E2F1 and MYC activated SNHG15 promotes gastric cancer progression via interacting with miR-24-3p to regulate cell cycle and ferroptosis

doi:10.21203/rs.3.rs-1766741/v1

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E2F1 and MYC activated SNHG15 promotes gastric cancer progression via interacting with miR-24-3p to regulate cell cycle and ferroptosis

https://doi.org/10.21203/rs.3.rs-1766741/v1

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Background: The regulatory loop between long noncoding RNAs (lncRNAs) and microRNAs (miRNAs) plays a dynamic role in transcriptional and translational regulation, and is involved in carcinogenesis. However, the regulatory circuitry between lncRNAs and miRNAs in gastric cancer (GC) remains elusive.

Methods: The novel SNHG15 was identifed by analyzing RNA-sequencing (RNA-seq) data of GC tissues from our cohort and TCGA dataset, and further validated by qRT-PCR in GC cells and tissues. Gain- and loss-of-function assays were performed to examine the role of SNHG15 on GC both in vitro and in vivo. The regulatory mechanisms of SNHG15 were investigated by bioinformatics, fuorescence in situ hybridization (FISH), luciferase reporter assay, and chromatin immunoprecipitation (ChIP) assay. Meanwhile, the oncogenic activity of SNHG15 was explored in both GC cells and animal xenograft studies.

Results: SNHG15 is highly expressed in GC. The enhanced SNHG15 exists in GC and positively correlates with malignant stage and poor prognosis in GC patients. Gain- and loss-of-function studies showed that SNHG15 was required for affecting cell growth, migration and invasion both in vitro and in vivo. Mechanistically, the oncogenic transcription factors E2F1 and MYC could bind to the SNHG15 promoter and enhance its expression. Meanwhile, SNHG15 increased E2F1, MYC and CDC25A mRNA expression by sponging miR-24-3p. In addition, SNHG15 could also inhibit ferroptosis through a miR-24-3p-mediated regulation of MDM2/p53/SLC7A11 signalling axis.

Conclusions: Our results support a novel model in which the E2F1, MYC/SNHG15/miR-24-3p crosstalk served as the critical effectors in GC tumorigenesis and progression, providing a new therapeutic direction in the treatment of GC.

Gastric cancer

E2F1

MYC

SNHG15

miR-24-3p

Cell cycle

Ferroptosis

With over 1 million new cases and an estimated 769000 deaths, gastric cancer (GC) is the fifth most diagnosed malignancy and the fourth leading cause of cancer-related deaths globally in 2020 [1]. GC ranks the sixth most common malignant tumor and the third-highest cancer mortality in China in 2020 [2]. Despite continuous progress in clinical detection and treatment strategies, the prognosis of GC patients remains poor largely due to late diagnosis, a high postoperative recurrence rate, and metastasis [3]. The mechanisms of tumorigensis and metastasis, which are the main causes of death, remain poorly characterized. Therefore, there is an urgent need to gain a better understanding of tumorigenesis and explore novel diagnostic and therapeutic strategies to improve the GC clinical outcomes.

Only 2% DNA sequences encode proteins and over 90% of them are most non-coding transcripts, which are defined as noncoding RNAs (ncRNAs) [4]. One subcategory of these transcripts, called long noncoding RNAs (lncRNAs), belonging to a class of RNA molecules consisting of more than 200 nucleotides with no or low protein-coding potential [5]. Increasing evidences have shown that lncRNAs play crucial functional roles in various biological processes, such as cell proliferation, cell differentiation, immune response, carcinogenesis and metastasis [6–8]. Accumulating evidence supports that lncRNAs have extremely complicated regulatory mechanisms, including miRNA sponging, epigenetic modification, and transcription regulation [9–11]. LncRNAs can interact with DNA, RNA, and protein molecules to modulate the local or global gene expression, and can be simultaneously regulated by transcription factors (TFs) [12, 13].

Growing evidence suggests that the dysregulation of lncRNAs expression play pivotal roles in the initiation and progression of human tumors [14–16]. The lncRNAs expression profile was frequently abnormal in GC and specific aberrant lncRNAs were related to tumorigenesis, such as SNHG10, GCMA, and COL1A1-014 had important roles in GC progression [17–19]. However, only a small fraction of the lncRNAs in the genome have been functionally characterized, and our current understanding on the expression profiles and biological functions of lncRNAs in GC is still at an early stage. Thus, further insight into the lncRNA-dependent gene-regulatory mechanisms will provide useful prognostic biomarkers and individualized therapy for GC.

In this study, through performing high-throughput RNA-sequencing (RNA-seq), we identified the contributions of the lncRNA small nucleolar RNA host gene 15 (SNHG15) in the development and progression of GC. Our results identified upregulation of SNHG15 was dramatically associated with the malignant biological behaviors of GC cells and poor outcomes in GC patients. Specifically, SNHG15 expression was induced by the oncogenic TFs E2F1 and MYC, and strongly correlated with E2F1, MYC, and CDC25A in GC tissues. Our findings also confirm that SNHG15 forms a molecular decoy for miR-24-3p, which targets E2F1 or MYC for degradation. Meanwhile, miR-24-3p can also silence the gene CDC25A. SNHG15 increased E2F1, MYC and CDC25A mRNA expression by sponging miR-24-3p to regulate cell cycle. In addition, SNHG15 could also inhibit ferroptosis through a miR-24-3p-mediated regulation of MDM2/p53/SLC7A11 signalling axis. These findings uncover a previously unknown tumorigenesis-regulatory mechanism in which a feedback loop comprising SNHG15 and miR-24-3p interplays with E2F1, MYC, CDC25A, and MDM2 to maintain SNHG15-mediated cell proliferation and migration in GC.

Clinical samples and ethical statement

Included in this study were 93 patients who underwent primary GC resection at the Shanghai Ninth People’s Hospital between 2016 and 2018 (SHNPH). Another cohort of patients involving 50 GC tissues and paired normal tissues collected between 2014 and 2018 were obtained from Sun Yat-sen University Cancer Center (SYSUCC). All specimens were snap-frozen in liquid nitrogen after removal from patients and were stored at −80 °C for further use. None of these patients had received chemotherapy or radiation therapy before. Written informed consents were acquired from all patients and this study was approved by the Ethics Committee of the hospitals. All patients were staged according to the criteria of the WHO Classification of Tumors of the Digestive System, 2010 edition.

Transcriptome sequencing and data processing

Total RNA was extracted using an RNeasy Mini Kit (QIAGEN, Valencia, CA). The library was subsequently subjected to high-throughput sequencing by the HiSeq X sequencer (Illumina HiSeq 2000 platform). Differentially expressed genes (DEGs) and pathway analysis were processed as described previously [20]. In addition, the available RNA-seq transcriptome data (371 GC and 32 normal samples) and clinicopathological information were downloaded from The Cancer Genome Atlas (TCGA) database (https://portal.gdc.cancer.gov/repository) for further validation.

