The effects of BMP2 and the mechanisms involved in the invasion and angiogenesis of IDH1 mutant glioma cells.

PURPOSE: This study investigated the effect of an isocitrate dehydrogenase 1 (IDH1) mutation (mutIDH1) on the invasion and angiogenesis of human glioma cells. METHODS: Doxycycline was used to induce the expression of mutIDH1 in glioma cells. Transwell and wound healing assays were conducted to assess glioma cell migration and invasion. Western blotting and cell immunofluorescence were used to measure the expression levels of various proteins. The influence of bone morphogenetic protein 2 (BMP2) on invasion, angiogenesis-related factors, BMP2-related receptor expression, and changes in Smad signaling pathway-related proteins were evaluated after treatment with BMP2. Differential gene expression and reference transcription analysis were performed. RESULTS: Successful infection with recombinant lentivirus expressing mutIDH1 was demonstrated. The IDH1 mutation promoted glioma cell migration and invasion while positively regulating the expression of vascularization-related factors and BMP2-related receptors. BMP2 exhibited a positive regulatory effect on the migration, invasion, and angiogenesis of mutIDH1-glioma cells, possibly mediated by BMP2-induced alterations in Smad signaling pathway-related factors.After BMP2 treatment, the differential genes of MutIDH1-glioma cells are closely related to the regulation of cell migration and cell adhesion, especially the regulation of Smad-related proteins. KEGG analysis confirmed that it was related to BMP signaling pathway and TGF-ß signaling pathway and cell adhesion. Enrichment analysis of gene ontology and genome encyclopedia further confirmed the correlation of these pathways. CONCLUSION: Mutation of isocitrate dehydrogenase 1 promotes the migration, invasion, and angiogenesis of glioma cells, through its effects on the BMP2-driven Smad signaling pathway. In addition, BMP2 altered the transcriptional patterns of mutIDH1 glioma cells, enriching different gene loci in pathways associated with invasion, migration, and angiogenesis.

A perioperative study of Safusidenib in patients with IDH1-mutated glioma.

This is a single arm, open label perioperative trial to assess the feasibility, pharmacokinetics and pharmacodynamics of treatment with safusidenib following biopsy, and prior to surgical resection in patients with IDH1 mutated glioma who have not received radiation therapy or chemotherapy. Fifteen participants will receive treatment in two parts. First, biopsy followed by one cycle (28 days) of safusidenib, an orally available, small molecular inhibitor of mutated IDH1, then maximal safe resection of the tumor (Part A). Second, after recovery from surgery, safusidenib until disease progression or unacceptable toxicity (Part B). This research will enable objective measurement of biological activity of safusidenib in patients with IDH1 mutated glioma. Anti-tumor activity will be assessed by progression free survival and time to next intervention.Clinical Trial Registration: NCT05577416 (ClinicalTrials.gov).

Adult low-grade gliomas (aLGG) are primary brain cancers, defined by mutations in IDH1 or IDH2. When the IDH gene becomes abnormal (mutated), production of a metabolite that causes cancer cells to grow is increased. These tumors grow slowly but invade the normal functioning brain, making them nearly impossible to cure. The current standard of care treatment includes surgery, followed by radiation therapy and chemotherapy, the timing of which depends on the risk of cancer regrowth. Some patients may be suitable for monitoring with MRI scans alone, however recurrences will inevitably occur. Recently developed targeted mutant IDH inhibitors for aLGG patients may be beneficial both at diagnosis and recurrence. Notably, early treatment prior to radiation therapy and chemotherapy delays growth of aLGG and the need for subsequent radiation therapy and chemotherapy. Nevertheless, most patients will eventually suffer further tumor growth and the optimal timing and sequencing of these therapies remains an area of active research. This research investigates the mutant IDH1 inhibitor safusidenib. The researchers are conducting an innovative clinical trial where patients with aLGG, who have not received radiation therapy or chemotherapy, are treated with safusidenib following a biopsy and prior to surgical removal of their tumor. In this study they investigate whether this trial design is safe and feasible, and how safusidenib works; with the goal to better understand the optimal use of IDH inhibitors for patients with aLGG.

IDH1 and IDH2 Gene Mutations in Omani Patients with Acute Myeloid Leukemia: Prognostic Significance and Clinic-pathologic Features.

