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08_merge-all-cells_post-Clustering.Rmd
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---
title: "R Notebook"
output: html_notebook
---
```{r, import libraries}
library(igraph)
library(SingleCellExperiment)
library(S4Vectors)
library(stringr)
library(DT)
library(dplyr)
library(tidyr)
library(mclust)
library(ggplot2)
library(RColorBrewer)
library(scater)
library(Rphenoannoy)
set.seed(101100)
```
```{r}
final.fibro <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL_Fibros.rds")
final.fibro.other <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_fibros_workingfile_OTHER.rds")
final.fibro.HEV <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_fibros_workingfile_HEV.rds")
final.vessel <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/vessel-VESSEL.rds")
final.vessel.tumour <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_vessel_workingfile_TUMOUR.rds")
final.stroma.tumour <- readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_stroma_workingfile_TUMOUR.rds")
final.immune.tumour <- readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_immune_workingfile_TUMOUR.rds")
final.immune.nonTcell <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/NonTcell_immune_afterStroma_Tcell-merge_workingfile.rds")
final.immune.Tcell <- readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL_Tcells-minusImmune_workingfile.rds")
final.tumour.tumour <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/merge_all-tumour_TUMOUR.rds")
final.nontumour.undefined <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/merge_all-NONtumour_final_workingfile_UNDEFINED.rds")
final.nontumour.tumour <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/merge_all-NONtumour_final_workingfile_TUMOUR.rds")
```
```{r}
#final.fibro
colData(final.fibro) %>% colnames()
final.fibro$cell_category <- "Fibroblast"
final.fibro$cell_type <- final.fibro$CAFtype
final.fibro$cell_subtype <- final.fibro$CAFsubtype
cur_DF <- colData(final.fibro)%>% data.frame() %>% select(-contains(c("som_","rp_","CAF"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.fibro$ImageNumber, final.fibro$CellNumber))
colData(final.fibro) <- cur_DF
#final.fibro.other
colData(final.fibro.other) %>% colnames()
final.fibro.other$cell_category <- "Other"
final.fibro.other$cell_type <- "Other"
final.fibro.other$cell_subtype <- "Other"
cur_DF <- colData(final.fibro.other)%>% data.frame() %>% select(-contains(c("som_","rp_","CAF"))) %>% DataFrame()
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.fibro.other$ImageNumber, final.fibro.other$CellNumber))
colData(final.fibro.other) <- cur_DF
#final.fibro.HEV
colData(final.fibro.HEV) %>% colnames()
final.fibro.HEV$cell_category <- "vessel"
final.fibro.HEV$cell_type <- "HEV"
final.fibro.HEV$cell_subtype <- "HEV"
cur_DF <- colData(final.fibro.HEV)%>% data.frame() %>% select(-contains(c("som_","rp_","CAF"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.fibro.HEV$ImageNumber, final.fibro.HEV$CellNumber))
colData(final.fibro.HEV) <- cur_DF
#final.vessel
colData(final.vessel) %>% colnames()
final.vessel$cell_category <- "vessel"
final.vessel$cell_type <- "vessel"
final.vessel$cell_subtype <- "vessel"
cur_DF <- colData(final.vessel)%>% data.frame() %>% select(-contains(c("som_","rp_","vessel","tumour"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.vessel$ImageNumber, final.vessel$CellNumber))
colData(final.vessel) <- cur_DF
#final.vessel.tumour.tumour
colData(final.vessel.tumour) %>% colnames()
