Combinational test of DGKi with PD-1 blocker
scTCR-seq + scRNA-seq
DGKi + Nivolumab combinational therapy
Introduction
- DGKs are physiological regulators of T-cell
development,differentiation, and function.
- Inhibition of DGK may restore the T-cell activation program
- Enhanced effects may be seen with combination therapy (Nivolumab +
DGKi)
Data
Sample name: P*****
Condition (1 control, 3 test conditions): CONTROL, CND1, CND2,
COMBO
Number of cells : ~9000 cells
Initial Processing
## perform default analysis
perform_default_analysis <- function(obj.srt, n_features = 2000, n_pcs = 20,
dims_for_neighbors = 1:10,
resolutions = c(0.2, 0.5),
umap_dims = 1:10) {
# Step 1: Find variable features
obj.srt <- FindVariableFeatures(obj.srt,
selection.method = 'vst',
nfeatures = n_features)
# Step 2: Scale and normalize data
all_genes <- rownames(obj.srt)
obj.srt <- NormalizeData(obj.srt)
obj.srt <- ScaleData(obj.srt, features = all_genes)
# Step 3: Run PCA
obj.srt <- RunPCA(obj.srt,
features = VariableFeatures(object = obj.srt), npcs = n_pcs)
# Step 4: Find neighbors
obj.srt <- FindNeighbors(obj.srt, dims = dims_for_neighbors)
# Step 5: Find clusters
obj.srt <- FindClusters(obj.srt, resolution = resolutions)
# Step 6: Run UMAP
obj.srt <- RunUMAP(obj.srt, dims = umap_dims)
# Return the Seurat object with analysis results
return(obj.srt)
}
# apply
obj.srt <- perform_default_analysis(obj.srt)
Numer of cells
UMAP (sample)
Colored by condition
Colored by each condition
Clustering
Clusters
- resolution : 0.2
- resolution : 0.5
cluster and condition (res 0.1)
TABLE1 : Number of cells (res 0.2)
HEATMAP1 : Percentage of cells (res 0.2)
cluster and condition (res 0.5)
TABLE2 : Number of cells (res 0.5)
HEATMAP2 : Percentage of cells (res 0.5)
Clonotype Diversity
CTRL
CND1
CND2
COMB
scRepertoire
Clonotype analysis using scRepertoire package
Analysis below represents the basic analysis using the package.
Quantify Clonotypes
Unique clonotypes by TRA chain
Unique clonotypes by TRB chain
Clonotype Abundance
Length of Clonotypes
Compare Clonotypes
Visualize Gene Usage
More Advanced Clonal Analysis
Diversity Analysis
Specific cell type + clonotype
Cytotoxic T Cell Markers and associated clonotypes
Purpose : Try to find the clonotypes with strong cytotoxic T cell marker gene expressions, expecting functional clonotypes to fight the disease.
Cytotoxic T cell markers :
Mostly enzymes that kill target cells.
- IFNG (Increase when infection)
- GZMA (Granzyme A)
- GZMB (Granzyme B)
- NKG7 (NK cell granule protein 7)
Reordered by clonotype version
Note: No clear pattern across cytotoxic gene expression by clonotypes.