Objectives: We sought to define the prevalence of isocitrate dehydrogenase (IDH) mutations, evaluate the clinicopathologic impact of IDH mutations, assess the effect of IDH mutations on the response to the currently offered treatment for acute myeloid leukemia (AML) cases, and determine the impact of other common concurrent mutations with IDH. Methods: A single-center retrospective cohort study was conducted at Sultan Qaboos University Hospital (SQUH) from October 2009 to October 2019. We included all Omani patients (pediatric and adult) treated at SQUH with the standard therapy, for whom DNA extraction was performed at diagnosis. The target mutations in both IDH1 and IDH2 genes were screened using the direct polymerase chain reaction product sequencing method. Statistical analysis was conducted using SPSS software. Survival differences were estimated using the log-rank test. Continuous variables were presented as median (IQRs), while categorical variables were presented as frequency. Results: A total of 61 patients treated, for whom DNA extraction was performed at diagnosis were evaluated. The median age was 40 (range = 25.5-65.5). The prevalence of IDH1 R132, IDH2 R140, and IDH2 R172 mutations among the study group was 6.6%, 3.3%, and 1.6%, respectively. Clinicopathologic characteristics associated with IDH mutations at diagnosis included older age, lower white blood cell count, higher median platelet counts, normal karyotype AML, and cytogenetics intermediate-risk group. The overall survival (OS) in patients harboring IDH mutations was poor, with a median OS of nine months. This analysis confirms that the response rate and OS for both IDH-mutated and IDH wild-type AML patients were comparable. This will provide contemporary data to be used for comparison with the results of novel investigational (e.g., selective IDH inhibitor) strategies. Conclusions: The current study results were consistent with the other international studies of IDH mutations in AML and demonstrate the poor prognosis associated with IDH mutations. Clinicopathologic features associated with IDH mutations included older age, lower white blood cell count, higher median platelet counts, normal karyotype AML, and cytogenetics intermediate-risk group.

Withaferin A Inhibits Liver Cancer Tumorigenesis by Suppressing Aerobic Glycolysis through the p53/IDH1/HIF-1α Signaling Axis.

BACKGROUND: The energy supply of certain cancer cells depends on aerobic glycolysis rather than oxidative phosphorylation. Our previous studies have shown that withaferin A (WA), a lactone compound derived from Withania somnifera, suppresses skin carcinogenesis at least partially by stabilizing IDH1 and promoting oxidative phosphorylation. Here, we have extended our studies to evaluate the anti-tumor effect of WA in liver cancer. METHODS: Differential expression of glycolysis-related genes between liver cancer tissues and normal tissues and prognosis were verified using an online database. Glycolysis-related protein expression was detected using western blot after overexpression and knockdown of IDH1 and mitochondrial membrane potential assay based on JC-1, and mitochondrial complex I activity was also detected. The inhibitory effect of WA on the biological functions of HepG2 cells was detected along with cell viability using MTT assay, scratch assay, clone formation assay, glucose consumption and lactate production assay. Western blot and qRT-PCR were used to detect the expression of proteins and genes related to IDH1, p53 and HIF1α signaling pathways. RESULTS: We first identified that IDH1 expression was downregulated in human liver cancer cells compared to normal liver cells. Next, we found that treatment of HepG2 cells with WA resulted in significantly increased protein levels of IDH1, accompanied by decreased levels of several glycolytic enzymes. Furthermore, we found that WA stabilized IDH1 proteins by inhibiting the degradation by the proteasome. The tumor suppressor p53 was also upregulated by WA treatment, which played a critical role in the upregulation of IDH1 and downregulation of the glycolysis-related genes. Under hypoxic conditions, glycolysis-related genes were induced, which was suppressed by WA treatment, and IDH1 expression was still maintained at higher levels under hypoxia. CONCLUSION: Taken together, our results indicated that WA suppresses liver cancer tumorigenesis by p53-mediated IDH1 upregulation, which promotes mitochondrial respiration, thereby inhibiting the HIF-1α pathway and blocking aerobic glycolysis.

Efficacy of Personalized Postoperative Epilepsy Management in Patients with Glioblastoma Utilizing IDH1 Gene Assessment.

