University of Virginia School of Medicine, Department of Pathology, Charlottesville, Virginia (K.H.D., A.M.H., J.C.Z.) and.
University of Colorado Anschutz Medical Campus, Department of Biochemistry and Molecular Genetics, Aurora, Colorado (J.A.R., A.D.).
J Pharmacol Exp Ther. 2023 Sep;386(3):323-330. doi: 10.1124/jpet.123.001634. Epub 2023 Jun 22.
Glucose 6-phosphate dehydrogenase (G6PD) deficiency is the most common enzymopathy in humans (∼5% of all individuals). G6PD deficiency (G6PDd) is caused by an unstable enzyme and manifests most strongly in red blood cells (RBCs) that cannot synthesize new protein. G6PDd RBCs have decreased ability to mitigate oxidative stress due to lower levels of NADPH, as a result of a defective pentose phosphate pathway. Accordingly, oxidative drugs can result in hemolysis and potentially life-threatening anemia in G6PDd patients. Dapsone is a highly useful drug for treating a variety of pathologies but oral dapsone is contraindicated in patients with G6PDd due to oxidative stress-induced anemia. Dapsone must be metabolized to become hemolytic. Dapsone hydroxylamine (DDS-NOH) has been implicated as the major hemolytic dapsone metabolite, but this has never been tested on G6PDd RBCs with in vivo circulation as a metric. Moreover, the metabolic lesion caused by DDS-NOH is unknown. We report that RBCs from a novel humanized mouse expressing the human Mediterranean G6PD-deficient variant have increased sensitivity to DDS-NOH. In addition, we show that DDS-NOH damaged RBCs can either undergo sequestration (with subsequent return to circulation) or permanent removal in a dose-dependent manner, with G6PD-sufficient RBCs mostly being sequestered, and G6PDd RBCs mostly being permanently removed. Finally, we characterize the metabolic lesion caused by DDS-NOH in G6PDd RBCs and report a blockage in terminal glycolysis resulting in a cellular accumulation of pyruvate. These findings confirm DDS-NOH as a hemolytic metabolite and elucidate metabolic effects of DDS-NOH on G6PDd RBCs. SIGNIFICANCE STATEMENT: These findings confirm that dapsone hydroxylamine, an active metabolite of dapsone, causes in vivo clearance of murine red blood cells expressing a human variant of deficient glucose 6-phosphate dehydrogenase (G6PD), an enzymopathy that affects half a billion individuals (G6PD deficiency). Both cellular mechanisms of clearance (sequestration versus destruction) and specific metabolic disturbances caused by dapsone hydroxylamine are elucidated, providing novel mechanistic understanding.
葡萄糖-6-磷酸脱氢酶（G6PD）缺乏症是人类最常见的酶病（∼所有个体的 5%）。G6PD 缺乏症（G6PDd）是由不稳定的酶引起的，主要表现在无法合成新蛋白质的红细胞（RBC）中。由于戊糖磷酸途径缺陷，G6PDd RBC 产生新的 NADPH 的能力下降，导致其减轻氧化应激的能力降低。因此，氧化性药物会导致 G6PDd 患者发生溶血和潜在的威胁生命的贫血。氨苯砜是一种非常有用的药物，可用于治疗多种疾病，但由于氧化应激诱导的贫血，口服氨苯砜在 G6PDd 患者中被禁用。氨苯砜必须代谢才能产生溶血作用。已发现氨苯砜羟胺（DDS-NOH）是主要的溶血氨苯砜代谢物，但这从未在具有体内循环的 G6PDd RBC 上作为指标进行过测试。此外，DDS-NOH 引起的代谢损伤尚不清楚。我们报告说，表达人类地中海 G6PD 缺乏变异体的新型人源化小鼠的 RBC 对 DDS-NOH 更为敏感。此外，我们表明，DDS-NOH 损伤的 RBC 可以以剂量依赖性方式进行隔离（随后返回循环）或永久性清除，G6PD 充足的 RBC 主要被隔离，而 G6PDd RBC 主要被永久性清除。最后，我们描述了 DDS-NOH 在 G6PDd RBC 中引起的代谢损伤，并报告了终末糖酵解的阻断导致丙酮酸在细胞内积累。这些发现证实了 DDS-NOH 是一种溶血代谢物，并阐明了 DDS-NOH 对 G6PDd RBC 的代谢影响。意义声明：这些发现证实，氨苯砜羟胺，即氨苯砜的一种活性代谢物，可清除表达人类葡萄糖-6-磷酸脱氢酶（G6PD）缺陷变异体的鼠红细胞，这种酶病影响了 5 亿人（G6PD 缺乏症）。阐明了 DDS-NOH 引起的清除的细胞机制（隔离与破坏）和特定的代谢紊乱，提供了新的机制理解。