final.vessel.tumour$cell_category <- "Tumour"
final.vessel.tumour$cell_type <- "Tumour"
final.vessel.tumour$cell_subtype <- "Tumour"
cur_DF <- colData(final.vessel.tumour)%>% data.frame() %>% select(-contains(c("som_","rp_","vessel","tumour"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.vessel.tumour$ImageNumber, final.vessel.tumour$CellNumber))
colData(final.vessel.tumour) <- cur_DF
#final.stroma.tumour.tumour
colData(final.stroma.tumour) %>% colnames()
final.stroma.tumour$cell_category <- "Tumour"
final.stroma.tumour$cell_type <- "Tumour"
final.stroma.tumour$cell_subtype <- "Tumour"
cur_DF <- colData(final.stroma.tumour)%>% data.frame() %>% select(-contains(c("som_","rp_","vessel","tumour"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.stroma.tumour$ImageNumber, final.stroma.tumour$CellNumber))
colData(final.stroma.tumour) <- cur_DF
#final.immune.tumour.tumour
colData(final.immune.tumour) %>% colnames()
final.immune.tumour$cell_category <- "Tumour"
final.immune.tumour$cell_type <- "Tumour"
final.immune.tumour$cell_subtype <- "Tumour"
cur_DF <- colData(final.immune.tumour)%>% data.frame() %>% select(-contains(c("som_","rp_","vessel","tumour"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.immune.tumour$ImageNumber, final.immune.tumour$CellNumber))
colData(final.immune.tumour) <- cur_DF
#final.immune.nonTcell.tumour
colData(final.immune.nonTcell) %>% colnames()
final.immune.nonTcell$cell_category <- "Immune"
final.immune.nonTcell$cell_type <- final.immune.nonTcell$immune_category
final.immune.nonTcell$cell_subtype <- final.immune.nonTcell$immune_category
cur_DF <- colData(final.immune.nonTcell)%>% data.frame() %>% select(-contains(c("som_","rp_","vessel","tumour","immune"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.immune.nonTcell$ImageNumber, final.immune.nonTcell$CellNumber))
colData(final.immune.nonTcell) <- cur_DF
#final.immune.Tcell
colData(final.immune.Tcell) %>% colnames()
final.immune.Tcell$cell_category <- "T cell"
final.immune.Tcell$cell_type <- final.immune.Tcell$TcellCategory
final.immune.Tcell$cell_subtype <- final.immune.Tcell$TcellType
cur_DF <- colData(final.immune.Tcell)%>% data.frame() %>% select(-contains(c("som_","rp_","immune","tumour","Tcell"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.immune.Tcell$ImageNumber, final.immune.Tcell$CellNumber))
colData(final.immune.Tcell) <- cur_DF
#final.tumour.tumour
colData(final.tumour.tumour) %>% colnames()
final.tumour.tumour$cell_category <- "Tumour"
final.tumour.tumour$cell_type <- "Tumour"
final.tumour.tumour$cell_subtype <- "Tumour"
cur_DF <- colData(final.tumour.tumour)%>% data.frame() %>% select(-contains(c("som_","rp_","immune","tumour","Tcell"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.tumour.tumour$ImageNumber, final.tumour.tumour$CellNumber))
colData(final.tumour.tumour) <- cur_DF
#final.nontumour.tumour
colData(final.nontumour.tumour) %>% colnames()
final.nontumour.tumour$cell_category <- "Tumour"
final.nontumour.tumour$cell_type <- "Tumour"
final.nontumour.tumour$cell_subtype <- "Tumour"
cur_DF <- colData(final.nontumour.tumour)%>% data.frame() %>% select(-contains(c("som_","rp_","immune","tumour","Tcell"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.nontumour.tumour$ImageNumber, final.nontumour.tumour$CellNumber))
colData(final.nontumour.tumour) <- cur_DF
#final.nontumour.undefined
colData(final.nontumour.undefined) %>% colnames()
final.nontumour.undefined$cell_category <- "Other"
final.nontumour.undefined$cell_type <- "Other"
