Background
Nasopharyngeal carcinoma is a rare type of head and neck cancer originates from the nasopharyngeal epithelium, with the highest incidence in southeast Asia [
1,
2]. Radiotherapy is the standard treatment for early disease, and radiotherapy combined with chemotherapy is the standard treatment for advanced NPC patients [
3]. Cell proliferation and tumor growth for NPC patients is generally acknowledged as the main reason for treatment failure [
1,
3,
4]. Therefore, more efforts are required for the development of novel biomarkers and targets for NPC diagnosis and therapy.
Glycolysis, which is also known as the Warburg effect, has been widely recognized as a central hallmark of human cancer [
5,
6]. The high frequency of high glycolysis rates in cancer cells remains an established feature of many human tumors. This energy production process provides metabolites for cancer cells and can be used as precursors of anabolic pathway, which supports the biosynthetic needs of malignant cell proliferation [
6‐
8]. A better understanding of the mechanistic links between glycolysis and cell proliferation may ultimately lead to better treatments for human cancer. Phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) is a glycolysis-regulatory enzyme, which efficiently catalyzes the production of F-2,6-BP and lactate [
9]. Recent studies revealed the unexpected role of PFKFB3 in promoting cell proliferation by regulating the expression of key cell cycle-related proteins [
10,
11]. Therefore, PFKFB3 inhibition may be a promising approach for cancer treatment.
Long non-coding RNAs (lncRNAs) are a class of RNA molecules consisting of more than 200 nucleotides without protein-coding potential [
12,
13]. LncRNAs are now recognized to constitute a regulatory system that function at the transcriptional and post-transcriptional levels [
12,
14,
15]. Recently, many lncRNAs have been identified to promote tumorigenesis through metabolic reprogramming, but the mechanisms remain elusive [
16,
17]. Thus, the roles of lncRNAs in metabolism reprogramming and the underlying mechanisms have attracted our interest.
In this study, we identify and functionally characterize a novel metabolism-related lncRNA in NPC. We demonstrate a strong relationship between LINC00930 dysregulation and NPC development. Functionally, LINC00930 has a pivotal role in glucose metabolism remodeling and cell proliferation. We provide evidence that LINC00930 serves as a molecular scaffold for the interaction of RBBP5 and GCN5, thus enhancing H3K4 trimethylation and H3K9 acetylation levels, which transactivates the target gene PFKFB3 in NPC. Therefore, our study reveals a previously unappreciated lncRNA, which connects the glycolytic remodeling and NPC progression and is a promising therapeutic target.
Methods
Patient specimens
For cohort 1 (Supplementary Table
S1), 71 cases of freshly-frozen tumor tissues and corresponding normal nasopharyngeal epithelium samples were obtained from NPC patients. For cohort 2 (Supplementary Table
S2), another 128 paraffin-embedded NPC biopsy tissues and adjacent tissues were collected. Patients of both cohort 1 and 2 were obtained from the Fourth Affiliated Hospital of Guangxi Medical University from January 2006 to December 2017 and had detailed clinical characteristics and long-term follow-up data. All patients were pathologically diagnosed with primary NPC, without receiving any antineoplastic therapy prior to biopsy.
A total of 120 nasopharyngeal brushing samples including normal nasopharyngeal epithelium and pre-cancerous lesions (SM or DYS) were archived and collected from August 2016 to December 2017 in the Fourth Affiliated Hospital of Guangxi Medical University. No corticosteroid, rhinitis spray or NSAIDs should be used within 4 weeks before sampling.
Cell lines
Eleven NPC cell lines (6-10B, 5-8F, HNE1, HNE2, HNE3, CNE1, CNE2, HONE1, SUNE1, C666–1 and HK-1) were maintained in RPMI-1640 (Invitrogen, Carlsbad, USA) supplemented with 10% fetal bovine serum (FBS, Gibco, Grand Island, USA). The human immortalized nasopharyngeal epithelial cell line (NP69) was cultured in keratinocyte/serum-free medium (Invitrogen) supplemented with bovine pituitary extract (BD Bioscience, CA, USA). All cell lines were genotyped for identity by GENEWIZ Biotechnology Co., Ltd. (Suzhou, China) and tested routinely for Mycoplasma contamination (Yeasen, # 40601ES20).
