Searching the Database
Under the ISH tab, there are several ways to search the database.
Specimen Search
To search for a specific glioblastoma specimen type, filter your search criteria by a specific gene, tumor feature, study or specific clinical conditions.
To refine your search by specific clinical conditions, click the “+” next to “Additional Filters By Clinical Conditions”.
If you are unsure of where to begin your search, you can browse by the tumor information where the columns are described below.
| Column | Definition |
|---|---|
| Column | Definition |
| Tumor | Specimen ID of the tumor, linking to the Specimen Detail Page |
| Molecular Subtype | Classical: chromosome 7 amplification (EGFR), paired with chromosome 10 loss. Mesenchymal: noted for focal hemizygous deletions of a region at 17q11.2 (NF1). Proneural: defined by alterations of PDGFRA and point mutations in IDH1. Neural: typified by the expression of neuronal markers such as NEFL, GABRA1, SYT1, and SLC12A5. |
| Extent of Resection | Degree to which tumor tissue was surgically removed |
| Surgery | Whether the tumor was a primary surgery, or removal of a subsequent recurrent tumor |
| MGMT Methylation | A diagnostic test result referring to whether a tumor’s O-6-methylguanine-DNA methyltransferase DNA repair gene was methylated |
| Survival Days | Number of days from the initial diagnosis |
| EGFR Amplification | A diagnostic test result referring to whether a particular tumor’s epidermal growth factor receptor (EGFR) gene had multiple copies |
| Initial KPS | Karnofsky Performance Status is a measure of functional impairment (on a scale of 0-100, 0 referring to dead, 50 to requiring considerable assistance and frequent medical care, and 100 referring to no evidence of disease) |
| Age | Age in years at the time of diagnosis |
| SNI Link | Link to the Swedish Neurological Institute page for this donor (Registration required) |
Gene Search
The genes chosen for this project were selected based upon current knowledge of GBM biology and what was learned from each of the individual Ivy GAP Studies. To see a list of genes for each study, please see the Gene Lists whitepaper in Documentation. Typing in the gene search text box will bring up a list of suggested genes. If your gene of interest was not assayed in this project, no results will be returned. Once you have chosen a gene, either by selecting a gene from the suggested list, or hitting “Enter” after typing in the name, symbol or ID, a list of specimen blocks that contain search results fulfilling your criteria will open.
Tumor Feature
Each of the tumor structural features was identified and labeled in ~12,000 H&E histological images using a semi-automated annotation application based on advanced statistical machine learning algorithms, and can be searched and visualized in the Ivy GAP web application.
Tumor features are described using a specific ontology. The Ivy GAP ontology is a hierarchical organization of glioblastoma anatomic structural features and associated transcriptomes from RNA-Seq samples that were identified by reference histology or reference gene expression patterns. It does not reflect ontological origins of the structural features; it is simply a representation of relationships among the glioblastoma features and gene expression patterns associated with putative cancer stem cell clusters that were identified in the project.
The structural features are commonly identified by neuropathologists in glioblastoma tissue sections stained with Hematoxylin and Eosin (H&E). The major structural regions are Leading Edge (LE), Infiltrating Tumor (IT), and Cellular Tumor (CT).
Within each of these regions, particular structural features can be found such as Microvascular Proliferation (MVP), Pseudopalisading Cells around Necrosis (PAN), Hyperplastic Blood Vessels (HBV), and Necrosis (NE). These features are routinely used to distinguish glioblastoma, or Grade IV glioma, from lower grades of glioma. The feature HBV can be observed occasionally in the LE and IT regions, but it as well as MVP, NE, and PAN are frequently identified in the CT region.
Specimen Search Results
| Column | Description |
|---|---|
| Specimen | Clicking on this link will take you to a Specimen Detail Page |
| Study | Indicates which study the samples were assayed |
| Donor | Donor age and gender |
| Location | Location in the brain from which the tumor was removed |
| Molecular Subtype | Classical: chromosome 7 amplification (EGFR), paired with chromosome 10 loss. Mesenchymal: noted for focal hemizygous deletions of a region at 17q11.2 (NF1). Proneural: defined by alterations of PDGFRA and point mutations in IDH1. Neural: typified by the expression of neuronal markers such as NEFL, GABRA1, SYT1, and SLC12A5. |
| Normalized Area | The fraction of the selected “structure” in the specimen sub-block |
Differential Search
Similar to other Atlases in the Allen Brain Atlas resources, you can search for enriched gene expression in a tumor feature or structure by comparing expression between your target structure and a contrast structure. By default, this search uses all the data as contrast, but changing the contrast structure to more define your search will produce different results. Changing the threshold on a differential search will restrict the list of returned results by excluding any experiment where the expression energy of the target structures is less than threshold.
