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Code for the analyses of the paper "High grade serous ovarian cancer organoids as models of chromosomal instability" (Vias and Morrill et al, eLife 2023)

Summary of paper

There are four main figures. All the figures are created using the scripts inside the folder figures.

Summary of folders and files

I only include the files which were finally used for the paper

  • copy_number_analysis_organoids Analysis of copy number exposures, and of segments
    • copy_number_analysis_organoids.Rmd: R markdown file
  • RNASeq_and_CN Comparison of copy number and gene expression - 20191218_ViasM_BJ_orgaBrs - Scripts - analyse_joint_counts_CN.R
  • RNASeq_DE_resistant_sensitive Analysis of DE for sensitive vs resistant in TCGA samples
    • analysis_scripts
      • 3_analysis_organoids_PCA_subset_samples.R: PCA for RNA-Seq data
      • DGE_Genepathways_RnaSeqPipAucWo3PBias_MVorganoids_CMS20210518.Rmd: pathway enrichment analysis
  • scDNAseq-Organoids Scripts to create single cell DNA plots
    • code
      • plotting_nosexchrom.R: plotting scDNA data without sex chromosomes
      • subclonal_structure.R: clustering of scDNA profiles
  • survival_analysis Scripts to create plots of the survival of organoids, and miscelaneous plots
    • code
      • organoid_survival2.rmd R markdown
      • response_drugs.R
      • Survival.Rmd
  • figures folder to create the final version of the figures; explained above
    • fig1.R
    • fig2.R
    • fig3.R
    • fig4.R

Order in which to run the files

I would run the files in the order they appear above.

Other folders for related analyses, but not part of the paper

  • not_used additional related analyses that haven't made it to the paper
    • cell_lines Analysis of cell lines (deprecated)

Input files

  • copy_number_analysis_organoids
    • copy_number_analysis_organoids.Rmd
      • data/organoid_exposures.rds
      • data/NewOrganoidNaming.csv
      • data/sig_data_unorm.RDS
      • data/Export-matrix_OV_Sigs_on_TCGA-OV_12112019.rds
      • data/summary.ascatTCGA.penalty70.txt
      • data/pcawg_CN_features.rds
      • data/tcga_CN_features.rds
      • ../../cnsignatures/manuscript_Rmarkdown/data/BriTROC_absolute_copynumber.rds
      • data/6_TCGA_Signatures_on_BRITROC/0_BRITROC_absolute_CN.rds
      • data/organoid_absolute_CN.rds
      • data/BriTROC_CN_features.rds
      • data/CN_Calls_ABSOLUTE_PCAWG/OV-AU.segments.raw.rds
      • data/CN_Calls_ABSOLUTE_PCAWG/OV-US.segments.raw.rds
      • ../RNASeq_DE_resistant_sensitive/files/20191218_ViasM_BJ_orgaBrs_tpm.csv
      • ../RNASeq_DE_resistant_sensitive/files/PDOnameProperSample_sWGS_RNAseq.xlsx
      • data/asorg_PDO.csv signature exposures for ascites and organoids
      • data/ascites_exposures_20210125.rds
  • RNASeq_DE_resistant_sensitive
    • analysis_scripts
      • 3_analysis_organoids_PCA_subset_samples.R
      • DGE_Genepathways_RnaSeqPipAucWo3PBias_MVorganoids_CMS20210518.Rmd
  • RNASeq_and_CN
    • 20191218_ViasM_BJ_orgaBrs
      • Scripts
        • analyse_joint_counts_CN.R
  • scDNAseq-Organoids
    • code
      • plotting_nosexchrom.R
      • subclonal_structure.R
  • survival_analysis
    • code
      • organoid_survival2.rmd
      • response_drugs.R
      • Survival.Rmd
  • figures
    • fig1.R
      • ../copy_number_analysis_organoids/robjects/exposures.RDS
      • ../copy_number_analysis_organoids/robjects/dendrograminputclr.RDS
      • ../copy_number_analysis_organoids/robjects/heatmapinputclr_with_ticks.RDS
      • ../copy_number_analysis_organoids/robjects/heatmapinputclr.RDS
      • ../copy_number_analysis_organoids/robjects/rank_nsegments.RDS
      • ../copy_number_analysis_organoids/robjects/rank_ploidy.RDS
      • ../copy_number_analysis_organoids/robjects/rank_nsegments_df.RDS
      • ../copy_number_analysis_organoids/robjects/rank_ploidy_df.RDS
      • ../survival_analysis/robjects/km_as_one.RDS
      • ../survival_analysis/data/OrganoidSurvival.csv
      • ../survival_analysis/data/OrganoidCulturesSurvival_091117OR.xlsx
      • ../survival_analysis/data/OrganoidCulturesSurvival_091117OR_17082021.xlsx
    • fig2.R
      • ../scDNAseq-Organoids/robjects/fig2_absCN.RDS
      • ../scDNAseq-Organoids/robjects/fig2_subclonal_hclust_nosexchromPDO2.RDS
      • ../scDNAseq-Organoids/robjects/fig2_subclonal_hclust_nosexchromPDO3.RDS
      • ../scDNAseq-Organoids/robjects/fig2_subclonal_hclust_nosexchromPDO6.RDS
      • ../scDNAseq-Organoids/robjects/fig2_subclonal_hclust_nosexchromPDO2_2.RDS
      • ../scDNAseq-Organoids/robjects/fig2_subclonal_hclust_nosexchromPDO3_2.RDS
      • ../scDNAseq-Organoids/robjects/fig2_subclonal_hclust_nosexchromPDO6_2.RDS
      • ../scDNAseq-Organoids/robjects/fig2_colours.RDS
      • ../copy_number_analysis_organoids/data/absolute_profiles/PDO2_annotated_2.pdf
      • ../copy_number_analysis_organoids/data/absolute_profiles/PDO3_annotated_2.pdf
      • ../copy_number_analysis_organoids/data/absolute_profiles/PDO6_annotated_2.pdf
      • fig2_chrom_annotation_crop_nosexchrom.png
    • fig3.R
      • ../RNASeq_and_CN/20191218_ViasM_BJ_orgaBrs/output/output_GRCh37/fig3_df_gene_characteristics.RDS
      • ../RNASeq_and_CN/20191218_ViasM_BJ_orgaBrs/output/output_GRCh37/fig3_df_average_bottomCN.RDS
      • ../RNASeq_and_CN/20191218_ViasM_BJ_orgaBrs/output/output_GRCh37/joint_counts_CN_subset.RDS
      • ../RNASeq_DE_resistant_sensitive/objects/fig4_pca_with_gsva_annotation_NC.RDS
      • ../RNASeq_DE_resistant_sensitive/objects/fig4_pca_with_gsva_annotation_NC_prcomp.RDS
      • ../RNASeq_DE_resistant_sensitive/objects/fig4_df_colmeans_deseqcounts_correlation_tcga_org.RDS
      • ../RNASeq_DE_resistant_sensitive/objects/fig3_ssgsea_repair.RDS
    • fig4.R
      • ../RNASeq_DE_resistant_sensitive/files/PDOnameProperSample_sWGS_RNAseq.xlsx
      • ../RNASeq_and_CN/20191218_ViasM_BJ_orgaBrs/Input/samples.csv
      • ../copy_number_analysis_organoids/data/Book1_ascites.xlsx
      • ../copy_number_analysis_organoids/robjects/fig4_ascites.RDS
      • ../survival_analysis/robjects/AUC_all_df.RDS
      • ../survival_analysis/robjects/fig4_PDS_PDO.RDS
      • ../copy_number_analysis_organoids/robjects/exposures.RDS
      • ../RNASeq_DE_resistant_sensitive/objects/fig4_fgseaResTidy.RDS

