RESUMO
Comprehensive sequencing of patient tumors reveals genomic mutations across tumor types that enable tumorigenesis and progression. A subset of oncogenic driver mutations results in neomorphic activity where the mutant protein mediates functions not engaged by the parental molecule. Here, we identify prevalent variant-enabled neomorph-protein-protein interactions (neoPPI) with a quantitative high-throughput differential screening (qHT-dS) platform. The coupling of highly sensitive BRET biosensors with miniaturized coexpression in an ultra-HTS format allows large-scale monitoring of the interactions of wild-type and mutant variant counterparts with a library of cancer-associated proteins in live cells. The screening of 17,792 interactions with 2,172,864 data points revealed a landscape of gain of interactions encompassing both oncogenic and tumor suppressor mutations. For example, the recurrent BRAF V600E lesion mediates KEAP1 neoPPI, rewiring a BRAFV600E/KEAP1 signaling axis and creating collateral vulnerability to NQO1 substrates, offering a combination therapeutic strategy. Thus, cancer genomic alterations can create neo-interactions, informing variant-directed therapeutic approaches for precision medicine.
Assuntos
Neoplasias , Proteínas Proto-Oncogênicas B-raf , Carcinogênese , Humanos , Proteína 1 Associada a ECH Semelhante a Kelch/genética , Proteína 1 Associada a ECH Semelhante a Kelch/metabolismo , Mutação , Fator 2 Relacionado a NF-E2/metabolismo , Neoplasias/genética , Proteínas Proto-Oncogênicas B-raf/genética , Proteínas Proto-Oncogênicas B-raf/metabolismoRESUMO
Glioblastoma (GBM) is one of the most lethal types of primary brain tumors in adults with a median survival of less than 15 months. Although comprehensive clinical treatment strategies including surgical resection followed by radiotherapy and chemotherapy are widely applied, the prognosis for GBM patients remains dismal. The Nuclear Factor-κB (NF-κB) signaling pathway is a complex network linking extracellular stimuli to cell survival and proliferation, and aberrant activation of NF-κB signaling has been implicated in the propagation of a wide range of cancers. However, the underlying mechanism of NF-κB activation still requires further investigation. Here, we report that crumbs homolog 2 (CRB2) is markedly up-regulated in human GBM relative to non-tumor tissues or normal astrocytes. Clinically, enriched CRB2 could be observed in high grade glioma with IDH IDH wild-type and 1p19q co-deletion and implied poor outcome in GBM. Consistent with this, malignant characteristics of GBM cells including proliferation, migration, invasion and tumorigenesis were significantly suppressed by lentivirus knock-down of CRB2. Furthermore, exogenous overexpression of CRB2 enhanced the malignant biological signatures of GBM cells as well as therapy resistance to temozolomide (TMZ). To further investigate the molecular mechanisms responsible, bioinformatics analysis was performed using 3 public databases, with the result that CRB2 was found to correlate closely with tumor necrosis factor α (TNFα)-NF-κB signaling. Mechanistically, elevated CRB2 increased the phosphorylation of IκB-kinase α (IKKα), thus activating NF-κB via reduction of Ikß protein. Taken together, these data suggest that CRB2 might be a reliable prognostic biomarker and potential therapeutic target for GBM.
