Anti-Fibrotic Effect of Combined mTORc1 and mTORc2 Inhibition in Bronchiolitos Obliterans

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Purpose The mammalian target of rapamycin (mTOR) pathway plays an important role in regulating protein translation. The aim of this study was to evaluate the contribution of mTORc1 and mTORc2 signaling in fibrotic functions of mesenchymal cell and the potential use of mTOR inhibitors as anti-fibrotic agents in bronchiolitis obliterans syndrome (BOS). Methods and Materials Lung-resident mesenchymal stromal cells (LR-MSCs) were isolated from bronchoalveolar lavage of patients±BOS. PI3K/PTEN/Akt/mTOR pathway was studied by western analysis of whole-cell lysates for total PTEN, phospho-Akt, and phospho-p70S6K1. Effect of mTOR inhibitors on collagen I and α-smooth muscle actin (α-SMA) expression was assessed. Results A significantly higher expression of p70S6Kinase (Thr389) and 4E-BP1 (Thr37/46) was noted in BOS LR-MSCs compared to non-BOS LR-MSCs. BOS LR-MSCs also exhibited increased expression of phosphorylated AKT (S473). A decrease in collagen I protein expression was seen in the presence of rapamycin, while α-SMA expression was not changed. Rapamycin inhibited p70S6Kinase but failed to decrease 4E-BP1 expression. An increased in pAKT(S473) in response to rapamycin was noted. A high baseline expression of pAKT(S473) in BOS LR-MSCs and its further upregulation by mTORc1 inhibition, led us to study AZD8055, an ATP-competitive mTOR inhibitor which inhibits downstream signaling from both mTORc1 and mTORc2. Treatment with AZD8055 led to a 96% collagen suppression compared to 62% decrease by rapamycin (p Conclusions mTOR signaling pathway is activated in BOS LR-MSCs. While treatment with rapamycin modestly inhibits fibrotic phenotype of LR-MSCs, its also results in augmented mTORc2 signaling. AZD8055, an ATP-competitive mTOR inhibitor, inhibits collagen, α-SMA, and mTOR substrate phosphorylation more significantly than rapamycin, suggesting AZD8055 as a promising therapeutic in the context of lung transplantation.