Intratumor morphologic and molecular genetic heterogeneity in astrocytomas of different grade of malignancy in the material from the first operation

Introduction. Intratumor heterogeneity is one of the key reasons for unfavourable prognosis in malignant tumors. Astrocytic tumors are known to develop therapy resistance inevitably during the course of disease. One of possible reason is tumor heterogeneity. Purpose. The aim of this work was to assess the intratumor morphologic and molecular heterogeneity in diffuse astrocytoma, anaplastic astrocytomas and primary glioblastomas. Material and methods. We conducted morphologic (n=22) and molecular-genetic (n=8) analysis of surgical specimens obtained from primarily operated glioblastoma giv (gb), anaplastic astrocytomas giii (aa) and diffuse astrocytoma gii (da) patients aged 18 years and older in whom total or subtotal tumor resection was performed. Tissue sampling for the analysis was performed from 5 equidistant areas of each tumor. Morphologic diagnosis was established according to who classification of central nervous system tumors (2007/2016). Mgmt, c-kit, top2a, pdgfr-α, ercc1, vegf genes mrnaexpression was assessed by rt-pcr. Idh1 and idh2 mutational status was evaluated by allele-specific pcr. Results. Morphologic heterogeneity was evident in 72,7 % tumors (16/22) overall. Heterogeneity was observed in 68,8 % (11/16) of gb, 80 % (4/5) of aa and in the only case of da. In 50 % of cases at least 3 different morphologic variants were seen in different areas of the tumor. This morphologic heterogeneity presented as the combination of different grades of anaplasia (gii – giv) in one tumor. Molecular profile was assessed in 48 expression analysis of genes: mgmt, c-kit, top2a, pdgfr-α, ercc1, vegf from 8 patients. Intratumoral molecular heterogeneity was revealed in 41,7 % of cases (20/48). Conclusion. The presence of intratumoral heterogeneity should be taken into account during surgery for adequate tumor sampling for histologic and molecular analysis which is critical for proper assessment of prognosis and following treatment planning.

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TB-9 An attempt to establish a patient-derived brain tumor culture model by organoid culture method

Neuro-Oncology Advances ◽

10.1093/noajnl/vdab159.025 ◽

2021 ◽

Vol 3 (Supplement_6) ◽

pp. vi7-vi7

Author(s):

Hideki Kuroda ◽

Noriyuki Kijima ◽

Tomoyoshi Nakagawa ◽

Ryuichi Hirayama ◽

Yoshiko Okita ◽

...

Keyword(s):

Brain Tumor ◽

Brain Tumors ◽

Tumor Resection ◽

Culture Method ◽

Molecular Heterogeneity ◽

Low Grade ◽

Diffuse Astrocytoma ◽

True Nature ◽

Culture Models ◽

Establishment Rate

Abstract Background: Molecular heterogeneity among and within tumors are one of the reasons for the poor survival rate of brain tumors even with the current standard therapy. However, monolayer culture and neuro-sphere culture (NS) use exogenous growth factors, so may not show the true nature of the tumor. And the culture establishment rate is low, especially low-grade tumors. Therefore, we used the glioblastoma organoid (GBO) culture method showed by Fadi to create culture models of various brain tumors and investigated their characteristics. Methods: We examined the establishment rate in pathological and genotypic types of 56 patients who underwent brain tumor resection at our hospital between January 2020 and June 2021 and were cultured with GBO or NS. If tumor cells are increased visually at 1 month after culture, we defined establishment. Results: There were 15 cases of glioblastoma, 7 cases of anaplastic astrocytoma, 7 cases of diffuse astrocytoma, 3 cases of diffuse midline glioma, 2 cases of anaplastic oligodendroglioma, 5 cases of oligodendroglioma, and 16 cases of others. The establishment rate was 76.5% by the GBO method and 40% by the N S method. By histological type, GBO: 80% in glioblastoma, NS: 58.3% in glioblastoma, GBO: 83.3% in AA, NS: 40% in AA, and GBO: 100% in DA. The IDH mutation and pTERT mutation were investigated in GBO: IDHwt/TERT+ 87.5%, IDHwt/TERT- 64.3%, IDHmt/TERT- 100%, and in NS: IDHwt/TERT+ 75%, IDHwt/TERT- 33.3%, IDHmt/ TERT- 20% in NS. In addition, establishment was observed in GBO 2 case in medulloblastoma, 1 case in ependymoma. Discussion and Conclusion: This suggest that GBO can be used to establish culture models for low-grade tumors. In addition, GBO can establish culture earlier, so it is expected to be applicable to personalized therapies such as preclinical drug efficacy studies tailored to individual patients.

