Fig. 3.

Tumor cell-intrinsic factors underlying treatment resistance to tipifarnib. a tSNE plot of total human tumor cells (dots) from the PDX model of BC159-T#3, color-coded by sample origin, circled by cluster (left panel). Comparison of HRAS mRNA expression level in tumor cells treated with tipifarnib or vehicle as control. Dots indicate individual cells, P = 0.0091 (right panel). b Comparison of RAS and MAPK pathway activity in tumor cells treated by tipifarnib and vehicle as control. P = 0.00011, P = 0.0066, respectively. Each box shows the median and IQR (interquartile range, 25th to 75th percentiles), whiskers indicate the highest and lowest value within 1.5 times the IQR, and outliers are marked as dots. c Single-cell western blots of HRAS protein and its downstream protein markers (p-AKT and p-ERK) in tumor cells treated with tipifarnib or vehicle. Dots indicate individual cells, and diamond and star shapes indicate means and median of peak area, respectively. d Prediction of cell cycle state in tumor cells from PDX at a single-cell level (dots) using G1/S and G2/M module score (left panel). Cells are colored by their assigned cell cycle state (cycling; orange, intermediate; yellow, non-cycling; gray, middle panel). The relative cellular composition for tumor cells treated with tipifarnib or vehicle. Chi-squared test, P = 0.021, *P < 0.05 (right panel). e Violin plots of significantly upregulated genes, IGFBP7, MDK, and B2M, in tumor cells treated by tipifarnib or vehicle (dots). P = 1.3e−07, P = 1.2e−05, P = 6.4e−06, respectively. f Validation of comparative upregulation of IGFBP7, MDK, and B2M in tumor cells treated with tipifarnib compared to those with vehicle by immunohistochemistry staining performed on PDX. Scale bar, 100 μm. g tSNE plot of total tumor cells from BC159-T#3 patient tumor, color-coded by cluster (left panel). Violin plot of HRAS mRNA expression level against distinct tumor cell clusters of BC159-T#3. Student’s t test, ****P < 0.0001, *P < 0.05. Abbreviation: BT, basal tumor (right panel). h Prediction of cell cycle state in tumor cells from BC159-T#3 at a single-cell level (dots) using G1/S and G2/M module score (left panel). Cells are colored by cell cycle as in d (middle panel). The relative cellular composition among basal tumor clusters. Chi-squared test, P < 2.2e−16, ****P < 0.0001 (right panel). i Violin plots of mRNA expression levels of IGFBP7, MDK, and B2M against distinct tumor cell clusters of BC159-T#3 (dots). ****P < 0.0001, *P < 0.05. j Comparison of IGFBP7, MDK, and B2M protein expression between fast (upregulated Ki-67) and slow (downregulated Ki-67) growing regions in BC159-T#3 by immunohistochemistry staining. Scale bar, 100 μm