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The cellular thermal shift assay (CETSA) and isothermal dose-response fingerprint assay (ITDRF CETSA) have been introduced as powerful tools for investigating target engagement by measuring ligand-triggered thermodynamic stabilization of cellular target proteins. Yet, these techniques have rarely been used to evaluate the thermal stability of RNA-binding proteins (RBPs) when exposed to ligands. Here, we present an adjusted approach using CETSA and ITDRFCETSA to determine the interaction between enasidenib and RBM45. Our assay is sensitive and time-efficient and can potentially be adapted for studying the interactions of RBM45 protein with other potential candidates. Key features ⢠This protocol builds upon the method developed by Molina et al. and extends its application to new protein classes, such as RBPs.
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BACKGROUND: Phosphatidylinositol-4-phosphate 5-kinase type 1 alpha (PIP5K1A) acts upstream of the Akt regulatory pathway and is abnormally expressed in many types of malignancies. However, the role and mechanism of PIP5K1A in colorectal cancer (CRC) have not yet been reported. In this study, we aimed to determine the association between PIP5K1A and progression of CRC and assess the efficacy and mechanism by which rupatadine targets PIP5K1A. METHODS: Firstly, expression and function of PIP5K1A in CRC were investigated by human colon cancer tissue chip analysis and cell proliferation assay. Next, rupatadine was screened by computational screening and cytotoxicity assay and interactions between PIP5K1A and rupatadine assessed by kinase activity detection assay and bio-layer interferometry analysis. Next, rupatadine's anti-tumor effect was evaluated by in vivo and in vitro pharmacodynamic assays. Finally, rupatadine's anti-tumor mechanism was explored by quantitative real-time reverse-transcription polymerase chain reaction, western blot, and immunofluorescence. RESULTS: We found that PIP5K1A exerts tumor-promoting effects as a proto-oncogene in CRC and aberrant PIP5K1A expression correlates with CRC malignancy. We also found that rupatadine down-regulates cyclin-dependent kinase 2 and cyclin D1 protein expression by inhibiting the PIP5K1A/Akt/GSK-3ß pathway, induces cell cycle arrest, and inhibits CRC cell proliferation in vitro and in vivo. CONCLUSIONS: PIP5K1A is a potential drug target for treating CRC. Rupatadine, which targets PIP5K1A, could serve as a new option for treating CRC, its therapeutic mechanism being related to regulation of the Akt/GSK-3ß signaling pathway.
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Proliferação de Células , Neoplasias Colorretais , Ciproeptadina , Fosfotransferases (Aceptor do Grupo Álcool) , Proteínas Proto-Oncogênicas c-akt , Transdução de Sinais , Humanos , Proliferação de Células/efeitos dos fármacos , Neoplasias Colorretais/tratamento farmacológico , Neoplasias Colorretais/patologia , Neoplasias Colorretais/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Animais , Transdução de Sinais/efeitos dos fármacos , Fosfotransferases (Aceptor do Grupo Álcool)/metabolismo , Ciproeptadina/farmacologia , Ciproeptadina/análogos & derivados , Camundongos Nus , Linhagem Celular Tumoral , Camundongos Endogâmicos BALB C , Masculino , Proto-Oncogene Mas , Ensaios Antitumorais Modelo de Xenoenxerto , Camundongos , Antineoplásicos/farmacologiaRESUMO
Invasive pituitary neuroendocrine tumors (PitNETs) are the most prevalent types of intracranial and neuroendocrine tumors. Their aggressive growth and difficulty in complete resection result in a high recurrence rate. Cystine transporter solute carrier family 7 member 11 (SLC7A11) is overexpressed in various cancers, which contributes to tumor growth, progression, and metastasis by promoting cystine uptake and glutathione biosynthesis. We identified SLC7A11 as an invasive biomarker based on three Gene Expression Omnibus cohorts. This study aimed to investigate the role of SLC7A11 in invasive PitNETs. Cell proliferation was assessed using CCK-8 and colony formation assays, while cell apoptosis was estimated with flow cytometry. Wound healing assays and transwell assays were utilized to evaluate migration and invasion ability. Our findings demonstrated that SLC7A11 was markedly upregulated in invasive PitNETs, and was associated with the invasiveness of PitNETs. Knockdown of SLC7A11 could largely suppress tumor cell proliferation, migration, and invasion, while inducing apoptosis. Furthermore, SLC7A11 depletion was implicated in regulating epithelial-mesenchymal transition and inactivating the PI3K/AKT signaling pathway. These insights suggest SLC7A11 as a potential therapeutic target for invasive PitNETs.