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NIMG-73. CAPTURING GLIOBLASTOMA HETEROGENEITY USING IMAGING AND DEEP LEARNING: APPLICATION TO MGMT PROMOTER METHYLATION

Neuro-Oncology ◽

10.1093/neuonc/noab196.570 ◽

2021 ◽

Vol 23 (Supplement_6) ◽

pp. vi146-vi146

Author(s):

Sanjay Saxena ◽

Anahita Fathi Kazerooni ◽

Erik Toorens ◽

Spyridon Bakas ◽

Hamed Akbari ◽

...

Keyword(s):

Deep Learning ◽

Promoter Methylation ◽

Tissue Sample ◽

Tumor Resection ◽

Mgmt Promoter Methylation ◽

Intratumor Heterogeneity ◽

Molecular Heterogeneity ◽

Mgmt Methylation ◽

Tissue Samples ◽

Mgmt Promoter

Abstract PURPOSE Intratumor heterogeneity is frequent in glioblastoma (GB), giving rise to the tumor’s resistance to standard therapies and, ultimately, poorer clinical outcomes. Yet heterogeneity is often not quantified when assessing the genomic or methylomic profile of a tumor, when a single tissue sample is analyzed. This study proposes a novel approach to non-invasively characterize heterogeneity across glioblastoma using deep learning analysis MRI scans, using MGMT promoter methylation (MGMTpm) as a test-case, and validates the imaging-derived heterogeneity maps with MGMTpm heterogeneity measured via multiple tissue samples. METHODS Multi-parametric MRI (mpMRI) scans (T1, T1-Gd, T2, T2-FLAIR) of 181 patients with newly diagnosed glioblastoma, who underwent surgical tumor resection and had MGMT methylation assessment results, were retrospectively collected. We trained a 5-fold cross-validated deep convolutional neural network with six convolutional layers for a discovery cohort of 137 patients by placing overlapping regional patches over the whole tumor on mpMRI scans to capture spatial heterogeneity of MGMTpm status in different regions within the tumor. Our approach effectively hypothesized that despite heterogeneity in the training examples, dominant imaging patterns would be captured by deep learning. Trained model was independently applied to an unseen replication cohort of 44 patients, with multiple tissue specimens chosen from different spatial regions within the tumor, allowing us to compare imaging- and tissue-based MGMTpm estimates. RESULTS Our model yielded AUC of 0.75 (95% CI: 0.65–0.79) for global MGMT status prediction, which reflected the heterogeneity in MGMTpm, but also that a dominant imaging pattern of MGMT methylation seemed to emerge. In methylated patients with multiple tissue samples, a significant Pearson's correlation coefficient of 0.64 (p< 0.05) was found between imaging-based heterogeneity maps and MGMTpm heterogeneity. CONCLUSION A novel method based on mpMRI and deep neural networks yielded imaging-based heterogeneity maps that strongly associated with intratumor molecular heterogeneity in MGMT promoter methylated tumors.

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Die Anämie bei malignen Tumorerkrankungen

Nuklearmedizin ◽

10.1055/s-0038-1629498 ◽

1989 ◽

Vol 28 (06) ◽

pp. 247-254

Author(s):

E. Aulbert

Keyword(s):

