Journal: Journal of Translational Medicine

Article Title: GX15-070 enhances niraparib efficacy in ovarian cancer by promoting a shift in Mcl1-mediated DNA repair pathway from HR to NHEJ

doi: 10.1186/s12967-025-07284-7

Figure Lengend Snippet: Inhibition of Mcl1 enhances sensitivity to niraparib and impairs HR activity. (A) GSEA analysis using TCGA-OV dataset showed the correlation between Mcl1 expression and HR and NHEJ pathways. (B-C) γH2AX foci was used to detect the influence of Mcl1 expression on repair of DNA damage induced by niraparib in A2780 and PEO1 cell lines. Cells were subjected to Mcl1 knockdown, followed by treatment with or without niraparib for 24 h. After removing niraparib, cells were cultured in normal medium for an additional 24 h. Immunofluorescence staining with γH2AX antibody was performed at different time points. (D-E) Representative immunofluorescence micrographs and quantification of RAD51 foci in A2780 and PEO1 cell lines after Mcl1 knockdown. (F) The effect of niraparib on cell viability in A2780 and PEO1 cell lines after Mcl1 knockdown. (G) Representative IHC micrographs of the Mcl1 protein expression in ovarian cancer tissues (Scale bar = 50 μm, 40 × magnification). (H-J) The influence of Mcl1 expression on clinical outcome in ovarian cancer patients treated with PARPi maintenance therapy. Experiments were repeated three times and represented as mean ± SD. *P < 0.05 , **P < 0.01 , ***P < 0.001

Article Snippet: After subjecting the lysates to SDS-PAGE electrophoresis, proteins were transferred onto a polyvinylidene difluoride membrane by electroblotting, blocked in 5% nonfat milk, and then blotted with anti-bodies targeting Mcl1 (16225-1-AP, Proteintech, 1:2000), Ku70 (10723-1-AP, Proteintech, 1:5000) and β-actin (ab8227, Abcam; 1:1,000).

Techniques: Inhibition, Activity Assay, Expressing, Knockdown, Cell Culture, Immunofluorescence, Staining

Journal: Journal of Translational Medicine

Article Title: GX15-070 enhances niraparib efficacy in ovarian cancer by promoting a shift in Mcl1-mediated DNA repair pathway from HR to NHEJ

doi: 10.1186/s12967-025-07284-7

Figure Lengend Snippet: GX15-070 enhances niraparib sensitivity and impairs HR activity by competitively inhibiting the interaction between Mcl1 and Ku70. (A) Co-IP analysis to measure the interaction of KU70 protein and Mcl1 protein in A2780 and PEO1 cells. IgG was used as a negative control. (B) Schematic representation of Mcl1 protein structure showing its PEST sequence, BH domains (BH3, BH1, BH2), and transmembrane (TM) domain. The ΔBH3 mutant lacks the BH3 domain. (C) Co-IP were used analyze the interaction of BH3 domain of Mcl1 and KU70 protein. FLAG antibody was used for IP, and KU70 expression was detected. (D) Co-IP were used to detect the influence of GX15-070 on the interaction of KU70 and Mcl1 protein. Mcl1 was immunoprecipitated and probed for KU70. (E-F) BH3 domain in Mcl1 is required to repair niraparib-induced DSBs. Red dots indicated γH2AX foci in A2780 and PEO1 cells. (G-J) BH3 domain in Mcl1 is required to promote the DNA repair pathway from NHEJ to HR. (K) BH3 domain in Mcl1 played a critical role in the sensitivity of niraparib in A2780 and PEO1 cell lines. Experiments were repeated three times and represented as mean ± SD. *P < 0.05 , **P < 0.01 , ***P < 0.001 (siCtrl vs. siMcl1); # P < 0.05 , ## P < 0.01 , ### P < 0.001 (Compared to siMcl1)

Article Snippet: After subjecting the lysates to SDS-PAGE electrophoresis, proteins were transferred onto a polyvinylidene difluoride membrane by electroblotting, blocked in 5% nonfat milk, and then blotted with anti-bodies targeting Mcl1 (16225-1-AP, Proteintech, 1:2000), Ku70 (10723-1-AP, Proteintech, 1:5000) and β-actin (ab8227, Abcam; 1:1,000).

Techniques: Activity Assay, Co-Immunoprecipitation Assay, Negative Control, Sequencing, Mutagenesis, Expressing, Immunoprecipitation

Journal: Journal of Translational Medicine

Article Title: GX15-070 enhances niraparib efficacy in ovarian cancer by promoting a shift in Mcl1-mediated DNA repair pathway from HR to NHEJ

doi: 10.1186/s12967-025-07284-7

Figure Lengend Snippet: GX15-070 combined with niraparib suppresses tumor growth in CDX and PDX models. (A-B) Tumor growth curves and photograph of the tumors harvested at the end of the experiments with the CDX models treated with vehicle, niraparib, GX15-070 or a combination. (C) Experimental design to evaluate combination effect of niraparib and GX15-070 in PDX models. (D) Tumor growth curves and quantification of the tumor weights in the different groups of PDX models. (E) Representative IHC micrographs of indicated markers in the different groups of CDX models and PDX models (Scale bar = 100 μm). (F) Schematic diagram for GX15-070 in enhancing niraparib sensitivity. Briefly, GX15-070 competitively disrupts the interaction between Mcl1 and Ku70, leading to promoting a shift in Mcl1-mediated DNA repair pathway from HR to NHEJ. *P < 0.05 , **P < 0.01 , ***P < 0.001

Article Snippet: After subjecting the lysates to SDS-PAGE electrophoresis, proteins were transferred onto a polyvinylidene difluoride membrane by electroblotting, blocked in 5% nonfat milk, and then blotted with anti-bodies targeting Mcl1 (16225-1-AP, Proteintech, 1:2000), Ku70 (10723-1-AP, Proteintech, 1:5000) and β-actin (ab8227, Abcam; 1:1,000).

Techniques: