Viswanath Pavithra, Radoul Marina, Izquierdo-Garcia Jose Luis, Luchman Hema Artee, Gregory Cairncross J, Pieper Russell O, Phillips Joanna J, Ronen Sabrina M
1Department of Radiology and Biomedical Imaging, University of California San Francisco, 1700 4th Street, Box 2532. Byers Hall 3rd Floor, Suite, San Francisco, CA 94143 USA.
2Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain.
Cancer Metab. 2018 Apr 3;6:3. doi: 10.1186/s40170-018-0178-3. eCollection 2018.
Magnetic resonance spectroscopy (MRS) studies have identified elevated levels of the phospholipid precursor phosphocholine (PC) and phosphoethanolamine (PE) as metabolic hallmarks of cancer. Unusually, however, PC and PE levels are reduced in mutant isocitrate dehydrogenase 1 (IDHmut) gliomas that produce the oncometabolite 2-hydroxyglutarate (2-HG) relative to wild-type IDH1 (IDHwt) gliomas. The goal of this study was to determine the molecular mechanism underlying this unusual metabolic reprogramming in IDHmut gliomas.
Steady-state PC and PE were quantified using P-MRS. To quantify de novo PC and PE synthesis, we used C-MRS and measured flux to C-PC and C-PE in cells incubated with [1,2-C]-choline and [1,2-C]-ethanolamine. The activities of choline kinase (CK) and ethanolamine kinase (EK), the enzymes responsible for PC and PE synthesis, were quantified using P-MR-based assays. To interrogate the role of 2-HG, we examined IDHwt cells incubated with 2-HG and, conversely, IDHmut cells treated with the IDHmut inhibitor AGI-5198. To examine the role of hypoxia-inducible factor 1-α (HIF-1α), we silenced HIF-1α using RNA interference. To confirm our findings in vivo and in the clinic, we studied IDHwt and IDHmut orthotopic tumor xenografts and glioma patient biopsies.
De novo synthesis of PC and PE was reduced in IDHmut cells relative to IDHwt. Concomitantly, CK activity and EK activity were reduced in IDHmut cells. Pharmacological manipulation of 2-HG levels established that 2-HG was responsible for reduced CK activity, EK activity, PC and PE. 2-HG has previously been reported to stabilize levels of HIF-1α, a known regulator of CK activity. Silencing HIF-1α in IDHmut cells restored CK activity, EK activity, PC and PE to IDHwt levels. Our findings were recapitulated in IDHmut orthotopic tumor xenografts and, most importantly, in IDHmut patient biopsies, validating our findings in vivo and in the clinic.
This study identifies, to our knowledge for the first time, a direct role for 2-HG in the downregulation of CK and EK activity, and thereby, PC and PE synthesis in IDHmut gliomas. These results highlight the unusual reprogramming of phospholipid metabolism in IDHmut gliomas and have implications for the identification of MRS-detectable metabolic biomarkers associated with 2-HG status.
磁共振波谱（MRS）研究已确定磷脂前体磷酸胆碱（PC）和磷酸乙醇胺（PE）水平升高是癌症的代谢特征。然而，不同寻常的是，相对于野生型异柠檬酸脱氢酶1（IDHwt）胶质瘤，产生致癌代谢物2-羟基戊二酸（2-HG）的突变型异柠檬酸脱氢酶1（IDHmut）胶质瘤中PC和PE水平降低。本研究的目的是确定IDHmut胶质瘤中这种异常代谢重编程的分子机制。
使用磷磁共振波谱（P-MRS）对稳态PC和PE进行定量。为了定量从头合成PC和PE，我们使用碳磁共振波谱（C-MRS）并测量了与[1,2-碳]-胆碱和[1,2-碳]-乙醇胺孵育的细胞中通向碳-PC和碳-PE的通量。使用基于磷磁共振波谱的测定法对负责PC和PE合成的胆碱激酶（CK）和乙醇胺激酶（EK）的活性进行定量。为了探究2-HG的作用，我们检测了与2-HG孵育的IDHwt细胞，反之，检测了用IDHmut抑制剂AGI-5198处理的IDHmut细胞。为了研究缺氧诱导因子1-α（HIF-1α）的作用，我们使用RNA干扰使HIF-1α沉默。为了在体内和临床中证实我们的发现，我们研究了IDHwt和IDHmut原位肿瘤异种移植以及胶质瘤患者活检样本。
相对于IDHwt，IDHmut细胞中PC和PE的从头合成减少。同时，IDHmut细胞中CK活性和EK活性降低。对2-HG水平的药理学操作表明，2-HG是CK活性、EK活性、PC和PE降低的原因。此前有报道称2-HG可稳定HIF-1α的水平，HIF-1α是CK活性的已知调节因子。在IDHmut细胞中使HIF-1α沉默可使CK活性、EK活性、PC和PE恢复到IDHwt水平。我们的发现在IDHmut原位肿瘤异种移植中得到重现，最重要的是，在IDHmut患者活检样本中得到重现，从而在体内和临床中验证了我们的发现。
据我们所知，本研究首次确定了2-HG在下调IDHmut胶质瘤中CK和EK活性以及由此下调PC和PE合成方面的直接作用。这些结果突出了IDHmut胶质瘤中磷脂代谢的异常重编程，并对鉴定与2-HG状态相关的磁共振波谱可检测代谢生物标志物具有启示意义。
