Meta-analysis of whole-genome gene expression datasets assessing the effects of IDH1 and IDH2 mutations in isogenic disease models

Mutations in isocitrate dehydrogenase 1 (IDH1) and IDH2 are oncogenic drivers to a variable extent in several tumors, including gliomas, acute myeloid leukemia (AML), cholangiocarcinoma, melanoma, and thyroid carcinoma. The pathobiological effects of these mutations vary considerably, impeding the identification of common expression profiles. We performed an expression meta-analysis between IDH-mutant (IDHmut) and IDH-wild-type (IDHwt) conditions in six human and mouse isogenic disease models. The datasets included colon cancer cells, glioma cells, heart tissue, hepatoblasts, and neural stem cells. Among differentially expressed genes (DEGs), serine protease 23 (PRSS23) was upregulated in four datasets, i.e., in human colon carcinoma cells, mouse heart tissue, mouse neural stem cells, and human glioma cells. Carbonic anhydrase 2 (CA2) and prolyl 3-hydroxylase 2 (P3H2) were upregulated in three datasets, and SOX2 overlapping transcript (SOX2-OT) was downregulated in three datasets. The most significantly overrepresented protein class was termed intercellular signal molecules. An additional DEG set contained genes that were both up- and downregulated in different datasets and included oxidases and extracellular matrix structural proteins as the most significantly overrepresented protein classes. In conclusion, this meta-analysis provides a comprehensive overview of the expression effects of IDH mutations shared between different isogenic disease models. The generated dataset includes biomarkers, e.g., PRSS23 that may gain relevance for further research or clinical applications in IDHmut tumors.

Isocitrate Dehydrogenase–Mutated Cholangiocarcinoma: Natural History and Clinical Outcomes

Purpose Clinical-pathologic features and natural history of patients with isocitrate dehydrogenase ( IDH)-mutant intrahepatic cholangiocarcinoma (CCA) are not well characterized. Here, we sought to describe the natural history, clinical phenotype, and prognostic impact of advanced, IDH-mutated CCA. METHODS We conducted a multicentric, retrospective analysis of patients with IDH-mutated (IDH1 or IDH2) CCA between 2010 and 2020. Median overall survival (OS) and progression-free survival (PFS) analyses were performed using the Kaplan-Meier method. Chi-square test was used to analyze disease control rate (DCR) and overall response rate (ORR). Matched controls were used for comparing survival between patients with and without IDH mutations (mIDH). RESULTS Sixty-five patients with IDH-mutated CCA were included. All patients had intrahepatic CCA. On first-line chemotherapy, median OS and median PFS were 21.2 months and 8.3 months, respectively. Notably, median OS (32.4 v 19.5 months, P = .12) and PFS (18.0 v 8.0 months, P = .12) were not significantly affected by disease status at presentation (locally advanced v metastatic, respectively). Median OS was significantly longer in patients with mIDH (21.2 v 10.5 months; P < .01). First-line gemcitabine-containing regimens had a significantly higher DCR and ORR than non–gemcitabine-containing regimens (DCR: 75% v 33%, P = .01; ORR: 39% v 0%, P = .02). In patients receiving IDH inhibitor therapy, median PFS was 4.6 months with a DCR of 29%. CONCLUSION CCA with m IDH confers a unique subtype resulting in a better survival compared with that of counterparts. IDH inhibitors represent a promising therapeutic option in later lines of therapy in this subgroup.

Phase I Study of High-Dose l-Methylfolate in Combination with Temozolomide and Bevacizumab in Recurrent IDH Wild-Type High-Grade Glioma

Isocitrate dehydrogenase (IDH) mutations in low-grade gliomas (LGG) result in improved survival and DNA hypermethylation compared with IDH wild-type LGGs. IDH-mutant LGGs become hypomethylated during progression. It is uncertain whether methylation changes occur during IDH wild-type GBM progression and whether the methylome can be reprogrammed. This phase I study evaluated the safety, tolerability, efficacy, and methylome changes after l-methylfolate (LMF) treatment, in combination with temozolomide and bevacizumab in patients with recurrent high-grade glioma. Fourteen patients total, 13 with GBM, one with anaplastic astrocytoma, all IDH wild-type were enrolled in the study. All patients received LMF at either 15, 30, 60, or 90 mg daily plus temozolomide (75 mg/m2 5 days per month) and bevacizumab (10 mg/kg every two weeks).No MTD was identified. LMF-treated patients had median overall survival of 9.5 months [95% confidence interval (CI), 9.1–35.4] comparable with bevacizumab historical control 8.6 months (95% CI, 6.8–10.8). Six patients treated with LMF survived more than 650 days. Across all treatment doses, the most adverse events were diarrhea (7%, 1 patient, grade 2), reflux (7%, 1 patient, grade 2), and dysgeusia (7%, 1 patient, grade 2). In the six brains donated at death, there was a 25% increase in DNA methylated CpGs compared with the paired initial tumor. LMF in combination with temozolomide and bevacizumab was well tolerated in patients with recurrent IDH wild-type high-grade glioma. This small study did not establish a superior efficacy with addition of LMF compared with standard bevacizumab therapy; however, this study did show methylome reprogramming in high-grade glioma. Significance: Glioblastoma (GBM) is a primary brain tumor with a poor prognosis. Therapies to date have failed to improve survival. LGGs, with IDH mutations, have increased global DNA methylation and increased survival compared with GBMs. GBMs lack this mutation and have less DNA methylation. Here we show that the DNA methylome can be modified in GBM with LMF. Such treatment might be useful in methylome priming prior to immunotherapy.

