IDH1R132H in Neural Stem Cells: Differentiation Impaired by Increased Apoptosis

Abstract

BackgroundThe high frequency of mutations in the isocitrate dehydrogenase 1 (IDH1) gene in diffuse gliomas indicates its importance in the process of gliomagenesis. These mutations result in loss of the normal function and acquisition of the neomorphic activity converting α-ketoglutarate to 2-hydroxyglutarate. This potential oncometabolite may induce the epigenetic changes, resulting in the deregulated expression of numerous genes, including those related to the differentiation process or cell survivability.MethodsNeural stem cells were derived from human induced pluripotent stem cells following embryoid body formation. Neural stem cells transduced with mutant IDH1R132H, empty vector, non-transduced and overexpressing IDH1WT controls were differentiated into astrocytes and neurons in culture. The neuronal and astrocytic differentiation was determined by morphology and expression of lineage specific markers (MAP2, Synapsin I and GFAP) as determined by real-time PCR and immunocytochemical staining. Apoptosis was evaluated by real-time observation of Caspase-3 activation and measurement of PARP cleavage by Western Blot.ResultsCompared with control groups, cells expressing IDH1R132H retained an undifferentiated state and lacked morphological changes following stimulated differentiation. The significant inhibitory effect of IDH1R132H on neuronal and astrocytic differentiation was confirmed by immunocytochemical staining for markers of neural stem cells. Additionally, real-time PCR indicated suppressed expression of lineage markers. High percentage of apoptotic cells was detected within IDH1R132H-positive neural stem cells population and their derivatives, if compared to normal neural stem cells and their derivatives. The analysis of PARP and Caspase-3 activity confirmed apoptosis sensitivity in mutant protein-expressing neural cells.ConclusionsOur study demonstrates that expression of IDH1R132H increases apoptosis susceptibility of neural stem cells and their derivatives. Robust apoptosis causes differentiation deficiency of IDH1R132H-expressing cells.

Mutant IDH1 Disrupts the Mouse Subventricular Zone and Alters Brain Tumor Progression

“…Collectively, these experiments suggest that mutant Idh1 likely affects NSCs both directly, as observed through defective differentiation, and indirectly, as observed through a disorganization of the microenvironment, two components with implications in tumor initiation and development (27). This data also corroborates a previous study investigating the role of mutant IDH1 on embryoid body induced human neural stem cells which showed that IDH1-R132H impaired the ability of these cells to differentiate, specifically into the astrocytic and neuronal lineage (28). …”

Low-Grade Astrocytoma Mutations in IDH1, P53, and ATRX Cooperate to Block Differentiation of Human Neural Stem Cells via Repression of SOX2

“…Expression of mutant IDH1 in progenitor cells inhibits differentiation (Figueroa et al, 2010; Lu et al, 2012; Pirozzi et al, 2017; Rosiak et al, 2016; Turcan et al, 2012; Xu et al, 2011). However, the effect of combined IDH mutation and P53/ATRX loss on NSC self-renewal and differentiation is not known.…”

“…Our model shows that the combination of the 3 oncogenic hits promotes gliomagenesis not by altering cell growth but by arresting differentiation of NSCs (Pirozzi et al, 2017; Rosiak et al, 2016; Sulkowski et al, 2017), which become locked in a self-renewing and brain-invasive state. Several lines of evidence support the notion that the IDH1 mutation is the inciting oncogenic hit in LGA.…”

“…Mutant IDH1 is a shared core mutation in both LGA and oligodendroglioma (Cancer Genome Atlas Research et al, 2015; Parsons et al, 2008), suggesting the cell of origin is a shared neuroglial progenitor (Bardella et al, 2016; Rosiak et al, 2016). …”

“…First, exome sequencing of IDH-mutated astrocytoma at initial diagnosis and recurrence showed that, while the IDH1 is preserved, distinct clonal mutations in P53 and ATRX emerge at recurrence (Johnson et al, 2014). Second, expression of mutant IDH1 alone in progenitor cells leads to pre-tumorigenic changes (Bardella et al, 2016; Figueroa et al, 2010; Lu et al, 2012; Pirozzi et al, 2017; Rosiak et al, 2016; Saha et al, 2014; Turcan et al, 2012). Third, germline mosaicism for neomorphic IDH mutations in Ollier disease can result in brain astrocytomas (Amary et al, 2011; Bonnet et al, 2016; Pansuriya et al, 2011).…”