Objective: We explored the correlation between the presence of isocitrate dehydrogenase-1 (IDH1) mutations and the incidence of postoperative epilepsy in patients with glioblastoma, as well as assessed the efficacy of preemptive administration of antiepileptic medications in mitigating the occurrence of postoperative epilepsy. Methods: Fifty-three patients who received a postoperative pathological diagnosis of glioblastoma, were enrolled in this study. Tumor specimens were subjected to IDH1 gene analysis. The patient cohort was stratified based on their IDH1 mutation status and the administration of prophylactic antiepileptic drugs during the postoperative phase. We subsequently conducted a comparative analysis of postoperative epileptic complications within each patient subgroup. Results: In the cohort of 53 patients under study, the occurrence of epilepsy was observed in 10 out of 21 patients carrying IDH1 mutations, while 5 out of 32 patients with wild-type IDH1 also experienced epilepsy, revealing a statistically significant difference (P < 0.05). Among the 27 patients who received prophylactic antiepileptic drugs, 6 of them developed epilepsy, whereas 9 out of 26 patients who did not receive prophylactic antiepileptic drugs exhibited concurrent epilepsy, with no statistically significant difference (P > 0.05). However, when performing a subgroup analysis, it was found that 3 out of 12 patients with IDH1 mutations who received prophylactic antiepileptic drugs experienced epilepsy, whereas 7 out of 9 patients who did not receive prophylactic antiepileptic drugs developed epilepsy, demonstrating a statistically significant difference (P < 0.05). Furthermore, within the group of 15 patients with wild-type IDH1, 3 patients who received prophylactic antiepileptic drugs developed epilepsy, while 2 cases of epilepsy occurred among the 17 patients who did not receive prophylactic antiepileptic drugs, with no statistically significant difference (P > 0.05). Conclusion: In individuals with IDH1 mutant glioblastoma who have undergone surgical resection, the implementation of preventive antiepileptic therapy demonstrates a potential to diminish the occurrence of postoperative epilepsy.

Suppression of antitumor immune signatures and upregulation of VEGFA as IDH-mutant gliomas progress to higher grade.

OBJECTIVE: Several studies have compared the immune microenvironment of isocitrate dehydrogenase (IDH)-wildtype glioma versus IDH-mutant glioma. The authors sought to determine whether histological tumor progression in a subset of IDH-mutant glioma was associated with concomitant alterations in the intratumoral immune microenvironment. METHODS: The authors performed bulk RNA sequencing on paired and unpaired samples from patients with IDH-mutant glioma who underwent surgery for tumor progression across multiple timepoints. They compared patterns of differential gene expression, overall inflammatory signatures, and transcriptomic measures of relative immune cell proportions. RESULTS: A total of 55 unique IDH-mutant glioma samples were included in the analysis. The authors identified multiple genes associated with progression and higher grade across IDH-mutant oligodendrogliomas and astrocytomas. Compared with lower-grade paired samples, grade 4 IDH-mutant astrocytomas uniquely demonstrated upregulation of VEGFA in addition to counterproductive alterations in inflammatory score reflective of a more hostile immune microenvironment. CONCLUSIONS: Here, the authors have provided a transcriptomic analysis of a progression cohort for IDH-mutant glioma. Compared with lower-grade tumors, grade 4 astrocytomas displayed alterations that may inform the timing of antiangiogenic and immune-based therapy as these tumors progress.

The biological function and mechanism of IDH1 gene in intrahepatic cholangiocarcinoma cell HuCCT1 / 解放军医学杂志