Assuntos
Neoplasias Encefálicas , Proteínas de Transporte , Glioblastoma , Glioma , Proteínas de Membrana , Neoplasias Encefálicas/patologia , Proteínas de Transporte/metabolismo , Linhagem Celular Tumoral , Regulação Neoplásica da Expressão Gênica , Glioblastoma/patologia , Glioma/patologia , Humanos , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , NF-kappa B/genética , NF-kappa B/metabolismo , Temozolomida/uso terapêuticoRESUMO
Nonglioblastomatous diffuse glioma (non-GDG) is a heterogeneous neuroepithelial tumor that exhibits a varied survival range from 4 to 13 years based on the diverse subtypes. Recent studies demonstrated novel molecular markers can predict prognosis for non-GDG patients; however, these findings as well as pathological classification strategies show obvious limitations on malignant transition due to the heterogeneity among non-GDGs. Therefore, developing reliable prognostic biomarkers and therapeutic targets have become an urgent need for precisely distinguishing non-GDG subtypes, illuminating the underlying mechanism. Nuclear factor κß (NF-κB) has been proved to be a significant nuclear transcriptional regulator with specific DNA-binding sequences to participate in multiple pathophysiological processes. However, the underlying mechanism of NF-κB activation still needs to be further investigated. Herein, our results indicated retinol-binding protein 1 (RBP1) was significantly upregulated in the IDHWT and 1p19qNon co-del non-GDG subtypes and enriched RBP1 expression was markedly correlated with more severe outcomes. Additionally, malignant signatures of the non-GDG cells including proliferation, migration, invasion, and self-renewal were significantly suppressed by lentiviral knockdown of RBP1. To further explore the underlying molecular mechanism, bioinformatics analysis was performed using databases, and the results demonstrated RBP1 was strongly correlated with tumor necrosis factor α (TNFα)-NF-κB signaling. Moreover, exogenous silencing of RBP1 reduced phosphorylation of IkB-kinase α (IKKα) and thus decreased NF-κB expression via decreasing the degradation of the IκBα protein. Altogether, these data suggested RBP1-dependent activation of NF-κB signaling promoted malignancy of non-GDG, indicating that RBP1 could be a reliable prognostic biomarker and potential therapeutic target for non-GDG.
Assuntos
Glioma/patologia , NF-kappa B/metabolismo , Proteínas Celulares de Ligação ao Retinol/metabolismo , Biomarcadores Tumorais/metabolismo , Movimento Celular , Proliferação de Células , Transformação Celular Neoplásica , Transição Epitelial-Mesenquimal , Glioma/genética , Glioma/metabolismo , Humanos , Quinase I-kappa B/metabolismo , Isocitrato Desidrogenase/metabolismo , Fosforilação , Prognóstico , Proteínas Celulares de Ligação ao Retinol/genética , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Low-grade gliomas (LGGs) are grade III gliomas based on the WHO classification with significant genetic heterogeneity and clinical properties. Traditional histological classification of gliomas has been challenged by the improvement of molecular stratification; however, the reproducibility and diagnostic accuracy of LGGs classification still remain poor. Herein, we identified fatty acid binding protein 5 (FABP5) as one of the most enriched genes in malignant LGGs and elevated FABP5 revealed severe outcomes in LGGs. Functionally, lentiviral suppression of FABP5 reduced malignant characters including proliferation, cloning formation, immigration, invasion and TMZ resistance, contrarily, the malignancies of LGGs were enhanced by exogenous overexpression of FABP5. Mechanistically, epithelial-mesenchymal transition (EMT) was correlated to FABP5 expression in LGGs and tumour necrosis factor α (TNFα)-dependent NF-κB signalling was involved in this process. Furthermore, FABP5 induced phosphorylation of inhibitor of nuclear factor kappa-B kinase α (IKKα) thus activated nuclear factor kappa-B (NF-κB) signalling. Taken together, our study indicated that FABP5 enhances malignancies of LGGs through canonical activation of NF-κB signalling, which could be used as individualized prognostic biomarker and potential therapeutic target of LGGs.
