Marine Giry

Detection, Characterization, and Inhibition of FGFR–TACC Fusions in IDH Wild-type Glioma

Purpose: Oncogenic fusions consisting of fibroblast growth factor receptor (FGFR) and TACC are present in a subgroup of glioblastoma (GBM) and other human cancers and have been proposed as new therapeutic targets. We analyzed frequency and molecular features of FGFR–TACC fusions and explored the therapeutic efficacy of inhibiting FGFR kinase in GBM and grade II and III glioma. Experimental Design: Overall, 795 gliomas (584 GBM, 85 grades II and III with wild-type and 126 with IDH1/2 mutation) were screened for FGFR–TACC breakpoints and associated molecular profile. We also analyzed expression of the FGFR3 and TACC3 components of the fusions. The effects of the specific FGFR inhibitor JNJ-42756493 for FGFR3–TACC3–positive glioma were determined in preclinical experiments. Two patients with advanced FGFR3–TACC3–positive GBM received JNJ-42756493 and were assessed for therapeutic response. Results: Three of 85 IDH1/2 wild-type (3.5%) but none of 126 IDH1/2-mutant grade II and III gliomas harbored FGFR3–TACC3 fusions. FGFR–TACC rearrangements were present in 17 of 584 GBM (2.9%). FGFR3–TACC3 fusions were associated with strong and homogeneous FGFR3 immunostaining. They are mutually exclusive with IDH1/2 mutations and EGFR amplification, whereas they co-occur with CDK4 amplification. JNJ-42756493 inhibited growth of glioma cells harboring FGFR3–TACC3 in vitro and in vivo. The two patients with FGFR3–TACC3 rearrangements who received JNJ-42756493 manifested clinical improvement with stable disease and minor response, respectively. Conclusions: RT-PCR sequencing is a sensitive and specific method to identify FGFR–TACC–positive patients. FGFR3–TACC3 fusions are associated with uniform intratumor expression of the fusion protein. The clinical response observed in the FGFR3–TACC3–positive patients treated with an FGFR inhibitor supports clinical studies of FGFR inhibition in FGFR–TACC–positive patients. Clin Cancer Res; 21(14); 3307–17. ©2015 AACR. See related commentary by Ahluwalia and Rich, p. 3105

Prevalence, clinico-pathological value, and co-occurrence of PDGFRA abnormalities in diffuse gliomas

PDGFRA is a critical gene in glioma biology. Similar to EGFR, PDGFRA has been shown to be overexpressed, amplified, mutated, or truncated in gliomas, particularly glioblastomas. In addition, PDGFRA has been recently shown to be rearranged in glioblastoma. However, the frequency, cooccurrence, and clinical value of PDGFRA abnormalities in diffuse gliomas remain unclear. We investigated PDGFRA abnormalities and their clinical impact on 619 primary diffuse gliomas, including 167 grade II, 168 grade III, and 284 grade IV gliomas, with use of BAC-aCGH and validated our findings by quantitative polymerase chain reaction (PCR). We studied PDGFRA expression using reverse-transcription quantitative PCR in 84 gliomas and 12 non-tumor samples. In 138 samples, we also screened PDGFRA point mutations in exons 5, 7, 8, 9, 10, 11, and 23; presence of KDR-PDGFRA fusion gene; and PDGFRA truncation. PDGFRA was amplified and gained in 5.2% and 1.9% of samples, respectively. In addition PDGFRA was point-mutated, rearranged, and truncated in 2.9%, 0%, and 0.7% of cases, respectively. PDGFRA point mutations were observed exclusively in grade IV gliomas and in 12.5% of PDGFRA-amplified tumors. High-level PDGFRA amplification was associated with PDGFRA overexpression, high malignancy grade, and older patient age. Of interest, high-level PDGFRA amplification has an independent negative prognostic value for progression-free survival and overall survival among patients with grade III tumors. PDGFRA is altered through various genetic mechanisms in a subset of high-grade gliomas in patients who might be ideal candidates for PDGFRA inhibitor treatment, and PDGFRA gene amplification could be used as a prognostic biomarker in anaplastic gliomas.

