Gliom oluşumunun erken aşamalarında IDH1 mutasyonlarının epigenomda yol açtığı değişimlerin karakterizasyonu

Isocitrate dehydrogenase (IDH1/2) mutations, which have been shown to develop before other mutations known to play a role in glioma development, are believed to trigger gliomagenesis. Studies to date have mostly used heterogeneous tumor samples or cells with oncogenic mutations when investigating the early stages of glioma development. This limits our knowledge of how gliomagenesis is triggered by IDH1/2 mutations. Moreover, examination of epigenomic changes caused by IDH1/2 mutations has not gone beyond classical histone methylation analyzes based on methylation arrays, RNA-seq and expression microarrays, and ChIP-seq. The aim of this study was to understand the permanent changes caused by the IDH1-R132H mutation, which is the earliest seen in gliomagenesis, in the epigenome of neural progenitor cells (NPCs) cultured in a 3D environment. For this, firstly, conditioned medium with Immortalized Human Astrocytes (IHAs) expressing doxycycline-inducible IDH1 (Mutant and wild type) was obtained. These media samples were then delivered for specified periods of time to NPCs differentiated from human pluripotent stem cells (hiPSC) in an alginate-based 3D matrix. RT-qPCR and RNA-seq analyzes were performed by isolating RNA from cells taken at three time points (day 0, day 14, day 17) from NPCs exposed to different media. Key genes reported to be altered in IDH1 mutant gliomas were checked by RT-qPCR. Then, epigenetic marks in histones and DNA were analyzed by ChIP-qPCR and MeDIP-qPCR. In selecting the loci for these analyses, we particularly prioritized the aim of furthering the mechanistic information regarding the IDH1-MYC relationship in the risk of glioma located at 8q24, which we brought to the literature for the first time in our group's previous studies, and directly demonstrating it in an in vitro model. Our findings indicate that one of the direct targets of the oncogenic changes that occur in the IDH1-MYC axis, which we think are initiators of gliomagenesis, is the ID1 (inhibitor of differentiation) gene, and that as a result of the induction of this gene, the cells take on a less differentiated phenotype.