RESUMO
MYCN amplification is an independent risk factor for poor prognosis in neuroblastoma (NB), but its protein product cannot be directly targeted because of protein structure. Thus, this study aimed to explore novel ways to indirectly target N-Myc by regulating its post-translational modifications (PTMs) and therefore protein stability. N-Myc coimmunoprecipitation combined with HPLC-MS/MS identified 16 PTM residues and 114 potential N-Myc-interacting proteins. Notably, both acetylation and ubiquitination were identified on lysine 199 of N-Myc. We then discovered that p300, which can interact with N-Myc, modulated the protein stability of N-Myc in MYCN-amplified NB cell lines and simultaneously regulated the acetylation level and ubiquitination level on lysine-199 of N-Myc protein in vitro. Furthermore, p300 correlated with poor prognosis in NB patients. Taken together, p300 can be considered as a potential therapeutic target to treat MYCN-amplified NB patients, and other identified PTMs and interacting proteins also provide potential targets for further study.
Assuntos
Lisina , Neuroblastoma , Humanos , Proteína Proto-Oncogênica N-Myc/genética , Proteína Proto-Oncogênica N-Myc/metabolismo , Proteína Proto-Oncogênica N-Myc/uso terapêutico , Lisina/metabolismo , Espectrometria de Massas em Tandem , Processamento de Proteína Pós-Traducional , Neuroblastoma/tratamento farmacológico , Neuroblastoma/genética , Neuroblastoma/metabolismo , Estabilidade Proteica , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão GênicaRESUMO
BACKGROUND: Despite the extensive study of MYCN-amplified neuroblastomas, there is a significant unmet clinical need in MYCN non-amplified cases. In particular, the extent of heterogeneity within the MYCN non-amplified population is unknown. METHODS: A total of 1566 samples from 16 datasets were identified in Gene Expression Omnibus (GEO) and ArrayExpress. Characterisation of the subtypes was analysed by ConsensusClusterPlus. Independent predictors for subgrouping were constructed from the single sample predictor based on the multiclassPairs package. Findings were verified using immunohistochemistry and CIBERSORTx analysis. RESULTS: We demonstrate that MYCN non-amplified neuroblastomas are heterogeneous and can be classified into 3 subgroups based on their transcriptional signatures. Within these groups, subgroup_2 has the worst prognosis and this group shows a 'MYCN' signature that is potentially induced by the overexpression of Aurora Kinase A (AURKA); whilst subgroup_3 is characterised by an 'inflamed' gene signature. The clinical implications of this subtype classification are significant, as each subtype demonstrates a unique prognosis and vulnerability to investigational therapies. A total of 420 genes were identified as independent subgroup predictors with average balanced accuracy of 0.93 and 0.84 for train and test datasets, respectively. CONCLUSION: We propose that transcriptional subtyping may enhance precision prognosis and therapy stratification for patients with MYCN non-amplified neuroblastomas.
Assuntos
Proteína Proto-Oncogênica N-Myc , Neuroblastoma , Humanos , Neuroblastoma/genética , Neuroblastoma/classificação , Neuroblastoma/patologia , Neuroblastoma/mortalidade , Proteína Proto-Oncogênica N-Myc/genética , Prognóstico , Aurora Quinase A/genética , Perfilação da Expressão Gênica/métodos , Regulação Neoplásica da Expressão Gênica , Biomarcadores Tumorais/genética , Amplificação de GenesRESUMO
BACKGROUND: Minichromosome maintenance complex component 6 (MCM6), as an important replication permission factor, is involved in the pathogenesis of various tumors. Here we studied the expression of MCM6 in neuroblastoma and its influence on tumor characteristics and prognosis. METHODS: Publicly available datasets were used to explore the influence of the differential expression of MCM6 on neuroblastoma tumor stage, risk and prognosis. In cell experiments, human neuroblastoma cell lines SK-N-SH and SK-N-BE [ (2)] were utilized to verify the ability of MCM6 to promote cell proliferation, migration and invasion. We further explored the possible molecular mechanism of MCM6 affecting the phenotype of neuroblastoma cells by mutual verification of RNA-seq and western blotting, and flow cytometry to inquire about its potential specific roles in the cell cycle. RESULTS: Through multiple datasets mining, we found that high expression of MCM6 was positively correlated with elevated tumor stage, high risk and poor prognosis in neuroblastoma. At the cellular level, neuroblastoma cell proliferation, migration and invasion were significantly inhibited after MCM6 was interfered by siRNA. Mutual verification of RNA-seq and western blotting suggested that the downstream cell cycle-related genes were differentially expressed after MCM6 interference. Flow cytometric analysis revealed that neuroblastoma cells were blocked in G1/S phase after MCM6 interference. CONCLUSION: MCM6 is considered to be the driving force of G1/S cell cycle progression, and it is also a prognostic marker and a potential novel therapeutic target in neuroblastoma.
