The molecular mechanism of gemcitabine in inhibiting the HIF-1α/VEGFB/FGF2/FGFR1 signaling pathway for ovarian cancer treatment

Abstract

Ovarian cancer is a common malignant tumor in women, exhibiting a certain sensitivity to chemotherapy drugs like gemcitabine (GEM). This study, through the analysis of ovarian cancer single-cell RNA sequencing (scRNA-seq) data and transcriptome data post-GEM treatment, identifies the pivotal role of hypoxia-inducible factor 1 alpha (HIF-1α) in regulating the treatment process. The results reveal that HIF-1α modulates the expression of VEGF-B, thereby inhibiting the fibroblast growth factor 2 (FGF2)/FGFR1 signaling pathway and impacting tumor formation. In vitro experiments validate the mechanistic role of HIF-1α in GEM treatment, demonstrating that overexpression of HIF-1α reverses the drug's effects on ovarian cancer cells while silencing fibroblast growth factor receptor 1 (FGFR1) can restore treatment efficacy. These findings provide essential molecular targets and a theoretical foundation for the development of novel treatment strategies for ovarian cancer in the future.

Keywords: Fibroblast growth factor 2; Fibroblast growth factor receptor 1; Gemcitabine; Hypoxia-inducible factor 1 alpha; Ovarian cancer; Vascular endothelial growth factor B.

Fig. 1

Analysis of ovarian cancer scRNA-seq data based on public databases. A Visualization of cell annotation results grouping on UMAP clustering; B Correlation heatmaps of top five genes in 13 cell types; C, D KEGG E and GO F functional enrichment plots of epithelial cell characteristic genes; E Cell communication network plot in samples, where thickness of lines in the left plot represents the number of pathways, and in the right plot represents interaction strength; F Communication plot between epithelial cells and other cells

Fig. 2

Identification of key genes in high-throughput transcriptome analysis. A Volcano plot of differential analysis, where red indicates significantly upregulated genes, blue indicates significantly downregulated genes, and grey indicates non-significant genes; B, C Enrichment analysis of significant DEGs using GO (B) and KEGG (C); D Venn diagram of differential genes between characteristic genes of epithelial cells in scRNA-seq dataset and high-throughput transcriptome data after GEM treatment; E Boxplots of expression of 18 intersecting genes in the transcriptome dataset; F Boxplot of HIF-1α expression levels in TCGA_GTEx-OV, where * indicates P ≤ 0.05 (TCGA_GTEx-OV dataset: Normal: n = 88, Tumor: n = 427); G UMAP plot of HIF-1α expression in scRNA-seq dataset

Fig. 3

Exploration of the Impact of GEM Treatment on Other Key Genes. A PPI network of genes interacting with HIF-1α among significant DEGs; B Correlation heatmap of genes interacting with HIF-1α; C Boxplots of expression levels of VEGF-B, PROX1, SOD2 (control-OVCAR3, n = 3, GEM-OVCAR3, n = 3); D–F Boxplots of expression levels of VEGF-B (D), PROX1 (E), SOD2 (F) in TCGA_GTEx-OV dataset, where * indicates P < 0.05, ** indicates P < 0.01, *** indicates P < 0.001 (TCGA_GTEx-OV dataset: Normal: n = 88, Tumor: n = 427); (G) PPI network of genes interacting with VEGF-B among significantly DEGs; (H) Boxplots of expression levels of FGF2 and FGFR1 (control-OVCAR3, n = 3, GEM-OVCAR3, n = 3)

Fig. 4

Validation of the upstream–downstream relationships of HIF-1α, VEGF-B, and FGF2/FGFR1. A Overexpression of HIF-1α followed by qPCR A and Western blot analysis B to assess the expression levels of HIF-1α, VEGF-B, FGF2, and FGFR1; C Overexpression of VEGF-B followed by qPCR C and Western blot analysis D to evaluate the expression levels of HIF-1α, VEGF-B, FGF2, and FGFR1; E Overexpression of FGFR1 followed by qPCR E and Western blot analysis F to determine the expression levels of HIF-1α, VEGF-B, FGF2, and FGFR1. Cell experiments were conducted in triplicate. *P < 0.05, **P < 0.01, ***P < 0.001

Fig. 5

Exploring the molecular mechanisms of Dioscorea nipponica Makino extract on inhibiting the proliferation, migration, invasion capabilities, and angiogenesis of ovarian cancer cells. A, B qPCR and WB analysis of HIF-1α, VEGF-B, FGF2, and FGFR1 expression levels in ISsh-80, SK-OV-3, and SK-OV-3/GEM cells; C, D qPCR and WB analysis of HIF-1α, VEGF-B, FGF2, and FGFR1 expression levels in the control (SK-OV-3), GEM, oe-NC + GEM, oe-HIF-1α + GEM, and oe-HIF-1α + sh-FGFR1 + GEM groups; E CCK-8 assay measuring the differences in proliferation across groups; F Scratch assay measuring the differences in migration across groups; G Transwell assay measuring the differences in invasion across groups; H Angiogenesis assay measuring the formation of tubular structures across groups. All experiments were repeated three times. Cell experiments were repeated three times. **P < 0.01, ***P < 0.001

Fig. 6

Molecular mechanisms of GEM treatment in breast cancer