Cell culture and transfection

Six GC cell lines (NCIN87, BGC823, MKN28, MGC803, MKN45, AGS) and normal gastric epithelial cells (GES1) were purchased from Chinese Academy of Sciences, Shanghai Institutes for Biological Sciences, and maintained in RPMI-1640 medium (Gibco, Shanghai, China) with 10% FBS (GIBCO, Brazil) at 37 °C with 5% CO₂. Small interfering RNAs (SNHG15 siRNA, E2F1 siRNA, MYC siRNA, and CDC25A siRNA), miR-24-3p mimics, miR-24-3p inhibitor, as well as their corresponding control oligonucleotides were purchased from GenePharma (GenePharma Corporation, Shanghai, China). SNHG15 pcDNA3.1 vector (SNHG15-OE), E2F1 pcDNA3.1 vector (E2F1-OE), MYC pcDNA3.1 vector (MYC-OE), and empty vector (vector) were subcloned into the expression vector pcDNA3.1 (Invitrogen, USA). Transfection was carried out by the reagent Lipofectamine RNAimax (Termo Fisher Scientifc Inc. Massachusetts, USA) based on the manufacturer’s instructions. The GC cells were seeded into six-well plates and cultured overnight until 80% confluence. Transfection was performed using Lipofectamine 2000 (Invitrogen, Carlsbad, CA, USA) according to the manufacturer’s instruction. The transfection efficiency was determined by quantitative reverse‑transcription PCR (qRT‑PCR) and Western blot. Sh-SNHG15 and sh-NC lentivirus were purchased from OBiO Technology (Shanghai, China) and constructed into MGC803 cell lines for further in vivo experiments. SiRNAs, shRNAs and miRNAs sequences are available in Additional file 1: Table S1, S2, and S3.

Total RNA extraction and qRT-PCR

Total RNA from tissues and cells was isolated by TRIzol reagent (Invitrogen), and reverse-transcribed into cDNA with the PrimeScript RT reagent Kit (Takara, Dalian, China). qRT-PCR was performed using SYBR Green PCR Master Mix (Applied Biosystems, Waltham, MA, USA) as our previously described [21, 22]. The sequences of primers used in this study are listed in Additional file 1: Table S4 and S5.

Cell proliferation assay and colony formation assay

The relative cell viability at 1, 2, 3, and 4 d after transfection was investigated using the Cell Counting Kit-8 (Dojindo, Kumamoto, Japan) according to the manufacturer’s protocol. Erastin (abs810744) and ferrostatin-1 (abs813072) were purchased from univ-bio (Absin, Shanghai, China). The stimuli concentrations were as follows: ferrostatin-1, 2 μM; erastin, 20 μM. Briefly, GC cells were cultured in a 96-well plate at 37 °C. After 10 μL CCK8 solution was added to each well, the plates were incubated at 37 °C for 1 h. And then the optical density at 450 nm (OD450) was measured for each sample.

As for colony formation assay, a total of 300 cells were seeded into 6-well plates and cultured in a humidified atmosphere containing 5% CO₂ at 37 °C for 2 weeks. Cell colonies were washed with PBS, fixed with 4% paraformaldehyde, and stained with 0.1% crystal violet (1 mg/mL) for 30 min.

Western blot analysis

Western blotting was performed as previously reported [23]. The protein expression was detected by using antibodies of E2F1, MYC, CDC25A, MDM2, p53, SLC7A11 and GAPDH. Details of antibodies are listed in Additional file 1: Table S6. The expression of GAPDH was used as internal reference.

Cell migration and invasion assay

Cell migration and invasion assays were performed as previously reported [21].

Apoptosis and cell cycle analysis

Apoptosis and cell cycle analysis were examined by flow cytometry as previously reported [21].

5′ and 3′ rapid amplification of cDNA ends (RACE) assay

5′ and 3′ RACE assays were performed to determine the full length of SNHG15 using a SMARTer RACE cDNA Amplification kit (Clontech, Takara, Japan) according to the manufacturer’s instructions.

Fluorescence in situ hybridization

Fluorescence in situ hybridization (FISH) analysis was performed according to a previously described method [11]. Briefly, specific fuorescently labeled probes were made for SNHG15 and miR-24-3p. The probe sequences are listed in Additional file 1: Table S7. The hybridization was performed using RNAscope Fluorescent Reagent Kit (Advanced Cell Diagnostics, Hayward, CA, USA) according to the manufacturer’s instructions. All images were acquired on confocal laser scanning microscopy (Leica, Germany).

Immunohistochemistry

Immunohistochemistry (IHC) was performed and measured as reported previously [24].

Luciferase reporter assay

To determine the effect of E2F1 or MYC on SNHG15 promoter, E2F1-OE, MYC-OE, vector was individually co-transfected into 293 T cells together with the pGL3-based construct containing SNHG15 wild-type (WT) or mutant (MUT) promoter sequences plus Renilla luciferase reporter plasmid. To evaluate the effect of miR-24-3p on SNHG15, MYC 3′UTR, E2F1 3′UTR, CDC25A, or MDM2 3′UTR, 293 T cells were co-transfected with the pmirGLO-SNHG15-WT/MUT, pmirGLO-E2F1-WT/MUT, pmirGLO-MYC-WT/MUT, pmirGLO-CDC25A-3′UTR-WT/MUT, or pmirGLO-MDM2-WT/MUT reporter plasmids individually together with miR-control or miR-24-3p mimics, respectively. Twenty-four hours after transfection, firefly and Renilla luciferase activity were measured by a Dual-Luciferase Reporter Assay System (Promega, Madison, WI, USA).

Chromatin immunoprecipitation assay

Chromatin immunoprecipitation (ChIP) was performed with a ChIP assay kit (Beyotime, Haimen, Jiangsu, China) according to our previous protocol [25]. In summary, MGC803 and MKN45 cells were cross-linked with 1% formaldehyde for 10 min and sonicated to shear DNA to lengths between 200 and 1000 base pairs. Cell lysates were precleared with protein A/G beads before they were incubated at 4 °C overnight with protein A/G beads coated with the anti-E2F1 antibody or anti-MYC antibody. Anti-rabbit immunoglobulin G (IgG) was used as a negative control. After washing, the bead-bound immunocomplexes were eluted by elution buffer. To reverse histone-DNA crosslinks, samples were added with 5M NaCl and then heated at 65 °C for 4 h, followed by treated with proteinase K and further incubated at 45 °C for 1 h. The bound DNA fragments were further purified by DNA Extraction Kit (TIANGEN, Beijing, China) and subjected to qRT-PCR using the specific primers, which are listed in Additional file 1: Table S8.

