Absence of R140Q mutation of isocitrate dehydrogenase 2 in gliomas and breast cancers

Somatic mutations of isocitrate dehydrogenase (IDH)-1 and IDH2 proteins have been described in gliomas. The mutations target the R132 amino acid residue and the R172 residue in IDH1 and IDH2, respectively. The same mutations were observed in acute myeloid leukemias with normal karyotype, but a new mutation in IDH2 (R140Q substitution) was detected in malignant myeloid diseases and appears to be the most frequent IDH mutation in these pathologies. To the best of our knowledge, no study thus far has reported the presence of this R140Q mutation in IDH2 in tumors of the nervous system and breast cancers. We evaluated IDH1 and IDH2 exon 4 in 48 low-grade gliomas, 58 primary glioblastomas and 94 breast cancers to evaluate the frequency of mutation and investigated the R140Q substitution in IDH2. The results were compared to our recently obtained results in hematopoietic diseases. The frequency of IDH1 and IDH2 mutations in our panel of gliomas was similar to previously reported mutations. No IDH2 R140 mutation was observed. Compared to hematopoietic diseases, the IDH2 R172 mutation was also more rare and IDH1 mutations more prominent in tumors of the nervous system. No IDH1 or IDH2 mutation was detected in the 94 breast cancer samples. Thus, the IDH2 R140 mutation appears to be restricted to hematopoietic diseases.     

Identification of additional IDH mutations associated with oncometabolite R(-)-2-hydroxyglutarate production

Mutations in cytosolic isocitrate dehydrogenase 1 (IDH1) or its mitochondrial homolog IDH2 can lead to R(-)-2-hydroxyglutarate (2HG) production. To date, mutations in three active site arginine residues, IDH1 R132, IDH2 R172 and IDH2 R140, have been shown to result in the neomorphic production of 2HG. Here we report on three additional 2HG-producing IDH1 mutations: IDH1 R100, which is affected in adult glioma, IDH1 G97, which is mutated in colon cancer cell lines and pediatric glioblastoma, and IDH1 Y139. All these new mutants stereospecifically produced 2HG's (R) enantiomer. In contrast, we find that the IDH1 SNPs V71I and V178I, as well as a number of other single-sample reports of IDH non-synonymous mutation, did not elevate cellular 2HG levels in cells and retained the wild-type ability for isocitrate-dependent NADPH production. Finally, we report the existence of additional rare, but recurring mutations found in lymphoma and thyroid cancer, which while failing to elevate 2HG nonetheless displayed loss of function, indicating a possible tumorigenic mechanism for a non-2HG-producing subset of IDH mutations in some malignancies. These data broaden our understanding of how IDH mutations may contribute to cancer through either neomorphic R(-)-2HG production or reduced wild-type enzymatic activity, and highlight the potential value of metabolite screening in identifying IDH-mutated tumors associated with elevated oncometabolite levels.     

Comparison of immunohistochemistry, DNA sequencing and allele-specific PCR for the detection of IDH1 mutations in gliomas

Previous studies have identified mutations of the isocitrate dehydrogenase 1 (IDH1) gene in more than 70% of World Health Organization (WHO) grade II and III gliomas. The most frequent mutation leads to a specific amino acid change from arginine to histidine at codon 132 (c.395G>A, p.R132H). IDH1 mutated tumors have a better prognosis than IDH1 non-mutated tumors. The aim of our study was to evaluate and compare the methods of mIDH1 R132H immunohistochemistry, allele-specific PCR and DNA sequencing for determination of IDH1 status. We performed a retrospective study of 91 patients with WHO grade II (n=43) and III (n=48) oligodendrogliomas. A fragment of exon 4 spanning the sequence encoding the catalytic domain of IDH1, including codon 132, was amplified and sequenced using standard conditions. Allele-specific amplification was performed using two forward primers with variations in their 3' nucleotides such that each was specific for the wild-type or the mutated variant, and one reverse primer. Immunohistochemistry was performed with mouse monoclonal mIDH1 R132H. DNA was extracted from FFPE sections following macrodissection. IDH1 mutations were found in 55/90 patients (61.1%) by direct sequencing. R132H mutations were found in 47/55 patients (85.4%). The results of the allele-specific PCR positively correlated with those from DNA sequencing. Other mutations (p.R132C, p.R132S and pR132G) were found by DNA sequencing in 3, 3 and 2 tumors, respectively (8/55 patients, 14.6%). mIDH1 R132H immunostaining was found in the 47 patients presenting the R132H mutation (sensitivity 47/47, 100% for this mutation). None of the tumors presenting a wild-type IDH1 gene were stained (specificity 35/35, 100%). Our results demonstrate that immunohistochemistry using the mIDH1 R132H antibody and allele-specific amplification are highly sensitive techniques to detect the most frequent mutation of the IDH1 gene.     