Malignant Tumors ◽

Gradient Centrifugation ◽

Tumor Resection ◽

Hemoglobin Concentration ◽

Limiting Factor ◽

Novikoff Hepatoma ◽

Tumor Mass ◽

Lysosomal Fraction ◽

Proliferation Activity ◽

Morris Hepatoma

The cellular uptake and lysosomal accumulation of 67Ga-labelled transferrin within tumors of different malignancy were examined using tissue fractionation and immunological techniques. As tumor models the slowly growing Morris hepatoma 5123C, the moderately growing Novikoff hepatoma and the fast and aggressive Yoshida hepatoma AH 130 were investigated. Isolation of subcellular fractions of tumor homogenates was performed by differential centrifugation and density-gradient centrifugation. The intracellular 67Gatransferrin was found to be highly concentrated within the purified lysosomes. The transferrin within the lysosomal fraction was identified by radial immunodiffusion technique using monospecific antiserum. The accumulation of 67Gatransferrin by the tumors resulted in a faster disappearance of 67Ga-transferrin from the blood. This loss of circulating 67Ga-transferrin correlated with the proliferation activity and the spread of the tumors. Since transferrin is indispensible for the utilization of iron by the heme-synthesizing red cell precursors, transferrin concentration in the blood is the limiting factor for the utilization of iron in hemoglobin synthesis. Thus, in a further series of experiments we investigated the development of anemia in tumor-bearing rats. With increasing tumor mass a progressive fall of hemoglobin concentration was found. The anemia was more severe in the faster growing Novikoff hepatoma than in the slowly growing Morris hepatoma. The most significant reduction of hemoglobin concentration was found in the very fast growing Yoshida hepatoma. After total tumor resection hemoglobin concentration and red blood cell count normalized completely within 6-8 weeks. We conclude from these data that the uptake of transferrin by the tumor cells results in a faster disappearance of transferrin from the blood. This loss of circulating transferrin correlates with tumor mass and proliferation activity and is one of the factors responsible for the anemia seen in patients with malignant tumors.

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Molecular diagnostics helps to identify distinct subgroups of spinal astrocytomas

Acta Neuropathologica Communications ◽

10.1186/s40478-021-01222-6 ◽

2021 ◽

Vol 9 (1) ◽

Author(s):

Annamaria Biczok ◽

Felix L. Strübing ◽

Julia M. Eder ◽

Rupert Egensperger ◽

Oliver Schnell ◽

...

Keyword(s):

Spinal Cord ◽

Molecular Markers ◽

Prognostic Significance ◽

The Elderly ◽

Molecular Profile ◽

Diffuse Astrocytoma ◽

Molecular Alterations ◽

High Grade Astrocytoma ◽

Dismal Prognosis ◽

Few Data

AbstractPrimary spinal cord astrocytomas are rare, hence few data exist about the prognostic significance of molecular markers. Here we analyze a panel of molecular alterations in association with the clinical course. Histology and genome sequencing was performed in 26 spinal astrocytomas operated upon between 2000 and 2020. Next-generation DNA/RNA sequencing (NGS) and methylome analysis were performed to determine molecular alterations. Histology and NGS allowed the distinction of 5 tumor subgroups: glioblastoma IDH wildtype (GBM); diffuse midline glioma H3 K27M mutated (DMG-H3); high-grade astrocytoma with piloid features (HAP); diffuse astrocytoma IDH mutated (DA), diffuse leptomeningeal glioneural tumors (DGLN) and pilocytic astrocytoma (PA). Within all tumor entities GBM (median OS: 5.5 months), DMG-H3 (median OS: 13 months) and HAP (median OS: 8 months) showed a fatal prognosis. DMG-H3 tend to emerge in adolescence whereas GBM and HAP develop in the elderly. HAP are characterized by CDKN2A/B deletion and ATRX mutation. 50% of PA tumors carried a mutation in the PIK3CA gene which is seemingly associated with better outcome (median OS: PIK3CA mutated 107.5 vs 45.5 months in wildtype PA). This exploratory molecular profiling of spinal cord astrocytomas allows to identify distinct subgroups by combining molecular markers and histomorphology. DMG-H3 tend to develop in adolescence with a similar dismal prognosis like GBM and HAP in the elderly. We here describe spinal HAP with a distinct molecular profile for the first time.

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Discovery of Spatial Peptide Signatures for Neuroblastoma Risk Assessment by MALDI Mass Spectrometry Imaging

Cancers ◽

10.3390/cancers13133184 ◽

2021 ◽

Vol 13 (13) ◽

pp. 3184

Author(s):

Zhiyang Wu ◽

Patrick Hundsdoerfer ◽

Johannes H. Schulte ◽

Kathy Astrahantseff ◽

Senguel Boral ◽

...