Objective To explore the role and possible molecular mechanism of Isocitrate dehydrogenase 1(IDH1)gene in proliferation and migration of intrahepatic cholangiocarcinoma(iCCA)cell HuCCT1.Methods HuCCT1 cells with IDH1 gene knockout(HuCCT1IDH1-/-)were constructed by CRISPR/Cas9 gene editing technology.To investigate the capacities of proliferation,migration and invasion of HuCCT1WT(HuCCT1 cells with wild-type IDH1 gene)and HuCCT1IDH1-/-cells,assays of CCK-8,clone formation,scratch and transwell were performed.Western blotting was used to detect the expression levels of epithelial-mesenchymal transition(EMT)associated proteins E-cadherin,N-cadherin,Vimentin,MMP-9,Wnt3a and β-catenin in two groups of cells.The transcriptome sequencing data of HuCCT1WT and HuCCT1IDH1-/-cells were analyzed by bioinformatics methods,Western blotting was used to verify the expression of signaling pathway-related proteins.Results Compared with HuCCT1WT cells,HuCCT1IDH1-/-cells showed the number of proliferation and clone formation significantly reduced(P<0.05),the proportion of cells blocked in G2/M phase was significantly increased(P<0.01),the rate of scratch healing was significantly decreased(P<0.01),and the number of migrated cells(P<0.001)and invaded cells(P<0.05)was significantly reduced.qRT-PCR assay showed that the expression levels of IDH1,Vimentin,MMP-9 and genes related to the regulation of G2/M cycle proliferation,Cyclin A2,Cyclin B1 and CDK1 mRNA were down-regulated in HuCCT1IDH1-/-cells(P<0.05),and the expression of CDH1 mRNA encoding E-cadherin was up-regulated(P<0.01);Western blotting assay showed that the expression level of E-cadherin in HuCCT1IDH1-/-cells was significantly increased(P<0.05),and the expression level of N-cadherin,Vimentin and MMP-9 protein was significantly decreased(P<0.05)than that in HuCCT1WT cells.Data of transcriptome sequencing revealed 1476 differentially expressed genes(DEGs)between two groups of HuCCT1 cells.Go enrichment analysis showed the DEGs were significantly enriched in cell biological processes associated with inflammatory response,cell signaling and cell metabolism.Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway analysis suggested that the DEGs may be involved in some signaling pathways such as Wnt,MAPK,Rap1,Hippo and TNF,which are closely related to the regulation of proliferation and invasion of tumor cells.Western blotting verification results showed that compared with HuCCT1WT cells,the relative expression of Wnt3a and β-catenin proteins of HuCCT1IDH1-/-cells was significantly decreased(P<0.05).Conclusions IDH1 gene may participate in the control of biological functions of HuCCT1 cells,including cell proliferation,migration,invasion and epithelial mesenchymal transition.The mechanism may be related to the activation of the Wnt/β-catenin signaling pathway.

Biomarkers in Adult-Type Diffuse Gliomas: Elevated Levels of Circulating Vesicular Heat Shock Protein 70 Serve as a Biomarker in Grade 4 Glioblastoma and Increase NK Cell Frequencies in Grade 3 Glioma.

The presence of circulating Hsp70 levels and their influence on the immunophenotype of circulating lymphocyte subsets were examined as diagnostic/prognostic biomarkers for the overall survival (OS) in patients with IDH-mutant WHO grade 3 oligodendroglioma, astrocytoma, and IDH-wildtype grade 4 glioblastoma (GBM). Vesicular and free Hsp70 in the plasma/serum was measured using the Hsp70-exo and R&D Systems DuoSet® Hsp70 ELISAs. The immunophenotype and membrane Hsp70 status was determined by multiparameter flow cytometry on peripheral blood lymphocytes and single-cell suspensions of tumor specimens and cultured cells. Compared to healthy controls, circulating vesicular Hsp70 levels were significantly increased in patients with GBM, concomitant with a significant decrease in the proportion of CD3+/CD4+ helper T cells, whereas the frequency of NK cells was most prominently increased in patients with grade 3 gliomas. Elevated circulating Hsp70 levels and a higher prevalence of activated CD3-/CD56+/CD94+/CD69+ NK cells were associated with an improved OS in grade 3 gliomas, whereas high Hsp70 levels and low CD3+/CD4+ frequencies were associated with an adverse OS in GBM. It is assumed that a reduced membrane Hsp70 density on grade 4 versus grade 3 primary glioma cells and reduced CD3+/CD4+ T cell counts in GBM might drive an immunosuppressive tumor microenvironment.

Prognostic Value of Immunohistochemical Expression of MTAP and AKIP1 in IDH1 Mutant Astrocytoma.

BACKGROUND: Definite treatment for glioma is not exist, and with increased drug resistance, more effort should be paid to identify new prognostic biomarkers and molecular targets for therapy for glioma patients. AIM: The current study aimed to evaluate the immunohistochemical (IHC) expression of MTAP and A-Kinase Interacting Protein 1 (AKIP1) in astrocytoma and to investigate their association with the clinicopathological characters of these cases. METHODS: Totally 66 cases of astrocytoma patients involved in this study. Cases underwent tumor resection and tissue sections were stained with MTAP, AKIP1 and IDH1 by IHC and evaluated in different grades of astrocytoma and their association with survival and response to therapy was investigated. RESULTS: High AKIP1 expression was positively correlated with treatment resistance and progressive disease. Positive IDH and retained MTAP expressions had shown better treatment response rather than negative IDH and lost MTAP. High AKIP, negative IDH and loss of MTAP expressions were significantly associated with poor survival outcome. CONCLUSION: Irrespective to grade and IDH status, the loss of MTAP immunoreactivity and high AKIP1 expression are predictive factors in astrocytoma, and they may be used as a biomarker for guiding astrocytoma management and prognosis surveillance.