Chordoid gliomas of the third ventricle share TTF-1 expression with organum vasculosum of the lamina terminalis.

Chordoid glioma of the third ventricle (CG3V) is a rare tumor developing in a stereotyped localization. It has been related to the circumventricular organ of the lamina terminalis, in the anterior part of the third ventricle, but its oncogenesis is poorly understood. TTF-1 transcription factor is involved in the development and adult physiology of the ventral forebrain. We studied the histopathologic and immunohistochemical features of a multicentric series of 17 cases of CG3V. We described additional histologic patterns (solid, fibrosing, and fusiform) to the typical chordoid pattern. TTF-1 was constantly expressed in CG3V, as in developing and adult lamina terminalis. The anti-TTF-1 SPT24 clone was more sensitive than the 8G7G3/1 clone. No mutation of IDH1 R132, IDH2 R172, or BRAF V600 codons was found. We showed TTF-1 as a useful marker for the diagnosis of CG3V and the understanding of its oncogenesis.

K27M mutation in H3F3A in ganglioglioma grade I with spontaneous malignant transformation extends the histopathological spectrum of the histone H3 oncogenic pathway

Here we describe the presence of the mutation p.K27M of H3F3A (H3.3K27M) in two tumours of young patients with classical histopathology of ganglioglioma (grade I WHO 2007), although H3.3K27M represents a hallmark of midline High Grade Glioma (HGG). Ganglioglioma grade I is a rare, circumscribed, glioneuronal tumour of the central nervous system (CNS) that occurs most often in young patients, most frequently in the temporal lobe and presents with seizures [1]. The neuronal component consists of ganglion cells, abnormally grouped and occasionally binucleated. The glial component consists of piloid or fibrillary astrocytic elements, or of pseudo-oligodendroglial elements. Mitoses are occasional and necrosis is absent. Eosinophilic granular bodies (EGB) and perivascular lymphocytes are associated [1]. The glial component variably expresses GFAP and OLIG2 and the neuronal component variably expresses synaptophysin, chromogranin A and MAP2. The progenitor marker CD34 often shows an extravascular stellar immunostaining [1]. Ki67 labelling index is usually less than 3% and p53 immunostaining is negative. The majority of gangliogliomas are benign grade I tumours, however 6% present as grade III anaplastic ganglioglioma or undergo malignant transformation. Markers to predict anaplastic transformation of grade I tumours are poorly defined [1]. The most frequent genetic alterations in ganglioglioma are BRAF p.V600E mutation (40–60%), gain of chromosomes 7 (23%) and 5 (18%) [1–4]. Recurrent mutations in the genes H3F3A and HIST1H3B (encoding histones H3.3 and H3.1 respectively) were described in paediatric and adult HGG and carry a dismal prognosis [5–7]. These mutations show a distinct anatomical segregation: K27M mutation of H3F3A is observed in thalamus, pons and spinal cord, K27M mutation of HIST1H3B is specifically observed in diffuse intrinsic pontine gliomas and G34R/V mutation of H3F3A is observed in the cerebral hemispheres [5–7]. At position 27, lysine is replaced by methionine (K27M) resulting in decreased K27 trimethylation (H3K27me3) [5]. H3.3K27M immunolabelling shows 100% sensitivity and specificity compared to sequencing [5]. Recently, five H3.3K27M paediatric tumours with histopathology other than HGG were reported: three tumours of unclassified histopathology and harbouring BRAF V600E mutation [8], one ganglioglioma harbouring BRAF V600E mutation which underwent a delayed malignant transformation [9], a spinal pilocytic astrocytoma which underwent a delayed malignant transformation after 10 years [10]. Here we extend the histopathological spectrum of H3.3K27M tumours and present two extratemporal grade I gangliogliomas with secondary malignant transformation. One paediatric midline ganglioglioma had combined H3.3K27M and BRAF V600E mutations and its long-term malignant relapse was BRAF wild type. The second case represented a cerebellar tumour with a rapid malignant transformation. The first case was a 12-year-old girl diagnosed with a partially resected right thalamic tumour. Histopathology was of a ganglioglioma grade I. The tumour was reticulin free and composed of astrocytic piloid elements, pseudo-oligodendroglial elements and ganglionic cells associated with EGBs, lymphocytes and CD34 extravascular stellar immunostaining (Figure 1A–C). Binucleated chromogranin A positive cells and mononucleated synaptophysin expressing cells were detected (Figure 1D). The neurofilament immunolabelling confirmed the circumscribed nature of the tumour. The Ki67 labelling index was 2% (Figure 1E). The residual tumour was stable for 7 years until the patient presented with intracranial hypertension. MRI showed an in situ relapse (Figure 1G). The surgical resection was subtotal. Histopathology demonstrated an anaplastic ganglioglioma. The lesion contained lowgrade areas (Figure 1H) and anaplastic areas with high cellularity, marked atypia, mitotic activity exceeding 5 mitoses per 10 high power field (HPF), microvascular proliferation, pseudopalisading necrosis, high p53 expression and Ki67 index reaching 20% (Figure 1I).

