Hexokinase 2 Axis Research Articles

Downregulation of hsa_circ_0006620 inhibits the malignant progression of prostate cancer by regulation of the miR-502-3p/HK2 axis mediated by aerobic glycolysis.

Circular RNAs (circRNA) are a class of covalently-closed, single-stranded RNAs that have been implicated in cancer progression due to their regulation of metabolism. However, the roles of circRNA in prostate cancer remain largely unknown. In this study, fluorescence in situ hybridization and RT-qPCR were used to investigate hsa_circ_0006620 expressions in both prostate cancer cells and tissues, after high-throughput sequencing. The luciferase reporter assay was used to identify hsa_circ_0006620 downstream targets. Transwell migration assays, 5-ethynyl-20-deoxyuridine assays, and Cell Counting Kit-8assays were used to investigate both proliferation and migration. In vivo tumorigenesis and metastasis assays were performed to investigate the role of hsa_circ_0006620 in prostate cancer. The results showed that hsa_circ_0006620 expression increased in prostate cancer cells and tissues. Hsa_circ_0006620 downregulation inhibited prostate cancer cell proliferation as well as in vivo and in vitro migrations. The luciferase results validated that miR-502-3p and hexokinase 2 (HK2) were hsa_circ_0006620 downstream targets. HK2 overexpression or miR-502-3p inhibition reversed prostate cancer cell migration after hsa_circ_0006620 silencing. The study also found that overexpression of HK2 or inhibition of prostate cancer reversed aerobic glycolysis after hsa_circ_0006620 silencing. In summary, the results showed thathsa_circ_0006620 downregulation inhibited prostate cancerby regulation of the miR-502-3p/HK2 axis mediated by aerobic glycolysis.

Hsa_circ_0087862 contributes to the progression of colorectal cancer through regulating miR-512-3p/HK2 axis

BackgroundColorectal cancer (CRC) theratened thousands of people every year. Emerging evidences suggested that circular RNAs (circRNAs) were involved in CRC malignancies. However, the underlying mechanisms have yet not been revealed. MethodsQuantitative real-time PCR (qRT-PCR) was used to determine the expression of circ_0087862 and microRNA-512–3p (miR-512–3p). Western blot was performed to measure the protein expression of hexokinase 2 (HK2), B-cell lymphoma-2 (Bcl-2), BCL2-associated X (Bax) and BCL2 antagonist/killer 1 (Bak). Moreover, 3-(4, 5-dimethyl-2-thiazolyl)-2, 5-diphenyl-2-H-tetrazolium bromide (MTT) assay, colony formation and 5-ethynyl-2’-deoxyuridine (EdU) assay were employed to assess CRC cell proliferation. Also, migration/invasion abilities and apoptosis rates were investigated by transwell assay and flow cytometry. Glucose consumption, lactate production and ATP production were detected using the corresponding kits. Dual-luciferase reporter analysis and RNA immunoprecipitation (RIP) experiments were utilized to analyze the target association of miR-512–3p and circ_0087862 or HK2. Finally, xenograft assay was carried out to analyze the function of circ_0087862 in tumor growth in vivo. ResultsCirc_0087862 expression was elevated in CRC tissues and cells. Circ_0087862 silencing repressed cell viabilities, proliferation, migration/invasion and glycolysis, and reinforced cell apoptosis. However, HK2 could weaken these impacts. Additionally, miR-512–3p targeted HK2, and circ_0087862 could regulate HK2 expression by miR-512–3p. Furthermore, circ_0087862 silencing decreased CRC cell xenograft tumor growth. ConclusionCollectively, our data suggested that circ_0087862 knockdown impeded cell viabilities, proliferation, and glycolysis, and contributed to cell apoptosis in CRC, indicating circ_0087862 as a promising tumor promoter.

