human rcc caki 1 cells  (ATCC)

Journal: Advanced Science

Article Title: TWF2 Drives Tumor Progression and Sunitinib Resistance in Renal Cell Carcinoma through Hippo Signaling Suppression

doi: 10.1002/advs.202506367

Figure Lengend Snippet: TWF2 is identified as a master regulator for RCC drug resistance and tumor progression. A) Schematic diagram illustrating the establishment of sunitinib‐resistant (786‐O‐R, 769‐P‐R) and sunitinib‐sensitive (786‐O‐S, 769‐P‐S) RCC cell lines. B) Flow chart of screening key regulators involved in RCC drug resistance and tumor progression. C) Venn diagram displaying the intersection of five datasets: MS Top100, RNA‐seq Top100, TCGA–KIRC OS (genes upregulated in tumors vs normal tissues), TCGA–KIRC Metastasis (genes upregulated in metastatic vs primary tumors), and TCGA–KIRC Prognosis (genes associated with poor prognosis). This analysis identified six overlapping genes: CTHRC1 , TWF2 , COL6A3 , SLC38A5 , IFI44 , and OASL . D) Volcano plot of differentially expressed genes in sunitinib‐resistant versus sunitinib‐sensitive 786‐O cells based on transcriptomic analysis. Genes upregulated in resistant cells are shown in red, and downregulated genes in blue. E) Relative TWF2 mRNA expression in paired tumor and adjacent normal tissues from the SYSU ccRCC cohort. F) Representative western blot (left) and the corresponding statistical analysis (right) of TWF2 protein expression levels in twelve paired ccRCC tumors (T) and adjacent normal tissues (N). G) Representative western blot (top) and mRNA expression analysis (bottom) of TWF2 in sunitinib‐sensitive (S) and ‐resistant (R) 786‐O cell. H) Representative immunohistochemical (IHC) staining showing TWF2 expression in ccRCC tumors and adjacent normal tissues. I) Representative IHC images of TWF2 expression in ccRCC tissues from patients classified as responders or nonresponders to sunitinib treatment. J) Overall survival (OS) in ccRCC patients with low ( n = 60) or high ( n = 60) TWF2 expression. K) Disease‐free survival (DFS) in ccRCC patients with low ( n = 60) or high ( n = 60) TWF2 . Data are presented as means ± SD and are analyzed by Student's t ‐test (E–G) or log‐rank test (J, K). ** p < 0.01; *** p < 0.001.

Article Snippet: The immortalized renal epithelial cell line (HK2) and human RCC cell lines (786‐O, 769‐P, A‐498, ACHN, Caki‐1, and Caki‐2) were obtained from the American Type Culture Collection.

Techniques: RNA Sequencing, Expressing, Western Blot, Immunohistochemical staining, Immunohistochemistry

Journal: NPJ Precision Oncology

Article Title: m6A reader IGF2BP2-stabilized lncRNA LHX1-DT inhibits renal cell carcinoma (RCC) cell proliferation and invasion by sponging miR-590-5p

doi: 10.1038/s41698-025-00958-x

Figure Lengend Snippet: A The m6A level was identified in RCC tissues and adjacent normal renal tissues by qRT-PCR. B The m6A level was identified in normal renal tubular epithelial cells, 293 T cells, and RCC cell lines by qRT-PCR. C , D The qRT-PCR ( C ) and western blot ( D ) analysis of the relative expression of 10 m6A regulatory factors in RCC tissues and adjacent normal renal tissues. E The qRT-PCR analysis of the relative expression of METTL14 in HK-2 cells, 293 T cells, and RCC cell lines. F The protein expression levels of METTL14 in RCC tissues and adjacent normal renal tissues. G The protein expression levels of METTL14 in HK-2 cells, 293 T cells, and RCC cell lines. H Verification of METTL14 overexpression by western blot assay. I The m6A level was detected after METTL14 overexpression by qRT-PCR.