MPC-2 Clinical course and prognosis of lower-grade glioma, IDH wildtype and pTERT mutant

Background and Purpose: In the cIMPACT-Now update 3, it was proposed that grade 2 astrocytic gliomas without IDH-mutations and grade 3 astrocytic gliomas with TERT promoter mutations should be designated as diffuse IDH wildtype astrocytic glioma with molecular features of WHO grade IV glioblastoma. Therefore, we investigated whether this group of tumors actually corresponds to grade IV prognostically in cases that we encountered ourselves. Cases and Methods: Among the 65 patients having primary astrocytic glioma who were operated in our hospital from January 2016 to March 2021, the prognostic values of seven patients with lower-grade glioma, IDH wildtype, and pTERT mutant were investigated. Results: Among the seven patients, the median age was 59 years (50–66 years). Four of them had anaplastic astrocytoma, two had diffuse astrocytoma, and no tumor lesion could be identified upon histological examination for one patient. The male-to-female ratio was 1:6. MGMT methylation was observed in two patients (29%). The median survival was 20 months, with a significantly worse prognosis when compared with lower-grade glioma without the TERT promoter mutation (13 patients: median survival 40 months), but a better prognosis when compared with glioblastoma (45 patients: median survival 13 months) (Log-rank p = 0.0051). Conclusion: Although EGFR amplification, combined whole chromosome 7 gain, and whole chromosome 10 loss were not examined, the prognostic value of lower-grade glioma, IDH wildtype, and pTERT mutant was not as poor as that of glioblastoma. Further investigation is required to confirm whether these groups of tumors should be treated in the same way as grade IV glioblastoma.

Clinical, Morphological, and Molecular Study of Diffuse WHO Grade II and III Astrocytomas: A Retrospective Analysis from a Single Tertiary Care Institute

Introduction Astrocytomas are the most common gliomas, classified on the basis of grade and IDH mutation status according to the World Health Organization (WHO) 2016 update. IDH mutations are seen in 70 to 80% of diffuse grade II and III astrocytomas and are associated with better outcome. They serve as predictive biomarker in IDH-targeted therapies such as small-molecule inhibitors or vaccines. Objective The aim of this study was to analyze the clinical, morphological, immunohistochemical, and molecular genetic characteristics of diffuse astrocytoma (DA: grades II and III). The IDH mutant and wild-type tumors are compared and contrasted with survival analysis on follow-up. Materials and Methods This was a retrospective study conducted on surgically resected tumor specimens. The hematoxylin and eosin-stained slides were examined for histologic features. Immunohistochemistry (IHC) was performed using IDH1R132H, ATRX, p53, and Ki67. All cases of negative immunohistochemical expression of IDH1R132H were subjected to IDH1 mutation analysis by Sanger sequencing. Overall survival was estimated by the Kaplan-Meier method using the log-rank (Mantel–Cox) test. Results The study included 51 cases of DA in the age of 17 to 66 years, mean ± standard deviation was 35.5 ± 9.7 years, and male:female ratio was 2:1.The IDH1R132H cytoplasmic immunopositivity was seen in 36 cases (70.5%), of which 63.6% were of grade II and 72.5% were of grade III. ATRX showed loss of expression in 50 cases (98%), and p53 showed diffuse strong immunohistochemical expression in all the cases of IDH mutant tumors. The difference in the age at presentation for IDH mutant (32.5 years) and wild type tumors (38 years) was statistically significant. Median survival was 55.3 months and 22.2 months in of IDH mutant and wild type cases, respectively. Conclusion IHC and sequencing for IDH mutations is helpful in making an integrated diagnosis and classifying definite molecular subgroups of astrocytic tumors. Mutations in IDH core-elate with survival. IDH mutant tumors showed longer survival duration and are good prognostic indicators.