The Long non-coding RNA MALAT1 functions as a competing endogenous RNA to regulate vascular remodeling by sponging miR-145-5p/HK2 in hypertension

ABSTRACT Long non-coding RNAs (LncRNAs) have been found to play a regulatory role in the pathophysiology of vascular remodeling-associated illnesses through the lncRNA-microRNA (miRNA) regulation axis. LncRNA metastasis-associated lung adenocarcinoma transcript 1 (MALAT1) is thought to be involved in proliferation, migration, apoptosis, and calcification of vascular smooth muscle cells (VSMCs). The purpose of this study was to investigate the regulatory role of MALAT1 on vascular remodeling in hypertension. Our data indicate that the expression of MALAT1 is significantly upregulated in hypertensive aortic smooth muscle. Knockdown of MALAT1 inhibited the proliferation, migration, and phenotypic transition of VSMCs induced by Ang II. Bioinformatics analysis was used to predict the complementary binding of miR-145-5p to the 3’-untranslated region of MALAT1. Besides, the expressions of MALAT1 and miR-145-5p were negatively correlated, while luciferase reporter assays and RNA immunoprecipitation assay validated the interaction between miR-145-5p and MALAT1. The proliferation, migration and phenotypic transformation of VSMCs induced by overexpression of MALAT1 were reversed in the presence of miR-145-5p. Furthermore, we verified that miR-145-5p could directly target and bind to hexokinase 2 (HK2) mRNA, and that HK2 expression was negatively correlated with miR-145-5p in VSMCs. Knockdown of HK2 significantly inhibited the effects of overexpression of MALAT1 on Ang II-induced VSMCs proliferation, migration and phenotypic transformation. Taken together, the MALAT1/miR-145-5p/HK2 axis may play a critical regulatory role in the vascular remodeling of VSMCs in hypertension.

The knockdown of lncRNA DLGAP1-AS2 suppresses osteosarcoma progression by inhibiting aerobic glycolysis via the miR-451a/HK2 axis.

Osteosarcoma (OS) is one of the most aggressive bone tumors worldwide. Emerging documents have shown that long noncoding RNAs (lncRNAs) elicit crucial regulatory functions in the process of tumorigenesis. LncRNA DLGAP1-AS2 is recognized as a regulator in several types of cancers, but its biological functions and molecular mechanisms in OS remain to be elucidated. RT-qPCR and In situ hybridization (ISH) were used to evaluate DLGAP1-AS2 expression in OS samples. Western blotting was used for the measurement of the protein levels of hexokinase 2 (HK2) and epithelial-mesenchymal transition (EMT)-related markers. The proliferation of OS cells was determined using a CCK-8 assay and EdU assay. TUNEL assay and flow cytometry were performed to assess OS cell apoptosis. Glucose metabolism in vitro assays were used. The binding relations among miR-451a, HK2, and DLGAP1-AS2 were validated by luciferase reporter assay. The cellular distribution of DLGAP1-AS2 in OS cells was determined by FISH and subcellular fractionation assays. Mouse xenograft models were established to perform the experiments in vivo. We found that DLGAP1-AS2 expression was upregulated in OS tissues and cells. Downregulation of DLGAP1-AS2 expression suppressed the malignancy of OS cells by restraining cell proliferation, the EMT process, invasiveness, migration, and aerobic glycolysis and accelerating apoptotic behaviors. Of note, silenced DLGAP1-AS2 restrained tumor growth and metastasis in vivo. However, DLGAP1-AS2 overexpression accelerated the progression of OS. We further found that DLGAP1-AS2 upregulation was induced by hypoxia and low glucose. Additionally, DLGAP1-AS2 bound to miR-451a to upregulate HK2 expression. Rescue assays revealed that the DLGAP1-AS2/miR-451a/HK2 axis contributed to OS cell malignancy by promoting aerobic glucose metabolism. Overall, these findings revealed a new regulatory pathway where DLGAP1-AS2 upregulated HK2 expression by sponging miR-451a to accelerate OS development.

Let-7b-5p inhibits breast cancer cell growth and metastasis via repression of hexokinase 2-mediated aerobic glycolysis

Hexokinase 2 (HK2), a critical rate-limiting enzyme in the glycolytic pathway catalyzing hexose phosphorylation, is overexpressed in multiple human cancers and associated with poor clinicopathological features. Drugs targeting aerobic glycolysis regulators, including HK2, are in development. However, the physiological significance of HK2 inhibitors and mechanisms of HK2 inhibition in cancer cells remain largely unclear. Herein, we show that microRNA-let-7b-5p (let-7b-5p) represses HK2 expression by targeting its 3′-untranslated region. By suppressing HK2-mediated aerobic glycolysis, let-7b-5p restrains breast tumor growth and metastasis both in vitro and in vivo. In patients with breast cancer, let-7b-5p expression is significantly downregulated and is negatively correlated with HK2 expression. Our findings indicate that the let-7b-5p/HK2 axis plays a key role in aerobic glycolysis as well as breast tumor proliferation and metastasis, and targeting this axis is a potential therapeutic strategy for breast cancer.

