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Article ; Online: P16 immunohistochemistry is a sensitive and specific surrogate marker for CDKN2A homozygous deletion in gliomas.

Vij, Meenakshi / Cho, Benjamin B / Yokoda, Raquel T / Rashidipour, Omid / Umphlett, Melissa / Richardson, Timothy E / Tsankova, Nadejda M

Acta neuropathologica communications

2023 Volume 11, Issue 1, Page(s) 73

Abstract: Molecular characterization of gliomas has uncovered genomic signatures with significant impact on tumor diagnosis and prognostication. CDKN2A is a tumor suppressor gene involved in cell cycle control. Homozygous deletion of the CDKN2A/B locus has been ... ...

Abstract	Molecular characterization of gliomas has uncovered genomic signatures with significant impact on tumor diagnosis and prognostication. CDKN2A is a tumor suppressor gene involved in cell cycle control. Homozygous deletion of the CDKN2A/B locus has been implicated in both gliomagenesis and tumor progression through dysregulated cell proliferation. In histologically lower grade gliomas, CDKN2A homozygous deletion is associated with more aggressive clinical course and is a molecular marker of grade 4 status in the 2021 WHO diagnostic system. Despite its prognostic utility, molecular analysis for CDKN2A deletion remains time consuming, expensive, and is not widely available. This study assessed whether semi-quantitative immunohistochemistry for expression of p16, the protein product of CDKN2A, can serve as a sensitive and a specific marker for CDKN2A homozygous deletion in gliomas. P16 expression was quantified by immunohistochemistry in 100 gliomas, representing both IDH-wildtype and IDH-mutant tumors of all grades, using two independent pathologists' scores and QuPath digital pathology analysis. Molecular CDKN2A status was determined using next-generation DNA sequencing, with homozygous CDKN2A deletion detected in 48% of the tumor cohort. Classifying CDKN2A status based on p16 tumor cell expression (0-100%) demonstrated robust performance over a wide range of thresholds, with receiver operating characteristic curve area of 0.993 and 0.997 (blinded and unblinded pathologist p16 scores, respectively) and 0.969 (QuPath p16 score). Importantly, in tumors with pathologist-scored p16 equal to or less than 5%, the specificity for predicting CDKN2A homozygous deletion was 100%; and in tumors with p16 greater than 20%, specificity for excluding CDKN2A homozygous deletion was also 100%. Conversely, tumors with p16 scores of 6-20% represented gray zone with imperfect correlation to CDKN2A status. The findings indicate that p16 immunohistochemistry is a reliable surrogate marker of CDKN2A homozygous deletion in gliomas, with recommended p16 cutoff scores of ≤ 5% for confirming and > 20% for excluding biallelic CDKN2A loss.
MeSH term(s)	Humans ; Immunohistochemistry ; Cyclin-Dependent Kinase Inhibitor p16/genetics ; Homozygote ; Sequence Deletion ; Glioma/diagnosis ; Glioma/genetics ; Glioma/pathology ; Biomarkers, Tumor/genetics ; Biomarkers, Tumor/analysis ; Gene Deletion
Chemical Substances	Cyclin-Dependent Kinase Inhibitor p16 ; Biomarkers, Tumor ; CDKN2A protein, human
Language	English
Publishing date	2023-05-03
Publishing country	England
Document type	Journal Article ; Research Support, N.I.H., Extramural
ZDB-ID	2715589-4
ISSN	2051-5960 ; 2051-5960
ISSN (online)	2051-5960
ISSN	2051-5960
DOI	10.1186/s40478-023-01573-2
Database	MEDical Literature Analysis and Retrieval System OnLINE

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Article: Isolation of adult human astrocyte populations from fresh-frozen cortex using fluorescence-activated nuclei sorting

Mussa, Zarmeen / Tome-Garcia, Jessica / Jiang, Yan / Akbarian, Schahram / Tsankova, Nadejda M.

