Ad-Delo3-RGD replicates in and kills brain CSC lines similarly to wild type adenovirus. (A) PCR analysis of Ad-Delo3-RGD preparation demonstrates the absence of E1A13S sequences and for this the absence of RCA in purified Ad-Delo3-RGD. (B) PCR-based assessment of virus replication of an E1A-deleted adenovirus (dl703), Ad-Delo3-RGD or Ad-WT in R11, R28 and R40 CSC. Ad-Delo3-RGD replicates around 130 fold (R11) or 345 fold (R28) better in CSC than dl703. In R40 brain CSC line, Ad-Delo3-RGD replicates around 620 fold better than dl703, and even 4.3 fold better than Ad-WT. Viral DNA for PCR amplification was isolated from infected cells 4 h (white bars) or 72 h (grey bars) post infection. Values are given as relation of adenoviral fiber DNA copies to β-actin copies. Columns represent the mean of 3 measurements (C) Tumorspheres of brain CSC lines are destroyed by the YB-1 dependent adenovirus Ad-Delo3-RGD. Brain CSCs were seeded in 6-well plates and 24 h later infected with 50 MOI of the indicated virus, or were treated with 100 μM TMZ. 5–7 days later morphology was studied microscopically (magnification = 100 x). Control: untreated CSC spheres.