Journal: International Journal of Oncology

Article Title: Cyclovirobuxine D inhibits growth and progression of non-small cell lung cancer cells by suppressing the KIF11-CDC25C-CDK1-CyclinB1 G 2 /M phase transition regulatory network and the NFκB/JNK signaling pathway

doi: 10.3892/ijo.2023.5505

Figure Lengend Snippet: KIF11, KIF11-CDK1-CDC25C-cyclinB1 oncogenic network and NF-kB/JNK signaling pathway are the potential CVB-D therapeutic targets in the NSCLC cells. (A) Venn diagram showed the overlap number of potential CVB-D target genes in NSCLC cells by intersecting four LUAD datasets. (B) Protein-protein interaction network analysis revealed interaction between the 10 overlapping DEGs. The disconnected nodes are hidden. (C) The interactive heatmap from GEPIA 2 database showed expression of the 6 DEGs in LUAD and normal lung tissues. KIF11 is the most differentially expressed gene between LUAD and normal lung tissues. (D) Venn diagram revealed 2 common genes (KIF11 and CDK1) based on intersection between the 10 overlapping DEGs and the 100 predicted CVB-D target genes extracted from the Swisstarget prediction database. (E) Western blot analysis identified the expression of KIF11, CDK1, CDC25C and cyclinB1 proteins in the control and CVB-D-treated NSCLC cells. (F and G) Immunohistochemical assay results showed the expression levels and localization of the KIF11 protein in the control and CVB-D-treated A549 xenograft tumor tissues. (H) Western blot assay results revealed the KIF11 protein levels in the si-NC and si-KIF11-transfected NSCLC cells. (I) Western blot analysis demonstrated the expression levels of the oncogenic signaling network proteins (KIF11, CDK1, CDC25C and CyclinB1) and the NF-κB/JNK signaling pathway proteins (p65, p-p65, JNK and p-JNK) in the si-NC- and si-KIF11-transfected NSCLC cells. ** P<0.01 vs. control group. KIF11, kinesin family member 11; CVB-D, cyclovirobuxine D; NSCLC, non-small cell lung cancer; LUAD, lung adenocarcinoma; DEGs, differentially expressed genes; si, small interfering; NC, negative control; p-, phosphorylated.

Article Snippet: Then, the slices were blocked for 30 min at 37°C with goat serum (included in the UltraSensitive™ SP IHC kit) and incubated with primary antibodies (diluted with 5% BSA) overnight at 4°C against the target proteins: anti-KIF11 (1:100; cat no. 23333-1-AP; from Proteintech Group, Inc.), anti-CDC25C (1:100; cat no. AF6258), anti-CDK1 (1:100; cat no. AF6108) and anti-cyclinB1 (1:100; cat no. AF6168; all from Affinity Biosciences, Ltd.); anti-Ki67 (1:100; cat no. A20018), anti-Bcl2 (1:100; cat no. A19693) and anti-N-Cadherin (1:100; cat no. A0433; all from ABclonal Biotech Co., Ltd.).

Techniques: Expressing, Western Blot, Immunohistochemistry, Transfection, Negative Control

Journal: International Journal of Oncology

Article Title: Cyclovirobuxine D inhibits growth and progression of non-small cell lung cancer cells by suppressing the KIF11-CDC25C-CDK1-CyclinB1 G 2 /M phase transition regulatory network and the NFκB/JNK signaling pathway

doi: 10.3892/ijo.2023.5505

Figure Lengend Snippet: CVB-D inhibits in vivo progression of A549 xenografts tumors. (A) Experimental design for tumor xenograft experiments of NSCLC in nude mice to evaluate the in vivo therapeutic effects of CVB-D. Specific time points for CVB-D treatment and experimental analysis are indicated. (B) Representative images of the NSCLC xenograft tumor models with or without CVB-D treatment. (C) Representative images of the NSCLC xenograft tumor tissues from control and CVB-D-treated mice. (D and E) NSCLC xenograft tumor weights and volumes in the control and CVB-D treatment groups of nude mice. (F) Representative H&E and TUNEL staining images of the A549 xenograft tumor sections in the control and CVB-D treatment groups of nude mice (magnification, ×200; scale bar, 50 µ m). (G) The proportion of apoptotic cells in the A549 xenograft tumors derived from the control and CVB-D-treatment groups of nude mice based on TUNEL staining. (H) The body weights of the xenograft tumor model mice in the control and CVB-D treatment groups. (I and J) Immunohistochemical assay data revealed the expression levels and distribution of CDK1, CDC25C, cyclinB1, Ki67, Bcl-2 and N-cadherin proteins in the A549 xenograft tumors derived from the control and CVB-D treatment groups (magnification, ×200; scale bar, 50 µ m). * P<0.05 and ** P<0.01 vs. control. CVB-D, cyclovirobuxine D; NSCLC, non-small cell lung cancer; H&E, hematoxylin and eosin.

