Evaluating Mechanisms of IDH1 Regulation through Site-Specific Acetylation Mimics

Isocitrate dehydrogenase (IDH1) catalyzes the reversible NADP+-dependent oxidation of isocitrate to α-ketoglutarate (αKG). IDH1 mutations, primarily R132H, drive > 80% of low-grade gliomas and secondary glioblastomas and facilitate the NADPH-dependent reduction of αKG to the oncometabolite D-2-hydroxyglutarate (D2HG). While the biochemical features of human WT and mutant IDH1 catalysis have been well-established, considerably less is known about mechanisms of regulation. Proteomics studies have identified lysine acetylation in WT IDH1, indicating post-translational regulation. Here, we generated lysine to glutamine acetylation mimic mutants in IDH1 to evaluate the effects on activity. We show that mimicking lysine acetylation decreased the catalytic efficiency of WT IDH1, with less severe catalytic consequences for R132H IDH1.

Extensive MET alterations confer clinical response to MET inhibitors in gliomas

Activating alterations of the MET gene are well-characterized oncogenic drivers, and MET inhibitors could successfully treat several tumor types with MET alterations, including gliomas with PTPRZ1-MET fusion. However, the full diversity and prevalence of MET alterations in gliomas are still lacking to accurately identify a subset of patients likely to benefit from MET inhibitor treatment. Here, we interrogated genomic profiles of 1,351 gliomas, and further identify 60 cases harboring MET alterations, including MET fusions and various MET exon skipping events. MET RNA alterations, but not MET amplification, are highly enriched in the secondary glioblastomas (sGBM) with significantly worse prognosis. Further molecular analysis has shown that MET RNA alterations acting an additive effects of MET overexpression are induced in the course of glioma evolution. In vitro and clinical studies indicate cells and patients harboring MET alterations have better response to MET inhibitors. Collectively, these data suggest that a subgroup of gliomas harboring MET alterations likely to have benefit from MET-targeted therapy.

PATH-23. GENOMIC LANDSCAPE OF IDH-MUTANT PRIMARY GLIOBLASTOMAS SHOWS DISTINCT CLINICAL AND MOLECULAR FEATURES AND THAT CDKN2A SHOULD BE SUPPLEMENTED WITH MGMTp AND G-CIMP FOR PRECISE PROGNOSTICATION

There have only been rare studies of IDH-mutant primary glioblastomas (IDH-mutant astrocytoma IV); there were one or two studies on secondary glioblastomas. In a cohort of 70 cases, we conducted clinical analysis, methylation profiling, RNA sequencing, targeted sequencing, and TERTp seqeuncing on available FFPE tissues. Median follow-up was 58.2 months (n= 60). IDH-mutant primary glioblastomas had longer median OS (30.4 months) and median PFS (25.9 months) than IDH-mutant secondary glioblastomas as in the literature or established databases. MGMTp methylated cases had better OS (p= 0.001) and it was an independent prognosticator. We previously showed G-CIMP to be an independent prognostic marker for IDH-mutant glioblastomas (NOA 2019). Although CDKN2A deletion was an independent prognostic marker for poorer OS (p= 0.036) and PFS (p= 0.005), MGMTp methylation had a trend of superseding CDKN2A deletion (p= 0.055) for prognostication and G-CIMP subgroups could similarly partially supersede CDKN2A deletion (p= 0.582). Hence, CDKN2A deletion should be supplemented with these two biomarkers for finer prognostication. Targeted sequencing (n= 55) showed that there were more ATRX (35/55, 64%), TP53 (31/55, 56%), KMT2D (18/55, 33%), POLE (11/55, 20%) and MSH6 (7/55, 13%) mutations, but fewer TERTp (3/69, 4%) and PTEN (1/55, 2%) mutations than IDH-wildtype glioblastomas as from literature and databases. CNVs revealed by methylomes (n= 53) and mutations (n= 55) showed that there were more PDGFRA (amplification: 9/53, 17%, mutation: 10/55, 18%) alterations, but fewer MET (amplification: 3/53, 6%, mutation: 4/55, 7%) alterations and hypermutated (6/55, 11%) cases than IDH-mutant secondary glioblastomas from literature. GISTIC analysis revealed amplifications of CCND2, CDK4, MYC, and PDGFRA, deletions of CDKN2A, RB1, and chromosome 10q to be significant CNVs (q< 0.05). There were few EGFR amplifications (2/53, 4%), which was different from regular glioblastomas. RNA sequencing (n= 42) showed few fusions (4/42, 10%), which was different from IDH-mutant secondary glioblastomas.

