Voxelotor treatment of a patient with sickle cell disease and very severe anemia
Authors: Arun S. Shet, Laurel Mendelsohn, Julia Harper, David Ostrowski, Eric R. Henry, Eveline Gwaabe, Jim Nichols, Abdu I. Alayash, William A. Eaton, Swee Lay Thein
Journal: American Journal of Hematology (2018)

The case involves a 38-year-old female of Nigerian descent with sickle cell anemia (SCA, βS/βS genotype), no α-globin gene deletion and no glucose 6-phosphate dehydrogenase deficiency. Over the past two decades, the patient received routine care at the National Institutes of Health (NIH) participating in several studies. During childhood, the patient's disease course was mild but after 24 years of age, she developed numerous sickle-related complications including osteomyelitis, acute chest syndrome (ACS), multi-organ system failure, chronic renal insufficiency, cholelithiasis, secondary iron overload, cardiomyopathy, and sickle hepatopathy. SCA specific therapies during this period included hydroxyurea (HU, between 2001 and 2009) with minimal clinical or hemoglobin improvement. Decitabine therapy was attempted to improve her hemoglobin but had to be discontinued largely due to thrombocytosis. A phase I trial of the anti-sickling agent AES-103 was also discontinued after 3 months due to unexplained aplastic crisis. Empiric trials of oral magnesium and omega-3 fatty acids provided no measurable clinical benefit. In February 2014, she was admitted for acute painful crisis that triggered ACS and multi-organ failure requiring hemodialysis for approximately 4 weeks and transfusion with 8 units of packed red blood cells (PRBCs, four with exchange transfusion and two separate simple transfusions of two units each). In 2015, she began a simple red-cell transfusion program with extended phenotyped blood for worsening anemia with a transfusion trigger set at <5 g/dL. In June 2016, 10 days after receiving two units of PRBCs, she was admitted with severe acute vasoocclusive pain crisis. A diagnosis of delayed hemolytic transfusion reaction was made on the basis of the acute posttransfusion hemoglobin drop accompanied by a marked increase in intravascular hemolysis markers and a positive direct Coombs test. Severe anemia prompted a retrial of HU that had to be discontinued again as she could not tolerate a dose greater than 300 mg/day (6 mg/kg). Concomitant erythropoietin not only failed to augment her HU response but triggered vaso-occlusive crises.1 In January 2017, deterioration in cardiac function mandated maintaining a hemoglobin >7 g/dL; only least incompatible PRBCs could be provided due to presence of a RBC autoantibody. However, hemoglobin increments obtained following transfusion were transient (lasting on average <10 days) and complicated by reactions (mostly intractable rigors) and acute pain. Transfusion under the cover of immunoglobulins and high-dose steroids did not impact the duration of hemoglobin increment and the patient declined rituximab. At this time, phase I studies of Voxelotor reported hemoglobin increments by ~1-2 g/dL and a phase III trial evaluating the utility of this drug in SCA patients with anemia is ongoing (NCT03036813; NCT02850406).2 Voxelotor is believed to inhibit sickling in vivo by preferential binding to the high-oxygen affinity R quaternary conformation of hemoglobin, thereby shifting the T-R quaternary conformational equilibrium toward R, which cannot enter the sickle fiber.3, 4 As a result, at a given oxygen pressure, the population of the R conformation is increased, the fraction of hemes with oxygen bound is increased, and the sickling frequency is decreased. A consequence of increasing the fraction of the high-affinity R quaternary conformations, is that these molecules deliver very little oxygen to the tissues. However, the net effect on tissue oxygenation is impossible to predict as there are two opposing effects—increased oxygen delivery from less vaso-occlusion if sickling is reduced and decreased oxygen delivery from a left shift in the oxygen dissociation curve.3 In any case, an incremental increase in hemoglobin concentration is not an indication of decreased anemia unless it is greater than the concentration of hemoglobin with drug bound.5 Occupancy of hemoglobin by Voxelotor and formation of an HbS-Voxelotor adduct results in a dose-dependent decrease in the p20, that is, the oxygen pressure at which oxygen is bound to 20% of the hemes. Using the measured Δp20, the percent hemoglobin modified by Voxelotor can be calculated and appears to correlate with percent hemoglobin modified determined by comparing the initial portions of the measured oxygen dissociation curve with the curve prior to starting the drug (day −1) and the curve for which there is 100% occupancy of the drug (see Figures S1 and S2 of Supporting Information).6 Using the Δp20, the median percentage of modified hemoglobin was found to be 25% in 11 SCA patients taking Voxelotor 1000 mg/day on day 28.6 Given the lack of success in treating this patient, we attempted a compassionate basis therapeutic trial of Voxelotor using an FDA approved eIND application.7 The patient began Voxelotor 1500 mg/day and experienced relatively minor side effects (headache and grade II diarrhea) while being adherent to the treatment (determined by pill counts and observation of a split HemoglobinS peak on cation exchange high-performance liquid chromatography).8 The predicted percent modification of hemoglobin at this dosage is 25%-30% (Supporting Information, Figure S2B).6 We used the HEMOX Analyzer (TCS Scientific Corp, New Hope, Pennsylvania) to record blood oxygen dissociation curves and determine the percent hemoglobin modified with Voxelotor. Most measurements at multiple time points were <10% and the maximum percentage hemoglobin modified was 18% (Table 1). These large differences between the precent modified hemoglobin measured and predicted based on drug pharmacokinetics persisted throughout the entire period of drug exposure despite documented adherence. The objective measures and lack of clinical or laboratory improvement led to a trial of split dosing and dose escalation. Neither split dosing nor dose escalation to 1800 g/day resulted in an clinical or laboratory change and the latter was accompanied by transiently elevated serum erythropoietin (Table 1). The drug was subsequently tapered and discontinued. In summary, a therapeutic trial of Voxelotor in