Several differential searches have already been calculated for you and are accessible by selecting from the “Browse by Differential Expression”.
Differential Search Results
Performing a differential search will return a list of specimen blocks that include the genes with the highest differential expression levels ranked by fold change in the target structure over the contrast structure.
| Column Header | Description |
|---|---|
| Column Header | Description |
| Entrez ID | Unique identifier from the National Center for Biotechnology Information (NCBI) database for gene-specific information |
| Gene Symbol | The gene symbol, which is a short abbreviated form of the gene name. Clicking on this link will take you to all GBM experiments performed on this gene |
| Specimen | Clicking on this link will take you to a Specimen Detail Page |
| Donor | Donor age and gender |
| Molecular Subtype | Classical: chromosome 7 amplification (EGFR), paired with chromosome 10 loss. Mesenchymal: noted for focal hemizygous deletions of a region at 17q11.2 (NF1). Proneural: defined by alterations of PDGFRA and point mutations in IDH1. Neural: typified by the expression of neuronal markers such as NEFL, GABRA1, SYT1, and SLC12A5. |
| Target Sum | Sum of gene expression energy over all the target structures |
| Fold Change | The measure describing how gene expression changes when comparing the target structure(s) expression energy to the contrast structure(s) expression energy |
Expression Search
To see gene expression ranked by expression energy within a single glioblastoma structure, use the Expression Search feature. Clicking on a tumor feature from the Glioblastoma Ontology or the “Browse by Expression” box will return a list of genes ranked by the average expression where each column is described below.
| Column | Description |
|---|---|
| Entrez ID | Unique identifier from the National Center for Biotechnology Information (NCBI) database for gene-specific information |
| Gene Symbol | The official gene symbol, which is a short abbreviated form of the gene name. Clicking on this link will take you to all GBM experiments performed on this gene |
| Specimen | Clicking on this link will take you to a Specimen Detail Page |
| Donor | Donor age and gender |
| Molecular Subtype | Classical: chromosome 7 amplification (EGFR), paired with chromosome 10 loss. Mesenchymal: noted for focal hemizygous deletions of a region at 17q11.2 (NF1). Proneural: defined by alterations of PDGFRA and point mutations in IDH1. Neural: typified by the expression of neuronal markers such as NEFL, GABRA1, SYT1, and SLC12A5. |
| Average Expression | The sum of expressing pixels divided by sum product of pixels of that tumor feature |
Clinical Conditions
When searching for specific images, one of the search tools available to you is to “Filter by Clinical Condition”. Each condition and its significance is outlined below. Listed references can be found in the Overview whitepaper in Documentation.
Molecular Subtype
Genomic characteristics of tissue block related to classification of patients. The molecular subtype was determined using all cellular tumor samples from a given tumor. An analysis of each cellular tumor sample was conducted with 840 transcripts from the RNA-Seq data as per Verhaak et al., 2010. For a given tumor, if multiple cellular tumor samples exhibited distinct subtypes, then the tumor was classified as a mixture of its subtypes. The Classical subtype refers to chromosome 7 amplification, specifically EGFR, paired with chromosome 10 loss. The Mesenchymal subtype is noted for focal hemizygous deletions of a region at 17q11.2, containing the gene NF1. The Proneural subtype is defined by alterations of PDGFRA and point mutations in IDH1. The Neural subtype is typified by the expression of neuronal markers such as NEFL, GABRA1, SYT1, and SLC12A5.
Extent of Resection
Degree to which tumor tissue was surgically removed. Complete resection is associated with increased survival rates (Keles et al., 1999).
Multifocality
Whether a tumor was comprised of single or multiple masses. Increased multifocality is associated with decreased rates of survival (Thomas et al., 2013).