Molecular Evaluation of DNMT3A and IDH1/2 Gene Mutation:Frequency,Distribution Pattern and Associations with Additional Molecular Markers in Normal Karyotype Indian Acute Myeloid Leukemia Patients

Mutations in the DNMT3A and IDH genes represent the most common genetic alteration after FLT3/NPM1in acute myeloid leukemia (AML). We here analyzed the frequency and distribution pattern of DNMT3A andIDH mutations and their associations with other molecular markers in normal karyotype AML patients. Fortyfivepatients were screened for mutations in DNMT3A (R882), IDH1 (R132) and IDH2 (R140 and R172) genesby direct sequencing. Of the 45 patients screened, DNMT3A and IDH mutations were observed in 6 (13.3%)and 7 (15.4%), respectively. Patients with isolated DNMT3A mutations were seen in 4 cases (9%), isolatedIDH mutations in 5 (11.1%), while interestingly, two cases showed both DNMT3A and IDH mutations (4.3%).Nucleotide sequencing of DNMT3A revealed missense mutations (R882H and R882C), while that of IDH revealedR172K, R140Q, R132H and R132S. Both DNMT3A and IDH mutations were observed only in adults, with ahigher frequency in males. DNMT3A and IDH mutations were significantly associated with NPM1, while trendstowards higher coexistence with FLT3 mutations were observed. This is the first study to evaluate DNMT3A/IDH mutations in Indian patients. Significant associations among the various molecular markers was observed,that highlights cooperation between them and possible roles in improved risk stratification.     

Analysis of IDH1 and IDH2 mutations in Japanese glioma patients

A recent study reported on mutations in the active site of the isocitrate dehydrogenase 1 ( IDH1 ) gene in several types of gliomas. All mutations detected resulted in an amino acid exchange at position 132. We analyzed the genomic region spanning wild-type R132 of IDH1 by direct sequencing in 125 glial tumors. A total of 39 IDH1 mutations were observed. Mutations of the IDH2 gene, homologous to IDH1 , were often detected in gliomas without IDH1 mutations. In the present study, R172 mutation of the IDH2 gene was detected in one anaplastic astrocytoma. IDH1 or IDH2 mutations were frequently in oligodendrogliomas (67%), anaplastic astrocytomas (62%), anaplastic oligoastrocytomas (75%), anaplastic oligodendrogliomas (50%), secondary glioblastomas (67%), gangliogliomas (38%), and anaplastic gangliogliomas (60%). Primary glioblastomas were characterized by a low frequency of mutations (5%) at amino acid position 132 of IDH1 . Mutations of the IDH1 or IDH2 genes were significantly associated with improved outcome in patients with anaplastic astrocytomas. Our data suggest that IDH1 or IDH2 mutation plays a role in early tumor progression of several types of glioma and might arise from a common glial precursor. The infrequency of IDH1 mutation in primary glioblastomas revealed that these subtypes are genetically distinct entities from other glial tumors. ( Cancer Sci  2009; 100: 1996–1998)     

Analysis of NADP+-dependent isocitrate dehydrogenase-1/2 gene mutations in pediatric brain tumors: report of a secondary anaplastic astrocytoma carrying the IDH1 mutation