final.nontumour.undefined$cell_subtype <- "Other"
cur_DF <- colData(final.nontumour.undefined)%>% data.frame() %>% select(-contains(c("som_","rp_","immune","tumour","Tcell"))) %>% DataFrame()
colnames(cur_DF)
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(final.nontumour.undefined$ImageNumber, final.nontumour.undefined$CellNumber))
colData(final.nontumour.undefined) <- cur_DF
```
```{r}
final.fibro %>%colData() %>%colnames() %>%length()
assay(final.fibro, "exprs") <-NULL
final.fibro.other %>%colData() %>%colnames()%>%length()
assay(final.fibro.other, "exprs") <-NULL
final.fibro.HEV %>%colData() %>%colnames()%>%length()
assay(final.fibro.HEV, "exprs") <-NULL
final.vessel %>%colData() %>%colnames()%>%length()
assay(final.vessel, "exprs") <-NULL
final.vessel.tumour %>%colData() %>%colnames()%>%length()
assay(final.vessel.tumour, "exprs") <-NULL
final.stroma.tumour %>%colData() %>%colnames()%>%length()
assay(final.stroma.tumour, "exprs") <-NULL
final.immune.tumour %>%colData() %>%colnames()%>%length()
assay(final.immune.tumour, "exprs") <-NULL
final.immune.nonTcell %>%colData() %>%colnames()%>%length()
assay(final.immune.nonTcell, "exprs") <-NULL
final.immune.Tcell %>%colData() %>%colnames()%>%length()
assay(final.immune.Tcell, "exprs") <-NULL
final.tumour.tumour %>%colData() %>%colnames()%>%length()
assay(final.tumour.tumour, "exprs") <-NULL
final.nontumour.undefined %>%colData() %>%colnames()%>%length()
assay(final.nontumour.undefined, "exprs") <-NULL
final.nontumour.tumour %>%colData() %>%colnames()%>%length()
assay(final.nontumour.tumour, "exprs") <-NULL
all.cells <- cbind(final.fibro,final.fibro.other,final.fibro.HEV,final.vessel,final.vessel.tumour,final.stroma.tumour,final.immune.tumour,final.immune.nonTcell,final.immune.Tcell,final.tumour.tumour,final.nontumour.undefined,final.nontumour.tumour)
saveRDS(all.cells,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_cells_combined_types.rds")
all.cells$cell_category[all.cells$cell_type=="other"] <-"Other"
all.cells$cell_type[all.cells$cell_type=="other"] <-"Other"
table(all.cells$cell_category)
rm(final.fibro,final.fibro.other,final.fibro.HEV,final.vessel,final.vessel.tumour,final.stroma.tumour,final.immune.tumour,final.immune.nonTcell,final.immune.Tcell,final.tumour.tumour,final.nontumour.undefined,final.nontumour.tumour)
```
#add clinical data
```{r}
wd <-dirname(getwd())
clinical.data <- read.csv(file=file.path(wd,"sce_objects","clinical_data", "clinical_data_ROI_ac_combined_CORRECT.csv"))
clinical.data$TMA <-clinical.data$TMA.x
clinical.data$TMA.x <-NULL
clinical.data$TMA.y <-NULL
clinical.data$X.1 <-NULL
clinical.data$X <-NULL
head(clinical.data)
unique(clinical.data$DX.name)
clinical.data$DX.name[clinical.data$Patient_ID=="Control"] <-"Control"
table(clinical.data$DX.name)
clinical.data$Patient_ID %>% unique() %>% length() #1071 PATIENTS IN TOTAL
area <- read.csv(file=file.path(wd,"clinical_data", "area.csv"))
area$X <- NULL
area$Tma_ac <- area$TMA_ImageID
area$TMA_ImageID <-NULL
area_clinical <- left_join(clinical.data, area, by="Tma_ac")
length(unique(area$Tma_ac))
length(unique(clinical.data$Tma_ac))
length(unique(area_clinical$Tma_ac))
length(unique(area_clinical$Patient_ID))
length(unique(clinical.data$Patient_ID))
head(area_clinical)
colnames(area_clinical)[colnames(area_clinical) %in% colnames(colData(all.cells))]
area_clinical$Patient_ID[!area_clinical$Patient_ID %in% unique(all.cells$Patient_ID)]
unique(all.cells$Patient_ID)[!unique(all.cells$Patient_ID) %in% area_clinical$Patient_ID]