LncRNA DGCR5 Silencing Enhances the Radio-Sensitivity of Human Esophageal Squamous Cell Carcinoma via Negatively Regulating the Warburg Effect.

Recently, long noncoding RNAs (lncRNAs) and the Warburg effect have been reported to play important roles in the radio-sensitivity of tumor cells. Survival correlates with pathologic responses to chemoradiotherapy and improving responses to radiation may translate into improved survival. This study aims to examine the effects and mechanisms of lncRNA DGCR5 and the Warburg effect on ESCC cell radiosensitivity. Levels of DGCR5, miR-195 and hexokinase 2 (HK2) expression in ESCC tissues and cells were determined and their clinical significance was analyzed. TE-1 and KYSE150 cells received a 6 Gy dose of X-ray radiation and their survival, proliferation and apoptosis were evaluated using colony formation assays, CCK-8 assays, and flow cytometry, respectively. Lactic acid production and glucose consumption were also examined in both cell types. Finally, the expression of apoptotic proteins was assessed using Western blotting. Analysis revealed that DGCR5 and HK2 were overexpressed in ESCC, while miR-195 was under expressed. Moreover, it was demonstrated that down-regulation of DGCR5 inhibited cell proliferation and promoted apoptosis, resulting in increased radiosensitivity by inhibition of the Warburg Effect. Conversely, overexpression of DGCR5 exhibited an opposite phenomenon in vitro. When investigating the mechanism, we identified that miR-195 was predicted to be a direct downstream target of DGCR5. Meanwhile, HK2 was predicted to be a direct downstream target of miR-195. Dual-luciferase reporter assays verified the direct interaction between these molecules. Finally, in vivo experiments were utilized to validate that knockdown of DGCR5 suppressed the Warburg effect via targeting of the miR-195/HK2 axis to increase the radiosensitivity of ESCC. Our study reveals that down-regulation of DGCR5 resulted in inhibition of the Warburg effect through interaction with the miR-195/HK2 axis increasing ESCC cell apoptosis after irradiation, thus enhancing cell radiosensitivity.

Circ-CCS regulates oxaliplatin resistance via targeting miR-874-3p/HK2 axis in colorectal cancer.

Colorectal cancer (CRC) is a malignancy that threatens the patient's life. Previous reports showed that circular RNAs (circRNAs) can affect CRC development. Herein, we demonstrated the characters of circular RNA copper chaperone for superoxide dismutase (circ-CCS) in CRC tissues and cells. Circ-CCS, CCS mRNA, microRNA-874-3p (miR-874-3p) and hexokinase 2 (HK2) were indicated by qRT-PCR and western blot in CRC. The cell roles were examined. Additionally, the interaction between miR-874-3p and circ-CCS or HK2 was forecasted by the bioinformatics method and assessed by dual-luciferase reporter assay. Finally, the mouse test was implemented to demonstrate the effect of circ-CCS in vivo. Circ-CCS and HK2 were increased, whereas miR-874-3p was diminished in CRC. Circ-CCS lack subdued the IC₅₀ value of oxaliplatin, cell proliferation, migration, invasion and glycolysis metabolism in CRC cells, while it endorsed cell apoptosis. Furthermore, miR-874-3p was validated as having a tumor repressive effect in CRC cells by restraining HK2. The results also showed that HK2 could regulate the development of CRC. In mechanism, circ-CCS targeted miR-874-3p to control HK2. In addition, circ-CCS knock-down also attenuated tumor growth in mice. Circ-CCS expedited CRC through miR-874-3p/HK2.

