Figure 5.

Effect of TMZ on OAd-mediated CPE in murine triple negative breast cancer cells. 4T1 Murine breast cancer cells were no treated (Mock) or treated with Adhz60 and TMZ at the following doses for Adhz60 and TMZ, respectively: (10 MOI and 400 µM). AdLacZ was used at 10 MOI. DMSO was added as a control. (A) Cytopathic effect was evaluated by crystal violet staining at 72 hours following treatment and reported as percentage of control (mock). (B) Cell viability was calculated by measuring the absorbance of solubilized dye at 590 nm. Results represent the mean of three repeated measurements ± standard deviation (SD; error bars) (*P < 0.05). (C) Seventy-two hours post-treatment, supernatants were collected and used to determine virus adenovirus yield from each cell line. Results represent the mean of three independent experiments ± standard deviation (SD; error bars) (**P < 0.01). (D) Whole cell protein lysates were collected 72h after indicated treatment. Western blot and bar graphs of LC3-I and II expressions. Bars represent mean ± SEM expressed as percentage of change from 3 separate experiments, (*P < 0.05) decrease in the level of LC3-I expression. Actin was used as a loading control. (E) Cells were stained with annexin V-PE and 7-aminoactinomycin D (7-AAD). Positive cells for annexin V-PE and 7-AAD staining were analyzed by FACScan flow cytometer. (F) Results represent the mean of three independent experiments ± standard deviation (SD; error bars) (*P < 0.05). (G) Whole cell protein lysates were collected 72h after indicated treatment. Expression of cleaved caspase-3 was detected by western blot; actin was used as a loading control. A representative experiment is shown from three performed.