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The therapeutic efficacy of adoptive T cell therapy is largely restricted by reduced viability and dysfunction of CD8+ T cells. Continuous antigen stimulation disrupts the expansion, effector function, and metabolic fitness of CD8+ T cells, leading to their differentiation into an exhausted state within the tumor microenvironment (TME). While the function of the cell cycle negative regulator p16 in senescent cells is well understood, its role in T cell exhaustion remains unclear. In this study, we demonstrated that TCR stimulation of CD8+ T cells rapidly upregulates p16 expression, with its levels positively correlating with TCR affinity. Chronic TCR stimulation further increased p16 expression, leading to CD8+ T cell apoptosis and exhaustion differentiation, without inducing DNA damage or cell senescence. Mechanistic investigations revealed that p16 downregulates mTOR, glycolysis, and oxidative phosphorylation (OXPHOS) associated gene expression, resulting in impaired mitochondrial fitness, reduced T cell viability, and diminished effector function. Furthermore, the deletion of p16 significantly enhances the persistence of CD8+ T cells within tumors and suppresses the terminal exhaustion of tumor-infiltrating T cells. Overall, our findings elucidate how increased p16 expression reshapes T cell intracellular metabolism, drives T cell apoptosis and exhaustion differentiation, and ultimately impairs T cell anti-tumor function.
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Apoptose , Linfócitos T CD8-Positivos , Inibidor p16 de Quinase Dependente de Ciclina , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/genética , Animais , Camundongos , Humanos , Camundongos Endogâmicos C57BL , Microambiente Tumoral/imunologia , Diferenciação Celular , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos do Interstício Tumoral/imunologia , Linfócitos do Interstício Tumoral/metabolismo , Glicólise , Exaustão das Células TRESUMO
Aim of the study: To investigate the anti-tumor effects of Lasiokaurin on breast cancer and explore its underlying molecular mechanism. Materials and methods: In this study, MTT assay, plate colony formation assays, soft agar assay, and EdU assay were employed to evaluate the anti-proliferation effects of LAS. Apoptosis and cell cycle distribution were detected by flow cytometry. The molecular mechanism was predicted by performing RNA sequencing and verified by using immunoblotting assays. Breast cancer organiods derived from patient-derived xenografts model and MDA-MB-231 xenograft mouse model were established to assess the effect of LAS. Results: Our study showed that LAS treatment significantly suppressed cell viability of 5 breast cancer cell lines, with the IC50 value of approximately 1-5 µM. LAS also inhibitied the clonogenic ability and DNA synthesis of breast cancer cells, Moreover, LAS induced apoptosis and G2/M cell cycle arrest in SK-BR-3 and MDA-MB-231 cells. Notably, transcriptomic analysis predicted the mechanistic involvement of PLK1 in LAS-suppressed breast cancer progression. Our experiment data further verified that LAS reduced PLK1 mRNA and protein expression in breast cancer, accompanied by downregulating CDC25C and AKT phosphorylation. Ultimately, we confirmed that LAS inhibit breast cancer growth via inhibiting PLK1 pathway in vivo. Conclusions: Collectively, our findings revealed that LAS inhibits breast cancer progression via regulating PLK1 pathway, which provids scientific evidence for the use of traditional Chinese medicine in cancer therapy.
