Evaluating the effectiveness of hepatocyte-targeted LVs for F.IX gene transfer. (A) Schematic representation of the third-generation self-inactivating LVs used for these studies. The miRNA-regulated LV was generated by incorporating 4 tandem copies of a sequence completely complementary to miR-142-3p. Wpre indicates woodchuck hepatitis post-regulatory element. (B) Measurement of hF.IX:Ag in normal C57BL/6 mice treated with 2 × 10⁸ IU of LV.ALB.FIX (open and filled gray squares) or LV.ET.FIX (open and filled black circles). Results are presented for individual mice. (C) Measurement of hF.IX:Ag in hemophilia B mice treated with 5 × 10⁸ IU of LV.ET.FIX. Results are presented as the mean plus or minus the standard error of the mean (SEM; n = 7). (D) T-cell proliferation response to hF.IX antigen. Splenocytes were isolated from untreated () and LV.ET.FIX-treated () hemophilia B mice at 7 days after injection, and cultured either in complete RPMI media alone or complete RPMI medium containing hF.IX antigen (10 μg/mL). T-cell proliferation was determined by measurement of thymidine incorporation. Results are presented as the mean plus or minus SEM (n = 3, *P < .05). CPM indicates counts per million.