Impact of CDK4/6 and MEK inhibition on the tumour microenvironment. (A)
The KPC-derived 4662 cell line labelled with H2B-GFP was treated with
palbociclib, trametinib or the combination. Cellular proliferation was
determined using IncuCyte live cell imaging over the indicated time frame
(***p<0.001, by Student t-test relative to the single-agent controls).
(B) The 4662 model was introduced into C57BL/6J mice subcutaneously, mice were
randomised at approximately 200 mm3 and tumour volume was measured by
callipers with mice on treatment (***p<0.001 for the combination relative
to vehicle control by Student t-test). (C) Schematic workflow for the
single-cell sequencing. Feature plots denote the expression of PTPRC (CD45 gene)
in all of the captured cells with treatment. (D) All of the cells from the
treated mice were filtered for quality and clustered using Seurat software in
the dimension plot. The general clustering of different immunological subtypes
is indicated. (E) The violin plots denote gene markers associated with myeloid,
neutrophil and macrophage populations. (F) Seurat heatmap identifying top
differences between the myeloid clusters (0, 2 and and representative feature
plots are shown). (G) Distribution of cells in the indicated myeloid clusters,
changes in clusters 0 and 2 with treatments are highly significant by
Ļ2statistic (***p<1Eā100). (H) Genes
differentially expressed in cells between clusters 2 and 0 are highlighted in
the violin plot (all genes are highly significant between the clusters
p<1Eā5).