Somatic mutations of the isocitrate dehydrogenase-1 gene (IDH1), most commonly resulting in replacement of arginine at position 132 by histidine (p.R132H), have been reported for WHO grade II and III diffuse gliomas and secondary glioblastomas. We investigated IDH1/2 mutations in a retrospective series of 165 pediatric brain tumors, including atypical teratoid/rhabdoid tumors (AT/RT) and choroid plexus tumors, which had not previously been investigated. Mutation analysis was performed by use of pyrosequencing and, additionally, data were validated for a cohort of 70 gliomas from among the series by use of the arrayed primer extension technique. We identified one tumor which harbored mutation of IDH1 at codon 132 and no alteration was identified in the matched-germline DNA. No IDH2 mutations were detected. Most noteworthy, the IDH1 mutant tumor was an anaplastic astrocytoma involving the cortex in the left frontal lobe which appeared seven years after radiation treatment for an extensive sellar/suprasellar craniopharyngioma. This anaplastic astrocytoma was regarded as secondary to radiation treatment because it seemed to originate within the irradiation field that received a dose varying from a maximum of 30.6 Gy of 4 MV X-rays down to very few Gy of lower-energy scattered radiation. In this work our observations agree with those in previous reports showing the rarity of IDH1/2 mutations in childhood tumors. The interesting identification of an IDH1 mutation in a radiation-induced secondary malignant glioma raises the likelihood that these types of tumor may develop IDH1/2 mutations. Thus, caution is needed when dealing with these tumors, and further genetic analysis is warranted.     

Frequent IDH1/2 mutations in intracranial chondrosarcoma: a possible diagnostic clue for its differentiation from chordoma

Mutations in the genes encoding isocitrate dehydrogenase (IDH) 1/2 have been detected in a significant proportion of diffuse gliomas and in a small fraction of acute myeloid leukemia (AML) cases. Recently, in an examination of various types of mesenchymal tumor, IDH1/2 mutations were only found in cartilaginous tumors including central conventional and periosteal enchondromas/chondrosarcomas. The frequency of IDH1/2 mutations was 56%, and the IDH1 R132C mutation, which is not common in diffuse gliomas or AML, accounted for 40% of these mutations. In this study, we investigated the IDH1/2 mutation status of intracranial chondrosarcomas and chordomas, which are morphologically similar and affect similar regions of the cranial cavity. Of the 13 chondrosarcomas analyzed, six (46.1%) displayed IDH1/2 mutations (the predominant type was IDH1 R132C). Also, an IDH2 mutation (R172S) was observed in one case. Conversely, none of the ten chordomas analyzed displayed any IDH1 or IDH2 mutations. Our data suggest that the IDH1/2 mutation status could be valuable for distinguishing intracranial chondrosarcomas from chordomas.     

Molecular Investigation of Isocitrate Dehydrogenase Gene (IDH) Mutations in Gliomas: First Report of IDH2 Mutations in Indian Patients

Recent genome wide sequencing has identified mutations in IDH1/IDH2 predominantly in grade II-III gliomasand secondary glioblastomas which are associated with favorable clinical outcome. These mutations have becomemolecular markers of significant diagnostic and prognostic relevance in the assessment of human gliomas. In thecurrent study we evaluated IDH1 (R132) and IDH2 (R172) in 32 gliomas of various grades and tumor subtypes.Sequencing analysis revealed R132H mutations in 18.7% tumors, while none of the cases showed IDH2 (R172)mutations. The frequency of IDH1 mutations was higher in females (21.4%) than males (11.1%), and it wassignificantly higher in younger patients. Histological analyses demonstrated presence of necrosis and microvascular proliferation in 69% and 75% respectively. Interestingly, IDH1 mutations were predominantly presentin non-necrotic tumors as well as in cases showing microvascular proliferation. Of the six IDH1 positive cases,three were glioblastomas (IV), and one each were anaplastic oligoastrocytoma (III), anaplastic oligodendrogliomaIII (n=1) and diffuse astrocytoma. In conclusion, IDH1 mutations are quite frequent in Indian glioma patientswhile IDH2 mutations are not observed. Since IDH mutations are associated with good prognosis, their use inroutine clinical practice will enable better risk stratification and management of glioma patients.     

