Fig 2: Pharmacodynamic effects of adavosertib in combination with T-DXd following 10-day treatment. Mice were treated for 10 days with adavosertib, T-DXd, or T-DXd + adavosertib, or were untreated (N = 5). A, PD marker assessment by IHC for adavosertib, T-DXd, and adavosertib + T-DXd tumors assessing phosphorylated Chk1, phosphorylated Kap1, γH2AX, Ki67, and cleaved caspase 3 (CC3). B, Differential expression of proteins assessed by RPPA revealed alterations in expression of several proteins involved in G2–M transition, including Aurora-A, cyclin B, PLK1, CDC25, and phosphorylated CDK1. C, Morphologic features on H&E staining observed following 10-day treatments with adavosertib, T-DXd, or T-DXd + adavosertib. CCNE1, Cyclin E; ERBB2, Erb-B2 receptor tyrosine kinase 2; H&E, hematoxylin and eosin; PD, pharmacodynamic.

Fig 3: Identification of lncRNA RAET1K function and biological mechanism. (A) The Kaplan‐Meier curve of the risk score for the overall survival of RAET1K in lung adenocarcinoma (LUAD). The blue line presents the lower expression level group of RAET1K, and the red presents the higher ones. Gene enrichment plots showed gene set enrichment analysis (GSEA) between high- and low-expressed RAET1K. (B) The nomogram of clinical features and RAET1K expression level for predicting the 1- and 3-year survival with risk score. (C) calibration plot indicated this nomogram model had a predictive power for overall survival. (D) The upper enrichment plots contain value of the genes' enrichment scores and the corresponding barcode plot shows the genes position. In the bottom heatmap red represents Spearman correlations with higher expression level of RAET1K, blue represents Spearman correlations with lower expression level of RAET1K. Expression of RAET1K and CCNE1 expression level were positive related with each other (E), while RAET1K (F) and CCNE1 (G) were negatively correlated with miR-135a. (H) RAET1K and CCNE1 expression were upregulated in both early and advance stage of LUAD, while miR-135a was downregulated, ***P < 0.001. (I) Construction of ceRNA network of lncRNA-miRNA-mRNA in blue module. The green node in diamond was lncRNA RAET1K, the blue circle nodes were mRNAs, and the pink circle nodes were miRNA. The line between nodes present their relation and the red lines shown RAET1K targeted miR-135a-5p and CCNE1.

Fig 4: Elevated c-MYC expression prevents T/CQ-mediated cell cycle arrest. (A) Representative cell cycle histograms with percentages of cells in G0/G1, S, and G2/M phases are shown for Panc 10.05, HPAF-II, and PDX220 cells expressing Tet-On c-MYC. Cells were treated for 7 d with DMSO (Ctrl), 2 μg/ml Dox, trametinib (T, at 5–100 nM) and CQ (at 5–20 μM; T/CQ), or D/T/CQ. (B) Average percentage of cells in S phase for Tet-On c-MYC–expressing Panc 10.05, HPAF-II, and PDX220 cells that were treated as above (n = 3 from two independent experiments for each cell type, mean ± SEM). *, P < 0.05; **, P < 0.01; Student’s two-sided t test. (C) Immunoblot analysis of lysates generated from Tet-On c-MYC–expressing cells that were treated as above (n = 1 for each cell type). Lysates were blotted for c-MYC, pERK1/2 (phospho-T202/Y204), ERK1/2, pRB (phospho-S807/S811), RB, E2F1, CCNB1, CCNA2, CCND2, CCNE1, CDK2, CDK4, CDK6, p27KIP1, p21CIP1, and ACTB. Densitometry quantitation of c-MYC and pRB protein levels normalized to actin are shown. Source data are available for this figure: SourceData F3.

Fig 5: CUL4A knockdown rescues elevated proliferation of livers of FTOL-KOmice. (A) Western blot analyses of CCNE1 in whole liver lysates isolated from ST-DEN injected Ctrl and FTOL−KO mice (n = 4 for each timepoint, two-way ANOVA). CCNE1/β-ACTIN ratio normalized to 0 h Ctrl expression. (B) Scheme of intravenous tail vain injections and different experimental mice groups. (C) qRT-PCR analysis of Cul4a and Fto expression in whole liver of 24 h ST-DEN injected Ctrl/AAV8shRNA-scrmbl, Ctrl/AAV8shRNA−Cul4a, FTOL−KO/AAV8shRNA-scrmb, and FTOL−KO/AAV8shRNA−Cul4a (n = 5, one-way ANOVA). (D) Western blot analysis of CUL4A and FTO in whole liver lysates isolated from 24 h ST-DEN-injected Ctrl/AAV8shRNA-scrmbl, Ctrl/AAV8shRNA−Cul4a, FTOL−KO/AAV8shRNA-scrmb, and FTOL−KO/AAV8shRNA−Cul4a (n = 5). (E) Ki67 ELISA with liver proteins isolated from 24 h ST-DEN-injected Ctrl/AAV8shRNA-scrmbl, Ctrl/AAV8shRNA−Cul4a, FTOL−KO/AAV8shRNA-scrmb, and FTOL−KO/AAV8shRNA−Cul4a (n = 4–5, one-way ANOVA). p∗ 0.05, p∗∗∗∗ 0.0001. Data are means with SD.