Journal: Advanced Science

Article Title: Long Non‐Coding RNA IGFRIL Couples with PTBP1 to Destabilize IGFBP3 mRNA to Promote the IGF1R‐AKT‐mTOR Axis and Hepatocellular Carcinoma

doi: 10.1002/advs.202507676

Figure Lengend Snippet: IGFRIL couples with PTBP1 to enhance the activation of AKT‐mTOR signaling in HCC cells. A) Overlapping DEGs upon IGFRIL and PTBP1 knockdown in HepG2 cells. B) GO and KEGG pathway enrichment analyses of the overlapping differentially expressed genes (DEGs). C) Spearman's correlation of gene set enrichment scores between IGFRIL and PTBP1 knockdown. NES, normalized enrichment score. D) GSEA plots of “PI3K_EVENTS_IN_ERBB4_SIGNALING” in IGFRIL ‐knocked‐down and PTBP1 ‐knocked‐down HepG2 cells. E) IB assays of key cascades of the receptor tyrosine kinases (RTK) and AKT‐mTOR signaling in HepG2 cells upon IGFRIL knockdown or overexpression. F) Knockdown of IGFRIL accelerates the autophagic flux in HepG2 cells. HepG2 cells stably expressing mRFP‐GFP‐LC3 were treated with DMSO, NVP‐AEW541 (2 µ m ), or Rapamycin (10 µ m ) for 12 h upon knockdown of IGFRIL , followed by staining with DAPI. Red puncta represent the autophagosomes; and yellow puncta in the merged picture represent the autolysosomes. Scale bars, 40 µm. G,H) Rapamycin attenuates the promoting effects of IGFRIL overexpression on cell malignant phenotypes and mTOR signaling. Data are shown as mean ± SEM, * P < 0.05, ** P < 0.01, *** P < 0.001 by 1‐way ANOVA (E,G,H). n.s., not significant.

Article Snippet: When the tumor volume reached ≈100 mm 3 after implantation, the mice were randomly divided into three treatment groups for assessing the sensitivity of PDXs with different IGFRIL levels to IGF1R or mTOR inhibitors: 1) DMSO (intraperitoneal injection); 2) BMS‐754807 (25 mg kg −1 every 5 days, intraperitoneal injection; Cat. s1124, Selleck, USA); and 3) rapamycin (10 mg kg −1 every 5 days, intraperitoneal injection; Cat. s1039, Selleck, USA).

Techniques: Activation Assay, Knockdown, Over Expression, Stable Transfection, Expressing, Staining

Journal: Advanced Science

Article Title: Long Non‐Coding RNA IGFRIL Couples with PTBP1 to Destabilize IGFBP3 mRNA to Promote the IGF1R‐AKT‐mTOR Axis and Hepatocellular Carcinoma

doi: 10.1002/advs.202507676

Figure Lengend Snippet: IGFRIL activates the IGF1R‐AKT‐mTOR signaling and plays oncogenic roles dependent on IGFBP3. A) The effects of knockdown or overexpression of IGFBP3 on the activities of the IGF1R‐AKT‐mTOR axis. B) The effects of IGFBP3 overexpression on IGFRIL ‐activated IGF1R‐AKT‐mTOR axis, and the effects of IGFBP3 knockdown on IGFRIL knockdown‐repressed IGF1R‐AKT‐mTOR axis. C,D) The effects of IGFBP3 knockdown or overexpression on the abilities of plate colony formation, migration, and invasion. E) The effects of IGFBP3 overexpression on the IGFRIL ‐enhanced plate colony formation, migration, and invasion. F) The effects of IGFBP3 knockdown on the IGFRIL knockdown‐reduced plate colony formation, migration, and invasion. G) The effects of IGF1R inhibition with its inhibitor AEW541 (2 µM) or siRNAs against IGF1R on the IGFRIL‐activated IGF1R‐AKT‐mTOR axis. H,I) The effects of IGF1R inhibition on the IGFRIL ‐enhanced plate colony formation, migration, and invasion. J) j Spearman's correlations between the levels of IGFRIL and levels of IGFBP3, p‐IGF1R, p‐AKT and p‐mTOR, respectively, in HCC tumor tissues ( n = 83) from the VALI1 cohort. Data are shown as mean ± SEM, * P < 0.05, ** P < 0.01, *** P < 0.001 by 1‐way ANOVA (C,E,F,G,I) or Student's t‐ test (D).

Article Snippet: When the tumor volume reached ≈100 mm 3 after implantation, the mice were randomly divided into three treatment groups for assessing the sensitivity of PDXs with different IGFRIL levels to IGF1R or mTOR inhibitors: 1) DMSO (intraperitoneal injection); 2) BMS‐754807 (25 mg kg −1 every 5 days, intraperitoneal injection; Cat. s1124, Selleck, USA); and 3) rapamycin (10 mg kg −1 every 5 days, intraperitoneal injection; Cat. s1039, Selleck, USA).