An integrative computational approach for the identification of dual inhibitors of isocitrate dehydrogenase 1 and 2 from phytocompounds of Phyllantus amarus.

Genetic alterations of the genes encoding the isocitrate dehydrogenase (IDH) enzymes have been identified in about 20% of acute myeloid leukemia (AML) cases as well as many other forms of cancers. Notable among these alterations are the neomorphic IDH1_R132H and IDH2_R140Q mutations which lead to the production of an oncometabolite. Hence, their inhibition is widely considered a therapeutic strategy in the treatment of many cancers. While many inhibitors of the mutant enzymes have been developed, an inhibitor that is capable of co-inhibiting both enzymes are currently lacking while drug resistance has also limited the clinical usage of previously identified mono inhibitors. Consequently, this study employed molecular modeling approaches, such as molecular docking, molecular mechanics generalized Born Surface area (MM/GBSA), molecular dynamics (MD) simulation, and density functional theory (DFT) analysis to identify potential dual inhibitors of the previously mentioned mutant IDH1/2 from the phytocompounds of Phyllantus amarus. Of the 31 phytocompounds identified, 20 showed good binding affinities for both IDH1 _R132H and IDH2 _R140Q (ranging from -5.2 Kca/mol to -9.6 Kcal/mol) and had desirable pharmacokinetic properties. However, ellagic acid and pinoresinol possessed better pharmacokinetic properties, rendering suitable hits. Investigation of the behavior of the IDH1_R132H and IDH2_R140Q complexes with ellagic acid and pinoresinol via the RMSD, RMSF, and contact map analyses showed that all the complexes-maintained stability throughout the simulation time. Ultimately, ellagic acid and pinoresinol were identified as promising hits for the development of IDH1_R132H and IDH2_R140Q dual inhibitors. However, further experimental studies are needed to confirm their potential as therapies.Communicated by Ramaswamy H. Sarma.

Paradigm shift of chemotherapy and systemic treatment for biliary tract cancer.

Biliary tract cancers (BTC) are frequently identified at late stages and have a poor prognosis due to limited systemic treatment regimens. For more than a decade, the combination of gemcitabine and cis-platin has served as the first-line standard treatment. There are few choices for second-line chemo-therapy. Targeted treatment with fibroblast growth factor receptor 2 inhibitors, neurotrophic tyrosine receptor kinase inhibitors, and isocitrate dehydrogenase 1 inhibitors has had important results. Immune checkpoint inhibitors (ICI) such as pembrolizumab are only used in first-line treatment for microsatellite instability high patients. The TOPAZ-1 trial's outcome is encouraging, and there are several trials underway that might soon put targeted treatment and ICI combos into first-line options. Newer targets and agents for existing goals are being studied, which may represent a paradigm shift in BTC management. Due to a scarcity of targetable mutations and the higher toxicity profile of the current medications, the new category of drugs may occupy a significant role in BTC therapies.

Exceptionally rare IDH1-mutant adult medulloblastoma with concurrent GNAS mutation revealed by in vivo magnetic resonance spectroscopy and deep sequencing.

Medulloblastoma (MB) is the most common malignant brain tumor occurring in childhood and rarely found in adults. Based on transcriptome profile, MB are currently classified into four major molecular groups reflecting a considerable biological heterogeneity: WNT-activated, SHH-activated, group 3 and group 4. Recently, DNA methylation profiling allowed the identification of additional subgroups within the four major molecular groups associated with different clinic-pathological and molecular features. Isocitrate dehydrogenase-1 and 2 (IDH1 and IDH2) mutations have been described in several tumors, including gliomas, while in MB are rarely reported and not routinely investigated. By means of magnetic resonance spectroscopy (MRS), we unequivocally assessed the presence the oncometabolite D-2-hydroxyglutarate (2HG), a marker of IDH1 and IDH2 mutations, in a case of adult MB. Immunophenotypical work-up and methylation profiling assigned the diagnosis of MB, subclass SHH-A, and molecular testing revealed the presence of the non-canonical somatic IDH1(p.R132C) mutation and an additional GNAS mutation, also rarely described in MB. To the best of our knowledge, this is the first reported case of MB simultaneously harboring both mutations. Of note, tumor exhibited a heterogeneous phenotype with a tumor component displaying glial differentiation, with robust GFAP expression, and a component with conventional MB features and selective presence of GNAS mutation, suggesting co-existence of two different major tumor subclones. These findings drew attention to the need for a deeper genetic characterization of MB, in order to get insights into their biology and improve stratification and clinical management of the patients. Moreover, our results underlined the importance of performing MRS for the identification of IDH mutations in non-glial tumors. The use of throughput molecular profiling analysis and advanced medical imaging will certainly increase the frequency with which tumor entities with rare molecular alterations will be identified. Whether these findings have any specific therapeutic implications or prognostic relevance requires further investigations.

