An oncolytic HAdV-5 with reduced surface charge combines diminished toxicity and improved tumor targeting

Human adenovirus type 5 (HAdV-5)-based oncolytic viruses hold significant promise for anti-cancer therapy. However, poor tumor-targeting and off-target organ transduction after systemic administration limit their therapeutic efficacy. In addition, the strong liver tropism of HAdV-5-based vectors poses the risk of hepatotoxicity. By genetic modification of the major capsid protein hexon we generated a HAdV-5-based oncolytic vector (HAdV-5-HexPos3) with reduced negative surface charge. Coxsackie and adenovirus receptor (CAR) binding-ablated (ΔCAR) HAdV-5-HexPos3_ΔCAR exhibited superior and CAR-independent transduction of various cancer cell lines <i>in vitro</i>, further enhanced in the presence of HAdV-5 naive murine plasma. Upon intravenous administration into tumor-bearing immunodeficient NSG mice, replication-deficient HAdV-5-HexPos3_ΔCAR vector particles showed significantly reduced off-target organ tropism in all tissues analyzed, including the liver. Moreover, we detected a significantly increased intratumoral vector load for HAdV-5-HexPos3_ΔCAR, leading to a 29-fold elevated tumor-to-liver ratio compared with a control vector with unmodified hexon. Intravenous injection of a conditionally replicating hexon-unmodified control vector induced severe hepatotoxicity in tumor-bearing NSG mice, while a conditionally replicating HAdV-5-HexPos3_ΔCAR vector was well tolerated and resulted in intratumoral vector presence for up to 56 days. HAdV-5-HexPos3_ΔCAR represents a promising vector platform for the generation of HAdV-5-based oncolytic viruses with reduced systemic toxicity and improved therapeutic efficacy.