Reagents and antibodies
Trichostatin A (#CSN12139), Anacardic Acid (#CSN13640) and Curcumin (#CSN19424) were purchased from CSNPharm. YF-2 (#S0022) and PFK15 (#S7289) were obtained from Selleck chemicals. Antisense oligonucleotide (ASO) of LINC00930 was provided from Takara Bio. Antibodies against p27 (#sc1641) was obtained from Santa cruz Biotechnology. Antibodies against CDK1 (#ab133327), PFKFB3 (#ab181861), RBBP5 (#ab154755), GCN5 (#ab217876) and β-tubulin (#ab6046) were purchased from Abcam. Antibodies against H3K4me3 (#9751), H3K9me3 (#13969), H3K27me3 (#9733) IgG (#2985) were provided by Cell Signaling Technology.
Plasmid constructs, lentivirus and siRNAs
The full-length cDNA of human LINC00930 was PCR-amplified and subcloned into the pCDNA3.1 vector (Invitrogen), and the final construct was verified by sequencing. The empty pcDNA3.1 vector was used as the control. The small hairpin RNA (shRNA) of the LINC00930 was provided by Addgene. The siRNAs targeting human LINC00930, PFKFB3, RBBP5, GCN5 and the scramble siRNA control were all purchased from Takara (Dalian, China). Details of shRNAs and siRNAs target sequences are listed in Supplementary Table
S3. N-Terminal FLAG-tagged RBBP5 and HA-tagged GCN5 expression vectors (for expression in mammalian cells) were provided by OBiO Technology (Shanghai, China). Point mutations (Supplementary Table
S4) in constructs were generated using a Site-Directed Mutagenesis Kit (Agilent). Plasmid vectors and siRNAs transfection was conducted using the Lipofectamine 3000 reagent (Life Technologies) according to manufacturer’s instructions. Cells were harvested 48–72 h post transfection for various assays.
Differentially expressed lncRNAs analysis
We profiled differentially expressed genes between NPC and normal tissue using two independent sets of microarray data, GSE64634 [
18] and GSE12452 [
19]. Both microarray data were performed and analysed based on Affymetrix Human Genome U133 Plus 2.0 platform. GSE64634 has 12 NPC and 4 normal nasopharyngeal samples; GSE12452 contains 31 NPC and 10 normal nasopharyngeal samples, respectively. GSE64634 and GSE12452 gene expression profiles date was analyzed by means of DESeq2 online software with fold change ≥2.0 and the false discovery ratio (FDR) < 0.01. Four hundred twenty-one and three hundred eighty-two differentially expressed genes were obtained in GSE64634 dataset GSE12452 dataset, respectively. The differentially expressed lncRNAs between NPC and non-tumor tissues were listed in Supplementary Table
S5.
In situ hybridization for LINC00930
We used three 30-base nucleotide probes from different regions of the LINC00930 corresponding to bp 550–579, 875–904 and 1211–1240 of the LINC00930. The sequences were 5′-CTC TTC ATT CAA CTG GTA CCT CAG TTG GAA-3′, 5′-TGT GAG GAC AAC TGG AAG GTG CCC CTC ACC-3′, and 5′-CTG AGG GGT CAC CAG TCA CAG CCA TGG CCT-3′. The probes were synthetized and labeled with DIG-dUTP at the 3′ end using a kit from Boster (# MK10098, Wuhan, China). In situ hybridization was performed as previously described [
20]. Briefly, After deparaffinized in xylene and re-hydrated in gradient ethanol solution, the slides were then fixed in 4% paraformaldehyde. After that, the slides were incubated with pre-hybridizing solution at 37 °C for 2 h, washed with PBS for 3 times, and incubated with specific targeted probes dissolved in sheared salmon sperm DNA (Invitrogen) and yeast tRNA solution, incubated at 4 °C overnight. The concentration of probe solution was 500 ng/mL. The slides were then washed in 37 °C gradient SSC solution. Finally, dig nucleic acid detection kit (Roche) was used for immunological detection of digoxin. In order to avoid the degradation of RNA by RNase, all glassware and solutions involved in ISH assay should be treated with RNase inhibitor DEPC.
A quantitative scoring criterion for in situ hybridization was used in which both the staining intensity and the number of positive cells were recorded as previously described [
21]. Specifically, the staining scores of LINC00930 was double-blind read by two experienced pathologists. LINC00930 was mainly located in the nucleus. The positive cell percentage and staining intensity of positive cells were scored respectively. Positive cell percentage evaluating criteria: Five high-power visual fields were observed on each slice, and the percentage of positive cells was counted. Less than 5% was 0, 5% ~ 25% was 1, 26% ~ 50% was 2, 51% ~ 75% was 3, and 76% ~ 100% was 4. Positive staining intensity evaluating criteria: colorless is 0, light yellow is 1, brownish yellow is 2 and dark brown is 3. The staining score was obtained by multiplying the two scores: 0–6 points were considered as low expression, 7–12 points were considered as high expression. Correlations between different clinical status and LINC00930 positive expression were analyzed using a chi-square test.
TRIzol reagent (Invitrogen) was utilized to isolate total RNA from NPC tissues and cells. The RNeasy serum/plasma kit (QIAGEN GmbH) was applied to extract total RNA from serum samples. The RNA quality and amount were evaluated by a NanoDrop 3300 spectrophotometer (Thermo Scientific). Reverse transcriptase (Promega) was used to perform reverse transcription. SYBR Green qPCR Super Mix-UDG (Thermo Fisher) was employed to conduct Quantitative real-time PCR (qRT-PCR). β-tubulin was utilized as the normalization control. Specific primers are shown in Supplementary Table
S4.
For the colony formation assay, 1000 cells in 2 ml medium were seeded into six-well plates and cultured for 6 or 11 days. Cell colonies were then successively fixed, stained, and counted. For the CCK-8 assay, 2 × 103 cells in 200 μl medium solution were cultured in 96-well plates. Then, a 20 μl CCK-8 reagent (DOJINDO) was added to each well. After the 96-well plates were incubated for 2–4 h at 37, we then measure the absorbance at 450 nm for each sample well.
Flow cytometry analysis
Flow cytometry analysis was performed as previously described [
22,
23]. In brief, 5 × 10
5 cells were plated into 6-well plates. After adhering to the well, the cells were treated with serum starvation for 24 h and harvested by trypsin after releasing for 48 h. Cells were trypsinized, washed three times with PBS, and fixed overnight in 75% pre-cooled ethanol at − 20 °C. Cells were washed three times with PBS, and PI/RNA reagent (BD, USA) was added and incubated for 15 min at room temperature in the dark. Single-cell suspension was obtained through a nylon membrane and detected by flow cytometer. The results were analyzed by Modfit 3.2 software. Three independent experiments were performed.
Seahorse Biosciences XF96 analyzer (North Billerica, MA, USA) was applied to determine ECAR (extracellular acidification rate) and OCR (oxygen consumption rate), as previously described [
15,
24]. Cells transfected with control siRNA, LINC00930 shRNA, empty vector, and LINC00930 overexpressing vector, were seeded in a XF96-well Assay plates and incubated overnight. ECAR was measured under basal conditions and in response to 10 mM glucose, 5 μM oligomycin, and 100 mM 2-deoxyglucose (all from Sigma-Aldrich). OCR was measured under basal conditions and in response to 5 μM oligomycin, and 50 mM FCCP and 20 mM Rota/AA (all from Sigma-Aldrich). For evaluation of the real-time ECAR and OCR, 3 min of mixture, 3 min of waiting, and 3 min of measurement was performed in turn. ECAR and OCR measurements were normalized to total protein content and reported as mpH/min.
13C-Labeled intracellular metabolites were detected as previously described [
24]. Briefly, cells (2 × 10
7) were incubated with 2 g/L
13C6-labeled glucose (Sigma-Aldrich, St. Louis, MO) for 2 h. Metabolites were extracted, and those including at least one
13C atom were analyzed using an LCsystem equipped with a TripleTOF 5600 mass spectrometer (SCIEX, Framingham, MA, USA). The concentrations of
13C6-labeled metabolites were normalized to cell number.
F-2,6-BP level measurement
NPC cells and xenograft tumor tissues were collected to detect F-2,6-BP levels using a coupled enzyme reaction as previously reported [
25]. Briefly, Cells and tissues samples were homogenized and lysed in NaOH and then heated for 10 min at 80 °C. After cooling, the samples were centrifuged and the supernatant neutralized with acetic acid. F-2,6-BP was measured by the stimulation of PP
i:PFK and assayed in the presence of 0.5 mM pyrophosphate and 1 mM fructose 6-phosphate. Finally, the F-2,6-BP concentration was normalized to total cellular protein according to the manufacturer’s instructions.
Lactate concentration detection
Lactate concentration in growth medium and xenograft tumor was measured using a Lactate Assay kit (Abcam) according to the manufacturer’s protocols. First, xenograft tumor tissues were powdered and homogenized in absolute methanol to obtalin tissue homogenate. Then, the cell medium or tissue homogenate was collected and then incubated with reaction mix at room temperature for 30 min. Absorbance at a wavelength of 450 nm was measured using a microplate reader (Bio-Tek). Lactate production = lactate in cultured medium (mM) − lactate in the fresh medium (mM).