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Background: Arnicolide C, which is isolated from Centipeda minima, has excellent antitumor effects. However, the potential impacts and related mechanisms of action of arnicolide C in breast cancer remain unknown. Methods: The viability of breast cancer cells was measured using MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and colony formation assays. For analysis of apoptosis and the cell cycle, flow cytometry was used. A molecular docking approach was used to explore the possible targets of arnicolide C. Western blot analysis was used to detect changes in the expression of 14-3-3θ and proteins in related pathways after arnicolide C treatment in breast cancer cells. The anti-breast cancer effect of arnicolide C in vivo was evaluated by establishing cell-derived xenograft (CDX) and patient-derived xenograft (PDX) models. Results: Arnicolide C inhibited proliferation, increased apoptosis, and induced G1 arrest. In particular, molecular docking analysis indicated that arnicolide C binds to 14-3-3θ. Arnicolide C reduced 14-3-3θ expression and inhibited its downstream signaling pathways linked to cell proliferation. Similar results were obtained in the CDX and PDX models. Conclusion: Arnicolide C can have an anti-breast cancer effect both in vitro and in vivo and can induce cell cycle arrest and increase apoptosis in vitro. The molecular mechanism may be related to the effect of arnicolide C on the expression level of 14-3-3θ. However, the specific mechanism through which arnicolide C affects 14-3-3θ protein expression still needs to be determined.
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Pancreatic cancer is one of the most lethal cancer types with 5-year survival rate of â¼10.8%. Various KRAS mutations exist in â¼85% pancreatic cancer cell lines. Mutated KRAS is a major cause that leads cancer cell proliferation. Chemotherapy is still the major treatment for pancreatic cancer. Alternatively, repositioning old drug to inhibit mutated KRAS may be a cost-effective way for pancreatic cancer treatment. In this study, we choose mutated KRAS (G12D) as a target. Based on mutated KRAS GTP binding domain (hydrolyze GTP to GDP), we perform virtual screening on FDA-approved drugs. Montelukast shows strong binding affinity to mutated KRAS as well as interfering both GTP and GDP binding to mutated KRAS. Furthermore, Montelukast shows very strong anti-proliferation effect on mutated KRAS pancreatic cancer cells both in vitro and in vivo. Our results support repositioning of Montelukast as single agent for pancreatic cancer treatment.
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Neoplasias Pancreáticas , Proteínas Proto-Oncogênicas p21(ras) , Humanos , Proteínas Proto-Oncogênicas p21(ras)/genética , Reposicionamento de Medicamentos , Neoplasias Pancreáticas/tratamento farmacológico , Neoplasias Pancreáticas/genética , Neoplasias Pancreáticas/metabolismo , Mutação , Proliferação de Células , Guanosina Trifosfato/uso terapêuticoRESUMO
BACKGROUND: The microtubule protein inhibitor C118P shows excellent anti-breast cancer effects. However, the potential targets and mechanisms of C118P in breast cancer remain unknown. METHODS: Real-time cellular analysis (RTCA) was used to detect cell viability. Apoptosis and the cell cycle were detected by flow cytometry. Computer docking simulations, surface plasmon resonance (SPR) technology, and microscale thermophoresis (MST) were conducted to study the interaction between C118P and alanine-serine-cysteine transporter 2 (ASCT2). Seahorse XF technology was used to measure the basal oxygen consumption rate (OCR). The effect of C118P in the adipose microenvironment was explored using a co-culture model of adipocytes and breast cancer cells and mouse cytokine chip. RESULTS: C118P inhibited proliferation, potentiated apoptosis, and induced G2/M cell cycle arrest in breast cancer cells. Notably, ASCT2 was validated as a C118P target through reverse docking, SPR, and MST. C118P suppressed glutamine metabolism and mediated autophagy via ASCT2. Similar results were obtained in the adipocyte-breast cancer microenvironment. Adipose-derived interleukin-6 (IL-6) promoted the proliferation of breast cancer cells by enhancing glutamine metabolism via ASCT2. C118P inhibited the upregulation of ASCT2 by inhibiting the effect of IL-6 in co-cultures. CONCLUSION: C118P exerts an antitumour effect against breast cancer via the glutamine transporter ASCT2.