PVT1/miR-145-5p/HK2 modulates vascular smooth muscle cells phenotype switch via glycolysis: The new perspective on the spiral artery remodeling

IntroductionVascular smooth muscle cells (VSMC) switched from a contractile phenotype to a synthetic phenotype during the decidual spiral artery (SPAs) remodeling process. The lncRNA plasmacytoma variant translocation 1 (PVT1) and glucose metabolism have been found to regulate the VSMC phenotype switch. This study aimed to analyze the dynamic expression of PVT1 and glycolytic key enzymes hexokinase2 (HK2) at different remodeling stages in early human pregnancy and elucidate the underlying mechanism of the PVT1/miR-145-5p/HK2 axis involved in the spiral artery remodeling. MethodsqRT-PCR, Western blot (WB) analysis, Immunohistochemistry (IHC) and fluorescence in situ hybridization (FISH) were used to detect the expression and localization of PVT1 and HK2 in decidual tissue. HA-VSMCs were transfected with specific siRNA, shRNA and plasmids to regulate corresponding genes. Extracellular lactate, cellular ATP, ROS, and intracellular NADPH levels were measured using the corresponding assay kits. Migration was measured by wound-healing and Transwell assays. Contractile phenotypic markers α-SMA, MYH11 with calponin and synthetic phenotypic markers OPN and vimentin were detected by WB. The PDC model was used to detect the degree of spiral arterial remodeling. ResultsPVT1 and HK2 were upregulated with gestational age (GA) increasing in decidual tissue during the early pregnancy. HK2 regulated the glycolytic activity and VSMC phenotype switch in vitro. PVT1 regulated the glycolytic activity and VSMC phenotype switch through HK2. PVT1 played a ceRNA role in regulating HK2 expression by sponging miR-145-5p. PVT1 and HK2 influenced spiral artery remodeling in the PDC model. DiscussionPVT1 and HK2 were upregulated, and miR-145-5p was downregulated in decidua with the GA increasing. Meanwhile, the PVT1/miR-145-5p/HK2 axis may be involved in regulating the phenotypic switch and migratory capacity of VSMCs by affecting glycolysis in decidual SPAs remodeling.

Downregulation of Circ-PITHD1 Suppressed Colorectal Cancer via Glycolysis Inhibition through miR-590-5p/HK2 Axis.

Colorectal cancer (CRC) is a frequent malignancy around the globe. Circular RNAs (circRNAs) are implicated in CRC development. Nevertheless, the regulatory mechanisms and biological functions regarding circRNAs in CRC progression are largely unclear. The present investigation employed next-generation sequencing (NGS) to study the abnormal circRNA expression in CRC tissues. The regulatory mechanism and targets were then analyzed by bioinformatics, luciferase reporter analysis, CCK8, colony formation, and Transwell migration. In vivo metastasis and tumorigenesis assays were conducted to elucidate circ-PITHD1 roles regarding CRC. The data showed that circ-PITHD1 expression increased in a CRC cell line and tissues, which indicated that circ-PITHD1 functioned in CRC progression. circ-PITHD1 downregulation inhibited CRC invasion and proliferation in the experiments. Luciferase reporter results confirmed that both miR-590-5p and hexokinase 2 (HK2) were circ-PITHD1 downstream targets. HK2 overexpression or miR-590-5p suppression reversed CRC cell proliferation and invasion after silencing of circ-PITHD1 by regulation of glycolysis. Taken together, this investigation discovered that circ-PITHD1 downregulation suppressed CRC progression by inhibiting glycolysis via the miR-590-5p/HK2 axis.

Circ_0002762 Accelerates Glycolysis Metabolism to Promote Cervical Cancer Progression via the miR-526b-5p/HK2 Axis

Objectives: The purpose of this study was to investigate the role and mechanism of circ_0002762 in CC. Design: This study was designed for silencing circ_0002762 in CC cells and xenograft tumor models to investigate the role of circ_0002762 in CC in vitro and in vivo. Materials and Methods: The relative expression levels of circ_0002762, miR-526b-5p, and hexokinase2 (HK2) in CC tissues and cells were detected by real-time quantitative polymerase chain reaction or Western blot. Glycolysis-related extracellular acidification rate, glucose production, lactic acid consumption, and ATP levels were measured using the appropriate kits. Cell proliferation was assessed by 5-ethynyl-2′-deoxyuridine and colony formation assay. Cell apoptosis was detected by flow cytometry. The binding relationship between miR-526b-5p and circ_0002762 or HK2 was verified by dual-luciferase reporter assay and RNA pull-down assay. Tumor growth in vivo was detected by xenograft tumor model. Results: The expressions of circ_0002762 and HK2 were up-regulated and miR-526b-5p was down-regulated in CC tissues and cells. Circ_0002762 knockdown inhibited glycolysis and proliferation and promoted apoptosis of CC cells. In addition, miR-526b-5p suppression reversed the inhibition of CC development induced by circ_0002762 silencing. HK2 overexpression eliminated the inhibition of miR-526b-5p on CC progression. Moreover, silencing of circ_0002762 inhibited CC tumor growth in vivo. Limitations: The practical application of circ_0002762 in clinical practice needs further investigation. Conclusion: Circ_0002762 knockdown inhibited CC progression by regulating miR-526b-5p/HK2 axis, suggesting that circ_0002762 was a promising therapeutic strategy for CC.

