Dual targeted gene delivery strategy mediated by GalNAc-modified lipid nanoparticles enhances liver regeneration through specific knockdown of MKK4.

Acute and chronic liver diseases are often related to the disorder of liver regeneration. However, there is no drug specifically approved for promoting liver regeneration especially in acute-on-chronic liver failure (ACLF). According to recent studies, inhibition of mitogen-activated protein kinase kinase 4 (MKK4) protein is critical for promoting hepatocyte proliferation and liver regeneration. However, MKK4 is widely present in various parts of the body as well with the similar binding pocket as MKK7 for small molecular inhibitors, and non-specific inhibition of MKK4 expression may pose an off-target risk in clinical practice. Therefore, MKK4-siRNA (siMKK4) was designed and optimized as a therapeutic gene for specific knockdown of MKK4 and avoid the off-target binding to mitogen-activated protein kinase kinase 7 (MKK7). Moreover, N-acetylgalactosamine (GalNAc)-modified lipid nanoparticles (LNPs) were used as gene delivery carriers to construct a dual targeted gene therapy system GalNAc-LNP-siMKK4 with liver tropism and active targeting to hepatocyte. GalNAc-LNP-siMKK4 can be efficiently constructed by the reverse phase evaporation method, with uniform particle size, good stability, biocompatibility, hepatocyte targeting ability, and high gene silence effect on the expression of MKK4.