{"database":"biostudies-arrayexpress","file_versions":[],"scores":null,"additional":{"submitter":["Maria Puschhof"],"organism":["Homo sapiens"],"full_dataset_link":["https://www.ebi.ac.uk/biostudies/studies/E-MTAB-16836"],"description":["Two human colorectal cancer organoids derived from liver metastasis of the same patient were grown for seven days, then treated with FOLFIRI chemotherapy in vitro and sampled at different timepoints for single-cell RNA-sequencing. Time points include 0 h, 12 h, 24 h, 48 h, 72 h and 120 h on-treatment as well as 12 h, 24 h, 48 h and 72 h washout after a 72 h treatment period. Samples were multiplexed using hashtag oligos prior to sequencing."],"repository":["biostudies-arrayexpress"],"sample_protocol":["Sample Collection - Organoids were harvested and washed twice with cold PBS (1500 rpm, 4 °C, 5 min). Dissociation of the organoids was achieved resuspending the pellet in TrypLE (Thermo Fisher Scientific, catalog #12605036) and incubated at 37 °C in water bath until a homogeneous cell suspension was obtained, pipetting the mix occasionally to mechanically accelerate the dissociation process. To stop the reaction, PBS + 2% FCS was added to the solution and spin down (1 500 rpm, 4 °C, 5 min). Eventually, cells were cryopreserved until all timepoints were collected for further processing.  For sample multiplexing, frozen samples were rapidly thawed in water bath 37 °C and transferred to a conical tube with added fresh organoid medium + 2% FBS and spin down. Cell pellet was then resuspended in 100 µL of PBS + 2% FCS with 1:50 FcR Blocking Reagent (Miltenyi Biotec, catalog #130-059-901) and stained with 1µL of hashing antibodies (Total-Seq-B, BioLegend, 0.5 µg antibody) for 30 minutes at 4 °C. Afterwards, cells were washed twice with PBS + 2% FCS. Cell pellet was then resuspended in PBS + 2% FCS added with a live/dead marker (DAPI, Invitrogen catalog # 62248 or Zombie NIR, BioLegend, catalog # 423106), filtered, and multiplexed samples were sorted for live cells in a single tube.","Library Construction - After cell lysis inside of GEMs, mRNA is reverse transcribed to cDNA with droplet-specific barcodes. After GEM breakdown, the library is being prepared according to the protocol including cDNA amplification by PCR.","Sample Treatment - Organoids were seeded as single cells at a density of 20 000 cells per 20 µL dome, counting for a total of 400 000 cells plated per timepoint. After seven days that spheres were generated, FOLFIRI treatment was started at a concentration representative of the IC50 and timepoints were collected at 0, 12, 24, 48, and 72 h treatment. After 72h, FOLFIRI was removed and changed with either fresh medium to let the organoids recover (“Release timeline”), or FOLFIRI was reapplied (“Continuous timeline”). Timepoints in the “Release timeline” were collected after 12, 24, 48 and 72 h, while the “Continuous timeline” included a timepoint collected after 48 h.","Nucleic Acid Extraction - A maximum of 40 000 cells from each pooled sample was loaded onto a well of a 10X Chromium Next GEM Chip G and scRNA-sequencing libraries were generated following the Chromium Next GEM Single Cell 3’ Reagent Kit v3.1 (Dual Index protocol) according to the manufacturer’s instructions. Each reaction generated a library for gene expression (GEX) and a library for hashtag oligos (HTO).","Sequencing - Next generation sequencing was performed at the German Cancer Research Center using an Illumina NovaSeq 6000 S4 instrument (paired-end, one lane per library)."],"figure_sub":["Organization","MINSEQE Score","Assays and Data","Processed Data","MAGE-TAB Files"],"data_protocol":["Data Transformation - Quality control and filtering was performed at library level using Seurat in R. Cells were filtered for having at least 200 genes, while genes were filtered for being expressed in at least 3 cells prior to demultiplexing. Then, high quality cells were extracted with a gene count higher than 2,000, total count higher than 20,000 and mitochondrial fraction  less than 20%. For downstream analysis, cell cycle-related effects were regressed out using the option vars.to.regress in Seurat’s ScaleData. Trajectory inference was performed using Palantir, provided as h5ad file \\\"Org_chemo_LM3_palantir.h5ad\\\".","Sequence Alignment - Reads were mapped to the human reference genome assembly refdata-gex-GRCh38-2020-A using the cellranger count pipeline of the 10X Genomics Cell Ranger (v6.1.0) with default settings, while providing the hashtag oligo (HTO) sequences as feature library. Demultiplexing was subsequently performed in R using the filtered_feature_bc_matrix as input. HTO counts were normalized using CLR normalization. Using Seurat’s HTODemux function with default settings, cells were assigned to samples using the \\\"HTO_maxID\\\" assignment. Only singlet cells were kept for downstream analysis. After quality control and filtering, counts were normalized and cell cycle effects were regressed out (provided as \\\"*counts_log1p_woCC.txt\\\")."],"omics_type":["Metabolomics","Unknown","Transcriptomics","Genomics","Proteomics"],"instrument_platform":["Illumina NovaSeq 6000"],"study_type":["RNA-seq of coding RNA from single cells"],"species":["Homo sapiens"],"pubmed_title":["TROP2 targeting reveals therapy-driven cell state dynamics and vulnerabilities in colorectal cancer"],"pubmed_authors":["Nuria Vaquero-Siguero, Nikolaos Georgakopoulos, Maria C Puschhof, Ioannis Chiotakakos, Jasmin Meier, Sigrid K Fey, Gabriele Diamante, Manuel Mastel, Aitana Guiseris Martinez, Guillaume Belthier, Nikolai Schleußner, Julia Volk, Carolin Artmann, Bryce Lim, Ronald Koschny, Cyrill Wehling, Kyanna S Ouyang, Michael Günther, Solveig Kuss, Paula Hoffmeister, Moritz Mall, Jens Neumann, Steffen Ormanns, Martin Schneider, Thomas Schmidt, Jens Puschhof, Andreas Trumpp, Jacco van Rheenen, Julio Saez-Rodriguez, Bruno C Köhler, Rene Jackstadt","Maria Puschhof"],"additional_accession":[]},"is_claimable":false,"name":"Timecourse analysis of human colorectal cancer organoids under FOLFIRI chemotherapy","description":"Two human colorectal cancer organoids derived from liver metastasis of the same patient were grown for seven days, then treated with FOLFIRI chemotherapy in vitro and sampled at different timepoints for single-cell RNA-sequencing. Time points include 0 h, 12 h, 24 h, 48 h, 72 h and 120 h on-treatment as well as 12 h, 24 h, 48 h and 72 h washout after a 72 h treatment period. Samples were multiplexed using hashtag oligos prior to sequencing.","dates":{"release":"2026-04-01T00:00:00Z","modification":"2026-05-18T12:43:14.039Z","creation":"2026-03-31T11:14:20.506Z"},"accession":"E-MTAB-16836","cross_references":{"ENA":["ERP191621"],"EFO":["EFO_0002944","EFO_0004170","EFO_0005684","EFO_0004917","EFO_0005518","EFO_0003816","EFO_0004184","EFO_0003969"]}}