Long noncoding RNA CASC7 is a novel regulator of glycolysis in oesophageal cancer via a miR-143-3p-mediated HK2 signalling pathway

Long noncoding RNAs have been proven to play a crucial role in many tumours. Here, we explored the role of the lncRNA cancer susceptibility candidate 7 (CASC7) in oesophageal cancer. LncRNA CASC7 was identified in our database analysis, and we found that it was significantly higher in oesophageal tumour tissue than in normal tissue and that high expression of lncRNA CASC7 predicted a poor prognosis. Furthermore, we verified through cell experiments that low expression of lncRNA CASC7 in oesophageal cancer cells significantly inhibited tumour proliferation, which could be explained by the effect of lncRNA CASC7 on aerobic glycolysis. Next, we found that the expression of CASC7 and hexokinase 2 (HK2) in oesophageal cancer was positively correlated in database analysis, and this conclusion was further verified in cell experiments. To determine the mechanism, we found that miR-143-3p can bind to both lncRNA CASC7 and HK2. In clinical specimens, we also found high expression of lncRNA CASC7 in tumours, and the expression levels of lncRNA CASC7 and HK2 were positively correlated. In conclusion, downregulating lncRNA CASC7 could inhibit tumour proliferation by reducing glycolysis through the miR-143-3p/HK2 axis.

Long Non-coding RNA LOXL1-AS1 Facilitates Colorectal Cancer Progression via Regulating miR-1224-5p/miR-761/HK2 Axis.

Colorectal cancer (CRC) is one of the most common cancers worldwide. Long non-coding RNAs (lncRNAs) play crucial roles in the development of malignant tumors. The present study aimed to explore the function and potential mechanism of lncRNA LOXL1 antisense RNA 1 (LOXL1-AS1) in CRC. The abundance of LOXL1-AS1, miR-1224-5p, miR-761, and hexokinase 2 (HK2) was detected by quantitative real-time PCR or western blot assay. Cell proliferation was assessed by Cell Counting Kit-8 and colony formation assays. Cell apoptosis, invasion, and migration were examined by flow cytometry, transwell assay, and wound healing assay. Glycolysis was evaluated by detecting glucose consumption, lactate production, and ATP/ADP ratios. Xenograft assay was used for in vivo tumor growth analysis. LOXL1-AS1 and HK2 levels were increased, while miR-1224-5p and miR-761 levels were reduced in CRC tissues and cells. Knockdown of LOXL1-AS1 suppressed CRC cell proliferation, invasion, migration, and glycolysis, and induced cell apoptosis. Silencing of LOXL1-AS1 blocked tumor growth in vivo. Moreover, LOXL1-AS1 accelerated CRC cell progression by absorbing miR-1224-5p/miR-761. Besides, miR-1224-5p and miR-761 inhibited CRC cell progression via targeting HK2. LOXL1-AS1 contributed to CRC progression via modulating miR-1224-5p/miR-761/HK2 pathway.

Disrupting Circadian Rhythm via the PER1-HK2 Axis Reverses Trastuzumab Resistance in Gastric Cancer.

In trastuzumab-resistant HER2-positive gastric cancer, glycolysis fluctuates with a circadian oscillation regulated by the BMAL1-CLOCK-PER1-HK2 axis, which can be disrupted with a metformin-based chronotherapy to overcome trastuzumab resistance.

RETRACTED ARTICLE: Circ-NUP214 Promotes Papillary Thyroid Carcinoma Tumorigenesis by Regulating HK2 Expression Through miR-15a-5p.

