The present disclosure provides lentiviral vectors comprising a nucleic acid sequence encoding a polypeptide with factor IX (FIX) activity, and methods of using such lentiviral vectors. The liver-targeted lentiviral vectors disclosed herein can be used for gene therapy, wherein the lentiviral gene delivery enables stable integration of the transgene expression cassette into the genome of targeted cells (e.g., hepatocytes) of pediatric (e.g., neonatal) or adult subjects, achieving an improvement in FIX expression at low lentiviral vector doses. The present disclosure also provides methods of treating bleeding disorders such as hemophilia (e.g., hemophilia B) comprising administering to a subject in need thereof a liver-targeted lentiviral vector comprising a nucleic acid sequence encoding a polypeptide with FIX activity sequence at low dosages.
