Cisplatin-induced GADD34 upregulation potentiates oncolytic viral therapy in the treatment of malignant pleural mesothelioma

Background: NV1066, a replication-competent oncolytic herpes simplex virus type 1 (HSV-1) attenuated by a deletion in the gene gamma(1)34.5, preferentially replicates in and kills malignant cells. gamma(1)34.5 encodes ICP34.5, a viral protein essential for productive replication, which has homology with mammalian stress response induced GADD34 (growth arrest and DNA damage-inducible protein). We hypothesized that cisplatin upregulates GADD34 expression, which enhances NV1066 replication and oncolysis. Methods: Ten human malignant pleural mesothelioma (MPM) cell lines were infected with NV1066 at multiplicities of infection (MOI; ratio of viral particles per tumor cell) 0.005 to 0.8 in vitro, with and without cisplatin (1 to 4 mu M). In the MPM cell line VAMT, viral replication was determined by plaque assay, cell kill by lactate dehydrogenase assay, and GADD34 induction by quantitative RT-PCR and Western blot. Synergistic efficacy was confirmed by the isobologram and combination index methods of Chou-Talalay. GADD34 upregulation by cisplatin was inhibited with GADD34 siRNA to further confirm the synergistic efficacy dependence with GADD34. Results: Combination therapy with NV1066 and cisplatin showed strong synergism in epithelioid (H-2452, H-Meso), sarcomatoid (H-2373, H-28), and biphasic (JMN, Meso-9, MSTO-211H) MPM cell lines, and an additive effect in others. In VAMT cells combination therapy enhanced viral replication 4 to 11-fold (p<0.01) and cell kill 2 to 3-fold (p<0.01). Significant dose reductions for both agents (2 to 600-fold) were achieved over a wide range of therapeutic-effect levels (LD50-LD99) without compromising cell kill. Synergistic cytotoxicity correlated with GADD34 upregulation (2 to 4-fold, p<0.01) and was eliminated following transfection with GADD34 siRNA. Conclusion: Cisplatin-induced GADD34 expression selectively enhanced the cytotoxicity of the gamma(1)34.5-deficient oncolytic virus, NV1066. This provides a cellular basis for combination therapy with cisplatin and NV1066 to treat MPM and achieve synergistic efficacy, while minimizing dosage and toxicity.