Keyword(s):

Mass Spectrometry ◽

Risk Assessment ◽

Mass Spectrometry Imaging ◽

Principal Component ◽

Therapy Resistance ◽

Risk Classification ◽

Intratumor Heterogeneity ◽

Ionization Mass ◽

Characteristic Analysis ◽

Analytical Strategy

Risk classification plays a crucial role in clinical management and therapy decisions in children with neuroblastoma. Risk assessment is currently based on patient criteria and molecular factors in single tumor biopsies at diagnosis. Growing evidence of extensive neuroblastoma intratumor heterogeneity drives the need for novel diagnostics to assess molecular profiles more comprehensively in spatial resolution to better predict risk for tumor progression and therapy resistance. We present a pilot study investigating the feasibility and potential of matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI-MSI) to identify spatial peptide heterogeneity in neuroblastoma tissues of divergent current risk classification: high versus low/intermediate risk. Univariate (receiver operating characteristic analysis) and multivariate (segmentation, principal component analysis) statistical strategies identified spatially discriminative risk-associated MALDI-based peptide signatures. The AHNAK nucleoprotein and collapsin response mediator protein 1 (CRMP1) were identified as proteins associated with these peptide signatures, and their differential expression in the neuroblastomas of divergent risk was immunohistochemically validated. This proof-of-concept study demonstrates that MALDI-MSI combined with univariate and multivariate analysis strategies can identify spatially discriminative risk-associated peptide signatures in neuroblastoma tissues. These results suggest a promising new analytical strategy improving risk classification and providing new biological insights into neuroblastoma intratumor heterogeneity.

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A Systems Approach to Brain Tumor Treatment

Cancers ◽

10.3390/cancers13133152 ◽

2021 ◽

Vol 13 (13) ◽

pp. 3152

Author(s):

James H. Park ◽

Adrian Lopez Garcia de Lomana ◽

Diego M. Marzese ◽

Tiffany Juarez ◽

Abdullah Feroze ◽

...

Keyword(s):

Brain Tumor ◽

Brain Tumors ◽

Rational Design ◽

Clinical Medicine ◽

Systems Approach ◽

Therapy Resistance ◽

High Rate ◽

Intratumor Heterogeneity ◽

Primary Therapy ◽

Molecular Features

Brain tumors are among the most lethal tumors. Glioblastoma, the most frequent primary brain tumor in adults, has a median survival time of approximately 15 months after diagnosis or a five-year survival rate of 10%; the recurrence rate is nearly 90%. Unfortunately, this prognosis has not improved for several decades. The lack of progress in the treatment of brain tumors has been attributed to their high rate of primary therapy resistance. Challenges such as pronounced inter-patient variability, intratumoral heterogeneity, and drug delivery across the blood–brain barrier hinder progress. A comprehensive, multiscale understanding of the disease, from the molecular to the whole tumor level, is needed to address the intratumor heterogeneity resulting from the coexistence of a diversity of neoplastic and non-neoplastic cell types in the tumor tissue. By contrast, inter-patient variability must be addressed by subtyping brain tumors to stratify patients and identify the best-matched drug(s) and therapies for a particular patient or cohort of patients. Accomplishing these diverse tasks will require a new framework, one involving a systems perspective in assessing the immense complexity of brain tumors. This would in turn entail a shift in how clinical medicine interfaces with the rapidly advancing high-throughput (HTP) technologies that have enabled the omics-scale profiling of molecular features of brain tumors from the single-cell to the tissue level. However, several gaps must be closed before such a framework can fulfill the promise of precision and personalized medicine for brain tumors. Ultimately, the goal is to integrate seamlessly multiscale systems analyses of patient tumors and clinical medicine. Accomplishing this goal would facilitate the rational design of therapeutic strategies matched to the characteristics of patients and their tumors. Here, we discuss some of the technologies, methodologies, and computational tools that will facilitate the realization of this vision to practice.

