Spatial determination and prognostic impact of the fibroblast transcriptome in pancreatic ductal adenocarcinoma
- Institute of Immunology and Immunotherapy, College of Medical and Dental Sciences, University of Birmingham, United Kingdom
- Centre for Computational Biology, University of Birmingham, United Kingdom
- Cancer Research Horizons, The Francis Crick Institute, United Kingdom
- University Hospitals Birmingham NHS Foundation Trust, Queen Elizabeth Hospital Birmingham, United Kingdom
Figures
Figure 1
Schematic representation of experimental approach.
Histological slides of 24 surgical resection specimens from patients with PDAC were stained with DAPI (‘nuclear’), anti-pan-CK (‘epithelial’), anti-α-SMA (‘fibroblast’) and anti-CD45 (‘immune’) to …
Figure 2 with 5 supplements
Overview of gene expression data from spatially-defined stromal and immune regions within the PDAC tumor microenvironment using NanoString GeoMx DSP.
(A) Expression profile of all endogenous probes across regions of interest (ROI) with hierarchical clustering of ROIs. (B) UMAP embedding from normalized count data showing all ROIs overlaid with …
Figure 2—figure supplement 1
Raw count expression profiles.
(A) Heatmap of raw expression profile for the complete probeset (colour scale = raw count). (B) Per patient (scan) expression distributions for PanCK+ (tumor) and PanCK- (non-tumor) regions of …
Figure 2—figure supplement 2
Housekeeping gene correlations and data normalisation.
(A) Pairwise correlation scatter plots for 6 housekeeping control probes. Raw housekeeping gene counts from all regions of interest (ROIs) were correlated against each other. Plots display the …
Figure 2—figure supplement 3
Correlation matrix of endogenous probes.
Pairwise correlations of normalized gene count data represented as a matrix of Pearson correlation coefficients.
Figure 2—figure supplement 4
Individual gene expression profiles on UMAP embeddings of all regions of interest (ROIs).
UMAP embedding generated from normalized count data of all probes in all REIs overlaid with ROI-specific normalized gene count.
Figure 2—figure supplement 5
High level cell type contexture of PDAC tumor microenvironment.
(A) (left), UMAP embedding of transcriptional data for 17,958 cells in the tumor microenvironment of 3 x PDAC patient tissue samples overlaid with high level cell type identified from unsupervised …
Figure 3 with 1 supplement
Expression signature of PDAC tumor-proximal and tumor-distal stromal cells.
(A) Differentially expressed genes (DEGs) between stroma regions proximal (P) or distal (D) from tumor. Colored points indicate differentially expressed genes (BH adjusted P<0.05 & absolute log2FC >0…
Figure 3—figure supplement 1
Correlations of proximity specific markers DKK3 and C3.
Pairwise Pearson correlations of tumor-proximity specific markers C3 and DKK3 against marker genes for Lymphocytes, Monocytes and Fibroblast cells. Coefficients are highlighted where there are …
Figure 4
Proximal and Distal Fibroblast populations identified in single cell transcriptome data of PDAC.
(A) Average expression of spatially defined tumor-proximal or tumor-distant stromal genes within cell types defined by scRNA-seq (n=3). (B) UMAP embedding of scRNA-Seq data from fibroblasts overlaid …
Figure 5 with 1 supplement
Survival expression signatures within spatially defined regions of PDAC.
(A) Differential gene expression from all regions in relation to poor (<1 year) or good (3+year) survival. Coloured points indicate differentially expressed genes (BH adjusted p<0.05 and absolute …
Figure 5—figure supplement 1
Profile of Survival expression signatures within PDAC TME.
(A) Average scRNA expression profile of Survival associated genes within high level cell types of scRNA-seq dataset (n=3). (B) Expression profile of Survival associated genes identified by 3 yr vs 1 …
Figure 6
Graphical summary of the transcriptional features of spatially defined regions in the PDAC tumor microenvironment.
Tables
Table 1
Morphology marker antibodies.
| Name | Channel | Host | Company | Clone # | Catalog # | Concentration used |
|---|---|---|---|---|---|---|
| SMA | 488 | Mouse | Invitrogen | 1A4 | 53-9760-82 | 1:200 |
| Syto83 | 532 | Thermo fisher | 400 nM | |||
| PanCk | 594 | Mouse | Novus | AE1/AE3 | NBP2-33200DL594 | 1:500 |
| CD45 | 647 | Mouse | Novus | 2B11+PD7/26 | NBP2-34528AF647 | 1:200 |
Additional files
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Supplementary file 1
NanoString nCounter RNA hybridisation probeset.
List of RNA probe panels used for Nanostring nCounter data collection including the immunoncology core panel, fibroblast-specific, housekeeping and negative control probe sets.
- https://cdn.elifesciences.org/articles/86125/elife-86125-supp1-v2.docx
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MDAR checklist
- https://cdn.elifesciences.org/articles/86125/elife-86125-mdarchecklist1-v2.docx
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Source data 1
Raw NanoString nCounter data.
- https://cdn.elifesciences.org/articles/86125/elife-86125-data1-v2.csv