MDA assay

The relative MDA concentration in cell lysates was assessed by a Lipid Peroxidation (MDA) Assay Kit (S0131S, Beyotime, Shanghai, China), according to the manufacturer’s instructions. This assay measures MDA reaction with thiobarbituric acid (TBA) that generate a MDA-TBA adduct in a sample. The MDA-TBA adduct can be detected by colorimetry (OD = 535 nm).

Cell death assay

For cell death assay, the GC cells were treated with different stimuli, collected using trypsin and without EDTA, and resuspended in 1 ml PBS. Trypan blue exclusion staining was then used to assess cell death during exposure to different stimuli as described previously [23]. Cell death was assessed using a cell viability analyzer (Beckman Coulter, Brea, CA, USA).

In vivo studies

Male BALB/c nude mice at the age of 28 days were purchased from the Institute of Zoology, Chinese Academy of Sciences, and housed in a specific pathogen-free environment. MGC803 cells (1 × 10⁶) stably expressing sh-NC (Scrambled), sh-SNHG15 (shRNA1) or MKN45 cells (1 × 10⁷) stably expressing SNHG15 pcDNA3.1 vector (LV-SNHG15), or empty vector (LEV) were established. Mice were randomly divided into 4 groups: sh-NC, sh-SNHG15; LEV, LV-SNHG15. The subcutaneous tumor formation experiment was performed according to a previously described method [21]. Mice were injected intraperitoneally with 1× 10⁶ cells. All mice were sacrificed under anesthesia at 35 days after injection. Laparotomy was performed to observe metastasis in the peritoneal cavity.

Statistical analysis

Statistical data were analyzed using SPSS 22.0 (IBM Corp., Armonk, NY, USA), R 4.0.5 and GraphPad Prism 8.0. Each experiment was repeated at least three times, and the data were shown as mean±standard deviation in the bar charts. The diferences among multiple groups were performed with the Student t-tests or one-way ANOVAs. The association of the expression of SNHG15 with the patient’s clinicopathological parameters was performed with the Chi–squared test. Overall survival (OS) curves were analyzed with the Kaplan–Meier method and compared by the log rank test. P values of <0.05 were considered signifcant.

Identifcation and characteristics of SNHG15 in GC

Firstly, 50 paired GC tissues and matched adjacent non-tumor tissues were selected to carry out the RNA-seq to analyze any alternative expression profiles of lncRNAs. Among them, 20 lncRNAs that have signifcant diference expression in GC tissues were chosen (15 upregulated circRNAs, 5 downregulated circRNAs) (Fig. 1A) and the same tendency was also found in GC tissues derived from TCGA (Additional file 2: Fig. S1A, B). We further analyzed the expression of the upregulated lncRNAs in 10 GC and the paired non-GC patient’s primary tissues from the SHNPH database and found that SNHG15 was the most signifcantly upregulated in tumor tissues compared with adjacent non-tumor tissues, and the expression of other 5 lncRNAs were also consistent with those observed in RNA-Seq (Additional file 2: Fig. S2). Notably, the results suggested that the expression of SNHG15 in GC tissues was signifcantly increased compared with corresponding normal mucosa sample in GC patients in SHNPH cohort (Fig. 1B) and SYSUCC cohort (Fig. 1C). Te correlation analysis in 93 cases of GC patients with clinical characteristics revealed that SNHG15 expression was correlated with tumor differentiation, lymph node metastasis and TNM stage using Chi-squared test (Table 1). Subsequently, Kaplan–Meier survival analysis with the log-rank test was performed. The data from our center revealed that GC patients with higher SNHG15 expression had poorer overall survival rates than those with lower SNHG15 expression (Fig. 1D). Next, we compared the SNHG15 expression in a tissue microarray containing 93 GC samples using FISH and demonstrated that the expression of SNHG15 was signifcantly upregulated in GC tissues compared with normal mucosa tissues (Fig. 1E); the increased expression of SNHG15 was positively correlated with advanced AJCC TNM stage (Fig. 1F, G). Collectively, these results suggested that the aberrantly high expression of SNHG15 is signifcantly associated with tumor progression in GC. Additionally, the coding potential of SNHG15 was predicted using several prediction software and the NCBI ORF finder (https://www.ncbi.nlm.nih.gov/orffinder/). The results showed that SNHG15 could not code any protein, indicating it a non-coding RNA (Additional file 2: Fig. S3A–D).

SNHG15 functions as a tumor inducer in vitro and in vivo

To elucidate the function of SNHG15 on cell biological behavior, we first explored the expression level of SNHG15 in six GC cell lines compared with normal gastric epithelial cells GES1 (Fig. 2A). Then, we transfected MGC803 and AGS cells with two different siRNAs and each siRNA could effectively inhibit the SNHG15 expression (Fig. 2B). The whole length of SNHG15-pcDNA overexpression vector was transfected into MKN45 cell. Meanwhile, to explore the function of SNHG15 in GC, SNHG15 was also knocked down by two of lentivirus-mediated short hairpin RNAs (shRNAs) in GC, and ectopic SNHG15 was transfected into GC with a lentivirus-carried SNHG15 (Additional file 2: Fig. S4A, B). Following, qRT-PCR verified that the expression of SNHG15 could increase over about 100 fold (Fig. 2B). To investigate the biological roles of SNHG15 in GC progression, the cell proliferation was analyzed using CCK8 assay and clone formation. All results showed that MKN45 cell proliferative capability was increased by SNHG15 overexpression, while knockdown of SNHG15 remarkably attenuated the proliferative effects in MGC803 and AGS cells (Fig. 2C, D). It can be concluded that SNHG15 is effective in promoting GC cell growth. We also tested the influence of SNHG15 on GC cell-cycle progression. As shown in Fig. 2E, knock down of SNHG15 significantly blocked the cell cycle at the G1–S phase, while upregulation of SNHG15 slightly facilitated cell cycle progression.

We then tested the proliferation-inducing effect of SNHG15 on an nude mouse model by injecting SNHG15 stable knockdown MGC803 cells, SNHG15 overexpressing MKN45 cells, or corresponding control cells into nude mice to determine whether SNHG15 could affect GC cell tumorigenesis in vivo. The results suggested that tumors grown from SNHG15 knockdown cells were smaller than the control cells (Fig. 3A–C); however, tumors grown from SNHG15 overexpressing cells were larger than those grown from control cells (Fig. 3 E–G). We further explored the role of SNHG15 in peritoneal dissemination by inoculating cells directly into the abdominal cavity of nude mice, and found that the abdominal metastatic burden was decreased in SNHG15 stable knockdown MGC803 cells, and increased in SNHG15-overexpressing MKN45 cells (Fig. 3D, H). To analyze the physiological function of SNHG15, we also generated additional SNHG15 stable knockdown BGC823 cells, and validated these data in vivo assays (Additional file 2: Fig. S5A–C).