Journal of visualized experiments. 2021 Apr. 16, , no. 170

2021

Abstract: The complexity of human astrocytes remains poorly defined in primary human tissue, requiring better tools for their isolation and molecular characterization. Fluorescence-activated nuclei sorting (FANS) can be used to successfully isolate and study human ...

Abstract	The complexity of human astrocytes remains poorly defined in primary human tissue, requiring better tools for their isolation and molecular characterization. Fluorescence-activated nuclei sorting (FANS) can be used to successfully isolate and study human neuronal nuclei (NeuN+) populations from frozen archival tissue, thereby avoiding problems associated with handling fresh tissue. However, efforts to similarly isolate astroglia from the non-neuronal (NeuN-) element are lacking. A recently developed and validated immunotagging strategy uses three transcription factor antibodies to simultaneously isolate enriched neuronal (NeuN+), astrocyte (paired box protein 6 (PAX6)+NeuN-), and oligodendrocyte progenitor (OLIG2+NeuN-) nuclei populations from non-diseased, fresh (unfixed) snap-frozen postmortem human temporal neocortex tissue. This technique was shown to be useful for the characterization of cell type-specific transcriptome alterations in primary pathological epilepsy neocortex. Transcriptomic analyses confirmed that PAX6+NeuN- sorted populations are robustly enriched for pan-astrocyte markers and capture astrocytes in both resting and reactive conditions. This paper describes the FANS methodology for the isolation of astrocyte-enriched nuclei populations from fresh-frozen human cortex, including tissue dissociation into single-nucleus (sn) suspension; immunotagging of nuclei with anti-NeuN and anti-PAX6 fluorescently conjugated antibodies; FANS gating strategies and quality control metrics for optimizing sensitivity and specificity during sorting and for confirming astrocyte enrichment; and recommended procurement for downstream transcriptome and chromatin accessibility sequencing at bulk or sn resolution. This protocol is applicable for non-necrotic, fresh-frozen, human cortical specimens with various pathologies and recommended postmortem tissue collection within 24 h.
Keywords	astrocytes ; chromatin ; cortex ; dissociation ; epilepsy ; humans ; neocortex ; neurons ; oligodendroglia ; quality control ; transcription factors ; transcriptome ; transcriptomics
Language	English
Dates of publication	2021-0416
Size	p. e62405.
Publishing place	Journal of Visualized Experiments
Document type	Article
ZDB-ID	2259946-0
ISSN	1940-087X
ISSN	1940-087X
DOI	10.3791/62405
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Article: Multiparametric Radiogenomic Model to Predict Survival in Patients with Glioblastoma.

Mahmoudi, Keon / Kim, Daniel H / Tavakkol, Elham / Kihira, Shingo / Bauer, Adam / Tsankova, Nadejda / Khan, Fahad / Hormigo, Adilia / Yedavalli, Vivek / Nael, Kambiz

Cancers

2024 Volume 16, Issue 3

Abstract: Background: Clinical, histopathological, and imaging variables have been associated with prognosis in patients with glioblastoma (GBM). We aimed to develop a multiparametric radiogenomic model incorporating MRI texture features, demographic data, and ... ...

Abstract	Background: Clinical, histopathological, and imaging variables have been associated with prognosis in patients with glioblastoma (GBM). We aimed to develop a multiparametric radiogenomic model incorporating MRI texture features, demographic data, and histopathological tumor biomarkers to predict prognosis in patients with GBM. Methods: In this retrospective study, patients were included if they had confirmed diagnosis of GBM with histopathological biomarkers and pre-operative MRI. Tumor segmentation was performed, and texture features were extracted to develop a predictive radiomic model of survival (<18 months vs. ≥18 months) using multivariate analysis and Least Absolute Shrinkage and Selection Operator (LASSO) regularization to reduce the risk of overfitting. This radiomic model in combination with clinical and histopathological data was inserted into a backward stepwise logistic regression model to assess survival. The diagnostic performance of this model was reported for the training and external validation sets. Results: A total of 116 patients were included for model development and 40 patients for external testing validation. The diagnostic performance (AUC/sensitivity/specificity) of the radiomic model generated from seven texture features in determination of ≥18 months survival was 0.71/69.0/70.3. Three variables remained as independent predictors of survival, including radiomics ( Conclusions: Results show that our radiogenomic model generated from radiomic features at baseline MRI, age, and
Language	English
Publishing date	2024-01-30
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers16030589
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Article ; Online: Isolation of Adult Human Astrocyte Populations from Fresh-frozen Cortex using Fluorescence-Activated Nuclei Sorting.

