Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: p27 is associated with the prognosis and chemotherapy response of EOC patients . A , representative IHC staining images of p27 in human epithelial ovarian cancer (EOC) tissues from 90 patients. This Scale bar represents 200 μm (100 × ), 50 μm (400 × ). B and C , Kaplan-Meier plots depicting progression-free survival (B) and overall survival (C) of EOC patients based on p27 expression (log-rank test). D , quantitative analysis of p27 expression in chemo-sensitive ( N = 68) and chemo-resistant ( N = 22) EOC tissues, based on IHC staining (unpaired two-tailed Student’s t test). E . receiver operating characteristic curve of p27 expression levels and chemotherapy response. F , Western blot analysis of p27 expression in parental and cisplatin-resistant SKOV3 and A2780 cells. β-Actin was used as a loading control. Three biologically independent experiments were performed. G , qRT-PCR analysis of CDKN1B (encoding p27) expression in parental and cisplatin-resistant SKOV3 and A2780 cells. β-Actin served as a loading control. Data are presented as mean ± SEM of three biologically independent experiments (unpaired two-tailed Student’s t test). ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. EOC, epithelial ovarian cancer; IHC, immunohistochemistry; qRT-PCR, quantitative real-time PCR.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: Immunohistochemistry, Expressing, Two Tailed Test, Western Blot, Control, Quantitative RT-PCR, Real-time Polymerase Chain Reaction

Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: Depletion of p27 contributes to cisplatin resistance of EOC cells in vitro and in viv o . A and B , Cell viability ( A ) and IC 50 values ( B ) of indicated EOC cells after treatment with various concentrations of cisplatin (cDDP) for 72 h. Data are presented as mean ± SEM of three biologically independent experiments. C and D , representative images ( C ) and quantification ( D ) of colony formation in the indicated EOC cells. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). E , schematic diagram of the experimental design used to establish the animal model. F , images of excised tumors. G and H , tumor volume ( G ) and tumor weight ( H ) of sgNC/sgp27 SKOV3 subcutaneous xenografts in nude mice after treatment with NS or cDDP. Data are presented as mean ± SEM (n = 6 mice per group, two-way ANOVA). I , body weights of nude mice in the indicated groups. Data represent mean ± SEM (n = 6 mice per group, two-way ANOVA). J , representative IHC staining images of indicated proteins in tumor tissues from subcutaneous xenografts in nude mice. This scale bar represents 50 μm. K and L , quantification of IHC staining for p27 (K) and Ki67 (L) in tumor tissues from subcutaneous xenografts in nude mice. Data represent mean ± SEM of five random fields from six different mice (two-way ANOVA). ns: not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. EOC, epithelial ovarian cancer; IHC, immunohistochemistry.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: In Vitro, Animal Model, Immunohistochemistry

Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: p27 is a key positive regulator of ferroptosis in EOC cells . A , volcano plot showing differentially expressed genes in sgp27-expressing versus sgNC-expressing SKOV3 cells. Red indicates upregulated genes ( p < 0.05, Log FC > 1), and blue indicates downregulated genes ( p < 0.05, Log FC < −1). B , KEGG enrichment analysis of differentially expressed genes in sgp27-expressing versus sgNC-expressing SKOV3 cells, showing the top 15 enriched pathways. C and D , representative images ( C ) and quantification ( D ) of IHC staining for 4-HNE in sgNC/sgp27-expressing SKOV3 subcutaneous xenografts in nude mice treated with NS or cDDP. Scale bar: 50 μm. Data represent mean ± SEM of five random fields from five different mice (two-way ANOVA). E , cell viability of indicated EOC cells treated with cDDP (20 μM) and ferrostatin-1 (10 μM) for 72 h. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). F and G , representative images ( F ) and statistical analysis ( G ) of intracellular ROS in the indicated EOC cells treated with NS or cDDP (20 μM), stained with DCFH-DA, and measured by flow cytometry. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). H and I , representative images ( H ) and statistical analysis ( I ) of intracellular lipid ROS in the indicated groups, stained with BODIPY 581/591-C11 and measured by flow cytometry. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). J and K , Levels of Fe 2+ ( J ) and GSH ( K ) in the indicated groups. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). ns: not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. EOC, epithelial ovarian cancer; IHC, immunohistochemistry; ROS, reactive oxygen species.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: Expressing, Immunohistochemistry, Staining, Flow Cytometry

Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: Depletion of p27 reduces Erastin-induced ferroptosis and attenuates its antitumor efficacy . A and D , levels of intracellular ROS ( A ), lipid ROS ( B ), Fe 2+ ( C ), and GSH ( D ) in the indicated groups. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). E , schematic diagram of the experimental design used to establish the animal model. F , images of removed tumors. G and H , tumor volume ( G ) and tumor weight ( H ) of sgNC/sgp27 SKOV3 subcutaneous xenografts in nude mice following solvent control or Erastin treatment. Data represent mean ± SEM (n = 4 mice per group, two-way ANOVA). I , body weights of nude mice in the indicated groups. Data represent mean ± SEM (n = 4 mice per group, two-way ANOVA). J , representative images of IHC staining of indicated proteins in tumor tissues from subcutaneous xenografts in nude mice. This scale bar represents 50 μm. K , quantification of IHC staining of p27, 4-HNE, and Ki67 in tumor tissues from subcutaneous xenografts in nude mice. Data represent mean ± SEM of five random fields of view from six different mice (two-way ANOVA). ns: not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. EOC, epithelial ovarian cancer; IHC, immunohistochemistry; ROS, reactive oxygen species.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: Animal Model, Solvent, Control, Immunohistochemistry

Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: p27 enhances the transcriptional activity of CYBB in EOC cells . A , venn diagram showing the intersection of differentially expressed genes in sgp27 versus sgNC SKOV3 cells with ferroptosis-related genes from the FerrDb database ( http://www.zhounan.org ) and the heatmap showing seven ferroptosis-related genes among the differentially expressed genes. B , qRT-PCR analysis of indicated ferroptosis-related genes in indicated EOC cells. β-Actin was used as the loading control. Data represent mean ± SEM of three biologically independent experiments (unpaired two-tailed Student’s t test). C , Western blot analysis of p27 and CYBB expression in indicated EOC cells. β-Actin was used as the loading control. Three biologically independent experiments were performed. D and E , representative images ( D ) and quantification ( E ) of IHC staining of CYBB in sgNC/sgp27 SKOV3 subcutaneous xenografts in nude mice treated with NS, cDDP, or solvent control and Erastin. This scale bar represents 50 μm. Data represent mean ± SEM of five random fields of view per mouse (two-way ANOVA). F , schematic diagram of the different fragments of the CYBB promoter. G , relative luciferase activity of the CYBB promoter in p27-overexpressing CAOV3 cells. Data represent mean ± SEM of three biologically independent experiments (unpaired two-tailed Student’s t test). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. CYBB, cytochrome b-245 heavy chain; EOC, epithelial ovarian cancer; IHC, immunohistochemistry.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: Activity Assay, Quantitative RT-PCR, Control, Two Tailed Test, Western Blot, Expressing, Immunohistochemistry, Solvent, Luciferase

Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: p27 enhances cisplatin sensitivity in EOC by partially increasing CYBB expression and activating ferroptosis . A , Western blot analysis of p27 expression in indicated EOC cells. β-Actin was used as the loading control. Three biologically independent experiments were performed. B and E , levels of intracellular ROS ( B ), lipid ROS ( C ), Fe 2+ ( D ), and GSH ( E ) in the indicated groups. Data represent mean ± SEM of three biologically independent experiments (one-way ANOVA). F , cell viability and IC 50 values of indicated EOC cells following treatment with cisplatin at various concentrations for 72 h. Data represent mean ± SEM of three biologically independent experiments. G and H , representative images ( G ) and quantification ( H ) of colony formation in indicated EOC cells. Data represent mean ± SEM of three biologically independent experiments (two-way ANOVA). I , representative IHC staining images of CYBB in human EOC tissues from 90 patients. The Scale bar represent: 200 μm (100 × ), 50 μm (400 × ). J , quantitative analysis of CYBB protein levels in EOC chemo-sensitive ( N = 68) and chemo-resistant ( N = 22) tissues based on IHC staining (unpaired two-tailed Student’s t test). K , correlation between the IHC scores of p27 and CYBB (Spearman’s correlation test). ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. EOC, epithelial ovarian cancer; ROS, reactive oxygen species.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: Expressing, Western Blot, Control, Immunohistochemistry, Two Tailed Test

Journal: The Journal of Biological Chemistry

Article Title: The cyclin-dependent kinase inhibitor p27 facilitates chemosensitivity by promoting ferroptosis in epithelial ovarian cancer

doi: 10.1016/j.jbc.2025.111011

Figure Lengend Snippet: SKPin C1 increases the expression of p27 and enhances the anti-tumor effect of cisplatin of EOC in vitro and in viv o . A , Western blot analysis of p27 expression in CAOV3 cells following treatment with SKPin C1 at various concentrations for 24 h. β-Actin was used as the loading control. Three biologically independent experiments were performed. B , percentage inhibition at each concentration of cisplatin, SKPin C1, or their combination in CAOV3 cells. C , combination index (CI) scores for CAOV3 cells treated with cisplatin in combination with SKPin C1 at the indicated concentrations. D , schematic diagram of the experimental design used to establish the animal model. E , images of removed tumors. F – G Tumor volume ( F ) and tumor weight ( G ) of CAOV3 subcutaneous xenografts in nude mice following treatment with solvent control, cisplatin, SKPin C1, or cisplatin + SKPin C1. Data represent mean ± SEM (n = 5 mice per group, one-way ANOVA). H , body weights of nude mice in the indicated groups. Data represent mean ± SEM (n = 5 mice per group, one-way ANOVA). I , representative IHC staining images of indicated proteins in tumor tissues from subcutaneous xenografts in nude mice. This scale bar represent 50 μm. J and L , quantification of IHC staining for p27 ( J ), Ki67 ( K ), and 4-HNE ( L ) in tumor tissues from subcutaneous xenografts in nude mice. Data represent mean ± SEM of five random fields of view from five different mice (one-way ANOVA). ns: not significant, ∗ p < 0.05, ∗∗ p < 0.01, ∗∗∗ p < 0.001, ∗∗∗∗ p < 0.0001. CYBB, cytochrome b-245 heavy chain; CI, combination index; EOC, epithelial ovarian cancer; IHC, immunohistochemistry.

Article Snippet: The primary antibodies used for Western blot included anti-p27 (Proteintech; 25614-1-AP; 1:1000 dilution), anti-CYBB (Proteintech; 19013-1-AP; 1:1000 dilution), anti-Skp2 (Proteintech; 15010-1-AP; 1:1000 dilution), anti-β-actin (ABclonal; AC026; 1:10,000 dilution), and anti-Vinculin (Proteintech; 26520-1-AP; 1:10,000 dilution).

Techniques: Expressing, In Vitro, Western Blot, Control, Inhibition, Concentration Assay, Animal Model, Solvent, Immunohistochemistry