Article Snippet: Human RCC cells (Caki-1, 786-O, and ACHN), human renal tubular epithelial cells (HK-2), and 293 T cells were purchased from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Quantitative RT-PCR, Western Blot, Expressing, Over Expression

Journal: NPJ Precision Oncology

Article Title: m6A reader IGF2BP2-stabilized lncRNA LHX1-DT inhibits renal cell carcinoma (RCC) cell proliferation and invasion by sponging miR-590-5p

doi: 10.1038/s41698-025-00958-x

Figure Lengend Snippet: A LHX1-DT was predicted to be located mainly in the cytoplasm using the bioinformatics tools in lncATLAS. B The qRT-PCR analysis of subcellular LHX1-DT expression in the nucleus and cytoplasm of RCC cells. C Subcellular localization of LHX1-DT in RCC cells detected by RNA-FISH. LHX1-DT was stained red (Cy3), and nuclei were stained blue (DAPI). D Schematic diagram of the Ago2-RIP process. E Fold enrichment of LHX1-DT in 786-O and Caki-1 cells. F Venn diagram showing the predicted target of LHX1-DT. G Relative expression of miR-590-5p in 52 paired RCC tissues compared with adjacent normal renal tissues by qRT-PCR. H RIP assay was performed using AGO2 antibody in 786-O and Caki-1 cells, then the enrichment of miR-590-5p was detected. I RIP assay was performed using AGO2 antibody in 786-O and Caki-1 cells transfected with miR-590-5p mimics or mimics NC, then the enrichment of LHX1-DT was detected. J The correlation between miR-590-5p and LHX1-DT was evaluated, and regression analysis was applied ( r = −0.2984, P = 0.0317). K Expression level of miR-590-5p in 786-O and Caki-1 cells after transfection with overexpression plasmids of LHX1-DT. L Expression level of miR-590-5p in 786-O and Caki-1 cells treated with siRNAs of LHX1-DT. M The binding sites of LHX1-DT and miR-590-5p. N , O Luciferase reporter activity of wild-type (WT) or mutated (MUT) LHX1-DT co-transfected with miR-590-5p mimics in 786-O ( N ) and Caki-1 ( O ) cells. P LHX1-DT was pulled down with biotinylated miR-590-5p, and miR-590-5p was also pulled down with biotinylated LHX1-DT in 786-O and Caki-1 cells. Q , R The addition of functional FOXQ1-cDNA partially rescued the growth ( Q ) and invasion ( R ) of 786-O and Caki-1 cells transfected with miR-590-5p mimics. S , T The addition of LHX1-DT-shRNA partially rescued the growth ( S ) and invasion ( T ) of 786-O and Caki-1 cells transfected with an miR-590-5p inhibitor.

Article Snippet: Human RCC cells (Caki-1, 786-O, and ACHN), human renal tubular epithelial cells (HK-2), and 293 T cells were purchased from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Quantitative RT-PCR, Expressing, Staining, Transfection, Over Expression, Binding Assay, Luciferase, Activity Assay, Functional Assay, shRNA

Journal: NPJ Precision Oncology

Article Title: m6A reader IGF2BP2-stabilized lncRNA LHX1-DT inhibits renal cell carcinoma (RCC) cell proliferation and invasion by sponging miR-590-5p

doi: 10.1038/s41698-025-00958-x

Figure Lengend Snippet: A , B The m6A ( A ) and expression of LHX1-DT ( B ) were detected after METTL14 overexpression by qRT-PCR. C The correlation between METTL14 and LHX1-DT was evaluated, and regression analysis was applied ( r = 0.09209, P = 0.0304). D , E Reduction of LHX1-DT RNA stability in METTL14 knockdown 786-O ( D ) and Caki-1 ( E ) cells as compared to control. Cells were treated with 5 μg/mL actinomycin-D and RNA was isolated at 0 h, 2 h and 4 h. F Relative expression of LHX1-DT was detected by RIP assay. G RIP analysis showing the enrichment of LHX1-DT on IGF2BP2 in the indicated cells. H , I Reduction of LHX1-DT RNA stability in IGF2BP2 knockdown 786-O ( H ) and Caki-1 ( I ) cells as compared to control. Cells were treated with 5 μg/mL actinomycin-D and RNA was isolated at 0 h, 2 h and 4 h. J Relative expression of IGF2BP2 in 523 RCC tissues compared with 100 adjacent normal renal tissues in the GEPIA2 dataset. K The putative wild-type m6A sites and designed mutant m6A sites in LHX1-DT. L The RIP analysis shows the enrichment of LHX1-DT on IgG and IGF2BP2 in the LHX1-DT-Wt or LHX1-DT-Mut RCC cells. M The qRT-PCR analysis of LHX1-DT expression in the LHX1-DT-Wt or LHX1-DT-Mut RCC cells with or without METTL14 or IGF2BP2 knockdown. N Schematic representation of mutated LHX1-DT of pmirGLO vector to investigate the m6A roles on LHX1-DT expression. O The luciferase activities of different mutated LHX1-DT reporters in the indicated groups.

