Sunitinib Synergizes with Cisplatin by Suppressing DNA Repair Pathways in High-Grade Serous Ovarian Carcinoma Cells

Sunitinib Synergizes with Cisplatin by Suppressing DNA Repair Pathways in High-Grade Serous Ovarian Carcinoma Cells | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Sunitinib Synergizes with Cisplatin by Suppressing DNA Repair Pathways in High-Grade Serous Ovarian Carcinoma Cells Yeernaer Hazaisihan, Yankun Yu, Li Wei, Xiaoning Li, Di Zhou, and 5 more This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-8439132/v1 This work is licensed under a CC BY 4.0 License Status: Under Revision Version 1 posted 9 You are reading this latest preprint version Abstract Background: High-grade serous ovarian carcinoma (HGSOC) frequently develops resistance to platinum-based chemotherapy such as cisplatin, leading to high mortality. Sunitinib, a multi-target Receptor tyrosine kinase inhibitor, has shown potential in the treatment of ovarian carcinoma (OC). However, the synergistic effects between cisplatin and sunitinib, and the sensitization mechanism of sunitinib in HGSOC are not yet understood. Methods: The anti-tumor effects of sunitinib on HGSOC cells were assessed using CCK-8, EdU, colony formation, wound healing, Transwell, and flow cytometry assays. RNA sequencing was performed on sunitinib-treated cells, followed by differential expression, enrichment, and protein-protein interaction network (PPI) analyses. Genes on cisplatin sensitivity were predicted using the CellMiner and GEPIA3 databases. Synergy between sunitinib and cisplatin was evaluated using SynergyFinder 3.0, and DNA damage was assessed by γ-H2AX expression. Results: Sunitinib significantly inhibited proliferation, migration, and invasion in HGSOC cells, while further inducing cell cycle arrest, promoting necrosis. Besides, sunitinib downregulated critical DNA damage repair pathways, including homologous recombination, fanconi anemia, and base excision repair. Furthermore, sunitinib synergizes with cisplatin in HGSOC cells, enhancing DNA damage compared to monotherapy. Additionally, we screened out 25 sunitinib-downregulated core genes (SDCs). Drug-sensitivity analyses showed that higher SDCs expression was significantly associated with cisplatin resistance in OC. Notably, in cisplatin-resistant HEY-A8/DDP cells, sunitinib displayed stronger cytotoxicity than cisplatin. Conclusion: Sunitinib induces cell-cycle arrest and necrosis. In addition, sunitinib synergized with cisplatin and enhanced cisplatin sensitivity by impairing DNA repair pathways. Drug-sensitivity analyses showed that SDCs are associated with cisplatin resistance in OC, suggesting that sunitinib may help overcome cisplatin chemoresistance. Notably, sunitinib retains substantial cytotoxic activity in cisplatin-resistant cells. Together, these findings suggest that the sunitinib–cisplatin combination is a promising strategy to overcome cisplatin resistance in HGSOC. HGSOC sunitinib cisplatin DNA repair combinatory therapy Figures Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 Backgrounds Ovarian carcinoma (OC) is a highly invasive malignant tumor of the female reproductive system. In China, it was estimated that 61,100 new ovarian carcinoma cases and 32,600 attributable deaths were reported in 2022 [1] . Although early-stage ovarian carcinoma has high treatment efficacy, the lack of effective screening methods means approximately 75% of patients are diagnosed at an advanced stage, resulting in a 5-year survival rate of less than 30% [2] . High-grade serous ovarian carcinoma (HGSOC) is the most common subtype of OC, which is classified as a subtype of epithelial ovarian carcinoma (EOC). The current standard treatment for EOC, primary tumor debulking surgery is the preferred option. For advanced EOC, neoadjuvant chemotherapy followed by interval tumor debulking surgery serves as an alternative for patients unsuitable for immediate surgery [2,3] . Despite the initial efficacy of platinum-based chemotherapy reaching up to 70%, most advanced-stage patients eventually relapse because of chemoresistance—with a cumulative relapse rate of 70% within three years [4,5] . To address this challenge, PARP inhibitors (PARPi) have shown promising benefits for BRCA-mutated and HR-deficient tumors [6,7] , but about 50% of HGSOC patients lack HR deficiency and gain limited benefit [8] . Besides, most patients with BRCA1/2-mutated tumors initially respond to PARP inhibitors but eventually develop resistance, resulting in relapse and disease progression [9] . Immune checkpoint inhibitors (ICIs), even though promising in other cancers, show low efficacy in platinum-resistant EOC due to limited tumor immunogenicity and overlapping resistance mechanisms [10,11] . These realities highlight the necessity of elucidating the molecular mechanisms of HGSOC and developing new therapeutic strategy. Sunitinib is an orally bioavailable, multi-targeted receptor tyrosine kinase (RTK) inhibitor approved by Food and Drug Administration (FDA) for adjuvant therapy of high-risk renal cell carcinoma following nephrectomy, as well as for gastrointestinal stromal tumors refractory to or intolerant of imatinib [12,13] . It selectively inhibits a variety of RTKs implicated in tumor angiogenesis and growth, including VEGFR-1/2, PDGFR-α/β, c-KIT, RET, and FLT3 [14,15] . By inhibiting the key RTKs, sunitinib suppresses angiogenic signaling, tumor progression, and metastasis, making it a promising agent in cancer therapy. In OC, the randomized AGO-OVAR 2.11 phase II trial demonstrated that sunitinib was feasible and moderately effective in patients with recurrent platinum-resistant OC [16] . Another phase II study evaluating sunitinib in patients with clear cell ovarian cancer demonstrated that the drug exhibits limited activity [17] . Although sunitinib has been clinically studied in OC, most studies have focused on its monotherapy, with limited exploration of its combination with cisplatin. Interestingly, in an Ehrlich ascites carcinoma model, sunitinib significantly enhanced the efficacy of cisplatin while attenuating cisplatin-induced nephrotoxicity in rats [18] , highlighting its potential as a cisplatin-sensitizing agent. Therefore, we hypothesized that sunitinib may have a similar effect on HGSOC cells, enhancing the sensitivity of cisplatin. In this study, we aim to investigate the synergistic effects of sunitinib and cisplatin, providing a basis for improving the treatment efficacy of HGSOC and overcoming cisplatin resistance via combination therapy. We further sought to predict the putative mechanisms by which sunitinib acts, with a particular focus on pathways that may sensitize HGSOC cells to cisplatin. Materials and methods Materials Sunitinib was provided by Sunitinib was provided by Liang Guo, Shihezi University. Cell culture HGSOC cell lines (HEY-A8, SKOV3) were purchased from Shanghai EK-Bioscience Biotechnology Co, Ltd. The cisplatin-resistant HEY-A8/DDP cell line was established by exposing parental HEY-A8 cells to escalating concentrations of cisplatin, followed by repeated selection and subculturing to achieve stable cisplatin resistance. HEY-A8 and HEY-A8/DDP cells were cultured in Dulbecco’s Modified Eagle Medium (Gibco, Thermo Fisher Scientific, USA) supplemented with 10% FBS (Gibco, Thermo Fisher Scientific, USA), 100 U/mL penicillin and 100 mg/mL streptomycin (Solarbio, China) in humidified air at 37 ℃with 5% CO2. SKOV3 were cultured in McCoy's 5A (PM150710, Wuhan, China) supplemented with 10% FBS, 100 U/mL penicillin and 100 mg/mL streptomycin in humidified air at 37 ℃ with 5% CO2. EdU staining assay In a 24-well plate, 1×10 5 cells in the logarithmic growth phase were inoculated and cultured to the normal growth stage. Cells were then treated with different concentrations of sunitinib for 24 hours. After treatment, Edu working solution (C0071S, beyotime, China) was added and incubated for 2 hours. The cells were fixed with 4% paraformaldehyde, permeabilized with 0.5% Triton X-100, and subjected to a Click reaction with fluorescent dye-conjugated azide for 30 minutes in the dark. Nuclei were counterstained with DAPI, and Edu-positive cells were visualized and quantified using fluorescence microscopy. Untreated and positive control groups were included for validation. Cell counting kit-8 assay Cells were seeded in 96-well plates at a density of 2,000 cells per well for proliferation assays and 5,000 cells per well for inhibition assays, followed by incubation at 37°C with 5% CO₂ for 6 h to allow cell attachment. After removing the culture medium, 100 µL of fresh culture medium containing varying concentrations of the drug was added to each well, and the cells were incubated for 24 hours. Following this, the medium in each well was replaced with 10% CCK-8 reagent (BS350B, Biosharp, China) in complete medium, and the cells were incubated for an additional 2 hours in the dark at 37°C with 5% CO₂. Absorbance was then measured at 450 nm using a microplate reader. Colony formation assay 1500 cells were evenly seeded in a six-well plate. After 24 hours of cell attachment, the cells were treated with varying concentrations of sunitinib for 48 hours. Subsequently, the medium containing sunitinib was removed, and the cells were maintained in normal culture conditions for 14 days. The colonies were then fixed with 4% paraformaldehyde, stained with 1% crystal violet solution, washed three times with distilled water, air-dried, and photographed. Flow cytometry assay for the cell cycle Cells were seeded in 6-well plates and allowed to grow until they reached approximately 70% confluence. The culture medium was then replaced with fresh medium containing various concentrations of sunitinib, and the cells were treated for 24 hours. After treatment, the cells were harvested by trypsinization, washed with PBS, and fixed in absolute ethanol for one week. The fixed cells were subsequently stained with 1 mL of DNA staining solution (MULTI SCIENCE, CCS012) at room temperature for 30 minutes in the dark. Cell cycle distribution was analyzed using a flow cytometer, and the data were processed to determine the percentages of cells in the G0/G1, S, and G2/M phases, as well as the proportion of apoptotic cells (sub-G1 peak). Annexin-V/PI assay for apoptosis and necrosis The YF®647A-Annexin V and PI Apoptosis Kit was purchased from PROTEINBIO (Nanjing, China). Cells were seeded into 6-well plates and cultured until reaching approximately 70% confluence. Then, the culture medium was replaced with fresh medium containing various concentrations of sunitinib, and the cells were incubated for 24 hours. Subsequently, the cells were digested, resuspended in 100 µL of 1× binding buffer, and stained with 5 µL of YF647A-Annexin V and 5 µL of PI. The samples were incubated at room temperature in the dark for 15 minutes. Thereafter, 400 µl of Annexin V binding buffer was added, the samples were mixed thoroughly, and flow cytometric analysis was performed. YF647A-Annexin V was excited at 647 nm and detected in the APC channel, while PI was excited at 617 nm. Western Blot Analysis Western Blot Analysis Cells were lysed in RIPA buffer (R0010, Solarbio, China) with PMSF and incubated on ice for 20 minutes. After centrifugation at 12,000 rpm for 20 min at 4°C, protein concentrations were determined. Protein samples were diluted to 1× with 5× SDS-PAGE Loading buffer (Beyotime, China), and equal amounts were loaded onto an SDS-PAGE gel for electrophoresis, followed by transfer to 0.45 µm PVDF membranes (Millipore, Merck KGaA, Germany). The membranes were blocked with 5% BSA and incubated with primary antibodies overnight at 4°C. Primary antibodies used were β-actin (1:3000, TA-09, ZSGB-BIO, China) and γ-H2AX (1:1000, sc-517348, Santa Cruz, USA). After washing, the membranes were incubated with a horseradish peroxidase-conjugated secondary antibody (1:16,000, BS1247, Bioworld, USA) and protein bands were visualized using SuperSignal™ West Pico PLUS chemiluminescent substrate (34580, Thermo Fisher, USA). Transwell assay The invasion assay was performed using Matrigel-coated Transwell chambers (LABSELECT, China). Matrigel (Corning, USA) and culture medium were mixed at an 8:1 ratio to create the coating solution, which was then added to the upper chambers of the Transwell system. The cells were treated with different concentrations of sunitinib for 48 hours before plating in the Transwell chambers. For each well, 10,000 cells were seeded. The upper chamber was supplemented with 600 µL of 20% serum-containing medium, while the lower chamber contained 200 µL of serum-free medium with 10,000 cells. The cells were allowed to migrate for 24 hours and invade for 48 hours. After the incubation period, the cells that had invaded the lower chamber were fixed and stained for analysis. Wound healing assay After plating the cells in the 6-well culture plates, a scratch was made across the cell monolayer using a sterile pipette tip when the cells reached the appropriate confluence. The cells were then treated with different concentrations of serum-free sunitinib. Images of the wound area were captured at 0 and 24 hours then the gap width was measured at both time points to evaluate the cell migration capacity. Statistical analysis Statistical analyses were performed using GraphPad Prism 9.5 software. Two-group comparisons were conducted using two-tailed Student’s t -tests, and comparisons among multiple groups were performed using one-way ANOVA. Statistical significance was set at p < 0.05. Results Sunitinib suppresses proliferation, migration, and invasion in HGSOC cells To evaluate the inhibitory effects of sunitinib on HGSOC cell proliferation, we performed CCK-8 assays to determine the IC50 values in HEY-A8 and SKOV3 cells after 24 h of treatment. The results showed that the IC 50 values of sunitinib in HEY-A8 and SKOV3 cells were 11.83 µM and 17.31 µM, respectively (Fig. 1 A-C). Colony formation assays showed that sunitinib suppresses the colony formation ability of HEY-A8 and SKOV3 cells (Fig. 1 D). EdU incorporation assays demonstrated that sunitinib inhibited proliferation of HEY-A8 and SKOV3 cells in a dose-dependent manner (Fig. 1 E-G). In addition, wound-healing and Transwell assays revealed significant suppression of both migration and invasion following sunitinib treatment in HEY-A8 and SKOV3 cells (Fig. 1 H and Supplementary Fig. 1A-C). These data indicate that sunitinib exerts strong anti-proliferative and anti-metastatic activity in HGSOC cells. Sunitinib induces cell cycle arrest and promotes necrosis in HGSOC cells To investigate how sunitinib suppresses growth and induces cytotoxicity in HGSOC, we exposed HEY-A8 and SKOV3 cells to increasing concentrations of sunitinib for 24 h and quantified cell-cycle distribution and apoptosis. Flow-cytometric analysis showed that sunitinib induced distinct cell-cycle alterations in the two HGSOC cell lines. In HEY-A8 cells, sunitinib treatment primarily caused a G0/G1-phase arrest accompanied by a marked reduction in the S-phase population. Unlike HEY-A8, SKOV3 cells exhibited a pronounced G2/M-phase arrest, which was similarly associated with a substantial decrease in S-phase fraction. (Fig. 2 A). Subsequently, we assessed cell death using flow cytometry. After sunitinib treatment, most cells accumulated in the Annexin V−/PI+ quadrant (Q1), suggesting that sunitinib primarily exerts its cytotoxic effects by inducing necrosis in HEY-A8 and SKOV3 cells (Fig. 2 B-C). In addition, sunitinib induced apoptosis in SKOV3 cells but not in HEY-A8 cells (Fig. 2 D). In brief, sunitinib induced cell cycle arrest and caused cell death predominantly through necrosis in HGSOC cells, while also eliciting a modest apoptotic response. Sunitinib downregulates DNA Damage Repair Pathways To elucidate the molecular mechanisms underlying sunitinib’s effects on HGSOC cells, HEY-A8 cells were treated with sunitinib and subjected to differential expression analysis (Fig. 3 A; Supplementary Tables 1 and 2). There were relatively few enriched pathways among the upregulated genes (Supplementary Fig. 2), while downregulated genes exhibit significant enrichment in several key biological processes, including cell-cycle regulation, DNA replication, and DNA damage repair (Fig. 3 B–D). Most notably, DNA repair mechanism is well recognized as central determinants of platinum responsiveness in HGSOC [19,20] . Gene Ontology (GO) enrichment analysis indicated that sunitinib downregulate DNA metabolic processes, DNA repair, and DNA damage response (DDR), with a particular emphasis on double-strand break repair via homologous recombination (HR) (Fig. 3 B). Consistently, KEGG pathway analysis revealed that sunitinib significantly suppresses key DNA damage repair pathways, including homologous recombination (HR), the Fanconi anemia (FA), base excision repair (BER), nucleotide excision repair (NER), and mismatch repair (MMR) (Fig. 3 C). These pathways are essential for the repair of DNA crosslinks and double-strand breaks induced by cisplatin and other platinum agents [21–23] . Reactome pathway analysis further underscored the downregulation of cell-cycle checkpoints, S-phase progression, and DNA replication (Fig. 3 D).Additionally, GSEA demonstrated that sunitinib downregulates the pathways: DNA replication, cell cycle progression, and DNA repair—HR and BER (Fig. 3 E). It is well established that increased DNA repair capacity is a key mechanism of cisplatin resistance in OC. Among these pathways, HR is now firmly established as a key determinant of platinum responsiveness in HGSOC [24] . Tumors with HR deficiency (HRD), including those harboring BRCA1/2 or other HRR gene mutations, exhibit higher platinum sensitivity and prolonged progression-free survival compared with HR-proficient tumors, whereas restoration of HR function is a hallmark of acquired platinum resistance [25] . BER has also been implicated in cisplatin-resistant phenotypes, serving as a key mechanism for repairing cisplatin-induced DNA crosslinks [26] . The central BER endonuclease APE1 is frequently overexpressed in ovarian tumors, and its high expression correlates with poor prognosis and chemoresistance [27,28] . Likewise, disruption of XRCC1 increases cisplatin-induced DNA damage and sensitizes cells to cisplatin, supporting the concept that attenuation of BER shifts the balance from damage tolerance toward cell death under platinum treatment [29] . The FA pathway, which orchestrates repair of DNA interstrand crosslinks during S phase and functionally intersects with HR, is also tightly linked to cisplatin sensitivity [30] . Activation of FA signaling through factors such as And-1 and the FANCM/FAAP24 complex promotes resolution of cisplatin-induced crosslinks and contributes to the resistant phenotype in OC cells, whereas suppression of FA components, such as FANCD2—impairs crosslink repair and re-sensitizes tumors to cisplatin in vitro and in vivo [31,32] . Taken together, previous evidence indicates that upregulation or rewiring of DNA repair programs is a central route to cisplatin resistance [33] . In this study, our data support sunitinib-mediated suppression of DNA repair pathways such as HR, BER and FA pathways in HGSOC cells. Therefore, our transcriptomic findings suggest that sunitinib functionally weakens the DDR, limiting the ability of HGSOC cells to repair cisplatin-induced DNA lesions. This provides a strong mechanistic rationale for combining sunitinib with cisplatin to overcome platinum resistance in HGSOC. Sunitinib Synergizes with Cisplatin in HGSOC Cells Considering cisplatin exerts its antitumor effects by inducing DNA crosslinks and DNA damage [34] , while sunitinib impairs DNA repair pathways, we next assessed their synergism. To assess the potential synergy, we used SynergyFinder 3.0 [35] to analyze the combination of sunitinib and cisplatin in HEY-A8 and SKOV3 cells. The computed Zero Interaction Potency (ZIP) score exceeded 10 in HEY-A8 and SKOV3 cells, indicating a strong synergistic effect between sunitinib and cisplatin in HGSOC cells. (Fig. 4 A-D). To investigate the effect of sunitinib on DNA damage, the level of DNA damage marker γ-H2AX [36] was assed using western blot assay. Our data indicate that, treatment with cisplatin and sunitinib, either alone or in combination, led to increased γ-H2AX expression in both HEY-A8 and SKOV3 cells. Most notably, the combined treatment with sunitinib and cisplatin induced the most significant increase in γ-H2AX levels, suggesting that the combination of these agents induces a higher extent of DNA damage compared to either treatment alone (Fig. 4 E-F). These results suggest that sunitinib can induce DNA damage in HGSOC cells. Notably, when combined with cisplatin, sunitinib enhances the DNA damage response, resulting in a more substantial DNA damage effect than cisplatin alone. Collectively, sunitinib exhibits synergistic effects with cisplatin in HGSOC cells by downregulating DNA damage repair pathways and enhancing the sensitivity of HGSOC cells to cisplatin. Sunitinib-regulated Core Genes are associated with prognosis in OC To further prioritize key mediators within these sunitinib-suppressed networks, we analyzed protein–protein interaction (PPI) using the STRING database [37] and Cytoscape [38] . We then identified 25 downregulated core genes from the PPI network (Fig. 5 A–C) and defined them as the sunitinib-downregulated core genes (SDCs) (Fig. 5 D). Subsequently, we performed correlation analysis using TNMplot [39] , which revealed significant positive correlations among the expression levels of these genes in ovarian serous cystadenocarcinoma (Fig. 5 D). We further analyzed the combined signature of SDCs via the GEPIA3 database [40] , and found that it was significantly higher in OC than in normal ovarian tissues, suggesting that these genes are overexpressed in OC (Fig. 5 E). Consistently, analysis using TNMplot confirmed that these genes were expressed at significantly higher levels in ovarian serous cystadenocarcinoma than in normal ovarian tissues. (Fig. 5 F). The prognostic significance of SDCs was further evaluated using the Kaplan–Meier Plotter by analyzing overall survival (OS) and progression-free survival (PFS). Most SDCs were associated with shorter OS and PFS in patients with OC (Fig. 5 G), whereas a few genes were linked to improved outcomes. We further constructed a combined signature model on the Kaplan–Meier Plotter platform using the average expression of SDCs to predict OS and PFS. Higher SDCs signature expression was significantly associated with shorter OS, while no statistically significant difference was observed for PFS (Fig. 5 H). Collectively, these findings suggest that sunitinib-mediated suppression of SDCs is associated with improved OS in OC patients, implying that sunitinib may prolong survival by downregulating SDCs. In addition, we applied the same workflow to sunitinib-upregulated genes and identified six sunitinib-upregulated core genes (SUCs) (Supplementary Fig. 3A-B). Unexpectedly, the SUCs signature was also expressed at higher levels in OC than in normal ovarian tissues (Supplementary Fig. 3C-D). However, Kaplan–Meier Plotter analysis indicated that higher SUCs signature expression was significantly associated with prolonged OS and PFS in OC patients (Supplementary Fig. 3E-F). These results suggest that SUCs are linked to a favorable prognosis in OC, and that sunitinib may improve patient outcomes by upregulating these genes. In summary, high expression of the SDCs signature is associated with poor prognosis in OC patients, while low expression of the SUCs signature correlates with poor prognosis as well. These findings suggest that sunitinib may improve the survival rate of OC patients by modulating the expression of these genes. Correlation between SDCs and Cisplatin Sensitivity in OC Next, we evaluated the association of SDCs and SUCs with cisplatin sensitivity. Using the CellMiner database [41] , we found that SDCs expression was positively correlated with cisplatinIC 50 , suggesting that higher expression of these genes is associated with increased resistance to cisplatin chemotherapy (Fig. 6 A and Supplementary Fig. 4). Subsequently, we used GEPIA 3 database to explore the effect of these genes on cisplatin chemotherapy. Our results showed that, in OC patients with high SDCs signature expression, cisplatin treatment did not improve OS, whereas in patients with low SDCs signature expression, cisplatin treatment significantly prolonged OS (Fig. 5 B). These findings suggest that high SDCs signature expression is associated with resistance to cisplatin-based chemotherapy. We further analyzed individual genes of SDCs and found that high expression of nine genes—CDT1, KIF11, BUB1B, MCM7, CDC45, MCM5, FEN1, PLK1, and RAD54L—was significantly correlated with cisplatin resistance in OC patients (Fig. 6 C-K). Additionally, we observed that only one of one gene was correlated with cisplatin sensitivity in SDCs, while the remaining genes showed no significant correlation with cisplatin chemotherapy outcomes (Supplementary Fig. 5). Besides, we assessed the cisplatin sensitivity of the SUCs by the same methods. Notably, our analysis in the CellMiner Database revealed that 3 of these genes were associated with cisplatin resistance, while 2 were linked to cisplatin sensitivity. (Supplementary Fig. 6A). Meanwhile, our survival analysis revealed that no significant association was observed between SDCs and OS in OC patients (Supplementary Fig. 6B-C). Taken together, our data demonstrate that the genes downregulated by sunitinib are associated with cisplatin resistance in OC. This observation supports the notion that sunitinib may reverse cisplatin resistance in HGSOC by downregulate these genes. Cytotoxic effects of sunitinib in cisplatin-resistant HGSOC cells Based on the results of our previous drug sensitivity analysis, we further assessed the cytotoxicity of sunitinib in cisplatin-resistant HEY-A8/DDP cells. Our data showed that the IC₅₀ of cisplatin were 15.81 µM in HEY-A8 cells and 33.42 µM in HEY-A8/DDP cells, corresponding to a resistance index (RI) of 2.11. In contrast, the IC₅₀ of sunitinib were 12.00 µM in HEY-A8 cells and 17.74 µM in HEY-A8/DDP cells, while the RI was 1.48 (Fig. 7 A-B). The results show that HEY-A8/DDP cells also respond less to sunitinib, although this resistance is not as strong as their resistance to cisplatin. Notably, in HEY-A8/DDP cells, the IC₅₀ of sunitinib (17.74 µM) was lower than cisplatin (33.42 µM), suggesting that sunitinib retains substantial cytotoxic efficacy and is less affected by the cisplatin-resistant phenotype compared with cisplatin. Given these findings, we assume that utilization of sunitinib may hold promise for reducing cisplatin dose and reverse cisplatin-resistance in HGSOC. Discussion HGSOC is one of the lethal gynecological cancers, with the major challenge being the development of resistance to platinum-based chemotherapy, particularly cisplatin [42] . Although most patients with HGSOC initially responded to platinum-based chemotherapy, the majority of individuals with advanced disease ultimately succumb to relapse driven by acquired drug resistance [43] . A key mechanism of drug resistance in HGSOC is the activation of DDR pathway, which repairs DNA damage induced by chemotherapeutic agents such as cisplatin [44] . In our study, our research demonstrates that sunitinib potentially inhibit DDR pathways to enhance cisplatin sensitivity. Our findings demonstrate that sunitinib treatment induces reduction in S phase in HGSOC cells. Reactome pathway analysis further supports these observations, showing that sunitinib downregulates S phase progression and the G1/S transition (Fig. 3 D). Cisplatin treatment leads to the formation of DNA lesions that, during DNA replication, result in double-strand breaks (DSBs) [45] . Among the various DNA repair pathways, HR plays a critical role in repairing DSBs with high fidelity by using a sister chromatid as a template [33] . Importantly, HR-mediated repair is most active during the S phase of the cell cycle, when sister chromatids are available, facilitating accurate repair of replication-associated DNA damage [46] . Therefore, we assume that sunitinib inhibits the G1/S transition, restricting cell entry into S phase and decreasing the proportion of cells undergoing active DNA replication, ultimately sensitizing HGSOC cells to cisplatin. Furthermore, pathway analysis reveals that sunitinib downregulates several DNA repair pathways, including HR, BER and FA pathway. Collectively, these results point to impaired DNA damage repair and a consequent increase in unrepaired lesions in HGSOC cells, providing a mechanistic rationale for combining sunitinib with cisplatin to improve treatment response. Meanwhile, our study shows that sunitinib and cisplatin act synergistically in HGSOC cells, leading to more pronounced DNA damage than either agent alone. The synergistic effect suggests that sunitinib may overcome cisplatin resistance by increasing DNA damage, offering a promising therapeutic approach for platinum-resistant HGSOC. What’s more, in HEY-A8/DDP cells, sunitinib shows a lower IC₅₀ and a lower resistance index than cisplatin, indicating limited cross-resistance in this cisplatin-resistant setting. These results indicate that sunitinib remains more active than cisplatin in cisplatin-resistant HGSOC cells, supporting its potential to help overcome cisplatin resistance. Mechanistically, we analyzed SDCs, which are associated with poor prognosis and cisplatin resistance. Among these SDCs, CDC45 and MCM2-7 are part of the Cdc45-MCM-GINS (CMG) helicase complex, which is essential for DNA replication and for coping with cisplatin-induced replication fork stalling. [47] . High expression of CMG components facilitates DNA replication bypass of cisplatin-damaged DNA, reducing cisplatin's cytotoxicity [48–51] . BUB1B, BUB1, TTK, and MAD2L1 are key components of the spindle assembly checkpoint, promote cisplatin resistance. Elevated BUB1B expression upregulates the RAD51 repair pathway, allowing tumor cells to evade cisplatin-induced apoptosis, while inhibiting BUB1B restores cisplatin sensitivity [52–55] . FEN1, an endonuclease in the BER pathway, enhances cisplatin resistance by promoting the repair of cisplatin-induced DNA lesions, while inhibiting FEN1 sensitizes tumor cells to cisplatin [56] . Besides, HR regulators such as RAD54L and BRCA1 facilitate RAD51 loading, and inhibition of HR—through targeting BRCA1 can sensitize tumors to cisplatin [58] . Moreover, suppressing EXO1, a HR-associated nuclease, enhances cisplatin sensitivity in OC [59] . KIF11 interacts with CaSR to upregulate BRCA1 and cyclin B1, contributing to cisplatin resistance. Inhibiting KIF11 reduces these proteins and restores cisplatin sensitivity in resistant cancers [60,61] . PLK1 signaling promotes chemoresistance, and inhibiting PLK1 with B4 enhances cisplatin efficacy in resistant tumors [62] . These results highlight the critical role of SDCs in cisplatin resistance and suggest that inhibiting them by sunitinib can restore cisplatin sensitivity. Moreover, the combination of sunitinib and cisplatin shows promising potential in overcoming resistance in HGSOC. Sunitinib enhances the DNA damage induced by cisplatin, leading to a stronger cytotoxic response. Importantly, sunitinib demonstrates efficacy in cisplatin-resistant HGSOC cells, with a smaller resistance shift compared to cisplatin alone. These findings support the use of sunitinib as a sensitizing agent to improve cisplatin-based therapies in resistant HGSOC, offering a potential strategy to overcome cisplatin-resistance and improve patient outcomes. Thus, the crosstalk among DNA damage repair, necrosis, and cisplatin resistance, as well as in vivo validation, demands further investigation. Conclusion In summary, our study highlights the synergistic of sunitinib and cisplatin in HGSOC cells. Sunitinib enhances cisplatin sensitivity by impairing key DNA damage repair pathways, inducing necrosis and causing cell cycle arrest. Drug sensitivity analysis also demonstrates that sunitinib-downregulated genes are associated with cisplatin chemoresistance. Notably, sunitinib exhibits higher cytotoxicity than cisplatin alone in HEY-A8/DDP cells. The combination of sunitinib with cisplatin has the potential to overcome platinum resistance, offering a promising therapeutic strategy for HGSOC patients, particularly those with platinum-resistant disease. Abbreviations Full form Abbreviation High-grade serous ovarian carcinoma HGSOC Ovarian carcinoma OC Epithelial ovarian carcinoma EOC Receptor tyrosine kinase RTK Vascular endothelial growth factor receptor VEGFR Platelet-derived growth factor receptor PDGFR Food and Drug Administration FDA PARP inhibitors PARPi Immune checkpoint inhibitors ICIs Cell counting kit-8 CCK-8 5-Ethynyl-2'-deoxyuridine EdU Phosphate-buffered saline PBS Sodium dodecyl sulfate-polyacrylamide gel electrophoresis SDS-PAGE Polyvinylidene fluoride PVDF Bovine serum albumin BSA Horseradish peroxidase HRP Protein-protein interaction PPI Gene Ontology GO Kyoto Encyclopedia of Genes and Genomes KEGG DNA damage response DDR Homologous recombination HR Fanconi anemia FA Base excision repair BER Nucleotide excision repair NER Mismatch repair MMR Gene set enrichment analysis GSEA Phosphorylated histone H2AX γ-H2AX Overall survival OS Progression-free survival PFS Sunitinib-downregulated core genes SDCs Sunitinib-upregulated core genes SUCs Homologous recombination deficiency HRD Apurinic/apyrimidinic endonuclease 1 APE1 Cdc45-MCM-GINS CMG Spindle assembly checkpoint SAC Resistance index RI National Cancer Institute NCI Cancer Genome Atlas TCGA National Center for Advancing Translational Sciences NCATS Chromatin licensing and DNA replication factor 1 CDT1 Kinesin family member 11 KIF11 BUB1 mitotic checkpoint serine/threonine kinase B BUB1B Minichromosome maintenance complex component 7 MCM7 Cell division cycle 45 CDC45 Minichromosome maintenance complex component 5 MCM5 Flap structure-specific endonuclease 1 FEN1 Polo-like kinase 1 PLK1 RAD54 like DNA repair and recombination protein RAD54L Declarations Ethics approval and consent to participate Not applicable. Consent for publication Not applicable. Competing interests The authors declare no competing interests. Clinical trial number Not applicable. Funding This work was supported by Science and Technology Program of XPCC (No. 2025ZD010, No. 2023CB008-02); the National Natural Science Foundation of China(No. 82360494, No. 82460492); the International Science and Technology Cooperation Promotion Plan of Shihezi University(No. GJHZ202301); the First Affiliated Hospital of Shihezi University Science and Technology Project(No. LC2025002); the Independent Project of Shihezi University(No. ZZZC2023046, No. ZZZC2023051); the Natural Science Foundation of Xinjiang Production and Construction Corps (No. 2025DA061); the Major Project of Xinjiang Uygur Autonomous Region (No. 2024A03004-4). Authors' contributions Conception and design: WJ, YKY, LG. Methodology design: YH, YKY, LW, XNL, XTG, LT. Provision of experimental platforms and materials: LG, WJ, SJW. Experimental implementation: YH, YKY, DZ. Manuscript writing: YH. Review & editing: WJ. Data Availability The datasets generated and analysed during the current study are available in the NCBI Gene Expression Omnibus (GEO) repository, https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE318963 (accession number: GSE318963). Acknowledgements We sincerely thank the Pathological Diagnosis Clinical Medical Research Center of Xinjiang Production and Construction Corps for their assistance. References Han B, Zheng R, Zeng H, Wang S, Sun K, Chen R, et al. Cancer incidence and mortality in China, 2022. J Natl Cancer Cent 2024;4(1):47–53. Zhang GN. [Pay attention to the selection and implementation of initial treatment for patients with advanced epithelial ovarian cancer]. Zhonghua Fu Chan Ke Za Zhi 2021;56(6):380–4. Siminiak N, Czepczyński R, Zaborowski MP, Iżycki D. Immunotherapy in Ovarian Cancer. Arch Immunol Ther Exp (Warsz) 2022;70(1):19. Li H, Sheng JJ, Zheng SA, Liu PW, Wu N, Zeng WJ, et al. 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Supplementary Files SupplementaryFigure1.jpg Supplementary Figure 1. Sunitinib inhibits proliferation, migration, and invasion of SKOV3 cells. (A-B) Transwell migration and Matrigel invasion assays indicate reduced migratory and invasive capacities of HGSOC cells following sunitinib treatment. (C) Wound-healing assays reveal impaired SKOV3 cell migration in the presence of sunitinib. (The notation “ns” signifies no significance; *p<0.05; **p<0.01; *** p<0.001; ****p<0.0001) SupplementaryFigure2.jpg Supplementary Figure 2. Enriched Pathways in Sunitinib-Upregulated Gene Set. (A) GO enrichment analysis (BP, CC, and MF) performed using TNMplot. (B) KEGG pathway enrichment analysis performed using TNMplot. (D) Reactome pathway enrichment analysis. SupplementaryFigure3.jpg Supplementary Figure 3. The OS and PFS of SDCs. (A-Y) The prediction of OS and PFS of SDCs via Kaplan-Meier database. The genes in sequence are: FANCD2, FANCI, RFC3, MCM10, CDK1, PCNA, BUB1B, MAD2L1, BUB1, CCNA2, BRCA1, KIF11, CDT1, RAD54L, MCM5, MCM4, TTK, MCM6, CDC45, EXO1, MCM7, PLK1, FEN1, MCM3 and CDC6. SupplementaryFigure4.jpg Supplementary Figure 4. Identification and clinical relevance of SDCs in ovarian carcinoma.(A) PPI network constructed from genes involved in the top 10 enriched GO terms (BP, CC, and MF) of the sunitinib-downregulated gene set. (B) The six hub genes with the highest interaction degree identified from the PPI network (defined as SDCs), and the correlation analysis of SDCs using the TNMplot database. (C) Differential expression analysis of SDCs in ovarian carcinoma predicted using the TNMplot database. (D) Differential expression analysis of the SDCs signature in ovarian carcinoma using the GEPIA3 database, showing higher expression in tumor tissues than normal tissues. (E) Analysis of the associations between individual SDC gene expression and overall survival (OS) and progression-free survival (PFS) in ovarian carcinoma via Kaplan–Meier Plotter. (F) Kaplan–Meier Plotter analysis showing that high SDCs signature expression is associated with prolonged OS and PFS in patients with ovarian carcinoma. SupplementaryFigure5.jpg Supplementary Figure 5. (A-Y) Correlation between gene expression and cisplatin sensitivity. The mRNA expression levels of SDCs were analyzed for their correlation with cisplatin IC50 values in the CCLE-BROAD-MIT cancer cell line panel using the CellMiner database. The genes in sequence are: MCM6, MCM3, BUB1B, MCM5, CDT1, RAD54L, MCM7, MCM10, RFC3, FANCD2, BRCA1, MCM4, EXO1, PCNA, MAD2L1, BUB1, KIF11, CCNA2, FEN1, CDC45, FANCI, TTK, PLK1, CDC6, CDK1 SupplementaryFigure6.jpg Supplementary Figure 6. GEPIA3-based survival analyses of SDCs in ovarian cancer (OV) patients receiving cisplatin treatment. (A-P) The associations between SDCs expression and OS under cisplatin therapy. SupplementaryFigure7.jpg Supplementary Figure 7. Association of SUCs with cisplatin sensitivity. (A) Correlation between SUCs expression and cisplatin IC50 predicted using the CellMiner database. (B) Association between SUCs signature expression and overall survival (OS) in patients receiving cisplatin therapy. (C) Associations between the expression of individual SUCs and OS under cisplatin therapy. originalwesternblots.jpg SupplementaryTable1.xls SupplementaryTable2.xlsx Cite Share Download PDF Status: Under Revision Version 1 posted Editorial decision: Revision requested 19 Mar, 2026 Reviews received at journal 15 Mar, 2026 Reviews received at journal 05 Mar, 2026 Reviewers agreed at journal 02 Mar, 2026 Reviewers agreed at journal 25 Feb, 2026 Reviewers invited by journal 25 Feb, 2026 Editor assigned by journal 13 Feb, 2026 Submission checks completed at journal 12 Feb, 2026 First submitted to journal 09 Feb, 2026 You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-8439132","acceptedTermsAndConditions":true,"allowDirectSubmit":false,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":598185622,"identity":"332e06e9-1088-41e2-9241-ca7647fceafe","order_by":0,"name":"Yeernaer Hazaisihan","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Yeernaer","middleName":"","lastName":"Hazaisihan","suffix":""},{"id":598185624,"identity":"3d906eb5-9ed3-4223-aaed-c23e61b5229f","order_by":1,"name":"Yankun Yu","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Yankun","middleName":"","lastName":"Yu","suffix":""},{"id":598185626,"identity":"4d87f0f3-3624-4ca0-96ae-d02a936c8d4f","order_by":2,"name":"Li Wei","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Li","middleName":"","lastName":"Wei","suffix":""},{"id":598185627,"identity":"20062ea4-478e-4f92-96a0-16c18c38c1f5","order_by":3,"name":"Xiaoning Li","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Xiaoning","middleName":"","lastName":"Li","suffix":""},{"id":598185628,"identity":"242c0dc7-38ee-4ac2-8676-99afb9e6550e","order_by":4,"name":"Di Zhou","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Di","middleName":"","lastName":"Zhou","suffix":""},{"id":598185629,"identity":"035d6207-6022-431a-9a4e-fff15ef9d6cc","order_by":5,"name":"Shaojie Wang","email":"","orcid":"","institution":"Kurle Hospital","correspondingAuthor":false,"prefix":"","firstName":"Shaojie","middleName":"","lastName":"Wang","suffix":""},{"id":598185631,"identity":"5d804bd0-cc19-4d35-b2b4-128c21287474","order_by":6,"name":"Xiangting Gao","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Xiangting","middleName":"","lastName":"Gao","suffix":""},{"id":598185633,"identity":"66e800c9-76e8-491a-921f-2572c192175a","order_by":7,"name":"Lin Tao","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Lin","middleName":"","lastName":"Tao","suffix":""},{"id":598185637,"identity":"35a9f874-d6ac-4534-a672-6f5c53d7c3f0","order_by":8,"name":"Liang Guo","email":"","orcid":"","institution":"Shihezi University","correspondingAuthor":false,"prefix":"","firstName":"Liang","middleName":"","lastName":"Guo","suffix":""},{"id":598185639,"identity":"e269322a-ea1c-4f5e-972d-73a0252f8533","order_by":9,"name":"Wei Jia","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAAAyElEQVRIiWNgGAWjYBACxmYQaWDDYADmshGvJY0ELVBwmAQtzO3MzyQ+FJy3N+c/Y8DwoewwA//sBkIOYzOTnGFwO3FnwxkDxhnnDjNI3DlASAuD2W0eg9sJBgd7DJh524AulEggpIX92+0/BufsDQ7zGDD/JU4Lj9ltBoMDjBuOAbUwEqml/GePQXLihjNsBQd7zqXzSNwgoMWw//hmgx9/7OwNzh/e+OBHmbUc/wxCWhqQOAeAmAe/eiCQJ6hiFIyCUTAKRgEAF8JAfrpvyEUAAAAASUVORK5CYII=","orcid":"","institution":"Shihezi University","correspondingAuthor":true,"prefix":"","firstName":"Wei","middleName":"","lastName":"Jia","suffix":""}],"badges":[],"createdAt":"2025-12-24 05:38:19","currentVersionCode":1,"declarations":"","doi":"10.21203/rs.3.rs-8439132/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-8439132/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":103731389,"identity":"209ff502-d443-49a9-86f2-43a8e619962a","added_by":"auto","created_at":"2026-03-02 09:15:01","extension":"jpg","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":2696404,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSunitinib suppresses proliferation, migration, and invasion of HGSOC cells. \u003c/strong\u003e(A) CCK-8 assays showed that sunitinib inhibits the proliferation of HGSOC cells. (B–C) The IC₅₀ of sunitinib was estimated by CCK-8 assay: 11.83 μM in HEY-A8 cells and 17.31 μM in SKOV3 cells. (D) Colony-formation assays demonstrated reduced clonogenic growth of HEY-A8 and SKOV3 cells upon sunitinib treatment. (E-G) EdU incorporation assays indicated that sunitinib suppresses HEY-A8 and SKOV3 cells proliferation. (H) Wound-healing assays revealed impaired migratory capacity of HEY-A8 cells upon sunitinib treatment. (The notation “ns” signifies no significance; *p\u0026lt;0.05; **p\u0026lt;0.01; *** p\u0026lt;0.001; ****p\u0026lt;0.0001)\u003c/p\u003e","description":"","filename":"Figure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/b9496b34d42a10b2c08c17bf.jpg"},{"id":104399939,"identity":"a04bd925-c9cb-4437-845b-5d995ccf3aba","added_by":"auto","created_at":"2026-03-11 12:08:15","extension":"jpg","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":1322342,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSunitinib induced cell cycle arrest and promoted necrosis in HGSOC cells.​\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A) Sunitinib induced G0/G1 phase arrest in HEY-A8 cells and G2/M phase arrest in SKOV3 cells, with a concomitant reduction in the S phase population in both cell lines.​ (B–D) Flow cytometry was used to detect apoptosis in HGSOC cell. (C) Sunitinib induced necrosis in HEY-A8 and SKOV3 cells in a dose-dependent manner; (D) Sunitinib induced apoptosis in SKOV3 cells, but failed to induce apoptosis in HEY-A8 cells. (The notation “ns” signifies no significance; *p\u0026lt;0.05; **p\u0026lt;0.01; *** p\u0026lt;0.001; ****p\u0026lt;0.0001).\u003c/p\u003e","description":"","filename":"Figure2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/f4bfeb50f1b7547f48d491f9.jpg"},{"id":103731392,"identity":"c7cabbec-6246-4a61-9712-2d719593c8c9","added_by":"auto","created_at":"2026-03-02 09:15:01","extension":"jpg","order_by":3,"title":"Figure 3","display":"","copyAsset":false,"role":"figure","size":3637031,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eAnalysis of differentially expressed genes and functional enrichment of sunitinib-downregulated gene sets in HEY-A8 cells.​\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A) The volcano map of DEGs. (B) Gene Ontology (GO) enrichment analysis of sunitinib-downregulated gene sets was conducted using the TNMplot database, including biological processes (BP), cellular components (CC), and molecular functions (MF). (C) Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analysis of sunitinib-downregulated gene sets was conducted via the TNMplot database.​ (D) Reactome pathway enrichment analysis was performed for the sunitinib-downregulated gene sets.​ (E) Gene Set Enrichment Analysis (GSEA) was conducted to analyze the sunitinib-downregulated genes.\u003c/p\u003e","description":"","filename":"Figure3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/948593e329f4ad5cc239ef5b.jpg"},{"id":103731394,"identity":"a39e082a-cfd5-40ee-8fc5-640cc58f0da3","added_by":"auto","created_at":"2026-03-02 09:15:01","extension":"jpg","order_by":4,"title":"Figure 4","display":"","copyAsset":false,"role":"figure","size":3984162,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSunitinib synergizes with cisplatin in HGSOC cells and augments DNA damage. \u003c/strong\u003e(A–B) The curve and dose- response matrix for cisplatin and sunitinib in HEY-A8 and SKOV3 cells. (C–D) Drug synergy was evaluated using the Zero Interaction Potency (ZIP) model implemented in SynergyFinder 3.0. In this model, synergy scores \u0026gt; 0 indicate synergistic interactions, whereas scores \u0026lt; 0 indicate antagonism; scores \u0026gt; 10 are considered strong synergy. In the synergy maps, red denotes synergy and green denotes antagonism. (E–F) Western blot analysis of HEY-A8 and SKOV3 cells treated for 24 h with DMSO, cisplatin, sunitinib, and the combination.\u003c/p\u003e","description":"","filename":"Figure4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/d4c278483abdaf65059ec6e0.jpg"},{"id":103731403,"identity":"41ef8dac-2c01-4737-b2e9-6db3f716ffd3","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"jpg","order_by":5,"title":"Figure 5","display":"","copyAsset":false,"role":"figure","size":10232920,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eComprehensive Analysis of the 25-Core Gene Signature in Ovarian Cancer.\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003e(A-C) PPI Network of Genes in the Top 10 Pathways from GO, KEGG, and Reactome Enrichment analyses: Cytoscape-based protein-protein interaction (PPI) networks of the gene sets in the top 10 pathways identified from GO (A), KEGG (B), and Reactome (C) enrichment analyses of the sunitinib-downregulated genes (D)The PPI network of sunitinib-downregulated core genes, which was screened out from Fig.5A-C. (E) Correlation analysis of the 25 core genes, showing strong positive correlations among all genes.(F) Expression analysis of individual genes in ovarian serous cystadenocarcinoma (OSCC) tissues, with higher expression in cancer tissues compared to normal tissues. (G) GEPIA3 analysis showing significant upregulation of the 25-gene signature in ovarian cancer tissues (p = 2.39 × 10⁻⁵). (H) KM plotter analysis indicating that high expression of most genes is associated with poorer overall survival in ovarian cancer patients. (I) KM plotter analysis of the combined 25-gene signature showing significant association with reduced overall survival in ovarian cancer patients.\u003c/p\u003e","description":"","filename":"Figure5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/41147fff104c6e155183d685.jpg"},{"id":103731404,"identity":"2379b7a1-125c-4442-b2e4-36d4aa8b27c2","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"jpg","order_by":6,"title":"Figure 6","display":"","copyAsset":false,"role":"figure","size":3271165,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eCorrelation analysis between SDCs expression and cisplatin sensitivity in ovarian cancer.\u003c/strong\u003e(A) The CellMiner platform was used to analyze the correlation between cisplatin IC₅₀ values and the expression levels of SDCs based on data from the CCLE (BROAD-MIT) and GDSC2 datasets. (B) Analysis of the GEPIA3 database indicated that high expression of the SDC signature is associated with cisplatin resistance in ovarian cancer. (C–K) The GEPIA3 database was further used to evaluate the OS of ovarian cancer patients receiving cisplatin-based chemotherapy according to the expression levels of CDT1, KIF11, BUB1B, MCM7, CDC45, MCM5, FEN1, PLK1, and RAD54L. Patients with high expression of these genes exhibited shorter overall survival, suggesting that elevated expression of these genes is linked to cisplatin chemoresistance.\u003c/p\u003e","description":"","filename":"Figure6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/de64faeaae76a4a18602336b.jpg"},{"id":103731395,"identity":"4470b3f8-3161-4f0d-9852-be265f6d91f2","added_by":"auto","created_at":"2026-03-02 09:15:01","extension":"jpg","order_by":7,"title":"Figure 7","display":"","copyAsset":false,"role":"figure","size":994185,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eIC\u003c/strong\u003e\u003csub\u003e\u003cstrong\u003e50\u003c/strong\u003e\u003c/sub\u003e\u003cstrong\u003e of cisplatin and sunitinib in HEY-A8and HEY-A8/DDP cells. \u003c/strong\u003e(A) IC\u003csub\u003e\u003cstrong\u003e50 \u003c/strong\u003e\u003c/sub\u003eof cisplatin in HEY-A8and HEY-A8/DDP cells. (B) IC\u003csub\u003e\u003cstrong\u003e50 \u003c/strong\u003e\u003c/sub\u003eof sunitinib in HEY-A8and HEY-A8/DDP cells.\u003c/p\u003e","description":"","filename":"Figure7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/c61591e552c88468e1abe322.jpg"},{"id":105562620,"identity":"dc1b7c27-0739-4fd8-b3ae-f08ae396d9ef","added_by":"auto","created_at":"2026-03-27 12:43:41","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":14493395,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/6ee016ce-d3a3-4646-9298-a684db5e8ebc.pdf"},{"id":104808227,"identity":"77039489-b666-4420-b429-c64a92b3a9fc","added_by":"auto","created_at":"2026-03-17 12:33:27","extension":"jpg","order_by":1,"title":"","display":"","copyAsset":false,"role":"supplement","size":2419371,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 1. Sunitinib inhibits proliferation, migration, and invasion of SKOV3 cells.\u003c/strong\u003e (A-B) Transwell migration and Matrigel invasion assays indicate reduced migratory and invasive capacities of HGSOC cells following sunitinib treatment. (C) Wound-healing assays reveal impaired SKOV3 cell migration in the presence of sunitinib. (The notation “ns” signifies no significance; *p\u0026lt;0.05; **p\u0026lt;0.01; *** p\u0026lt;0.001; ****p\u0026lt;0.0001)\u003c/p\u003e","description":"","filename":"SupplementaryFigure1.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/d90dc692106c4141b15f9adf.jpg"},{"id":104400106,"identity":"12cd8d86-8e98-4ae0-8733-718b9a206fff","added_by":"auto","created_at":"2026-03-11 12:08:53","extension":"jpg","order_by":2,"title":"","display":"","copyAsset":false,"role":"supplement","size":2207048,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 2. Enriched Pathways in Sunitinib-Upregulated Gene Set\u003c/strong\u003e. (A) GO enrichment analysis (BP, CC, and MF) performed using TNMplot. (B) KEGG pathway enrichment analysis performed using TNMplot. (D) Reactome pathway enrichment analysis.\u003c/p\u003e","description":"","filename":"SupplementaryFigure2.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/2a7eeb06a38c6ff6bc8f2cc5.jpg"},{"id":103731398,"identity":"b77ee7cc-5590-4f6d-8920-3a1f79da18cb","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"jpg","order_by":3,"title":"","display":"","copyAsset":false,"role":"supplement","size":8346002,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 3. The OS and PFS of SDCs.\u003c/strong\u003e (A-Y) The prediction of OS and PFS of SDCs via Kaplan-Meier database. The genes in sequence are: FANCD2, FANCI, RFC3, MCM10, CDK1, PCNA, BUB1B, MAD2L1, BUB1, CCNA2, BRCA1, KIF11, CDT1, RAD54L, MCM5, MCM4, TTK, MCM6, CDC45, EXO1, MCM7, PLK1, FEN1, MCM3 and CDC6.\u003c/p\u003e","description":"","filename":"SupplementaryFigure3.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/6edb232a2595d454e2854d94.jpg"},{"id":103731405,"identity":"b3b1a6a9-0e2c-4803-9f1c-159cde9ac3ba","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"jpg","order_by":4,"title":"","display":"","copyAsset":false,"role":"supplement","size":5188657,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 4. Identification and clinical relevance of SDCs in ovarian carcinoma.\u003c/strong\u003e(A) PPI network constructed from genes involved in the top 10 enriched GO terms (BP, CC, and MF) of the sunitinib-downregulated gene set. (B) The six hub genes with the highest interaction degree identified from the PPI network (defined as SDCs), and the correlation analysis of SDCs using the TNMplot database. (C) Differential expression analysis of SDCs in ovarian carcinoma predicted using the TNMplot database. (D) Differential expression analysis of the SDCs signature in ovarian carcinoma using the GEPIA3 database, showing higher expression in tumor tissues than normal tissues. (E) Analysis of the associations between individual SDC gene expression and overall survival (OS) and progression-free survival (PFS) in ovarian carcinoma via Kaplan–Meier Plotter. (F) Kaplan–Meier Plotter analysis showing that high SDCs signature expression is associated with prolonged OS and PFS in patients with ovarian carcinoma.\u003c/p\u003e","description":"","filename":"SupplementaryFigure4.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/833638525419ee8cd15f4c08.jpg"},{"id":103731400,"identity":"87a0fb08-4f3f-49df-a2d1-05e76d512d87","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"jpg","order_by":5,"title":"","display":"","copyAsset":false,"role":"supplement","size":9123221,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 5. (A-Y) Correlation between gene expression and cisplatin sensitivity. \u003c/strong\u003eThe mRNA expression levels of SDCs were analyzed for their correlation with cisplatin IC50 values in the CCLE-BROAD-MIT cancer cell line panel using the CellMiner database. The genes in sequence are: MCM6, MCM3, BUB1B, MCM5, CDT1, RAD54L, MCM7, MCM10, RFC3, FANCD2, BRCA1, MCM4, EXO1, PCNA, MAD2L1, BUB1, KIF11, CCNA2, FEN1, CDC45, FANCI, TTK, PLK1, CDC6, CDK1\u003c/p\u003e","description":"","filename":"SupplementaryFigure5.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/eeaf3b512d0a9b53aa8f03df.jpg"},{"id":103731397,"identity":"58a96641-8f8f-402d-b943-454e4513a3ca","added_by":"auto","created_at":"2026-03-02 09:15:01","extension":"jpg","order_by":6,"title":"","display":"","copyAsset":false,"role":"supplement","size":4284221,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 6. GEPIA3-based survival analyses of SDCs in ovarian cancer (OV) patients receiving cisplatin treatment. \u003c/strong\u003e(A-P) The associations between SDCs expression and OS under cisplatin therapy.\u003c/p\u003e","description":"","filename":"SupplementaryFigure6.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/691d9a941578c9dbf6933272.jpg"},{"id":104400003,"identity":"6602a77f-e0d4-4936-ae33-f4f1310bce27","added_by":"auto","created_at":"2026-03-11 12:08:29","extension":"jpg","order_by":7,"title":"","display":"","copyAsset":false,"role":"supplement","size":4245782,"visible":true,"origin":"","legend":"\u003cp\u003e\u003cstrong\u003eSupplementary Figure 7. Association of SUCs with cisplatin sensitivity.\u003c/strong\u003e\u003cbr\u003e\n(A) Correlation between SUCs expression and cisplatin IC50 predicted using the CellMiner database. (B) Association between SUCs signature expression and overall survival (OS) in patients receiving cisplatin therapy. (C) Associations between the expression of individual SUCs and OS under cisplatin therapy.\u003c/p\u003e","description":"","filename":"SupplementaryFigure7.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/e609226242160f087c1fe742.jpg"},{"id":103731401,"identity":"1cc7ea71-b1c1-4aae-8c61-3a086c45ea2c","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"jpg","order_by":8,"title":"","display":"","copyAsset":false,"role":"supplement","size":2205051,"visible":true,"origin":"","legend":"","description":"","filename":"originalwesternblots.jpg","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/42aab1488d0f0381caa1ddfd.jpg"},{"id":103731406,"identity":"9ea30421-acbf-493d-a1b4-87c8a405a577","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"xls","order_by":9,"title":"","display":"","copyAsset":false,"role":"supplement","size":8686709,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable1.xls","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/c3a948516921dbe60d1bf447.xls"},{"id":103731402,"identity":"4cf76990-caff-4a6f-adce-326e329c2e09","added_by":"auto","created_at":"2026-03-02 09:15:02","extension":"xlsx","order_by":10,"title":"","display":"","copyAsset":false,"role":"supplement","size":219683,"visible":true,"origin":"","legend":"","description":"","filename":"SupplementaryTable2.xlsx","url":"https://assets-eu.researchsquare.com/files/rs-8439132/v1/dc83e45ac07135fd139533bd.xlsx"}],"financialInterests":"No competing interests reported.","formattedTitle":"Sunitinib Synergizes with Cisplatin by Suppressing DNA Repair Pathways in High-Grade Serous Ovarian Carcinoma Cells","fulltext":[{"header":"Backgrounds","content":"\u003cp\u003eOvarian carcinoma (OC) is a highly invasive malignant tumor of the female reproductive system. In China, it was estimated that 61,100 new ovarian carcinoma cases and 32,600 attributable deaths were reported in 2022 \u003csup\u003e[1]\u003c/sup\u003e. Although early-stage ovarian carcinoma has high treatment efficacy, the lack of effective screening methods means approximately 75% of patients are diagnosed at an advanced stage, resulting in a 5-year survival rate of less than 30% \u003csup\u003e[2]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eHigh-grade serous ovarian carcinoma (HGSOC) is the most common subtype of OC, which is classified as a subtype of epithelial ovarian carcinoma (EOC). The current standard treatment for EOC, primary tumor debulking surgery is the preferred option. For advanced EOC, neoadjuvant chemotherapy followed by interval tumor debulking surgery serves as an alternative for patients unsuitable for immediate surgery \u003csup\u003e[2,3]\u003c/sup\u003e. Despite the initial efficacy of platinum-based chemotherapy reaching up to 70%, most advanced-stage patients eventually relapse because of chemoresistance\u0026mdash;with a cumulative relapse rate of 70% within three years \u003csup\u003e[4,5]\u003c/sup\u003e. To address this challenge, PARP inhibitors (PARPi) have shown promising benefits for BRCA-mutated and HR-deficient tumors \u003csup\u003e[6,7]\u003c/sup\u003e, but about 50% of HGSOC patients lack HR deficiency and gain limited benefit \u003csup\u003e[8]\u003c/sup\u003e. Besides, most patients with BRCA1/2-mutated tumors initially respond to PARP inhibitors but eventually develop resistance, resulting in relapse and disease progression \u003csup\u003e[9]\u003c/sup\u003e. Immune checkpoint inhibitors (ICIs), even though promising in other cancers, show low efficacy in platinum-resistant EOC due to limited tumor immunogenicity and overlapping resistance mechanisms \u003csup\u003e[10,11]\u003c/sup\u003e. These realities highlight the necessity of elucidating the molecular mechanisms of HGSOC and developing new therapeutic strategy.\u003c/p\u003e \u003cp\u003eSunitinib is an orally bioavailable, multi-targeted receptor tyrosine kinase (RTK) inhibitor approved by Food and Drug Administration (FDA) for adjuvant therapy of high-risk renal cell carcinoma following nephrectomy, as well as for gastrointestinal stromal tumors refractory to or intolerant of imatinib \u003csup\u003e[12,13]\u003c/sup\u003e. It selectively inhibits a variety of RTKs implicated in tumor angiogenesis and growth, including VEGFR-1/2, PDGFR-α/β, c-KIT, RET, and FLT3 \u003csup\u003e[14,15]\u003c/sup\u003e. By inhibiting the key RTKs, sunitinib suppresses angiogenic signaling, tumor progression, and metastasis, making it a promising agent in cancer therapy.\u003c/p\u003e \u003cp\u003eIn OC, the randomized AGO-OVAR 2.11 phase II trial demonstrated that sunitinib was feasible and moderately effective in patients with recurrent platinum-resistant OC \u003csup\u003e[16]\u003c/sup\u003e. Another phase II study evaluating sunitinib in patients with clear cell ovarian cancer demonstrated that the drug exhibits limited activity \u003csup\u003e[17]\u003c/sup\u003e. Although sunitinib has been clinically studied in OC, most studies have focused on its monotherapy, with limited exploration of its combination with cisplatin. Interestingly, in an Ehrlich ascites carcinoma model, sunitinib significantly enhanced the efficacy of cisplatin while attenuating cisplatin-induced nephrotoxicity in rats \u003csup\u003e[18]\u003c/sup\u003e, highlighting its potential as a cisplatin-sensitizing agent. Therefore, we hypothesized that sunitinib may have a similar effect on HGSOC cells, enhancing the sensitivity of cisplatin.\u003c/p\u003e \u003cp\u003eIn this study, we aim to investigate the synergistic effects of sunitinib and cisplatin, providing a basis for improving the treatment efficacy of HGSOC and overcoming cisplatin resistance via combination therapy. We further sought to predict the putative mechanisms by which sunitinib acts, with a particular focus on pathways that may sensitize HGSOC cells to cisplatin.\u003c/p\u003e"},{"header":"Materials and methods","content":"\u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003eMaterials\u003c/h2\u003e \u003cp\u003eSunitinib was provided by Sunitinib was provided by Liang Guo, Shihezi University.\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eCell culture\u003c/h3\u003e\n\u003cp\u003eHGSOC cell lines (HEY-A8, SKOV3) were purchased from Shanghai EK-Bioscience Biotechnology Co, Ltd. The cisplatin-resistant HEY-A8/DDP cell line was established by exposing parental HEY-A8 cells to escalating concentrations of cisplatin, followed by repeated selection and subculturing to achieve stable cisplatin resistance. HEY-A8 and HEY-A8/DDP cells were cultured in Dulbecco\u0026rsquo;s Modified Eagle Medium (Gibco, Thermo Fisher Scientific, USA) supplemented with 10% FBS (Gibco, Thermo Fisher Scientific, USA), 100 U/mL penicillin and 100 mg/mL streptomycin (Solarbio, China) in humidified air at 37 ℃with 5% CO2. SKOV3 were cultured in McCoy's 5A (PM150710, Wuhan, China) supplemented with 10% FBS, 100 U/mL penicillin and 100 mg/mL streptomycin in humidified air at 37 ℃ with 5% CO2.\u003c/p\u003e\n\u003ch3\u003eEdU staining assay\u003c/h3\u003e\n\u003cp\u003eIn a 24-well plate, 1\u0026times;10\u003csup\u003e5\u003c/sup\u003e cells in the logarithmic growth phase were inoculated and cultured to the normal growth stage. Cells were then treated with different concentrations of sunitinib for 24 hours. After treatment, Edu working solution (C0071S, beyotime, China) was added and incubated for 2 hours. The cells were fixed with 4% paraformaldehyde, permeabilized with 0.5% Triton X-100, and subjected to a Click reaction with fluorescent dye-conjugated azide for 30 minutes in the dark. Nuclei were counterstained with DAPI, and Edu-positive cells were visualized and quantified using fluorescence microscopy. Untreated and positive control groups were included for validation.\u003c/p\u003e\n\u003ch3\u003eCell counting kit-8 assay\u003c/h3\u003e\n\u003cp\u003eCells were seeded in 96-well plates at a density of 2,000 cells per well for proliferation assays and 5,000 cells per well for inhibition assays, followed by incubation at 37\u0026deg;C with 5% CO₂ for 6 h to allow cell attachment. After removing the culture medium, 100 \u0026micro;L of fresh culture medium containing varying concentrations of the drug was added to each well, and the cells were incubated for 24 hours. Following this, the medium in each well was replaced with 10% CCK-8 reagent (BS350B, Biosharp, China) in complete medium, and the cells were incubated for an additional 2 hours in the dark at 37\u0026deg;C with 5% CO₂. Absorbance was then measured at 450 nm using a microplate reader.\u003c/p\u003e\n\u003ch3\u003eColony formation assay\u003c/h3\u003e\n\u003cp\u003e1500 cells were evenly seeded in a six-well plate. After 24 hours of cell attachment, the cells were treated with varying concentrations of sunitinib for 48 hours. Subsequently, the medium containing sunitinib was removed, and the cells were maintained in normal culture conditions for 14 days. The colonies were then fixed with 4% paraformaldehyde, stained with 1% crystal violet solution, washed three times with distilled water, air-dried, and photographed.\u003c/p\u003e \u003cdiv id=\"Sec8\" class=\"Section2\"\u003e \u003ch2\u003eFlow cytometry assay for the cell cycle\u003c/h2\u003e \u003cp\u003eCells were seeded in 6-well plates and allowed to grow until they reached approximately 70% confluence. The culture medium was then replaced with fresh medium containing various concentrations of sunitinib, and the cells were treated for 24 hours. After treatment, the cells were harvested by trypsinization, washed with PBS, and fixed in absolute ethanol for one week. The fixed cells were subsequently stained with 1 mL of DNA staining solution (MULTI SCIENCE, CCS012) at room temperature for 30 minutes in the dark. Cell cycle distribution was analyzed using a flow cytometer, and the data were processed to determine the percentages of cells in the G0/G1, S, and G2/M phases, as well as the proportion of apoptotic cells (sub-G1 peak).\u003c/p\u003e \u003c/div\u003e\n\u003ch3\u003eAnnexin-V/PI assay for apoptosis and necrosis\u003c/h3\u003e\n\u003cp\u003eThe YF\u0026reg;647A-Annexin V and PI Apoptosis Kit was purchased from PROTEINBIO (Nanjing, China). Cells were seeded into 6-well plates and cultured until reaching approximately 70% confluence. Then, the culture medium was replaced with fresh medium containing various concentrations of sunitinib, and the cells were incubated for 24 hours. Subsequently, the cells were digested, resuspended in 100 \u0026micro;L of 1\u0026times; binding buffer, and stained with 5 \u0026micro;L of YF647A-Annexin V and 5 \u0026micro;L of PI. The samples were incubated at room temperature in the dark for 15 minutes. Thereafter, 400 \u0026micro;l of Annexin V binding buffer was added, the samples were mixed thoroughly, and flow cytometric analysis was performed. YF647A-Annexin V was excited at 647 nm and detected in the APC channel, while PI was excited at 617 nm.\u003c/p\u003e\n\u003ch3\u003eWestern Blot Analysis\u003c/h3\u003e\n\u003cdiv class=\"Heading\"\u003eWestern Blot Analysis\u003c/div\u003e \u003cp\u003eCells were lysed in RIPA buffer (R0010, Solarbio, China) with PMSF and incubated on ice for 20 minutes. After centrifugation at 12,000 rpm for 20 min at 4\u0026deg;C, protein concentrations were determined. Protein samples were diluted to 1\u0026times; with 5\u0026times; SDS-PAGE Loading buffer (Beyotime, China), and equal amounts were loaded onto an SDS-PAGE gel for electrophoresis, followed by transfer to 0.45 \u0026micro;m PVDF membranes (Millipore, Merck KGaA, Germany). The membranes were blocked with 5% BSA and incubated with primary antibodies overnight at 4\u0026deg;C. Primary antibodies used were β-actin (1:3000, TA-09, ZSGB-BIO, China) and γ-H2AX (1:1000, sc-517348, Santa Cruz, USA). After washing, the membranes were incubated with a horseradish peroxidase-conjugated secondary antibody (1:16,000, BS1247, Bioworld, USA) and protein bands were visualized using SuperSignal\u0026trade; West Pico PLUS chemiluminescent substrate (34580, Thermo Fisher, USA).\u003c/p\u003e \u003cdiv id=\"Sec11\" class=\"Section2\"\u003e \u003ch2\u003eTranswell assay\u003c/h2\u003e \u003cp\u003eThe invasion assay was performed using Matrigel-coated Transwell chambers (LABSELECT, China). Matrigel (Corning, USA) and culture medium were mixed at an 8:1 ratio to create the coating solution, which was then added to the upper chambers of the Transwell system. The cells were treated with different concentrations of sunitinib for 48 hours before plating in the Transwell chambers. For each well, 10,000 cells were seeded. The upper chamber was supplemented with 600 \u0026micro;L of 20% serum-containing medium, while the lower chamber contained 200 \u0026micro;L of serum-free medium with 10,000 cells. The cells were allowed to migrate for 24 hours and invade for 48 hours. After the incubation period, the cells that had invaded the lower chamber were fixed and stained for analysis.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec12\" class=\"Section2\"\u003e \u003ch2\u003eWound healing assay\u003c/h2\u003e \u003cp\u003eAfter plating the cells in the 6-well culture plates, a scratch was made across the cell monolayer using a sterile pipette tip when the cells reached the appropriate confluence. The cells were then treated with different concentrations of serum-free sunitinib. Images of the wound area were captured at 0 and 24 hours then the gap width was measured at both time points to evaluate the cell migration capacity.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec13\" class=\"Section2\"\u003e \u003ch2\u003eStatistical analysis\u003c/h2\u003e \u003cp\u003eStatistical analyses were performed using GraphPad Prism 9.5 software. Two-group comparisons were conducted using two-tailed Student\u0026rsquo;s \u003cem\u003et\u003c/em\u003e-tests, and comparisons among multiple groups were performed using one-way ANOVA. Statistical significance was set at \u003cem\u003ep\u003c/em\u003e\u0026thinsp;\u0026lt;\u0026thinsp;0.05.\u003c/p\u003e \u003c/div\u003e"},{"header":"Results","content":"\u003cdiv id=\"Sec15\" class=\"Section2\"\u003e \u003ch2\u003eSunitinib suppresses proliferation, migration, and invasion in HGSOC cells\u003c/h2\u003e \u003cp\u003eTo evaluate the inhibitory effects of sunitinib on HGSOC cell proliferation, we performed CCK-8 assays to determine the IC50 values in HEY-A8 and SKOV3 cells after 24 h of treatment. The results showed that the IC\u003csub\u003e50\u003c/sub\u003e values of sunitinib in HEY-A8 and SKOV3 cells were 11.83 \u0026micro;M and 17.31 \u0026micro;M, respectively (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eA-C). Colony formation assays showed that sunitinib suppresses the colony formation ability of HEY-A8 and SKOV3 cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eD). EdU incorporation assays demonstrated that sunitinib inhibited proliferation of HEY-A8 and SKOV3 cells in a dose-dependent manner (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eE-G). In addition, wound-healing and Transwell assays revealed significant suppression of both migration and invasion following sunitinib treatment in HEY-A8 and SKOV3 cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003eH and Supplementary Fig.\u0026nbsp;1A-C). These data indicate that sunitinib exerts strong anti-proliferative and anti-metastatic activity in HGSOC cells.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec16\" class=\"Section2\"\u003e \u003ch2\u003eSunitinib induces cell cycle arrest and promotes necrosis in HGSOC cells\u003c/h2\u003e \u003cp\u003eTo investigate how sunitinib suppresses growth and induces cytotoxicity in HGSOC, we exposed HEY-A8 and SKOV3 cells to increasing concentrations of sunitinib for 24 h and quantified cell-cycle distribution and apoptosis. Flow-cytometric analysis showed that sunitinib induced distinct cell-cycle alterations in the two HGSOC cell lines. In HEY-A8 cells, sunitinib treatment primarily caused a G0/G1-phase arrest accompanied by a marked reduction in the S-phase population. Unlike HEY-A8, SKOV3 cells exhibited a pronounced G2/M-phase arrest, which was similarly associated with a substantial decrease in S-phase fraction. (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eA).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eSubsequently, we assessed cell death using flow cytometry. After sunitinib treatment, most cells accumulated in the Annexin V\u0026minus;/PI+ quadrant (Q1), suggesting that sunitinib primarily exerts its cytotoxic effects by inducing necrosis in HEY-A8 and SKOV3 cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eB-C). In addition, sunitinib induced apoptosis in SKOV3 cells but not in HEY-A8 cells (Fig.\u0026nbsp;\u003cspan refid=\"Fig2\" class=\"InternalRef\"\u003e2\u003c/span\u003eD).\u003c/p\u003e \u003cp\u003eIn brief, sunitinib induced cell cycle arrest and caused cell death predominantly through necrosis in HGSOC cells, while also eliciting a modest apoptotic response.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec17\" class=\"Section2\"\u003e \u003ch2\u003eSunitinib downregulates DNA Damage Repair Pathways\u003c/h2\u003e \u003cp\u003eTo elucidate the molecular mechanisms underlying sunitinib\u0026rsquo;s effects on HGSOC cells, HEY-A8 cells were treated with sunitinib and subjected to differential expression analysis (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eA; Supplementary Tables\u0026nbsp;1 and 2). There were relatively few enriched pathways among the upregulated genes (Supplementary Fig.\u0026nbsp;2), while downregulated genes exhibit significant enrichment in several key biological processes, including cell-cycle regulation, DNA replication, and DNA damage repair (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB\u0026ndash;D). Most notably, DNA repair mechanism is well recognized as central determinants of platinum responsiveness in HGSOC \u003csup\u003e[19,20]\u003c/sup\u003e .\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eGene Ontology (GO) enrichment analysis indicated that sunitinib downregulate DNA metabolic processes, DNA repair, and DNA damage response (DDR), with a particular emphasis on double-strand break repair via homologous recombination (HR) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eB). Consistently, KEGG pathway analysis revealed that sunitinib significantly suppresses key DNA damage repair pathways, including homologous recombination (HR), the Fanconi anemia (FA), base excision repair (BER), nucleotide excision repair (NER), and mismatch repair (MMR) (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eC). These pathways are essential for the repair of DNA crosslinks and double-strand breaks induced by cisplatin and other platinum agents \u003csup\u003e[21\u0026ndash;23]\u003c/sup\u003e. Reactome pathway analysis further underscored the downregulation of cell-cycle checkpoints, S-phase progression, and DNA replication (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eD).Additionally, GSEA demonstrated that sunitinib downregulates the pathways: DNA replication, cell cycle progression, and DNA repair\u0026mdash;HR and BER (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eE).\u003c/p\u003e \u003cp\u003eIt is well established that increased DNA repair capacity is a key mechanism of cisplatin resistance in OC. Among these pathways, HR is now firmly established as a key determinant of platinum responsiveness in HGSOC \u003csup\u003e[24]\u003c/sup\u003e. Tumors with HR deficiency (HRD), including those harboring BRCA1/2 or other HRR gene mutations, exhibit higher platinum sensitivity and prolonged progression-free survival compared with HR-proficient tumors, whereas restoration of HR function is a hallmark of acquired platinum resistance \u003csup\u003e[25]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eBER has also been implicated in cisplatin-resistant phenotypes, serving as a key mechanism for repairing cisplatin-induced DNA crosslinks \u003csup\u003e[26]\u003c/sup\u003e. The central BER endonuclease APE1 is frequently overexpressed in ovarian tumors, and its high expression correlates with poor prognosis and chemoresistance \u003csup\u003e[27,28]\u003c/sup\u003e. Likewise, disruption of XRCC1 increases cisplatin-induced DNA damage and sensitizes cells to cisplatin, supporting the concept that attenuation of BER shifts the balance from damage tolerance toward cell death under platinum treatment \u003csup\u003e[29]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eThe FA pathway, which orchestrates repair of DNA interstrand crosslinks during S phase and functionally intersects with HR, is also tightly linked to cisplatin sensitivity \u003csup\u003e[30]\u003c/sup\u003e. Activation of FA signaling through factors such as And-1 and the FANCM/FAAP24 complex promotes resolution of cisplatin-induced crosslinks and contributes to the resistant phenotype in OC cells, whereas suppression of FA components, such as FANCD2\u0026mdash;impairs crosslink repair and re-sensitizes tumors to cisplatin in vitro and in vivo \u003csup\u003e[31,32]\u003c/sup\u003e.\u003c/p\u003e \u003cp\u003eTaken together, previous evidence indicates that upregulation or rewiring of DNA repair programs is a central route to cisplatin resistance \u003csup\u003e[33]\u003c/sup\u003e. In this study, our data support sunitinib-mediated suppression of DNA repair pathways such as HR, BER and FA pathways in HGSOC cells. Therefore, our transcriptomic findings suggest that sunitinib functionally weakens the DDR, limiting the ability of HGSOC cells to repair cisplatin-induced DNA lesions. This provides a strong mechanistic rationale for combining sunitinib with cisplatin to overcome platinum resistance in HGSOC.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec18\" class=\"Section2\"\u003e \u003ch2\u003eSunitinib Synergizes with Cisplatin in HGSOC Cells\u003c/h2\u003e \u003cp\u003eConsidering cisplatin exerts its antitumor effects by inducing DNA crosslinks and DNA damage \u003csup\u003e[34]\u003c/sup\u003e, while sunitinib impairs DNA repair pathways, we next assessed their synergism. To assess the potential synergy, we used SynergyFinder 3.0 \u003csup\u003e[35]\u003c/sup\u003e to analyze the combination of sunitinib and cisplatin in HEY-A8 and SKOV3 cells. The computed Zero Interaction Potency (ZIP) score exceeded 10 in HEY-A8 and SKOV3 cells, indicating a strong synergistic effect between sunitinib and cisplatin in HGSOC cells. (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eA-D).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eTo investigate the effect of sunitinib on DNA damage, the level of DNA damage marker γ-H2AX \u003csup\u003e[36]\u003c/sup\u003e was assed using western blot assay. Our data indicate that, treatment with cisplatin and sunitinib, either alone or in combination, led to increased γ-H2AX expression in both HEY-A8 and SKOV3 cells. Most notably, the combined treatment with sunitinib and cisplatin induced the most significant increase in γ-H2AX levels, suggesting that the combination of these agents induces a higher extent of DNA damage compared to either treatment alone (Fig.\u0026nbsp;\u003cspan refid=\"Fig4\" class=\"InternalRef\"\u003e4\u003c/span\u003eE-F). These results suggest that sunitinib can induce DNA damage in HGSOC cells. Notably, when combined with cisplatin, sunitinib enhances the DNA damage response, resulting in a more substantial DNA damage effect than cisplatin alone. Collectively, sunitinib exhibits synergistic effects with cisplatin in HGSOC cells by downregulating DNA damage repair pathways and enhancing the sensitivity of HGSOC cells to cisplatin.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec19\" class=\"Section2\"\u003e \u003ch2\u003eSunitinib-regulated Core Genes are associated with prognosis in OC\u003c/h2\u003e \u003cp\u003eTo further prioritize key mediators within these sunitinib-suppressed networks, we analyzed protein\u0026ndash;protein interaction (PPI) using the STRING database \u003csup\u003e[37]\u003c/sup\u003e and Cytoscape \u003csup\u003e[38]\u003c/sup\u003e. We then identified 25 downregulated core genes from the PPI network (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eA\u0026ndash;C) and defined them as the sunitinib-downregulated core genes (SDCs) (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eD). Subsequently, we performed correlation analysis using TNMplot \u003csup\u003e[39]\u003c/sup\u003e, which revealed significant positive correlations among the expression levels of these genes in ovarian serous cystadenocarcinoma (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eD).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eWe further analyzed the combined signature of SDCs via the GEPIA3 database \u003csup\u003e[40]\u003c/sup\u003e, and found that it was significantly higher in OC than in normal ovarian tissues, suggesting that these genes are overexpressed in OC (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eE). Consistently, analysis using TNMplot confirmed that these genes were expressed at significantly higher levels in ovarian serous cystadenocarcinoma than in normal ovarian tissues. (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eF).