Assuntos
Biomarcadores Tumorais/metabolismo , Neoplasias Encefálicas/patologia , Proteínas de Ligação a Ácido Graxo/metabolismo , Regulação Neoplásica da Expressão Gênica , Glioma/patologia , NF-kappa B/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Neoplasias Encefálicas/genética , Neoplasias Encefálicas/metabolismo , Proliferação de Células , Proteínas de Ligação a Ácido Graxo/genética , Glioma/genética , Glioma/metabolismo , Humanos , NF-kappa B/genética , Invasividade Neoplásica , Prognóstico , Transdução de Sinais , Taxa de Sobrevida , Células Tumorais Cultivadas , CicatrizaçãoRESUMO
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RESUMO
Background: Lower-grade gliomas (LGGs) are characterized by remarkable genetic heterogeneity and different clinical outcomes. Classification of LGGs is improved by the development of molecular stratification markers including IDH mutation and 1p/19q chromosomal integrity, which are used as a hallmark of survival and therapy sensitivity of LGG patients. However, the reproducibility and sensitivity of the current classification remain ambiguous. This study aimed to construct more accurate risk-stratification approaches. Methods: According to bioinformatics, the sequencing profiles of methylation and transcription and imaging data derived from LGG patients were analyzed and developed predictable risk score and radiomics score. Moreover, the performance of predictable models was further validated. Results: In this study, we determined a cluster of 6 genes that were correlated with IDH mutation/1p19q co-deletion status. Risk score model was calculated based on 6 genes and showed gratifying sensitivity and specificity for survival prediction and therapy response of LGG patients. Furthermore, a radiomics risk score model was established to noninvasively assist judgment of risk score in pre-surgery. Taken together, a predictable nomogram that combined transcriptional signatures and clinical characteristics was established and validated to be preferable to the histopathological classification. Our novel multi-omics nomograms showed a satisfying performance. To establish a user-friendly application, the nomogram was further developed into a web-based platform: https://drw576223193.shinyapps.io/Nomo/, which could be used as a supporting method in addition to the current histopathological-based classification of gliomas. Conclusions: Our novel multi-omics nomograms showed the satisfying performance of LGG patients and assisted clinicians to draw up individualized clinical management.
RESUMO
Tumor suppressor genes represent a major class of oncogenic drivers. However, direct targeting of loss-of-function tumor suppressors remains challenging. To address this gap, we explored a variant-directed chemical biology approach to reverse the lost function of tumor suppressors using SMAD4 as an example. SMAD4, a central mediator of the TGF-ß pathway, is recurrently mutated in many tumors. Here, we report the development of a TR-FRET technology that recapitulated the dynamic differential interaction of SMAD4 and SMAD4R361H with SMAD3 and identified Ro-31-8220, a bisindolylmaleimide derivative, as a SMAD4R361H/SMAD3 interaction inducer. Ro-31-8220 reactivated the dormant SMAD4R361H-mediated transcriptional activity and restored TGF-ß-induced tumor suppression activity in SMAD4 mutant cancer cells. Thus, demonstration of Ro-31-8220 as a SMAD4R361H/SMAD3 interaction inducer illustrates a general strategy to reverse the lost function of tumor suppressors with hypomorph mutations and supports a systematic approach to develop small-molecule protein-protein interaction (PPI) molecular glues for biological insights and therapeutic discovery.
Assuntos
Indóis/metabolismo , Proteína Smad4/metabolismo , Bibliotecas de Moléculas Pequenas/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Linhagem Celular , Feminino , Transferência Ressonante de Energia de Fluorescência , Genes Supressores de Tumor , Humanos , Indóis/química , Masculino , Ligação Proteica , Transdução de Sinais/genética , Proteína Smad4/química , Proteína Smad4/genética , Bibliotecas de Moléculas Pequenas/química , Fator de Crescimento Transformador beta/genéticaRESUMO
INTRODUCTION: Glioblastoma (GBM) is the most lethal primary malignant brain tumor in adults with poor survival due to acquired therapeutic resistance and rapid recurrence. Currently, the standard clinical strategy for glioma includes maximum surgical resection, radiotherapy, and temozolomide (TMZ) chemotherapy; however, the median survival of patients with GBM remains poor despite these comprehensive therapies. Therefore, the identification of new prognostic biomarkers is urgently needed to evaluate the malignancy and long-term outcome of glioma. AIMS: To further investigate prognostic biomarkers and potential therapeutic targets for GBM. RESULTS: In this study, we identified tribbles pseudokinase 2 (TRIB2) as one of the genes that is most correlated with pathological classification, radioresistance, and TMZ resistance in glioma. Additionally, the expression of mitogen-activated protein kinase kinase kinase 1 (MAP3K1) showed a strong correlation with TRIB2. Moreover, a combined increase in TRIB2 and MAP3K1 was observed in GBM and indicated a poor prognosis of patients with glioma. Finally, enriched TRIB2 expression and MAP3K1 expression were shown to be associated with resistance to TMZ and radiotherapy. CONCLUSION: Combined elevation of TRIB2 and MAP3K1 could be novel prognostic biomarkers and potential therapeutic targets to evaluate the malignancy and long-term outcomes of GBM.
