Oncolytic viruses represent a novel
cancer treatment strategy. Despite their promising preclinical data, however, corresponding clinical trials have disappointed. To aid preclinical analyses, we hypothesized that three-dimensional
tumor cell clusters or spheroids might provide an assay system superior to conventional monolayer cell cultures. Spheroids show
viral infection, replication and oncolytic patterns distinct from conventional monolayer assays. Therefore, viral
tumor penetration and oncolysis measurements may be improved with such three-dimensional models. Also, preclinical analyses of oncolytic viruses frequently measure mitochondrial activity, but more accurate measures of oncolysis might involve quantitation of intracellular
protein release. Therefore, we measured
luciferase released from
luciferase-expressing spheroids and found unique patterns that maintained consistency with various viruses and doses. The relative variations between viruses and doses may represent temporal differences in oncolysis dynamics. Analysis of five recombinant replicative adenoviruses with promise for clinical application showed that Ad5/3-Delta24 produced the most
luciferase release 1 week after
infection and achieved the earliest and highest peak
luciferase release level. Ad5/3-Delta24 also effected the earliest subtotal spheroid cell death. These findings closely parallel monolayer oncolysis assays with these agents. Therefore, the
luciferase-expressing
tumor spheroid assay represents a promising three-dimensional model for preclinical analysis of replicative oncolytic agents.