In vivo antimyeloma effect of Zalypsis. (A) CB17-SCID mice subcutaneously inoculated with 3 × 106 MM1S or OPM1 cells in the right flank were assigned to the different treatment groups receiving Zalypsis at a dose of 0.8 mg/kg and 1 mg/kg intravenously once weekly for 3 doses (↑ indicates days of treatment) or to the control group receiving the vehicle alone. Tumor diameters were measured every other day, and tumor volume was estimated as the volume of an ellipse. (A,B) The evolution of the volume of MM1S and OPM-1 plasmacytomas for the indicated days. Statistical differences in each time point were analyzed for each dose compared with the vehicle control group with one-way analysis of variance and Bonferroni post hoc tests, and statistical significance was defined as P < .05. *First day in which differences were statistically significant for each dose. Bars represent SE. (C,D) For survival evaluation, mice were killed when their tumor diameters reached 2 cm or when they became moribund. Time to endpoint was defined as the time from the day of initiation of treatment to death as a result of toxicity, tumor growth, or any other cause. Statistical differences were analyzed in a Kaplan-Meier curve. *Significance was compared with the vehicle control group, defined as P < .05 in the log rank test. (E) Immunohistochemical analyses were performed with different apoptosis-related markers in selected control and treated tumors after the death of mice bearing MM1S and OPM-1 plasmacytomas. Panels (original magnification ×100) show that Zalypsis treatment induced PARP and caspase-3 expression (first 2 rows), an increase of H2AX phosphorylation (third row), and nuclear accumulation of p53 in tumors derived from both cell lines (fourth row), although basal p53 expression was higher in OPM-1 tumors.