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Longitudinal single-cell profiling reveals molecular heterogeneity and tumor-immune evolution in refractory mantle cell lymphoma

Nature Communications ◽

10.1038/s41467-021-22872-z ◽

2021 ◽

Vol 12 (1) ◽

Author(s):

Shaojun Zhang ◽

Vivian Changying Jiang ◽

Guangchun Han ◽

Dapeng Hao ◽

Junwei Lian ◽

...

Keyword(s):

Mantle Cell Lymphoma ◽

Single Cell ◽

Communication Networks ◽

Immune Cell ◽

Cell Lymphoma ◽

Survivin Expression ◽

Therapy Resistance ◽

Molecular Heterogeneity ◽

Multiple Cancer ◽

Mantle Cell

AbstractThe mechanisms driving therapeutic resistance and poor outcomes of mantle cell lymphoma (MCL) are incompletely understood. We characterize the cellular and molecular heterogeneity within and across patients and delineate the dynamic evolution of tumor and immune cell compartments at single cell resolution in longitudinal specimens from ibrutinib-sensitive patients and non-responders. Temporal activation of multiple cancer hallmark pathways and acquisition of 17q are observed in a refractory MCL. Multi-platform validation is performed at genomic and cellular levels in PDX models and larger patient cohorts. We demonstrate that due to 17q gain, BIRC5/survivin expression is upregulated in resistant MCL tumor cells and targeting BIRC5 results in marked tumor inhibition in preclinical models. In addition, we discover notable differences in the tumor microenvironment including progressive dampening of CD8+ T cells and aberrant cell-to-cell communication networks in refractory MCLs. This study reveals diverse and dynamic tumor and immune programs underlying therapy resistance in MCL.

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Transcriptome profiles of stem-like cells from primary breast cancers allow identification of ITGA7 as a predictive marker of chemotherapy response

British Journal of Cancer ◽

10.1038/s41416-021-01484-w ◽

2021 ◽

Author(s):

Noha Gwili ◽

Stacey J. Jones ◽

Waleed Al Amri ◽

Ian M. Carr ◽

Sarah Harris ◽

...

Keyword(s):

Breast Cancer ◽

Cancer Cells ◽

Breast Cancer Cells ◽

Predictive Marker ◽

Therapy Resistance ◽

Differentially Expressed ◽

Molecular Profile ◽

Chemotherapy Response ◽

Breast Cancers

Abstract Background Breast cancer stem cells (BCSCs) are drivers of therapy-resistance, therefore are responsible for poor survival. Molecular signatures of BCSCs from primary cancers remain undefined. Here, we identify the consistent transcriptome of primary BCSCs shared across breast cancer subtypes, and we examine the clinical relevance of ITGA7, one of the genes differentially expressed in BCSCs. Methods Primary BCSCs were assessed using immunohistochemistry and fluorescently labelled using Aldefluor (n = 17). Transcriptomes of fluorescently sorted BCSCs and matched non-stem cancer cells were determined using RNA-seq (n = 6). ITGA7 expression was examined in breast cancers using immunohistochemistry (n = 305), and its functional role was tested using siRNA in breast cancer cells. Results Proportions of BCSCs varied from 0 to 9.4%. 38 genes were significantly differentially expressed in BCSCs; genes were enriched for functions in vessel morphogenesis, motility, and metabolism. ITGA7 was found to be significantly downregulated in BCSCs, and low expression significantly correlated with reduced survival in patients treated with chemotherapy, and with chemoresistance in breast cancer cells in vitro. Conclusions This study is the first to define the molecular profile of BCSCs from a range of primary breast cancers. ITGA7 acts as a predictive marker for chemotherapy response, in accordance with its downregulation in BCSCs.

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Intratumor Heterogeneity in Uveal Melanoma BAP-1 Expression

Cancers ◽

10.3390/cancers13051143 ◽

2021 ◽

Vol 13 (5) ◽

pp. 1143

Author(s):

Gustav Stålhammar ◽

Hans E. Grossniklaus

Keyword(s):