Augmenting Venetoclax Activity Through Signal Transduction in AML.

Venetoclax, a small-molecule B-cell lymphoma 2 (BCL-2) inhibitor, selectively eradicates leukemic stem cells (LSCs). While venetoclax has revolutionized the treatment of acute myeloid leukemia (AML), treatment failure and disease relapse are common. Mechanisms underlying venetoclax resistance are surprisingly heterogeneous. Venetoclax resistance encompasses a spectrum of genetic and epigenetic changes, with numerous pathways contributing to the upregulation of additional anti-apoptotic proteins. In this review, we address the mechanisms of venetoclax resistance in the context of signal transduction. We emphasize how aberrant cell signaling impairs apoptosis and predisposes to venetoclax failure. Commonly activated pathways, such as FLT3, PI3K/AKT/mTOR, and RAS, contribute to upregulated anti-apoptotic mediators and are frequently responsible for refractory disease or disease relapse. We highlight novel combination strategies aimed at disabling constitutively active signal transduction to augment response and overcome venetoclax resistance.

Urinary 8-hydroxy-2'-deoxyguanosine levels are elevated in patients with IDH1-wildtype glioblastoma and are associated with tumor recurrence in gliomas.

2021 World Health Organization (WHO) Central Nervous System (CNS) Tumor Classification includes molecular diagnostic parameters such as isocitrate dehydrogenase (IDH) mutation or 1p19q codeletion status, in addition to the classical histological classification. Several studies have revealed that patients with IDH1 mutation have a longer survival rate compared to wildtype individuals. In glioma cells, increased oxidative stress has been identified. However, till now, the relation between oxidative stress levels and IDH1 mutation status in those patients was not examined. Therefore, the aim of this study was to investigate the urinary levels of oxidatively induced DNA damage products, 8-hydroxy-2'- deoxyguanosine (8-OH-dG), (5'R) and (5'S)-8,5'-cyclo-2'-deoxyadenosines (R-cdA and S-cdA) as reliable oxidative stress markers in patients with IDH1-wildtype (n = 20) and IDH1-mutant (n = 22) glioma. Absolute quantification of 8-OH-dG, R-cdA and S-cdA was achieved by liquid chromatography-tandem mass spectrometry with isotope dilution. The levels of 8-OH-dG were significantly greater in IDH1-wildtype glioma patients than those in IDH1-mutant ones (p = 0.017). No statistically significant difference was observed for R-cdA and S-cdA levels. 8-OH-dG levels were positively correlated with patients' tumor recurrence in all patients (r = 0.382, p = 0.014). The mutation status of glioma is well correlated with oxidative stress. Examination of noninvasively measured oxidative DNA damage products along with IDH1 mutation status in glioma patients, might be particularly important in terms of evaluating and monitoring the effectiveness of treatment.

Characterization of an IDH1 R132H Rabbit Monoclonal Antibody, MRQ-67, and Its Applications in the Identification of Diffuse Gliomas.

The current diagnosis of diffuse glioma involves isocitrate dehydrogenase (IDH) mutation testing. Most IDH mutant gliomas carry a G-to-A mutation at IDH1 position 395, resulting in the R132H mutant. R132H immunohistochemistry (IHC), therefore, is used to screen for the IDH1 mutation. In this study, the performance of MRQ-67, a recently generated IDH1 R132H antibody, was characterized in comparison with H09, a frequently used clone. Selective binding was demonstrated by an enzyme-linked immunosorbent assay for MRQ-67 to the R132H mutant, with an affinity higher than that for H09. By Western and dot immunoassays, MRQ-67 was found to bind specifically to the IDH1 R1322H, with a higher capacity than H09. IHC testing with MRQ-67 demonstrated a positive signal in most diffuse astrocytomas (16/22), oligodendrogliomas (9/15), and secondary glioblastomas tested (3/3), but not in primary glioblastomas (0/24). While both clones demonstrated a positive signal with similar patterns and equivalent intensities, H09 exhibited a background stain more frequently. DNA sequencing on 18 samples showed the R132H mutation in all IHC positive cases (5/5), but not in negative cases (0/13). These results demonstrate that MRQ-67 is a high-affinity antibody suitable for specific detection of the IDH1 R132H mutant by IHC and with less background as compared with H09.

