The Integrin-TGFβ Axis: Inhibition of Integrin αvβ6 Prevents Radiation-Induced Lung Fibrosis

Article Outline

• Purpose/Objective(s)
• Materials/Method
• Results
• Conclusions
• Copyright

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Purpose/Objective(s) 

Dose escalation for thoracic neoplasms is limited by the risk of permanent pulmonary radiation injury such as pulmonary fibrosis. There is an urgent need to identify new molecular targets and therapeutics for radiation-induced lung fibrosis (RILF). TGFβ is a key cytokine involved in the pathogenesis of radiation fibrosis. TGFβ triggers inflammatory and pro-fibrotic responses involving a fibroblast phenotype, matrix production, protease activity and apoptosis. TGFβ is secreted in a latent form due to an interaction with its propeptide, and TGFβ activators release TGFβ from this complex. In the lung, a major TGFβ activator is the integrin αvβ6. We have previously shown that integrin αvβ6 is necessary for radiation-induced lung fibrosis (ASTRO, 2005).

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Materials/Method 

Itgb6−/− mice and control C57BL/6 wildtype mice were anesthetized positioned in a dedicated plexiglass tray that enabled blocking of the rest of the body to selectively treat the lungs with a 14 Gy single dose fraction delivered by a ⁶⁰Co source. We assessed fibrosis in mice sacrificed at 26 weeks by calculating the percent fibrosis area of paraffin-embedded, formalin-fixed tissue sections that were stained with Masson trichrome. To assess the effects of an inhibitory monoclonal antibody to integrin αvβ6, we further injected C57BL/6 mice with various doses of anti-αvβ6 antibodies (Biogen Idec; 1, 3, 6, 10 mg/kg/wk), soluble TGFβ receptor (5 mg/kg), or appropriate controls (PBS and IgG), starting 15 weeks after 14 Gy thoracic radiation.

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Results 

Mice lacking integrin αvβ6, Itgb6−/−, are protected against RILF. At 27 weeks, wildtype control mice (21/23) developed 17% ± 3% mean area of lung fibrosis, while none of the 17 Itgb6−/− mice revealed any fibrotic areas. In addition, we show that inhibition of integrin αvβ6 with a monoclonal antibody beginning 16 weeks after radiation prevents RILF. C57BL/6 mice were injected with a neutralizing integrin αvβ6 monoclonal antibody in escalating dosage (1, 3, 6, 10 mg/kg/wk). Among control IgG-treated mice, 35/42 mice had histologically evident areas of fibrosis. In contrast 25/42, 10/42, 8/42 and 9/41 of the mice treated with 1-, 3-, 6- or 10-mg/kg/wk neutralizing integrin αvβ6 antibody, respectively, had evident areas of fibrosis. Profile of bronchoalveolar lavage (BAL) proteins showed reduced levels of fibrosis-associated proteins in the BAL fluid of integrin αvβ6 monoclonal antibody-treated mice, even at the low dose of inhibitor (1 mg/kg/week).

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Conclusions 

Our results demonstrate a robust requirement for integrin αvβ6 in the radiation-induced lung fibrosis model, likely due to its function as a TGFβ-1 activator. Integrin αvβ6 upregulation is a late event post-radiation, and is temporally and spatially linked to fibrosis. Inhibition of integrin αvβ6-mediated TGFβ activation is a promising new approach for anti-fibrosis therapy.