Mussa, Zarmeen / Tome-Garcia, Jessica / Jiang, Yan / Akbarian, Schahram / Tsankova, Nadejda M

Journal of visualized experiments : JoVE

2021 , Issue 170

Abstract	The complexity of human astrocytes remains poorly defined in primary human tissue, requiring better tools for their isolation and molecular characterization. Fluorescence-activated nuclei sorting (FANS) can be used to successfully isolate and study human neuronal nuclei (NeuN+) populations from frozen archival tissue, thereby avoiding problems associated with handling fresh tissue. However, efforts to similarly isolate astroglia from the non-neuronal (NeuN-) element are lacking. A recently developed and validated immunotagging strategy uses three transcription factor antibodies to simultaneously isolate enriched neuronal (NeuN+), astrocyte (paired box protein 6 (PAX6)+NeuN-), and oligodendrocyte progenitor (OLIG2+NeuN-) nuclei populations from non-diseased, fresh (unfixed) snap-frozen postmortem human temporal neocortex tissue. This technique was shown to be useful for the characterization of cell type-specific transcriptome alterations in primary pathological epilepsy neocortex. Transcriptomic analyses confirmed that PAX6+NeuN- sorted populations are robustly enriched for pan-astrocyte markers and capture astrocytes in both resting and reactive conditions. This paper describes the FANS methodology for the isolation of astrocyte-enriched nuclei populations from fresh-frozen human cortex, including tissue dissociation into single-nucleus (sn) suspension; immunotagging of nuclei with anti-NeuN and anti-PAX6 fluorescently conjugated antibodies; FANS gating strategies and quality control metrics for optimizing sensitivity and specificity during sorting and for confirming astrocyte enrichment; and recommended procurement for downstream transcriptome and chromatin accessibility sequencing at bulk or sn resolution. This protocol is applicable for non-necrotic, fresh-frozen, human cortical specimens with various pathologies and recommended postmortem tissue collection within 24 h.
MeSH term(s)	Adult ; Antigens, Nuclear ; Astrocytes ; Cell Nucleus ; Cerebral Cortex/cytology ; Cytological Techniques ; Fluorescence ; Freezing ; Gene Expression Profiling ; Humans ; Nerve Tissue Proteins ; PAX6 Transcription Factor
Chemical Substances	Antigens, Nuclear ; Nerve Tissue Proteins ; PAX6 Transcription Factor ; PAX6 protein, human ; neuronal nuclear antigen NeuN, human
Language	English
Publishing date	2021-04-16
Publishing country	United States
Document type	Journal Article ; Research Support, N.I.H., Extramural ; Video-Audio Media
ZDB-ID	2259946-0
ISSN	1940-087X ; 1940-087X
ISSN (online)	1940-087X
ISSN	1940-087X
DOI	10.3791/62405
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Article: FACS-based Isolation of Neural and Glioma Stem Cell Populations from Fresh Human Tissues Utilizing EGF Ligand.

Tome-Garcia, Jessica / Doetsch, Fiona / Tsankova, Nadejda M

Bio-protocol

2018 Volume 7, Issue 24

Abstract: Direct isolation of human neural and glioma stem cells from fresh tissues permits their biological study without prior culture and may capture novel aspects of their molecular phenotype in their native state. Recently, we demonstrated the ability to ... ...