Isocitrate Dehydrogenase Mutations in Glioma: Genetics, Biochemistry, and Clinical Indications

Mutations in isocitrate dehydrogenase (IDH) are commonly observed in lower-grade glioma and secondary glioblastomas. IDH mutants confer a neomorphic enzyme activity that converts α-ketoglutarate to an oncometabolite D-2-hydroxyglutarate, which impacts cellular epigenetics and metabolism. IDH mutation establishes distinctive patterns in metabolism, cancer biology, and the therapeutic sensitivity of glioma. Thus, a deeper understanding of the roles of IDH mutations is of great value to improve the therapeutic efficacy of glioma and other malignancies that share similar genetic characteristics. In this review, we focused on the genetics, biochemistry, and clinical impacts of IDH mutations in glioma.

Molecular Characterization of Astrocytoma Progression Towards Secondary Glioblastomas Utilizing Patient-Matched Tumor Pairs

Astrocytomas are primary human brain tumors including diffuse or anaplastic astrocytomas that develop towards secondary glioblastomas over time. However, only little is known about molecular alterations that drive this progression. We measured multi-omics profiles of patient-matched astrocytoma pairs of initial and recurrent tumors from 22 patients to identify molecular alterations associated with tumor progression. Gene copy number profiles formed three major subcluters, but more than half of the patient-matched astrocytoma pairs differed in their gene copy number profiles like astrocytomas from different patients. Chromosome 10 deletions were not observed for diffuse astrocytomas, but occurred in corresponding recurrent tumors. Gene expression profiles formed three other major subclusters and patient-matched expression profiles were much more heterogeneous than their copy number profiles. Still, recurrent tumors showed a strong tendency to switch to the mesenchymal subtype. The direct progression of diffuse astrocytomas to secondary glioblastomas showed the largest number of transcriptional changes. Astrocytoma progression groups were further distinguished by signaling pathway expression signatures affecting cell division, interaction and differentiation. As expected, IDH1 was most frequently mutated closely followed by TP53, but also MUC4 involved in the regulation of apoptosis and proliferation was frequently mutated. Astrocytoma progression groups differed in their mutation frequencies of these three genes. Overall, patient-matched astrocytomas can differ substantially within and between patients, but still molecular signatures associated with the progression to secondary glioblastomas exist and should be analyzed for their potential clinical relevance in future studies.