all.cells[, is.na(all.cells$Patient_ID)]$Tma_ac %>% unique()
saveRDS(all.cells,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_cells_combined_types_CLINICAL-DATA.rds")
all.cells.filtered <- all.cells[, all.cells$Tma_ac!="88A_1"&all.cells$Tma_ac!="88A_2"&all.cells$Tma_ac!="88A_3"&all.cells$Tma_ac!="88A_4"&all.cells$Tma_ac!="86B_24"]
all.filtered <-all.cells[, all.cells$Tma_ac!="88A_1"&all.cells$Tma_ac!="88A_2"&all.cells$Tma_ac!="88A_3"&all.cells$Tma_ac!="88A_4"&all.cells$Tma_ac!="86B_24"]
all.cells[, all.cells$Tma_ac=="88A_1"|
all.cells$Tma_ac=="88A_2"|
all.cells$Tma_ac=="88A_3"|
all.cells$Tma_ac=="88A_4"|
all.cells$Tma_ac=="86B_24"]
saveRDS(all.filtered,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/all_cells_combined_types_CLINICAL-DATA_FILTERED.rds")
```
```{r, add clustering results to subset tumour}
cur_DF <- as_tibble(colData(all.cells)) %>% left_join(area_clinical, by = c("Tma_ac","TMA","acID")) %>% DataFrame()
all.equal(paste(cur_DF$ImageNumber, cur_DF$CellNumber), paste(all.cells$ImageNumber, all.cells$CellNumber))
colData(all.cells) <- cur_DF
rownames(colData(all.cells)) <-all.cells$CellID
unique(all.cells$DX.name)
head(colData(all.cells))
all.cells$Patient_ID %>% unique() %>% length()
length(unique(all.filtered[,all.filtered$Patient_ID!="Control" ]$Patient_ID))
#1056 patients in total!
length(unique(clinical.data$Patient_ID))
length(unique(all.cells$Patient_ID))
length(unique(all.filtered$Patient_ID))
length(unique(clinical.data$Patient_ID))
tcell.sce$DX.name[is.na(tcell.sce$DX.name)]<-"NA"
table(all.filtered$cell_category)
```
```{r}
all.fibro <- all.filtered[, all.filtered$cell_category=="Fibroblast"]
all.vessel <- all.filtered[, all.filtered$cell_category=="vessel"]
all.tumour <- all.filtered[, all.filtered$cell_category=="Tumour"]
all.immune <- all.filtered[, all.filtered$cell_category=="Immune"|all.filtered$cell_category=="T cell"]
all.tcell<- all.filtered[, all.filtered$cell_category=="T cell"]
all.immune.nonT <- all.filtered[, all.filtered$cell_category=="Immune"]
all.other <- all.filtered[, all.filtered$cell_category=="Other"]
unique(all.filtered$cell_category)
table(all.filtered$cell_category)
saveRDS(all.fibro,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/FIBRO_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.vessel,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/VESSEL_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.tumour,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/TUMOUR_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.other,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/OTHER_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.immune,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/IMMUNE_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.immune.nonT,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/IMMUNE_nonT_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.tcell,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/TCELL-only_CLINICAL-DATA_FILTERED.rds")
saveRDS(all.filtered,"~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/all_cells_combined_types_CLINICAL-DATA_FILTERED.rds")
all.filtered <-readRDS("~/data/lena_processed2/NSCLC_NEW/sce_objects/final objects with categories/FINAL/all_cells_combined_types_CLINICAL-DATA_FILTERED.rds")
length(unique(all.filtered$Patient_ID))
```
```{r}
df <- colData(all.filtered)%>% data.frame() %>% select(c(CellID, cell_category, Compartment))
df$Mask <- ifelse(df$Compartment>0, "Tumour","Stroma")
i <-"Tumour"
table(df$cell_category, df$Mask)
```