PFK activity
PFK activity in of NPC cells was determined as reported before [
26]. Cells were lysated in the basic buffer containing 50 mM Tris-HCl (pH = 7.4), 5 mM MgCl
2, 5 mM (NH
4)
2SO
4, 1 mM F-6-P, 1 mM ATP, 0.5 mM NADH, 2 mU/ml aldolase, 2 mU/ml triosephosphate isomerase, 2 mU/ml α-glycerophosphate dehydrogenase. PFK activity was assayed in the presence increasing concentrations of F-2,6-BP (1 μM, 10 μM, and 50 μM) or AMP (1 μM, 2 μM, 10 μM, 50 μM, and 150 μM) added to buffer. Enzymatic activity was measured in triplicate spectrophotometrically at 340 nm.
RNA pull-down assay
Biotinylated lncRNAs were refolded in NEB enzyme buffer with RNaseOUT (Invitrogen). To prepare cell lysates, NPC cells were harvested into RNA pull-down buffer containing 0.25% NP-40, 10 mM Tris-HCl (pH = 7.0), 1.5 mM MgCl2, 10 mM KCl, 0.5 mM DTT, 1 mM PMSF, and protease inhibitor complex (PIC). Cell suspension was gently pipetted up and down 15 times and then centrifuged at 12000 rpm for 30 s immediately. The pellets were dissolved in 800 μl of RNA pull-down buffer as the nuclear protein. For the pull-down incubations, nuclear lysates were precleared with streptavidin beads and then incubated with 2 μg of biotinylated RNA and 40 μl of streptavidin beads for 4 h at 4 °C. Beads were collected by centrifugation and RNA-associated proteins were eluted and subjected to MS analysis or Western blotting.
RNA immunoprecipitation (RIP)
RIP assay was conducted with a Magna RNA-binding protein immunoprecipitation kit (Millipore, Bedford, MA) as previously described [
24]. Briefly, cells were crosslinked with formaldehyde and then lysed with RIP buffer. Then cell lysate was incubated with protein beads and antibody complex overnight at 4 °C. Anti-RBBP5 antibody (1:100, Abcam), anti-GCN5 antibody (1:50, Abcam) and negative control IgG (1:100, Cell Signaling Technology) were used in this study. RNA samples were extracted and subjected to Northern blot and qRT-PCR analysis. RNA levels were normalized to the input (20%).
Co-immunoprecipitation (co-IP)
Co-immunoprecipitation (Co-IP) was performed as previously described [
27]. Briefly, cells were lysed in 1 mL of IP lysis buffer (50 mM Tris-HCl at pH 7.4, 1 mM EDTA at pH 8.0, 150 mM NaCl, 1% NP-40, PIC, PhosSTOP, and PMSF). Cell lysates were immunoprecipitated with indicated primary antibodies overnight at 4 °C and then Protein A/G (5:1) for 4 h. The beads were washed three times with the lysis buffer and eluted in SDS sample buffer. The eluted immunocomplexes were resolved by SDS-PAGE, followed by Western blotting.
Chromatin immunoprecipitation (ChIP) and chromatin isolation by RNA purification (ChIRP)
ChIP assay was employed to determine the binding of the chromatin-modifying complexes to the promoter regions of PFKFB3. Cells were fixed and immunoprecipitated using the EZ-Magna ChIP assay kit as recommended by the manufacturer (Millipore, USA). Antibodies anti-H3K4me3, anti-H3K9me3 and anti-H3K27me3 were utilized to immunoprecipitate purified chromatin. Primers used to amplify promoter regions of PFKFB3 are shown in Supplementary Table
S4. For ChIRP assay, a 3′ end Biotin-TEG modified-DNA probe targeting LINC00930 was synthesized and purified by HPLC. Probes were divided into the odd number group (1, 3, 5, etc.) and the even number group (2, 4, 6, etc.), and probes targeting LacZ were selected as nonspecific controls.
Western blot
Total protein was extracted from cultured cells by using RIPA lysis buffer supplemented with 1% proteinase inhibitor complex (PIC, Roche) and 1% PhosStop (Roche). Proteins were separated by 10% SDS-PAGE gel electrophoresis and then transferred to PVDF membranes (Millipore). The membranes were blocked with 5% BSA for 1 h and then incubated with primary antibodies and horseradish peroxidase–conjugated secondary antibodies successively, and the signals were detected with ECL Substrate Kit (Thermo Scientific).