Article Snippet: Then, the slices were blocked for 30 min at 37°C with goat serum (included in the UltraSensitive™ SP IHC kit) and incubated with primary antibodies (diluted with 5% BSA) overnight at 4°C against the target proteins: anti-KIF11 (1:100; cat no. 23333-1-AP; from Proteintech Group, Inc.), anti-CDC25C (1:100; cat no. AF6258), anti-CDK1 (1:100; cat no. AF6108) and anti-cyclinB1 (1:100; cat no. AF6168; all from Affinity Biosciences, Ltd.); anti-Ki67 (1:100; cat no. A20018), anti-Bcl2 (1:100; cat no. A19693) and anti-N-Cadherin (1:100; cat no. A0433; all from ABclonal Biotech Co., Ltd.).

Techniques: In Vivo, TUNEL Assay, Staining, Derivative Assay, Immunohistochemistry, Expressing

Journal: International Journal of Oncology

Article Title: Cyclovirobuxine D inhibits growth and progression of non-small cell lung cancer cells by suppressing the KIF11-CDC25C-CDK1-CyclinB1 G 2 /M phase transition regulatory network and the NFκB/JNK signaling pathway

doi: 10.3892/ijo.2023.5505

Figure Lengend Snippet: Molecular docking results of CVB-D in non-small cell lung cancer. (A-D) Molecular docking of CVB-D into the protein crystal structures of KIF11, CDC25C, CDK1 and cyclinB1. CVB-D, cyclovirobuxine D; KIF11, kinesin family member 11.

Article Snippet: Then, the slices were blocked for 30 min at 37°C with goat serum (included in the UltraSensitive™ SP IHC kit) and incubated with primary antibodies (diluted with 5% BSA) overnight at 4°C against the target proteins: anti-KIF11 (1:100; cat no. 23333-1-AP; from Proteintech Group, Inc.), anti-CDC25C (1:100; cat no. AF6258), anti-CDK1 (1:100; cat no. AF6108) and anti-cyclinB1 (1:100; cat no. AF6168; all from Affinity Biosciences, Ltd.); anti-Ki67 (1:100; cat no. A20018), anti-Bcl2 (1:100; cat no. A19693) and anti-N-Cadherin (1:100; cat no. A0433; all from ABclonal Biotech Co., Ltd.).

Techniques:

Journal: International Journal of Oncology

Article Title: Cyclovirobuxine D inhibits growth and progression of non-small cell lung cancer cells by suppressing the KIF11-CDC25C-CDK1-CyclinB1 G 2 /M phase transition regulatory network and the NFκB/JNK signaling pathway

doi: 10.3892/ijo.2023.5505

Figure Lengend Snippet: Chemical structure of CVB-D and its mechanism in the NSCLC cells. (A) The 2D structure and 3D conformation of CVB-D. (B) CVB-D inhibits NSCLC progression by suppressing activation of the NF-κB/JNK signaling pathway and the KIF11-CDK1-CDC25C-cyclinB1 network. CVB-D, cyclovirobuxine D; NSCLC, non-small cell lung cancer; KIF11, kinesin family member 11.

Article Snippet: Then, the slices were blocked for 30 min at 37°C with goat serum (included in the UltraSensitive™ SP IHC kit) and incubated with primary antibodies (diluted with 5% BSA) overnight at 4°C against the target proteins: anti-KIF11 (1:100; cat no. 23333-1-AP; from Proteintech Group, Inc.), anti-CDC25C (1:100; cat no. AF6258), anti-CDK1 (1:100; cat no. AF6108) and anti-cyclinB1 (1:100; cat no. AF6168; all from Affinity Biosciences, Ltd.); anti-Ki67 (1:100; cat no. A20018), anti-Bcl2 (1:100; cat no. A19693) and anti-N-Cadherin (1:100; cat no. A0433; all from ABclonal Biotech Co., Ltd.).

Techniques: Activation Assay