Asparagine depletion by the antileukemic agent PEGylated asparaginase induces hepatic stellate cell activation and fibrosis

Abstract

<b>Abstract ID 15650</b> <b>Poster Board 451</b> <b>Background:</b> One of the leading therapies for acute lymphoblastic leukemia (ALL) is the chemotherapeutic agent PEGylated asparaginase (PEG-ASNase). However, PEG-ASNase therapy is avoided in adults due to the high risk of liver injury, which presents clinically as elevated hepatic transaminases, bilirubin levels, and fatty liver. In our previous studies, we developed a murine model of PEG-ASNase-induced fatty liver that recapitulates clinical liver injury observations. We demonstrated, using this model, that PEG-ASNase-induced liver injury is associated with the activation of adipose triglyceride lipase (ATGL), which leads to adipocyte lipolysis, free fatty acid (FFA) mobilization, and fatty liver. However, whether the fatty liver induced by PEG-ASNase treatment progresses to advanced liver injury such as to hepatic stellate cell (HSC) activation and hepatic fibrosis is unknown. <b>Hypothesis:</b> PEG-ASNase treatment activates HSCs and induces hepatic fibrosis. <b>Methods:</b> Nine-week-old wild-type C57BL/6 mice received a 1,500 IU/kg single dose of PEG-ASNase or vehicle control intraperitoneally. Blood and liver samples were collected five days after administration. Alpha-SMA immunochemistry and Sirius Red staining were performed to assess the expression of the fibrogenic marker and collagen deposition, respectively. LX-2 (human HSC line) and isolated primary murine HSCs were treated with 1IU/mL of PEG-ASNase, and the mRNA and protein levels of the liver fibrosis markers (a-SMA, TGF-b, COL1A1, and COL1A2) were assessed. <b>Results:</b> We show that PEG-ASNase treatment was associated with increased immunostaining of a-SMA and collagen deposition in the liver, suggesting that PEG-ASNase activates HSCs. Consistent with our in&nbsp;vivo findings, PEG-ASNase treatment induced the protein expression of a-SMA and the mRNA expression of liver fibrosis marker genes, a-SMA, TGF-b, COL1A1, and COL1A2 in LX2 cells and primary murine HSCs. Similar to our previous findings using adipocytes, we found that PEG-ASNase treatment induced the mRNA and protein expressions of ATGL, supporting that PEG-ASNase-induced HSC lipolysis may be associated with HSC activation and the development of liver fibrosis. <b>Conclusion:</b> Our results support that PEG-ASNase therapy induces HSC activation and hepatic fibrosis. We hypothesize that PEG-ASNase therapy leads to ATGL-induced-HSC lipolysis, retinol release, and thereby HSC activation. <b>Support/Funding Information:</b> National Institutes of Health grants CA216815 and TL1TR001858 (USA), the Pittsburgh Liver Research Center, and American Foundation for Pharmaceutical Education (USA), and the University of Pittsburgh School of Pharmacy (USA).