\u003c/p\u003e \u003cp\u003eThe prognostic significance of SDCs was further evaluated using the Kaplan\u0026ndash;Meier Plotter by analyzing overall survival (OS) and progression-free survival (PFS). Most SDCs were associated with shorter OS and PFS in patients with OC (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eG), whereas a few genes were linked to improved outcomes. We further constructed a combined signature model on the Kaplan\u0026ndash;Meier Plotter platform using the average expression of SDCs to predict OS and PFS. Higher SDCs signature expression was significantly associated with shorter OS, while no statistically significant difference was observed for PFS (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eH). Collectively, these findings suggest that sunitinib-mediated suppression of SDCs is associated with improved OS in OC patients, implying that sunitinib may prolong survival by downregulating SDCs.\u003c/p\u003e \u003cp\u003eIn addition, we applied the same workflow to sunitinib-upregulated genes and identified six sunitinib-upregulated core genes (SUCs) (Supplementary Fig.\u0026nbsp;3A-B). Unexpectedly, the SUCs signature was also expressed at higher levels in OC than in normal ovarian tissues (Supplementary Fig.\u0026nbsp;3C-D). However, Kaplan\u0026ndash;Meier Plotter analysis indicated that higher SUCs signature expression was significantly associated with prolonged OS and PFS in OC patients (Supplementary Fig.\u0026nbsp;3E-F). These results suggest that SUCs are linked to a favorable prognosis in OC, and that sunitinib may improve patient outcomes by upregulating these genes.\u003c/p\u003e \u003cp\u003eIn summary, high expression of the SDCs signature is associated with poor prognosis in OC patients, while low expression of the SUCs signature correlates with poor prognosis as well. These findings suggest that sunitinib may improve the survival rate of OC patients by modulating the expression of these genes.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec20\" class=\"Section2\"\u003e \u003ch2\u003eCorrelation between SDCs and Cisplatin Sensitivity in OC\u003c/h2\u003e \u003cp\u003eNext, we evaluated the association of SDCs and SUCs with cisplatin sensitivity. Using the CellMiner database \u003csup\u003e[41]\u003c/sup\u003e, we found that SDCs expression was positively correlated with cisplatinIC\u003csub\u003e50\u003c/sub\u003e, suggesting that higher expression of these genes is associated with increased resistance to cisplatin chemotherapy (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eA and Supplementary Fig.\u0026nbsp;4).\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003eSubsequently, we used GEPIA 3 database to explore the effect of these genes on cisplatin chemotherapy. Our results showed that, in OC patients with high SDCs signature expression, cisplatin treatment did not improve OS, whereas in patients with low SDCs signature expression, cisplatin treatment significantly prolonged OS (Fig.\u0026nbsp;\u003cspan refid=\"Fig5\" class=\"InternalRef\"\u003e5\u003c/span\u003eB). These findings suggest that high SDCs signature expression is associated with resistance to cisplatin-based chemotherapy. We further analyzed individual genes of SDCs and found that high expression of nine genes\u0026mdash;CDT1, KIF11, BUB1B, MCM7, CDC45, MCM5, FEN1, PLK1, and RAD54L\u0026mdash;was significantly correlated with cisplatin resistance in OC patients (Fig.\u0026nbsp;\u003cspan refid=\"Fig6\" class=\"InternalRef\"\u003e6\u003c/span\u003eC-K). Additionally, we observed that only one of one gene was correlated with cisplatin sensitivity in SDCs, while the remaining genes showed no significant correlation with cisplatin chemotherapy outcomes (Supplementary Fig.\u0026nbsp;5).\u003c/p\u003e \u003cp\u003eBesides, we assessed the cisplatin sensitivity of the SUCs by the same methods. Notably, our analysis in the CellMiner Database revealed that 3 of these genes were associated with cisplatin resistance, while 2 were linked to cisplatin sensitivity. (Supplementary Fig.\u0026nbsp;6A). Meanwhile, our survival analysis revealed that no significant association was observed between SDCs and OS in OC patients (Supplementary Fig.\u0026nbsp;6B-C).\u003c/p\u003e \u003cp\u003eTaken together, our data demonstrate that the genes downregulated by sunitinib are associated with cisplatin resistance in OC. This observation supports the notion that sunitinib may reverse cisplatin resistance in HGSOC by downregulate these genes.\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec21\" class=\"Section2\"\u003e \u003ch2\u003eCytotoxic effects of sunitinib in cisplatin-resistant HGSOC cells\u003c/h2\u003e \u003cp\u003eBased on the results of our previous drug sensitivity analysis, we further assessed the cytotoxicity of sunitinib in cisplatin-resistant HEY-A8/DDP cells. Our data showed that the IC₅₀ of cisplatin were 15.81 \u0026micro;M in HEY-A8 cells and 33.42 \u0026micro;M in HEY-A8/DDP cells, corresponding to a resistance index (RI) of 2.11. In contrast, the IC₅₀ of sunitinib were 12.00 \u0026micro;M in HEY-A8 cells and 17.74 \u0026micro;M in HEY-A8/DDP cells, while the RI was 1.48 (Fig.\u0026nbsp;\u003cspan refid=\"Fig8\" class=\"InternalRef\"\u003e7\u003c/span\u003eA-B). The results show that HEY-A8/DDP cells also respond less to sunitinib, although this resistance is not as strong as their resistance to cisplatin. Notably, in HEY-A8/DDP cells, the IC₅₀ of sunitinib (17.74 \u0026micro;M) was lower than cisplatin (33.42 \u0026micro;M), suggesting that sunitinib retains substantial cytotoxic efficacy and is less affected by the cisplatin-resistant phenotype compared with cisplatin. Given these findings, we assume that utilization of sunitinib may hold promise for reducing cisplatin dose and reverse cisplatin-resistance in HGSOC.\u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003cp\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"Discussion","content":"\u003cp\u003eHGSOC is one of the lethal gynecological cancers, with the major challenge being the development of resistance to platinum-based chemotherapy, particularly cisplatin \u003csup\u003e[42]\u003c/sup\u003e. Although most patients with HGSOC initially responded to platinum-based chemotherapy, the majority of individuals with advanced disease ultimately succumb to relapse driven by acquired drug resistance \u003csup\u003e[43]\u003c/sup\u003e. A key mechanism of drug resistance in HGSOC is the activation of DDR pathway, which repairs DNA damage induced by chemotherapeutic agents such as cisplatin \u003csup\u003e[44]\u003c/sup\u003e. In our study, our research demonstrates that sunitinib potentially inhibit DDR pathways to enhance cisplatin sensitivity.\u003c/p\u003e \u003cp\u003eOur findings demonstrate that sunitinib treatment induces reduction in S phase in HGSOC cells. Reactome pathway analysis further supports these observations, showing that sunitinib downregulates S phase progression and the G1/S transition (Fig.\u0026nbsp;\u003cspan refid=\"Fig3\" class=\"InternalRef\"\u003e3\u003c/span\u003eD). Cisplatin treatment leads to the formation of DNA lesions that, during DNA replication, result in double-strand breaks (DSBs) \u003csup\u003e[45]\u003c/sup\u003e. Among the various DNA repair pathways, HR plays a critical role in repairing DSBs with high fidelity by using a sister chromatid as a template \u003csup\u003e[33]\u003c/sup\u003e. Importantly, HR-mediated repair is most active during the S phase of the cell cycle, when sister chromatids are available, facilitating accurate repair of replication-associated DNA damage \u003csup\u003e[46]\u003c/sup\u003e. Therefore, we assume that sunitinib inhibits the G1/S transition, restricting cell entry into S phase and decreasing the proportion of cells undergoing active DNA replication, ultimately sensitizing HGSOC cells to cisplatin.\u003c/p\u003e \u003cp\u003eFurthermore, pathway analysis reveals that sunitinib downregulates several DNA repair pathways, including HR, BER and FA pathway. Collectively, these results point to impaired DNA damage repair and a consequent increase in unrepaired lesions in HGSOC cells, providing a mechanistic rationale for combining sunitinib with cisplatin to improve treatment response.\u003c/p\u003e \u003cp\u003eMeanwhile, our study shows that sunitinib and cisplatin act synergistically in HGSOC cells, leading to more pronounced DNA damage than either agent alone. The synergistic effect suggests that sunitinib may overcome cisplatin resistance by increasing DNA damage, offering a promising therapeutic approach for platinum-resistant HGSOC. What\u0026rsquo;s more, in HEY-A8/DDP cells, sunitinib shows a lower IC₅₀ and a lower resistance index than cisplatin, indicating limited cross-resistance in this cisplatin-resistant setting. These results indicate that sunitinib remains more active than cisplatin in cisplatin-resistant HGSOC cells, supporting its potential to help overcome cisplatin resistance.\u003c/p\u003e \u003cp\u003eMechanistically, we analyzed SDCs, which are associated with poor prognosis and cisplatin resistance. Among these SDCs, CDC45 and MCM2-7 are part of the Cdc45-MCM-GINS (CMG) helicase complex, which is essential for DNA replication and for coping with cisplatin-induced replication fork stalling. \u003csup\u003e[47]\u003c/sup\u003e. High expression of CMG components facilitates DNA replication bypass of cisplatin-damaged DNA, reducing cisplatin's cytotoxicity \u003csup\u003e[48\u0026ndash;51]\u003c/sup\u003e. BUB1B, BUB1, TTK, and MAD2L1 are key components of the spindle assembly checkpoint, promote cisplatin resistance. Elevated BUB1B expression upregulates the RAD51 repair pathway, allowing tumor cells to evade cisplatin-induced apoptosis, while inhibiting BUB1B restores cisplatin sensitivity \u003csup\u003e[52\u0026ndash;55]\u003c/sup\u003e. FEN1, an endonuclease in the BER pathway, enhances cisplatin resistance by promoting the repair of cisplatin-induced DNA lesions, while inhibiting FEN1 sensitizes tumor cells to cisplatin \u003csup\u003e[56]\u003c/sup\u003e. Besides, HR regulators such as RAD54L and BRCA1 facilitate RAD51 loading, and inhibition of HR\u0026mdash;through targeting BRCA1 can sensitize tumors to cisplatin \u003csup\u003e[58]\u003c/sup\u003e. Moreover, suppressing EXO1, a HR-associated nuclease, enhances cisplatin sensitivity in OC\u003csup\u003e[59]\u003c/sup\u003e. KIF11 interacts with CaSR to upregulate BRCA1 and cyclin B1, contributing to cisplatin resistance. Inhibiting KIF11 reduces these proteins and restores cisplatin sensitivity in resistant cancers \u003csup\u003e[60,61]\u003c/sup\u003e. PLK1 signaling promotes chemoresistance, and inhibiting PLK1 with B4 enhances cisplatin efficacy in resistant tumors \u003csup\u003e[62]\u003c/sup\u003e. These results highlight the critical role of SDCs in cisplatin resistance and suggest that inhibiting them by sunitinib can restore cisplatin sensitivity.\u003c/p\u003e \u003cp\u003eMoreover, the combination of sunitinib and cisplatin shows promising potential in overcoming resistance in HGSOC. Sunitinib enhances the DNA damage induced by cisplatin, leading to a stronger cytotoxic response. Importantly, sunitinib demonstrates efficacy in cisplatin-resistant HGSOC cells, with a smaller resistance shift compared to cisplatin alone. These findings support the use of sunitinib as a sensitizing agent to improve cisplatin-based therapies in resistant HGSOC, offering a potential strategy to overcome cisplatin-resistance and improve patient outcomes. Thus, the crosstalk among DNA damage repair, necrosis, and cisplatin resistance, as well as in vivo validation, demands further investigation.\u003c/p\u003e"},{"header":"Conclusion","content":"\u003cp\u003eIn summary, our study highlights the synergistic of sunitinib and cisplatin in HGSOC cells. Sunitinib enhances cisplatin sensitivity by impairing key DNA damage repair pathways, inducing necrosis and causing cell cycle arrest. Drug sensitivity analysis also demonstrates that sunitinib-downregulated genes are associated with cisplatin chemoresistance. Notably, sunitinib exhibits higher cytotoxicity than cisplatin alone in HEY-A8/DDP cells. The combination of sunitinib with cisplatin has the potential to overcome platinum resistance, offering a promising therapeutic strategy for HGSOC patients, particularly those with platinum-resistant disease.