Gene silencing of ANGPTL3 suppresses α1-antitrypsin and lectin mannose-binding 1 expression in the hepatoma Huh7 cell line

Background: Angiopoietin-like 3 (ANGPTL3) is a hepatokine acting as negative regulator of hepatic and endothelial lipases. Vupanorsen, an antisense oligonucleotide targeting ANGPTL3, reduced triglycerides up to 60% in a phase 2b trial, but caused dose-dependent increases in hepatic fat fraction. Proprotein convertase subtilisin/kexin type 9 (PCSK9) plasma levels also correlates with liver fat accumulation. Methods: To further elucidate the biological activity of ANGPTL3 on lipid metabolism in the liver we performed a RNAseq analysis from human hepatoma Huh7 cells transfected with siRNA-ANGPTL3, siRNA-PCSK9 and double siRNA-ANGPTL3/PCSK9. Results: RNA-seq analysis detected 192 genes out of 13,945 transcripts differential ex-pressed in response to ANGPTL3-siRNA compared to scramble-siRNA controls. In cells transfected with PCSK9-siRNA, 88 genes were differentially expressed, while dual transfection with both ANGPTL3- and PCSK9-siRNA resulted in 219 differentially ex-pressed genes. When ANGPTL3 gene expression was silenced we observed a compensa-tory induction of proprotein convertase subtilisin/kexin type 9 (PCSK9) mRNA and pro-tein expression. Moreover, PCSK9 silencing increased ANGPTL3 mRNA expression. Bi-oinformatic analysis revealed that gene silencing of ANGPTL3 or both ANGPTL3/PCSK9 suppresses SERPINA1, gene encoding α1-antitrypsin, and lectin mannose-binding 1 ex-pression (LMAN1). These data were confirmed by western blot and qRT-PCR analysis. Conclusions: Our data identified two key genes SERPINA1 and LMAN1, involved in endoplasmic reticulum stress, lipid accumulation and liver damage, that are downregu-lated in response to gene silencing of ANGPTL3 in Huh7 hepatoma cell line.