In summary, SNHG15 was confirmed to play important roles on GC cell migration, invasion, and peritoneal colonization, signifying that the effect of SNHG15 on GC patients prognosis might be probably associated with the role in metastasis.

Oncogenic transcription factors contribute to SNHG15 overexpression

To further identify the potential mechanism involved in SNHG15 overexpression in GC, we analyzed the data available from TCGA dataset by cBioPortal [26]. As shown in Fig. 4A, there is a gene copy number co-amplification among SNHG15, E2F1, and MYC. We hypothesized that TFs may play important roles in modulating SNHG15 gene transcription. To confirm this hypothesis, we first obtained ChIP-seq of TFs from the ENCODE project by using UCSC Genome Bioinformatics Site (http://genome.ucsc.edu/) and JASPAR database (https://jaspar.genereg.net/), and found high enrichment of oncogenic TFs E2F1 and MYC at the promoter of SNHG15. Meanwhile, correlation analysis revealed that SNHG15 was significantly and positively correlated with E2F1 and MYC in our cohort (Fig. 4B) and the TCGA dataset (Fig. 4C). In addition, our ChIP assays showed that E2F1 and MYC could bind to the SNHG15 promoter DNA regions (Fig. 4D, left). To further evaluate the biologic roles of E2F1 and MYC in SNHG15 expression, we performed gain-of-function method, and revealed that the overexpression (Fig. 4D, right) upregulated E2F1 and MYC enrichment on the SNHG15 promoter. Meanwhile, overexpression and downregulation of E2F1 and MYC could lead to upregulated or downregulated expression of SNHG15 in GC cells, respectively (Fig. 4E). In addition, luciferase report assays indicated that E2F1 bound to the B (−610 bp) binding site, and MYC bound to the A (−1354 bp) binding site (Fig. 4F).

E2F1, MYC and CDC25A are key downstream targets of SNHG15

To explore the biologic pathways involved in GC pathogenesis, SNHG15 expression level was stratified by the median, and GSEA analysis was applied in our cohort RNA-seq datasets. Enrichment plots of GSEA showed that the gene signatures of cell cycle, nucleotide excision repair, and DNA replication were more involved in GC patients with SNHG15 higher expression relative to patients with lower expression (Fig. 5A) and TCGA datasets (Additional file 2: Fig. S6A). To gain further insights into SNHG15-related pathway in regulating GC progression, we evaluated the gene profiles in GC tissues with altered SNHG15 expression. RNA transcriptome sequencing analysis was used to detect the altered gene expression profiles from high and low expression of SNHG15 in our cohort (Fig. 5B) and TCGA (Additional file 2: Fig. S6B). Gene Ontology analysis revealed that changes in gene sets were primarily related to cell cycle in high-expressed-SNHG15 tissues (Fig. 5C). Further qRT-PCR data and Western blot validated that alterations in SNHG15 expression markedly affected the key gene signatures involved in tumorigenesis and GC progression (Fig. 5D, E), indicating that SNHG15 maybe a significant regulator for modulating the malignant phenotypes of GC by regulating cell cycle.

To further elucidate the potential genes involved in the function of SNHG15, we analyzed the RNA-sequencing and qPCR data, and confirmed that alterations in SNHG15 expression greatly affected E2F1, MYC and CDC25A gene enrichment (Fig. 5B, D, E). Therefore, we tested their expressions in human GC samples. IHC and qRT-PCR showed that E2F1, MYC and CDC25A expressions were increased in GC tumors with high expression of SNHG15 compared with GC tumors with low SNHG15 expression (Fig. 6A). Meanwhile, correlation analysis demonstrated that SNHG15 was significantly and positively correlated with E2F1, MYC and CDC25A in our GC cohort (Fig. 6B). To determine whether E2F1, MYC and CDC25A participated in increased GC cell proliferation and invasion induced by SNHG15, we performed a rescue experiment by cotransfected MGC803 cells with SNHG15-OE, siE2F1, or siMYC, or siCDC25A (Fig. 6C). Transwell assays demonstrated that downregulation of E2F1, MYC, and CDC25A partially rescued the SNHG15-induced cell invasion (Fig. 6D), and colony formation assays and CCK8 also showed that interference of E2F1, MYC and CDC25A could partially rescue the SNHG15-induced cell proliferation in vitro (Fig. 6E, F).

SNHG15 regulates E2F1, MYC, and CDC25A expression by competing for miR-24-3p

To elaborate the mechanism for SNHG15-regulated E2F1, MYC, and CDC25A expression, we firstly examined SNHG15 distribution in GC cells using RNA-fluorescence in situ hybridization and subcellular distribution. It was found that SNHG15 RNAs were more distributed in the cytoplasm (Fig. 1E, F), suggesting that SNHG15 was more likely to play a post-transcriptional regulation function. Knowing that lncRNAs can function as natural “microRNA sponges” that protect mRNAs by competing for the targeting miRNAs, it is worth investigating whether SNHG15 playes a similar role. We predicted miRNAs targeting sites on SNHG15 using the starBase tools prediction algorithm [27]. Of these miRNAs, miR-17-3p, miR-31-5p, miR-103b, and miR-24-3p have been reported to be associated with tumor proliferation and invasion [28, 29], and with significant binding energy predicted by the DIANA tools (Fig. 7A). Next, we focused on these miRNAs and explored the interaction between SNHG15 in GC cell lines. Upregulation of SNHG15 significantly decreased the expression of miR-24-3p, but not the expression of other miRNAs in GC cells (Fig. 7B). Interestingly, miR-24-3p, which was reported to target E2F2, MYC, and other cell-cycle related genes [30, 31], was also found to act as a tumor suppressor in GC in our previous study [21]. To investigate whether SNHG15 acted as a sponge of miR-24-3p, we constructed luciferase reporter vectors containing the wild type known as pLUC-SNHG15-WT or mutated miR-24-3p binding sites known as pLUC-SNHG15-mut. It was found that the upregulation of miR-24-3p decreased the luciferase activities of the reporter vector pLUC-SNHG15-WT but not mutant reporter vector in 293T cells (Fig. 7C). To illustrate whether SNHG15 was modulated by miR-24-3p, we conducted transiently upregulation or inhibition of miR-24-3p by mimics or inhibitor in MGC803 cells. We found SNHG15 levels were inhibited or upregulated after overexpression or inhibition of miR-24-3p (Additional file 2: Fig. S7). These findings indicated that miR-24-3p was associated with the expression of SNHG15. To verify the ceRNA network in GC tissues, miR-24-3p expression level was also analyzed and Pearson analysis showed a negative correlation between the expression score of miR-24-3p and SNHG15 (Additional file 2: Fig. S8).

To determine whether SNHG15 regulated E2F1, MYC, and CDC25A expression by competing for miR-24-3p, we performed rescue assays. Ectopic expression of SNHG15 increased E2F1, MYC, and CDC25A protein levels and upregulation of miR-24-3p reduced this increase, while inhibition of SNHG15 decreased the expression of E2F1, MYC, and CDC25A, and the use of miR-24-3p inhibitor reversed this decrease (Fig. 7D). We also found that the expression of miR-24-3p was negatively correlated with the expression of E2F1, MYC, and CDC25A in our GC tissues (Additional file 2: Fig. S9). CCK8 also showed that miR-24-3p partially rescued the SNHG15-induced cell proliferation in vitro (Additional file 2: Fig. S10). Ectopic expression of miR-24-3p remarkably inhibited cell migration and invasion, and re-expression of miR-24-3p was able to reverse the effect of SNHG15 on cell migration, invasion and colony formation in vitro (Fig. 7E, F). The results illustrated that SNHG15 regulated E2F1, MYC, and CDC25A expression by interacting with miR-24-3p.

To test the effect of miR-24-3p on the predicted target genes, we constructed three luciferase reporters to examine miR-24-3p activity by inserting the 3′ untranslated region (UTR) of human E2F1, MYC and CDC25A into 3′UTR of Renilla luciferase. As expected, miR-24-3p could significantly decrease the luciferase reporter signals of 3′UTR of E2F1, MYC and CDC25A (Fig. 7G). Meanwhile, these results also suggest that miR-24-3p regulates SNHG15 expression by targeting E2F1 and MYC. Furthermore, we conducted FISH in GC cells to examine the subcellular localization of miR-24-3p and SNHG15, and found a large extent of overlap in the cytoplasm, suggesting co-localization of miR-24-3p and SNHG15 (Fig. 7H).

SNHG15 acts as a repressor of ferroptosis in GC cells

Gene Ontology analysis demonstrated that changes in gene sets were mainly correlated to the regulation of signal transduction by p53 in addition to cell cycle regulation in high-expressed-SNHG15 tissues (Fig. 5C). Given that tumor growth arrest, DNA repair and apoptosis are believed to constitute the core mechanisms of p53-dependent tumour suppression [32], we examined the effects of SNHG15 on apoptosis by flow cytometry. However, we found that upregulation or downregulation of SNHG15 had no effect on apoptosis of GC cells (Additional file 2: Fig. S11A, B). LncRNAs are involved in the regulation of reactive oxygen species (ROS) levels, and ferroptosis is a form of cell death resulting from the catastrophic accumulation of lipid ROS [33]. Further analyses in our cohort (Additional file 2: Fig. S12A, B) and the TCGA database (Additional file 2: Fig. S12C, D) revealed that SNHG15 expression was significantly correlated with key regulatory gene sets of ferroptosis. We further confirm the effect of SNHG15 on ferroptosis, and found that downregulation of SNHG15 increased erastin-induced ferroptosis (Fig. 8A), which was rescued by treatments of the cells with ferroptosis inhibitor ferrostatin-1 (Fig. 8B). Conversely, SNHG15 overexpressing GC cells were robustly resistant to erastin-induced ferroptosis (Fig. 8C). Moreover, downregulation of SNHG15 in GC cells exhibited an increase in MDA production, while SNHG15 overexpression decreased MDA production (Fig. 8D, E).

Although p53-mediated growth arrest and apoptosis serve as critical barriers to cancer development, emerging evidence suggests that ferroptosis regulation of p53 is also important [34]. As shown in Fig. 5C, p53 may play an important role in SNHG15-mediated carcinogenesis. To test the effect of miR-24-3p on the regulation of signal transduction by p53, we constructed luciferase reporter containing the 3′UTR of human MDM2, which is considered to be a negative regulator of p53 signaling pathway [35]. As expected, miR-24-3p could significantly decrease the luciferase reporter signals of 3′UTR of MDM2 (Fig. 8F). Previous study showed p53 sensitized cells to ferroptosis by repressing expression of SLC7A11, a key regulator of ferroptosis[34]. Rescue assays revealed that ectopic expression of SNHG15 increased MDM2 protein level and upregulation of miR-24-3p reduced this increase, and SNHG15 suppressed ferroptosis by miR-24-3p-mediated regulation of MDM2/p53/SLC7A11 signal axis (Fig. 8G). Meanwhile, we found that miR-24-3p inhibitor could deteriorate ferroptosis promotion induced by downregulation of SNHG15 (Fig. 8H), while upregulation of miR-24-3p could reverse the ferroptosis inhibition induced by SNHG15 (Fig. 8I). Taken together, these findings indicate that the SNHG15/miR-24-3p/MDM2/p53/SLC7A11 signaling inhibits ferroptosis.

Accumulating evidences have identified that lncRNAs are aberrantly expressed in various tumors, where they hold potential applications for cancer diagnosis, monitoring, cancer therapy or prognosis [36-38]. In this study, we compared the lncRNA profiles of GC and adjacent normal control tissues by RNA-seq analysis from our cohort and TCGA datasats, and found a novel lncRNA SNHG15 that demonstrated significant overexpression in GC. We further confirmed this expression pattern in another GC cohort and found that high SNHG15 expression was correlated with GC differentiation, lymph node metastasis, TNM stage, and poor prognosis. More concretely, SNHG15 upregulation could serve as a biomarker to differentiate between cancerous and normal gastric tissues. These fndings suggest that SNHG15 could be served as a prominently diagnostic and prognostic marker or therapeutic target for GC.

To explore the biological functions of SNHG15, we first confirmed that cell cycle, DNA replication and mismatch repair were remarkably enriched as a response to SNHG15 alteration in GC by GO analysis and GSEA. Knock down of SNHG15 repressed GC cell growth and invasion in vitro and in vivo, while upregulation of SNHG15 promoted cell proliferation and metastasis. These findings consistently support that aggressive GC is more prone to show higher SNHG15 expression, which may interpretate the key roles that SNHG15 plays in the progression of GC.

The TFs such as YAP1, LEF1, MYC, and MYB are often activated during gastric tumorigenesis, leading to abnormal genes regulation by copy number amplifications and upregulation of their expressions [39-41]. In the current study, the results suggested that genomic-level alterations contributed noticeably to the aberrant expression of lncRNAs in GC. Importantly, we found that oncogenic TFs E2F1 and MYC, as well as SNHG15, were upregulated through gene copy number amplification in GC by analyzing TCGA datasets. In this study, our data manifested that SNHG15 could be activated by E2F1 and MYC in GC cells. By combining high-throughput data analysis and functional assays in SNHG15 higher or lower expression GC patients, we identified three key target genes (E2F1, MYC, and CDC25A), which are known as well characterized genes involved in cancer proliferation and migration by modulating cell cycle [42-44]. Meanwhile, these results were also confirmed in the tissue samples from patients with GC.