Circular RNA nucleoporin 214 (circ-NUP214) is a novel-identified circRNA, and has been demonstrated to promote cell tumorigenesis in papillary thyroid carcinoma (PTC). However, the detailed roles and molecular mechanism underlying circ-NUP214 in PTC remain unclear. The expression levels of circ-NUP214, microRNA (miR)-15a-5p, and hexokinase 2 (HK2) were detected by quantitative real-time polymerase chain reaction, western blot or immunohistochemical (IHC) assay. Cell proliferation, apoptosis, migration and invasion were analyzed using cell counting kit-8 assay, 5-Ethynyl-2'-Deoxyuridine (EdU), flow cytometry, and transwell assay, respectively. Glucose metabolism was assessed by analyzing glucose consumption, lactate production, and adenosine triphosphate (ATP)/adenosine diphosphate (ADP) ratios. Murine xenograft models were established for in vivo experiments. The interaction between miR-15a-5p and circ-NUP214 or HK2 was analyzed by the dual-luciferase reporter assay. A significant increase of circ-NUP214 expression in PTC tissues and cells was found. Circ-NUP214 knockdown inhibited cell proliferation, migration, invasion and glycolysis but induced apoptosis in PTC in vitro and reduced tumor growth in vivo. MiR-15a-5p was a target of circ-NUP214, and silencing of miR-15a-5p reversed the inhibitory effects of circ-NUP214 knockdown on PTC cell malignant phenotypes and glycolysis. MiR-15a-5p directly targeted HK2, miR-15a-5p-mediated inhibition of cell proliferation, migration, invasion and glycolysis and the promotion of cell apoptosis in PTC were abolished by HK2 overexpression. Besides that, circ-NUP214 indirectly regulated HK2 expression via miR-15a-5p. Circ-NUP214 promoted PTC tumorigenesis by regulating miR-15a-5p/HK2 axis, suggesting a potential therapeutic target for PTC treatment.

Methylation-associated silencing of miR-9-1 promotes nasopharyngeal carcinoma progression and glycolysis via HK2.

Characteristically, cancer cells metabolize glucose through aerobic glycolysis, known as the Warburg effect. Accumulating evidence suggest that during cancer formation, microRNAs (miRNAs) could regulate such metabolic reprogramming. In the present study, miR‐9‐1 was identified as significantly hypermethylated in nasopharyngeal carcinoma (NPC) cell lines and clinical tissues. Ectopic expression of miR‐9‐1 inhibited NPC cell growth and glycolytic metabolism, including reduced glycolysis, by reducing lactate production, glucose uptake, cellular glucose‐6‐phosphate levels, and ATP generation in vitro and tumor proliferation in vivo. HK2 (encoding hexokinase 2) was identified as a direct target of miR‐9‐1 using luciferase reporter assays and Western blotting. In NPC cells, hypermethylation regulates miR‐9‐1 expression and inhibits HK2 translation by directly targeting its 3' untranslated region. MiR‐9‐1 overexpression markedly reduced HK2 protein levels. Restoration of HK2 expression attenuated the inhibitory effect of miR‐9‐1 on NPC cell proliferation and glycolysis. Fluorescence in situ hybridization results indicated that miR‐9‐1 expression was an independent prognostic factor in NPC. Our findings revealed the role of the miR‐9‐1/HK2 axis in the metabolic reprogramming of NPC, providing a potential therapeutic strategy for NPC.

Silencing of circHIPK3 Sensitizes Paclitaxel-Resistant Breast Cancer Cells to Chemotherapy by Regulating HK2 Through Targeting miR-1286.

BackgroundResistance development to paclitaxel (PTX) has become a major obstacle in the successful treatment of breast cancer (BC). Circular RNAs (circRNAs) have been identified as essential regulators in PTX resistance of BC. Here, we explored the precise roles of circRNA homeodomain interacting protein kinase 3 (circHIPK3, circ_0000284) in PTX resistance of BC.MethodsThe expression levels of circHIPK3, microRNA (miR)-1286, and hexokinase 2 (HK2) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot. Ribonuclease R (RNase R) assay was used to confirm the stability of circHIPK3. Cellular localization of circHIPK3 was assessed by subcellular localization assay. The half maximal inhibitory concentration (IC50) value for PTX was measured by Cell Counting Kit-8 (CCK-8) assay. Cell colony formation, cell cycle distribution, and apoptosis were gauged by colony formation assay and flow cytometry, respectively. Animal studies were performed to evaluate the role of circHIPK3 in vivo. The direct relationship between miR-1286 and circHIPK3 or HK2 was verified by dual-luciferase reporter and RNA immunoprecipitation (RIP) assays.ResultsOur results showed that circHIPK3 was up-regulated in PTX-resistant BC tissues and cells compared with the sensitive counterparts. The silencing of circHIPK3 promoted PTX sensitivity of PTX-resistant BC cells in vitro and in vivo. CircHIPK3 directly targeted miR-1286, and miR-1286 acted as a downstream mediator of circHIPK3 function in vitro. HK2 was a direct target of miR-1286, and circHIPK3 modulated HK2 expression through miR-1286. The increased expression of miR-1286 sensitized PTX-resistant BC cells to PTX in vitro by down-regulating HK2.ConclusionOur findings demonstrated that the silencing of circHIPK3 sensitized PTX-resistant BC cells to PTX therapy at least in part via the regulation of the miR-1286/HK2 axis.