Tumor cells with neuronal intermediate progenitor features define a subgroup of 1p/19q co-deleted anaplastic gliomas

The integrated diagnosis of anaplastic oligodendroglioma, IDH mutant and 1p/19q co‐deleted, grade III (O3id) is a histomolecular entity that WHO 2016 classification distinguished from other diffuse gliomas by specific molecular alterations. In contrast, its cell portrait is less well known. The present study is focused on intertumor and intratumor, cell lineage‐oriented, heterogeneity in O3id. Based on pathological, transcriptomic and immunophenotypic studies, a novel subgroup of newly diagnosed O3id overexpressing neuronal intermediate progenitor (NIP) genes was identified. This NIP overexpression pattern in O3id is associated with: (i) morphological and immunohistochemical similarities with embryonic subventricular zone, (ii) proliferating tumor cell subpopulation with NIP features including expression of INSM1 and no expression of SOX9, (iii) mutations in critical genes involved in NIP biology and, (iv) increased tumor necrosis. Interestingly, NIP tumor cell subpopulation increases in O3id recurrence compared with paired newly diagnosed tumors. Our results, validated in an independent cohort, emphasize intertumor and intratumor heterogeneity in O3id and identified a tumor cell subpopulation exhibiting NIP characteristics that is potentially critical in oncogenesis of O3id. A better understanding of spatial and temporal intratumor cell heterogeneity in O3id will open new therapeutic avenues overcoming resistance to current antitumor treatments.

A recurrent point mutation in PRKCA is a hallmark of chordoid gliomas

Chordoid glioma (ChG) is a characteristic, slow growing, and well-circumscribed diencephalic tumor, whose mutational landscape is unknown. Here we report the analysis of 16 ChG by whole-exome and RNA-sequencing. We found that 15 ChG harbor the same PRKCAD463H mutation. PRKCA encodes the Protein kinase C (PKC) isozyme alpha (PKCα) and is mutated in a wide range of human cancers. However the hot spot PRKCAD463H mutation was not described in other tumors. PRKCAD463H is strongly associated with the activation of protein translation initiation (EIF2) pathway. PKCαD463H mRNA levels are more abundant than wild-type PKCα transcripts, while PKCαD463H is less stable than the PCKαWT protein. Compared to PCKαWT, the PKCαD463H protein is depleted from the cell membrane. The PKCαD463H mutant enhances proliferation of astrocytes and tanycytes, the cells of origin of ChG. In conclusion, our study identifies the hallmark mutation for chordoid gliomas and provides mechanistic insights on ChG oncogenesis.Chordoid glioma is a slow growing diencephalic tumor whose mutational landscape is poorly characterized. Here, the authors perform whole-exome and RNA-sequencing and find that 15 of 16 chordoid glioma cases studied harbor the same PRKCA mutation which results in enhanced proliferation.