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Targeting metabolic remodeling represents a potentially promising strategy for hepatocellular carcinoma (HCC) therapy. In-depth understanding on the regulation of the glutamine transporter alanine-serine-cysteine transporter 2 (ASCT2) contributes to the development of novel promising therapeutics. As a developmentally regulated RNA binding protein, RBM45 is capable to shuttle between nucleus and cytoplasm, and directly interacts with proteins. By bioinformatics analysis, we screened out that RBM45 was elevated in the HCC patient specimens and positively correlated with poor prognosis. RBM45 promoted cell proliferation, boosted xenograft tumorigenicity and accelerated HCC progression. Using untargeted metabolomics, it was found that RBM45 interfered with glutamine metabolism. Further results demonstrated that RBM45 positively associated with ASCT2 in human and mouse specimens. Moreover, RBM45 enhanced ASCT2 protein stability by counteracting autophagy-independent lysosomal degradation. Significantly, wild-type ASCT2, instead of phospho-defective mutants, rescued siRBM45-suppressed HCC cell proliferation. Using molecular docking approaches, we found AG-221, a mutant isocitrate dehydrogenase 2 (mIDH2) inhibitor for acute myeloid leukemia therapy, pharmacologically perturbed RBM45-ASCT2 interaction, decreased ASCT2 stability and suppressed HCC progression. These findings provide evidence that RBM45 plays a crucial role in HCC progression via interacting with and counteracting the degradation of ASCT2. Our findings suggest a novel alternative structural sites for the design of ASCT2 inhibitors and the agents interfering with RBM45-ASCT2 interaction may be a potential direction for HCC drug development.
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Hepatocellular carcinoma (HCC) represents one of the primary liver malignancies with poor prognosis. RHNO1, which implicated in the ATR-CHK1 signaling pathway thus functions in the DNA replication stress response. However, the role and molecular mechanisms of RHNO1 in HCC remain largely elusive. Here, we imply that RHNO1 is elevated in HCC tumor tissues and that high expression of RHNO1 predicts poor prognosis of HCC patients. Moreover, RHNO1 mRNA, especially protein levels were significantly increased in most HCC cells. Knockdown of RHNO1 through small interfering RNAs (siRNAs) inhibited the proliferation and triggered cell apoptosis of HCC cells both in vitro and in vivo. Specifically, we find that RHNO1 deficiency confers apoptosis via mitochondrial-mediated pathway. Mechanistically, silencing of RHNO1 impeded HCC proliferation and induced apoptosis by inactivating the PI3K/Akt pathway. Overall, these findings unravel that RHNO1 functions as an oncogene in HCC, and involved in regulating mitochondrial apoptosis to promote HCC thus may serve as a therapeutic and diagnostic target for HCC.
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Carcinoma Hepatocelular , Neoplasias Hepáticas , Humanos , Apoptose , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Proliferação de Células/genética , Regulação Neoplásica da Expressão Gênica , Neoplasias Hepáticas/patologia , Fosfatidilinositol 3-Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , RNA Interferente PequenoRESUMO
Alanine-serine-cysteine transporter 2 (ASCT2) is reported to participate in the progression of tumors and metabolic diseases. It is also considered to play a crucial role in the glutamate-glutamine shuttle of neuroglial network. However, it remains unclear the involvement of ASCT2 in neurological diseases such as Parkinson's disease (PD). In this study, we demonstrated that high expression of ASCT2 in the plasma samples of PD patients and the midbrain of MPTP mouse models is positively correlated with dyskinesia. We further illustrated that ASCT2 expressed in astrocytes rather than neurons significantly upregulated in response to either MPP+ or LPS/ATP challenge. Genetic ablation of astrocytic ASCT2 alleviated the neuroinflammation and rescued dopaminergic (DA) neuron damage in PD models in vitro and in vivo. Notably, the binding of ASCT2 to NLRP3 aggravates astrocytic inflammasome-triggered neuroinflammation. Then a panel of 2513 FDA-approved drugs were performed via virtual molecular screening based on the target ASCT2 and we succeed in getting the drug talniflumate. It is validated talniflumate impedes astrocytic inflammation and prevents degeneration of DA neurons in PD models. Collectively, these findings reveal the role of astrocytic ASCT2 in the pathogenesis of PD, broaden the therapeutic strategy and provide a promising candidate drug for PD treatment.
