Cancer Metabolism Laboratory, The Francis Crick Institute, London, UK.
Drug Transport and Tumour Metabolism Lab, MRC London Institute of Medical Sciences, London, UK.
Commun Biol. 2022 Aug 26;5(1):877. doi: 10.1038/s42003-022-03805-y.
α-ketoglutarate (αKG) is a central metabolic node with a broad influence on cellular physiology. The αKG analogue N-oxalylglycine (NOG) and its membrane-permeable pro-drug derivative dimethyl-oxalylglycine (DMOG) have been extensively used as tools to study prolyl hydroxylases (PHDs) and other αKG-dependent processes. In cell culture media, DMOG is rapidly converted to MOG, which enters cells through monocarboxylate transporter MCT2, leading to intracellular NOG concentrations that are sufficiently high to inhibit glutaminolysis enzymes and cause cytotoxicity. Therefore, the degree of (D)MOG instability together with MCT2 expression levels determine the intracellular targets NOG engages with and, ultimately, its effects on cell viability. Here we designed and characterised a series of MOG analogues with the aims of improving compound stability and exploring the functional requirements for interaction with MCT2, a relatively understudied member of the SLC16 family. We report MOG analogues that maintain ability to enter cells via MCT2, and identify compounds that do not inhibit glutaminolysis or cause cytotoxicity but can still inhibit PHDs. We use these analogues to show that, under our experimental conditions, glutaminolysis-induced activation of mTORC1 can be uncoupled from PHD activity. Therefore, these new compounds can help deconvolute cellular effects that result from the polypharmacological action of NOG.
α-酮戊二酸（αKG）是一个中央代谢节点，对细胞生理有广泛的影响。αKG 的类似物草酰甘氨酸（NOG）及其膜通透前药衍生物二甲基草酰甘氨酸（DMOG）已被广泛用作研究脯氨酰羟化酶（PHD）和其他αKG 依赖性过程的工具。在细胞培养基中，DMOG 迅速转化为 MOG，MOG 通过单羧酸转运蛋白 MCT2 进入细胞，导致细胞内的 NOG 浓度足够高，从而抑制谷氨酰胺分解酶并引起细胞毒性。因此，（D）MOG 的不稳定性程度以及 MCT2 的表达水平决定了 NOG 与之结合的细胞内靶标，并最终决定其对细胞活力的影响。在这里，我们设计并表征了一系列 MOG 类似物，旨在提高化合物的稳定性，并探索与 MCT2 相互作用的功能要求，MCT2 是 SLC16 家族中一个相对研究较少的成员。我们报告了能够通过 MCT2 进入细胞的 MOG 类似物，并确定了不抑制谷氨酰胺分解或引起细胞毒性但仍能抑制 PHD 的化合物。我们使用这些类似物表明，在我们的实验条件下，谷氨酰胺分解诱导的 mTORC1 激活可以与 PHD 活性解耦。因此，这些新化合物可以帮助阐明由于 NOG 的多药理学作用而导致的细胞效应。
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