ACTR-51. REMARKABLE RESPONSE OF A PATIENT WITH SECONDARY GBM TO A HISTONE DEACETYLASE INHIBITOR

Glioblastomas are highly aggressive, grade IV tumors of glial cells that arise either as de novo primary tumors or as secondary tumors, which malignantly transformed from lower grade gliomas. Secondary glioblastomas have a relatively low incidence making up 5–10% of all glioblastoma diagnoses and tend to occur in younger patients. However, these tumors are still quite aggressive with survival outcomes that do not differ substantially from primary glioblastomas. Secondary glioblastomas also often harbor mutation of isocitrate dehydrogenase (IDH) enzyme, which produces the oncometabolite 2-hydroxyglutarate (2-HG) from alpha-ketoglutarate (α-KG). The accumulation of 2-HG has several downstream effects due to its competitive inhibition of α-KG-dependent enzymes, including epigenetic modification via hypermethylation of histones. Histone deacetylase inhibitors (HDACi) are a class of molecules that inhibit histone deacetylation and have been shown to have anti-tumor effect in part due to this epigenetic modification. Thus, because of their shared targets with regard to histone modification, it is plausible that HDACi could counter the oncometabolite effects of accumulated 2-HG in IDH1 mutant tumors. Since belinostat is a pan-HDACi that has improved blood-brain barrier penetration compared to many other HDACis, we conducted a pilot study that enrolled 15 patients examining its upfront use for the treatment of glioblastomas and found that it was well tolerated. One patient in particular, likely with secondary glioblastoma harboring the typical IDH1 mutation, underwent treatment with standard chemoradiation as well as belinostat on this study. For this case, a remarkable improvement in tumor burden with very significant decrease in enhancing residual tumor and restoration of magnetic resonance spectroscopy (MRS)-detectable metabolism was noted. Furthermore, improved neurocognitition and quality-of-life were also observed in this patient during the 18-month follow-up period. Collectively, these outcomes potentially support the use of belinostat as an adjuvant therapy for patients with secondary glioblastoma that harbor a mutant IDH enzyme.

329 Primary and Secondary Glioblastomas are Driven by Distinct Forms of Fusion Oncoproteins

INTRODUCTION The role of chromosomal rearrangement in neoplastic transformation has been well-studied in number of cancers. However, the area remains under-studied in high-grade gliomas. METHODS We performed RNA-seq of 272 gliomas, identifying 214 fusion transcripts. Additional review of the literature identified an additional 772 fusion transcripts in the published literature. Analysis was performed pertaining to rearrangement hot-spots and recurrent fusion transcripts. RESULTS >The most common form of fusion transcript arose from joining of sequences from the same chromosome (76%) rather than sequences from difference chromosomes. Frequency of fusion transcript increased with advanced tumor grade. More fusion transcripts are found in the classical subtype of glioblastoma (P = 0.012), particularly in tumors with amplification of EGFR. Fusion transcripts were most commonly mapped to chromosomes 7 and 12, suggesting these chromosomes contain hots-spots for chromosomal rearrangement. For primary glioblastomas, the most prevalent fusion transcripts involved 1) segments of the EGFR sequence fused to other segments of EGFR or to sequences derived from non-EGFR genes (5.6%) or 2) fusion between FGFR and TACC (FGFR3-TACC3 (3.8%) and FGFR1-TACC1 (0.5%)). PTPRZ1-MET fusions are unique in that they are predominantly found in secondary glioblastomas. All three classes of the encoded fusion proteins have been shown to modulate aspects of glioblastoma biology, including tumorigenesis and invasion. CONCLUSION While ∼50% of glioblastoma harbor fusion transcripts, the occurrence of “driver” fusion transcripts is a relative rarity (<5%). Primary and secondary glioblastomas harbor distinct forms of “driver” fusion transcripts.

Isocitrate dehydrogenase status and molecular subclasses of glioma and glioblastoma

Diffuse gliomas and secondary glioblastomas (GBMs) that develop from low-grade gliomas are a common and incurable class of brain tumor. Mutations in the metabolic enzyme glioblastomas (IDH1) represent a distinguishing feature of low-grade gliomas and secondary GBMs. IDH1 mutations are one of the most common and earliest detectable genetic alterations in low-grade diffuse gliomas, and evidence supports this mutation as a driver of gliomagenesis. Here, the authors highlight the biological consequences of IDH1 mutations in gliomas, the clinical and therapeutic/diagnostic implications, and the molecular subtypes of these tumors. They also explore, in brief, the non-IDH1–mutated gliomas, including primary GBMs, and the molecular subtypes and drivers of these tumors. A fundamental understanding of the diversity of GBMs and lower-grade gliomas will ultimately allow for more effective treatments and predictors of survival.