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Hepatocellular carcinoma (HCC) is the most common form of primary liver cancer, and it usually occurs following chronic liver disease. Although some progress has been made in the treatment of HCC, the prognosis of patients with advanced HCC is not optimistic, mainly because of the inevitable development of drug resistance. Therefore, multi-target kinase inhibitors for the treatment of HCC, such as sorafenib, lenvatinib, cabozantinib, and regorafenib, produce small clinical benefits for patients with HCC. It is necessary to study the mechanism of kinase inhibitor resistance and explore possible solutions to overcome this resistance to improve clinical benefits. In this study, we reviewed the mechanisms of resistance to multi-target kinase inhibitors in HCC and discussed strategies that can be used to improve treatment outcomes.
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Abnormal energy metabolism is a characteristic of tumor cells, and mitochondria are important components of tumor metabolic reprogramming. Mitochondria have gradually received the attention of scientists due to their important functions, such as providing chemical energy, producing substrates for tumor anabolism, controlling REDOX and calcium homeostasis, participating in the regulation of transcription, and controlling cell death. Based on the concept of reprogramming mitochondrial metabolism, a range of drugs have been developed to target the mitochondria. In this review, we discuss the current progress in mitochondrial metabolic reprogramming and summarized the corresponding treatment options. Finally, we propose mitochondrial inner membrane transporters as new and feasible therapeutic targets.
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Mitocôndrias , Neoplasias , Humanos , Mitocôndrias/metabolismo , Metabolismo Energético/fisiologia , Neoplasias/metabolismo , Membranas Mitocondriais/metabolismo , OxirreduçãoRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Baipuhuang Keli (BPH, constituted by Bai Tou Weng (Pulsatilla chinensis (Bunge) Regel), Pu Gong Ying (Taraxacum mongolicum Hand.-Mazz.), Huang Qin (Scutellaria baicalensis Georgi), Huang Bo (Phellodendron amurense Rupr.)) is a Chinese herbal formula with clearing heat and cooling blood, and removing toxin effects, which is suit for the case of breast cancer. AIM OF THE STUDY: Here, we aim to explore the effects of BPH on triple-negative breast cancer (TNBC) and its potential mechanisms. MATERIALS AND METHODS: In this study, cell viability assay, colony formation assay, soft agar assay, cell proliferation curve assay, and EdU assay were employed to determine the anti-proliferation effect induced by BPH. Cell cycle distribution was detected by flow cytometry. DNA damage in cells treated with BPH was indicated by comet assay, immunofluorescence, and Western Blot. Both the 4T1 orthotopic tumor model and the MDA-MB-231 subcutaneous tumor model were used to assess in vivo effect of BPH (312.5, and 625 mg/kg). The protein expression levels of the DNA damage response (DDR) pathway and the MAPK/ERK pathway were detected by Western Blot. RESULTS: Our results indicated that TNBC cells were more sensitive to BPH than mammary epithelial cells. Cell proliferation of TNBC cells was significantly inhibited by BPH in a dose-dependent manner. Moreover, BPH induced DNA damage in TNBC cells in a concentration and time-dependent manner. DDR of TNBC cells was inhibited by BPH. MAPK/ERK pathway was inhibited in cells treated with BPH, and DNA damage can be reversed while EGF was added to activate MAPK/ERK pathway. The 4T1 orthotopic tumor model and the MDA-MB-231 subcutaneous tumor model further confirmed that BPH inhibited TNBC proliferation via inhibition of DDR and MAPK/ERK pathway in vivo. CONCLUSIONS: Collectively, we proved that BPH is a potential anticancer Chinese herbal formula for TNBC in the manner of in vitro and in vivo experiments.
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Neoplasias de Mama Triplo Negativas , Humanos , Apoptose , Linhagem Celular Tumoral , Proliferação de Células , Dano ao DNA , Sistema de Sinalização das MAP Quinases , Medicina Tradicional Chinesa , Neoplasias de Mama Triplo Negativas/patologia , FemininoRESUMO
Although the Hedgehog (Hh) pathway plays an evolutionarily conserved role from Drosophila to mammals, some divergences also exist. Loss of Sufu, an important component of the Hh pathway, does not lead to an obvious developmental defect in Drosophila. However, in mammals, loss of SUFU results in serious disorder, even various cancers. This divergence suggests that SUFU plays additional roles in mammalian cells, besides regulating the Hh pathway. Here, we identify that the transcription factor ZNF281 is a novel binding partner of SUFU. Intriguingly, the Drosophila genome does not encode any homologs of ZNF281. SUFU is able to suppress ZNF281-induced tumor cell migration and DNA damage repair by inhibiting ZNF281 activity. Mechanistically, SUFU binds ZNF281 to mask the nuclear localization signal of ZNF281, culminating in ZNF281 cytoplasmic retention. In addition, SUFU also hampers the interactions between ZNF281 and promoters of target genes. Finally, we show that SUFU is able to inhibit ZNF281-induced tumor cell migration using an in vivo model. Taken together, these results uncover a Hh-independent mechanism of SUFU exerting the anti-tumor role, in which SUFU suppresses tumor cell migration through antagonizing ZNF281. Therefore, this study provides a possible explanation for the functional divergence of SUFU in mammals and Drosophila.
