPK signaling
  • ex vivo tissue explant model

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