Effects of N-Acetylcysteine in Patients with Sickle Cell Disease

We previously showed that the plasma of patients with sickle cell disease (SCD) contains an elevated quantity of von Willebrand factor (VWF) with high specific adhesive activity (as measured by the binding of nanobody AU/VWFa-11). Total active VWF (VWF antigen multiplied by VWF specific activity) correlated with the extent of hemolysis in these patients (Chen J. et al., Blood, 2011, 117:3680). In another study, we showed that N-acetylcysteine (NAC) reduced VWF size and activity ex vivo and broke down platelet–VWF aggregates in vivo in mice (Chen J. et al, JCI, 2011, 121:522). Given the possibility that VWF is involved in the pathophysiology of sickle cell disease, we are examining whether NAC can benefit these patients. We are first conducting a pilot clinical trial to determine the safety profile of infused NAC and its effects on laboratory endpoints. Five SCD patients at disease baseline and not recently transfused will be enrolled. To date, two patients have completed intravenous infusion of NAC at a low dose of 150 mg/kg with a bolus infusion of 75 mg/kg for the 1^st hr and 75 mg/kg for 7 hr. Approximately one month after the low-dose infusion, these patients received another infusion of 300 mg/kg given as a bolus infusion of 150 mg/kg for the 1^st hr and 150 mg/kg for 7 hr. Blood was collected for analysis immediately before the infusion, at 1 hr (after bolus infusion), 8 hr, 24 hr, and 72 hr after infusion. Among the parameters examined were plasma VWF, red blood cell concentration, density, and size (to look for fragments), and the concentrations of NAC, cysteine and glutathione and their oxidized and mixed disulfide forms. In these two patients, the NAC infusion was well tolerated except that both patients experienced pruritus during the higher-dose bolus infusion. We measured the concentrations of reduced and oxidized NAC, cysteine, and glutathione in whole blood in the first study subject by mass spectrometry. NAC concentrations were 725 μM and 1.58 mM at the 1 hr time point at the low and high doses, respectively. Compared to the baseline (before NAC infusion), the concentration of total cysteine in blood was increased 2.4 fold for low dose and 2.9 fold for high dose at 1 hr and returned to baseline at 8 hr. The concentration of total glutathione in whole blood was increased 1.5 fold and 1.3 fold at 24, and 72 hr, respectively, for the high dose infusion but did not change much at the low dose infusion. The size of high molecular weight VWF multimers decreased with the high dose infusion, the effect being obvious at 1 hr. In addition, NAC infusion markedly reduced the concentration of red cell fragments and dense cells. Both of these effects were very rapid, being observable at the 1 hr time point. In summary, NAC infusion in sickle cell patients at disease baseline appears safe. NAC increases the concentrations of total cysteine and glutathione in blood, reduces high molecular weight VWF multimers, and decreases the number of dense red blood cells and the extent of red cell fragmentation.