Safety evaluation of DT388IL3, a diphtheria toxin/interleukin 3 fusion protein, in the cynomolgus monkey

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Abstract
We developed a fusion toxin, DT388IL3, consisting of the catalytic and translocation domains of diphtheria toxin (DT388) linked to interleukin 3 (IL3) for the treatment of patients with acute myeloid leukemia (AML). Our goal in this study was to estimate a range for the maximum tolerated dose (MTD) and to evaluate the dose-limiting toxicity (DLT) of DT388IL3 in cynomolgus monkeys (Macaca fasicularis), which possess cross-reactive IL3 receptors. In our previous study, we administered up to six infusions of DT388IL3 at 40, 60, or 100 μg/kg every other day to three pairs (one male monkey and one female monkey) of young adult monkeys. In five of six monkeys, results showed a dose-dependent increase in malaise and anorexia but no consistent abnormalities in serum chemistries or blood counts. There was no evidence of organ damage by blood tests or histopathology. However, the female treated at 100 μg/kg, died of moderate to severe vasculitis of multiple tissues. Based on these findings, this study repeated the 100 μg/kg group and added a group that received 150 μg/kg in an effort to confirm a dose response. Two female monkeys were treated with up to six infusions of DT388IL3 at 100 μg/kg or 150 μg/kg every other day. One additional female monkey was treated as a negative control. Monkeys in the 100 μg/kg group showed moderate malaise and anorexia, but no consistent abnormalities in blood counts or serum chemistries. Moderate elevations of liver enzymes were noted in the 150 μg/kg group in addition to severe malaise and anorexia. No significant findings were revealed at gross necropsy. The histopathological findings revealed regenerative myeloid hyperplasia and hepatic degeneration and regeneration in the 150 μg/kg group. Similar lesions of less severity were detected in the 100 μg/kg group. DT388IL3 plasma half-life was approximately 20 min with a peak concentration of approximately 2 μg/ml (30,000 pM). The IC50 for AML blasts in vitro was 6 pM. Collectively, our results suggest that DT388IL3 can be tolerated at doses up to 100 μg/kg in a nonhuman primate, which is higher than previously reported for other AML directed diphtheria toxin fusion proteins, and should in principle allow for dose escalation with reduced toxic side effects. Based on these findings a phase I clinical trial has recently been initiated with DT388IL3 for the treatment of AML.