ACT-5 Prognosis of IDH-mut lower-grade gliomas in Hokkaido University Hospital

Background: WHO grade 2 and 3 adult gliomas are nowadays getting together as lower-grade gliomas (LrGGs), but we had been recognized grade 3 (G3) tumors as high-grade and grade 2 (G2) tumors as low-grade. In this report, we investigate the treatment and prognosis of the patients with LrGG harboring IDH mutations in our institutions. Methods:We retrospectively review primary treatments and their prognosis for LrGG patients with IDH mutation since 2003. They categorized as astrocytomas and oligodendrogliomas according to 1p/19q loss-of-heterozygosity status. Prognosis were evaluated by overall survival. Postoperative primary treatments applied chemo-radiotherapy (CRT), radiotherapy only (RT), chemotherapy only (CT), and observation (Ob). Results: 36 astrocytomas and 60 oligodendrogliomas were identified. In astrocytomas, the patients with G3 (N=16) were treated by CRT (N=14) or CT (N=2), and the patients with G2 (N=20) were treated by CRT (N=2), RT (N=3), CT (N=3), or Ob (N=12). In oligodendrogliomas, the patients with G3 (N=34) were treated by CRT (N=32) or CT (N=2), and the patients with G2 (N=26) were treated by CRT (N=3), RT (N=1), CT (N=5), or Ob (N=17). 10-year survival rate (10yOS) of astrocytomas and oligodendrogliomas are 54% and 90%, respectively (p=0.002). According to histological malignancy, 10yOS of G3 and G2 astrocytomas were 54% and 54%, respectively (p=0.97) and that of G3 and G2 oligodendrogliomas were 86% and 100%, respectively (p=0.64). In both group, there are no different of prognosis according to histological malignancy. Discussion: There was no prognostic different between G2 and G3 astrocytomas in our institution. Since the treatment intensity for G2 and G3 astrocytomas were clearly different, the primary treatment for G2 astrocytomas might be insufficient. On the other hand, there were no prognostic different between G2 and G3 oligodendrogliomas in our institution, as with recent reports, so the primary treatment intensity for oligodendrogliomas should be appropriate.

MEOX2 Transcription Factor Is Involved in Survival and Adhesion of Glioma Stem-like Cells

The high expression of MEOX2 transcription factor is closely associated with poor overall survival in glioma. MEOX2 has recently been described as an interesting prognostic biomarker, especially for lower grade glioma. MEOX2 has never been studied in glioma stem-like cells (GSC), responsible for glioma recurrence. The aim of our study was to investigate the role of MEOX2 in GSC. Loss of function approach using siRNA was used to assess the impact of MEOX2 on GSC viability and stemness phenotype. MEOX2 was localized in the nucleus and its expression was heterogeneous between GSCs. MEOX2 expression depends on the methylation state of its promoter and is strongly associated with IDH mutations. MEOX2 is involved in cell proliferation and viability regulation through ERK/MAPK and PI3K/AKT pathways. MEOX2 loss of function correlated with GSC differentiation and acquisition of neuronal lineage characteristics. Besides, inhibition of MEOX2 is correlated with increased expression of CDH10 and decreased pFAK. In this study, we unraveled, for the first time, MEOX2 contribution to cell viability and proliferation through AKT/ERK pathway and its potential involvement in phenotype and adhesion properties of GSC.

Detection of 2-Hydroxyglutarate by 3.0-Tesla Magnetic Resonance Spectroscopy in Gliomas with Rare IDH Mutations: Making Sense of “False-Positive” Cases

We have previously published a study on the reliable detection of 2-hydroxyglutarate (2HG) in lower-grade gliomas by magnetic resonance spectroscopy (MRS). In this short article, we re-evaluated five glioma cases originally assessed as isocitrate dehydrogenase (IDH) wildtype, which showed a high accumulation of 2HG, and were thought to be false-positives. A new primer was used for the detection of IDH2 mutation by Sanger sequencing. Adequate tissue for DNA analysis was available in 4 out of 5 cases. We found rare IDH2 mutations in two cases, with IDH2 R172W mutation in one case and IDH2 R172K mutation in another case. Both cases had very small mutant peaks, suggesting that the tumor volume was low in the tumor samples. Thus, the specificity of MRS for detecting IDH1/2 mutations was higher (81.3%) than that originally reported (72.2%). The detection of 2HG by MRS can aid in the diagnosis of rare, non-IDH1-R132H IDH1 and IDH2 mutations in gliomas.

