Abstract
Fluorescence-guided surgery using 5-aminolevulinic acid (5-ALA) has become the main treatment modality in malignant gliomas. However unlike glioblastomas, there are inconsistent result about fluorescence status in WHO grade III gliomas. Here, we show that mutational status of IDH1 is linked to 5-ALA fluorescence. Using genetically engineered malignant glioma cells harboring wild type (U87MG-IDH1WT) or mutant (U87MG-IDH1R132H) IDH1, we demonstrated a lag in 5-ALA metabolism and accumulation of protoporphyrin IX (PpIX) in U87MG-IDH1R132Hcells. Next, we used liquid chromatography-mass spectrometry (LC-MS) to screen for tricarboxylic acid (TCA) cycle-related metabolite changes caused by 5-ALA exposure. We observed low baseline levels of NADPH, an essential cofactor for the rate-limiting step of heme degradation, in U87MG-IDH1R132H cells. High levels of NADPH are required to metabolize excessive 5-ALA, giving a plausible reason for the temporarily enhanced 5-ALA fluorescence in mutant IDH1 cells. This hypothesis was supported by the results of metabolic screening in human malignant glioma samples. In conclusion, we have discovered a relationship between enhanced 5-ALA fluorescence and IDH1 mutations in WHO grade III gliomas. Low levels of NADPH in tumors with mutated IDH1 is responsible for the enhanced fluorescence.
Highlights
Among recent advances in malignant glioma surgery, fluorescence-guided surgery is becoming popular due to its ability to improve the extent of resection while preserving patient functional status [1, www.impactjournals.com/oncotarget2]. 5-aminolevulinic acid (5-ALA) highlights otherwise indistinguishable malignant areas with a temporarily red fluorescence under an appropriate blue light source with a specific wavelength [3]. 5-ALA itself is a nonfluorescent endogenous compound that can be metabolized intracellularly into protoporphyrinogen IX (PpIX), a fluorescent intermediate product of the heme synthesis pathway [4]
Among 35 patients with WHO grade III gliomas who were operated upon using 5-ALA fluorescenceguided surgery, isocitrate dehydrogenase 1 (IDH1) mutations were observed in 24 and intraoperative fluorescence was observed in tumor tissues of 19 patients (Table 1)
Intracellular PpIX fluorescence increased rapidly up to 1 hour after incubation, diminished in U87MG-IDH1WT cells (Figure 3A), whereas there was a delay in 5-ALA metabolism in U87MGIDH1R132H cells; PpIX fluorescence peaked at 2 hours after incubation, at which time there was a significant difference in PpIX concentration between U87MG-IDH1R132H and U87MG-IDH1WT cells (Figure 3B)
Summary
Among recent advances in malignant glioma surgery, fluorescence-guided surgery is becoming popular due to its ability to improve the extent of resection while preserving patient functional status [1, www.impactjournals.com/oncotarget2]. 5-aminolevulinic acid (5-ALA) highlights otherwise indistinguishable malignant areas with a temporarily red fluorescence under an appropriate blue light source with a specific wavelength [3]. 5-ALA itself is a nonfluorescent endogenous compound that can be metabolized intracellularly into protoporphyrinogen IX (PpIX), a fluorescent intermediate product of the heme synthesis pathway [4]. The 5-ALA-induced fluorescence rate is lower in WHO grade III gliomas (83% in anaplastic oligodendroglioma/oligoastrocytomas, and 22% in anaplastic astrocytomas) than grade IV gliomas (96% in glioblastomas) [6]. The reason why grade III gliomas have a lower 5-ALA fluorescence rate than grade IV gliomas is unknown. The identification of isocitrate dehydrogenase 1 (IDH1) mutations in glioma is one of the major discoveries mediating metabolomics and oncogenesis [7]. IDH1 mutations in glioma cells diminish the canonical enzymatic activity of IDH1 while conferring a neomorphic enzymatic activity: the production of R-2hydoxyglutarate (2-HG) via the consumption of NADPH [13,14,15,16]. The IDH1 mutation is observed in 6% of WHO grade IV glioblastomas and 55% of grade III gliomas [17]
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