Comparative study of IDH1 mutations in gliomas by immunohistochemistry and DNA sequencing

Background

Mutations involving isocitrate dehydrogenase 1 (IDH 1) occur in a high proportion of diffuse gliomas, with implications on diagnosis and prognosis. About 90% involve exon 4 at codon 132, replacing amino acid arginine with histidine (R132H). Rarer ones include R132C, R132S, R132G, R132L, R132V, and R132P. Most authors have used DNA-based methods to assess IDH1 status. Preliminary studies comparing imunohistochemistry (IHC) with IDH1-R132H mutation-specific antibodies have shown concordance with DNA sequencing and no cross-reactivity with wild-type IDH1 or other mutant proteins. The present study compares results of IHC with DNA sequencing in diffuse gliomas.

Materials and methods

Fifty diffuse gliomas with frozen tissue samples for DNA sequencing and adequate tissue in paraffin blocks for IHC using IDH1-R132H specific antibody were assessed for IDH1 mutations.

Results

Concordance of findings between IHC and DNA sequencing was noted in 88% (44/50) cases. All 6 cases with discrepancy were immunopositive with DIA-H09 antibody. While in 3 of these 6 cases, DNA sequencing failed to reveal any mutations, R132L (arginine replaced by leucine) mutation was found in the rest 3 cases. Interestingly, of the immunopositive cases, 46.6% (14/30) showed immunostaining in only a fraction of tumor cells.

Conclusions

IHC is an easy and quick method of detecting IDH1-R132H mutations, but there may be some discrepancies between IHC and DNA sequencing. Although there were no false-negative cases, cross-reactivity with IDH1-R132L was seen in 3, a finding not reported thus far. Because of more universal availability of IHC over genetic testing, cross-reactivity and staining heterogeneity may have bearing over its use in detecting IDH1-R132H mutation in gliomas.     

Pharmacological characterization of TQ05310, a potent inhibitor of isocitrate dehydrogenase 2 R140Q and R172K mutants

Isocitrate dehydrogenase 2 (IDH2), an important mitochondrial metabolic enzyme involved in the tricarboxylic acid cycle, is mutated in a variety of cancers. AG‐221, an inhibitor primarily targeting the IDH2‐R140Q mutant, has shown remarkable clinical benefits in the treatment of relapsed or refractory acute myeloid leukemia patients. However, AG‐221 has weak inhibitory activity toward IDH2‐R172K, a mutant form of IDH2 with more severe clinical manifestations. Herein, we report TQ05310 as the first mutant IDH2 inhibitor that potently targets both IDH2‐R140Q and IDH2‐R172K mutants. TQ05310 inhibited mutant IDH2 enzymatic activity, suppressed (R)‐2‐hydroxyglutarate (2‐HG) production and induced differentiation in cells expressing IDH2‐R140Q and IDH2‐R172K, but not in cells expressing wild‐type IDH1/2 or mutant IDH1. TQ05310 bound to both IDH2‐R140Q and IDH2‐R172K, with Q316 being the critical residue mediating the binding of TQ05310 with IDH2‐R140Q, but not with IDH2‐R172K. TQ05310 also had favorable pharmacokinetic characteristics and profoundly inhibited 2‐HG production in a tumor xenografts model. The results of the current study establish a solid foundation for further clinical investigation of TQ05310, and provide new insight into the development of novel mutant IDH2 inhibitors.     

Immunohistochemical detection of IDH1 mutation, p53, and internexin as prognostic factors of glial tumors

Isocitrate dehydrogenase 1 (IDH1) mutations, which are early and frequent genetic alterations in astrocytomas, oligodendrogliomas, oligoastrocytomas, and secondary glioblastomas, are specific to arginine 132 (R132). Recently, we established monoclonal antibodies (mAbs) against IDH1 mutations: anti-IDH1-R132H and anti-IDH1-R132S. However, the importance of immunohistochemistry using the combination of those mAbs has not been elucidated. For this study, 164 cases of glioma were evaluated immunohistochemically for IDH1 mutations (R132H and R132S) using anti-IDH1 mAbs (HMab-1 and SMab-1). IDH1 mutation was detected, respectively, in 9.7%, 63.6%, 51.7%, and 77.8% of primary grade IV, secondary grade IV, grade III, and grade II gliomas. For each grade of glioma, prognostic factors for progression-free survival and overall survival were evaluated using clinical and pathological parameters in addition to IDH1 immunohistochemistry. IDH1 mutation, p53 overexpression, and internexin expression, as evaluated using immunohistochemistry with clinical parameters such as degree of surgical removal and preoperative Karnofsky Performance Status (KPS), might be of greater prognostic significance than histological grading alone in grade III as well as IDH1 mutation in grade IV gliomas.     