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Neoplasias , Fatores de Transcrição , Animais , Fatores de Transcrição/metabolismo , Proteínas Repressoras/metabolismo , Proteínas Hedgehog/metabolismo , Transdução de Sinais/fisiologia , Proteína GLI1 em Dedos de Zinco/metabolismo , Drosophila/metabolismo , Movimento Celular , Mamíferos/metabolismoRESUMO
BACKGROUND: The incidence of melanoma rapidly increased in the past decades, and the clinical treatment of melanoma met huge challenges because of tumor heterogeneity and drug resistance. C118P, a novel tubulin polymerization inhibitor, exhibited strong anticancer effects in many tumors. However, there was no data regarding the potential effects of C118P in melanoma cells. OBJECTIVE: To investigate of the efficacy and potential target of C118P in melanoma cells. METHODS: Human melanoma cells were treated with C118P, followed by assessments of proliferation, apoptosis and cell cycle distribution. Subsequently, RNA sequencing was performed to further identify the drug targets of C118P in melanoma cells. GO analysis and protein-protein interaction networks analysis were used to screen the potential targets, and verified by a series of assays. Finally, the anti-growth activity of C118P was evaluated in A375-xenografted nude mice, and the expression of BUB1B (BUB1 mitotic checkpoint serine/threonine kinase B), Ki67 and Tunel were determined. RESULTS: We found that C118P concentration-dependently inhibited proliferation of melanoma cells. Moreover, C118P simultaneously triggered dramatic G2/M arrest and apoptosis via independent mechanisms in melanoma cells in vitro. C118P exerted anti-melanoma effects by inducing potent G2/M arrest, which was mechanistically related to downregulation of the expression of BUB1B. Importantly, C118P inhibited the tumor growth in A375-xenografted nude, and increased the staining of Ki-67 and Tunel and suppressed the expression of BUB1B in melanoma tissues, which was consistent with in vitro study. CONCLUSION: C118P might provide a novel strategy for the clinical treatment of melanoma by inhibition of BUB1B.
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Apoptose , Melanoma , Camundongos , Animais , Humanos , Camundongos Nus , Pontos de Checagem da Fase G2 do Ciclo Celular/genética , Linhagem Celular Tumoral , Melanoma/patologia , Proteínas Serina-Treonina Quinases , Proliferação de Células , Proteínas de Ciclo CelularRESUMO
The occurrence and development of tumors depend on the tumor microenvironment (TME), which consists of various types of cellular and acellular components. Tumor-associated macrophages (TAMs) are the most abundant stromal cell types in the TME. The competition for nutrients between tumor cells and macrophages leads to a limited supply of nutrients, such as glucose, lipids, and amino acids, to immune cells, which affects the differentiation and function of macrophages. Other factors in the TME, such as cytokines, chemokines, and immune checkpoints, also affect the polarization and function of macrophages. Remodeling the tumor microenvironment induces changes in macrophage nutrient uptake and polarization status, which enhance anti-tumor immunity and oxidative stress resistance and suppress immune escape. This review summarizes the influence factors on tumor progression and immune function under different conditions of macrophages. It also demonstrates the metabolic heterogeneity and phenotypic plasticity of macrophages, which provides novel strategies for anti-tumor treatment.
