Effect of Intravenous and Oral N-Acetylcysteine Treatment in a Patient with Sickle Cell Disease at Disease Baseline
Authors: Ozpolat Hasan Tahsin , Chen Junmei , Fu Xiaoyun , Osborn Warren , Le Jennie , Ling Minhua , Schmitter Sophie , Ruuska Sarah , Chung Dominic W. , Konkle Barbara A , López José A. 
Published: October 2019

Abstract
Background: We previously showed that patients with sickle cell disease (SCD) have elevated levels of von Willebrand factor (VWF) with high adhesive activity and the level of total active VWF correlated with hemolysis at disease baseline. These findings suggest that VWF plays a role in the pathophysiology of SCD [Chen et al, Blood. 2011; 117:3680‐3]. We also showed that N-acetylcysteine (NAC), a mucolytic and antioxidant drug, can reduce the level of large VWF multimers and platelet adhesion in human plasma in vitro and in mice in vivo [Chen et al, JCI, 2011, 121:522]. Based on these findings, we hypothesized that administration of NAC in SCD patients might reduce VWF multimers in vivo and reduce oxidative stress by scavenging reactive oxygen species (ROS). We started a pilot study to evaluate the effects of intravenous (IV) and oral NAC administration in SCD patients at disease baseline (NCT01800526). In this report, we administered NAC to one patient by IV and orally and compared the effect on biomarkers by the two routes of NAC administration.
Patient and protocol: The subject is a 51-year-old male with SS genotype, a history of leg ulcers and acute chest syndrome, and who was not taking hydroxyurea at the time of the study. NAC was infused at a low dose (150 mg/kg over 8 hours) and infusion was repeated at high dose (300 mg/kg over 8 hrs) after 4 weeks. Blood samples were collected before (0), during (4 hr) and after the NAC infusions (8, 24 and 72 hr). Five months after the high dose IV infusion, during which parameters returned to baseline levels, oral NAC was administered for 4 weeks at 2.4g/day. Weekly blood samples were collected four weeks before the start of oral NAC, during oral NAC, and one week after oral NAC administration. We measured RBC parameters and RBC fragments, dense cells and percentage of platelet-monocyte complexes (PMC) by flow cytometry. We also analyzed VWF antigen, VWF multimers, ADAMTS13 antigen and activity levels in plasma. We also measured total, oxidized and reduced forms of NAC, cysteine and glutathione levels in plasma and whole blood by mass spectrometry.
Results: 1) The subject did not experience adverse side effect during and after the treatment period. 2) RBC, hemoglobin and hematocrit levels did not significantly change with either IV or oral NAC treatment. Oral and IV NAC decreased dense cell and RBC fragments. NAC reduced the number of dense cells 13% during low dose IV, 79% during high dose IV and 73% with the oral administration. Compared to baseline, RBC fragments decreased 40% during low dose IV, 28% during high dose IV, and 27% with the oral NAC treatment. 3) NAC reduced formation of PMC from 71% to 24% (low dose IV), from 72% to 35% (high dose IV), and from 62% to 27% (oral). 4) VWF multimer size significantly decreased during low and high dose NAC infusions but did not change with oral NAC administration. VWF antigen and ADAMTS13 activity levels did not change with either routes of NAC administration. 5) The concentration of NAC in whole blood increased during treatments: elevated to 0.5 mM and 1 mM at the end of low- and high-dose IV infusions, respectively. At the conclusion of oral administration, the NAC concentration was 4 µM. Total glutathione concentrations in whole blood did not change significantly during and after NAC treatment at either dose or by either route. Free Cys concentrations in plasma increased compared to respective baseline levels 10-fold for low dose infusion (from 10 µM to 101 µM), 20-fold for high dose infusion (from 9 µM to 200 µM), but the level did not change significantly for oral administration (from 5 µM to 7 µM). Protein-bound Cys levels in plasma as markers of oxidative stress were significantly decreased with low- and high-dose IV infusion. However, the level did not change significantly with oral NAC administration.
Summary: Both IV and oral administration of NAC were safe and well tolerated in the patient with SCD. Both IV and oral administration reduced the number of dense cells, RBC fragmentation, and formation of PMCs. IV, but not oral NAC, reduced VWF multimer size. These finding suggest that IV administration resulted in higher concentration and had more biologic effects; however both approaches may have a role in treatment of patients with SCD.

Disclosures
Konkle: Sangamo: Research Funding; Gilead: Consultancy; Shire: Research Funding; Genentech: Consultancy; CSL Behring: Consultancy; Bioverativ: Research Funding; BioMarin: Consultancy; Spark: Consultancy, Research Funding; Pfizer: Research Funding.