Differential Prognostic Impact of IDH1 and IDH2 Mutations in Chronic Myelomonocytic Leukemia

Introduction: Mutations involving isocitrate dehydrogenase 1/2 (IDH) are known oncogenic drivers in hematological malignancies, conferring neomorphic enzymatic activity to IDH 1/2, resulting in the oncometabolite, 2-hydroxyglutarae (2-HG). 2-HG in turn suppresses TET activity, making IDH and TET2 mutations synthetically lethal and almost mutually exclusive. The frequency of IDH mutations in CMML is &lt;10% and their prognostic impact remains unclear. We carried out this study in a large database of molecularly annotated CMML patients to better define the clinical profile and prognostic impact of these mutations. Methods: After IRB approval, CMML patients from the Mayo Clinic, Minnesota and the Moffitt Cancer Center (MCC), Tampa, Florida, were included in the study. All patients had bone marrow (BM) biopsies with cytogenetics and molecular genetics done either at diagnosis, or at first referral. Clinical and mutational data were abstracted and retrospectively analyzed. Overall survival (OS) was calculated from date of CMML diagnosis to date of death/last follow, while AML-free survival (AML-FS) was calculated from date of CMML diagnosis to date of leukemic transformation (LT). Patients that had undergone allogeneic HCT were excluded from the study (n=3). Statistical analysis was carried out using the Blue Sky software. Results: Six hundred and forty four patients were included in the study (Mayo Clinic-357, MCC- 287), median age 71 years (range, 20-95 years), 67.8% being male. Forty-three (6.7%) patients had IDH mutations, 35 (82%) IDH2 and 8 (18%) IDH1; of which, 34 (97%) involved the IDH2R140 hotspot and 5 (62.5%) involved the IDH1R132 hotspot, respectively. The median variant allele fractions (VAF) for IDH1 mutations was 41% (range, 8-46%) and for IDH2 mutations was 46% (range, 7-70%). There were no significant demographic or clinical differences between IDH mutant and wild type CMML patients, with the exception that IDH mutant CMML patients were less likely to be thrombocytopenic (p=0.006), were less likely to have TET2 co-mutations (14% vs 53.2%; p&lt;0.001) and were more likely to have SRSF2 co-mutations (69.8% VS 40.3%; p&lt;0.001). Importantly there were no differences in proliferative or dysplastic subtypes (p=0.3), cytogenetic (p=0.12) and molecular risk stratifications (p=045). There were also no significant demographic or clinical differences between IDH1 vs IDH2 mutant CMML patients. Six (14%) IDH mutant CMML patients had TET2 co-mutations; 5 (83%) with IDH2R140Q (median VAF-28%;all male) and 1 (17%) with IDH1R132H (VAF-44%;female) (Figure 1). Five (11%) IDH2 mutant patients were treated with enasidenib (IDH2 inhibitor), none with a durable response, while none of the IDH1 mutant patients received targeted therapy. At last follow up (median 18 months), 337 (52%) deaths and 119 (18.5%) LT have been documented, with IDH mutant patients having a higher LT rate (30.2% vs 17.6%, p=0.04) compared to wildtype patients. The median OS of the entire cohort was 35 months, with no difference in OS between IDH mutant and wild type patients (34.5 vs 35 months, p=0.12), with IDH1 mutant patients having a shorter OS in comparison to IDH2 mutant patients (31 vs 37 months; p=0.005- Figure 1). IDH mutant CMML patients also had a shorter AML-FS in comparison to wild type patients (36.6 vs 210 months, p=0.005), with there being no differential impact on AML-FS of IDH1 vs IDH2 mutations (p=0.26, Figure 1). Conclusions: IDH mutations are infrequent in CMML (7%), with IDH2 mutations being more common than IDH1 mutations (80 vs 20%). IDH mutations co-occur very infrequently with TET2 mutations (14%), with IDH mutant patients being less likely to have thrombocytopenia and more likely to have SRSF2 co-mutations. IDH mutations negatively impacting AML-FS without a significant impact on OS. Prospective clinical trials testing the safety and efficacy of IDH1/2 inhibitors in CMML are much needed. Figure 1 Figure 1. Disclosures Komrokji: AbbVie: Consultancy; PharmaEssentia: Membership on an entity's Board of Directors or advisory committees; Taiho Oncology: Membership on an entity's Board of Directors or advisory committees; Novartis: Consultancy, Honoraria, Membership on an entity's Board of Directors or advisory committees; Jazz: Consultancy, Speakers Bureau; Acceleron: Consultancy; BMSCelgene: Consultancy, Membership on an entity's Board of Directors or advisory committees, Speakers Bureau; Geron: Consultancy. Al-Kali: Novartis: Research Funding; Astex: Other: Research support to institution. Padron: BMS: Research Funding; Stemline: Honoraria; Taiho: Honoraria; Kura: Research Funding; Incyte: Research Funding; Blueprint: Honoraria. Patnaik: StemLine: Research Funding; Kura Oncology: Research Funding.