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Neoplasias , Microambiente Tumoral , Diferenciação Celular , Citocinas/metabolismo , Humanos , Macrófagos/metabolismo , Neoplasias/patologiaRESUMO
BACKGROUND: Glioma is one of the most fatal types of malignant tumours, the cause of which is mostly unknown. Orphan GPCRs (GPRs) have been previously implicated in tumour growth and metastasis. Therefore, these GPRs could prove to be alternative and promising therapeutic targets for cancer treatment. OBJECTIVE: The role of GPR160 in glioma has not yet been assessed. This study aims to explore the association of GPR160 with glioma progression and investigate its role in epithelial-to-mesenchymal transition (EMT) and metastasis. METHODS: Changes in protein expression were assessed using western blot analysis and immunofluorescent staining assays, while mRNA expression changes were evaluated using qRT-PCR. To detect the changes in progression and metastasis, MTT, EdU proliferation, wound healing, transwell migration, and flow cytometry assays were carried out in vitro. An epithelial to mesenchymal phenotypic analysis was performed to detect EMT. RESULTS: We demonstrated that knockdown of GPR160 inhibited proliferation, colony formation, and cell viability and promoted apoptosis. Pro-apoptotic biomarkers were upregulated, while anti-apoptotic biomarkers were downregulated. Cell lines with GPR160 knockdown (GPR160 KD) showed a slowed migration rate and decreased invasion ability. EMT mesenchymal biomarkers were downregulated in GPR160 KD cell lines, while epithelial biomarkers were upregulated. CONCLUSION: This study provides evidence that GPR160 is a potential therapeutic target in glioma for the first time. These findings can be used to discover in detail the molecular mechanism and pathways through which GPR160 promotes glioma progression.
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Transição Epitelial-Mesenquimal , Glioma , Biomarcadores , Linhagem Celular Tumoral , Movimento Celular , Proliferação de Células/genética , Regulação para Baixo , Regulação Neoplásica da Expressão Gênica , Glioma/genética , Humanos , RNA MensageiroRESUMO
Metabolic reprogramming is considered to be a hallmark of cancer, and increased glutamine metabolism plays an important role in the progression of many tumors, including colorectal cancer (CRC). Targeting of glutamine uptake via the transporter protein ASCT2/SLC1A5 (solute carrier family 1 member 5) is considered to be an effective strategy for the treatment of malignant tumors. Here, we demonstrate that Ag120 (ivosidenib), a mutant isocitrate dehydrogenase 1 (IDH1) inhibitor approved for the treatment of certain cancers, acts as an ASCT2 inhibitor in CRC cells. Ag120 blocked glutamine uptake and metabolism, leading to reduced cell proliferation, elevated autophagy, and increased oxidative stress in CRC cells in vitro and in vivo, potentially via the ERK and mTOR signaling pathways. These effects occurred independently of mutant IDH1 activity and were supported by experiments with ASCT2-depleted or -overexpressing cells. These data identify a novel mechanism of Ag120 anti-tumor activity and support further exploration of ASCT2 inhibitors for cancer therapy.
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Purpose: Orphan GPCRs (GPRs) play important roles in the malignant progression of cancer and have the potential to develop into anti-tumor drug targets. However, the biological processes and molecular mechanisms of GPR27 have not been properly assessed in cancer. Our objective was to reveal the effect of GPR27 on the progression of hepatocellular carcinoma (HCC). Methods: GPR27 levels were detected in HCC cell lines using quantitative reverse transcriptase-polymerase chain reaction and Western blot analysis. Next, the changes of phenotypes after GPR27 knockdown or overexpression were evaluated using in vitro methods. Finally, the mechanism of GPR27 in HCC was tested using RNA-seq and in vivo mouse xenograft model. Results: In the present study, we reported that suppression of GPR27 expression inhibited proliferation, colony formation, cell viability, and induced cell S phase arrest of HCC cells, whereas GPR27 overexpression led to the opposite outcomes. Moreover, suppression of GPR27 expression resulted in blocking MAPK/ERK signal pathway which indicated the inhibition of HCC cells proliferation. Further study in vivo confirmed that GPR27 can affect the proliferation of HCC cells through the MAPK/ERK pathway. Conclusion: Taken together, the findings of the present study uncover biological functions of GPR27 in HCC cells, and delineate preliminary molecular mechanisms of GPR27 in modulating HCC development and progression.