Assuntos
Proteínas de Ciclo Celular/metabolismo , Componente 6 do Complexo de Manutenção de Minicromossomo/efeitos adversos , Neuroblastoma/genética , Animais , Modelos Animais de Doenças , Progressão da Doença , Humanos , Camundongos , Camundongos Nus , Neuroblastoma/patologia , Prognóstico , Transfecção , Resultado do TratamentoAssuntos
Hiperplasia do Linfonodo Gigante/diagnóstico , Histiocitoma Fibroso Maligno/diagnóstico , Linfonodos/patologia , Hiperplasia do Linfonodo Gigante/diagnóstico por imagem , Hiperplasia do Linfonodo Gigante/terapia , Criança , Histiocitoma Fibroso Maligno/diagnóstico por imagem , Histiocitoma Fibroso Maligno/terapia , Humanos , Masculino , Tomografia por Emissão de Pósitrons combinada à Tomografia Computadorizada , Proteína EWS de Ligação a RNA/genéticaRESUMO
Background: Aidi injection, a classic traditional Chinese medicine (TCM) formula, has been used on a broader scale in treating a variety of cancers. In this study, we aimed to explore the potential anti-tumor effects of Aidi injection in the treatment of neuroblastoma (NB) using network pharmacology (NP). Methods: To elucidate the anti-NB mechanism of Aidi injection, an NP-based approach and molecular docking validation were employed. The compounds and target genes were collected from the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database and Bioinformatics Analysis Tool for Molecular mechANism of Traditional Chinese Medicine (BATMAN-TCM) database. The protein-protein interaction network was constructed using the STRING database. clusterProfiler (R package) was utilized to annotate the bioinformatics of hub target genes. The gene survival analysis was performed on R2, a web-based genomic analysis application. iGEMDOCK was used for molecular docking validation, and GROMACS was utilized to validate molecular docking results. Furthermore, we investigated the anticancer effects of gomisin B and ginsenoside Rh2 on human NB cells using a cell viability assay. The Western blot assay was used to validate the protein levels of target genes in gomisin B- and ginsenoside Rh2-treated NB cells. Results: A total of 2 critical compounds with 16 hub target genes were identified for treating NB. All 16 hub genes could potentially influence the survival of NB patients. The top three genes (EGFR, ESR1, and MAPK1) were considered the central hub genes from the drug-compound-hub target gene-pathway network. The endocrine resistance and estrogen signaling pathways were identified as the therapeutic pathways using the Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis. Gomisin B and ginsenoside Rh2 showed a good binding ability to the target protein in molecular docking. The results of cell experiments showed the anti-NB effect of gomisin B and ginsenoside Rh2. In addition, the administration of gomisin B over-regulated the expression of ESR1 protein in MYCN-amplified NB cells. Conclusion: In the present study, we investigated the potential pharmacological mechanisms of Aidi against NB and revealed the anti-NB effect of gomisin B, providing clinical evidence of Aidi in treating NB and establishing baselines for further research.
RESUMO
Studies have shown that PCNA clamp associated factor (PCLAF) plays a paramount role in a variety of cancers; however, the expression profile and the specific molecular mechanism of PCLAF in cancer remains unclear, as is its value in the human pancancer analysis. Based on the publicly available datasets of The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO), a comprehensive analysis of the probable carcinogenic effects of the PCLAF gene was performed in 33 human cancers. It was found that PCLAF is highly expressed in cancer tissues compared with normal tissues, and is significantly correlated with poor prognosis. We found that the eight tumors with significantly high PCLAF expression presented with decreased DNA methylation levels of PCLAF, including cholangiocarcinoma (CHOL), cervical squamous cell carcinoma and endocervical adenocarcinoma (CESC), glioblastoma multiforme (GBM), pheochromocytoma and paraganglioma (PCPG), sarcoma (SARC), testicular germ cell tumor (TGCT), stomach adenocarcinoma (STAD), and uterine corpus endometrial carcinoma (UCEC). The expression of PCLAF was found to be positively correlated with activated CD4 T cells (Act CD4) and type 2 T helper (Th2) cells, suggesting that PCLAF may play a particular role in tumor immune infiltration. In addition, the functional mechanism of PCLAF also involves the mitotic cell cycle process, cell division, and DNA replication. Our first pancancer study provides a relatively extensive understanding of the carcinogenic effects of PCLAF in miscellaneous tumors.
Assuntos
Adenocarcinoma , Regulação Neoplásica da Expressão Gênica , Humanos , Prognóstico , Linfócitos TRESUMO
PCLAF (PCNA clamp-associated factor), also known as PAF15/ KIAA0101, is overexpressed in most human cancers and is a predominant regulator of tumor progression. However, its biological function in neuroblastoma remains unclear. PCLAF is extremely overexpressed in neuroblastoma and is associated with poor prognosis. Through the analysis of various data sets, we found that the high expression of PCLAF is positively correlated with increased stage and high risk of neuroblastoma. Most importantly, knocking down PCLAF could restrict the proliferation of neuroblastoma cells in vitro and in vitro. By analyzing RNA-seq data, we found that the enrichment of cell cycle-related pathway genes was most significant among the differentially expressed downregulated genes after reducing the expression of PCLAF. In addition, PCLAF accelerated the G1/S transition of the neuroblastoma cell cycle by activating the E2F1/PTTG1 signaling pathway. In this study, we reveal the mechanism by which PCLAF facilitates cell cycle progression and recommend that the PCLAF/E2F1/PTTG1 axis is a therapeutic target in neuroblastoma.