Phua Yu Leng, D'Annibale Olivia M, Karunanidhi Anuradha, Mohsen Al-Walid, Kirmse Brian, Dobrowolski Steven F, Vockley Jerry
Department of Pediatrics, Division of Genetic and Genomic Medicine, UPMC Children's Hospital of Pittsburgh, Pittsburgh, PA, USA.
Department of Pathology, Clinical Biochemical Genetics Laboratory, UPMC Children's Hospital of Pittsburgh, Pittsburgh, PA, USA.
bioRxiv. 2023 Feb 3:2023.02.02.526527. doi: 10.1101/2023.02.02.526527.
Combined D, L-2-Hydroxyglutaric Aciduria (D,L-2HGA) is a rare genetic disorder caused by recessive mutations in the gene that encodes the mitochondrial citrate carrier protein (CIC). deficiency leads to a secondary increase in mitochondrial 2-ketoglutarate that, in turn, is reduced to neurotoxic 2-hydroxyglutarate. Clinical symptoms of Combined D,L-2HGA include neonatal encephalopathy, respiratory insufficiency and often with death in infancy. No current therapies exist, although replenishing cytosolic stores by citrate supplementation to replenish cytosolic stores has been proposed. In this study, we demonstrated that patient derived fibroblasts exhibited impaired cellular bioenergetics that were worsened with citrate supplementation. We hypothesized treating patient cells with phenylbutyrate, an FDA approved pharmaceutical drug, would reduce mitochondrial 2-ketoglutarate, leading to improved cellular bioenergetics including oxygen consumption and fatty acid oxidation. Metabolomic and RNA-seq analyses demonstrated a significant decrease in intracellular 2-ketoglutarate, 2-hydroxyglutarate, and in levels of mRNA coding for citrate synthase and isocitrate dehydrogenase. Consistent with the known action of phenylbutyrate, detected levels of phenylacetylglutamine was consistent with the drug acting as 2-ketoglutarate sink in patient cells. Our pre-clinical studies suggest citrate supplementation is unlikely to be an effective treatment of the disorder. However, cellular bioenergetics suggests phenylbutyrate may have interventional utility for this rare disease.
联合型D，L-2-羟基戊二酸尿症（D，L-2HGA）是一种罕见的遗传性疾病，由编码线粒体柠檬酸载体蛋白（CIC）的基因突变引起，该突变呈隐性。CIC缺乏导致线粒体2-酮戊二酸继发性增加，进而还原为具有神经毒性的2-羟基戊二酸。联合型D，L-2HGA的临床症状包括新生儿脑病、呼吸功能不全，且常在婴儿期死亡。目前尚无有效治疗方法，尽管有人提出通过补充柠檬酸盐来补充胞质储存。在本研究中，我们发现患者来源的成纤维细胞表现出线粒体生物能量代谢受损，补充柠檬酸盐后情况恶化。我们推测，用美国食品药品监督管理局（FDA）批准的药物苯丁酸钠治疗患者细胞，会减少线粒体2-酮戊二酸，从而改善包括耗氧和脂肪酸氧化在内的细胞生物能量代谢。代谢组学和RNA测序分析表明，细胞内2-酮戊二酸、2-羟基戊二酸以及编码柠檬酸合酶和异柠檬酸脱氢酶的mRNA水平均显著降低。与苯丁酸钠的已知作用一致，检测到的苯乙酰谷氨酰胺水平表明该药物在患者细胞中充当2-酮戊二酸的吸收剂。我们的临床前研究表明，补充柠檬酸盐不太可能有效治疗该疾病。然而，细胞生物能量代谢研究表明，苯丁酸钠可能对这种罕见疾病具有干预作用。
