Research Article: Oral tetrahydrouridine and decitabine for non-cytotoxic epigenetic gene regulation in sickle cell disease: A randomized phase 1 study

Date Published: September 7, 2017

Publisher: Public Library of Science

Author(s): Robert Molokie, Donald Lavelle, Michel Gowhari, Michael Pacini, Lani Krauz, Johara Hassan, Vinzon Ibanez, Maria A. Ruiz, Kwok Peng Ng, Philip Woost, Tomas Radivoyevitch, Daisy Pacelli, Sherry Fada, Matthew Rump, Matthew Hsieh, John F. Tisdale, James Jacobberger, Mitch Phelps, James Douglas Engel, Santhosh Saraf, Lewis L. Hsu, Victor Gordeuk, Joseph DeSimone, Yogen Saunthararajah, David Rees

Abstract: BackgroundSickle cell disease (SCD), a congenital hemolytic anemia that exacts terrible global morbidity and mortality, is driven by polymerization of mutated sickle hemoglobin (HbS) in red blood cells (RBCs). Fetal hemoglobin (HbF) interferes with this polymerization, but HbF is epigenetically silenced from infancy onward by DNA methyltransferase 1 (DNMT1).Methods and findingsTo pharmacologically re-induce HbF by DNMT1 inhibition, this first-in-human clinical trial (NCT01685515) combined 2 small molecules—decitabine to deplete DNMT1 and tetrahydrouridine (THU) to inhibit cytidine deaminase (CDA), the enzyme that otherwise rapidly deaminates/inactivates decitabine, severely limiting its half-life, tissue distribution, and oral bioavailability. Oral decitabine doses, administered after oral THU 10 mg/kg, were escalated from a very low starting level (0.01, 0.02, 0.04, 0.08, or 0.16 mg/kg) to identify minimal doses active in depleting DNMT1 without cytotoxicity. Patients were SCD adults at risk of early death despite standard-of-care, randomized 3:2 to THU–decitabine versus placebo in 5 cohorts of 5 patients treated 2X/week for 8 weeks, with 4 weeks of follow-up. The primary endpoint was ≥ grade 3 non-hematologic toxicity. This endpoint was not triggered, and adverse events (AEs) were not significantly different in THU-decitabine—versus placebo-treated patients. At the decitabine 0.16 mg/kg dose, plasma concentrations peaked at approximately 50 nM (Cmax) and remained elevated for several hours. This dose decreased DNMT1 protein in peripheral blood mononuclear cells by >75% and repetitive element CpG methylation by approximately 10%, and increased HbF by 4%–9% (P < 0.001), doubling fetal hemoglobin-enriched red blood cells (F-cells) up to approximately 80% of total RBCs. Total hemoglobin increased by 1.2–1.9 g/dL (P = 0.01) as reticulocytes simultaneously decreased; that is, better quality and efficiency of HbF-enriched erythropoiesis elevated hemoglobin using fewer reticulocytes. Also indicating better RBC quality, biomarkers of hemolysis, thrombophilia, and inflammation (LDH, bilirubin, D-dimer, C-reactive protein [CRP]) improved. As expected with non-cytotoxic DNMT1-depletion, platelets increased and neutrophils concurrently decreased, but not to an extent requiring treatment holds. As an early phase study, limitations include small patient numbers at each dose level and narrow capacity to evaluate clinical benefits.ConclusionAdministration of oral THU-decitabine to patients with SCD was safe in this study and, by targeting DNMT1, upregulated HbF in RBCs. Further studies should investigate clinical benefits and potential harms not identified to date.Trial, NCT01685515

Partial Text: Sickle cell disease (SCD) is a congenital hemolytic anemia caused by polymerization and precipitation of mutated sickle hemoglobin (HbS, α2βS2) in red blood cells (RBCs). This decreases RBC life span by >90%, causing severe anemia. The unhealthy RBCs also adhere to and occlude blood vessels. Ensuing tissue hypoxia damages all organs, triggers severe pain, and compromises immunity. In low-income countries, most children with SCD do not survive to adulthood. Even in high-income countries, morbidity can be severe and life spans are reduced by several decades—the median life expectancy for people with SCD in the United States is approximately 45 years [1,2].

This first-in-human clinical trial evaluated safety of oral THU-decitabine, and identified an appropriate dose, for DNMT1 depletion and HbF induction in SCD [70]. Oral THU-decitabine was safe and well-tolerated in this study and produced a wide decitabine concentration-time profile (low Cmax, long Tmax) ideal for non-cytotoxic DNMT1 depletion, since DNMT1 depletion can occur at low nanomolar concentrations but depends on exposure timing [46,73–75]. Increases in fetal and total hemoglobin expected to be clinically significant were produced at the highest decitabine dose administered, 0.16 mg/kg, accompanied by improvements in laboratory biomarkers of hemolysis, coagulation, and inflammation. The side effects were a concurrent increase in platelets and decrease in neutrophils, expected with non-cytotoxic DNMT1 depletion.



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