Transcriptomic immune profiling of ovarian cancers in paraneoplastic cerebellar degeneration associated with anti-Yo antibodies

BackgroundParaneoplastic neurological syndromes are rare conditions where an autoimmune reaction against the nervous system appears in patients suffering from a tumour, but not linked to the spreading of the tumour. A break in the immune tolerance is thought to be the trigger.MethodsThe transcriptomic profile of 12 ovarian tumours (OT) from patients suffering from paraneoplastic cerebellar degeneration (PCD) linked to anti-Yo antibodies (anti-Yo PCD OT) was compared with 733 ovarian tumours (OT control) from different public databases using linear model analysis.ResultsA prominent significant transcriptomic over-representation of CD8+ and Treg cells was found in anti-Yo PCD OT, as compared to the OT control. However, the overall degree of immune cell infiltration was similar, according to the ESTIMATE immune score. We also found an under-representation of M2 macrophages in anti-Yo PCD OT. Furthermore, the differentially expressed genes were enriched for AIRE-related genes, a well-known transcription factor associated with a broad range of autoimmune diseases. Finally, we found that the differentially expressed genes were correlated to the transcriptomic profiling of the cerebellar structures.ConclusionsOur data pinpointed the enrichment of acquired immune response, particularly high density of CD8+ lymphocytes, and high-level expression of CDR-related antigens in anti-Yo PCD OT.

Diffuse gliomas with FGFR3-TACC3 fusion have characteristic histopathological and molecular features

Adult glioblastomas, IDH‐wildtype represent a heterogeneous group of diseases. They are resistant to conventional treatment by concomitant radiochemotherapy and carry a dismal prognosis. The discovery of oncogenic gene fusions in these tumors has led to prospective targeted treatments, but identification of these rare alterations in practice is challenging. Here, we report a series of 30 adult diffuse gliomas with an in frame FGFR3‐TACC3 oncogenic fusion (n = 27 WHO grade IV and n = 3 WHO grade II) as well as their histological and molecular features. We observed recurrent morphological features (monomorphous ovoid nuclei, nuclear palisading and thin parallel cytoplasmic processes, endocrinoid network of thin capillaries) associated with frequent microcalcifications and desmoplasia. We report a constant immunoreactivity for FGFR3, which is a valuable method for screening for the FGFR3‐TACC3 fusion with 100% sensitivity and 92% specificity. We confirmed the associated molecular features (typical genetic alterations of glioblastoma, except the absence of EGFR amplification, and an increased frequency of CDK4 and MDM2 amplifications). FGFR3 immunopositivity is a valuable tool to identify gliomas that are likely to harbor the FGFR3‐TACC3 fusion for inclusion in targeted therapeutic trials.

EGFR Amplification and IDH Mutations in Glioblastoma Patients of the Northeast of Morocco

Glioblastomas are the most frequent and aggressive primary brain tumors which are expressing various evolutions, aggressiveness, and prognosis. Thus, the 2007 World Health Organization classification based solely on the histological criteria is no longer sufficient. It should be complemented by molecular analysis for a true histomolecular classification. The new 2016 WHO classification of tumors of the central nervous system uses molecular parameters in addition to histology to reclassify these tumors and reduce the interobserver variability. The aim of this study is to determine the prevalence of IDH mutations and EGFR amplifications in the population of the northeast region of Morocco and then to compare the results with other studies. Methods. IDH1 codon 132 and IDH2 codon 172 were directly sequenced and the amplification of exon 20 of EGFR gene was investigated by qPCR in 65 glioblastoma tumors diagnosed at the University Hospital of Fez between 2010 and 2014. Results. The R132H IDH1 mutation was observed in 8 of 65 tumor samples (12.31%). No mutation of IDH2 was detected. EGFR amplification was identified in 17 cases (26.15%). Conclusion. A systematic search of both histological and molecular markers should be requisite for a good diagnosis and a better management of glioblastomas.