Dextromethorphan inhibits secretion of Collagens, Latent transforming growth factor proteins (LTBP1, LTBP2) and like cargoes to elicit an anti-fibrotic response.
Excessive deposition of fibrillar collagen in the interstitial extracellular matrix (ECM) of human lung tissue causes fibrosis, which can ultimately lead to organ failure. Despite our understanding of the molecular mechanisms underlying the disease, no cure for pulmonary fibrosis has not yet been found. In this study, we screened an FDA- approved drug library containing 712 drugs and found that Dextromethorphan (DXM), a cough expectorant, significantly reduces the amount of excess fibrillar collagen deposited in the ECM in in-vitro cultured primary human lung fibroblasts (NHLF) and ex-vivo cultured human precision-cut lung slice (hPCLS) models of lung fibrosis. The reduced extracellular fibrillar collagen levels upon DXM treatment are due to a reversible trafficking inhibition of collagen type I (COL1) in the endoplasmic reticulum (ER) in TANGO1 and HSP47 positive structures. Mass spectrometric analysis shows that DXM causes hyper-hydroxylation of proline and lysine residues on various collagen (COL1, COL3, COL4, COL5, COL7, COL12) and Latent-transforming growth factor beta-binding protein (LTBP1 and LTBP2) peptides coinciding with their secretion  block. In addition, thermal proteome profiling of cells treated with DXM shows increased thermal stability of prolyl- hydroxylases such as P3H2, P3H3, P3H4, P4HA1 and P4HA2, suggesting a change in activity. Transcriptome analysis of pro-fibrotic stimulated NHLFs and hPCLS upon DXM treatment showed activation of an anti- fibrotic program via regulation of pathways such as those involved in the MMP- ADAMTS axis, WNT, and fibroblast-to-myofibroblast differentiation. Taken together, the data obtained from in-vitro, in-vivo and ex-vivo models of fibrogenesis show that DXM has potent anti-fibrotic activity by efficient inhibition of COL1 membrane trafficking in the ER.