Pre-Treatment Steady-State Rheology Predicts Response to Hydroxyurea in Patients with Sickle Cell Disease
Authors: Baldan Alessandro , Summerall Carly Ginter , James Loretta , Dinu Bogdan R. , Flanagan Jonathan M. , Sheehan Vivien A. 
Published: October 2019

Abstract
Background: Patients with sickle cell disease (SCD) have altered blood rheology due to erythrocyte abnormalities, such as increased aggregation and RBC density. The hematocrit to viscosity ratio (HVR) is a measure of RBC oxygen carrying capacity, and is reduced in SCD. Higher percent dense red blood cells (%DRBC) and lower HVR have been associated with higher rates of disease related complications. The aim of this study was to elucidate the effects of hydroxyurea (HU), fetal hemoglobin (HbF) and transfusion therapy (TF) on the rheology of patients with SCD.
Methods: We measured whole blood viscosity of samples from children with SCD at low shear (45 s-1) and high shear (225 s-1) using a Brookfield cone and plate viscometer under oxygenated conditions. Venous blood samples in EDTA were analyzed within 4 hours of phlebotomy. Laboratory values were obtained with a Siemens ADVIA120 Hematology System; Hb profiles by high performance liquid chromatography. Samples were collected at routine clinic visits from SCD patients prior to initiation of therapy (n=105 for HbSS, n=77 for HbSC), then on HU at MTD, (n=30 for HbSS, n=13 for HbSC), or at least one week after TF (n=32). A paired T-test was used to assess treatment changes; correlation/trend tests were used to test association of hematological and rheology parameters. Results: Blood viscosities at low and high shear rates were significantly higher in untreated patients with HbSC compared to untreated patients with HbSS (p<0.001), while HVR was similar between patients with HbSC or HbSS. High endogenous HbF levels were significantly associated with higher HVR 225 s-1 in patients with HbSS but not HbSC. Patients with HbSS after TF had a significant improvement (rise) in their HVR at high shear (p=0.04) but not at low shear. This TF mediated rise in HVR 225 s-1 correlated with increased Hb (r=0.56, p<0.002), and decreased WBC and ANC (r= - 0.46 and r=- 0.36 respectively, p<0.05). Patients with greatest decreases in %HbS had most improvement in their HVR 225 s-1 (r = -0.36, p=0.15). HbSS patients treated with HU also had a rise in their HVR 225 s-1 (p=0.01) but not HVR 45 s-1. This improved HVR 225 s-1 was associated with typical HU induced changes in Hb, Hct, HbF and decreases in WBC and ANC.
For HbSS patients treated with either HU or TF, a baseline HVR (at either shear rate) below the group mean experienced a greater rise in HVR, while a baseline HVR above the mean experienced a decline (p<0.001; Figure 1). HbSS patients with a pre-treatment %DRBC above the mean experienced a decline in %DRBC on HU at MTD, while those with a %DRBC below the mean did not (p=0.001).A similar trend was seen in HbSC patients treated with HU to MTD, but did not reach statistical significance.
Patients with HbSC had significantly higher %DRBC compared to patients with HbSS (p<0.001). HbSC patients placed on HU had significantly higher baseline %DRBC than HbSC patients not treated with HU (15.8 ±13 versus 8.2± 6.5, p=0.001). However, HbSC patients treated to HU MTD did not experience a significant reduction in %DRBC. Individuals with HbSS and high endogenous HbF had significantly lower %DRBC (r= -0.36, p=0.002), and greater production of HbF on HU also resulted in lower %DRBC (p=0.036) in HbSS (n=25), but not HbSC patients (n=13).
Conclusions: In HbSS patients, high endogenous HbF levels were significantly associated with higher HVR at high shear and with lower %DRBC, but not in HbSC patients. In HbSS patients, HU and TF therapy improved HVR at high shear rates but not low shear, possibly indicating persistent red cell adhesion at low shear rates. This improvement correlated with HbF levels in HU treated patients, and HbS levels in TF treated patients. Patients with SCD and greater than average impairment of whole blood rheology at pre-treatment steady-state experienced greater rheological improvement, demonstrated clinically by a reduction in %DRBC or a rise in HVR on HU. A cohort of 13 HbSC patients treated prospectively with HU did not demonstrate statistically significant rheological improvement, either a result of small sample size or the minimal rise in HbF despite HU therapy.
Figure 1. Effect of baseline rheology on rheological changes at MTD of HU. Horizontal dotted line denotes mean baseline values. A. Patients with HbSS and a baseline HVR 45 s-1 below the mean showed an increase in HVR 45 s-1 at MTD (green dots). B. Patients with HbSS and a baseline HVR 225 s-1 below the mean showed an increase in HVR 225 s-1 at MTD (green dots). Figure 1. Effect of baseline rheology on rheological changes at MTD of HU. Horizontal dotted line denotes mean baseline values. A. Patients with HbSS and a baseline HVR 45 s-1 below the mean showed an increase in HVR 45 s-1 at MTD (green dots). B. Patients with HbSS and a baseline HVR 225 s-1 below the mean showed an increase in HVR 225 s-1 at MTD (green dots).

Disclosures
No relevant conflicts of interest to declare.