Ad5_NULL-A20: A tropism-modified, avb6 integrin-selective oncolytic adenovirus for epithelial ovarian cancer therapies

Purpose: Virotherapies are maturing in the clinical setting. Adenoviruses (Ad) are excellent vectors for the manipulability and tolerance of transgenes. Poor tumor selectivity, off-target sequestration, and immune inactivation hamper clinical efficacy. We sought to completely redesign Ad5 into a refined, tumor-selective virotherapy targeted to avb6 integrin, which is expressed in a range of aggressively transformed epithelial cancers but nondetectable in healthy tissues. Experimental Design: Ad5_NULL-A20 harbors mutations in each major capsid protein to preclude uptake via all native pathways. Tumor-tropism via avb6 targeting was achieved by genetic insertion of A20 peptide (NAVPNLRGDLQVLAQKVART) within the fiber knob protein. The vector's selectivity in vitro and in vivo was assessed. Results: The tropism-ablating triple mutation completely blocked all native cell entry pathways of Ad5_NULL-A20 via coxsackie and adenovirus receptor (CAR), avb3/5 integrins, and coagulation factor 10 (FX). Ad5_NULL-A20 efficiently and selectively transduced avb6^þ cell lines and primary clinical ascites-derived EOC ex vivo, including in the presence of preexisting anti-Ad5 immunity. In vivo biodistribution of Ad5_NULL-A20 following systemic delivery in non–tumor-bearing mice was significantly reduced in all off-target organs, including a remarkable 10⁷-fold reduced genome accumulation in the liver compared with Ad5. Tumor uptake, transgene expression, and efficacy were confirmed in a peritoneal SKOV3 xenograft model of human EOC, where oncolytic Ad5_NULL-A20–treated animals demonstrated significantly improved survival compared with those treated with oncolytic Ad5. Conclusions: Oncolytic Ad5_NULL-A20 virotherapies represent an excellent vector for local and systemic targeting of avb6-overexpressing cancers and exciting platforms for tumor-selective overexpression of therapeutic anticancer modalities, including immune checkpoint inhibitors.