Dependencies

Specific packages

  • You have to download the bitbucket repo CNsignatures and save it in the same folder where this repo is, i.e.
	                 |--- this folder
	Mother folder ---| 
	                 |--- CNsignatures
  • CompSign package
  • rnaseqRpkg: internal CRUK RNA-Seq pipeline package. Send me (lm687 at cam.ac.uk) an email about it

General R packages

library(AnnotationHub) ## Bioconductor
library(Biobase) ## Bioconductor
library(biomaRt) ## Bioconductor
library(CNTools) ## Bioconductor
library(CompSign) ## github
library(DESeq2) ## Bioconductor
library(EnvStats) ## CRAN
library(GSVA) ## Bioconductor
library(GSVAdata) ## Bioconductor
library(GenomicRanges) ## Bioconductor
library(MASS) ## CRAN
library(QDNAseq) ## Bioconductor
library(RColorBrewer) ## CRAN
library(ReactomePA) ## Bioconductor
library(biomaRt) ## Bioconductor
library(compositions) ## CRAN
library(cowplot) ## CRAN
library(dendextend) ## CRAN
library(dplyr) ## CRAN
library(fgsea) ## Bioconductor
library(ggdendro) ## CRAN
library(ggh4x) ## CRAN
library(ggplot2) ## CRAN
library(ggplotify) ## CRAN
library(ggrepel) ## CRAN
library(ggthemr) ## github
library(grid) ## CRAN
library(gridExtra) ## CRAN
library(jcolors) ## CRAN
library(latex2exp) ## CRAN
library(lsa) ## CRAN
library(parallel) ## CRAN
library(pheatmap) ## CRAN
library(readxl) ## CRAN
library(reshape2) ## CRAN
library(tidyverse) ## CRAN
library(viridis) ## CRAN

Scripts that create each figure

Main figures

Main figures are assembled in the figures/ folder - Figure 1: figures/fig1.R -

Supplementary figures

  • Supplementary Figure 1 Sample collection workflow and survival analysis. survival_analysis/code/organoid_survival2.rmd
  • Supplementary Figure 2 Clinical data. Carolin's script
  • Supplementary Figure 3 Orthotopic implantation of PDOs. No code.
  • Supplementary Figure 4 PDOs and patient samples mutation analysis. No code.
  • Supplementary Figure 5 Tissue and PDOs morphological structures and p53 status. No code.
  • Supplementary Figure 6 compare_to_shWGS.R
  • Supplementary Figure 7 PDOs genome-wide absolute copy number alteration analysis figures/supplementary_figures/suppl7.R
  • Supplementary Figure 8 Genomic features comparison between tumours and PDOs copy_number_analysis_organoids/copy_number_analysis_organoids.Rmd
  • Supplementary Figure 9 Absolute gene copy number in PDOs RNASeq_and_CN/20191218_ViasM_BJ_orgaBrs/scripts/analyse_joint_counts/.R
  • Supplementary Figure 10 Whole genome correlation between absolute gene copy number and expression RNASeq_and_CN/20191218_ViasM_BJ_orgaBrs/scripts/analyse_joint_counts/.R
  • Supplementary Figure 11 Drug sensitivity curves survival_analysis/code/response_drugs.R
  • Supplementary Figure 12 Subclonal heterogeneity ribbon plot subclonal_structure.R
  • Supplementary Figure 13 Single cell clade analyses of CN figures/supplementary_figures/supp_clades_scDNA.R, from data created in subclonal_structure.R

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