TGF-β1/Smad3-IN-1 (Compound 5aa) is an inhibitor of the TGF-β1/Smad3 signaling pathway, with an IC50 value of 1.07 μM. It exhibits anti-fibrotic properties and is effective orally.

TGF-β1/Smad3-IN-1, at concentrations of 100-500 nM for 48 hours, effectively reduces TGF-β1 levels in H2228 cells, showing superior inhibition compared to the same concentrations of Nintedanib. When used at 2-6 μM for 24 hours, it dose-dependently suppresses the expression of p-Smad3 and α-SMA, significantly hindering the migration of NIH3T3 cells. At 3-10 μM for 72 hours, TGF-β1/Smad3-IN-1 increases Cleave-caspase3 expression in NIH3T3 cells, inducing apoptosis in a dose-dependent manner. It has an IC50 of 1.07 μM for NIH3T3 cells and 2.86 ± 0.014 μM for TGFβ1-activated HFL1 cells, effectively inhibiting the expression of the activation marker α-SMA.

TGF-β1/Smad3-IN-1 exhibits higher bioavailability in SD rats compared to Nintedanib. Administration of TGF-β1/Smad3-IN-1 (p.o.; 100 mg/kg/day; days 2–20) in a bleomycin-induced pulmonary fibrosis mouse model suppresses the expression of lung TGFβ1 and HYP induced by bleomycin, reduces extracellular matrix deposition, and alleviates pulmonary fibrosis.