Assuntos
Neoplasias Encefálicas/metabolismo , Proteínas Quinases Dependentes de Cálcio-Calmodulina/biossíntese , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Glioblastoma/metabolismo , MAP Quinase Quinase Quinase 1/biossíntese , Temozolomida/uso terapêutico , Adulto , Antineoplásicos Alquilantes/uso terapêutico , Neoplasias Encefálicas/tratamento farmacológico , Neoplasias Encefálicas/patologia , Resistencia a Medicamentos Antineoplásicos/fisiologia , Feminino , Regulação Neoplásica da Expressão Gênica , Glioblastoma/tratamento farmacológico , Glioblastoma/patologia , Humanos , Masculino , Pessoa de Meia-Idade , PrognósticoRESUMO
OBJECTIVES: The aim of this study was to evaluate the neuroprotective effects of SalB on high glucose (HG)-induced excessive autophagy and apoptosis in vitro. METHODS: The proliferation and apoptosis of RSC96 cells were determined using the MTT assay and flow cytometry, respectively. Western blot analysis was performed to examine the expression of autophagy and apoptosis-related proteins. RT-PCR and flow cytometry were manipulated to examine the level of Bcl-2. The signals of autophagy markers were detected using immunofluorescence methods. KEY FINDINGS: We found that HG significantly reduced RSC96 cell's proliferation and induced apoptosis. What's more, HG increased the level of autophagy and apoptosis-related proteins. However, these effects were reversed by SalB. In addition, we also found that 3-MA decreased the expression of LC3A/B and Beclin1, while the JNK inhibitor SP600125 reduced the levels of phosphorylated JNK, LC3A/B and Beclin1. CONCLUSIONS: High glucose not only induced apoptosis but also caused autophagic cell death by activating the JNK pathway. These effects prevented by SalB in an opposite manner.
Assuntos
Apoptose/efeitos dos fármacos , Autofagia/efeitos dos fármacos , Benzofuranos/farmacologia , Neuropatias Diabéticas/prevenção & controle , Doenças do Sistema Nervoso Periférico/prevenção & controle , Animais , Antracenos/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Proteína Beclina-1/metabolismo , Linhagem Celular , Proliferação de Células/efeitos dos fármacos , Neuropatias Diabéticas/metabolismo , Glucose/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Doenças do Sistema Nervoso Periférico/metabolismo , Fosforilação/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Transdução de Sinais/efeitos dos fármacosRESUMO
AIMS: To investigate the underlying mechanism by which glioblastoma (GBM) cells gain temozolomide (TMZ) resistance and to clarify novel therapeutic targets and new prognostic biomarkers for GBM. MAIN METHODS: A genome-wide hierarchical bi-clustering based on previously published microarray databases identified Nuclear Factor I A (NFIA) as one of the most significantly upregulated genes correlated to TMZ resistance in GBM. Then, the potential biological functions of NFIA in oncogenesis and chemoresistance were clarified by qRT-PCR, Western blotting and in vivo xenograft models with artificially induced TMZ-resistant U87â¯cells. Additionally, immunohistochemistry (IHC) assays were performed to explore the clinical significance of NFIA in glioma patients. Last, luciferase reporter assay was performed to study the transcriptional regulation of NFIA on the nuclear factor κb (NF-kB) pathway. KEY FINDINGS: NFIA was correlated with TMZ resistance in GBM. Clinically, elevated NFIA expression was significantly correlated with adverse outcomes of glioma patients, especially in GBM patients. Moreover, NFIA contributed to the acquired TMZ resistance of GBM cells, while suppression of NFIA via lentivirus reduced cell proliferation, tumorigenesis and resistance to TMZ of GBM. Additionally, NFIA promoted transcription activity that regulated the expression of NF-kB. Last, NFIA induced phosphorylation of NF-kB p65 at serine 536, thus inducing TMZ resistance in GBM cells. Altogether, our study suggests that NFIA-dependent transcriptional regulation of NF-kB contributes to acquired TMZ resistance in GBM. SIGNIFICANCE: Abnormally activated NFIA-NF-kB signaling was strongly correlated with acquired TMZ resistance and poor prognosis in GBM, and it could be a new therapeutic target for TMZ-resistant GBM.