GBT1118 Diminishes Vaso-Occlusion in Sickle Cell Disease Mice

Recurrent, painful vaso-occlusive events are one of the most debilitating complications of sickle cell disease (SCD). The process involves occlusion of blood flow through small vessels by heterogeneous cell clusters comprised of red blood cells (RBCs), neutrophils, and platelets. Activated endothelial cells localize and facilitate vaso-occlusion. The central triggering event in the process is hemoglobin S (HbS) polymerization and the consequent damage to RBCs. Many recent clinical efforts to prevent vaso-occlusive events have focused on downstream events mediated by inflammatory molecules. The present work however studies the triggering event by assessing the direct impact of allosteric modulators of hemoglobin oxygen (Hb-O₂) affinity on vaso-occlusion in a murine model of SCD.

In this study, we used GBT1118 (a structural analog of GBT440) in a modified dorsal skin fold model in an HbSS knock-in mouse model of SCD (Townes; Wu et al., Blood 2006). GBT1118 binds covalently and reversibly via an imine intermediate to the N-terminal valine of the hemoglobin α chain, and allosterically increases the intracellular hemoglobin affinity for oxygen (Dufu et al., Am J Physiol Heart Circ Physiol 2017). The dorsal skin fold chamber model has been used previously to study vaso-occlusion in SCD mice (Kalambur et al., Am J Hematol 2004). In this present study, mice were treated acutely by oral gavage with GBT1118 or vehicle control in a blinded fashion. Two hours after drug treatment, a baseline of vascular patency at ambient air was established using intravital microscopy to count blood vessels with blood flow. Subsequently animals were exposed to 1 h hypoxia (7.0% oxygen) and 1 h reoxygenation at ambient air. At that time the previously observed blood vessels were monitored again, and the percentage of occluded vessels was recorded. We observed a significant reduction of vaso-occlusion in animals that had been treated with GBT1118 (p<0.05).

In summary, while improved survival has been described in hypoxia-challenged SCD mice treated with allosteric modulators of hemoglobin (Abdulmalik et al., Br J Haematol 2005), our study demonstrates for the first time that a hemoglobin allosteric modulator can reduce vaso-occlusion in a mouse model of SCD. The protective effect elicited by GBT1118 may either be attributed to improved oxygenation or allosteric modulation of HbS. Both mechanisms may directly delay hypoxia-triggered polymerization of HbS, sufficiently for RBCs to transit the microcirculation without sickling, without adhering to vessel wall, and therefore without causing vaso-occlusion.

The results of this study expand on the potential mechanisms by which allosteric modulators of Hb-O₂ affinity may contribute to the treatment of SCD, in addition to previously described hematological benefits.