Abstract	Direct isolation of human neural and glioma stem cells from fresh tissues permits their biological study without prior culture and may capture novel aspects of their molecular phenotype in their native state. Recently, we demonstrated the ability to prospectively isolate stem cell populations from fresh human germinal matrix and glioblastoma samples, exploiting the ability of cells to bind the Epidermal Growth Factor (EGF) ligand in fluorescence-activated cell sorting (FACS). We demonstrated that FACS-isolated EGF-bound neural and glioblastoma populations encompass the sphere-forming colonies
Language	English
Publishing date	2018-03-04
Publishing country	United States
Document type	Journal Article
ZDB-ID	2833269-6
ISSN	2331-8325
ISSN	2331-8325
DOI	10.21769/BioProtoc.2659
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Article ; Online: Emerging interplay of genetics and epigenetics in gliomas: a new hope for targeted therapy.

Yong, Raymund L / Tsankova, Nadejda M

Seminars in pediatric neurology

2015 Volume 22, Issue 1, Page(s) 14–22

Abstract: Diffusely infiltrating gliomas are inherently heterogeneous tumors, and there are ongoing efforts to establish a classification scheme that incorporates new molecular and traditional histologic features. In less than a decade, high-throughput sequencing ... ...

Abstract	Diffusely infiltrating gliomas are inherently heterogeneous tumors, and there are ongoing efforts to establish a classification scheme that incorporates new molecular and traditional histologic features. In less than a decade, high-throughput sequencing of gliomas has transformed the field, uncovering several pivotal, highly prevalent genetic alterations that stratify patients into different prognostic and treatment-response categories. We highlight the genetic aberrations recently discovered in isocitrate dehydrogenase, alpha thalassemia/mental retardation syndrome X-linked, death-domain-associated protein, histone H3.3, and telomerase reverse transcriptase and discuss how these mutations lead to unexpected changes in the epigenetic landscape in gliomas. We describe the opportunities these discoveries might provide for the development of novel targeted therapy aimed at reversing early epigenetic aberrations in glioma precursor cells. Finally, we discuss the challenges for effective treatment of this fatal disease posed by intratumoral heterogeneity and clonal evolution.
MeSH term(s)	Animals ; Brain Neoplasms/genetics ; Brain Neoplasms/therapy ; Epigenesis, Genetic/genetics ; Glioma/genetics ; Glioma/therapy ; Humans ; Molecular Biology
Language	English
Publishing date	2015-03
Publishing country	United States
Document type	Journal Article ; Review
ZDB-ID	1290000-x
ISSN	1558-0776 ; 1071-9091
ISSN (online)	1558-0776
ISSN	1071-9091
DOI	10.1016/j.spen.2014.12.004
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Article ; Online: Advances in genetic and epigenetic analyses of gliomas: a neuropathological perspective.

Tsankova, Nadejda M / Canoll, Peter

Journal of neuro-oncology

2014 Volume 119, Issue 3, Page(s) 481–490

Abstract: Gliomas, the most common malignant primary brain tumors, are universally fatal once they progress from low-grade into high-grade neoplasms. In recent years, we have accumulated unprecedented data about the genetic and epigenetic abnormalities in gliomas; ...