Impact of IDH1 mutation on clinical course of patients with intrahepatic cholangiocarcinoma: a retrospective analysis from a German tertiary center.

PURPOSE: IDH1 mutation is a known biomarker for targeted therapy of intrahepatic cholangiocarcinoma (iCCA), while its prognostic relevance for current palliative chemotherapy is still unclear. Aim of this study was to analyze clinicopathological characteristics of patients with IDH1 mutations and to outline a potential impact on the outcome after state-of-the-art palliative chemotherapy regimens. METHODS: All patients with iCCA receiving large panel molecular profiling and follow-up treatment at Frankfurt University Hospital until 04/2022 were retrospectively analyzed. Clinicopathological characteristics were assessed for IDH1 mutated (mut) and IDH1 wild type (wt) patients, and progression-free survival (PFS) and overall survival (OS) were determined. RESULTS: In total, 75 patients with iCCA received molecular profiling. Of the patients with available DNA data, pathogenic mutations in IDH1 were found in 14.5% (n = 10). IDH1 mut status was associated with lower serum CA-19/9 (p = 0.023), lower serum lactate dehydrogenase (p = 0.006), and a higher proportion of primary resectability (p = 0.028) as well as response to chemotherapy after recurrence (p = 0.009). Median PFS was 5.9 months (95% CI 4.4-7.3 months) for IDH1 wt in comparison to 9.8 months (95% CI 7.7-12 months) for patients with IDH1 mut (p = 0.031). IDH1 wt was a significant risk factor for shortened PFS in univariate (p = 0.043), but not in multivariate analysis (p = 0.061). There was no difference in OS between both groups. CONCLUSION: Patients with IDH1 mutated iCCA seem to have a favorable tumor biology including a longer PFS for palliative chemotherapy regimens compared to IDH1 wild type.

Low expression of isocitrate dehydrogenase 1 (IDH1) R132H is associated with advanced pathological features in laryngeal squamous cell carcinoma.

INTRODUCTION: Recent developments in genomic sequencing have led to the identification of somatic mutations in isocitrate dehydrogenase 1 (IDH1) in various malignancies. IDH1 R132H is the most common mutation of IDH1, which affects codon 132 and results in the conversion of amino acid residue arginine (R) to histidine (H). This study is designed to evaluate the association between the expression of IDH1 R132H and clinicopathological characteristics in laryngeal squamous cell carcinoma (LSCC). METHODS: The expression pattern and clinical significance of IDH1 R132H were investigated in tissue microarrays (TMAs) of 50 LSCC tumors as well as adjacent normal tissues using immunohistochemistry. Then the exons of the 12 tumor samples with negative/weak positive staining were sequenced by applying polymerase chain reaction (PCR). RESULTS: The results demonstrated that the cytoplasmic expression of IDH1 R132H was downregulated in tumor cells compared to adjacent normal tissues. A statistically significant association was found between a low level of cytoplasmic expression of IDH1 R132H protein and an increase in histological grade (p < 0.001), perineural invasion (p = 0.019), and lymph node involvement (p < 0.001). The exon4 sequencing results showed that only one sample was positive for IDH1 R132H mutation. IDH1 R132H expression was observed in 39 (78.0%) LSCC samples. CONCLUSION: These findings indicate that low cytoplasmic expression of IDH1 R132H may have clinical significance in LSCC patients and is associated with more aggressive tumor behavior and progression of the disease, which can help improve potential treatment in patients with LSCC. Further investigations are needed to understand the biological function of IDH1 R132H and larger sample size to confirm our findings.

Development of a novel glycolysis-related genes signature for isocitrate dehydrogenase 1-associated glioblastoma multiforme.

