Fig 2: FBN1 knockout inhibits progression of ovarian cancer and sensitizes response to cisplatin in vivo. (A) Images of tumors generated by FBN1-knockout and control cisplatin-resistant ovarian cancer organoids with or without cisplatin and/or apatinib. (B) Growth curves of xenograft tumors in mice. (C) Average tumor weights in nude mice. (D) Representative images of PET-CT used for detection of glucose uptake. Each group contained 5 mice. (E) Average SUVmax values of nude mice bearing tumors. (F) Effect of FBN1 morpholino in zebrafish model was tested by qRT-PCR. (G) Zebrafish model treated with or without cisplatin (0.2 mmol/L) and/or apatinib (40 µmol/L). (H) VEGFA mRNA in zebrafish models. (I) Immunofluorescence of FBN1 and specific angiogenesis marker CD31 in the xenograft tumors of FBN1 knockout and control groups with cisplatin treatment. (J) qRT-PCR analysis of the indicated genes in FBN1-knockout group and the control in nude mouse tumor tissues without drug treatment. Error bars, 95% CIs. *, P < 0.05, **, P < 0.01. (K) Heatmap showing that FBN1-affected genes are involved in glycolysis and angiogenesis with cisplatin treatment. Abbreviations: FBN1, fibrillin-1; CR, cisplatin-resistant; KO, knockout; NC, negative control; OE, overexpression; VEGFA, vascular endothelial growth factor A; qRT-PCR, quantitative real-time PCR; CD31, platelet endothelial cell adhesion molecule-1

Fig 3: FBN1 knockout decreases glycolysis and angiogenesis. (A) FBN1-knockout cisplatin-resistant ovarian cancer organoids and cell lines were verified by Western blotting. (B) GSEA analysis was performed using FBN1-knockout and control cisplatin-resistant ovarian cancer organoids (CR-organoid/FBN1-KO1 and NC). The signature was defined by genes showing significant expression changes. (C) Gene ontology of mass spectrum analysis in CR-organoid/FBN1-KO1 and CR-organoid/NC. (D) Pathway examination of mass spectrum analysis in CR-organoid/FBN1-KO1 and CR-organoid/NC. (E) Metabolites in pathways of glucose metabolism. (F) mRNA levels of genes altered by FBN1 knockout in cisplatin-resistant ovarian cancer organoids and cell lines. Abbreviations: FBN1, fibrillin-1; GSEA, Gene Set Enrichment Analysis; CR, cisplatin-resistant; KO, knockout; NC, negative control

Fig 4: FBN1 expression level is associated with cisplatin-resistance in patients with ovarian cancer. (A) Images of cisplatin-resistant and -sensitive organoids derived from ovarian cancer patients with or without 5 µg/L cisplatin treatment for 21 days. (B) Cell viability assay of organoids treated with 5 µg/L cisplatin in different time intervals. The data of 4 pairs of ovarian cancer organoids are shown. Data are presented as mean ±SD of triplicate measurements repeated three times. (C) GSEA analysis was performed using 6 cisplatin-resistant and 6 -sensitive organoids derived from ovarian cancer patients. (D) Immunofluorescence assay was performed to detect the association between FBN1 and cisplatin-sensitive ovarian cancer organoids. (E) FBN1 mRNA expression in 10 pairs of cisplatin-resistant and -sensitive organoids of ovarian cancer. (F) Representative images of immunohistochemistry staining of FBN1 in 132 cisplatin-resistant and 129 -sensitive ovarian cancer tissues. (G) High-low expression ratio of FBN1 in cisplatin-resistant and -sensitive ovarian cancer tissues. (H) H-score of FBN1 in cisplatin-resistant and -sensitive ovarian cancer tissues. (I) qRT-PCR analysis of FBN1 mRNA expression in 45 pairs of cisplatin-resistant and -sensitive ovarian cancer tissues. (J) Immunofluorescence of FBN1 in cisplatin-resistant and -sensitive ovarian cancer tissues. Red signals, FBN1; blue signals, DAPI. **, P < 0.01. Abbreviations: FBN1, fibrillin-1; SD, standard deviation; GSEA, Gene Set Enrichment Analysis; qRT-PCR, quantitative real-time PCR; DAPI, 2-(4-Amidinophenyl)-6-indolecarbamidine dihydrochloride

Fig 5: Immunohistochemical staining and immunofluorescence of FBN1, p-VEGFR2, and p-STAT2. (A) Representative images of immunohistochemistry of FBN1, p-VEGFR2 (Tyr1054), and p-STAT2 (Tyr690) in ovarian cancer tissues. (B) Human serum VEGFA concentration in 50 pairs of ovarian cancer patients measured by ELISA kit. (C-D) Association between SUVmax of PET/CT technology and FBN1 expression in the lesions of adnexal carcinomas from 100 ovarian cancer patients. (E) The relationship between SUVmax of PET/CT image and overall survival of 100 ovarian cancer patients. (F) mRNA expression levels of genes associated with glycolysis and angiogenesis assessed via qRT-PCR in 45 pairs of ovarian cancer samples with high or low FBN1 expression. (G) Representative images of immunofluorescence of FBN1, p-VEGFR2 (Tyr1054), and p-STAT2 (Tyr641) in ovarian cancer patients’ tissues. Green signals, FBN1 or p-VEGFR2 (Tyr1054); red signals, p-VEGFR2 (Tyr1054) or p-STAT2 (Tyr641); blue signals, DAPI. (H) Schematic model on the proposed role of the FBN1/VEGFR2/STAT2 signaling axis in modulating glycolysis, angiogenesis, and cisplatin sensitivity. Abbreviations: FBN1, fibrillin-1; ELISA, enzyme-linked immunosorbent assay; SUVmax, maximum of standardized uptake value; PET-CT, positron emission tomography-computed tomography; VEGFA, vascular endothelial growth factor A; STAT2, signal transducer and activator of transcription 2; VEGFR2, vascular endothelial growth factor receptor 2; DAPI, 2-(4-amidinophenyl)-6-indolecarbamidine dihydrochloride