Prevalence of IDH1/2 Mutations in Different Subtypes of Glioma in the North-East Population of Morocco

Background: Genetic alterations in gliomas have increasing importance for classification purposes. Thus, we are especially interested in studying IDH mutations which may feature potential roles in diagnosis, prognosis and response to treatment. Our aim was to investigate IDH mutations in diffuse glioma patients diagnosed in university hospital centre of Fez in Morocco. Materials and Methods: IDH1 codon 132 and IDH2 codon 172 were direct-sequenced in 117 diffuse glioma samples diagnosed and treated in University Hospital Hassan II between 2010 and 2014. Results: The R132H IDH1 mutation was identified in 43/117 tumor samples and R172K IDH2 mutation was detected in only one anaplastic oligodendroglioma. IDH mutations were observed in 63.2% of astrocytomas, 73.3% of diffuse oligodendrogliomas and 12.90% of glioblastomas. Conclusions: Our results confirmed other studies published earlier for other populations with some small discrepancies.     

Mutational analysis of IDH1 codon 132 in glioblastomas and other common cancers

Missense somatic mutations in IDH1 gene affecting codon 132 have recently been reported in glioblastoma multiforme (GBM) and other gliomas. The recurrent nature of the IDH1 mutations in the same amino acid strongly suggests that the mutations may play important roles in the pathogenesis of glial tumors. The aim of this study was to see whether the IDH1 codon 132 mutations occur in other human cancers besides glial tumors. We also attempted to confirm the occurrence of the IDH1 mutations in GBM of Korean patients. We have analyzed 1,186 cancer tissues from various origins, including carcinomas from breast, colon, lung, stomach, esophagus, liver, prostate, urinary bladder, ovary, uterine cervix, skin and kidney, and malignant mesotheliomas, primary GBM, malignant meningiomas, multiple myelomas and acute leukemias by single‐strand conformation polymorphism analysis. We found four IDH1 codon 132 mutations in the GBM (4/25; 16.0%), two in the prostate carcinomas (2/75; 2.7%) and one in the B‐acute lymphoblastic leukemias (B‐ALL) (1/60; 1.7%), but none in other cancers. The IDH1 mutations consisted of five p.R132H and two p.R132C mutations. The data indicate that IDH1 codon 132 mutations occur not only in GBM, but also in prostate cancers and B‐ALL. This study suggests that despite the infrequent incidence of the IDH1 mutations in prostate cancers and B‐ALL, mutated IDH1 could be therapeutically targeted in these cancers and in glial tumors with the IDH1 mutations. © 2009 UICC     

Association of The IDH1 C.395G>A (R132H) Mutation with Histological Type in Malay Brain Tumors

Background: Brain tumors, constituting one of the most deadly forms of cancer worldwide, result from the accumulation of multiple genetic and epigenetic alterations in genes and signaling pathways. Isocitrate dehydrogenase enzyme isoform 1 (IDH1) mutations are frequently identified in primary brain tumors and acute myeloid leukemia. Studies on IDH1 gene mutations have been extensively performed in various populations worldwide but not in Malaysia. This work was conducted to study the prevalence of IDH1 c.395G>A (R132H) hotspot mutations in a group of Malaysian patients with brain tumors in order to gain local data for the IDH1 mutation profile in our population. Methods: Mutation analysis of c.395G>A (R132H) of IDH1 was performed in 40 brain tumor specimens by the polymerase chain reaction-restriction fragment length polymorphism method (PCR-RFLP) and then verified by direct sequencing. Associations between the IDH1 c.395G>A (R132H) mutation and clinicopathologic characteristics were also analyzed. Results: The IDH1 c.395G>A (R132H) mutation was detected in 14/40 patients (35%). A significant association was found with histological tumor types, but not with age, gender and race. Conclusions: IDH1 is frequently mutated and associated with histological subtypes in Malay brain tumors.     

