RESUMO
BACKGROUND: Viruses are obligate parasites that depend on host cells to provide the energy and molecular precursors necessary for successful infection. The main component of virus-induced metabolic reprogramming is the activation of glycolysis, which provides biomolecular resources for viral replication. However, little is known about the crosstalk between oncolytic viruses and host glycolytic processes. METHODS: A MTT assay was used to detect M1 virus-induced cell killing. Flow cytometry was used to monitor infection of M1 virus expressing the GFP reporter gene. qPCR and western blotting were used to detect gene expression. RNA sequencing was performed to evaluate gene expression under different drug treatments. Scanning electron microscopy was performed to visualize the endoplasmic reticulum (ER). Caspase activity was detected. Last, a mouse xenograft model was established to evaluate the antitumor effect in vivo. Most data were analyzed with a two-tailed Student's t test or one-way ANOVA with Dunnett's test for pairwise comparisons. Tumor volumes were analyzed by repeated measures of ANOVA. The Wilcoxon signed-rank test was used to compare nonnormally distributed data. RESULTS: Here, we showed that the glucose analog 2-deoxy-D-glucose (2-DG) inhibited infection by M1 virus, which we identified as a novel type of oncolytic virus, and decreased its oncolytic effect, indicating the dependence of M1 replication on glycolysis. In contrast, lonidamine, a reported hexokinase 2 (HK2) inhibitor, enhanced the infection and oncolytic effect of M1 virus independent of HK2. Further transcriptomic analysis revealed that downregulation of the antiviral immune response contributes to the lonidamine-mediated potentiation of the infection and oncolytic effect of M1 virus, and that MYC is the key factor in the pool of antiviral immune response factors inhibited by lonidamine. Moreover, lonidamine potentiated the irreversible ER stress-mediated apoptosis induced by M1 virus. Enhancement of M1's oncolytic effect by lonidamine was also identified in vivo. CONCLUSIONS: This research demonstrated the dependence of M1 virus on glycolysis and identified a candidate synergist for M1 virotherapy.
RESUMO
Oncolytic viruses are potent anticancer agents that replicate within and kill cancer cells rather than normal cells, and their selectivity is largely determined by oncogenic mutations. M1, a novel oncolytic virus strain, has been shown to target cancer cells, but the relationship between its cancer selectivity and oncogenic signaling pathways is poorly understood. Here, we report that RAS mutation promotes the replication and oncolytic effect of M1 in cancer, and we further provide evidence that the inhibition of the RAS/RAF/MEK signaling axis suppresses M1 infection and the subsequent cytopathic effects. Transcriptome analysis revealed that the inhibition of RAS signaling upregulates the type I interferon antiviral response, and further RNA interference screen identified CDKN1A as a key downstream factor that inhibits viral infection. Gain- and loss-of-function experiments confirmed that CDKN1A inhibited the replication and oncolytic effect of M1 virus. Subsequent TCGA data mining and tissue microarray (TMA) analysis revealed that CDKN1A is commonly deficient in human cancers, suggesting extensive clinical application prospects for M1. Our report indicates that virotherapy is feasible for treating undruggable RAS-driven cancers and provides reliable biomarkers for personalized cancer therapy.
Assuntos
Neoplasias/metabolismo , Neoplasias/virologia , Vírus Oncolíticos/fisiologia , Transdução de Sinais , Proteínas ras/metabolismo , Animais , Antivirais/farmacologia , Biomarcadores Tumorais/metabolismo , Butadienos/farmacologia , Linhagem Celular , Linhagem Celular Tumoral , Inibidor de Quinase Dependente de Ciclina p21/deficiência , Inibidor de Quinase Dependente de Ciclina p21/metabolismo , Feminino , Regulação Viral da Expressão Gênica/efeitos dos fármacos , Humanos , Camundongos Endogâmicos BALB C , Camundongos Nus , Mutação/genética , Neoplasias/patologia , Nitrilas/farmacologia , Vírus Oncolíticos/efeitos dos fármacos , Vírus Oncolíticos/genética , RNA Interferente Pequeno/metabolismo , Transdução de Sinais/efeitos dos fármacos , Regulação para Cima/efeitos dos fármacos , Regulação para Cima/genética , Replicação Viral/efeitos dos fármacos , Replicação Viral/genéticaRESUMO
Avoiding immune destruction is essential for tumorigenesis. Current research into the interaction between tumor and immunological niches complement tumor pathology beyond cancer genetics. Intrinsic host defense immunity is a specialized innate immunity component to restrict viral infection. However, whether intrinsic immunity participates in tumor pathology is unclear. Previously, we identified a zinc-finger antiviral protein ZAP that is commonly downregulated in a panel of clinical cancer specimens. However, whether ZAP has an impact on tumor development was unknown. Here we report ZAP as a genuine tumor suppressor. Pan-caner analysis with TCGA data from 712 patients and large-scale immunohistochemistry in tissue microarrays from 1552 patients reveal that ZAP is prevalently downregulated, and associated with poor survival in liver, colon, and bladder cancer patients. Ectopic over-expression of ZAP inhibits the malignant phenotypes of colorectal tumor by cell cycle arrest. Using RNA immunoprecipitation and RNA decay assays, we demonstrate that ZAP directly and specifically binds to and degrades the transcript of TRAILR4, which in turn represses TRAILR4 expression and inhibits the aggressiveness of colorectal cancer cells. Furthermore, our CRISPR-engineered mice models show that loss-of-function of ZAP synergizes with APC-deficiency to drive malignant colorectal cancer in vivo. Overall, we identify a previously unknown function of the antiviral factor ZAP in colorectal tumorigenesis, linking intrinsic immunity to tumor pathogenetics.
Assuntos
Neoplasias Colorretais/etiologia , Proteínas Supressoras de Tumor/genética , Dedos de Zinco/genética , Animais , Biomarcadores Tumorais , Linhagem Celular Tumoral , Transformação Celular Neoplásica/genética , Transformação Celular Neoplásica/metabolismo , Neoplasias Colorretais/metabolismo , Neoplasias Colorretais/mortalidade , Neoplasias Colorretais/patologia , Modelos Animais de Doenças , Feminino , Expressão Gênica , Técnicas de Silenciamento de Genes , Humanos , Mutação com Perda de Função , Camundongos , Fenótipo , Prognóstico , Ligação Proteica , Estabilidade de RNA , RNA Mensageiro/genética , Proteínas de Ligação a RNA/genética , Proteínas Repressoras/genética , Receptores Chamariz do Fator de Necrose Tumoral/genética , Proteínas Supressoras de Tumor/metabolismo , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Oncolytic virotherapy is a promising therapeutic strategy that uses replication-competent viruses to selectively destroy malignancies. However, the therapeutic effect of certain oncolytic viruses (OVs) varies among cancer patients. Thus, it is necessary to overcome resistance to OVs through rationally designed combination strategies. Here, through an anticancer drug screening, we show that DNA-dependent protein kinase (DNA-PK) inhibition sensitizes cancer cells to OV M1 and improves therapeutic effects in refractory cancer models in vivo and in patient tumour samples. Infection of M1 virus triggers the transcription of interferons (IFNs) and the activation of the antiviral response, which can be abolished by pretreatment of DNA-PK inhibitor (DNA-PKI), resulting in selectively enhanced replication of OV M1 within malignancies. Furthermore, DNA-PK inhibition promotes the DNA damage response induced by M1 virus, leading to increased tumour cell apoptosis. Together, our study identifies the combination of DNA-PKI and OV M1 as a potential treatment for cancers.
