8-NH₂-Ado promotes cell death and inhibits cell growth but with only minor cell-cycle effects in MCL cell lines. (A) Cell death after 24 hours of continuous incubation was quantified by loss of mitochondrial membrane potential as measured by DiOC₆ staining, at various concentrations of 8-NH₂-Ado. For each cell line, the dose-response curve was estimated by nonlinear regression using a variable Hill Slope model. The EC₅₀ was defined as the drug concentration required to achieve 50% of the maximum response. The standard errors of the estimated EC₅₀ values for JeKo, Mino, and SP-53 were less than 3%; the EC₅₀ value could not be accurately estimated for Granta 519. (B) Cell death was also evaluated by PARP cleavage. The arrow denotes cleaved PARP. The cells were continuously incubated with 3μM 8-NH₂-Ado for 24 hours. (C) The growth inhibition of MCL cells was determined after 24 hours of continuous exposure to multiple concentrations of 8-NH₂-Ado. The cell concentrations were quantified using a particle count and size analyzer (Beckman Coulter). The IC₅₀ values were estimated by nonlinear regression using a variable Hill Slope model. The IC₅₀ value was defined as the drug concentration required to achieve 50% of the maximum growth inhibition. The standard errors for the IC₅₀ value estimates were less than 8% for all cell lines. (D) The cell-cycle effects were determined by flow cytometry for JeKo and Granta 519 after continuous treatment with 3μM 8-NH₂-Ado at various time points up to 24 hours. Independent experiments were performed in triplicate shown with SD error bars.