Background: The significant difference in prognosis between IDH1 wild-type and IDH1 mutant glioblastoma multiforme (GBM) may be attributed to their metabolic discrepancies. Hence, we try to construct a prognostic signature based on glycolysis-related genes (GRGs) for IDH1-associated GBM and further investigate its relationships with immunity. Methods: Differentially expressed GRGs between IDH1 wild-type and IDH1 mutant GBM were screened based on the TCGA database and the Molecular Signature Database (MSigDB). Consensus Cluster Plus analysis and KEGG pathway analyses were used to establish a new GRGs set. WGCNA, univariate Cox, and LASSO regression analyses were then performed to construct the prognostic signature. Then, we evaluated association of the prognostic signature with patients' survival, clinical characteristics, tumor immunogenicity, immune infiltration, and validated one hub gene. Results: 956 differentially expressed genes (DEGs) between IDH1 wild-type and mutant GBM were screened out and six key prognostically related GRGs were rigorously selected to construct a prognostic signature. Further evaluation and validation showed that the signature independently predicted GBM patients' prognosis with moderate accuracy. In addition, the prognostic signature was also significantly correlated with clinical traits (sex and MGMT promoter status), tumor immunogenicity (mRNAsi, EREG-mRNAsi and HRD-TAI), and immune infiltration (stemness index, immune cells infiltration, immune score, and gene mutation). Among six key prognostically related GRGs, CLEC5A was selected and validated to potentially play oncogenic roles in GBM. Conclusion: Construction of GRGs prognostic signature and identification of close correlation between the signature and immune landscape would suggest its potential applicability in immunotherapy of GBM in the future.

Shuffle-ResNet: Deep learning for predicting LGG IDH1 mutation from multicenter anatomical MRI sequences.

Background and Purpose.The world health organization recommended to incorporate gene information such as isocitrate dehydrogenase 1 (IDH1) mutation status to improve prognosis, diagnosis, and treatment of the central nervous system tumors. We proposed our Shuffle Residual Network (Shuffle-ResNet) to predict IDH1 gene mutation status of the low grade glioma (LGG) tumors from multicenter anatomical magnetic resonance imaging (MRI) sequences including T2-w, T2-FLAIR, T1-w, and T1-Gd.Methods and Materials.We used 105 patient's dataset available in The Cancer Genome Atlas LGG project where we split them into training and testing datasets. We implemented a random image patch extractor to leverage tumor heterogeneity where about half a million image patches were extracted. RGB dataset were created from image concatenation. We used random channel-shuffle layer in the ResNet architecture to improve the generalization, and, also, a 3-fold cross validation to generalize the network's performance. The early stopping algorithm and learning rate scheduler were employed to automatically halt the training.Results.The early stopping algorithm terminated the training after 131, 106, and 96 epochs in fold 1, 2, and 3. The accuracy and area under the curve (AUC) of the validation dataset were 81.29% (95% CI (79.87, 82.72)) and 0.96 (95% CI (0.92, 0.98)) when we concatenated T2-FLAIR, T1-Gd, and T2-w to produce an RGB dataset. The accuracy and AUC values of the test dataset were 85.7% and 0.943.Conclusions.Our Shuffle-ResNet could predict IDH1 gene mutation status using multicenter MRI. However, its clinical application requires more investigation.

Integrated analysis of the genomic and transcriptional profile of gliomas with isocitrate dehydrogenase-1 and tumor protein 53 mutations.

Background: The gene mutation of isocitrate dehydrogenase-1 (IDH1) is commonly found in LGG and some GBM patients and usually carries tumor protein 53 (TP53) mutations. However, the underlying mechanisms on both mutations of glioma patients in IDH1 and TP53 are still unclear. Aim: To find the potential target markers in GBM and LGG patients with IDH1 and TP53 mutation.Method: A total of 1122 glioma patients from The Cancer Genome Atlas were enrolled and divided as wild-type (without IDH1 and TP53 mutations) or both mutant (both IDH1 and TP53 mutations). The data of clinicopathological characteristics, mRNA, mutations, and copy number alteration were analyzed. Results: IDH1 and TP53 mutations, not gene expression, affect the survival probability of GBM and LGG patients, which might be related to neuron function, immune function, tumor invasion, and metastasis. The effects of the selected gene (EMILIN3, SAA1, VSTM2A, HAMP, IFT80, and CHIC2) on glioma patients could be regulated by IDH1 and TP53 mutations and had a higher survival possibility in these patients. Conclusions: The selected genes in GBM and LGG patients with IDH1 and TP53 mutations could be a potential prognosis marker in the future.