Fig 1: AREG regulates the activity of hEMT program in HNSCC.(A) Western blotting of indicated antibodies upon either knockdown of AREG with siRNAs or addition of recombinant AREG protein in SNU-1066 cells. (B-C) hockey stick plots showing the correlation of the mRNA expression of AREG with other genes, rank ordered by correlation coefficient. hEMT program genes are highlighted in red. Only genes with nonnegative correlation are shown. (B) is the TCGA dataset and (C) is CCLE+HPA combined dataset. (D-F) heatmaps showing changes in expression of hEMT program genes upon either AREG knockdown, or addition of recombinant AREG protein, in HNSCC cell lines or PDO samples. (G) A schematic graph of the experiment design, created using Biorender. (H) Growth curves and (I) images of xenograft tumors. (J) Representative IHC images of p-ERK and p-AKT on each group. Scale Bar; 100 μm
Fig 2: AREG, a top hEMT program gene, regulates PDO responsiveness to cisplatin.(A) A scatter plot showing the Pearson correlation R value (y) and p value (x) of each hEMT gene with IC50 values of cisplatin. Top 5 genes are highlighted. (B) Pearson correlation between AREG expression and IC50 values of cisplatin. Each dot is one PDO sample. (C) A column plot showing the expression fold change of seven EGFR ligands between HNSCC tumors with normal samples using TCGA bulk RNA-seq data. *, P < 0.05. (D) A Kaplan meier survival plot of TCGA HNSCC patients (n=520), grouped according to the mRNA expression of AREG. (E) multivariate survival analysis of indicated factors using the TCGA HNSCC cohort. (F) A box plot showing the protein levels of AREG across various HPA human tumor samples. (G) Box plots showing the mRNA expression of AREG in indicated HNSCC tumor samples from the dataset GSE9349. (H) dose-response curves of scramble control and AREG-knockdown samples to cisplatin treatment in HNSCC cell lines (A253 and SNU-1066) or (I) PDOs (PDO-42 and PDO-49). (J) Images of PDOs upon cisplatin treatment. (K) dose-response curves and (L) images of PDO-51 to cisplatin treatment in the presence and absence of recombinant AREG protein. Scale bar; 40 μm
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