Knockdown of circBFAR inhibits proliferation and glycolysis in gastric cancer by sponging miR-513a-3p/hexokinase 2 axis

The relationship between circular RNAs (circRNAs) and many types of cancer has been of great interest. A novel circRNA, circBFAR, has been identified, but the functions of circBFAR and its underlying mechanism in gastric cancer (GC) have not been reported. This study was designed to investigate the role of circBFAR in GC and its downstream miRNA targets. Quantitative real-time polymerase reaction was used to detect the expression of circBFAR and miRNAs. Cell counting kit-8 and EdU were used to detect the proliferation of GC cells. Measurement of the extracellular acidification rate, oxygen consumption rate and lactate acid production were performed to assess the glycolysis levels. The results showed that circBFAR exhibited higher expression in GC tissues and cell lines. circBFAR was proven to promote GC proliferation by targeting the miR-513a-3p/hexokinase 2 (HK2) axis. Inhibition of circBFAR also led to a significant decrease in the glycolysis levels. In this study, we found a circBFAR/miR-513a-3p/HK2 axis in GC and revealed the relationship between circBFAR and glycolysis for the first time. circBFAR may serve as a novel target of GC individualized therapy.

Long Noncoding RNA LINC01410 Suppresses Tumorigenesis and Enhances Radiosensitivity in Neuroblastoma Cells Through Regulating miR-545-3p/HK2 Axis.

BackgroundAbnormal expression of long noncoding RNAs (lncRNAs) was often involved in tumorigenesis and radiosensitivity of various cancers. The aim of this study was to explore the biological function and regulatory mechanism of lncRNA long intergenic non-protein coding RNA 1410 (LINC01410) in tumorigenesis and radiosensitivity of neuroblastoma (NB).MethodsThe expression of LINC01410, microRNA-329-3p (miR-545-3p) and hexokinase 2 (HK2) was detected by quantitative real-time polymerase chain reaction (qRT-PCR). Methylthiazolyldiphenyl tetrazolium bromide (MTT) assay, colony formation assay and transwell assay were utilized to detect cell viability, colony formation and cell invasion abilities. Glucose consumption or lactate production was measured by glucose assay kit or lactate assay kit, respectively. The interaction between miR-545-3p and LINC01410 or HK2 was predicted by starBase v2.0 and verified by dual-luciferase reporter, RNA Immunoprecipitation (RIP) and RNA pull-down assays. Western blot was used to measure the protein expression of HK2. The mice xenograft model was established to investigate the role of LINC01410 in vivo.ResultsLINC01410 and HK2 were highly expressed while miR-545-3p was lowly expressed in NB tissues and cells. LINC01410 knockdown inhibited tumorigenesis by repressing cell proliferation and invasion, and increased the radiosensitivity via inhibiting colony formation rates and glycolysis. LINC01410 knockdown also suppressed tumor growth in vivo. Moreover, miR-545-3p could bind to LINC01410 and its downregulation reversed the effects of LINC01410 knockdown on tumorigenesis and radiosensitivity. Additionally, HK2 was a direct target of miR-545-3p and its overexpression attenuated the effects of miR-545-3p restoration on suppression of tumorigenesis and promotion of radiosensitivity. Besides, LINC01410 functioned as a molecular sponge of miR-545-3p to regulate HK2 expression.ConclusionLINC01410 interference inhibited tumorigenesis and increased radiosensitivity via regulating miR-545-3p/HK2 axis, providing a novel therapeutic strategy for NB.