Abstract	Gliomas, the most common malignant primary brain tumors, are universally fatal once they progress from low-grade into high-grade neoplasms. In recent years, we have accumulated unprecedented data about the genetic and epigenetic abnormalities in gliomas; yet, our appreciation of how these deadly tumors arise is still rudimentary. One of the major deterrents in understanding gliomagenesis is the remarkably complex and heterogeneous molecular composition of gliomas, as well as their ability to change phenotypically as they progress and recur. In the past decade, several monumental studies have begun to define better glioma heterogeneity. Four distinct molecular subgroups have emerged: proneural, classical, mesenchymal, and neural; which have unique gene expression signatures and prognostic significance. Of these, gliomas of the proneural subtype, which encompass most grade II/III diffuse gliomas and secondary glioblastomas and often carry isocitrate dehydrogenase (IDH) mutations, have emerged as a distinct tumor subclass with a notably superior prognosis. Important molecular markers with prognostic relevance, such as mutant IDH1/2, have already been incorporated into clinical neuropathological practice. The recent molecular discoveries in gliomas have also emphasized the intimate link between epigenetics and genetics in gliomagenesis. Several of the novel genetic mutations described are responsible for distinct epigenetic remodeling in gliomas, the mechanisms of which are currently being elucidated. Importantly, these epigenetic and genomic alterations represent new and exciting drug targets for future therapeutic interventions in our continuous fight with this fatal malignancy.
MeSH term(s)	Biomarkers ; Brain Neoplasms/genetics ; Brain Neoplasms/pathology ; Carcinogenesis/genetics ; Carcinogenesis/pathology ; Epigenesis, Genetic ; Glioma/genetics ; Glioma/pathology ; Humans
Chemical Substances	Biomarkers
Language	English
Publishing date	2014-06-25
Publishing country	United States
Document type	Journal Article ; Review
ZDB-ID	604875-4
ISSN	1573-7373 ; 0167-594X
ISSN (online)	1573-7373
ISSN	0167-594X
DOI	10.1007/s11060-014-1499-x
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Zs.A 1825: Show issues

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Article ; Online: Multi-omic profiling of the developing human cerebral cortex at the single-cell level.

Zhu, Kaiyi / Bendl, Jaroslav / Rahman, Samir / Vicari, James M / Coleman, Claire / Clarence, Tereza / Latouche, Ovaun / Tsankova, Nadejda M / Li, Aiqun / Brennand, Kristen J / Lee, Donghoon / Yuan, Guo-Cheng / Fullard, John F / Roussos, Panos

Science advances

2023 Volume 9, Issue 41, Page(s) eadg3754

Abstract: The cellular complexity of the human brain is established via dynamic changes in gene expression throughout development that is mediated, in part, by the spatiotemporal activity of cis-regulatory elements (CREs). We simultaneously profiled gene ... ...

Abstract	The cellular complexity of the human brain is established via dynamic changes in gene expression throughout development that is mediated, in part, by the spatiotemporal activity of cis-regulatory elements (CREs). We simultaneously profiled gene expression and chromatin accessibility in 45,549 cortical nuclei across six broad developmental time points from fetus to adult. We identified cell type-specific domains in which chromatin accessibility is highly correlated with gene expression. Differentiation pseudotime trajectory analysis indicates that chromatin accessibility at CREs precedes transcription and that dynamic changes in chromatin structure play a critical role in neuronal lineage commitment. In addition, we mapped cell type-specific and temporally specific genetic loci implicated in neuropsychiatric traits, including schizophrenia and bipolar disorder. Together, our results describe the complex regulation of cell composition at critical stages in lineage determination and shed light on the impact of spatiotemporal alterations in gene expression on neuropsychiatric disease.
MeSH term(s)	Humans ; Multiomics ; Chromatin/genetics ; Chromatin/metabolism ; Regulatory Sequences, Nucleic Acid ; Cell Differentiation/genetics ; Brain/metabolism
Chemical Substances	Chromatin
Language	English
Publishing date	2023-10-12
Publishing country	United States
Document type	Journal Article
ZDB-ID	2810933-8
ISSN	2375-2548 ; 2375-2548
ISSN (online)	2375-2548
ISSN	2375-2548
DOI	10.1126/sciadv.adg3754
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Article ; Online: The prognostic impact of subclonal IDH1 mutation in grade 2-4 astrocytomas.

