Figure 1 from Epigenetic Heritability of Cell Plasticity Drives Cancer Drug Resistance through a One-to-Many Genotype-to-Phenotype Paradigm

Experimental design of long-term drug resistance evolution in colorectal cancer organoids. A, Workflow of lentiviral barcoding in colorectal cancer (CRC) organoid single cells as an evolutionary tracking tool. MOI, multiplicity of infection; RFP, red fluorescent protein. B, Experimental design of long-term drug resistance evolution in an MSS AKT–mutant organoid. Bulk DNA profiling was performed for genomic characterization and barcode measurement, as well as scRNA-seq and corresponding single-cell barcode extraction of five “solid” time points over 5 months: parental, under drug 1, regrowth after drug 1, under drug 2, and regrowth after drug 2. Additionally, floating DNA was collected every 2 days from the supernatant to profile barcodes as a “liquid biopsy.” C, Cells were exposed to four different sequences of drugs with first- and second-line treatments. D, In a second experiment, both organoid lines (MSS and MSI) were exposed to an ERK inhibitor and oxaliplatin. Before drug pressure, CIN was induced with CENP-E inhibitors alone or in combination with the MPS1 inhibitor to assess CIN effects on drug resistance. AKTi, AKT inhibitor; ERKi, ERK inhibitor; MEKi, MEK inhibitor. Created in BioRender. Sottoriva, A. (2025) https://BioRender.com/7toyszr.