LncRNAs characteristically perform their biological functions by physically interacting with regulatory proteins or acting as miRNA sponges [45]. For instance, lncRNA GSTM3TV2 upregulates LAT2 and OLR1 through competitively sponging let-7 to promote gemcitabine resistance in pancreatic cancer [46]. On the other hand, lncRNA ITGB8-AS1 modulates cell proliferation and tumor growth of colorectal cancer via sponging miR-33b-5p and let-7c-5p/let-7d-5p to regulate the expression of ITGA3 and ITGB3 [47]. LncRNAs have distinct functions on the basis of their subcellular distribution. We found that SNHG15 was a cytoplasmic lncRNA in GC cells, suggesting that it plays a role in post-transcriptional regulation. LncRNA can interfere with miRNAs by competing for shared miRNA response elements and affect post-transcriptional regulation. It was found in our study that SNHG15 shared miR-24-3p response elements with E2F1, MYC and CDC25A. Then we applied FISH and luciferase reporter assays to confirm that SNHG15 sponged miR-24-3p and protected E2F1, MYC and CDC25A against degradation by miR-24-3p, leading to the upregulation of E2F1, MYC, and CDC25A.

Ferroptosis is a form of lipid peroxidation-induced cell death that can be involved in tumorigenesis [48, 49], also highlighting the great potential of pharmacologic regulation of ferroptosis for cancer therapy. The p53 tumor suppressor not only participates in the control of cell survival and division under various stresses, but also regulates ferroptosis through a transcriptional or posttranslational mechanism [50]. The p53-mediated transcriptional repression of SLC7A11 is critical for ROS-induced ferroptosis [34]. LncRNAs are considered master regulators of the ferroptosis process, and lncRNA-based therapy can be served as an exciting choice to modulate ferroptosis for cancer treatment [51]. However, the molecular mechanism underlying ferroptosis resistance that is mediated by the aberrant activation of SNHG15 and the subsequent regulation of p53 signaling pathway, remains unclear. In the present study, we found that the SNHG15/miR-24-3p/MDM2/p53 signaling pathway decreased the levels of cellular lipid peroxidation by upregulating the expression of SLC7A11, resulting in the suppression of GC cells’ ferroptosis. Taken together, we investigated a novel model in which the E2F1, MYC/SNHG15/miR-24-3p feedback loop facilitated GC malignant progression in this study (Fig. 9).

In summary, our report identifed that an oncogenic driver, SNHG15, was elevated in GC tissues and associated with GC proliferation, migration and invasion. Additionally, higher expression of SNHG15 in GC patients was positively associated with metastasis and poorer survival. We proposed a mechanism that SNHG15 could be transcriptionally activated by E2F1 and MYC, and further blocked the inhibition activity of miR-24-3p on E2F1, MYC, and CDC25A; therefore, overexpression of SNHG15 could upregulate the expression of CDC25A, accompanied with the activation of E2F1 and MYC to regulate cell cycle. Meanwhile, SNHG15 could also inhibit ferroptosis through a miR-24-3p-mediated regulation of MDM2/p53/SLC7A11 signalling axis. Importantly, our results indicate that SNHG15 is a valuable prognostic predictor of GC and appears to be a promising target for GC therapy.

LncRNA: Long noncoding RNA; miRNA: MicroRNA; GC: Gastric cancer; OE: overexpression; SNHG15: small nucleolar RNA host gene 15; TFs: transcription factors; CCK8: Cell counting kit-8; FISH: Fluorescence in situ hybridization; TCGA: The Cancer Genome Atlas; qRT-PCR: Quantitative reverse transcription polymerase chain reaction; ChIP: Chromatin immunoprecipitation; shRNA: Short hairpin RNA; siRNA: Short interfering RNA; IHC: Immunohistochemistry; 3′-UTR: 3′-Untranslated region; RACE: rapid amplification of cDNA ends; CDS: Coding sequence; GEO: Gene expression omnibus; KEGG: Kyoto Encyclopedia of Genes and Genomes; GO: Gene Ontology; RNA-seq: RNA-sequencing; HRP: Horseradish peroxidase; DEGs: Diferentially expressed genes; ROS: reactive oxygen species; MDA: malondialdehyde.

Acknowledgements

We thank all our patients for their participation in this study and the staff at the Department of Gastric Surgery, Fudan University Shanghai Cancer Center. We thank the TCGA research network for providing the data analyzed in this article.

Authors’ contributions

DX, GW, QC, WK, YD, and YD designed the experiments and wrote the manuscript. YD, KL, YD, YL, BL, BH, ML, PK, JG, and XC performed the experiments. YD and YL analyzed the data. YD, ML, PK, JG, and XC collected patient samples. DX, QC, and GW revised this manuscript. All authors read and approved the final manuscript.

Funding

This work was supported by grants from the National Natural Science Foundation of China Grant (81802303).

Availability of data and materials

All data generated during this study are included in this published article and its supplementary files.

Ethics approval and consent to participate

All human tumor tissue samples were collected in accordance with national and institutional ethical guidelines.

Consent for publication

All authors approved the manuscript for submission and consented for publication.

Competing interests

The authors declare no confict of interests.

Author details

¹Department of Gastric Surgery, Fudan University Shanghai Cancer Center, Shanghai 200032, China. ²Department of Thoracic Surgery, Shanghai Pulmonary Hospital, Tongji University School of Medicine, Shanghai 200433, China. ³Breast Cancer Center, Shandong Cancer Hospital and Institute, Shandong First Medical University and Shandong Academy of Medical Science, Jinan 250117, China. ⁴Department of General Surgery, Shanghai Ninth People's Hospital, Shanghai JiaoTong University School of Medicine, Shanghai 200011, China. ⁵Department of Pathology, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011, China. ⁶Department of Laboratory Medicine, Shanghai Ninth People's Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai 200011, China. ⁷Department of Anatomical and Cellular Pathology, The Chinese University of Hong Kong, Hong Kong 999077, China. ⁸Fudan University Shanghai Cancer Center, School of Basic Medical Sciences, Department of Biochemistry and Molecular Biology, Shanghai Medical College of Fudan University, Shanghai 200032, China. ⁹MOE&NHC&CAMS Key Laboratory of Medical Molecular Virology, Shanghai Institute of Infections Disease and Biosecurity & School of Basic Medical Science, Shanghai Medical College, Fudan University, Shanghai 200032, China.