Techniques: Knockdown, Over Expression, Migration, Inhibition

Journal: Advanced Science

Article Title: Long Non‐Coding RNA IGFRIL Couples with PTBP1 to Destabilize IGFBP3 mRNA to Promote the IGF1R‐AKT‐mTOR Axis and Hepatocellular Carcinoma

doi: 10.1002/advs.202507676

Figure Lengend Snippet: Upregulation of IGFRIL sensitizes HCC cells to IGF1R and mTOR inhibitors. A) Spearman's correlations between the levels of IGFRIL and sensitivities to inhibitors against IGF1R (BMS‐754807) and mTOR (rapamycin) in eight types of HCC cell lines. The effects of BMS‐754807 or rapamycin on HCC cell growth were determined by CCK‐8 assays. B) The effects of IGFRIL knockdown on the sensitivity to BMS‐754807 or rapamycin in HepG2 cells. C) The effects of IGFRIL overexpression on the sensitivity to BMS‐754807 or rapamycin in JHH‐2 cells. D) The effects of BMS‐754807 or rapamycin treatment on tumor growth in patient‐derived xenografts (PDX) mouse models. The mice were treated with control (DMSO), BMS‐754807 (25 mg kg −1 every 5 days, intraperitoneal injection) or rapamycin (10 mg kg −1 every 5 days, intraperitoneal injection) ( n = 5 mice/group). E–G) The effects of IGFRIL knockdown or overexpression on the cell growth, migration, and invasion (E,F), and the activities of the IGF1R‐AKT‐mTOR axis (G) in IGFRIL high and IGFRIL low patient‐derived primary tumor cells (PDCs), respectively. H) The effects of BMS‐754807 (left) or rapamycin (right) treatment on the growth of IGFRIL high or IGFRIL low PDCs. I) The effects of IGFRIL knockdown or overexpression on the sensitivities to BMS‐754807 and rapamycin in IGFRIL high and IGFRIL low PDCs, respectively. J) Schematic diagram of the function and mechanism of IGFRIL in HCC tumorigenesis and drug sensitivity. Data are shown as mean ± SEM, * P < 0.05, ** P < 0.01, *** P < 0.001 by 1‐way ANOVA (B,D,E,I) or Student's t test (C,F,H).

Article Snippet: When the tumor volume reached ≈100 mm 3 after implantation, the mice were randomly divided into three treatment groups for assessing the sensitivity of PDXs with different IGFRIL levels to IGF1R or mTOR inhibitors: 1) DMSO (intraperitoneal injection); 2) BMS‐754807 (25 mg kg −1 every 5 days, intraperitoneal injection; Cat. s1124, Selleck, USA); and 3) rapamycin (10 mg kg −1 every 5 days, intraperitoneal injection; Cat. s1039, Selleck, USA).

Techniques: CCK-8 Assay, Knockdown, Over Expression, Derivative Assay, Control, Injection, Migration

Journal: Brazilian Journal of Medical and Biological Research

Article Title: Knockdown of long non-coding RNA ANRIL inhibits tumorigenesis in human gastric cancer cells via microRNA-99a-mediated down-regulation of BMI1

doi: 10.1590/1414-431x20186839

Figure Lengend Snippet: Figure 6. BMI1 inhibited the apoptotic pathway and activated the Notch and mTOR pathways. Protein expression was determined by western blotting. A and B, BMI1 was abnormally expressed after cell transfection. C and D, Bcl-2 expression was down-regulated while expressions of p16, cleaved caspase-9, and cleaved caspase-3 were up-regulated by BMI1 silence. E and F, Phosphorylated levels of key kinases in the Notch and mTOR pathways were increased by BMI1 overexpression. BMI1: B-lymphoma Mo-MLV insertion region 1; pEX-BMI1: recombined overexpression vector of BMI1; shBMI1: pENTRTM/U6 vector carrying small hairpin RNA targeting BMI1; shNC: pENTRTM/U6 vector carrying a non-targeting sequence; p16: multiple tumor suppressor 1; Bcl-2: B-cell lymphoma 2; mTOR: mam- malian target of rapamycin; p70S6K: p70 ribosomal protein S6 kinase; p-: phosphorylated.

Article Snippet: The primary antibodies used in this study were as follows: BMI1 (sc-10745); multiple tumor suppressor 1 (p16; sc-166760); B-cell lymphoma 2 (Bcl-2; sc-7328); caspase-3 (sc-271759); cleaved caspase-3 (sc-22171); caspase-9 (sc-56076); cleaved caspase-9 (sc-22182); Notch 1 (sc-376403); mTOR (sc-293089); phosphorylated mTOR (p-mTOR; s-c29313); p70 ribosomal protein S6 kinase (p70S6K; sc-9027), and phosphorylated p70S6K (p-p70S6K; sc-8416, all from Santa Cruz Biotechnology).

Techniques: Expressing, Western Blot, Transfection, Over Expression, Plasmid Preparation, Sequencing