Fig 1: Combination of reovirus and CD3-bispecific antibodies (bsAbs) is effective in a human, orthotopic human epidermal growth factor receptor (HER2)+ breast cancer model. (A) HER2 expression percentages on BT474 cells, as analyzed by flow cytometry using a two-step protocol. (B) Number of reovirus genomic segment 4 (S4) copies in BT474 cells after reovirus infection. BT474 cells (200,000/well) were infected with reovirus multiplicities of infection (MOI) 10 or phosphate-buffered saline (PBS) (Mock) as a control. Samples (n=3) were harvested 24 hours after infection and the number of viral S4 copies was determined by quantitative reverse transcription PCR. (C) Design of experiment described in (D and E). Mice (n=6/group) with established BT474 tumors were intravenously (i.v.) injected with 5×106 human PBMCs, and thereafter intratumorally (i.t.) injected with reovirus (107 plaque-forming units (pfu)) on 2 consecutive days. After 4 days, mice received intraperitoneal (i.p.) injections of 12.5 µg CD3xHER2 bsAbs (CD3xHER2) or PBS as control. (D) Individual growth curves of BT474-bearing mice receiving indicated treatments. Lines indicate timing of injection with PBMCs (orange), reovirus (blue) or CD3xHER2 (red). (E) Average relative changes (±SEM) in tumor volume from start of CD3xHER2 bsAb treatment. Significance versus PBS on day 42 was calculated using one-way analysis of variance with Dunnett’s post hoc test. Significance level: **p<0.01.