GBT440 Demonstrates High Specificity for Red Blood Cells in Nonclinical Species

Abstract

Sickle cell disease (SCD) is caused by a point mutation in the β-globin gene leading to production of hemoglobin S (HbS) that polymerizes upon deoxygenation with subsequent formation of sickled red blood cells (RBCs). GBT440 modulates O2 affinity of hemoglobin (Hb) by binding to the N-terminal α chain of Hb via a reversible Schiff base. We previously demonstrated that GBT440 preventedsickling of RBCs from sickle cell patients, in vitro. Also, in a murine model of sickle cell disease (Townes SS mice), GBT440 prevented ex vivo sickling of RBCs and prolonged RBC half-life. Pharmacokinetic (PK) studies of GBT440 were conducted in mouse, rat, dog and monkey following IV and oral administration. Both blood and plasma samples were collected and assayed for GBT440 concentration using LCMS. Following IV and oral administrations, GBT440 quickly partitions into the RBC with a high blood/plasma ratio of ~70:1 which corresponded to a RBC/plasma ratio of ~150:1. Volume of distribution (Vss) was small in whole blood (0.041-0.171 L/kg) but much larger in plasma (1.44-8.45 L/kg) indicating that RBCs are a reservoir of GBT440. Systemic clearance (CLs) was low in both blood (0.016-0.113 mL/min/kg) and plasma (0.943-3.16 mL/min/kg) indicating that GBT440 was mostly bound to hemoglobin and only a small fraction of unbound GBT440 re-distributed into the plasma and was available for clearance. Terminal elimination half-life (t1/2) was similar between whole blood and plasma for each species and was long, ranging from 6.4 hours in mouse plasma to 93.5 hours in dog plasma. GBT440 was well absorbed and absolute oral bioavailability ranged from 33% to 70% in four species. A quantitative whole body autoradiography study to determine tissue distribution of GBT440 was conducted in male rats following an oral dose of 14C-GBT440 (10 mg/kg; 150 µCi/kg PO). The data showed that GBT440 is co-located in hematopoietic tissues as expected for a molecule whose target is hemoglobin, including blood, spleen, liver and bone marrow. A mass balance study of 14C-GBT440 (10 mg/kg; 150 µCi/kg PO) was conducted in rats to determine route of elimination of GBT440. The 14C-GBT440-derived radioactivity was well absorbed and rapidly excreted after oral administration. By 240 hours postdose, mean values of 79.0 ± 3.86 and 9.74 ± 3.02% of the administered radioactivity were excreted in feces and urine, respectively. The mean overall recovery of radioactivity was 92.4 ± 0.875%. Metabolism via both Phase I and Phase II pathways was the major route of elimination of GBT440. These data indicate that despite its high affinity binding toward Hb, GBT440 could be released from the hemoglobin complex and completely eliminated from the body. To further correlate PK to pharmacological activity (hemoglobin modification based on changes in the oxygen equilibrium curve), mice were given an oral dose of 30, 50 and 100 mg/kg and blood were collected at 4 and 6 hr postdose for hemoximetry analysis. Data showed good correlation between blood concentrations and changes in p50. Blood concentrations following 30, 50 and 100 mg/kg at 4 hr were 243, 446, and 806 µM, which resulted in changes in p50 of 11%, 25% and 55%, respectively, indicating that GBT440 elicits an ex vivo dose dependent increase in Hb-O2 affinity following increasing dosage to mice. Based on PK data from 4 animal species, PK profile of GBT440 in human was predicted using a simple allometric scaling technique. The predicted PK profile following an oral administration was highly concordant with actual data from healthy subjects for Cmax, AUC and T½, which suggested that the disposition kinetics of GBT440 in humans were consistent to that in animals. In summary, nonclinical PK studies support previous findings that GBT440 partitions to RBCs, binds specifically to Hb with a slow off rate, modulates Hb-O2 affinity, and is completely eliminated from the body. GBT440 is in clinical trials for the treatment of SCD. Disclosures Hutchaleelaha: Global Blood Therapeutics: Employment, Equity Ownership. Patel:Global Blood Therapeutics: Employment, Equity Ownership. Silva:Global Blood Therapeutics: Employment, Equity Ownership. Oksenberg:Global Blood Therapeutics: Employment, Equity Ownership. Metcalf:Global Blood Therapeutics: Consultancy, Equity Ownership.

Voxelotor (GBT440), a first‐in‐class hemoglobin oxygen‐affinity modulator, has promising and reassuring preclinical and clinical data

“…In in vitro studies, voxelotor modified HbS (voxelotor‐HbS) maintains an oxyhemoglobin state under hypoxic conditions and delays polymerization of unmodified HbS and imparts similar delay times as fetal hemoglobin (HbF) (Figure ) . The Hebbel model assumed a left shift of the p50 to 18 mm Hg for 30% voxelotor‐HbS, however, the p50 at 1000 mg per day is more modest than this at 28 mm Hg with 27% modification . In adults with SCD who received 500 mg ( n = 10) or 700 mg of voxelotor for 28 days, irreversibly sickled cells were reduced, and five individuals continued 700 mg for 90 days and had a sustained reduction in irreversibly sickled cells and irreversibly sickled cells present at 28 days was sustained (Figure )…”

New developments in anti‐sickling agents: can drugs directly prevent the polymerization of sickle haemoglobin in vivo?

“…One series of compounds includes GTx011 (or GBT440). Although publicly reported details in the literature are limited, GTx011 also appears to be an orally bioavailable small molecule that modifies haemoglobin oxygen affinity by binding to the N‐terminal α‐chain of Hb and forming a reversible Schiff base (Hutchaleelaha et al , ). Binding of the compound is also proposed to allosterically influence the intra‐dimer interface of Hb (Hisα122 and Hisα103) and the distal valine surrounding haem pockets of both the α and β chains (Patel et al , ).…”

“…It has been reported to be a potent and direct anti‐sickling agent with high specificity for Hb; estimated Hb modification of 10–30% was safe in animal studies and effective at preventing Hb S polymerization. Pharmacokinetics studies conducted in four animal species (mouse, rat, dog, monkey) demonstrated that GBT440 is well absorbed following IV and oral administration, quickly partitions in to the RBC with a small fraction re‐distributed into the plasma (Hutchaleelaha et al , ). In male rats, it distributes into Hb, blood, spleen, liver and bone marrow (Hutchaleelaha et al , ).…”

“…Pharmacokinetics studies conducted in four animal species (mouse, rat, dog, monkey) demonstrated that GBT440 is well absorbed following IV and oral administration, quickly partitions in to the RBC with a small fraction re‐distributed into the plasma (Hutchaleelaha et al , ). In male rats, it distributes into Hb, blood, spleen, liver and bone marrow (Hutchaleelaha et al , ). Despite its high affinity binding toward Hb, it could be completely released and eliminated in faeces and urine; the major route of elimination was via both Phase I and Phase II pathways (Hutchaleelaha et al , ).…”

“…In male rats, it distributes into Hb, blood, spleen, liver and bone marrow (Hutchaleelaha et al , ). Despite its high affinity binding toward Hb, it could be completely released and eliminated in faeces and urine; the major route of elimination was via both Phase I and Phase II pathways (Hutchaleelaha et al , ). There is good correlation between blood concentration and changes in P 50 , eliciting an ex vivo dose‐dependent increase in Hb oxygen affinity following increasing dosage in mice (Hutchaleelaha et al , ).…”