Serum Exosomes-Based Biomarker circ_0008928 Regulates Cisplatin Sensitivity, Tumor Progression, and Glycolysis Metabolism by miR-488/HK2 Axis in Cisplatin-Resistant Nonsmall Cell Lung Carcinoma.

Background: Nonsmall cell lung carcinoma (NSCLC) is a major cause of cancer-related death worldwide. The resistance of NSCLC to chemical drugs, such as cisplatin (CDDP), poses a heavy burden for NSCLC therapy. Herein, the effects of circular_0008928 (circ_0008928) on the CDDP sensitivity and biological behavior of CDDP-resistant NSCLC cells and underlying mechanism are revealed. Materials and Methods: The expression of circ_0008928 and microRNA-488 (miR-488) was detected by quantitative real-time polymerase chain reaction. The expression of hexokinase 2 (HK2) protein and exosome-specific proteins was determined by Western blot. The half-maximal inhibitory concentration (IC50) value of CDDP was detected by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Cell proliferation and migratory and invasive abilities were illustrated by cell counting kit-8 and transwell assays. Cell glycolysis metabolism was illustrated by extracellular acidification rate assay, glucose kit and lactate kit assays and Western blot analysis. The binding sites between miR-488 and circ_0008928 or HK2 were predicted by starbase or microT-CDS online database, and identified by dual-luciferase reporter and RNA immunoprecipitation assays. Results: Circ_0008928 expression and HK2 protein expression were significantly upregulated, while miR-488 expression was obviously downregulated in NSCLC cells and CDDP-resistant NSCLC cells. Circ_0008928 expression was increased in serum exosomes of CDDP-resistant NSCLC patients compared with CDDP-sensitive NSCLC patients. In addition, circ_0008928 silencing improved CDDP sensitivity and attenuated CDDP-induced cell proliferation, migration, invasion, and glycolysis metabolism. Circ_0008928 was a sponge of miR-488, and miR-488 bound to HK2 in CDDP-resistant NSCLC cells. Furthermore, both miR-488 inhibitor and HK2 overexpression attenuated circ_0008928 absence-mediated impacts on CDDP sensitivity and tumor process in CDDP-resistant NSCLC. Conclusions: Circ_0008928 knockdown improved CDDP sensitivity and hindered cell proliferation, migration, invasion, and glycolysis metabolism by miR-488/HK2 axis in CDDP-resistant NSCLC. This finding provides a new mechanism for studying CDDP-resistant therapy in NSCLC.

LncRNA ZFAS1/miR-1271-5p/HK2 Promotes Glioma Development Through Regulating Proliferation, Migration, Invasion and Apoptosis.

Glioma is the common type of malignant tumor with high mortality worldwide. Survival rate of patients with glioma remains poor, and almost half of patients died within 15 months. The increasing researches indicated that long non-coding RNA (lncRNA) Zinc finger antisense 1 (ZFAS1) played essential roles in tumor initiation and progression. Therefore, it is worth clarifying potential role of ZFAS1 in glioma. The expression levels of ZFAS1, miR-1271-5p, and Hexokinase 2 (HK2) in glioma tissues and cells were examined by real-time quantitative polymerase chain reaction (RT-qPCR). Kaplan-Meier curve was employed to analyze the relationship between cumulative survival time of glioma patients and expression level of ZFAS1. The cell proliferation, apoptosis, and mobility ability were assessed with 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl-2H-tetrazol-3-ium bromide (MTT), flow cytometry, and transwell assays. The relationship among ZFAS1, miR-1271-5p, and HK2 was analyzed by bioinformatics assay, dual-luciferase reporter and Pearson's correlation analyses. The protein expression level of HK2 was examined by western blot assay. Finally, a xenograft experiment was established to assess the effects of ZFAS1 silencing in vivo. ZFAS1 was highly expressed in glioma tissues and cells, besides, the expression level of ZFAS1 was associated with survival time of glioma patients. Functional experiments suggested that knockdown of ZFAS1 or upregulation of miR-1271-5p constrained proliferation, migration and induced apoptosis of glioma cells. In addition, miR-1271-5p, interacted with HK2, was a target of ZFAS1. The gain of HK2 could overturn ZFAS1 silencing-induced effects on glioma cells. Besides, deficiency of ZFAS1 hindered tumor growth in vivo. ZFAS1 was involved in proliferation, migration, and apoptosis of glioma cells via regulating miR-1271-5p/HK2 axis.