Sung H, Ferlay J, Siegel RL, Laversanne M, Soerjomataram I, Jemal A, et al. Global Cancer Statistics 2020: GLOBOCAN Estimates of Incidence and Mortality Worldwide for 36 Cancers in 185 Countries. CA Cancer J Clin. 2021;71:209–249.
Cao W, Chen HD, Yu YW, Li N, Chen WQ. Changing profiles of cancer burden worldwide and in China: a secondary analysis of the global cancer statistics 2020. Chin Med J (Engl). 2021;134:783–791.
Smyth EC, Nilsson M, Grabsch HI, van Grieken NC, Lordick F. Gastric cancer. Lancet. 2020;396:635–648.
Djebali S, Davis CA, Merkel A, Dobin A, Lassmann T, Mortazavi A, et al. Landscape of transcription in human cells. Nature. 2012;489:101–108.
Fatica A, Bozzoni I. Long non-coding RNAs: new players in cell differentiation and development. Nat Rev Genet. 2014;15:7–21.
Battistelli C, Sabarese G, Santangelo L, Montaldo C, Gonzalez FJ, Tripodi M, et al. The lncRNA HOTAIR transcription is controlled by HNF4alpha-induced chromatin topology modulation. Cell Death Differ. 2019;26:890–901.
Jiang M, Zhang S, Yang Z, Lin H, Zhu J, Liu L, et al. Self-Recognition of an Inducible Host lncRNA by RIG-I Feedback Restricts Innate Immune Response. Cell. 2018;173:906–919 e913.
Wang Z, Yang B, Zhang M, Guo W, Wu Z, Wang Y, et al. lncRNA Epigenetic Landscape Analysis Identifies EPIC1 as an Oncogenic lncRNA that Interacts with MYC and Promotes Cell-Cycle Progression in Cancer. Cancer Cell. 2018;33:706–720 e709.
Zhang E, Han L, Yin D, He X, Hong L, Si X, et al. H3K27 acetylation activated-long non-coding RNA CCAT1 affects cell proliferation and migration by regulating SPRY4 and HOXB13 expression in esophageal squamous cell carcinoma. Nucleic Acids Res. 2017;45:3086–3101.
Liang Y, Chen X, Wu Y, Li J, Zhang S, Wang K, et al. LncRNA CASC9 promotes esophageal squamous cell carcinoma metastasis through upregulating LAMC2 expression by interacting with the CREB-binding protein. Cell Death Differ. 2018;25:1980–1995.
Liu S, Sun Y, Hou Y, Yang L, Wan X, Qin Y, et al. A novel lncRNA ROPM-mediated lipid metabolism governs breast cancer stem cell properties. J Hematol Oncol. 2021;14:178.
Jariwala N, Sarkar D. Emerging role of lncRNA in cancer: a potential avenue in molecular medicine. Ann Transl Med. 2016;4:286.
Xu TP, Liu XX, Xia R, Yin L, Kong R, Chen WM, et al. SP1-induced upregulation of the long noncoding RNA TINCR regulates cell proliferation and apoptosis by affecting KLF2 mRNA stability in gastric cancer. Oncogene. 2015;34:5648–5661.
Yuan JH, Yang F, Wang F, Ma JZ, Guo YJ, Tao QF, et al. A long noncoding RNA activated by TGF-beta promotes the invasion-metastasis cascade in hepatocellular carcinoma. Cancer Cell. 2014;25:666–681.
Ding X, Jia X, Wang C, Xu J, Gao SJ, Lu C. A DHX9-lncRNA-MDM2 interaction regulates cell invasion and angiogenesis of cervical cancer. Cell Death Differ. 2019;26:1750–1765.
Li B, Kang H, Xiao Y, Du Y, Xiao Y, Song G, et al. LncRNA GAL promotes colorectal cancer liver metastasis through stabilizing GLUT1. Oncogene. 2022;41:1882–1894.
Lan T, Yuan K, Yan X, Xu L, Liao H, Hao X, et al. LncRNA SNHG10 Facilitates Hepatocarcinogenesis and Metastasis by Modulating Its Homolog SCARNA13 via a Positive Feedback Loop. Cancer Res. 2019;79:3220–3234.
Tian Y, Ma R, Sun Y, Liu H, Zhang H, Sun Y, et al. SP1-activated long noncoding RNA lncRNA GCMA functions as a competing endogenous RNA to promote tumor metastasis by sponging miR-124 and miR-34a in gastric cancer. Oncogene. 2020;39:4854–4868.
Dong XZ, Zhao ZR, Hu Y, Lu YP, Liu P, Zhang L. LncRNA COL1A1-014 is involved in the progression of gastric cancer via regulating CXCL12-CXCR4 axis. Gastric Cancer. 2020;23:260–272.
Huang B, Li Q, Geng Q, Lao J, Guo J, Huang S, et al. ASTE1 frameshift mutation triggers the immune response in Epstein-Barr virus-associated gastric cancer. Signal Transduct Target Ther. 2022;7:4.
Duan Y, Hu L, Liu B, Yu B, Li J, Yan M, et al. Tumor suppressor miR-24 restrains gastric cancer progression by downregulating RegIV. Mol Cancer. 2014;13:127.
Chen X, Cai Y, Hu X, Ding C, He L, Zhang X, et al. Differential metabolic requirement governed by transcription factor c-Maf dictates innate γδT17 effector functionality in mice and humans. Sci Adv. 2022;8:eabm9120.
Duan Y, Gao Y, Zhang J, Chen Y, Jiang Y, Ji J, et al. Mitochondrial aldehyde dehydrogenase 2 protects gastric mucosa cells against DNA damage caused by oxidative stress. Free Radic Biol Med. 2016;93:165–176.
Li L, Liang Y, Kang L, Liu Y, Gao S, Chen S, et al. Transcriptional Regulation of the Warburg Effect in Cancer by SIX1. Cancer Cell. 2018;33:368–385.e367.
Gao YH, Wu ZX, Xie LQ, Li CX, Mao YQ, Duan YT, et al. VHL deficiency augments anthracycline sensitivity of clear cell renal cell carcinomas by down-regulating ALDH2. Nat Commun. 2017;8:15337.
Cerami E, Gao J, Dogrusoz U, Gross BE, Sumer SO, Aksoy BA, et al. The cBio cancer genomics portal: an open platform for exploring multidimensional cancer genomics data. Cancer Discov. 2012;2:401–404.
Li JH, Liu S, Zhou H, Qu LH, Yang JH. starBase v2.0: decoding miRNA-ceRNA, miRNA-ncRNA and protein-RNA interaction networks from large-scale CLIP-Seq data. Nucleic Acids Res. 2014;42:D92-97.
