Fig 2: SCLC ex vivo tumors have distinct phenotypic states regulated by the epigenome.(A) Whole transcriptome associations in matched CCX1 primary tumor, PDX, and ex vivo cells. (B) Scatter star plot of gene expression [transcript per million (TPM)] for program defining genes in five matched PDX and ex vivo cells: CCX1, CCX280, CBX41, CBX163, and CBX140. Hubs = means; spokes = individual sample values. (C) Phase contrast images (×20) of CCX1 ex vivo cells. Arrows mark representative populations of cells in each stratum. (D) Strata profiled by immunoblot. (E) Flow cytometry data show CD44- and EpCAM-expressing cells in each stratum. APC, allophycocyanin; PE, phycoerythrin. (F) Invasion assay of strata or cells sorted via fluorescence-activated cell sorting (FACS) for CD44 or EpCAM. Values are an average of the number of stained cells from four high-power fields. Error bars represent SEM of at least three different experiments. The student t test with Welch’s correction was used for statistical analysis. *P < 0.05. (G) CCX1 ex vivo cells (5 × 106) were treated with chemotherapy (EP; etoposide (1.5 µg/ml) and cisplatin (0.5 µg/ml)] for 5 days, followed by drug washout or irradiated to a total dose of 10 Gy (2 Gy per fraction daily). CD44- and EpCAM-expressing cells were quantified by flow cytometry at the indicated times, and the change in the percentage of CD44 and EpCAM (%?) calculated relative to t = 0. (H and I) Cells with differential expression of EpCAM and CD44 expression were FACS-sorted and tested for clonogenicity in vitro or in vivo (limiting dilution assay). Tumor growth was followed for ~8 weeks in NOD.Cg-Prkdcscid Il2rgtm1Wjl/SzJ (NSG) mice. *Nonoverlapping CIs of the proportions as calculated by the Clopper and Pearson exact test. (J) Track plots of ATAC-seq signals at the ASCL1, NEUROD1, and YAP1 loci in each stratum of CCX1. Red arrowheads and yellow shading mark accessibility and the designated promoters, respectively.