Metabolic fate of glucose in atypical meningioma studied by 13C NMR isotopomer analysis
Kumar Pichumani1, Omkar B Ijare1, and David S Baskin1

1Neurosurgery, Houston Methodist Hospital and Research Institute, Houston, TX, United States


Meningiomas are tumors arising from meninges, the membranes surrounding the brain and the spinal cord. Majority of meningiomas (grade-1) are benign and grow slowly. However, atypical meningiomas (grade-2) exhibit increased cellular abnormalities, and grow at a faster rate than benign meningiomas. Moreover, atypical meningiomas prone to recurrence and show resistance to radiotherapy. Atypical meningiomas show higher 18F-FDG uptake in PET scans. No prior reports are available on investigating metabolic fate of glucose in atypical meningiomas. The goal of this study is to probe the metabolic fate of glucose using NMR based [U-13C]glucose isotopic tracing methods in patient-derived atypical meningioma cells.


Meningiomas are the most frequently diagnosed CNS tumors that arise from meningeal membranes surrounding the brain and spinal cord. They represent about 30% of all CNS tumors. World Health Organization (WHO) classifies meningiomas into three types based on histology [1,2]. Grade I meningiomas are benign growths and are slow growing. Grade-II, known as “atypical” meningiomas represent 10-15% and show high index of recurrence. Grade III meningiomas are anaplastic malignancies and represent about 1-3% of meningiomas. Atypical menigiomas exhibit high uptake of 18F-FDG by PET imaging. Lactate and alanine levels are higher in atypical meningiomas indicating altered glucose metabolism similar to malignant brain tumors [3,4]. However, metabolic fate of glucose in human atypical meningiomas is largely unknown. In this study, we investigate [U-13C]glucose metabolism in a patient-derived atypical meningioma cell line using NMR spectroscopy and isotopomer analysis.


Tumor tissues were collected from atypical meningioma patients undergoing craniotomy at the Houston Methodist Hospital following an approved IRB protocol. Cells were isolated from the tumor tissue and were initially grown in ABMTM Astrocyte Basal Medium, incubated at 37°C under humidified air with 5% CO2. During the final 6 hours, 15.0 mM [U-13C]glucose was introduced to the glucose-free Dulbecco’s modified Eagle’s medium (DMEM). After 6 hours, the medium was removed, cells were washed with PBS buffer and the cell pellets were snap-frozen in liquid nitrogen. Prior to NMR data collection, the cell pellets were extracted in 5% perchloric acid, extracts were lyophilized and were reconstituted in 180 µL D2O containing 1.0 mM DSS-d6 (pH = 7.4). The sample-solutions were transferred to a 3-mm NMR tubes, and 1D 1H and 1H-decoupled 13C NMR spectra were collected on a Bruker 600 MHz spectrometer equipped with a 5-mm cryo-probe optimized for direct 13C detection.

Glycolysis is highly active in these cells leading to the generation of [U-13C]lactate and [U-13C]alanine (Figure 1). 35.0% of total lactate pool is 13C enriched. Further, 13C labeled glutamate isotopomers were also detected indicating oxidation of glucose in TCA cycle (Figure 2). Glucose-derived pyruvate carbons can enter TCA cycle through two different pathways: 1. [U-13C]glucose is converted to [U-13C]pyruvate via glycolysis leading to the production [1,2-13C]acetyl-CoA through pyruvate dehydrogenase (PDH) which further generates [4,5-13C] labeled α-ketoglutarate (α-KG)/glutamate upon condensation with unlabeled (12C)oxaloacetate (OAA) in the first turn of the cycle. After multiple turns of the cycle, [3,4,5-13C]α-KG and glutamate are generated. 2. Pyruvate carboxylase (PC) generates [1,2,3-13C]oxaloacetate from [U-13C]pyruvate which enters the TCA cycle leading to the generation of [2,3-13C] α-KG and glutamate isotopomers. Figure 2 shows the portion of the 13C NMR spectrum of C4 and C2 multiplet signals of glutamate. The singlet (C4S) is due to natural abundance C4 glutamate. The doublet (C4D45) and the quartet (C4Q) corresponds to [4,5-13C]glutamate, and [3,4,5-13C]glutamate isotopomers respectively. These two together measure the activity/flux of PDH enzyme. Similarly, the singlet (C2S) is due to natural abundance C2 glutamate. The doublet (C2D23) in C2 glutamate signal is generated by [2,3-13C]glutamate which is produced only through PC activity. Figure 3 (A) shows charts of normalized peak areas of C4S, C4D45 and C4Q peaks of C4 glutamate signal; Figure 3 (B) shows normalized peak areas of C2S and C2D23 of C2 glutamate signal. The relative activities of PDH and PC in meningioma cells are determined using [1,2-13C]acetyl-CoA values and the ratio of singlet (C2S) and C2D23 peak areas of C2 glutamate signal [3,4]. In this study, 18.3% of [1,2-13C]acetyl-CoA is produced from [U-13C]glucose metabolism in atypical meningioma cells. The ratio (C2S/C2D23) was found to be 0.26 which indicates a significant PC activity in atypical meningioma cells. Figure 4 is a Schema summarizing the metabolism of [U-13C]glucose in the meningioma tumor cell.


