Human adenovirus type 5 (HAdV-5)-based oncolytic viruses (OVs) hold promise as efficient anti-cancer therapy. Systemic OV administration is the preferable administration route as it allows to target difficult-to-address tumours and metastases. Poor tumour targeting and off-target organ transduction, however, limit the therapeutic efficacy of HAdV-5-based oncolytic virotherapy. Especially the strong inherent liver tropism of HAdV-5-based vectors poses a risk of severe hepatotoxicity.

We generated a novel HAdV-5-based oncolytic vector with reduced overall net negative surface‑charge by genetic modification of the major capsid protein Hexon (HexPos3). HAdV‑5‑HexPos3 exhibited superior and CAR-independent transduction efficiency in various cancer cell lines in vitro, further enhanced in the presence of HAdV-5 naïve murine plasma. Upon single i.v. administration into tumour-bearing NSG mice, replication-deficient HAdV-5-HexPos3 vector particles had a significantly reduced off-target organ tropism in all tissues analysed, including the liver. Moreover, we detected significantly increased intratumoural vector amounts for HAdV‑5-HexPos3 after a single i.v. injection, leading to a 29-fold elevated tumour‑to‑liver ratio compared to a Hexon-unmodified control vector. Single i.v. injection of a conditionally replicating Hexon-unmodified control vector induced severe hepatotoxicity in tumour-bearing NSG mice. In contrast, i.v. injection of a conditionally replicating HAdV-5-HexPos3 vector was well tolerated, led to prolonged survival of respective animals and resulted in intratumoural vector amounts detectable for up to 56 days post treatment. Thus, the HAdV-5-HexPos3 represents a favourable oncolytic vector and a promising platform for the generation of novel HAdV-5-based oncolytic vectors with reduced systemic toxicity and improved therapeutic efficacy.