Recombinant human endostatin (rEndostatin or endostar) has been shown to inhibit endothelial cells proliferation, migration, and angiogenesis and exhibits a broad spectrum of activities against solid tumors. However, rEndostatin is easily degradable and evenly distributed to all tissues. Selectively delivering rEndostatin to the lesion site might be more potent. The circumsporozoite protein (CSP) coats the malarial sporozoite and targets the liver for infection; I-plus of N end of CSP could specifically bind to the liver. Based on this, we hypothesize the fusion protein with introducing the CSP I-plus sequence into rEndostatin (rES-CSP) of which not only targets the liver, but also inhibits endothelial cells proliferation, migration, and tube formation. Therefore, it selectively reduces angiogenesis of hepatocellular carcinoma (HCC) and improves the anti-HCC effect. In this study, we synthesized a novel rES-CSP fusion gene by SOE-PCR and expressed the fusion protein in Escherichia coli BL2l (DE3). The suitable conditions were optimized by an orthogonal test (L(25)(5)(4)). The yields were 12 mg/l culture medium following refolding and purification on nickel-nitrilotriacetic acid (Ni-NTA) metal affinity chromatography matrices. The purified rES-CSP is specifically targeted to the hepatocyte and inhibited the proliferation and migration of human umbilical vein endothelial cells (HUVECs) in a dose-dependent manner and showed potent antiangiogenic capability on HUVECs tube formation assay and chick embryo chorioallantoic membrane (CAM) assay. These results lay the foundation for the further study of its targeting and anti-HCC in vivo and provide a feasible and convenient approach to produce liver-targeting drugs for treatment of the liver diseases.