Mutation Analysis of IDH1/2 Genes in Unselected De novo Acute Myeloid Leukaemia Patients in India - Identification of A Novel IDH2 Mutation

IDH1/2 mutations which result in alternation in DNA methylation pattern are one of the most commonmethylation associated mutations in Acute myeloid leukaemia. IDH1/2 mutations frequently associated withhigher platelet level, normal cytogentics and NPM1 mutations. Here we analyzed IDH1/2 mutations in 200 newlydiagnosed unselected Indian adult AML patients and investigated their correlation with clinical, cytogeneticparameters along with cooperating NPM1 mutation. We detected 5.5% and 4% mutations in IDH1/2 genes,respectively. Except IDH2 c.515_516GG>AA mutation, all the other identified mutations were reported mutations.Similar to reported c.515G>A mutation, the novel c.515_516GG>AA mutation replaces 172nd arginine to lysinein the active site of the enzyme. Even though there was a preponderance of IDH1/2 mutations in NK-AML,cytogenetically abnormal patients also harboured IDH1/2 mutations. IDH1 mutations showed significant higherplatelet count and NPM1 mutations. IDH2 mutated patients displayed infrequent NPM1 mutations and lowerWBC count. All the NPM1 mutations in the IDH1/2 mutated cases showed type A mutation. The present datasuggest that IDH1/2 mutations are associated with normal cytogenetics and type A NPM1 mutations in adultIndian AML patients.     

Detection of 2-hydroxyglutaric acid in vivo by proton magnetic resonance spectroscopy in U87 glioma cells overexpressing isocitrate dehydrogenase–1 mutation

The arginine 132 (R132) mutation of isocitrate dehydrogenase –1 (IDH1^R132) results in production of 2-hydroxyglutarate (2-HG) and is associated with a better prognosis compared with wild-type (WT) in glioma patients. The majority of lower-grade gliomas express IDH1^R132, whereas this mutation is rare in grade IV gliomas. The aim of this study was to noninvasively investigate metabolic and physiologic changes associated with the IDH1 mutation in a mouse glioma model. Using a 7T magnet, we compared MRI and proton magnetic resonance spectroscopy (MRS) in U87 glioma cells overexpressing either the mutated IDH1^R132 or IDH1 wild-type (IDH1^WT) gene in a mouse flank xenograft model. Flank tumors overexpressing IDH1^R132 showed a resonance at 2.25 ppm corresponding to the 2-HG peak described for human IDH1^R132 gliomas. WT tumors lacked this peak in all cases. IDH1 mutant tumors demonstrated significantly reduced glutamate by in vivo MRS. There were no significant differences in T₂, apparent diffusion coefficient (ADC), or perfusion values between the mutant and IDH1^WT tumors. The IDH1^R132 mutation results in 2-HG resonance at 2.25 ppm and a reduction of glutamate levels as determined by MRS. Our results establish a model system where 2-HG can be monitored noninvasively, which should be helpful in validating 2-HG levels as a prognostic and/or predictive biomarker in glioma.     

Isocitrate Dehydrogenase 1 (IDH1) Mutation in Breast Adenocarcinoma Is Associated With Elevated Levels of Serum and Urine 2‐Hydroxyglutarate

Mutations in the IDH1 and IDH2 (isocitrate dehydrogenase) genes have been discovered across a range of solid‐organ and hematologic malignancies, including acute myeloid leukemia, glioma, chondrosarcoma, and cholangiocarcinoma. An intriguing aspect of IDH‐mutant tumors is the aberrant production and accumulation of the oncometabolite 2‐hydroxyglutarate (2‐HG), which may play a pivotal oncogenic role in these malignancies. We describe the first reported case of an IDH1 p.R132L mutation in a patient with hormone receptor‐positive (HR+) breast adenocarcinoma. This patient was initially treated for locally advanced disease, but then suffered a relapse and metastasis, at which point an IDH1‐R132 mutation was discovered in an affected lymph node. The mutation was subsequently found in the primary tumor tissue and all metastatic sites, but not in an uninvolved lymph node. In addition, the patient's serum and urine displayed marked elevations in the concentration of 2‐HG, significantly higher than that measured in six other patients with metastatic HR+ breast carcinoma whose tumors were found to harbor wild‐type IDH1. In summary, IDH1 mutations may impact a rare subgroup of patients with breast adenocarcinoma. This may suggest future avenues for disease monitoring through noninvasive measurement of 2‐HG, as well as for the development and study of targeted therapies against the aberrant IDH1 enzyme.     