Article Snippet: Human RCC cells (Caki-1, 786-O, and ACHN), human renal tubular epithelial cells (HK-2), and 293 T cells were purchased from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Expressing, Over Expression, Quantitative RT-PCR, Knockdown, Control, Isolation, Mutagenesis, Plasmid Preparation, Luciferase

Journal: NPJ Precision Oncology

Article Title: m6A reader IGF2BP2-stabilized lncRNA LHX1-DT inhibits renal cell carcinoma (RCC) cell proliferation and invasion by sponging miR-590-5p

doi: 10.1038/s41698-025-00958-x

Figure Lengend Snippet: A Venn diagram showing the predicted target genes of miR-590-5p. B The qRT-PCR analysis of the relative expression of 11 possible targets of miR-590-5p in RCC tissues and adjacent normal renal tissues. C Relative expression of PDCD4 in 52 paired RCC tissues compared with adjacent normal renal tissues by qRT-PCR. D Kaplan–Meier analysis of the correlation between PDCD4 expression and OS in RCC patients according to the GEPIA2 dataset. E The correlation between PDCD4 and miR-590-5p was evaluated, and regression analysis was applied ( r = −0.2868, P = 0.0393). F The expression of PDCD4 was detected after silencing or overexpression of miR-590-5p by qRT-PCR. G Western blot assay showing protein levels of PDCD4 after silencing or overexpression of miR-590-5p. H The binding sites of PDCD4 and miR-590-5p. I , J Luciferase reporter activity of wild-type (WT) or mutated (MUT) PDCD4 co-transfected with miR-590-5p mimics in 786-O ( I ) and Caki-1 ( J ) cells. K , L The addition of functional PDCD4-cDNA partially rescued the growth ( K ) and invasion ( L ) of 786-O and Caki-1 cells transfected with miR-590-5p mimics. M , N The addition of PDCD4-shRNA partially rescued the growth ( M ) and invasion ( N ) of 786-O and Caki-1 cells transfected with an miR-590-5p inhibitor. O The correlation between LHX1-DT and PDCD4 was evaluated, and regression analysis was applied ( r = 0.8933, P < 0.0001). P Western blot assay showing protein levels of PDCD4 after overexpression or silencing of LHX1-DT. Q The expression of PDCD4 was detected after overexpression or silencing of LHX1-DT by qRT-PCR. R Protein level of PDCD4 was determined by western blot assay after co-transfection with functional LHX1-DT-cDNA and miR-590-5p mimics. S Protein level of PDCD4 was determined by western blot assay after co-transfection with LHX1-DT-shRNA and an miR-590-5p inhibitor. T The expression of PDCD4 was detected after co-transfection with functional LHX1-DT-cDNA and miR-590-5p mimics. U The expression of PDCD4 was detected after co-transfection with LHX1-DT-shRNA and an miR-590-5p inhibitor.

Article Snippet: Human RCC cells (Caki-1, 786-O, and ACHN), human renal tubular epithelial cells (HK-2), and 293 T cells were purchased from American Type Culture Collection (ATCC, Manassas, USA).

Techniques: Quantitative RT-PCR, Expressing, Over Expression, Western Blot, Binding Assay, Luciferase, Activity Assay, Transfection, Functional Assay, shRNA, Cotransfection