Vij, Meenakshi / Yokoda, Raquel T / Rashidipour, Omid / Tran, Ivy / Vasudevaraja, Varshini / Snuderl, Matija / Yong, Raymund L / Cobb, William S / Umphlett, Melissa / Walker, Jamie M / Tsankova, Nadejda M / Richardson, Timothy E

Neuro-oncology advances

2023 Volume 5, Issue 1, Page(s) vdad069

Abstract: Background: Isocitrate dehydrogenase (IDH) mutations are thought to represent an early oncogenic event in glioma evolution, found with high penetrance across tumor cells; however, in rare cases, IDH mutation may exist only in a small subset of the total ...

Abstract	Background: Isocitrate dehydrogenase (IDH) mutations are thought to represent an early oncogenic event in glioma evolution, found with high penetrance across tumor cells; however, in rare cases, IDH mutation may exist only in a small subset of the total tumor cells (subclonal IDH mutation). Methods: We present 2 institutional cases with subclonal Results: Immunohistochemistry (IHC) performed on 2 institutional World Health Organization grade 4 IDH-mutant astrocytomas revealed only a minority of tumor cells in each case with IDH1 R132H mutant protein, and next-generation sequencing (NGS) revealed remarkably low Conclusions: While rare, subclonal
Language	English
Publishing date	2023-05-29
Publishing country	England
Document type	Journal Article
ZDB-ID	3009682-0
ISSN	2632-2498 ; 2632-2498
ISSN (online)	2632-2498
ISSN	2632-2498
DOI	10.1093/noajnl/vdad069
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Article: Multi-Parametric Radiomic Model to Predict 1p/19q Co-Deletion in Patients with IDH-1 Mutant Glioma: Added Value to the T2-FLAIR Mismatch Sign.

Kihira, Shingo / Derakhshani, Ahrya / Leung, Michael / Mahmoudi, Keon / Bauer, Adam / Zhang, Haoyue / Polson, Jennifer / Arnold, Corey / Tsankova, Nadejda M / Hormigo, Adilia / Salehi, Banafsheh / Pham, Nancy / Ellingson, Benjamin M / Cloughesy, Timothy F / Nael, Kambiz

Cancers

2023 Volume 15, Issue 4

Abstract: Purpose: The T2-FLAIR mismatch sign has shown promise in determining IDH mutant 1p/19q non-co-deleted gliomas with a high specificity and modest sensitivity. To develop a multi-parametric radiomic model using MRI to predict 1p/19q co-deletion status in ... ...

Abstract	Purpose: The T2-FLAIR mismatch sign has shown promise in determining IDH mutant 1p/19q non-co-deleted gliomas with a high specificity and modest sensitivity. To develop a multi-parametric radiomic model using MRI to predict 1p/19q co-deletion status in patients with newly diagnosed IDH1 mutant glioma and to perform a comparative analysis to T2-FLAIR mismatch sign+. Methods: In this retrospective study, patients with diagnosis of IDH1 mutant gliomas with known 1p/19q status who had preoperative MRI were included. T2-FLAIR mismatch was evaluated independently by two board-certified neuroradiologists. Texture features were extracted from glioma segmentation of FLAIR images. eXtremeGradient Boosting (XGboost) classifiers were used for model development. Leave-one-out-cross-validation (LOOCV) and external validation performances were reported for both the training and external validation sets. Results: A total of 103 patients were included for model development and 18 patients for external testing validation. The diagnostic performance (sensitivity/specificity/accuracy) in the determination of the 1p/19q co-deletion status was 59%/83%/67% (training) and 62.5%/70.0%/66.3% (testing) for the T2-FLAIR mismatch sign. This was significantly improved ( Conclusion: The proposed radiomic model provides much needed sensitivity to the highly specific T2-FLAIR mismatch sign in the determination of the 1p/19q non-co-deletion status and improves the overall diagnostic performance of neuroradiologists when used as an assistive tool.
Language	English
Publishing date	2023-02-07
Publishing country	Switzerland
Document type	Journal Article
ZDB-ID	2527080-1
ISSN	2072-6694
ISSN	2072-6694
DOI	10.3390/cancers15041037
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