IDH mutations in primary myelofibrosis predict leukemic transformation and shortened survival: clinical evidence for leukemogenic collaboration with JAK2V617F

Isocitrate dehydrogenase (IDH) mutations are frequent in blast-phase myeloproliferative neoplasms and might therefore contribute to leukemic transformation. We examined this possibility in 301 consecutive patients with chronic-phase primary myelofibrosis (PMF). The mutant IDH was detected in 12 patients (4%): 7 IDH2 (5 R140Q, 1 R140W and 1 R172G) and 5 IDH1 (3 R132S and 2 R132C). In all, 6 (50%) of the 12 IDH-mutated patients also expressed JAK2V617F. Overall, 18 (6%) patients displayed only MPL and 164 (54.3%) only JAK2 mutations. Multivariable analysis that accounted for conventional risk factors disclosed inferior overall survival (OS; P=0.03) and leukemia-free survival (LFS; P=0.003) in IDH-mutated patients: OS hazard ratio (HR) was 0.39 (95% confidence interval (95% CI) 0.2-0.75), 0.50 (95% CI 0.27-0.95) and 0.53 (95% CI 0.23-1.2) for patients with no, JAK2 or MPL mutations, respectively. Further analysis disclosed a more pronounced effect for the mutant IDH on OS and LFS in the presence (P=0.0002 and P<0.0001, respectively) as opposed to the absence (P=0.34 and P=0.64) of concomitant JAK2V617F. Analysis of paired samples obtained during chronic- and blast-phase disease revealed the presence of both IDH and JAK2 mutations at both time points. Our observations suggest that IDH mutations in PMF are independent predictors of leukemic transformation and raise the possibility of leukemogenic collaboration with JAK2V617F.     

皮肤黑色素瘤中IDH1基因突变分析

目的:应用癌症基因组图谱(TCGA)数据库分析皮肤黑色素瘤中IDH1基因突变情况.方法:从癌症基因组图谱(TCGA)数据库收集皮肤黑色素瘤数据集,统计患者临床信息、应用cBioPortal网站对数据集进行突变、离散基因、基因共表达、生存期等分析.结果:287个病例中,IDH1基因突变15例,其中,R132C突变10例,R132L突变2例,P33S/E361K突变各1例,X174缺失1例.结论:IDH1基因突变在皮肤黑色素瘤中有一定发生率,占比5.2%,且以R132突变为主,此位点有可能成为靶向药物作用的靶点.     

Interlaboratory comparison of IDH mutation detection

Isocitrate dehydrogenase (IDH) mutational testing is becoming increasingly important. For this, robust and reliable assays are needed. We tested the variation of results between six laboratories of testing for IDH mutations. Each laboratory received five unstained slides from 31 formalin-fixed paraffin-embedded (FFPE) glioma samples, and followed its own standard IDH diagnostic routine. All laboratories used immunohistochemistry (IHC) with an antibody against the most frequent IDH1 mutation (R132H) as a first step. Three laboratories then sequenced only IHC negative cases while the others sequenced all cases. Based on the overall analysis, 13 samples from 11 tumors had an R132H mutation and one tumor showed an R132G mutation. Results of IHC for IDH1 R132H mutations in all six laboratories were completely in agreement, and identified all R132H mutations. Upon sequencing the results of two laboratories deviated from those of the others. After a review of the entire diagnostic process, on repeat (blinded) testing one laboratory was completely in agreement with the overall result. A change in technique did only partially improve the results in the other laboratory. IHC for the IDH1 R132H mutation is very reliable and consistent across laboratories. IDH sequencing procedures yielded inconsistent results in 2 out of 6 laboratories. Quality assurance is pivotal before IDH testing is made part of clinical management of patients.