Targeting Pro-Oxidant Iron with Deferoxamine as a Treatment for Ischemic Stroke: Safety and Optimal Dose Selection in a Randomized Clinical Trial

Abstract

A role of iron as a target to prevent stroke-induced neurodegeneration has been recently revisited due to new evidence showing that ferroptosis inhibitors are protective in experimental ischemic stroke and might be therapeutic in other neurodegenerative brain pathologies. Ferroptosis is a new form of programmed cell death attributed to an overwhelming lipidic peroxidation due to excessive free iron and reactive oxygen species (ROS). This study aims to evaluate the safety and tolerability and to explore the therapeutic efficacy of the iron chelator and antioxidant deferoxamine mesylate (DFO) in ischemic stroke patients. Administration of placebo or a single DFO bolus followed by a 72 h continuous infusion of three escalating doses was initiated during the tPA infusion, and the impact on blood transferrin iron was determined. Primary endpoint was safety and tolerability, and secondary endpoint was good clinical outcome (clinicalTrials.gov NCT00777140). DFO was found safe as adverse effects were not different between placebo and DFO arms. DFO (40-60 mg/Kg/day) reduced the iron saturation of blood transferrin. A trend to efficacy was observed in patients with moderate-severe ischemic stroke (NIHSS > 7) treated with DFO 40-60 mg/Kg/day. A good outcome was observed at day 90 in 31% of placebo vs. 50-58% of the 40-60 mg/Kg/day DFO-treated patients.

Keywords: antioxidant; deferoxamine; ferroptosis; iron; neuroprotection; outcome.

Conflict of interest statement

The authors declare that there is no conflict of interest regarding the publication of this paper.

Figures

Figure 1

Inclusion and exclusion criteria and CONSORT flow diagram of the three-dose tier sub-studies (DTS). In each DTS early termination cases due to mortality, discontinuation due to serious adverse events (SAE) possibly related to treatment, and patients excluded are indicated.

Figure 2

Deferoxamine (DFO) reduces TSAT time- and dose-dependently. (A) Time-course of serum DFO levels along the infusion in AIS patients treated with a 10 mg/Kg bolus of DFO IV followed by a 72 h continuous IV infusion of DFO in escalating dose tiers of 20, 40, or 60 mg/Kg/day (blood DFO levels at time = 0 of infusion in this graph are the result of the initial previous 10 mg/Kg bolus of DFO IV). Mean ± SD are shown; no significant effects were found (repeated measures ANOVA). (B) U-PAGE/WB depicting the bands of the iron-devoid form of human transferrin standard (Std) (apotransferrin, ATf) and human diferric transferrin (diFe-Tf) standard (holotransferrin, HTf). The iron load of transferrin determines the electrophoretic mobility of the different Tf forms in these urea gels. Arrows indicate the different electrophoretic pattern of ATf, the two monoferric forms of transferrin (mFe-Tf), and the diferric transferrin (diFe-Tf) form in serum samples of stroke patients. Placebo and DFO depict bands of transferrin of two representative patients (one of the placebo group and one of the DFO 60 group) before the onset of treatment (0), and 24 and 72 h after administration. In each lane, optical density of the bands allow calculation of the % TSAT for a given patient at a given time point using the formula: TSAT (%) = (0.5 *mFe·Tf + diFe·Tf)*100/(ATf + mFe·Tf + diFe·Tf). (C–F) % TSAT before the onset of treatment (0 h), and 24 and 72 h after administration of placebo (C), 20 mg/Kg/day DFO (D), 40 mg/Kg/day DFO (E), or 60 mg/Kg/day DFO (F). Values are presented as mean ± SD. * p ≤ 0.05, ** p ≤ 0.005 (repeated measures one-way ANOVA plus the post-hoc Benjamini–Krieger–Yekutieli test).

Figure 3

Exploratory analysis to examine whether the TSAT-modifier deferoxamine (DFO) doses favors a good outcome. (A) Median and quartiles of baseline NIHSS in a subpopulation of the TANDEM-1 study with NIHSS at admission > 7. Baseline NIHSS were found balanced between the four treatment groups when considering this patient subpopulation (NIHSS > 7) (p = 0.24, one-way ANOVA). (B) In this subpopulation (NIHSS > 7) we calculated the percentage of neurological improvement with the formula: % reduction of NIHSS at 90 days = ((NIHSS at admission-NIHSS at a given time)*100/NIHSS at admission). As DFO 40 and DFO 60, but not DFO 20, reduce TSAT, these two groups were pooled and compared to the placebo group. We observed that half of the patients in the 40 + 60 DFO group showed a 100% reduction of their initial neurological impairment, in contrast to those in the placebo group (p = 0.0546, Mann–Whitney U test). (C) Graph depicting the proportion of AIS patients showing good functional outcome (in black) in the placebo and DFO groups. DFO dose tiers of 40 and 60 mg/Kg/day have higher proportion of AIS classified as good outcome patients (mRS ≤ 2) at 7 and 90 days. Values are presented as median and quartiles and compared with one-way ANOVA (A) or Mann–Whitney U test (B).