Figure 1. Primary patient derived cell lines exhibit differential sensitivity to CDK4/6 inhibition:

(A) The genetics of the employed cell models are summarized in the oncoprint (somatic single nucleotide variant: green, homozygous deletion: blue, small insertion/deletion: orange, amplification: red). The percent aberrations of the same genes as reported in the TCGA PDAC cohort is provided in the table. (B) The cell lines were subjected to RNA sequencing and the log2-normalized transcript levels for the indicated genes are shown. (C) The indicated cell lines were treated with increasing dose of palbociclib (PD) (0, 100, 250, 1000 nM). Forty-eight hours post-treatment cells were labeled with BrdU and the relative incorporation was determined. All experiments were performed in triplicate with multiple wells measured. Bars indicate the means and standard-deviation. (D) The indicated pancreatic cancer cell lines expressing H2B-GFP were treated with 100 nM palbociclib and proliferation was determined by live cell imaging. (E) Immunoblot analysis of the indicated proteins from 519, 827, 3226 and 1222 cell lines that were treated with palbociclib (PD) for 48 hours. (F) Cyclin D1 and cyclin E1 levels were determined at the indicated time points following CHX release in 1222 and 3226 cell lines. Cells were pre-treated with palbociclib (200 nM). Protein expression was analyzed by western blotting, with actin level used as loading control and the band intensities were quantified. The mean and SD are shown (*p< 0.5, **p < 0.01, ***p < 0.001 as determined by t test). (G) Immunofluorescence staining of cyclin D1 in 519 cell line following 48 hours exposure with palbociclib (scale bar 50 μm). (H) Immunoblot and BrdU analysis from the indicated cell lines that were transfected with KRAS (red bars) or non-target (black bars) RNAi in the presence and absence of palbociclib (PD). The mean and SD are shown (*p<0.05, **p<0.01, ***p < 0.001 as determined by t test).