MGMT Methylation
A diagnostic test result referring to whether a tumor’s O-6-methylguanine-DNA methyltransferase DNA repair gene was methylated. Methylated MGMT is associated with increased rates of survival (Hegi et al., 2013).
Survival Days
Patient’s lifespan since initial diagnosis
EGFR Amplification
A diagnostic test result referring to whether a particular tumor’s epidermal growth factor receptor gene had multiple copies. The gene is frequently amplified in glioblastoma (Hobbs et al., 2012).
Initial KPS
Karnofsky Performance Status when first diagnosed with first tumor. Karnofsky Performance Status is a measure of functional impairment on a scale of 0-100, 0 referring to dead, 50 to requiring considerable assistance and frequent medical care, and 100 referring to no evidence of disease. High scores are associated with increased survival rates (Ening et al., 2015).
Age
Patient age in years at time of diagnosis. Younger patients tend to have increased survival rates (Ening et al., 2015).
Specimen Detail Page
Donor Page
Clicking on a Specimen ID will bring you to the specimen detail page.
- Tumor Subdivision - Navigation of tumor subdivisions and button for viewing patient and tumor summary information. the color in a tissue sub-block indicates the study in which the sub-block was used.
- Tumor Features in this Sub-Block - The glioblastoma tumor features that were identified and assayed for in the yellow highlighted sub-block. The color coding corresponds to the glioblastoma ontology.
- ISH Image - An image viewer showing the in situ hybridization experiment, and related features.
- Nearest H&E Image - Image and annotated anatomic tumor features of a nearby H&E stained tissue section.
- Section Information - Lists the Gene, Experiment ID, Section #, Treatments, and the study.
- Genes Surveyed in this Sub-Block - All genes assayed in this specimen sub-block are listed. To view a specific gene, select only that gene from the list.
- Selection Cart - When viewing an ISH image, select the checkbox to add the image to your viewing cart. Images selected in your cart are available to view until the cache is cleared or you click “Clear Selections”.
Patient, Tumor and MRI Summary
This panel displays information on patient and tumor, and on how the tumor was subdivided into blocks. This panel is arrived at by toggling the “View Patient, Tumor and MRI Summary” button in the Tumor Subdivision section of the Specimen Detail Page.
Description of Donor ID
The tumor names (or IDs) identify tumors across the different resources associated with the Ivy Glioblastoma Atlas Project. In the name W1-1-2, W1 identifies the patient, and -1-2 indicate that this tumor was the second mass removed during the first surgery.
The names of the tumor blocks were assigned as indicated in the Resected Tumor Image, and the names of the sub-blocks reflect their position within the tumor block.
Image Viewer
The GBM image viewer allows you to navigate through a series of specimen sub-block ISH images to view at higher resolution. Once a gene image is selected, you can manipulate it with your mouse and use the keyboard or on screen navigation tools to take additional actions.
The gene symbol and gene name are displayed in the title bar along with a dropdown menu and a “Sync” feature to allow you to navigate and keep the adjacent H&E viewer synched with your movements. The dropdown menu allows you to view the ISH image, the Expression Mask, the annotated features, or the annotated feature boundaries. If all the genes are selected in Box #6, the entire specimen sub-block can be viewed using the onscreen navigation tools or the keyboard commands. The current selection is outlined in black.
To view images at high resolution, in the Section Information box, click on the Experiment link; then, click the icon in the upper right corner of the viewer. Use the dropdown menu to select the type of image you want to see.
Alternatively, to view all the images available for a given gene, in the Section Information box, click on the gene symbol link.
Scale Bar
Shows the current viewing resolution of the image, in microns. This value dynamically changes as you zoom in/out of the image. You can position the scale bar anywhere on the main image by dragging the scale bar by its ruler.
You can toggle the orientation of the scale bar from horizontal to vertical by clicking on the scale bar text.
Keyboard Commands
| Key | Description |
|---|---|
| Key | Description |
| F | Advance to the next image from the specimen |
| D | Go back to the previous image from the specimen |
| R | Advance to the last image in the series |
| E | Go back to the first image in the series |
| A | Zoom in |
| Z | Zoom out |
Expression Mask Colors
The Expression mask image display highlights those cells that have the highest probability of gene expression using a heatmap color scale (from low/blue to high/red).