\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003ctable border=\"1\" cellspacing=\"0\" cellpadding=\"0\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\" style=\"width: 400px;\"\u003e\n \u003cp\u003eFull form\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\" style=\"width: 153px;\"\u003e\n \u003cp\u003eAbbreviation\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHigh-grade serous ovarian carcinoma\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHGSOC\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eOvarian carcinoma\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eOC\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eEpithelial ovarian carcinoma\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eEOC\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eReceptor tyrosine kinase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eRTK\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eVascular endothelial growth factor receptor\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eVEGFR\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePlatelet-derived growth factor receptor\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePDGFR\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eFood and Drug Administration\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eFDA\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePARP inhibitors\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePARPi\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eImmune checkpoint inhibitors\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eICIs\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCell counting kit-8\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCCK-8\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e5-Ethynyl-2\u0026apos;-deoxyuridine\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eEdU\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePhosphate-buffered saline\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePBS\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSodium dodecyl sulfate-polyacrylamide gel electrophoresis\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSDS-PAGE\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePolyvinylidene fluoride\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePVDF\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eBovine serum albumin\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eBSA\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHorseradish peroxidase\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHRP\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eProtein-protein interaction\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePPI\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eGene Ontology\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eGO\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eKyoto Encyclopedia of Genes and Genomes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eKEGG\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eDNA damage response\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eDDR\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHomologous recombination\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHR\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eFanconi anemia\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eFA\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eBase excision repair\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eBER\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNucleotide excision repair\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNER\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eMismatch repair\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eMMR\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eGene set enrichment analysis\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eGSEA\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePhosphorylated histone H2AX\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003e\u0026gamma;-H2AX\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eOverall survival\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eOS\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eProgression-free survival\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePFS\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSunitinib-downregulated core genes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSDCs\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSunitinib-upregulated core genes\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSUCs\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHomologous recombination deficiency\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eHRD\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eApurinic/apyrimidinic endonuclease 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eAPE1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCdc45-MCM-GINS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCMG\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSpindle assembly checkpoint\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eSAC\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eResistance index\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eRI\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNational Cancer Institute\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNCI\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCancer Genome Atlas\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eTCGA\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNational Center for Advancing Translational Sciences\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eNCATS\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eChromatin licensing and DNA replication factor 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCDT1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eKinesin family member 11\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eKIF11\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eBUB1 mitotic checkpoint serine/threonine kinase B\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eBUB1B\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eMinichromosome maintenance complex component 7\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eMCM7\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCell division cycle 45\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eCDC45\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eMinichromosome maintenance complex component 5\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eMCM5\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eFlap structure-specific endonuclease 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eFEN1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePolo-like kinase 1\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003ePLK1\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eRAD54 like DNA repair and recombination protein\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd valign=\"top\"\u003e\n \u003cp\u003eRAD54L\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eEthics approval and consent to participate\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConsent for publication\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eCompeting interests\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe authors declare no competing interests.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eClinical trial number\u0026nbsp;\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eNot applicable.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis work was supported by Science and Technology Program of XPCC (No. 2025ZD010, No. 2023CB008-02); the National Natural Science Foundation of China(No. 82360494, No. 82460492); the International Science and Technology Cooperation Promotion Plan of Shihezi University(No. GJHZ202301); the First Affiliated Hospital of Shihezi University Science and Technology Project(No. LC2025002); the Independent Project of Shihezi University(No. ZZZC2023046, No. ZZZC2023051); the Natural Science Foundation of Xinjiang Production and Construction Corps (No. 2025DA061); the Major Project of Xinjiang Uygur Autonomous Region (No. 2024A03004-4).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAuthors\u0026apos; contributions\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eConception and design: WJ, YKY, LG. Methodology design: YH, YKY, LW, XNL, XTG, LT. Provision of experimental platforms and materials: LG, WJ, SJW. Experimental implementation: YH, YKY, DZ. Manuscript writing: YH. Review \u0026amp; editing: WJ.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets generated and analysed during the current study are available in the NCBI Gene Expression Omnibus (GEO) repository, https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE318963 (accession number: GSE318963).\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eAcknowledgements\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eWe sincerely thank the Pathological Diagnosis Clinical Medical Research Center of Xinjiang Production and Construction Corps for their assistance.\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eHan B, Zheng R, Zeng H, Wang S, Sun K, Chen R, et al. Cancer incidence and mortality in China, 2022. 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Cell Death Dis 2025;16(1):374. \u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":false,"hideJournal":false,"highlight":"","institution":"","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"journal-of-ovarian-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jovr","sideBox":"Learn more about [Journal of Ovarian Research](http://ovarianresearch.biomedcentral.com)","snPcode":"13048","submissionUrl":"https://submission.nature.com/new-submission/13048/3","title":"Journal of Ovarian Research","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true},"keywords":"HGSOC, sunitinib, cisplatin, DNA repair, combinatory therapy","lastPublishedDoi":"10.21203/rs.3.rs-8439132/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-8439132/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003ch2\u003eBackground:\u003c/h2\u003e \u003cp\u003eHigh-grade serous ovarian carcinoma (HGSOC) frequently develops resistance to platinum-based chemotherapy such as cisplatin, leading to high mortality. Sunitinib, a multi-target Receptor tyrosine kinase inhibitor, has shown potential in the treatment of ovarian carcinoma (OC). However, the synergistic effects between cisplatin and sunitinib, and the sensitization mechanism of sunitinib in HGSOC are not yet understood.\u003c/p\u003e\u003ch2\u003eMethods:\u003c/h2\u003e \u003cp\u003eThe anti-tumor effects of sunitinib on HGSOC cells were assessed using CCK-8, EdU, colony formation, wound healing, Transwell, and flow cytometry assays. RNA sequencing was performed on sunitinib-treated cells, followed by differential expression, enrichment, and protein-protein interaction network (PPI) analyses. Genes on cisplatin sensitivity were predicted using the CellMiner and GEPIA3 databases. Synergy between sunitinib and cisplatin was evaluated using SynergyFinder 3.0, and DNA damage was assessed by γ-H2AX expression.\u003c/p\u003e\u003ch2\u003eResults:\u003c/h2\u003e \u003cp\u003eSunitinib significantly inhibited proliferation, migration, and invasion in HGSOC cells, while further inducing cell cycle arrest, promoting necrosis. Besides, sunitinib downregulated critical DNA damage repair pathways, including homologous recombination, fanconi anemia, and base excision repair. Furthermore, sunitinib synergizes with cisplatin in HGSOC cells, enhancing DNA damage compared to monotherapy. Additionally, we screened out 25 sunitinib-downregulated core genes (SDCs). Drug-sensitivity analyses showed that higher SDCs expression was significantly associated with cisplatin resistance in OC. Notably, in cisplatin-resistant HEY-A8/DDP cells, sunitinib displayed stronger cytotoxicity than cisplatin.\u003c/p\u003e\u003ch2\u003eConclusion:\u003c/h2\u003e \u003cp\u003eSunitinib induces cell-cycle arrest and necrosis. In addition, sunitinib synergized with cisplatin and enhanced cisplatin sensitivity by impairing DNA repair pathways. Drug-sensitivity analyses showed that SDCs are associated with cisplatin resistance in OC, suggesting that sunitinib may help overcome cisplatin chemoresistance. Notably, sunitinib retains substantial cytotoxic activity in cisplatin-resistant cells. Together, these findings suggest that the sunitinib\u0026ndash;cisplatin combination is a promising strategy to overcome cisplatin resistance in HGSOC.\u003c/p\u003e","manuscriptTitle":"Sunitinib Synergizes with Cisplatin by Suppressing DNA Repair Pathways in High-Grade Serous Ovarian Carcinoma Cells","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2026-03-02 09:14:56","doi":"10.21203/rs.3.rs-8439132/v1","editorialEvents":[{"type":"communityComments","content":0},{"type":"decision","content":"Revision requested","date":"2026-03-19T18:11:06+00:00","index":"","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-15T16:50:08+00:00","index":"hide","fulltext":""},{"type":"editorInvitedReview","content":"","date":"2026-03-05T15:31:31+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"180059827333312000048285537561119786606","date":"2026-03-02T12:53:31+00:00","index":"hide","fulltext":""},{"type":"reviewerAgreed","content":"282449315785734860809721867542591560901","date":"2026-02-25T15:32:05+00:00","index":"hide","fulltext":""},{"type":"reviewersInvited","content":"","date":"2026-02-25T12:35:29+00:00","index":"","fulltext":""},{"type":"editorAssigned","content":"","date":"2026-02-13T07:42:59+00:00","index":"","fulltext":""},{"type":"checksComplete","content":"","date":"2026-02-12T11:59:44+00:00","index":"","fulltext":""},{"type":"submitted","content":"Journal of Ovarian Research","date":"2026-02-10T01:49:52+00:00","index":"","fulltext":""}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"journal-of-ovarian-research","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":false,"externalIdentity":"jovr","sideBox":"Learn more about [Journal of Ovarian Research](http://ovarianresearch.biomedcentral.com)","snPcode":"13048","submissionUrl":"https://submission.nature.com/new-submission/13048/3","title":"Journal of Ovarian Research","twitterHandle":"@BioMedCentral","acdcEnabled":true,"dfaEnabled":true,"editorialSystem":"em","reportingPortfolio":"BMC/SO AJ","inReviewEnabled":true,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"256e8714-c0f9-4fca-9432-e064a95cd707","owner":[],"postedDate":"March 2nd, 2026","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"in-revision","subjectAreas":[],"tags":[],"updatedAt":"2026-03-19T18:23:56+00:00","versionOfRecord":[],"versionCreatedAt":"2026-03-02 09:14:56","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-8439132","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-8439132","identity":"rs-8439132","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}