AT2R agonists buloxibutid (Compound 21) and NAc inhibit fibrogenesis in human precision cut lung slices ex vivo.

Abstract

1. Buloxibutid, also known as Compound 21 (C21), is an angiotensin type 2 receptor AT2R agonist in early clinical trials for IPF, having shown antifibrotic efficacy in a bleomycin mouse model (PMID 29636695; 2018). 2. To compare C21 with an experimental AT2R agonist NAc and the IPF drug pirfenidone (PFD), using human precision cut lung slices (hPCLS). 3. Matched hPCLS from agarose-inflated lungs were left untreated or stimulated with fibrotic cocktail (FC = TGFp1, TNFa, LPA, PDGF; PMID 28314802; 2017) +/- C21 (10uM), NAc (10uM) or PFD (500uM). In situ fibrosis was assessed after 120h by Masson's trichrome staining. Secreted cytokines and extracellular matrix proteins were measured by ELISA in hPCLS-conditioned media collected at 48h and 120h respectively. 4. FC did not induce collagen deposition in hPCLS but increased secretion of both procollagen 1a1 (ng/mL: vehicle 40+/-12; FC 209+/-28, n=21, p<0.01, paired ttest) and fibronectin (mg/mL: vehicle 1.6+/-0.2; FC 4.4+/-0.3, n=15, p<0.01). C21 and NAc, but not PFD, significantly reduced secretion of both matrix proteins (p<0.05, one-way ANOVA, n=17, 14). Both AT2R agonists performed as well as PFD in inhibiting FC-induced IL-6 and IL-8 secretion (p<0.05, one-way ANOVA, n=5). 5. Fibrogenesis and inflammation can be modelled ex vivo in hPCLS using a cocktail of IPF-relevant mediators. C21 and NAc, at 50-fold lower concentrations than PFD, significantly reduced secretion of both fibrogenic and inflammatory markers. Establishing reversal of fibrosis by AT2R agonists in hPCLS from IPF lung would address a major limitation of current therapy and further support clinical translation of this novel drug class for IPF.