E1A-expressing adenoviral E3B mutants act synergistically with chemotherapeutics in immunocompetent tumor models

Abstract

The majority of clinical trials evaluating replication-selective oncolytic adenoviruses utilized mutants with immunomodulatory E3B genes deleted, likely contributing to the attenuated efficacy. We investigated whether an intact immune response could contribute to the observed improved efficacy in response to combinations with chemotherapeutics. Seven carcinoma cell lines were evaluated by combining viral mutants; dl309 (DE3B), dl704 (DE3gp19K), dl312 (DE1A) or wild-type Ad5 with the commonly used clinical drugs cisplatin and paclitaxel. Synergistic effects on cell death were determined by generation of combination indexes in cultured cells. In vivo tumor growth inhibition was achieved by virotherapy alone and was most efficacious with wild-type virus and least with the DE3B mutant. Significantly higher efficacy was observed when the viruses were combined with drugs. The greatest enhancement of tumor inhibition was in combination with the DE3B mutant restoring potency to that of Ad5 wild-type levels, observed only in animals with intact immune response. Increases in infectivity, viral gene expression and replication were identified as potential mechanisms contributing to the synergistic effects. Our results suggest that the attenuation of DE3B mutants can be overcome by low doses of chemotherapeutics only in the presence of an intact immune response indicating a role for T-cellmediated functions.

The Novel Oncolytic Adenoviral Mutant Ad5-3Δ-A20T Retargeted to αvβ6 Integrins Efficiently Eliminates Pancreatic Cancer Cells

“…Complete elimination of tumors was not possible with either Ad5-3D-A20T or Ad5wt because of limitations of the in vivo model. Murine tissues do not support productive infection with human adenovirus, preventing spread within the murine tumor microenvironment in addition to the rapid xenograft growth and the absence of an immune response (49). Despite the rapid hepatic elimination of adenovirus in mice, we confirmed that intravenous delivery of Ad5-3D-A20T resulted in high levels of viral gene expression in Suit-2 tumors.…”

An oncolytic adenovirus defective in pRb-binding (dl922–947) can efficiently eliminate pancreatic cancer cells and tumors in vivo in combination with 5-FU or gemcitabine

“…46,47 We speculate that early viral E1A expression was responsible for most of the synergistic effects based on the fact that E1A is a potent apoptosis inducer and synergizes with apoptosis-inducing cytotoxic drugs. [34][35][36]48 Adenoviral E1A proteins can interact with numerous cellular effector molecules such as tumor necrosis factora, NO, TNF-related apoptosis-inducing ligand, Fas ligand (reviewed in ref. 49 ) and immune cells, including NK cells, macrophages and T cells, promoting apoptosis.…”

“…Potent E1A expression was a requirement even in cells not supporting viral replication, in agreement with earlier reports. [34][35][36] Importantly, we demonstrate that E1A with the pRb-binding region deleted in dl922-947 can synergistically interact with 5-FU and gemcitabine in pancreatic cancer cells. Both replicationdependent and -independent mechanisms were indicated in the synergistic responses.…”

“…Synergy was determined by construction of isobolograms and calculation of combination indices (CIs) as described previously. 31,36,37 The interactions were defined as synergistic when CI values were less than the theoretical additive value, with data points below the line of additivity and antagonistic for data points above the line. CI values p0.9 indicate synergism, CIX1.1 antagonism and 0.9oCIo1.1 additive effects.…”

“…The concentration of each agent killing 50% of cells (50% effective concentration (EC 50 )) was calculated through the generation of dose-response curves using untreated cells as control as described previously. 35,36 Fivefold dilutions were prepared starting at 1 Â 10 5 particles per cell (ppc) for all viruses and at 400 mM for gemcitabine (Eli Lilly, Basingstoke, Hampshire, UK) and 5-FU (Sigma Chemicals). Each data point was generated from triplicate samples in each assay, repeated 3-5 times.…”