Immunohistochemistry (IHC)
IHC was performed using an EnVision HRP kit (Dako) as previously described [
22]. Briefly, paraffin-embedded mouse and human tissues were deparaffinized in xylene, re-hydrated in gradient ethanol. Then, antigen was retrieved by boiling the sample for 60 min. Samples were incubated with anti-PFKFB3 antibody at 4 °C overnight. The sections were stained with secondary antibody for 30 min at room temperature and then stained DAB reagent. The staining score of PFKFB3 was obtained by multiplying the the percentage and staining intensity of positive cells. The results were scored by two independent investigators. A score of 0–6 was considered to represent low expression and a score of 7–12 was considered to represent high expression.
Subcellular fractionation
A Cytoplasmic & Nuclear RNA Purification Kit (Norgen Biotek Corp, Canada) was used to detect LINC00930 expression in cytoplasmic and nuclear fractions. According to the manufacturer’s instructions, RNA was extracted from the cytoplasmic and nuclear fractions and subjected to qRT-PCR. β-Actin (ACTB) and GAPDH were applied as a cytoplasmic marker, and U1 was used as a nuclear marker.
Luciferase reporter assay
Different promoter regions of PFKFB3 (Supplementary Table
S4) was amplified and cloned into pGL3 basic luciferase reporter vectors. NPC cells were grown and transfected with vectors of promoter-firefly LUC, internal Renilla LUC and other relevant siRNAs. Forty-eight hours post-transfection, cells were washed with PBS. The luciferase reporter assay was conducted using a Dual-Luciferase Reporter Assay System (Promega, E1910) according to the manufacturer’s instructions. Luminescence was measured using a Gen5 microplate reader (BIOTEK, USA).
Xenografts
For cell-derived xenograft (CDX), NPC cells (2 × 106) were injected subcutaneously into the dorsal flanks of 4-week-old female BALB/c nu/nu mice (8 mice per group). Tumor volume was measured at the indicated time points and calculated as length × width2/2. For patient-derived xenograft (PDX), fresh NPC tumor samples were immediately inoculated subcutaneously into both flanks of nude mice. When the successfully established PDXs (F1) reached ~ 500 mm3, the tumors were transplanted to other mice (F2). Eventually, the mice bearing F3 grafts were used for radiotherapy and chemotherapy experiments.
Drugs treatment
For in vitro cell culture model, LINC00930 inhibitor (ASO LINC, Takara) and PFKFB3 inhibitor (PFK15, Selleck) was dissolved in physiological saline buffer and DMSO, respectively. Five thousand cells per well were seeded in a 96-well plate and cultured for 24 h. ASO LINC or/and PFK15 were added to indicated concentrations. Then, cells were subjected to different irradiation intensity (0, 2, 4, 6 and 8 Gy) with 6 MV X-rays and cultured for another 48 h. Relative survival was calculated as absorbance of the drug-treated cells/absorbance of the corresponding cells without drug treatment × 100%.
For xenograft animal models, PFK15 was dissolved in the solvent solution containing 5% DMSO, 40% PEG-300, 5% Tween-80, 5% Propylene glycol and 45% H2O (50 mg/kg, intraperitoneal administration). In vivo-optimized LINC00930 inhibitor (ASO LINC) was dissolved in physiological saline buffer (5 nM per injection, intratumoral administration). Once tumor sizes reached 100–150 mm3, mice were randomly assigned into five groups (8 mice per group), including vehicle group, vehicle + radiotherapy group, LINC00930 inhibitor + radiotherapy group, PFKFB3 inhibitor + radiotherapy group, ASO LINC + PFK15 + radiotherapy group. Mice for radiotherapy treatment were exposed to irradiation by 6 MV X-ray, with 2 Gy per day, twice a week. After 4 weeks, all mice were sacrificed, and the tumor weights were measured and calculated as length×width2/2. The animal experimental protocols were approved by the Medical Experimental Animal Care Commission of Guangxi medical university and Jining Medical University, and performed in accordance with the institutional ethical guidelines for animal experiments.