this “untransfusable” SCA patient did not increase hemoglobin. However, hemoglobin occupancy failed to reach the 25%-30% percent modification required to be therapeutic in sickle cell patients with less severe anemia.9 When compared with other severe SCD patients treated on a compassionate basis with Voxelotor,7 it is possible that autoantibody mediated hemolysis shortened an already reduced red cell half-life in our patient. As a result, rapid red cell clearance would negatively impact drug pharmacokinetics and account for the low Voxelotor occupancy in our patient although her hemolysis parameters do not overwhelmingly favor this hypothesis (Table 1). Based on our single patient experience, for patients with hemoglobin levels <6 g/dL and rapid red cell turnover, more pharmacologically effective dosage schedules may be required to achieve the desired therapeutic effect from this drug. Moreover, in treating such patients, we strongly recommend monitoring the fraction of hemoglobin modified by the drug in order to make an informed assessment of its efficacy. This work was supported by the Intramural Research Programs of the National Heart, Lung, and Blood Institute and the National Institute of Diabetes and Digestive and Kidney Diseases. We thank Global Blood Therapeutics for the gift of Voxelotor used in this study, the Department of Transfusion Medicine (DTM) at the NIH for transfusion support and Dr. Joshua Lehrer for helpful discussion and sharing results of unpublished work. We thank Dr. Jay Lozier and Frank H. Bunn for helpful discussions. Figure S1 Oxygen binding curve of patient's red cells. Measurements were made at 37°C with a HEMOX analyzer (TCS Scientific Corporation, New Hope, PA). Whole blood was diluted 1 to 50 in the HEMOX buffer supplied by TCS Scientific, which yields a p50 of 28 Torr for normal blood. The green curve in each panel is the same and was measured 1 day prior to first the treatment with Voxelotor. Its p50 is 44 Torr, which is higher than normal mainly because of the formation of fibers that have a much lower oxygen affinity than the free hemoglobin tetramer in solution—about 3-fold lower than the solution T quaternary conformation of hemoglobin (Eaton and Hofrichter, Adv. Prot. Chem. 1990). In addition, with such severe anemia, the level of red cell 2,3-DPG in the patient's red cells is likely to markedly elevated, further lowering oxygen affinity. Measurements were made on 18 different days during the 10-month treatment period (dates in Table 1 of main text). The curves in panels a-d are representative of these 18 measurements. The orange curve in each panel is the oxygen binding curve for the 1:50 dilution of the patient's whole blood on the day of the blood draw. The cyan curve in each panel was measured in the HEMOX buffer containing 200 micromolar Voxelotor, which is sufficient to modify almost 100% of the hemoglobin molecules with the drug (See Supporting Information Figure S13 in Li et al., PNAS 2017). The lowest measurable pressure of the HEMOX instrument is 2 Torr, which produces only a small error in the fractional saturation with oxygen at low pressures for red cells containing normal or sickle cell hemoglobin. However, it does result in oxygen saturations that are too low and p50's that are too high for hemoglobins with high affinity, such as hemoglobin with 100% drug bound. Based on the recorded optical density changes, 2 Torr corresponds to a fractional saturation of about 0.25. The black dashed curve is the linear combination of the cyan and green curves that is the best least squares fit to the orange curve between 2 and 20 Torr (see Figure S13 in Supporting Information in Li et al., PNAS 2017). Consequently, the fitted fraction of the cyan curve corresponds to the average fraction of hemoglobin molecule in the patient's red cells that have been modified with the drug (listed in Table 1 of main text). The fractions modified in panels a-d are 0.01, 0.05, 0.11, and 0.18, respectively. Unlike the oxygen binding measurements in Li et al., where there was a measurement of the binding curve in the presence and absence of Voxelotor on the same sample, we have only a single binding curve of the patient's blood before treatment. Oxygen binding curves are sensitive to the fraction polymerized, which could easily show changes with time, but only above 20 Torr (See Figures V.5 and V.6 in Eaton and Hofrichter, 1990). This sensitivity could account for the deviation of the fitted (black dashed) curve to the orange curve above 20 Torr and justifies obtaining the fraction modified by least-squares fitting the sum of the green and cyan curves to the orange curve only between 2 and 20 Torr. Measurements of the time-dependence of the binding curves showed less than a 10% change in the fraction modified after 4 hours at 37 C (D. Ostrowski, unpublished results) Figure S2. (A) Observed percent modification of hemoglobin after 28 days of treatment of SCD patients with a daily dose of 0, 500, 700, or 1000 mg of Voxelotor. The percent modified was calculated from the change in p20 of oxygen dissociation curves between Day 28 and Day 1 of treatment from results reported by Patel et al using the method described in ref. 4 as follows: % modified = (p20 + 0.16)/0.27 The reported values are median ± interquartile range. From weighted linear regression, the following relationship was determined with an R2 of 0.94: %Hb Modified = 0.0179 × Dose + 0.9233. Extrapolating this relationship, the expected modifications at daily doses of 1500 mg and 1800 mg were approximately 28% and 33%, respectively (B) Percent modification of hemoglobin calculated with the Δp20 method compared with the percent modification calculated via linear combination (LC) as described in Figure S1 for red cells from AA and SS donors treated with varying concentrations of Voxelotor. Using linear regression, the following relationship was found with an R2 of 0.89: %Hb Modified (Δp20) = 1.46 × % Hb Modified (LC) + 2.05. From this result, it was concluded that, in the range of expected modification, the two methods agreed given the large variation reported by Patel et al in the measurements Δp20 and percent hemoglobin modified.4 Please note: The publisher is not responsible for the content or functionality of any supporting information supplied by the authors. Any queries (other than missing content) should be directed to the corresponding author for the article.