Figure 6.

Ab-31 reduces TGFβ-induced αSMA expression in lung fibroblasts. (A) Normal human lung fibroblasts were stimulated with TGFβ (5 ng/mL) and stained with anti-αSMA antibody for immunofluorescence analysis. Cells were pre-treated with or without MK-0429 (10 μM) or the ALK5 inhibitor SB-525334 (10 μM) 30 min before the addition of TGFβ. 48 h after the treatment, cells were imaged with Opera Phenix high-content screening system for αSMA expression (fluorescence intensity) and αSMA-associated morphological changes (STAR program). (B) The effects of Ab-31, MK-0429, and benchmarking antibody mAb-24 on TGFβ-associated αSMA induction in IPF patient lung fibroblasts. C) Structural modeling predicts a distinct integrin binding mode for Ab-31. The structures of two therapeutic monoclonal antibodies, Abituzumab (17E6, RCSB PDB: 4O02) and LM609 (RCSB PDB: 6AVQ), in complex with αvβ3 integrin were shown to highlight the difference in binding modes for each molecule. A model of Ab-31 and αvβ3 integrin complex was determined by docking of related antibody sequence to αvβ3 structure (see “Methods”). For visualization, only the Fv region of each antibody were shown.