Xiang P, Yeung YT, Wang J, Wu Q, Du R, Huang C, et al. miR-17-3p promotes the proliferation of multiple myeloma cells by downregulating P21 expression through LMLN inhibition. Int J Cancer. 2021;148:3071–3085.
Chen Y, Zhao H, Li H, Feng X, Tang H, Qiu C, et al. LINC01234/MicroRNA-31-5p/MAGEA3 Axis Mediates the Proliferation and Chemoresistance of Hepatocellular Carcinoma Cells. Mol Ther Nucleic Acids. 2020;19:168–178.
Lal A, Navarro F, Maher CA, Maliszewski LE, Yan N, O'Day E, et al. miR-24 Inhibits cell proliferation by targeting E2F2, MYC, and other cell-cycle genes via binding to "seedless" 3'UTR microRNA recognition elements. Mol Cell. 2009;35:610–625.
Lee SH, Chen TY, Dhar SS, Gu B, Chen K, Kim YZ, et al. A feedback loop comprising PRMT7 and miR-24-2 interplays with Oct4, Nanog, Klf4 and c-Myc to regulate stemness. Nucleic Acids Res. 2016;44:10603–10618.
Tanikawa C, Matsuda K, Fukuda S, Nakamura Y, Arakawa H. p53RDL1 regulates p53-dependent apoptosis. Nat Cell Biol. 2003;5:216–223.
Badgley MA, Kremer DM, Maurer HC, DelGiorno KE, Lee HJ, Purohit V, et al. Cysteine depletion induces pancreatic tumor ferroptosis in mice. Science. 2020;368:85–89.
Jiang L, Kon N, Li T, Wang SJ, Su T, Hibshoosh H, et al. Ferroptosis as a p53-mediated activity during tumour suppression. Nature. 2015;520:57–62.
Haupt Y, Maya R, Kazaz A, Oren M. Mdm2 promotes the rapid degradation of p53. Nature. 1997;387:296–299.
Li Y, Gan Y, Liu J, Li J, Zhou Z, Tian R, et al. Downregulation of MEIS1 mediated by ELFN1-AS1/EZH2/DNMT3a axis promotes tumorigenesis and oxaliplatin resistance in colorectal cancer. Signal Transduct Target Ther. 2022;7:87.
Liu SJ, Dang HX, Lim DA, Feng FY, Maher CA. Long noncoding RNAs in cancer metastasis. Nat Rev Cancer. 2021;21:446–460.
Chang KC, Diermeier SD, Yu AT, Brine LD, Russo S, Bhatia S, et al. MaTAR25 lncRNA regulates the Tensin1 gene to impact breast cancer progression. Nat Commun. 2020;11:6438.
Zhou LQ, Li SH, Wu Y, Xin L. Establishment of a prognostic model of ten transcription factors in gastric cancer. Genomics. 2021;113:4075–4087.
Zhou Y, Zhang J, Li H, Huang T, Wong CC, Wu F, et al. AMOTL1 enhances YAP1 stability and promotes YAP1-driven gastric oncogenesis. Oncogene. 2020;39:4375–4389.
Choi W, Kim J, Park J, Lee DH, Hwang D, Kim JH, et al. YAP/TAZ Initiates Gastric Tumorigenesis via Upregulation of MYC. Cancer Res. 2018;78:3306–3320.
Mandigo AC, Yuan W, Xu K, Gallagher P, Pang A, Guan YF, et al. RB/E2F1 as a Master Regulator of Cancer Cell Metabolism in Advanced Disease. Cancer Discov. 2021;11:2334–2353.
Lourenco C, Resetca D, Redel C, Lin P, MacDonald AS, Ciaccio R, et al. MYC protein interactors in gene transcription and cancer. Nat Rev Cancer. 2021;21:579–591.
Biswas K, Philip S, Yadav A, Martin BK, Burkett S, Singh V, et al. BRE/BRCC45 regulates CDC25A stability by recruiting USP7 in response to DNA damage. Nat Commun. 2018;9:537.
Xue ST, Zheng B, Cao SQ, Ding JC, Hu GS, Liu W, et al. Long non-coding RNA LINC00680 functions as a ceRNA to promote esophageal squamous cell carcinoma progression through the miR-423-5p/PAK6 axis. Mol Cancer. 2022;21:69.
Xiong G, Liu C, Yang G, Feng M, Xu J, Zhao F, et al. Long noncoding RNA GSTM3TV2 upregulates LAT2 and OLR1 by competitively sponging let-7 to promote gemcitabine resistance in pancreatic cancer. J Hematol Oncol. 2019;12:97.
Lin X, Zhuang S, Chen X, Du J, Zhong L, Ding J, et al. lncRNA ITGB8-AS1 functions as a ceRNA to promote colorectal cancer growth and migration through integrin-mediated focal adhesion signaling. Mol Ther. 2022;30:688–702.
Chen X, Kang R, Kroemer G, Tang D. Broadening horizons: the role of ferroptosis in cancer. Nat Rev Clin Oncol. 2021;18:280–296.
Tang D, Chen X, Kang R, Kroemer G. Ferroptosis: molecular mechanisms and health implications. Cell Res. 2021;31:107–125.
Hong T, Lei G, Chen X, Li H, Zhang X, Wu N, et al. PARP inhibition promotes ferroptosis via repressing SLC7A11 and synergizes with ferroptosis inducers in BRCA-proficient ovarian cancer. Redox Biol. 2021;42:101928.
Balihodzic A, Prinz F, Dengler MA, Calin GA, Jost PJ, Pichler M. Non-coding RNAs and ferroptosis: potential implications for cancer therapy. Cell Death Differ. 2022;29:1094–1106.

Table 1 Correlation of the expression of SNHG15 in GC with clinicopathologic features

Characteristics	lncRNA SNHG15		p value
Characteristics	Low	High	p value
Gender
Male	23	25	0.581
Female	19	26
Age (years)
<60	20	23	0.808
≥60	22	28
Tumor location
Proximal third	10	9	0.512
Middle third	9	16
Distal third	23	26
WHO classification
Adenocarcinoma	33	43	0.375
Signet-ring cell carcinoma	3	5
Mucinous adenocarcinoma	6	3
Differentiation
Well, moderately	23	16	0.023
Poorly	19	35
Borrmann classification
Ⅰ/Ⅱ	17	28	0.166
Ⅲ/Ⅳ	25	23
Local invasion
T1、T2	12	22	0.147
T3、T4	30	29
Lymph node metastasis
No	19	10	0.008
Yes	23	41
Distant metastasis
No	39	44	0.308
Yes	3	7
TNM stage
Ⅰ/Ⅱ	18	11	0.027
Ⅲ/Ⅳ	24	40

No competing interests reported.

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E2F1 and MYC activated SNHG15 promotes gastric cancer progression via interacting with miR-24-3p to regulate cell cycle and ferroptosis

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