Statistical analysis
Statistical analysis was carried out using SPSS 19.0 software and GraphPad Prism 5 and Image J software. Two-tailed and unpaired Student’s t-tests were used for two group comparisons. One-way ANOVA tests were applied for multiple groups comparisons. Wilcoxon test and nonparametric Mann-Whitney-Wilcoxon test were both utilized for evaluating the differences of tumor tissues and paired controls. Pearson correlation analysis was performed to analyze the correlation of two molecules. Survival curves were estimated using the Kaplan-Meier method and compared using the log-rank test. Data are shown as the mean ± SD. P < 0.05 was considered statistically significant.
Discussion
Metabolic reprogramming is one of the key characteristics of malignant tumor. Recently, a flow of researches have implicated that lncRNAs were involved in reprogramming energy metabolism and regulated cell proliferation and tumor progression [
5,
12,
14,
17,
41,
42]. However, how lncRNA regulates cellular energy metabolism, especially glucose metabolism in NPC, remains largely unknown. Here, we report a novel metabolism-related and clinically relevant lncRNA, LINC00930, which significantly influences tumor glycolysis and cell proliferation by modulating the interaction with the RBBP5 and GCN5 epigenetic remodeling complex to further alter the pattern of histone modification and transactivating the target gene PFKFB3 in NPC.
The Warburg effect, which represents a shift in the way tumor cells utilize glucose from oxidative phosphorylation to glycolysis, is now considered a major feature of tumors. This change in energy metabolism is regulated by complex factors [
43,
44]. When we explored the mechanisms by which LINC00930 regulates glycolysis, we found the involvement of PFKFB3. One of the critical role of PFKFB3 is to catalyze the conversion of F-6-P to F-2,6-BP, thus activating glycolysis and promoting tumor progression [
11,
31]. Apart from the well-documented role in glycolysis activation, recent observations have established roles of PFKFB3 beyond glycolysis. The product of PFKFB3, F-2,6-BP increases cell cycle inhibitor p27 phosphorylation at Thr-187, which activating p27 ubiquitination and degradation, and thus promoting G1/S transition [
10,
11]. In this study, we demonstrated that LINC00930 epigenetically upregulated PFKFB3 and activating glycolysis process and cell cycle progression at the G1/S phase transition, thus regulating NPC cell proliferation and tumor growth. The data consistently suggest that LINC00930 is an tumor-promotive lncRNA in NPC.
LncRNAs are involved in the regulation of gene expression at the transcriptional and post-transcriptional levels. First, lncRNAs regulate transcriptional expression by blocking promoter regions, interacting with RNA-binding proteins, or modulating the activity of transcription factors. Second, lncRNAs recruit a chromatin remodeling complex to specific sites and regulate expression processes [
5,
21,
45]. In this study, we demonstrated that LINC00930 was mainly located in the nucleus and directly interacted with chromatin remodeling proteins, RBBP5 and GCN5. Further, we established specific region within LINC00930 and key amino acid residues within RBBP5/GCN5 involved in this RNA-protein intact complex. We found that LINC00930 activated the transcription of downstream target gene PFKFB3 by regulating histone modification. This notion was verified by three lines of experimental evidence: (i) LINC00930 physically interacted with RBBP5 and GCN5; (ii) pharmacological inhibitors of histone methylation and acetylation reduced the luciferase activity of PFKFB3; (iii) LINC00930 knockdown decreased the H3K4me3 and H3K9ac levels at the promoter region of PFKFB3. Further, PFKFB3 was functionally responsible for LINC00930-mediated nasopharyngeal carcinogenesis. Collectively, our findings establish a novel mechanism of lncRNA regulating tumor metabolism, of which LINC00930 regulates the key glycolytic gene PFKFB3 by epigenetic modification.
Enhancing the understanding of the molecular mechanisms underlying NPC may promote the development of effective target therapy and improve the overall prognosis of patients with this disease [
3,
46]. In recent years, novel molecular targeted therapy has achieved remarkable curative effects in clinical practice, which shows the correctness and feasibility of tumor targeted therapy theory, thus pushing NPC therapy to a new stage. Molecular targeted therapy in combination with radiotherapy appears to be a promising event for NPC comprehensive therapy strategy [
47]. Our study reported that high expression of LINC00930 and PFKFB3 was significantly associated with a poor prognosis in NPC. Further, we demonstrated via the in vivo therapeutic models that treatment with LINC00930 inhibitor and PFKFB3 inhibitor in combination with radiotherapy induced tumor regression, suggesting that targeting LINC00930 and PFKFB3 could be an effective approach to enhance radiosensitivity of NPC patients. We are actively pursuing a clinical strategy to treat nasopharyngeal carcinoma by interfering with LINC00930 and the target gene PFKFB3.
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