Two major findings in this study are: First, in addition to producing lactate, atypical meningioma cells are capable of oxidizing glucose in the TCA cycle to generate energy via acetyl-CoA production through PDH activity. Second, these cells show active anaplerosis through the presence of PC activity. Proliferating cells need high energy to produce carbon precursors/intermediates to synthesize macromolecules that are generated within the TCA cycle in the mitochondria. The flux through PC in these cells replenishes OAA in order to maintain continuous flow of TCA cycle intermediates for anabolic processes [5].


We thank Peak Center for financial support. We thank Blessy John and Sophie Lopez for technical assistance.


  1. Monleon D, Morales JM, Gonzales-Darder J et al., Benign and atypical meningioma metabolic signatures by HR-MAS molecular profiling. J Proteome Res. 7 (2008) 2882-2888.

  2. Monleón D, Morales JM, Gonzalez-Segura A, Gonzalez-Darder JM, Gil-Benso R, Cerdá-Nicolás M, López-Ginés C. Metabolic aggressiveness in benign meningiomas with chromosomal instabilities. Cancer Res. 70 (2010) 8426-8434.

  3. E.A. Maher, I. Marin-Valencia, R.M. Bachoo, T. Mashimo, J. Raisanen, K.J. Hatanpaa, A. Jindal, F.M. Jeffrey, C. Choi, C. Madden, D. Mathews, J.M. Pascual, B.E. Mickey, C.R. Malloy, R.J. DeBerardinis, Metabolism of [U-13 C]glucose in human brain tumors in vivo, NMR. Biomed. 25 (2012) 1234-1244.

  4. K. Pichumani, T. Mashimo, H-M. Baek, J. Ratnakar, B. Mickey, R.J. DeBerardinis, E.A. Maher, R.M. Bachoo, C.R. Malloy, Z. Kovacs, Conditions for 13C NMR detection of 2-Hydroxyglutarate in tissue extracts from isocitrate dehydrogenase-mutated gliomas, Anal. Biochem. 481 (2015) 4-6.

  5. Cheng T, Sudderth J, Yang C, Mullen AR, Jin ES, Matés JM, DeBerardinis RJ. Pyruvate carboxylase is required for glutamine-independent growth of tumor cells. Proc Natl Acad Sci U S A. 108 (2011) 8674-8679.


13C NMR spectral regions of C2 carbon of lactate and alanine with various spin-spin coupled multiplets. The singlet (S) is due to natural abundant C2 lactate whereas the quartet (Q) is due to [U-13C]lactate, which are arising from the metabolism of [U-13C]glucose in atypical meningioma cells. Similar assignments apply for C2 alanine.

13C NMR spectral regions of C4 glutamate and C2 glutamate signals with various spin-spin coupled multiplets. In C4 glutamate, the singlet (C4S) is due to natural abundance whereas the doublet (C4D45) and the quartet (C4Q) are due to [4,5-13C]glutamate and [3,4,5-13C]glutamate respectively which are generated from [U-13C]glucose-derived [1,2-13C]acetyl-CoA. In C2 glutamate, the singlet (C2S) is due to natural abundance, the doublets C2D23, and C2D12 are arising from spin-coupling interactions of [2,3-13C]glutamate and [1,2-13C]glutamate respectively; and the quartet (C2Q) is due to [1,2,3-13C]glutamate respectively, all arising from the metabolism of [U-13C]glucose. In particular, C2D23 is produced through pyruvate carboxylase (PC) activity.

(A) Normalized 13C NMR peak areas of various C4 glutamate multiplet signal. The singlet (C4S) appears due to natural abundance of C4 glutamate. The doublet (C4D45) and quartert (C4Q) are due to [4,5-13C]glutamate and [3,4,5-13C]glutamate isotopomers respectively that are derived from the metabolism of [U-13C]glucose in atypical meningioma cells. (B) Normalized 13C NMR peak areas of various multiplet signals of C2 glutamate. The singlet (C2S) and the doublet (C2D23) are due to natural abundance of C2 glutamate and [2,3-13C]glutamate respectively. (C) Fc3 represents the [1,2-13C]acetyl-CoA derived from [U-13C]pyruvate via PDH activity. The ratio (C2S/C2D23) is a measure of PC activity.

Schema summarizing metabolism of [U-13C]glucose in an atypical meningioma cell. In addition to glycolysis, both PDH and PC enzymatic pathways are active which generate [1,2-13C]acetyl-CoA and [1,2,3-13C]OAA respectively (depicted by red and blue filled circles). [1,2-13C]acetyl-CoA enters TCA cycle to produce [4,5-13C]α-KG and [4,5-13C]GLU in the first turn of the cycle. On the other hand, [1,2,3-13C]OAA upon entering TCA cycle generates [2,3-13C] α-KG /[2,3-13C]GLU. Abbreviations: LDH: lactate dehydrogenase; ALT: alanine aminotransferase; PDH: pyruvate dehydrogenase; PC: pyruvate carboxylase; CIT: citrate; OAA: oxaloacetate; α-KG: α-ketoglutarate; GLU-glutamate.

Proc. Intl. Soc. Mag. Reson. Med. 27 (2019)