Gene Expression ◽

Uveal Melanoma ◽

Tumor Size ◽

Hot Spots ◽

Malignant Tumors ◽

Intratumor Heterogeneity ◽

Prognostic Information ◽

Size Category ◽

Cold Spots ◽

Metastatic Risk

Malignant tumors are rarely homogenous on the morphological, genome, transcriptome or proteome level. In this study, we investigate the intratumor heterogeneity of BAP-1 expression in uveal melanoma with digital image analysis of 40 tumors. The proportion of BAP-1 positive cells was measured in full tumor sections, hot spots, cold spots and in scleral margins. The mean difference between hot spots and cold spots was 41 percentage points (pp, SD 29). Tumors with gene expression class 1 (associated with low metastatic risk) and 2 (high metastatic risk) had similar intratumor heterogeneity. Similarly, the level of intratumor heterogeneity was comparable in tumors from patients that later developed metastases as in patients that did not. BAP-1 measured in any tumor region added significant prognostic information to both American Joint Committee on Cancer (AJCC) tumor size category (p ≤ 0.001) and gene expression class (p ≤ 0.04). We conclude that there is substantial intratumor heterogeneity in uveal melanoma BAP-1 expression. However, it is of limited prognostic importance. Regardless of region, analysis of BAP-1 expression adds significant prognostic information beyond tumor size and gene expression class.

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P06.01 Pediatric non-pontine diffuse midline glioma H3K27M mutant: a monoinstitutional experience

Neuro-Oncology ◽

10.1093/neuonc/noab180.079 ◽

2021 ◽

Vol 23 (Supplement_2) ◽

pp. ii24-ii24

Author(s):

E Schiavello ◽

V Biassoni ◽

L De Cecco ◽

E Pecori ◽

S Filippo ◽

...

Keyword(s):

Liquid Biopsy ◽

P53 Mutation ◽

New Drugs ◽

Molecular Profile ◽

Histopathological Diagnosis ◽

Diffuse Astrocytoma ◽

Number Of Patients ◽

Pathological Evaluation ◽

Diffuse Midline Glioma ◽

Astrocytoma Grade

Abstract BACKGROUND The new entity “diffuse-midline-glioma H3K27M-mutant”(DMG) was defined in the 2016 WHO classification. This tumor subtype, regardless of histological features, is associated with an aggressive clinical behavior and poor prognosis, sharing the same outcome as DIPG. MATERIAL AND METHODS We report our experience from 2016 to 2018 in treating patients with radiotherapy and concomitant Nimotuzumab/Vinorelbine, as already published for DIPG’s patients (DOI10.1007/s11060-014-1428-z). All patients’ parents provided written informed consent for treatment. RESULTS Patients were 9: 7/9 females, median age at diagnosis 13 years (range 1–26); 8 with localized disease, 1 with spinal fluid (CSF) dissemination. Five patients had thalamic disease (1 bithalamic), 2 ponto-cerebellar (one with 2 lesions), 1 pineal gland and one with disseminated CSF disease at diagnosis. 3/9 were biopsed in thalamus, the others had partial resection. Central neuropathological review with confirmed histopathological diagnosis of DMG mutated was necessary for inclusion. At the first pathological evaluation tumors had been classified according to the fourth edition of the WHO 2007 as glioblastoma (n=4), anaplastic astrocytoma grade III (n=1), diffuse astrocytoma grade II (n=1), and 1 glial tumor with oligodendroglial features; 2 patients already had DMG at the first pathological evaluation. Immunohistochemical analysis revealed p53 mutation in 3 patients. All 9 patients were evaluated on serial plasma samples with NGS targeted panel (Pan-Cancer) and 6/9 presented p53 mutation in at least one of the samples. 8/9 expressed MAP2K1 gain of function at liquid biopsy. The small number of patients did not allow correlations with the prognosis. Two patients completed the scheduled 2 years of treatment: one had a local relapse at 37 months after diagnosis, the other is alive without evidence of progression at 30 months. With a median follow up of 14.4 months, PFS and OS were 33% and 77% at 1 year; OS was 22% at 2 years, with 2 patients long survivors at 36 and 40 months after diagnosis. CONCLUSION The molecular and phenotypic pattern of DMG allows to include patients in clinical trials/registry shared with patients suffering from DIPG. Our knowledge is still limited about this entity; new insights into molecular profile should help us to study new drugs directed against the effects of the H3K27M mutations and to define new diagnostic/prognostic approach as liquid biopsy able to correlate treatments and prognosis

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