LAUSR

N‐Acetyl‐seryl‐aspartyl‐lysyl‐proline (Ac‐SDKP) is a critical negative regulator of fibrosis development in the liver. However, its extremely short half‐life in vivo greatly compromises its potential applications. Here, we report an Ac‐SDKP analog peptide with d‐amino acid replacement (Ac‐SDDKDP). The stability of Ac‐SDDKDP and its prevention of liver fibrosis were investigated in vitro and in vivo. The stabilities of Ac‐SDKP and Ac‐SDDKDP exposed to angiotensin‐1‐converting enzyme (ACE) and their half‐lives in rats and human sera were determined by high‐performance liquid chromatography. The inhibitory effects of Ac‐SDKP and Ac‐SDDKDP on the proliferation and activation of hepatic stellate cells (HSC‐T6) were evaluated using the Cell Counting Kit‐8, Western blotting, reverse transcription quantitative polymerase chain reaction, and immunofluorescence assays. Finally, the protective effects of Ac‐SDKP and Ac‐SDDKDP on carbon tetrachloride (CCl4)‐induced liver fibrosis in rats were compared. d‐Amino acid replacement significantly enhanced the stability of the peptide to ACE and prolonged the half‐life of Ac‐SDKP in rats and human sera. The Ac‐SDKP‐mediated inhibition of HSC‐T6 cell proliferation was well preserved, and Ac‐SDDKDP exerted inhibitory effects comparable to Ac‐SDKP on α‐smooth muscle actin (α‐SMA), collagen I and III expression, and phosphorylated‐Smad‐2 expression. After intraperitoneal (i.p.) administration, Ac‐SDDKDP exhibited significantly greater protection than Ac‐SDKP against CCl4‐induced liver fibrosis in rats. The serum alanine aminotransferase, aspartate aminotransferase, albumin, and total protein levels of the Ac‐SDDKDP‐treated rats were significantly lower than those of the Ac‐SDKP‐treated rats. α‐SMA, CD45, and collagen I and III expression, as well as Smad‐2 phosphorylation were significantly attenuated in the livers of the Ac‐SDDKDP‐treated rats compared to those of the Ac‐SDKP‐treated rats. Furthermore, we showed that the Ac‐SDDKDP concentration in the rat liver increased to a physiological level of 60 min after i.p. administration, although i.p. administration of Ac‐SDKP failed to enhance the peptide concentration in the rat liver. Our findings indicate that d‐amino acid replacement is a simple and effective method to enhance the stability of Ac‐SDKP. Ac‐SDDKDP represents potential application of Ac‐SDKP in fibrosis treatment and provides a new potential treatment strategy for liver fibrosis. © 2019 IUBMB Life, 71(9):1302–1312, 2019